Advantages of a Centrifugal Pump in the Development of a Swine Model of an Artificial Placenta

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Charest-Pekeski, Steven K.S. Cho, Tanroop Aujla, Liqun Sun, and 17 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1251735/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract The recent demonstration of normal development of preterm sheep in an artificial extrauterine environment has renewed interest in artificial placenta (AP) systems as a potential treatment strategy for extremely preterm human infants. However, the feasibility of translating this technology to the human preterm infant remains unknown. Here we report the support of 13 preterm fetal pigs delivered at 102±4 days (d) gestation, weighing 616±139g with a circuit consisting of an oxygenator and a centrifugal pump, comparing these results with our previously reported pumpless circuit (n=12; 98±4d; 743±350g). The umbilical vessels were cannulated, and fetuses were supported for 46.4±46.8 hours using the pumped AP versus 11±13 hours on the pumpless AP circuit. Upon initiation of AP support on the pumped system, we observed supraphysiologic circuit flows, tachycardia, and hypertension, while animals maintained on a pumpless AP circuit exhibited subphysiologic flows. On the pumped AP circuit, there was a progressive decline in UV flow and oxygen delivery. We conclude that the addition of a centrifugal pump to the AP circuit improves survival of preterm pigs by augmenting UV flow through the reduction of right ventricular afterload. However, we continued to observe the development of heart failure within a matter of days. General Cell Biology & Physiology Artificial placenta preterm pig cannulation centrifugal pump oxygenator tachycardia hypertension fetal development Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Introduction In resource rich nations, extreme prematurity, defined as delivery prior to 28 weeks’ gestation, remains the leading cause of childhood mortality and morbidity 1,2 . Survival rates decline with decreasing gestational age (GA), with only 6% surviving at 22 weeks GA compared to >90% surviving at 28 weeks GA 3 . In Canada alone, the economic burden of caring for extremely preterm infants approaches $600 million nationally per annum 4 . Unfortunately, despite advances in medical technology and neonatal intensive care, improvements in the outcomes of children born extremely preterm over the last 15 years have been limited 5 . At the biological limit of viability (22-25 weeks GA), preterm infants commence pulmonary gas exchange during the late-canalicular and early saccular stages of lung development. Exposure to the positive pressure mechanical ventilation and high partial pressure of inspired oxygen required to achieve adequate gas exchange results in cessation of alveolarization and pulmonary microvascular injury, which is associated with high rates of chronic lung disease and pulmonary hypertension 6 . Furthermore, approximately 50% of children born at the threshold of viability exhibit neurological disabilities at 30 months corrected age, with half of these cases being classified as severe 7 . Despite major advances in the outcomes of prenatal infants resulting from the widespread administration of prenatal steroids and exogenous surfactant, recent improvements in the morbidity and mortality have been more incremental, emphasizing the need for new innovative approaches to supporting the fragile physiology and development of extremely preterm infants 1-3 . An artificial placenta (AP) represents a novel approach that aims to maintain the innate fetal circulation while promoting normal prenatal development. Gas exchange is achieved with a low-resistance hollow-fiber membrane oxygenator connected to the fetus via the umbilical vasculature, while incubating the fetus in a fluid filled environment. The first attempt to support previable human infants was reported in the late 1950s 8 ; with subsequent progress made using animal models. These early experiments were complicated by the development of heart failure and infections and concurrent improvements in conventional neonatal intensive care led to diminished enthusiasm for AP technology 8,9 , 10 . However, over recent years, several research teams have demonstrated the feasibility of supporting preterm goat and sheep fetuses with a variety of pumpless and pumped arteriovenous and venovenous AP systems using different approaches to establishing vascular access 8,9,11–24 . Two research teams have successfully demonstrated physiologic fetal sheep hemodynamics with normal organ maturation and minimal injury for periods of up to one month on pumpless arteriovenous AP systems 17–19,23 . Despite these advances, AP is yet to be proven in a realistic animal model of the extremely preterm human infant. At a comparable stage of lung development to a previable human fetus, the preterm sheep is approximately twice the weight 25,26 . Fetal size determines important anatomical considerations for establishing an extracorporeal membrane oxygenation (ECMO) system such as blood vessel diameter and blood pressure. In addition, fetal sheep possess two umbilical arteries (UA) and two umbilical veins (UV). This may allow a more stable transition to the AP circuit using one pair of UA and UV while maintaining native gas exchange via the placenta using the other pair of umbilical vessels, thus minimizing the cessation of oxygenation from the mother 27,28 . By contrast, human and pig fetuses usually have one UV and two UAs, which may represent a more technically challenging approach to the initiation of AP support. We have previously proposed 27 that Yucatan miniature pigs delivered at approximately 95 days gestation represent a realistic model of human infants born at the lower limit of viability in terms of their body weight and stage of lung development 25 . We recently demonstrated the feasibility of cannulating the umbilical vessels of preterm minipigs and transferring them to an AP system consisting of a warm, fluid environment and a pumpless ECMO circuit comprising a commercial neonatal oxygenator 27 . However, our experiments were characterized by subphysiologic circuit flows, tachycardia, and the development of hydrops. Echocardiography further revealed evidence of diminished right ventricular function, which we attributed to excessive ventricular afterload resulting from the large priming volume of the circuit and small umbilical cannulas 27 . We hypothesized that the addition of a pump might improve the hemodynamics of our animals on the circuit by reducing afterload through the generation of negative pressure downstream of the umbilical arterial cannulas, thereby improving venous return and cardiac output. Herein, we sought to investigate the hemodynamics of a pumped AP system, comparing our findings to those obtained using the pumpless circuit 27 . Methods Experimental groups. The results presented in this report are comprised from fetal pigs studied in utero and while maintained using a pumped AP circuit and of our previously published findings using a pumpless AP system 27 . Fetal Large White Landrace pigs were studied in utero at 98, 106, and 112 days GA (n = 24 ) and Yucatan miniature pigs at 107 ± 3 days (n = 16), providing reference physiologic data regarding heart rate (HR), blood pressure, blood gases, electrolytes, lactate and glucose concentrations, and UV flow 27 . Fetal Yucatan miniature pigs (n =13) were maintained using an umbilical arteriovenous AP circuit consisting of a centrifugal pump and oxygenator and using a pumpless AP circuit (n = 12) 27 . We included AP subjects that survived on the system for a minimum of 3 hours. Animals and approvals . In utero BP, HR, blood gases, electrolytes, glucose, and lactate measurements were obtained in White Landrace Cross sows (n = 6; term = 115 days) at the Preclinical Imaging and Research Laboratories, South Australian Health and Medical Research Institute (SAHMRI). All procedures were approved by the SAHMRI Animal Ethics Committee 27 . Sows were individually housed with environmental and social enrichment. The AP experiments were conducted in the Lab Animal Services (LAS) facility at The Hospital for Sick Children (SickKids), Peter Gilgan Center for Research and Learning in Toronto, Ontario. All maternal and fetal surgeries were approved by the SickKids Animal Care Committee and all procedures complied with the Canadian Council on Animal Care, Ontario Ministry of Agriculture, Food and Rural Affairs, Animals for Research Act guidelines, and the Care and Use of Animals for Scientific Purposes. All research and LAS staff members acted in accordance with ARRIVE guidelines proposed by Kilkenny et al. 54 . Pregnant Yucatan miniature pigs (n = 46; term = 115 days) were acquired from Memorial University of Newfoundland and Sinclair Bioresources and transported as per the Health of Animals Act of Canada 27 . Yucatan pigs were housed in pairs for at least two weeks prior to surgery to increase socialization between the animals and allow for acclimation to human handling and to their new environment. Sows were provided with ad libitum food and water, and environmental enrichments as per SickKids standard operating procedures. Protocol for in utero studies. Large White Landrace Cross gilts (n = 6; 98 ± 7 days GA; term = 115 days) were anaesthetised with an intramuscular injection (I.M.) of 20 mg/kg ketamine and inhalation of isoflurane. Gilts were intubated and general anaesthesia was maintained using isoflurane with 2 L/min O 2 and 4 L/min medical air. Gilts were positioned on the operating table on their backs, an incision was made along the abdomen, the uterus was incised, and a fetal head was exposed. Fetal pigs (n = 24) were cannulated via the UV, and venous blood was sampled for partial pressure of oxygen (PO 2 ), partial pressure of carbon dioxide (PCO 2 ), oxygen saturation (SO 2 ), pH, hemoglobin (Hb), bicarbonate (HCO 3 - ), base excess (BE), sodium (Na + ), potassium (K + ), and calcium (Ca 2+ ) as previously described 27,28 . In a subset of fetuses (n = 21, 105 ± 7 days GA), the carotid artery (CA) was instrumented and fetal BP, and HR were measured and continuously recorded in LabChart 8 Pro (ADInstruments Inc., Colorado Springs, USA) 27 . Following in utero experiments gilts and their fetal pigs were humanely euthanized with an intravenous overdose of sodium pentobarbital (Virbac, New South Wales, Australia). Surgical protocol for pumped AP studies. Pregnant Yucatan pigs (n = 19; 101 ± 4 days GA; term = 115 days) were anaesthetized with an I.M. injection of 10 mg/kg ketamine hydrochloride, 0.20 mg/kg acepromazine, and 0.015 mg/kg atropine sulfate (CDMV Inc., Saint-Hyacinthe, Canada), with maintenance of general anaesthesia with inhalation of 2-3% isoflurane (Fresenius Kabi Canada, Toronto, Canada). To prevent aortocaval compression, anaesthetized sows were positioned on the operating table in the left lateral position. Umbilical blood flow was measured in utero in 4 Yucatan sows in Toronto using cine phase contrast MRI as described in our previous publication. 27 A lower antero-lateral laparotomy was performed for a caesarean section. Following the delivery of all fetal pigs, sows were humanely euthanized with 106 mg/kg Euthanyl (CDMV Inc., Saint-Hyacinthe, Canada). Fetal surgical procedures. In a subset of fetuses (n = 11/13 successful pumped AP experiments), a small incision was made along the right side of the neck to expose the jugular vein (JV) and CA 28,55 . A size-matched custom-made PVC tubing was then inserted in the JV and CA for monitoring of central venous pressure (CVP) and fetal mean arterial pressure (MAP), respectively. The tubing was stabilized using silk sutures, and the neck incision sutured closed. Fetuses were delivered to minimize excessive torsion and stretching of the umbilical cord and were positioned on the maternal abdomen and subsequently weighed. Fetal pigs were then anaesthetized using an I.M. injection of 1 mg/kg rocuronium bromide (Sandoz Inc., Mississauga, Canada), and 5 mg/kg ketamine hydrochloride (CDMV Inc., Saint-Hyacinthe, Canada). Fetal normothermia was maintained by continuously bathing the fetus and the umbilical cord in warmed normal saline. The umbilical cord was treated with a topical application of 100 mg/kg papaverine hydrochloride (Sandoz Inc., Boucherville, Canada). In all AP studies, umbilical catheters were then placed in both UAs and the UV (12-gauge and 10-gauge custom-made cannulas, respectively), and secured into place using silk sutures (Ethicon Inc., New Jersey, USA) at the insertion of the cannula within the vessel. In smaller fetuses, UA cannulas were downsized to 14 GA angiocaths (Becton Dickinson Canada Inc., Mississauga, Canada). Cannulas were then connected to the AP circuit described below, via modified ¼” perfusion adapters (Medtronic of Canada Ltd., Brampton, Canada) to facilitate the initiation of UV flows. Re-positioning of the umbilical catheters was performed as needed to assist in the establishment of flow. AP circuit design. The arteriovenous AP circuit consisted of a commercially available centrifugal pump, connected to a low-resistance membrane oxygenator, and modified commercially available intravenous umbilical arterial and venous cannulas via ¼” internal diameter x 1/16” wall thickness, 3/16” x 1/16” TYGON PVC tubing with P.h.i.s.i.o coating (LivaNova PLC., London, England). In two experiments, a commercially available cardiopulmonary bypass oxygenator was used (Table 2); however, this was subsequently changed to another commercially available ECMO oxygenator (n = 11; Table 2). Total priming volumes for these neonatal membrane oxygenators were 40 and 65 ml, respectively. Circuits were then initially primed with plasmalyte (Baxter Inc., Mississauga, Canada) and then replaced with heparinized maternal blood, which was continuously recirculated to prevent clotting within the circuit and warm the blood prior to connection to the fetus. UA blood passed through the oxygenator inlet, exiting through the outlet before returning to the heart via the UV (Figure 1). The sweep gas supplied the oxygenator with a mixture of medical air and oxygen and was titrated to achieve normal physiologic levels of UV PO 2 of 35-40 mmHg and a PCO 2 of 50-60 mmHg 29 based on in utero fetal pig data. Fluid incubation Following cannulation, fetal pigs were enclosed in a custom-made silicone infused thermoplastic polyurethane film ‘biobag’. Approximately 2-4 litres of lactated ringer (LR; Baxter Inc., Mississauga, Canada) crystalloid solution was warmed to 39 ± 1°C in a large fluid reservoir and circulated via the inflow and outflow ports of the Biobag every 10 hours. The Biobag was equipped with a temperature port that facilitated monitoring of fluid temperature using a temperature probe (ADInstruments Inc., Colorado Springs, USA). The biobag was covered to prevent transmission of light to the fetus to better simulate uterine conditions and maintained at an appropriate temperature using a contact heat pad underneath the biobag and an overhead heater. Fetal pig maintenance on the AP circuit. Following cannulation and transition to the AP, fetal pigs received a maintenance infusion of 6 µg/kg/h prostaglandin E1 (Pfizer Canada Inc., Kirkland, Canada) with 100 units/kg/h heparin (Fresenius Kabi Canada, Toronto, Canada) to maintain patency of the ductus arteriosus, and prevent clotting in the AP circuit 19 . Dextrose (418 mg/kg/h; Pfizer Canada Inc., Kirkland, Canada) was delivered via the circuit for the first 8 hours of AP support, which was then exchanged for total parenteral nutrition solution with the goal of supporting fetal energy requirements and maintaining fetal glucose concentrations of > 100 mg/dL. Calcium chloride (80 mg/kg; Omega Laboratories Ltd., Montreal, Canada) and heparin (100 units/kg) were dosed empirically to achieve a target Ca ++ concentration of > 1.4 mmol/L, and an activated clotting time (ACT) of 250-300 seconds, respectively. In early experiments (n = 7/13 successful experiments), 6 mg/kg/h of papaverine was administered I.V. to prevent vasospasm of the umbilical vessels and augment circuit flows. In a subset of experiments (n = 3/13), this was subsequently changed to a maintenance infusion of 30 µg/kg/h milrinone lactate (Aurobindo Pharma, Hyderabad, India). Reconstituted hydrocortisone (8 mg/kg/day, Pfizer Canada Inc., Kirkland, Canada) and a broad-spectrum empirical antibiotic (piperacillin/tazobactam; 300 mg/kg/day, Sandoz Inc., Boucherville, Canada) was administered I.V., every 6 hours, and 8 hours, respectively. Whole maternal blood was transfused (10 ml/kg) to replete circulating blood volume following circuit phlebotomy. In addition, rocuronium bromide (1 mg/kg) and ketamine hydrochloride (5 mg/kg) were given for excessive fetal movements or perceived fetal agitation. Albumin (25%; CSL Behring Canada Inc., Ottawa, Canada) and furosemide (0.5 mg/kg; Pfizer Canada Inc., Kirkland, Canada) were given intermittently during longer experiments to address perivascular edema and hydrops fetalis. Fetal CA, and umbilical arterial and venous blood gases including PO 2 , PCO 2 , pH, Hb, HCO 3 - , SO 2 , BE, Na + , K + , lactate, glucose and ACT were sampled every 1-3 hours and analyzed using a handheld blood analyzer (Abbott Point of Care Inc., Nepean, Canada). Fetal oxygen delivery (DO 2 ) and consumption (VO 2 ) were calculated based on the combination of UV and UA oxygen carrying capacity and indexed UV flow indexed to fetal weight (measured at surgery). Physiologic monitoring. UV flow was continuously measured using a HXL 3/16” tubing flow probe (Transonic) and CVP, fetal BP, pre-pump circuit pressure, pre-oxygenator circuit pressure, and post-oxygenator circuit pressure were measured using Deltran fluid filled blood pressure transducers (ADInstruments Inc., Colorado Springs, USA). Data were sampled at 1000 Hz, digitized, and continuously recorded using LabChart Pro 8 (ADInstruments Inc., Colorado Springs, USA). At the end of each study, the data was extracted in consecutive 30-second intervals and analyzed in Excel (Microsoft Corporation, Washington, USA). Quantification of UV blood flow in utero using MRI: 3-D Volumetry and cine phase-contrast. In utero fetal weight and UV flow were measured in sixteen fetuses from 4 pregnant Yucatan pigs (n = 4; gestational age 107 ± 3 days GA) as previously described using a 3 Tesla magnetic resonance imaging (MRI) system 27 . Statistical analysis. Comparisons of anthropometric data between pumpless and pumped AP circuits were analyzed using a two-way ANOVA with Bonferroni correction for multiple comparisons. Comparison of the duration of AP support between pumpless and pumped circuits was analyzed using a Mann-Whitney U test. Changes in temperature, HR, and UV flow between pumpless and pumped AP circuits over the first 3 hours of support were analyzed using a mixed-effect model, with a Bonferroni correction for multiple comparisons. Differences in mean indexed and absolute UV flow and HR between in utero , pumpless and pumped AP circuits were compared using a repeated measures one-way ANOVA with a Bonferroni correction for multiple comparisons. Analysis of fetal HR and indexed UV flow were performed using a linear regression. Comparisons of mean blood gases, electrolytes, lactate, and glucose concentrations between animals studied in utero , or on pumpless and pumped AP circuits were analyzed using a repeated measure one-way ANOVA, and a Kruskal-Wallis test (when appropriate) with Bonferroni correction for multiple comparisons. UV flow, HR, fetal BP, CVP, and temperature are presented in 5-minute averages. Time post-cannulation versus UV flow is presented in 1 hour averages and analyzed using a one-way ANOVA with Bonferroni correction for multiple comparisons. Differences in MAP between fetal pigs studied in utero and those maintained using a pumped AP were analyzed using a mixed-effect model with Bonferroni correction for multiple comparisons. Indexed UV flow versus oxygen extraction was analysed using a linear regression. * P < 0.05 was considered statistically significant. All statistical analyses were performed using Prism 9 (GraphPad, San Diego, USA). Data are presented as mean ± standard deviation (SD), unless otherwise indicated. Results The addition of a centrifugal pump improves successful transition to AP and support times. Nineteen sows and 89 fetal pigs were used for pumped AP experiments (Table 1). A total of 32 fetal pigs were cannulated and transitioned to the AP. Types of perioperative complications preventing us from maintaining AP support were similar to those experienced in the pumpless AP circuit group and included the inability to establish circuit flows upon connection to the circuit (n = 11), accidental decannulation (n = 4), persistent vasospasm of the umbilical cord (n = 3), and excessive spiralling of the umbilical cord, which prevented cannulation or adequate cannula position (n = 1; Table 1). We successfully maintained 13 fetal pigs at an initial GA of 102 ± 4 days (range = 93-107 days) with a body weight of 616 ± 139 g (range = 390-820g) on a pumped AP circuit for 46.4 ± 46.8 hours (range = 3.4-177.8 hours). This represented a significant improvement in the duration of support over the 12 fetal pigs (98 ± 4 days GA; 743 ± 350 g; 11 ± 13 hours) maintained using a pumpless AP circuit ( P = 0.009; Table 1 and Figure 2). There was also a marked improvement in the rate of successful AP runs per litter using a pumped AP circuit compared to the pumpless AP circuit (62% pumped AP vs. 28% pumpless AP; Table 1). Successfully supported fetuses were similar in weight and GA in both pumped and pumpless groups (616 ± 139g vs. 743 ± 350g, P = 0.41; 102 ± 4 days vs. 98 ± 4 days, P = 0.24). On the pumped and pumpless AP circuits, fetal body weights were similar between successfully and unsuccessfully supported animals (616 ± 139g vs. 523 ± 145g, P = 0.47; 743 ± 350g vs. 644 ± 184g, P = 0.53). However, unsuccessfully supported fetuses or fetuses that we did not attempt to cannulate were significantly smaller in the pumped group compared to the pumpless group (523 ± 145g vs. 644 ± 184g, P = 0.04; 557 ± 124g vs. 892 ± 362g, P < 0.0001). Fetal weight at the time of termination of the AP study was significantly higher compared to pre-cannulation weight. Fetal weight gain correlated strongly with length of AP support (r = 0.89), with weight at the end of the experiment exceeding the expected growth trajectory of these fetuses. Individual data on supported fetuses, support times, and reasons for termination are shown in Table 2. Table 1. Summary of fetal pigs from pumped and pumpless AP circuits 27 . Pumped AP circuit Pumpless AP circuit Intervention Intervention Parameter Maintained on AP Cannulated but not maintained on AP Did not attempt to cannulate Maintained on AP Cannulated but not maintained on AP Did not attempt to cannulate N 13 AP success rate per litter (%) 19 57 12 AP success rate per litter (%) 56 9 P-values : Pumpless vs. Pumped AP circuit (ANOVA results) 62 28 Circuit type Intervention Interaction Body weight (g) 616 ± 139 (390-890) 523 ± 145 (244-710) * 557 ± 124 (372-941; n = 24) * 743 ± 350 (500-1500; n = 7) ab 644 ± 184 (343-1390; n = 52) a 892 ± 362 (531-1470) b <0.0001 0.002 0.02 GA (days) 102 ± 4 (93-107) 103 ± 4 (93-108) 102 ± 5 (93-108) 98 ± 4 (93-107) 101 ± 7 (93-112) 101 ± 7 (97-110) 0.04 0.34 0.60 Two-way ANOVA. Multiple comparisons for fetal body weight and GA within pumped and pumpless AP circuits and between the intervention groups: ab P < 0.05. * P < 0.05 body weight is significantly different between pumped and pumpless AP circuits within intervention groups (i.e., body weight maintained on a pumped AP circuit vs. body weight maintained on a pumpless AP circuit). Interpretation of numbers for multiple comparisons: a represents a statistically significant difference compared with b . Table 2. Summary data of 13 fetal pigs successfully cannulated and maintained using a pumped AP circuit. Data are presented by increasing GA and are expressed as mean ± SD. GA before (days) Weight before (g) Duration on AP (hours) GA after (days) Weight after (g) Sex (M, F) Reason for termination Oxygenator 93 390 36.7 95 450 F Fetal movement occluded UV flow O 98 450 3.4 98 450 M Heart failure O 100 505 14.1 101 NA M Mechanical failure (pump) O 100 520 15.1 101 620 M Heart failure BP 100 590 49.4 102 750 F Heart failure O 101 762 70.0 104 1190 M Mechanical failure (oxygenator) BP 102 480 177.8 109 1176 M Hydrops, cardiac dysfunction O 105 610 68.5 108 NA NA Mechanical failure (oxygenator) O 105 676 17.1 105 NA M Fetal movement occluded UV flow O 105 690 53.5 108 NA M Fetal movement occluded UV flow O 105 730 4.2 105 730 M Thrombosis development in circuit O 105 780 21.1 106 879 M Equipment failure (sweep-gas supply) O 107 820 72.5 110 876 F Fetal movement occluded UV flow O 102 ± 4 616 ± 139 46.4 ± 46.8 104 ± 4 792 ± 272 (n = 9) Abbreviations: NA, not available; M, male; F, female; GA, gestational age; O, commercially available ECMO oxygenator; BP, commercially available bypass oxygenator. Pumped AP circuits achieve higher umbilical venous flow. Animals on a pumped AP circuit maintained a physiologic core body temperature throughout the experiment. The temperature was lower in fetal pigs supported on a pumped system than pigs supported on a pumpless system ( P < 0.0001, Figure 3A). UV flow rates of fetuses supported by the pumped AP circuit were comparable to in utero controls, while UV flow on the pumpless AP system was subphysiologic ( in utero ; 108 ± 24 ml/min vs. pumped; 87 ± 28 ml/min vs. pumpless; 70 ± 18 ml/min, P = 0.001; in utero ; 173 ± 45 ml/min/kg vs. pumped; 143 ± 40 ml/min/kg vs. pumpless; 97 ± 39 ml/min/kg, P = 0.005; Figure 3C, E and F). Absolute UV flow measured within the first 3 hours post-cannulation was positively correlated to HR in pumped AP circuits (r 2 = 0.44; P < 0.0001; Figure 3G), but negatively correlated in fetuses maintained on a pumpless circuit (r 2 = 0.45; P < 0.0001; Figure 3G), with differences between the slopes being extremely significant ( P < 0.0001). Figures 5A-M demonstrate the changes in UV flow, HR, temperature, MAP, and CVP over time for the 13 fetal pigs successfully maintained using a pumped AP circuit. Upon initiation of AP support, we observed supraphysiologic UV flow, despite a low pump rate setting (Figure 5A-N). In longer trial runs, this period of supraphysiologic UV flow persisted for approximately 6 hours, then decreasing to subphysiologic flow, plateauing at ~12 hours of support (Figure 5O). In several experiments (n = 8/13) excessive spasmodic fetal movements resulted in a complete cessation of UV flow and consequently led to the termination of the experiment (Table 2). In four studies, development of a thrombosis in one of the UAs in the first 12 hours of support was associated with a significant reduction in UV flow. Fetal pigs were tachycardic for a large proportion of the AP studies (Figure 3B and D; and Figures 5A-M). Both pumped (205 ± 28 bpm; P < 0.0001; Figure 3D) and pumpless (206 ± 38 bpm; P < 0.0001; Figure 3D) fetal pig groups were similarly tachycardic throughout the experiment compared to in utero controls (130 ± 10 bpm; Figure 3D). Compared to in utero control animals, fetal pigs were hypertensive on the pumped AP system ( P = 0.013; Figure 6). Approximately 20 minutes after initiation of a milrinone infusion, there was a marked increase in UV flow, followed by a steady decline over a 3 hour period (Figure 7A). MAP and HR decreased following the start of the milrinone infusion, which persisted for ~1 hour before returning to a steady state (Figure 7B and C.). Upon initiation of AP support, fetal oxygen delivery was physiologic 29,30 . We observed a decline in oxygen delivery and increase in oxygen consumption over time (Figure 8A). UV flow and oxygen extraction fraction correlated negatively (r 2 = 0.45; P = 0.0003; Figure 8B). Fetal blood gas analysis indicates supraphysiologic oxygen tension and fetal anemia While there was no significant difference in SO 2 between fetal pigs studied in utero and those maintained using a pumped or pumpless AP circuit (Figure 4A), both AP circuits resulted in a significantly higher PO 2 compared to in utero controls ( in utero ; 53 ± 16 mmHg vs. pumped; 141± 129 mmHg, P = 0.0001, vs. pumpless; 280 ± 176 mmHg, P = 0.003, Figure 4B). Conversely, PCO 2 was significantly lower in the pumped AP group compared to in utero controls ( in utero ; 67 ± 11 mmHg vs. pumped; 49 ± 11 mmHg, P = 0.0009, Figure 4E). Animals on the pumped AP were significantly more alkalotic than both pumpless AP fetuses and in utero controls ( in utero ; 7.29 ± 0.08 vs. pumped; 7.36 ± 0.07, P = 0.03, vs. pumpless; 7.30 ± 0.06, P = 0.03, Figure 4F). BE and lactate were similar between groups (Figure 4G, H). We identified significantly lower hematocrit ( in utero; 28 ± 3%; vs. pumped; 24 ± 3%, P = 0.02, Figure 4C) and hemoglobin in the pumped AP group ( in utero; 96 ± 9 g/L vs. pumped: 82 ± 11 g/L, P = 0.01, Figure 4D), as well as higher Na + in both AP supported groups compared to intrauterine fetuses ( in utero ; 126 ± 2 mmol/L vs. pumped; 133 ± 5 mmol/L, P < 0.0001; vs. pumpless; 133 vs. 3 mmol/L, P < 0.0001, Figure 4L). Ca 2+ concentrations were subphysiologic in both pumped and pumpless groups ( in utero ; 1.56 ± 0.087 vs. pumped; 1.44 ± 0.092 mmol/L, P = 0. 003, vs. pumpless; 1.29 ± 0.21 mmol/L, P = 0. 001, Figure 4K). Glucose, administered continuously either directly or as part of total parenteral nutrition, was significantly higher in both AP groups, compared to in utero controls ( in utero ; 2.1 ± 0.43 mmol/L vs. pumped; 8.5 ± 4.8 mmol/L, P < 0.0001, vs. pumpless; 9.8 ± 5.8 mmol/L, P < 0.0001, Figure 4I). There was no difference in K + concentrations between in utero fetal pigs and those maintained using a pumped and pumpless AP circuit (Figure 4J). Discussion To our knowledge, this is the first study to examine the hemodynamics and blood gas status of preterm pigs supported using an AP system incorporating a small centrifugal pump. We previously showed the feasibility of supporting our animal model using a pumpless AP circuit connected to the fetal circulation via the umbilical arteries and vein 27 ; however, our experiments were characterized by significant hemodynamic decompensation within hours. We attributed this to afterload imbalance, elicited by supraphysiologic circuit resistance and further exacerbated by impaired umbilical venous return, which we surmised resulted in higher sympathetic tone, further exacerbating the increase in afterload 27 . Although other research groups have highlighted potential limitations of pumped ECMO circuits, including an increased risk of pump-induced hemolysis 17 , afterload imbalance, myocardial strain, and impaired autoregulation of UV blood flow 31 , we hypothesized that the addition of a small pump might reduce right ventricular afterload and sustain adequate UV flow, thereby improving survival on the AP. The present study examined differences in hemodynamics, blood gases, electrolytes, biochemistry, and survival of fetal pigs supported on a pumped versus pumpless AP system 27 . The increase in AP support to 46.4 ± 46.8 hours represented a significant increase in survival over fetal pigs supported without a pump possibly due to more reliable and physiologic UV flow compared to a pumpless system. Despite the pump support, following a period of supraphysiologic UV flow, the animals continued to experience a slower but consistent decline in UV flow and the development of signs of circulatory deterioration. We have yet to establish an AP system capable of meeting the long-term physiologic needs of animals that are similar in size to the extremely preterm human infants that are the proposed target of this approach. In a recent report, Partridge et al. 17 demonstrated AP support of fetal sheep weighing 1-2 kg using a pumpless system for up to 28 days without the use of vasopressors. Using a similar AP circuit, Usada et al. 18 reported survival of fetal sheep for up to one week. To investigate the potential utility of AP systems for extremely preterm infants born at 22-25 weeks GA, two research groups cannulated sheep born at 85-95 days GA weighing ~0.5-0.8 kg. Although cannulation was technically feasible, one study reported the development of hydrops fetalis following 5-8 days of support 32 , whereas the other required the use of aggressive pharmacological interventions in order to achieve hemodynamic stability 19 . Although fetal sheep delivered at 85-95 days GA share similar body weight to human fetuses delivered at the biological limit of viability, they are developmentally immature and their pulmonary development is analogous to a human fetus delivered at 18 weeks GA 10,26,33,34 . By contrast, fetal minipigs delivered at 93-107 days GA weighing 0.39-0.82 kg are in their canalicular and saccular stages of lung development and comparable to preterm human fetuses delivered at 22-28 weeks GA. Thus, fetal pigs delivered at 95 days GA would therefore be equivalent in body weight and lung maturity to human fetuses born at 22-25 weeks GA, and may represent a more appropriate model for the development of an AP system 27,33,34 . Upon initiation of AP support, we observed supraphysiologic UV flow in all pumped AP experiments. Umbilical vein blood flow was higher in fetuses supported with a pumped circuit than in fetuses supported using a pumpless system. Our experiments revealed a positive correlation between fetal HR and UV flow on the pumped AP, with persistent tachycardia seen with both systems. Contrary to our speculations regarding supraphysiologic resistance in the pumpless circuit 27 , we propose that the negative pressure generated by the centrifugal pump may have diminished right ventricular afterload and minimized resistance to flow across the UA cannulas, resulting in higher than normal flow rates. The association we observed between fetal HR and UV flow in animals supported on the pumped circuit may be attributable to the Frank-Starling mechanism and Anrep effect 35 , whereby increased ventricular preload enhances myocardial stretch and tension, resulting in increased contractility. Although this relationship holds true for slight increases in end-diastolic filling pressures, the fetal heart has little preload reserve 36–39 . Thus, further augmentation of cardiac output and contractility are also driven by the Bowditch effect 40 through increases in fetal HR 41 . In contrast to the physiology seen in a pumpless circuit, we observed a shift of placental-to-systemic blood flow in favor of the AP, effectively volume loading the fetal heart. Therefore, we speculate that these supraphysiologic circuit flows lead to increased preload, tachycardia, sympathetic nervous system activation, and likely higher cardiac output within the first hours of AP support. After ~24 hours of AP support, we observed marked reductions in UV flow compared to the high flows seen at the start of AP support and in many experiments, UV flows then remained subphysiologic. The drop in UV blood flow and persistent tachycardia could be explained by increased sympathetic nervous activation and continuous peripheral vasoconstriction. Although the introduction of milrinone improved UV flow and reduced BP, its inotropic and systemic vasodilatory effects appeared to be short-lived, indicating the presence of overwhelming perturbation of fetal cardiac loading conditions. However, our findings suggest that phosphodiesterase inhibitors may have a positive impact on the circulation of fetal animals supported on the AP 19,42 . Despite similar SO 2 , blood gases showed higher PO 2 , lower PCO 2 , as well as a more alkalotic pH in fetuses on the pumped AP compared to in utero controls. We were able to manage O 2 uptake and CO 2 elimination better on the pumped circuit than pumpless. However, the gas exchange capacity of the oxygenator used in both groups easily outperforms the native placenta, resulting in supraphysiologic UV PO 2 . High oxygen tension is a known contributor to umbilical cord and ductal constriction and may have negatively affected our experiments 43,44 . Gradual umbilical vessel spasm or constriction at the umbilicus, where the cord passes through the abdominal wall, could explain the gradual decline in UV flow we observed. Mean blood pressure in the UV is 6-8 mmHg in near-term sheep and human fetuses 29,30 . In our pumped AP studies, we observed pre- and post-oxygenator pressures of 25 mmHg and 18 mmHg, respectively (Supplementary Figure 1.). Although we did not measure blood pressure in the UV, it is likely that the pumped circuit exposed the umbilical vein to elevated blood pressure, even if there was some degree of pressure difference across the venous cannula. Berman et al. 45 advanced balloon catheters into the common UV of fetal sheep and demonstrated marked reductions in placental blood flow with increasing UV pressures. This was attributed to a decreased pressure gradient across the umbilical circulation, resulting in diminished umbilical flows. We hypothesize that a similar phenomenon may be occurring in the early phase of our AP experiments, with UV constriction induced by the high pressures and shear stress resulting from the elevated post-pump pressures. We observed a steady widening of the veno-arterial saturation gradient across the umbilical circulation during our AP runs that was negatively correlated with UV flow, indicating higher oxygen extraction at the tissues in the setting of a net reduction in UV flow and oxygen delivery. One factor that might additionally contribute to the progressive reduction in oxygen delivery was the low hemoglobin concentration we observed in animals supported on the pumped system. The large priming volume of the circuit and oxygenator contributes to fetal anemia because the circuit is primed with maternal blood. During pregnancy, maternal blood volume increases by ~40% and there is a progressive decline in hemoglobin concentrations 46–48 . Conversely, fetal hemoglobin increases with GA to sustain fetal tissue oxygenation in the setting of declining PO 2 29,49,50 . Thus, the mixing of maternal and fetal blood in the AP circuit contributes to hemodilution of fetal blood, while ongoing blood sampling further depletes fetal red blood cells. To circumvent this problem in future experiments, it may be possible to hemoconcentrate the circuit prime. Furthermore, pump-induced hemolysis may have also contributed to fetal anemia in these experiments. Other groups have recommended the administration of daily doses of erythropoietin to minimize the need for successive maternal donor blood transfusions 17 . Miniaturization of the oxygenator and pump head could also help to reduce fetal anemia by reducing circuit volume. Study limitations. There are several limitations of the present study. Firstly, we have not yet characterized the etiology of the variation in UV flows on the pumped AP circuit. We speculate that a combination of UV constriction and supraphysiologic UV pressures, as well as supraphysiologic oxygen tension may be contributing to progressively diminished UV flows. However, this hypothesis should be tested with direct measurements of UV pressure. Similarly, future experiments would be strengthened by the simultaneous measurement of umbilical artery pressure. We speculate that adrenergic drive may be contributing to tachycardia and peripheral vasoconstriction on the AP that would be enhanced by the measurement of circulating catecholamines. Secondly, although the addition of a centrifugal pump to the AP improved fetal pig hemodynamics, we periodically encountered air-entrainment into the circuit, as well as cavitation of fetal blood at excessively high negative pressures, resulting in diminished circuit flows. Although we did not detect pump-induced hemolysis, this could be a limiting factor during long-term AP experiments and could explain the fetal anemia on the circuit. Finally, systemic inflammation may have also contributed to the hemodynamic instability and cardiovascular decompensation we observed in pumped AP fetal pigs as centrifugal pumps have been reported to induce low-level systemic inflammation 51 . The analysis of inflammatory cytokines may be helpful to delineate the role of inflammation in fetal pig circulatory physiology on the AP. Future directions. Recent developments in AP technology have enabled sustained extrauterine fetal life on a pumpless arteriovenous ECMO circuit. Of note, these experiments were conducted using customized hollow-fiber membrane oxygenators that are unavailable for commercial use. Our initial approach in developing a pumpless AP system involved cannulas with the largest luminal diameter to wall thickness ratio possible to minimize circuit resistance. Our failure to demonstrate hemodynamic stability using a pumpless circuit encouraged our team to explore the use of mechanical support in the extremely preterm fetal pig. While the addition of a pump to the circuit improved survival, we observed supraphysiologic circuit flows, evidence of adrenergic drive, and high cardiac output at the start of support, followed by diminished UV flows with persistent tachycardia and hypertension in keeping with diminished cardiac output. This pattern may indicate a progressive increase in circuit resistance resulting from constriction in the cord vessels. An alternative approach to normalizing UV pressures and minimizing the pressure difference across the UV cannula would be to introduce a pressure drop in the circuit through restriction of the circuit or cannula lumen. According to Hagan-Poiseuille’s law, reducing the calibre of the venous side of the circuit would increase circuit resistance and limit the supraphysiologic circuit blood flow seen at the onset of our AP experiments. Miniaturization of the oxygenator and pump head may also help to achieve hemodynamic stability in our animal model. In sheep, placental blood volume at ~116 days GA (term = 145 days) is ~60-80 ml/kg, which represents ~30-40 ml for a 0.5 kg fetus 52,53 . The AP circuit used in the present study has a priming volume that is nearly double the expected placental blood volume for a fetal sheep delivered at 116 days gestation, which likely accounts for the increased afterload we seemed to induce in our pumpless AP experiments, as well as contributing to fetal anemia. Reductions in the oxygenator membrane surface area could also reduce the need for excessive heparin administration. Thus, modifications to the circuit through miniaturization of the oxygenator and centrifugal pump may be helpful for establishing an AP system capable of supporting the long-term physiological requirements of extremely preterm fetal pigs. Conclusion Over the past decade, important advances have been achieved in the field of artificial womb/AP technology. In experiments using a miniature pig model of the AP in which commercial neonatal ECMO oxygenators were connected the fetal circulation via the umbilical cord we demonstrated a marked improvement in survival, UV flow, and oxygen delivery in circuits that incorporated a small centrifugal pump compared with animals maintained on a pumpless circuit. However, despite the addition of the pump, we observed a progressive diminution of UV flow with persistent tachycardia and hypertension, which we attribute to preload imbalance, increased sympathetic tone, and UV hypertension. Thus, despite observing a clear short-term benefit with the addition of a centrifugal pump in supporting the fetal pig hemodynamics, we conclude that further modifications to the AP circuit are needed. A reduction to the size of the UV cannula could represent an alternative approach for mediating physiologic circuit flows and venous pressures. Given the limitations of neonatal intensive care therapies in preventing iatrogenic organ injury and neonatal death, we remain hopeful that the AP could provide reductions in the mortality and morbidity associated with preterm birth by maintaining a fetal circulation while allowing for normal growth and development of fetal organ systems. Declarations Competing interests The authors have no competing interests to disclose. Author Contributions Conception or design of the work: MS, JLM, CH. Data acquisition: AJCP, SKSC, TA, LS, AAF, MJM, AS, ME, LCL, CF, DM, JB, BSS, JL, OJM, FL, JRTD, MS, JLM, CH. Data analysis or interpretation: AJCP, SKSC, TA, AAF, MJM, MQ, JRTD, MS, JLM, CH. Drafting the work or revising it critically for important intellectual content: AJCP, SKSC, TA, LS, AAF, MJM, JB, JRTD, MS, JLM, CH. All authors approved the final version of the manuscript, agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved, and all persons designated as authors qualify for authorship, and all those who qualify for authorship are listed. Funding The artificial placenta experiments were generously supported by funding from the Canadian Institutes of Health Research and Breakfast of Champions and BMO Financial Group Chair in Cardiology to MS. Landrace pig studies and JLM were funded by an ARC Future Fellowship (Level 3; FT170100431). BSS was funded by SickKids Research Institute’s graduate scholarship program, Restracomp. 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DISTRIBUTION OF BLOOD BETWEEN INFANT AND PLACENTA AFTER BIRTH. The Lancet 294 , 871–873 (1969). Kilkenny, C., Browne, W. J., Cuthill, I. C., Emerson, M. & Altman, D. G. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 8 , e1000412 (2010). Schrauben, E. M. et al. Open or closed: Changes in ductus arteriosus flow patterns at birth using 4D flow MRI in newborn piglets. Physiol. Rep. 9 , e14999 (2021). Additional Declarations No competing interests reported. Supplementary Files SupplementaryFigure1PumpedAPPaper.pdf Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-1251735","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":78171235,"identity":"920b3df3-4d3a-42b1-a9c5-fbb353efa667","order_by":0,"name":"Alex J. 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Morrison","email":"","orcid":"","institution":"University of South Australia","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Janna","middleName":"L.","lastName":"Morrison","suffix":""},{"id":78171255,"identity":"270d487d-f9ca-4a08-b85e-30b2eac6c1f0","order_by":20,"name":"Christoph Haller","email":"data:image/png;base64,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","orcid":"","institution":"The Hospital for Sick Children","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Christoph","middleName":"","lastName":"Haller","suffix":""}],"badges":[],"createdAt":"2022-01-11 23:29:05","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-1251735/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-1251735/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":17605626,"identity":"a09472c4-364f-4392-925f-e7cc9b035eb6","added_by":"auto","created_at":"2022-01-24 18:37:48","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":94037,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eLayout of the pumped AP circuit. \u003c/strong\u003eFetal pigs (n = 13) were delivered via caesarean section, cannulated via the UA’s and UV, and maintained using a pumped AP circuit. Fetal pigs (n = 12) were also maintained using the same circuit design but without a centrifugal pump. Blood passed through the UAs and the centrifugal pump before entering the inflow port of the oxygenator. Oxygenated blood then passes through the oxygenator outflow port before returning to the heart via the UV. Sweep gas supplied the oxygenator with medical air and oxygen and the Biobag was filled with warmed LR solution to maintain fetal normothermia.\u0026nbsp;\u0026nbsp;\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig01.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/9deb764c564aefae9b9bf210.png"},{"id":17605624,"identity":"53406212-1d02-4c98-a10b-b7e69be731cd","added_by":"auto","created_at":"2022-01-24 18:37:48","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":38094,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eDuration of AP support between fetal pigs supported on a pumped and pumpless\u003c/strong\u003e\u003csup\u003e27\u003c/sup\u003e\u003cstrong\u003e AP circuit.\u003c/strong\u003e Fetal pigs maintained using a pumped AP circuit are shown as black circles (n = 13), and fetuses supported using a pumpless circuit as open circles (n = 12).\u0026nbsp;\u003c/p\u003e","description":"","filename":"Fig02.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/6984b66ef733c302b4313c91.png"},{"id":17605926,"identity":"e45613e4-f7ab-44a0-a37c-4fa584592d14","added_by":"auto","created_at":"2022-01-24 18:40:48","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":109913,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eFetal temperature, HR, and UV flow data for \u003cem\u003ein utero\u003c/em\u003e, pumped and pumpless AP fetal pigs\u003c/strong\u003e\u003csup\u003e27\u003c/sup\u003e\u003cstrong\u003e. (A) \u003c/strong\u003eTemperature; \u003cstrong\u003e(B) \u003c/strong\u003eFetal HR; and \u003cstrong\u003e(C) \u003c/strong\u003eUV flow vs. time post-cannulation. \u003cstrong\u003e(D)\u003c/strong\u003e Mean fetal HR; \u003cstrong\u003e(E) \u003c/strong\u003emean indexed UV flow; and \u003cstrong\u003e(F) \u003c/strong\u003emean absolute UV flow over the entire duration of support. \u003cstrong\u003e(G) \u003c/strong\u003eCorrelations between HR and absolute UV flow for fetal pigs supported using a pumpless and pumped AP circuit.\u003cstrong\u003e \u003c/strong\u003eFetal pigs maintained on a pumpless AP are represented as open circles and dashed lines (n = 10 for temperature; n = 9 for HR; n = 5 for UV flow), and fetal pigs maintained on a pumped AP circuit (n = 13 for temperature and UV flow; n = 12 for HR) as black circles and solid line. \u003cem\u003eIn utero\u003c/em\u003e fetal pigs are represented as open triangles (n = 6 for HR; n = 16 for indexed and absolute UV), pumpless AP fetal pigs as open circles (n = 10 for absolute UV flow) and pumped AP fetal pigs as black circles (n = 13 for unindexed UV flow\u003c/p\u003e","description":"","filename":"Fig03.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/1351ed7a3e0024f3323e30bc.png"},{"id":17605623,"identity":"24276f12-7e89-4432-a528-722f8d84dad0","added_by":"auto","created_at":"2022-01-24 18:37:48","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":106457,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eBlood gases, electrolytes, lactate, and glucose concentrations in fetal pigs studied \u003cem\u003ein utero\u003c/em\u003e versus fetal pigs supported using a pumped and pumpless AP circuit\u003c/strong\u003e\u003csup\u003e27\u003c/sup\u003e\u003cstrong\u003e. \u003c/strong\u003e\u003cem\u003eIn utero \u003c/em\u003efetal pig data represented by open triangles (n = 24), fetal pigs supported on a pumped AP circuit as black circles (n = 13), and fetuses supported on a pumpless AP circuit as open circles (n = 9).\u0026nbsp;If two \u003cem\u003eP-\u003c/em\u003evalues are presented, the first \u003cem\u003eP-\u003c/em\u003evalues represents the pumped AP circuit, and the second \u003cem\u003eP-\u003c/em\u003evalue represents the pumpless AP circuit.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"Fig04.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/ea9202deb7d73757ec723601.png"},{"id":17605928,"identity":"3c6f7057-5c8d-4abd-9111-9def7291fa05","added_by":"auto","created_at":"2022-01-24 18:40:49","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":248849,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eHemodynamic data in pumped AP fetal pigs. \u003c/strong\u003eData from n = 13 fetal pigs.\u003cstrong\u003e (A-M) \u003c/strong\u003eChanges in UV flow, HR, temperature, MAP, and CVP over time for each individual animal supported using a pumped AP circuit. \u003cstrong\u003e(N) \u003c/strong\u003eIndividual UV flow patterns for fetal pigs. \u003cstrong\u003e(O)\u003c/strong\u003e Aggregate UV flow data for pumped AP fetal pigs. Pumped AP fetal pigs are represented as black circles, and \u003cem\u003ein utero\u003c/em\u003e UV flow as the dashed grey line.\u0026nbsp;Data are presented in 5-minute \u003cstrong\u003e(A-N)\u003c/strong\u003e and 1 hour epochs \u003cstrong\u003e(O). *\u003c/strong\u003e Significantly different from the first recorded data point.\u003c/p\u003e","description":"","filename":"Fig05.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/473b60a26296ee60301975ba.png"},{"id":17605929,"identity":"c0f4d151-5762-42e9-b67e-99ae8e88ad6a","added_by":"auto","created_at":"2022-01-24 18:40:49","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":43439,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMAP in fetal pigs studied \u003cem\u003ein utero\u003c/em\u003e and maintained using a pumped AP circuit. \u003c/strong\u003eFetal pigs studied \u003cem\u003ein utero \u003c/em\u003eare represented in open triangles (n = 21) and fetal pigs maintained using a pumped AP as black circles (n = 6).\u003c/p\u003e","description":"","filename":"Fig06.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/b3523d8b21b12753632c630a.png"},{"id":17605927,"identity":"9148ffef-9830-4069-9c50-8a2e9ddb4a50","added_by":"auto","created_at":"2022-01-24 18:40:48","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":52653,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eEffects of milrinone lactate infusion on fetal pig hemodynamics while maintained using a pumped AP circuit. \u003c/strong\u003eMaintenance infusion of milrinone lactate was used in successful AP experiments (n = 5) for hemodynamic support. ‘Milrinone infusion start’ indicates the time point at which milrinone lactate IV infusion began. Data are presented as 5-minute epochs and expressed as mean ± SEM.\u0026nbsp;\u003c/p\u003e","description":"","filename":"Fig07.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/d7d4fb3cbdb10581f0291036.png"},{"id":17605628,"identity":"8ba956ea-4d2f-480c-a416-20eba778a4a4","added_by":"auto","created_at":"2022-01-24 18:37:48","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":61202,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eFetal oxygen delivery, oxygen consumption, and oxygen extraction fraction on a pumped AP circuit. (A)\u003c/strong\u003e Fetal oxygen delivery on the pumped AP circuit is represented as black circles (n = 13) and open circles for the pumpless AP circuit (n = 5)\u003csup\u003e27\u003c/sup\u003e, whereas oxygen consumption is shown as red squares with a dashed line (n = 11). \u003cstrong\u003e(B)\u003c/strong\u003e Correlation between UV flow and oxygen extraction fraction (n = 11). Data are presented as 1 hour averages over the first 24 hours of AP support.\u0026nbsp;\u003c/p\u003e","description":"","filename":"Fig08.png","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/ad833585058a9c188786090f.png"},{"id":19273888,"identity":"66ae1802-1f70-478e-acfc-3d071a219603","added_by":"auto","created_at":"2022-03-16 06:59:27","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1444886,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/ea0f64a6-1560-4e34-a3fb-b8ef48ad3c87.pdf"},{"id":17605631,"identity":"cd13e482-8127-4fc1-8e2e-fb73f54a6150","added_by":"auto","created_at":"2022-01-24 18:37:49","extension":"pdf","order_by":12,"title":"","display":"","copyAsset":false,"role":"supplement","size":1035251,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryFigure1PumpedAPPaper.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1251735/v1/674f28202903fc3ed1156d57.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eAdvantages of a Centrifugal Pump in the Development of a Swine Model of an Artificial Placenta\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eIn resource rich nations, extreme prematurity, defined as delivery prior to 28 weeks\u0026rsquo; gestation, remains the leading cause of childhood mortality and morbidity\u003csup\u003e1,2\u003c/sup\u003e. Survival rates decline with decreasing gestational age (GA), with only 6% surviving at 22 weeks GA compared to \u0026gt;90% surviving at 28 weeks GA\u003csup\u003e3\u003c/sup\u003e. In Canada alone, the economic burden of caring for extremely preterm infants approaches $600 million nationally per annum\u003csup\u003e4\u003c/sup\u003e. Unfortunately, despite advances in medical technology and neonatal intensive care, improvements in the outcomes of children born extremely preterm over the last 15 years have been limited\u003csup\u003e5\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAt the biological limit of viability (22-25 weeks GA), preterm infants commence pulmonary gas exchange during the late-canalicular and early saccular stages of lung development. Exposure to the positive pressure mechanical ventilation and high partial pressure of inspired oxygen required to achieve adequate gas exchange results in cessation of alveolarization and pulmonary microvascular injury, which is associated with high rates of chronic lung disease and pulmonary hypertension\u003csup\u003e6\u003c/sup\u003e. Furthermore, approximately 50% of children born at the threshold of viability exhibit neurological disabilities at 30 months corrected age, with half of these cases being classified as severe\u003csup\u003e7\u003c/sup\u003e. Despite major advances in the outcomes of prenatal infants resulting from the widespread administration of prenatal steroids and exogenous surfactant, recent improvements in the morbidity and mortality have been more incremental, emphasizing the need for new innovative approaches to supporting the fragile physiology and development of extremely preterm infants\u003csup\u003e1-3\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAn artificial placenta (AP) represents a novel approach that aims to maintain the innate fetal circulation while promoting normal prenatal development. Gas exchange is achieved with a low-resistance hollow-fiber membrane oxygenator connected to the fetus via the umbilical vasculature, while incubating the fetus in a fluid filled environment. The first attempt to support previable human infants was reported in the late 1950s\u003csup\u003e8\u003c/sup\u003e; with subsequent progress made using animal models. These early experiments were complicated by the development of heart failure and infections and concurrent improvements in conventional neonatal intensive care led to diminished enthusiasm for AP technology\u003csup\u003e8,9\u003c/sup\u003e\u003csup\u003e,\u003c/sup\u003e\u003csup\u003e10\u003c/sup\u003e. However, over recent years, several research teams have demonstrated the feasibility of supporting preterm goat and sheep fetuses with a variety of pumpless and pumped arteriovenous and venovenous AP systems using different approaches to establishing vascular access\u003csup\u003e8,9,11\u0026ndash;24\u003c/sup\u003e. Two research\u0026nbsp;teams have successfully demonstrated physiologic fetal sheep hemodynamics with normal organ maturation and minimal injury for periods of up to one month on pumpless arteriovenous AP systems\u003csup\u003e17\u0026ndash;19,23\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eDespite these advances, AP is yet to be proven in a realistic animal model of the extremely preterm human infant.\u0026nbsp;At a comparable stage of lung development to a previable human fetus, the preterm sheep is approximately twice the weight\u003csup\u003e25,26\u003c/sup\u003e. Fetal size determines important anatomical considerations for establishing an extracorporeal membrane oxygenation (ECMO) system such as blood vessel diameter and blood pressure. In addition, fetal sheep possess two umbilical arteries (UA) and two umbilical veins (UV). This may allow a more stable transition to the AP circuit using one pair of UA and UV while maintaining native gas exchange via the placenta using the other pair of umbilical vessels, thus minimizing the cessation of oxygenation from the mother\u003csup\u003e27,28\u003c/sup\u003e. By contrast, human and pig fetuses usually have one UV and two UAs, which may represent a more technically challenging approach to the initiation of AP support.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eWe have previously proposed\u003csup\u003e27\u003c/sup\u003e that Yucatan miniature pigs delivered at approximately 95 days gestation represent a realistic model of human infants born at the lower limit of viability in terms of their body weight and stage of lung development\u003csup\u003e25\u003c/sup\u003e. We recently demonstrated the feasibility of cannulating the umbilical vessels of preterm minipigs and transferring them to an AP system consisting of a warm, fluid environment and a pumpless ECMO circuit comprising a commercial neonatal oxygenator\u003csup\u003e27\u003c/sup\u003e. However, our experiments were characterized by subphysiologic circuit flows, tachycardia, and the development of hydrops. Echocardiography further revealed evidence of diminished right ventricular function, which we attributed to excessive ventricular afterload resulting from the large priming volume of the circuit and small umbilical cannulas\u003csup\u003e27\u003c/sup\u003e. We hypothesized that the addition of a pump might improve the hemodynamics of our animals on the circuit by reducing afterload through the generation of negative pressure downstream of the umbilical arterial cannulas, thereby improving venous return and cardiac output. Herein, we sought to investigate the hemodynamics of a pumped AP system, comparing our findings to those obtained using the pumpless circuit\u003csup\u003e27\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003e\u003cstrong\u003e\u003cem\u003eExperimental groups.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe results presented in this report are comprised from fetal pigs studied \u003cem\u003ein utero\u003c/em\u003e and while maintained using a pumped AP circuit and of our previously published findings using a pumpless AP system\u003csup\u003e27\u003c/sup\u003e. Fetal Large White Landrace pigs were studied \u003cem\u003ein utero\u003c/em\u003e at 98, 106, and 112 days GA (n = 24 ) and Yucatan miniature pigs at 107 \u0026plusmn; 3 days (n = 16), providing reference physiologic data regarding heart rate (HR), blood pressure, blood gases, electrolytes, lactate and glucose concentrations, and UV flow\u003csup\u003e27\u003c/sup\u003e.\u0026nbsp;Fetal Yucatan miniature pigs (n =13) were maintained using an umbilical arteriovenous AP circuit consisting of a centrifugal pump and oxygenator and using a pumpless AP circuit (n = 12)\u003csup\u003e27\u003c/sup\u003e. We included AP subjects that survived on the system for a minimum of 3 hours.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAnimals and approvals\u003c/em\u003e\u003c/strong\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eIn utero\u003c/em\u003e BP, HR, blood gases, electrolytes, glucose, and lactate measurements were obtained in White Landrace Cross sows (n = 6; term = 115 days) at the Preclinical Imaging and Research Laboratories, South Australian Health and Medical Research Institute (SAHMRI). All procedures were approved by the SAHMRI Animal Ethics Committee\u003csup\u003e27\u003c/sup\u003e. Sows were individually housed with environmental and social enrichment. The AP experiments were conducted in the Lab Animal Services (LAS) facility at The Hospital for Sick Children (SickKids), Peter Gilgan Center for Research and Learning in Toronto, Ontario. All maternal and fetal surgeries were approved by the SickKids Animal Care Committee and all procedures complied with the Canadian Council on Animal Care, Ontario Ministry of Agriculture, Food and Rural Affairs, Animals for Research Act guidelines, and the Care and Use of Animals for Scientific Purposes. All research and LAS staff members acted in accordance with ARRIVE guidelines proposed by Kilkenny \u003cem\u003eet al.\u003c/em\u003e\u003csup\u003e54\u003c/sup\u003e. Pregnant Yucatan miniature pigs (n = 46; term = 115 days) were acquired from Memorial University of Newfoundland and Sinclair Bioresources and transported as per the Health of Animals Act of Canada\u003csup\u003e27\u003c/sup\u003e. Yucatan pigs were housed in pairs for at least two weeks prior to surgery to increase socialization between the animals and allow for acclimation to human handling and to their new environment. Sows were provided with \u003cem\u003ead libitum\u0026nbsp;\u003c/em\u003efood and water, and environmental enrichments as per SickKids standard operating procedures.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eProtocol for in utero studies.\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eLarge White Landrace Cross gilts (n = 6; 98 \u0026plusmn; 7 days GA; term = 115 days) were anaesthetised with an intramuscular injection (I.M.) of 20 mg/kg ketamine and inhalation of isoflurane. Gilts were intubated and general anaesthesia was maintained using isoflurane with 2 L/min O\u003csub\u003e2\u003c/sub\u003e and 4 L/min medical air. Gilts were positioned on the operating table on their backs, an incision was made along the abdomen, the uterus was incised, and a fetal head was exposed. Fetal pigs (n = 24) were cannulated via the UV, and venous blood was sampled for partial pressure of oxygen (PO\u003csub\u003e2\u003c/sub\u003e), partial pressure of carbon dioxide (PCO\u003csub\u003e2\u003c/sub\u003e), oxygen saturation (SO\u003csub\u003e2\u003c/sub\u003e), pH, hemoglobin (Hb), bicarbonate (HCO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e), base excess (BE), sodium (Na\u003csup\u003e+\u003c/sup\u003e), potassium (K\u003csup\u003e+\u003c/sup\u003e), and calcium (Ca\u003csup\u003e2+\u003c/sup\u003e) as previously described\u003csup\u003e27,28\u003c/sup\u003e. \u003cem\u003e\u0026nbsp;\u003c/em\u003eIn a subset of fetuses (n = 21, 105 \u0026plusmn; 7 days GA), the carotid artery (CA) was instrumented and fetal BP, and HR were measured and continuously recorded in LabChart 8 Pro (ADInstruments Inc., Colorado Springs, USA)\u003csup\u003e27\u003c/sup\u003e. Following \u003cem\u003ein utero\u003c/em\u003e experiments gilts and their fetal pigs were humanely euthanized with an intravenous overdose of sodium pentobarbital (Virbac, New South Wales, Australia).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eSurgical protocol for pumped AP studies.\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePregnant Yucatan pigs (n = 19; 101 \u0026plusmn; 4 days GA; term = 115 days) were anaesthetized with an I.M. injection of 10 mg/kg ketamine hydrochloride, 0.20 mg/kg acepromazine, and 0.015 mg/kg atropine sulfate (CDMV Inc., Saint-Hyacinthe, Canada), with maintenance of general anaesthesia with inhalation of 2-3% isoflurane (Fresenius Kabi Canada, Toronto, Canada). To prevent aortocaval compression, anaesthetized sows were positioned on the operating table in the left lateral position. Umbilical blood flow was measured \u003cem\u003ein utero\u003c/em\u003e in 4 Yucatan sows in Toronto using cine phase contrast MRI as described in our previous publication.\u003csup\u003e27\u003c/sup\u003e A lower antero-lateral laparotomy was performed for a caesarean section. Following the delivery of all fetal pigs, sows were humanely euthanized with 106 mg/kg Euthanyl (CDMV Inc., Saint-Hyacinthe, Canada).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eFetal surgical procedures.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn a subset of fetuses (n = 11/13 successful pumped AP experiments), a small incision was made along the right side of the neck to expose the jugular vein (JV) and CA\u003csup\u003e28,55\u003c/sup\u003e. A size-matched custom-made PVC tubing was then inserted in the JV and CA for monitoring of central venous pressure (CVP) and fetal mean arterial pressure (MAP), respectively. The tubing was stabilized using silk sutures, and the neck incision sutured closed. Fetuses were delivered to minimize excessive torsion and stretching of the umbilical cord and were positioned on the maternal abdomen and subsequently weighed. Fetal pigs were then anaesthetized using an I.M. injection of 1 mg/kg rocuronium bromide (Sandoz Inc., Mississauga, Canada), and 5 mg/kg ketamine hydrochloride (CDMV Inc., Saint-Hyacinthe, Canada). Fetal normothermia was maintained by continuously bathing the fetus and the umbilical cord in warmed normal saline. The umbilical cord was treated with a topical application of 100 mg/kg papaverine hydrochloride (Sandoz Inc., Boucherville, Canada).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn all AP studies, umbilical catheters were then placed in both UAs and the UV (12-gauge and 10-gauge custom-made cannulas, respectively), and secured into place using silk sutures (Ethicon Inc., New Jersey, USA) at the insertion of the cannula within the vessel. In smaller fetuses, UA cannulas were downsized to 14 GA angiocaths (Becton Dickinson Canada Inc., Mississauga, Canada). Cannulas were then connected to the AP circuit described below, via modified \u0026frac14;\u0026rdquo; perfusion adapters (Medtronic of Canada Ltd., Brampton, Canada) to facilitate the initiation of UV flows. Re-positioning of the umbilical catheters was performed as needed to assist in the establishment of flow.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAP circuit design.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe arteriovenous AP circuit consisted of a commercially available centrifugal pump, connected to a low-resistance membrane oxygenator, and modified commercially available intravenous umbilical arterial and venous cannulas via \u0026frac14;\u0026rdquo; internal diameter x 1/16\u0026rdquo; wall thickness, 3/16\u0026rdquo; x 1/16\u0026rdquo; TYGON PVC tubing with P.h.i.s.i.o coating (LivaNova PLC., London, England). In two experiments, a commercially available cardiopulmonary bypass oxygenator was used (Table 2); however, this was subsequently changed to another commercially available ECMO oxygenator (n = 11; Table 2). Total priming volumes for these neonatal membrane oxygenators were 40 and 65 ml, respectively. Circuits were then initially primed with plasmalyte (Baxter Inc., Mississauga, Canada) and then replaced with heparinized maternal blood, which was continuously recirculated to prevent clotting within the circuit and warm the blood prior to connection to the fetus. UA blood passed through the oxygenator inlet, exiting through the outlet before returning to the heart via the UV (Figure 1). \u0026nbsp;The sweep gas supplied the oxygenator with a mixture of medical air and oxygen and was titrated to achieve normal physiologic levels of UV PO\u003csub\u003e2\u003c/sub\u003e of 35-40 mmHg and a PCO\u003csub\u003e2\u003c/sub\u003e of 50-60 mmHg\u003csup\u003e29\u003c/sup\u003e based on \u003cem\u003ein utero\u0026nbsp;\u003c/em\u003efetal pig data. \u0026nbsp;\u003cstrong\u003e\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eFluid incubation\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFollowing cannulation, fetal pigs were enclosed in a custom-made silicone infused thermoplastic polyurethane film \u0026lsquo;biobag\u0026rsquo;. Approximately 2-4 litres of lactated ringer (LR; Baxter Inc., Mississauga, Canada) crystalloid solution was warmed to 39 \u0026plusmn; 1\u0026deg;C in a large fluid reservoir and circulated via the inflow and outflow ports of the Biobag every 10 hours. The Biobag was equipped with a temperature port that facilitated monitoring of fluid temperature using a temperature probe (ADInstruments Inc., Colorado Springs, USA). The biobag was covered to prevent transmission of light to the fetus to better simulate uterine conditions and maintained at an appropriate temperature using a contact heat pad underneath the biobag and an overhead heater.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eFetal pig maintenance on the AP circuit.\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFollowing cannulation and transition to the AP, fetal pigs received a maintenance infusion of 6 \u0026micro;g/kg/h prostaglandin E1 (Pfizer Canada Inc., Kirkland, Canada) with 100 units/kg/h heparin (Fresenius Kabi Canada, Toronto, Canada) to maintain patency of the ductus arteriosus, and prevent clotting in the AP circuit\u003csup\u003e19\u003c/sup\u003e. Dextrose (418 mg/kg/h; Pfizer Canada Inc., Kirkland, Canada) was delivered via the circuit for the first 8 hours of AP support, which was then exchanged for total parenteral nutrition solution with the goal of supporting fetal energy requirements and maintaining fetal glucose concentrations of \u003cu\u003e\u0026gt;\u003c/u\u003e100 mg/dL. Calcium chloride (80 mg/kg; Omega Laboratories Ltd., Montreal, Canada) and heparin (100 units/kg) were dosed empirically to achieve a target Ca\u003csup\u003e++\u003c/sup\u003e concentration of \u003cu\u003e\u0026gt;\u003c/u\u003e1.4 mmol/L, and an activated clotting time (ACT) of 250-300 seconds, respectively. In early experiments (n = 7/13 successful experiments), 6 mg/kg/h of papaverine was administered I.V. to prevent vasospasm of the umbilical vessels and augment circuit flows. In a subset of experiments (n = 3/13), this was subsequently changed to a maintenance infusion of 30 \u0026micro;g/kg/h milrinone lactate (Aurobindo Pharma, Hyderabad, India). Reconstituted hydrocortisone (8 mg/kg/day, Pfizer Canada Inc., Kirkland, Canada) and a broad-spectrum empirical antibiotic (piperacillin/tazobactam; 300 mg/kg/day, Sandoz Inc., Boucherville, Canada) was administered I.V., every 6 hours, and 8 hours, respectively. Whole maternal blood was transfused (10 ml/kg) to replete circulating blood volume following circuit phlebotomy. In addition, rocuronium bromide (1 mg/kg) and ketamine hydrochloride (5 mg/kg) were given for excessive fetal movements or perceived fetal agitation. Albumin (25%; CSL Behring Canada Inc., Ottawa, Canada) and furosemide (0.5 mg/kg; Pfizer Canada Inc., Kirkland, Canada) were given intermittently during longer experiments to address perivascular edema and hydrops fetalis.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFetal CA, and umbilical arterial and venous blood gases including PO\u003csub\u003e2\u003c/sub\u003e, PCO\u003csub\u003e2\u003c/sub\u003e, pH, Hb, HCO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e-\u003c/sup\u003e, SO\u003csub\u003e2\u003c/sub\u003e, BE, Na\u003csup\u003e+\u003c/sup\u003e, K\u003csup\u003e+\u003c/sup\u003e, lactate, glucose and ACT were sampled every 1-3 hours and analyzed using a handheld blood analyzer (Abbott Point of Care Inc., Nepean, Canada). Fetal oxygen delivery (DO\u003csub\u003e2\u003c/sub\u003e) and consumption (VO\u003csub\u003e2\u003c/sub\u003e) were calculated based on the combination of UV and UA oxygen carrying capacity and indexed UV flow indexed to fetal weight (measured at surgery).\u003c/p\u003e\n\u003cp\u003e\u003cimg 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EyPLD1/kIkcxUEyyqHGwER8ddA7zdcDjf15amOdFgzEZ60Tx1eEnkJsHtmKktUC/vh8B+1dFVnY46EeHQ5wVa2Ln9iAbf30t+J4xdpiDDsbxXXFQsZ6gfcoTQMYUE/nntm7ZNUH8kOF20cazAj2uV2A/fVsQk9UeQ17OR2aOxzZfjyMgQ37KbrWtYqb4pk20p00ae2n88eNSXxiPLuUL9uV6KdeO2jOti3xSvJxJp9sE0/xpXinO4UsAya2kI7HygFMC6oBzVoclc7ThNX/v8QRP9KsyD0PZ7nPZGL6B6Pd3Dn/aHBUDHuaziadf6HnQivQReR4v9Xuc6N8rWsQwD1rAxmk9ZNu0jltMcXSQl7YyPm3ALq07MIa1Q74uT5LDmkw+CPp45A+fKq7MRfYE7W6DIzkT2C7b9tYFsCFlyS7lG3J4z/0h1J8wn0fr4o9g7spOxcyRrulZ2Qf0pSzZpfWgn3f5nag/4zXZSdueTRNb8djC6wW28O5cag823PoSoHj7/GleKc7hSwBw0PDLjqSakj1/wQnapkTMTfBQkDcdhmwU/zWtzaLLwrT5Vn8ZyGI0gR34m4cXc5GxIvuxDzl7OrEzbZVP0zrRjm3IzgvTFszZsh8b3FZdpDyusssvT/KR+VwGeGhjTP4lYwVzVhfTKQbIX+VfxtLRGpFTe+sCq3G+rrJzGqdcnfqO7B/l4gQyc20m37dsEIyZ5qLf25Gzii92rmQkitmlIH+lfwvmKFbS7Xl1qT3IS78kQ3ocYp/ryFi/BEw2TPNKcS7aRToEVoWZhKSfYqzDRXMysRk7Jfu1oG91CWGzuc2yiTnYqiJHu9p4mMdBRDvoEgSMY4PlwcMcfKOd8ZIH+kuFDkD8x14VE8WPftqYRz/tW/FCJxtdchiH7rRNoIPn0vhPFyOH+BMTbMVXbwP08Z7FD5nYr3bGYZ/H5lpWfirWfDroTPsc5QrPEdsmPWrz+cQox0nXFHPGH9EvGekT8/Nw4H2VE8xPm51Jj9rcJ61t2oNu2hPmTjZh/1YurmDOpfOwIedgv7fxXXl+BMZnDBSblb+5XhlH+tOGaV4pzkWXAG1qFfgJ+jiAGMfDwZFJzaZS8vJ9VVgugU0lnV50gE2ggxywh8MIO3Ugc2AyFzm06cD2g5R2FSsdZhzygE7apNt1aAxIDw99HkvGyQ/JZrzLnWAccZbcqRg46FhdELbAji25ssHHYLditrILXz1OfDJ+K8+OgmzlWqKi68+WfwJ5xPAI+E9+M8f1TDmv/eXPtO7YKDuP2AspN+33vpSZtk86sR1beXzsZD8oJ/RMa8RctfPdY4b9K9kTU2xdHu+KCf75uMk2YLzGHF0HwVyX67J4ZAt4O4/7vWoX2EWsS1lx0SXgFuQG82QvzxcuQKyXLk0vCez2gv+SyQN5dUC9dLbqBJe3l36w5SXgsegloOzx1y8B5WXCr3x+rb9E9Csw/yJVXh782mUtXzq9BJTnQi8BZRf9EnuJfwVwWgxfNlziXstfDvFDf+V4jLx8bPnl9dBLQCmllHJSegkopZRSTkovAaWUUspJ6SWglFJKOSm9BJRSSiknpZeAUkop5aT0ElBKKaWclF4CSimllJPSS0AppZRyUnoJKKWUUk5KLwGllFLKSekloJRSSjkpvQSUUkopJ6WXgFJKKeWk9BJQSimlnJReAkoppZST0ktAKaWUclJ6CSillFJOSi8BpZRSyknpJaCUUko5Kb0ElFJKKSell4BSSinlpPQSUEoppZyUXgLKo/Hz58+7Dx8+3L158+af5/fv3/c95+HHjx93Hz9+/CcW1/Lly5e7T58+3f369eu+5fFgnRJ0f/v27f7t/+Cd9nfv3v1ZXx5sFYzxvv/85z//eudxn3w8clegV+MUF9crUj/PtbhOHvJaoN/7GJuQB/RdsoZTPCbZqZ8HfXsgCx0O75MOh1inH8RjWl/Hc4XvWzEFrx3T+kL6zvvK/q1cXbGKB/NfCxd5QhEncArm27dvDwWyPD2sHWv2/fv3+5bHBX3kx9evX/98fw58/vz57v379/dvjwsFBF1++aGNWPBkAQb2E5eGhKLOvnvIZWKL1SGFztzjFEUv2PRvFcUs7oxF7gQxyfFO6mI872mjDhBHc6e4H0ExWh2wxCXjJ3TgpZ0rUpcOu4yNfHKbNHd1GDrIy3HIXK0P5Hh0eUx5X/nJOB+7Wj+BfVvrlXN1NjmKR+qgbctPMcVD6+FxfwjooDZN+I8p6sY0jrou3/ncilly+BJAIaOg8chxBZdCX/4NG2WV2I8Ji5+FArRWj3WIJNPG+Zuge9qgxGY6ZG8NutmwfgGgcNDGJ+vAevihoX6f4yATv1b9DyFtgSmHp/xSIV+xGp/6gL6pXeB/2kRcvA27V/IZt+rbgznMnfIK0DvJpQ27icPRPYGsaR97m+LIZ4KNq74EmTmOOGFDglwfO82d1kgwNscjY/IVVnIAOfiYuCyt2eQLbPU5Uzwk+yFQBzhTkTP5yl6nJki31tXH5lrTx/sqT5PDHlA4pwK1tYBnhYUlVn/rV7dgbUio1Y3yb4EdFIIjm+sxUHF4jMPyKOyJvBxjl+8VvvtGneYkrO9WYbwG1inXSoUlmYrLqhiLqT4wfvJjzzdkbenaK/p7/Vto7qq4IpMxCT7xKE5HijPj98ayx7YuFXv9YnWYMTf1e9zwZ5JP22odmcPjKC4ZO95zrINtzNvKGeydZIu9fmeKB7qnXKIG79V/dHKuUqfI68kPxcZrGXM87uinLjh+cdjj0CVAv1qmBcF43+SMwQAZJQdldPaDFjMdYS6OMJ5Hf8r1gPgYZMgW6aWNxcAHbOB9K2lgJRMkhz4lDnYxjndk890f2nIecZA9IvUiF93pL+0awyeyFUN/ZLfiwDyHeSSUxishIddJPmLz1oaZ7PCDTXL4RJf0+5q4XdjAfF8DQA+2SYcSXr76w1i3y+MJ6NZa8Kk8Z5zkXZJDxIcxGSfm+DxkYxdMPk4wDhu2mGKQj+viXXYI4qmYCuVsQjsyVkx+ITtlaV9s4euY9oH6t9YHvdIt23kyBonWdTUOe6a94X6u7J6QXVPMgb6tnEEPY7BJtvNMsUFHxp75aau/o3slaxV/dExrPNm1tYbCc32KvfpXaP1lk8ZPMZ/ioXwT9NOG7frcu9jDKpbIy3ORca4TW7O2I4/nCOvoGCigGE9ggAKjA0oJR+HEEIqn+hUk+n0zTUHAeR4trt4F8rBLBwp6JJc23pGLTo3hcys4WzJB7Up0ZGdS855JpAMWWZqDTPcn/cVO7wcfI1v5BMVd8x3a/WbKGOZiC/PlE/7R5+uEn3yX/YrFCtmhdRf4zIM8fJBu/JRtk11aD8FY5GtzIc/7mcuTSK/DPOTLJ2QiG730XZND+IiMhHafx3fFFRvwHdnMdZsc2mTfLZA8dDvKb4cYTH6v/BXTHOllrpjWbAVz9bidsiVtd/BN+1M6mZd7NlFeT+sCyMo4Itf94jsyjkLs5KfHUbZMsRXSxVjZoHlpJ/1uJ2RMeOcRyPF3MeWOSBliikvaswJdzNXjYEu2OdhCv+zS2hLX9IH+KUeYr3nsS+YxfysOyaQPaM81lr/Syfecy5wtv51DoyZDQAk1LaoOqzwIRAaIsV7YKMYUQm/DBoqkIEk4XAE9vOci8a4xwJit5DoiE7CD9sn3lQ7Fa7oZTv5ih/uLrhzjrJJUelN2Hoi5ztKnZFsVkIR5WwmomE5+YFfmGjb4BYa5W5tra108nqwvdrpsfKVNPgPyLskhbJv2C/rwhfjpOzFANnPQqX2wWsvJvoewWivaMob4NPm9t95TLIB279uK6YRiwaO5atvLjym2U5uj/F/FHhtyb+Cfj5d9U35uIT/dRt5XsQXsYUzaRHv6QFvKynVVjGErFti4ij8yJ98lT318ruK8AvuR4T6rbYV8TJvQ7f6CxiY+X/uZfcxnyl2BnVPMaM91YRw6qSHA95zLHOrLEdbRMSYlQJBUyBKKbRrvECAVVgKVwWJuLkIeBtilh6Cj022ZEpUE3VqYPZlCv0YnsHPSQdtqYY74O41xiOfUj9488LE9LyO5eZHla4icvUIJOS9BxnQRAuzy2Omg1hqwlrxnYRMaP/Wj1+OJr7ke6Ga+uCaHkLvynz7kKUeQq5jS5/Gf/JD/bk+CbsZsPbIv/RW0pY/MWeUX41drsoqF5uEL31PfxKRf/oLWa6VT/Ss5oDXSo7Gau7KTcR4DjZ+elX0CG1wWyC6tPXnD+wr6lVtO2gm0pU1aH+Exk29THqLTxzpb64x+2bsXH1DeOLRhl/YRdqzsBPVnPBjvexEyHoI22YE8zeWTOreqdQ7+pj7QhcJRHghqif+4AeQdiSGsM8iYlKjYTobTh+FZYB0CwxiK+5QwKVuO52GwWlxgYdwGJe50qMMRmcB8Foexuo2JrUMIP3XxSZjj/hIf2pwck+hgSdDr6ycb3U/57v6wIT2BU84K7FjZqTXIuMEUf+Llyaw8WIG9U+Gb1j5lA+++RpfmEGBjbtwJZGCr/GWexy1jAVOMHoLkZb5iV64h6z8Vli2bkDvtb8G8ldwJxqWt6QPyJp9A+ZN9rPORmDI34yLSB+yY9uPKBocx01zmqZ1Pf3cUk+xD52Q/tuY6MVd7ie8Zn5Vu4rBaT3Ss/EaWZKYtE9gzjUO32tGFzJW8Vd/U5vFwkJGxoU76D449sHlaF50DXm+wzesL9SrrDXYeqdVw6BKAUoRq8XCOwjgFiuKOQwo+Y3EuF15JiowpKdCnYkwgZIMgKMzXIYUMxnhS8u4F3QM6BeiITMbgH34SeOTgi8bIL8ZlkmK/ZCfpLzGjzW1Fn8bINo8deok3bb4JkaOiosT0gxo7cz2RgTw/rDUHOcyZYDzzVv3M9UPVkU78l9882IVc2rHf7eLd1xL7tCEYr3ijl3nIlZ/0ERv0qh3b3GfaLskhkK495J/Adq0JSI/j+XUrkJfrhd+Kk8Av4jVBvqXPWs9VLgA6GJO6Vkx6aPN8B+zMccSWNvxIjuqXjPRJe0zQv4oVICNtdiY9anMUPx/Hd9omn1Z+YmvGhbEaP83DfvdZSH/2IWPLZ2Be+rNi0qM2n6/9mPbQNq0R4yf9Hg8hfQ+FmjXFmD3ktRl9vKuOg2qi1o8axRjmHuGQ9V4gUYbB00YioCyyChTF041LtvpoRx8ycXJaAI1BDuNykWnTIQAKIPZ7oXe2ZKKfNs3VgeB2EXjG0E4sNFYFcU8vc/FXi+/xZK5k44MnAjBWdnuf1sF9QZ9k8Zmxkz3OJCdRTFYQKz9UEw5F5it2SnA/LDUG+5DnByLjtX7eN609ffKJh++5PsTm0hzSWm9tQuYix23XmtCGH3xPiP3U/hCIk+cwrIobbfRNIEOx1LMVA1CsVjIT5Z7rIFcmGOvjeCa8fSu3BeuUcmkT2Z8x8L6pH2SH9rSeCa2VP1M8kaV291NrkJBnjKd/iovydUIy/ckcm0DP0fzGNh7muJ7Jd/AxPCufZCfffW0UDwcZR/xawZogVzZRV7zWgWoB/Xx6PRLYobqHjFUMJuas+gt4sB2CnptaCeUHW3mecJBuHfJngRyeioyYLnGgCw6bfbpkMG9L7rWg0wsesD9TF/s29+dLZTqoMgavBT9oeFhH8ZB1ZozLeskQA4+R+7+KB+Nees48ySWAm8zq9sRCkLD6hUSAKXw85XnDocWm6GXtv38x8F/6D0W/CG4pU+hAzF8QFL7cq9qj5eVzi/VlbF4iXhNTPFb75SXyVy8B/MqhMG4lDAXO/0TLeJL0MQpfuR2s6d7ang1+PXB5vUXucsGiEK3+CeJWTMWfNc1fexS/1UW+vAxYv1xX3q9ZV+a8hgMxWcWDs+m18Ho8KeUZQmGc/q/BJegvZ6/1T9WllKejl4BSSinlpPQSUEoppZyUXgJKKaWUk9JLQCmllHJSegkopZRSTkovAaWUUspJ6SWglFJKOSm9BJRSSiknpZeAUkop5aT0ElBKKaWclF4CSimllFNyd/c/Sc9bmPQS6voAAAAASUVORK5CYII=\"\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003ePhysiologic monitoring.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eUV flow was continuously measured using a HXL 3/16\u0026rdquo; tubing flow probe (Transonic) and CVP, fetal BP, pre-pump circuit pressure, pre-oxygenator circuit pressure, and post-oxygenator circuit pressure were measured using Deltran fluid filled blood pressure transducers (ADInstruments Inc., Colorado Springs, USA). Data were sampled at 1000 Hz, digitized, and continuously recorded using LabChart Pro 8 (ADInstruments Inc., Colorado Springs, USA). At the end of each study, the data was extracted in consecutive 30-second intervals and analyzed in Excel (Microsoft Corporation, Washington, USA).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eQuantification of UV blood flow in utero\u003c/em\u003e \u003cem\u003eusing MRI: 3-D Volumetry and cine phase-contrast.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eIn utero\u0026nbsp;\u003c/em\u003efetal weight and UV flow were measured in sixteen fetuses from 4 pregnant Yucatan pigs (n = 4; gestational age 107 \u0026plusmn; 3 days GA) as previously described using a 3 Tesla magnetic resonance imaging (MRI) system\u003csup\u003e27\u003c/sup\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eStatistical analysis.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eComparisons of anthropometric data between pumpless and pumped AP circuits were analyzed using a two-way ANOVA with Bonferroni correction for multiple comparisons. Comparison of the duration of AP support between pumpless and pumped circuits was analyzed using a Mann-Whitney \u003cem\u003eU\u0026nbsp;\u003c/em\u003etest. Changes in temperature, HR, and UV flow between pumpless and pumped AP circuits over the first 3 hours of support were analyzed using a mixed-effect model, with a Bonferroni correction for multiple comparisons. Differences in mean indexed and absolute UV flow and HR between \u003cem\u003ein utero\u003c/em\u003e, pumpless and pumped AP circuits were compared using a repeated measures one-way ANOVA with a Bonferroni correction for multiple comparisons. Analysis of fetal HR and indexed UV flow were performed using a linear regression. Comparisons of mean blood gases, electrolytes, lactate, and glucose concentrations between animals studied \u003cem\u003ein utero\u003c/em\u003e, or on pumpless and pumped AP circuits were analyzed using a repeated measure one-way ANOVA, and a Kruskal-Wallis test (when appropriate) with Bonferroni correction for multiple comparisons. UV flow, HR, fetal BP, CVP, and temperature are presented in 5-minute averages. Time post-cannulation versus UV flow is presented in 1 hour averages and analyzed using a one-way ANOVA with Bonferroni correction for multiple comparisons. Differences in MAP between fetal pigs studied \u003cem\u003ein utero\u003c/em\u003e and those maintained using a pumped AP were analyzed using a mixed-effect model with Bonferroni correction for multiple comparisons. Indexed UV flow versus oxygen extraction was analysed using a linear regression. *\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.05 was considered statistically significant. All statistical analyses were performed using Prism 9 (GraphPad, San Diego, USA). Data are presented as mean \u0026plusmn; standard deviation (SD), unless otherwise indicated. \u0026nbsp;\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003e\u003cem\u003eThe addition of a centrifugal pump improves successful transition to AP and support times.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNineteen sows and 89 fetal pigs were used for pumped AP experiments (Table 1). A total of 32 fetal pigs were cannulated and transitioned to the AP.\u0026nbsp;Types of perioperative complications preventing us from maintaining AP support were similar to those experienced in the pumpless AP circuit group and included the inability to establish circuit flows upon connection to the circuit (n = 11), accidental decannulation (n = 4), persistent vasospasm of the umbilical cord (n = 3), and excessive spiralling of the umbilical cord, which prevented cannulation or adequate cannula position (n = 1; Table 1). We successfully maintained\u0026nbsp;13 fetal pigs at an initial GA of 102 \u0026plusmn; 4 days (range = 93-107 days) with a body weight of 616 \u0026plusmn; 139 g (range = 390-820g) on a pumped AP circuit for 46.4 \u0026plusmn; 46.8 hours (range = 3.4-177.8 hours). This represented a significant improvement in the duration of support over the 12 fetal pigs (98 \u0026plusmn; 4 days GA; 743 \u0026plusmn; 350 g; 11 \u0026plusmn; 13 hours) maintained using a pumpless AP circuit (\u003cem\u003eP =\u003c/em\u003e 0.009; Table 1 and Figure 2). There was also a marked improvement in the rate of successful AP runs per litter using a pumped AP circuit compared to the pumpless AP circuit (62% pumped AP vs. 28% pumpless AP; Table 1). Successfully supported fetuses were similar in weight and GA in both pumped and pumpless groups (616 \u0026plusmn; 139g vs. 743 \u0026plusmn; 350g, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.41; 102 \u0026plusmn; 4 days vs. 98 \u0026plusmn; 4 days, \u003cem\u003eP\u003c/em\u003e = 0.24). On the pumped and pumpless AP circuits, fetal body weights were similar between successfully and unsuccessfully supported animals (616 \u0026plusmn; 139g vs. 523 \u0026plusmn; 145g, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.47; 743 \u0026plusmn; 350g vs. 644 \u0026plusmn; 184g, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.53). However, unsuccessfully supported fetuses or fetuses that we did not attempt to cannulate were significantly smaller in the pumped group compared to the pumpless group (523 \u0026plusmn; 145g vs. 644 \u0026plusmn; 184g, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.04; 557 \u0026plusmn; 124g vs. 892 \u0026plusmn; 362g, \u003cem\u003eP\u003c/em\u003e \u0026lt; 0.0001). Fetal weight at the time of termination of the AP study was significantly higher compared to pre-cannulation weight. Fetal weight gain correlated strongly with length of AP support (r = 0.89), with weight at the end of the experiment exceeding the expected growth trajectory of these fetuses. Individual data on supported fetuses, support times, and reasons for termination are shown in Table 2.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1. Summary of fetal pigs from pumped and pumpless AP circuits\u003c/strong\u003e\u003csup\u003e27\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"8.365384615384615%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" valign=\"top\" width=\"27.98076923076923%\"\u003e\n \u003cp\u003ePumped AP circuit\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" valign=\"top\" width=\"27.5%\"\u003e\n \u003cp\u003ePumpless AP circuit\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" width=\"36.15384615384615%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"8.365384615384615%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" valign=\"top\" width=\"27.98076923076923%\"\u003e\n \u003cp\u003eIntervention\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" valign=\"top\" width=\"27.5%\"\u003e\n \u003cp\u003eIntervention\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" width=\"36.15384615384615%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"8.365384615384615%\"\u003e\n \u003cp\u003eParameter\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"9.711538461538462%\"\u003e\n \u003cp\u003eMaintained on AP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.423076923076923%\"\u003e\n \u003cp\u003eCannulated but not maintained on AP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"8.846153846153847%\"\u003e\n \u003cp\u003eDid not attempt to cannulate\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"9.711538461538462%\"\u003e\n \u003cp\u003eMaintained on AP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.038461538461538%\"\u003e\n \u003cp\u003eCannulated but not maintained on AP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"8.75%\"\u003e\n \u003cp\u003eDid not attempt to cannulate\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" width=\"36.15384615384615%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" valign=\"top\" width=\"8.365384615384615%\"\u003e\n \u003cp\u003eN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"3.3653846153846154%\"\u003e\n \u003cp\u003e13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003eAP success rate per litter (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" valign=\"top\" width=\"9.423076923076923%\"\u003e\n \u003cp\u003e19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" valign=\"top\" width=\"8.846153846153847%\"\u003e\n \u003cp\u003e57\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"3.076923076923077%\"\u003e\n \u003cp\u003e12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.634615384615385%\"\u003e\n \u003cp\u003eAP\u0026nbsp;\u003c/p\u003e\n \u003cp\u003esuccess rate per litter (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" valign=\"top\" width=\"9.038461538461538%\"\u003e\n \u003cp\u003e56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" valign=\"top\" width=\"8.75%\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"top\" width=\"36.15384615384615%\"\u003e\n \u003cp\u003e\u003cem\u003eP-values\u003c/em\u003e: Pumpless vs. Pumped AP circuit (ANOVA results)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"6.055363321799308%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.418685121107266%\"\u003e\n \u003cp\u003e62\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.536332179930795%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.937716262975778%\"\u003e\n \u003cp\u003e28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"19.377162629757784%\"\u003e\n \u003cp\u003eCircuit type\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"22.837370242214533%\"\u003e\n \u003cp\u003eIntervention\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"22.837370242214533%\"\u003e\n \u003cp\u003eInteraction\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"8.365384615384615%\"\u003e\n \u003cp\u003eBody weight (g)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"9.711538461538462%\"\u003e\n \u003cp\u003e616 \u0026plusmn; 139 (390-890)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.423076923076923%\"\u003e\n \u003cp\u003e523 \u0026plusmn; 145 (244-710) \u003cstrong\u003e*\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"8.846153846153847%\"\u003e\n \u003cp\u003e557 \u0026plusmn; 124 (372-941; n = 24) *\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"9.711538461538462%\"\u003e\n \u003cp\u003e743 \u0026plusmn; 350 (500-1500; n = 7)\u003cem\u003e\u0026nbsp;\u003cstrong\u003eab\u003c/strong\u003e\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.038461538461538%\"\u003e\n \u003cp\u003e644 \u0026plusmn; 184 (343-1390; n = 52) \u003cstrong\u003e\u003cem\u003ea\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"8.75%\"\u003e\n \u003cp\u003e892 \u0026plusmn; 362 (531-1470) \u003cstrong\u003e\u003cem\u003eb\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76923076923077%\"\u003e\n \u003cp\u003e\u0026lt;0.0001\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"12.692307692307692%\"\u003e\n \u003cp\u003e0.002\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"12.692307692307692%\"\u003e\n \u003cp\u003e0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"8.365384615384615%\"\u003e\n \u003cp\u003eGA (days)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"9.711538461538462%\"\u003e\n \u003cp\u003e102 \u0026plusmn; 4 (93-107)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.423076923076923%\"\u003e\n \u003cp\u003e103 \u0026plusmn; 4 (93-108)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"8.846153846153847%\"\u003e\n \u003cp\u003e102 \u0026plusmn; 5 (93-108)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"9.711538461538462%\"\u003e\n \u003cp\u003e98 \u0026plusmn; 4 (93-107)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"9.038461538461538%\"\u003e\n \u003cp\u003e101 \u0026plusmn; 7 (93-112)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"8.75%\"\u003e\n \u003cp\u003e101 \u0026plusmn; 7 (97-110)\u003csup\u003e\u0026nbsp;\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.76923076923077%\"\u003e\n \u003cp\u003e0.04\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"12.692307692307692%\"\u003e\n \u003cp\u003e0.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"12.692307692307692%\"\u003e\n \u003cp\u003e0.60\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eTwo-way ANOVA. Multiple comparisons for fetal body weight and GA within pumped and pumpless AP circuits and between the intervention groups: \u003cstrong\u003e\u003cem\u003eab\u003c/em\u003e\u003c/strong\u003e \u003cem\u003eP\u003c/em\u003e \u0026lt; 0.05. *\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.05 body weight is significantly different between pumped and pumpless AP circuits within intervention groups (i.e., body weight maintained on a pumped AP circuit vs. body weight maintained on a pumpless AP circuit). Interpretation of numbers for multiple comparisons: \u003cem\u003ea\u003c/em\u003e represents a statistically significant difference compared with \u003cem\u003eb\u003c/em\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2. Summary data of 13 fetal pigs successfully cannulated and maintained using a pumped AP circuit.\u0026nbsp;\u003c/strong\u003eData are presented by increasing GA and are expressed as mean \u0026plusmn; SD.\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGA before\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(days)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e\u003cstrong\u003eWeight before\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(g)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e\u003cstrong\u003eDuration on AP\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(hours)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e\u003cstrong\u003eGA after\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(days)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e\u003cstrong\u003eWeight after\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(g)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003e\u003cstrong\u003eSex\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(M, F)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003e\u003cstrong\u003eReason for termination\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e\u003cstrong\u003eOxygenator\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e93\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e390\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e36.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e95\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e450\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eFetal movement occluded UV flow\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e98\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e450\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e3.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e98\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e450\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eHeart failure\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e505\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e14.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e101\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eNA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eMechanical failure (pump)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e520\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e15.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e101\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e620\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eHeart failure\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eBP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e590\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e49.4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e102\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e750\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eHeart failure\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e101\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e762\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e70.0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e104\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e1190\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eMechanical failure (oxygenator)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eBP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e102\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e480\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e177.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e109\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e1176\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eHydrops, cardiac dysfunction\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e610\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e68.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e108\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eNA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eNA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eMechanical failure (oxygenator)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e676\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e17.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eNA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eFetal movement occluded UV flow\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e690\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e53.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e108\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eNA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eFetal movement occluded UV flow\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e730\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e4.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e730\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eThrombosis development in circuit\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e105\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e780\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e21.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e106\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e879\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eEquipment failure (sweep-gas supply)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e107\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e820\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e72.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e110\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e876\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003eF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003eFetal movement occluded UV flow\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003eO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" width=\"7.3076923076923075%\"\u003e\n \u003cp\u003e\u003cstrong\u003e102 \u0026plusmn; 4\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"10.865384615384615%\"\u003e\n \u003cp\u003e\u003cstrong\u003e616 \u0026plusmn; 139\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"11.826923076923077%\"\u003e\n \u003cp\u003e\u003cstrong\u003e46.4 \u0026plusmn; 46.8\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"1.8269230769230769%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"6.346153846153846%\"\u003e\n \u003cp\u003e\u003cstrong\u003e104 \u0026plusmn; 4\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e\u003cstrong\u003e792 \u0026plusmn; 272 (n = 9)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"5.480769230769231%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"29.03846153846154%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" width=\"13.653846153846153%\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eAbbreviations: NA, not available; M, male; F, female; GA, gestational age; O, commercially available ECMO oxygenator; BP, commercially available bypass oxygenator.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003ePumped AP circuits achieve higher umbilical venous flow.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAnimals on a pumped AP circuit maintained a physiologic core body temperature throughout the experiment. The temperature was lower in fetal pigs supported on a pumped system than pigs supported on a pumpless system (\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.0001, Figure 3A). UV flow rates of fetuses supported by the pumped AP circuit were comparable to \u003cem\u003ein utero\u003c/em\u003e controls, while UV flow on the pumpless AP system was subphysiologic (\u003cem\u003ein utero\u003c/em\u003e; 108 \u0026plusmn; 24 ml/min vs. pumped; 87 \u0026plusmn; 28 ml/min vs. pumpless; 70 \u0026plusmn; 18 ml/min, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.001; \u003cem\u003ein utero\u003c/em\u003e;\u003cem\u003e\u0026nbsp;\u003c/em\u003e173 \u0026plusmn; 45 ml/min/kg\u003cem\u003e\u0026nbsp;\u003c/em\u003evs. pumped; 143 \u0026plusmn; 40 ml/min/kg vs. pumpless; 97 \u0026plusmn; 39 ml/min/kg, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.005; Figure 3C, E and F).\u0026nbsp;Absolute UV flow measured within the first 3 hours post-cannulation was positively correlated to HR in pumped AP circuits (r\u003csup\u003e2\u003c/sup\u003e = 0.44; \u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.0001; Figure 3G), but negatively correlated in fetuses maintained on a pumpless circuit (r\u003csup\u003e2\u003c/sup\u003e = 0.45; \u003cem\u003eP\u003c/em\u003e \u0026lt; 0.0001; Figure 3G), with differences between the slopes being extremely significant (\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.0001). \u0026nbsp;Figures 5A-M demonstrate the changes in UV flow, HR, temperature, MAP, and CVP over time for the 13 fetal pigs successfully maintained using a pumped AP circuit. Upon initiation of AP support, we observed supraphysiologic UV flow, despite a low pump rate setting (Figure 5A-N). In longer trial runs, this period of supraphysiologic UV flow persisted for approximately 6 hours, then decreasing to subphysiologic flow, plateauing at ~12 hours of support (Figure 5O). In several experiments (n = 8/13) excessive spasmodic fetal movements resulted in a complete cessation of UV flow and consequently led to the termination of the experiment (Table 2). In four studies, development of a thrombosis in one of the UAs in the first 12 hours of support was associated with a significant reduction in UV flow.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eFetal pigs were tachycardic for a large proportion of the AP studies (Figure 3B and D; and Figures 5A-M). Both pumped (205 \u0026plusmn; 28 bpm; \u003cem\u003eP \u0026lt;\u0026nbsp;\u003c/em\u003e0.0001; Figure 3D) and pumpless (206 \u0026plusmn; 38 bpm; \u003cem\u003eP \u0026lt;\u0026nbsp;\u003c/em\u003e0.0001;\u003cem\u003e\u0026nbsp;\u003c/em\u003eFigure 3D) fetal pig groups were similarly tachycardic throughout the experiment compared to \u003cem\u003ein utero\u0026nbsp;\u003c/em\u003econtrols (130 \u0026plusmn; 10 bpm; Figure 3D). Compared to \u003cem\u003ein utero\u0026nbsp;\u003c/em\u003econtrol animals, fetal pigs were hypertensive on the pumped AP system (\u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.013; Figure 6). Approximately 20 minutes after initiation of a milrinone infusion, there was a marked increase in UV flow, followed by a steady decline over a 3 hour period (Figure 7A). MAP and HR decreased following the start of the milrinone infusion, which persisted for ~1 hour before returning to a steady state (Figure 7B and C.).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eUpon initiation of AP support, fetal oxygen delivery was physiologic\u003csup\u003e29,30\u003c/sup\u003e. We observed a decline in oxygen delivery and increase in oxygen consumption over time (Figure 8A). UV flow and oxygen extraction fraction correlated negatively (r\u003csup\u003e2\u003c/sup\u003e = 0.45; \u003cem\u003eP\u003c/em\u003e = 0.0003; Figure 8B).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eFetal blood gas analysis indicates supraphysiologic oxygen tension and fetal anemia\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWhile there was no significant difference in SO\u003csub\u003e2\u003c/sub\u003e between fetal pigs studied \u003cem\u003ein utero\u003c/em\u003e and those maintained using a pumped or pumpless AP circuit (Figure 4A), both AP circuits resulted in a significantly higher PO\u003csub\u003e2\u003c/sub\u003e compared to \u003cem\u003ein utero\u003c/em\u003e controls (\u003cem\u003ein utero\u003c/em\u003e; 53 \u0026plusmn; 16 mmHg vs. pumped; 141\u0026plusmn; 129 mmHg, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.0001, vs. pumpless; 280 \u0026plusmn; 176 mmHg, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.003, Figure 4B). Conversely, PCO\u003csub\u003e2\u003c/sub\u003e was significantly lower in the pumped AP group compared to \u003cem\u003ein utero\u003c/em\u003e controls (\u003cem\u003ein utero\u003c/em\u003e; 67 \u0026plusmn; 11 mmHg vs. pumped; 49 \u0026plusmn; 11 mmHg, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.0009, Figure 4E). Animals on the pumped AP were significantly more alkalotic than both pumpless AP fetuses and \u003cem\u003ein utero\u003c/em\u003e controls (\u003cem\u003ein utero\u003c/em\u003e; 7.29 \u0026plusmn; 0.08 vs. pumped; 7.36 \u0026plusmn; 0.07, \u003cem\u003eP =\u0026nbsp;\u003c/em\u003e0.03, vs. pumpless; 7.30 \u0026plusmn; 0.06, \u003cem\u003eP\u003c/em\u003e = 0.03, Figure 4F). BE and lactate were similar between groups (Figure 4G, H). We identified significantly lower hematocrit (\u003cem\u003ein\u0026nbsp;\u003c/em\u003eutero; 28 \u0026plusmn; 3%; vs. pumped; 24 \u0026plusmn; 3%, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.02, Figure 4C)\u0026nbsp;and hemoglobin in the pumped AP group (\u003cem\u003ein\u0026nbsp;\u003c/em\u003eutero; 96 \u0026plusmn; 9 g/L vs. pumped: 82 \u0026plusmn; 11 g/L, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e= 0.01, Figure 4D), as well as higher Na\u003csup\u003e+\u003c/sup\u003e in both AP supported groups compared to \u003cem\u003eintrauterine\u003c/em\u003e fetuses (\u003cem\u003ein utero\u003c/em\u003e; 126 \u0026plusmn; 2 mmol/L vs. pumped; 133 \u0026plusmn; 5 mmol/L, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e \u0026lt; 0.0001; vs. pumpless; 133 vs. 3 mmol/L, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.0001,\u0026nbsp;Figure 4L). Ca\u003csup\u003e2+\u0026nbsp;\u003c/sup\u003econcentrations were subphysiologic in both pumped\u003cem\u003e\u0026nbsp;\u003c/em\u003eand pumpless groups (\u003cem\u003ein utero\u003c/em\u003e; 1.56 \u0026plusmn; 0.087 vs. pumped; 1.44 \u0026plusmn; 0.092 mmol/L, \u003cem\u003eP =\u0026nbsp;\u003c/em\u003e0. 003, vs. pumpless; 1.29 \u0026plusmn; 0.21 mmol/L, \u003cem\u003eP =\u0026nbsp;\u003c/em\u003e0. 001, Figure 4K). Glucose, administered continuously either directly or as part of total parenteral nutrition, was significantly higher in both AP groups, compared to \u003cem\u003ein utero\u003c/em\u003e controls (\u003cem\u003ein utero\u003c/em\u003e; 2.1 \u0026plusmn; 0.43 mmol/L vs. pumped; 8.5 \u0026plusmn; 4.8 mmol/L, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.0001, vs. pumpless; 9.8 \u0026plusmn; 5.8 mmol/L, \u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u0026lt; 0.0001, Figure 4I). There was no difference in K\u003csup\u003e+\u0026nbsp;\u003c/sup\u003econcentrations between\u0026nbsp;\u003cem\u003ein utero\u003c/em\u003e fetal pigs and those maintained using a pumped and pumpless AP circuit (Figure 4J). \u0026nbsp;\u0026nbsp;\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eTo our knowledge, this is the first study to examine the hemodynamics and blood gas status of preterm pigs supported using an AP system incorporating a small centrifugal pump. We previously showed the feasibility of supporting our animal model using a pumpless AP circuit connected to the fetal circulation via the umbilical arteries and vein\u003csup\u003e27\u003c/sup\u003e; however, our experiments were characterized by significant hemodynamic decompensation within hours. We attributed this to afterload imbalance, elicited by supraphysiologic circuit resistance and further exacerbated by impaired umbilical venous return, which we surmised resulted in higher sympathetic tone, further exacerbating the increase in afterload\u003csup\u003e27\u003c/sup\u003e. Although other research groups have highlighted potential limitations of pumped ECMO circuits, including an increased risk of pump-induced hemolysis\u003csup\u003e17\u003c/sup\u003e, afterload imbalance, myocardial strain, and impaired autoregulation of UV blood flow\u003csup\u003e31\u003c/sup\u003e, we hypothesized that the addition of a small pump might reduce right ventricular afterload and sustain adequate UV flow, thereby improving survival on the AP. The present study examined differences in hemodynamics, blood gases, electrolytes, biochemistry, and survival of fetal pigs supported on a pumped versus pumpless AP system\u003csup\u003e27\u003c/sup\u003e. The increase in AP support to 46.4 \u0026plusmn; 46.8 hours represented a significant increase in survival over fetal pigs supported without a pump possibly due to more reliable and physiologic UV flow compared to a pumpless system. Despite the pump support, following a period of supraphysiologic UV flow, the animals continued to experience a slower but consistent decline in UV flow and the development of signs of circulatory deterioration. \u0026nbsp;We have yet to establish an AP system capable of meeting the long-term physiologic needs of animals that are similar in size to the extremely preterm human infants that are the proposed target of this approach.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eIn a recent report, Partridge \u003cem\u003eet al.\u003c/em\u003e\u003csup\u003e17\u003c/sup\u003e demonstrated AP support of fetal sheep weighing 1-2 kg using a pumpless system for up to 28 days without the use of vasopressors. Using a similar AP circuit, Usada \u003cem\u003eet al.\u003c/em\u003e\u003csup\u003e18\u003c/sup\u003e reported survival of fetal sheep for up to one week. To investigate the potential utility of AP systems for extremely preterm infants born at 22-25 weeks GA, two research groups cannulated sheep born at 85-95 days GA weighing ~0.5-0.8 kg. Although cannulation was technically feasible, one study reported the development of hydrops fetalis following 5-8 days of support\u003csup\u003e32\u003c/sup\u003e, whereas the other required the use of aggressive pharmacological interventions in order to achieve hemodynamic stability\u003csup\u003e19\u003c/sup\u003e. Although fetal sheep delivered at 85-95 days GA share similar body weight to human fetuses delivered at the biological limit of viability, they are developmentally immature and their pulmonary development is analogous to a human fetus delivered at 18 weeks GA\u003csup\u003e10,26,33,34\u003c/sup\u003e. By contrast, fetal minipigs delivered at 93-107 days GA weighing 0.39-0.82 kg are in their canalicular and saccular stages of lung development and comparable to preterm human fetuses delivered at 22-28 weeks GA. Thus, fetal pigs delivered at 95 days GA would therefore be equivalent in body weight and lung maturity to human fetuses born at 22-25 weeks GA, and may represent a more appropriate model for the development of an AP system\u003csup\u003e27,33,34\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eUpon initiation of AP support, we observed supraphysiologic UV flow in all pumped AP experiments. Umbilical vein blood flow was higher in fetuses supported with a pumped circuit than in fetuses supported using a pumpless system. Our experiments revealed a positive correlation between fetal HR and UV flow on the pumped AP, with persistent tachycardia seen with both systems. Contrary to our speculations regarding supraphysiologic resistance in the pumpless circuit\u003csup\u003e27\u003c/sup\u003e, we propose that the negative pressure generated by the centrifugal pump may have diminished right ventricular afterload and minimized resistance to flow across the UA cannulas, resulting in higher than normal flow rates. The association we observed between fetal HR and UV flow in animals supported on the pumped circuit may be attributable to the Frank-Starling mechanism and Anrep effect\u003csup\u003e35\u003c/sup\u003e, whereby increased ventricular preload enhances myocardial stretch and tension, resulting in increased contractility. Although this relationship holds true for slight increases in end-diastolic filling pressures, the fetal heart has little preload reserve\u003csup\u003e36\u0026ndash;39\u003c/sup\u003e. Thus, further augmentation of cardiac output and contractility are also driven by the Bowditch effect\u003csup\u003e40\u003c/sup\u003e through increases in fetal HR\u003csup\u003e41\u003c/sup\u003e. In contrast to the physiology seen in a pumpless circuit, we observed a shift of placental-to-systemic blood flow in favor of the AP, effectively volume loading the fetal heart. Therefore, we speculate that these supraphysiologic circuit flows lead to increased preload, tachycardia, sympathetic nervous system activation, and likely higher cardiac output within the first hours of AP support.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAfter ~24 hours of AP support, we observed marked reductions in UV flow compared to the high flows seen at the start of AP support and in many experiments, UV flows then remained subphysiologic. The drop in UV blood flow and persistent tachycardia could be explained by increased sympathetic nervous activation and continuous peripheral vasoconstriction. Although the introduction of milrinone improved UV flow and reduced BP, its inotropic and systemic vasodilatory effects appeared to be short-lived, indicating the presence of overwhelming perturbation of fetal cardiac loading conditions. However, our findings suggest that phosphodiesterase inhibitors may have a positive impact on the circulation of fetal animals supported on the AP\u003csup\u003e19,42\u003c/sup\u003e. Despite similar SO\u003csub\u003e2\u003c/sub\u003e, blood gases showed higher PO\u003csub\u003e2\u003c/sub\u003e, lower PCO\u003csub\u003e2\u003c/sub\u003e, as well as a more alkalotic pH in fetuses on the pumped AP compared to \u003cem\u003ein utero\u003c/em\u003e controls. We were able to manage O\u003csub\u003e2\u003c/sub\u003e uptake and CO\u003csub\u003e2\u003c/sub\u003e elimination better on the pumped circuit than pumpless. However, the gas exchange capacity of the oxygenator used in both groups easily outperforms the native placenta, resulting in supraphysiologic UV PO\u003csub\u003e2\u003c/sub\u003e. High oxygen tension is a known contributor to umbilical cord and ductal constriction and may have negatively affected our experiments\u003csup\u003e43,44\u003c/sup\u003e. Gradual umbilical vessel spasm or constriction at the umbilicus, where the cord passes through the abdominal wall, could explain the gradual decline in UV flow we observed. Mean blood pressure in the UV is 6-8 mmHg in near-term sheep and human fetuses\u003csup\u003e29,30\u003c/sup\u003e. In our pumped AP studies, we observed pre- and post-oxygenator pressures of 25 mmHg and 18 mmHg, respectively (Supplementary Figure 1.). Although we did not measure blood pressure in the UV, it is likely that the pumped circuit exposed the umbilical vein to elevated blood pressure, even if there was some degree of pressure difference across the venous cannula. Berman \u003cem\u003eet al.\u003c/em\u003e\u003csup\u003e45\u003c/sup\u003e advanced balloon catheters into the common UV of fetal sheep and demonstrated marked reductions in placental blood flow with increasing UV pressures. This was attributed to a decreased pressure gradient across the umbilical circulation, resulting in diminished umbilical flows. We hypothesize that a similar phenomenon may be occurring in the early phase of our AP experiments, with UV constriction induced by the high pressures and shear stress resulting from the elevated post-pump pressures.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eWe observed a steady widening of the veno-arterial saturation gradient across the umbilical circulation during our AP runs that was negatively correlated with UV flow, indicating higher oxygen extraction at the tissues in the setting of a net reduction in UV flow and oxygen delivery. One factor that might additionally contribute to the progressive reduction in oxygen delivery was the low hemoglobin concentration we observed in animals supported on the pumped system. The large priming volume of the circuit and oxygenator contributes to fetal anemia because the circuit is primed with maternal blood. During pregnancy, maternal blood volume increases by ~40% and there is a progressive decline in hemoglobin concentrations\u003csup\u003e46\u0026ndash;48\u003c/sup\u003e. Conversely, fetal hemoglobin increases with GA to sustain fetal tissue oxygenation in the setting of declining PO\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e29,49,50\u003c/sup\u003e. Thus, the mixing of maternal and fetal blood in the AP circuit contributes to hemodilution of fetal blood, while ongoing blood sampling further depletes fetal red blood cells. To circumvent this problem in future experiments, it may be possible to hemoconcentrate the circuit prime. Furthermore, pump-induced hemolysis may have also contributed to fetal anemia in these experiments. Other groups have recommended the administration of daily doses of erythropoietin to minimize the need for successive maternal donor blood transfusions\u003csup\u003e17\u003c/sup\u003e. Miniaturization of the oxygenator and pump head could also help to reduce fetal anemia by reducing circuit volume.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eStudy limitations.\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThere are several limitations of the present study. Firstly, we have not yet characterized the etiology of the variation in UV flows on the pumped AP circuit. We speculate that a combination of UV constriction and supraphysiologic UV pressures, as well as supraphysiologic oxygen tension may be contributing to progressively diminished UV flows. However, this hypothesis should be tested with direct measurements of UV pressure. Similarly, future experiments would be strengthened by the simultaneous measurement of umbilical artery pressure. We speculate that adrenergic drive may be contributing to tachycardia and peripheral vasoconstriction on the AP that would be enhanced by the measurement of circulating catecholamines. Secondly, although the addition of a centrifugal pump to the AP improved fetal pig hemodynamics, we periodically encountered air-entrainment into the circuit, as well as cavitation of fetal blood at excessively high negative pressures, resulting in diminished circuit flows. Although we did not detect pump-induced hemolysis, this could be a limiting factor during long-term AP experiments and could explain the fetal anemia on the circuit. Finally, systemic inflammation may have also contributed to the hemodynamic instability and cardiovascular decompensation we observed in pumped AP fetal pigs as centrifugal pumps have been reported to induce low-level systemic inflammation\u003csup\u003e51\u003c/sup\u003e. The analysis of inflammatory cytokines may be helpful to delineate the role of inflammation in fetal pig circulatory physiology on the AP.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eFuture directions.\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRecent developments in AP technology have enabled sustained extrauterine fetal life on a pumpless arteriovenous ECMO circuit. Of note, these experiments were conducted using customized hollow-fiber membrane oxygenators that are unavailable for commercial use. Our initial approach in developing a pumpless AP system involved cannulas with the largest luminal diameter to wall thickness ratio possible to minimize circuit resistance. Our failure to demonstrate hemodynamic stability using a pumpless circuit encouraged our team to explore the use of mechanical support in the extremely preterm fetal pig. While the addition of a pump to the circuit improved survival, we observed supraphysiologic circuit flows, evidence of adrenergic drive, and high cardiac output at the start of support, followed by diminished UV flows with persistent tachycardia and hypertension in keeping with diminished cardiac output. This pattern may indicate a progressive increase in circuit resistance resulting from constriction in the cord vessels. An alternative approach to normalizing UV pressures and minimizing the pressure difference across the UV cannula would be to introduce a pressure drop in the circuit through restriction of the circuit or cannula lumen. According to Hagan-Poiseuille\u0026rsquo;s law, reducing the calibre of the venous side of the circuit would increase circuit resistance and limit the supraphysiologic circuit blood flow seen at the onset of our AP experiments.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMiniaturization of the oxygenator and pump head may also help to achieve hemodynamic stability in our animal model. In sheep, placental blood volume at ~116 days GA (term = 145 days) is ~60-80 ml/kg, which represents ~30-40 ml for a 0.5 kg fetus\u003csup\u003e52,53\u003c/sup\u003e. The AP circuit used in the present study has a priming volume that is nearly double the expected placental blood volume for a fetal sheep delivered at 116 days gestation, which likely accounts for the increased afterload we seemed to induce in our pumpless AP experiments, as well as contributing to fetal anemia. Reductions in the oxygenator membrane surface area could also reduce the need for excessive heparin administration. Thus, modifications to the circuit through miniaturization of the oxygenator and centrifugal pump may be helpful for establishing an AP system capable of supporting the long-term physiological requirements of extremely preterm fetal pigs. \u0026nbsp;\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eOver the past decade, important advances have been achieved in the field of artificial womb/AP technology. In experiments using a miniature pig model of the AP in which commercial neonatal ECMO oxygenators were connected the fetal circulation via the umbilical cord we demonstrated a marked improvement in survival, UV flow, and oxygen delivery in circuits that incorporated a small centrifugal pump compared with animals maintained on a pumpless circuit. However, despite the addition of the pump, we observed a progressive diminution of UV flow with persistent tachycardia and hypertension, which we attribute to preload imbalance, increased sympathetic tone, and UV hypertension. Thus, despite observing a clear short-term benefit with the addition of a centrifugal pump in supporting the fetal pig hemodynamics, we conclude that further modifications to the AP circuit are needed. A reduction to the size of the UV cannula could represent an alternative approach for mediating physiologic circuit flows and venous pressures. Given the limitations of neonatal intensive care therapies in preventing iatrogenic organ injury and neonatal death, we remain hopeful that the AP could provide reductions in the mortality and morbidity associated with preterm birth by maintaining a fetal circulation while allowing for normal growth and development of fetal organ systems. \u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no competing interests to disclose.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConception or design of the work: MS, JLM, CH.\u003c/p\u003e\n\u003cp\u003eData acquisition: AJCP, SKSC, TA, LS, AAF, MJM, AS, ME, LCL, CF, DM, JB, BSS, JL, OJM, FL, JRTD, MS, JLM, CH.\u003c/p\u003e\n\u003cp\u003eData analysis or interpretation: AJCP, SKSC, TA, AAF, MJM, MQ, JRTD, MS, JLM, CH.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eDrafting the work or revising it critically for important intellectual content: AJCP, SKSC, TA, LS, AAF, MJM, JB, JRTD, MS, JLM, CH.\u003c/p\u003e\n\u003cp\u003eAll authors approved the final version of the manuscript, agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved, and all persons designated as authors qualify for authorship, and all those who qualify for authorship are listed.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe artificial placenta experiments were generously supported by funding from the Canadian Institutes of Health Research and Breakfast of Champions and\u0026nbsp;BMO Financial Group Chair in Cardiology to MS.\u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eLandrace pig studies and JLM were funded by an ARC Future Fellowship (Level 3; FT170100431). BSS was funded by SickKids Research Institute\u0026rsquo;s graduate scholarship program, Restracomp.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data that support the findings of this study are available from the corresponding author upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors would like to acknowledge Luke Itani for his illustrations of Figures 1 and Supplementary Figure 1. The authors would also like to acknowledge the staff members in the Lab Animal Services at The Hospital for Sick Children, Peter Gilgan Center for Research and Learning, Early Origins of Adult Health Research Group and PIRL, SAHMRI.\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003ePatel, R. 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Rep. \u003cb\u003e9\u003c/b\u003e, e14999 (2021).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Artificial placenta, preterm, pig, cannulation, centrifugal pump, oxygenator, tachycardia, hypertension, fetal development","lastPublishedDoi":"10.21203/rs.3.rs-1251735/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1251735/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe recent demonstration of normal development of preterm sheep in an artificial extrauterine environment has renewed interest in artificial placenta (AP) systems as a potential treatment strategy for extremely preterm human infants. However, the feasibility of translating this technology to the human preterm infant remains unknown. Here we report the support of 13 preterm fetal pigs delivered at 102±4 days (d) gestation, weighing 616±139g with a circuit consisting of an oxygenator and a centrifugal pump, comparing these results with our previously reported pumpless circuit (n=12; 98±4d; 743±350g). The umbilical vessels were cannulated, and fetuses were supported for 46.4±46.8 hours using the pumped AP versus 11±13 hours on the pumpless AP circuit. Upon initiation of AP support on the pumped system, we observed supraphysiologic circuit flows, tachycardia, and hypertension, while animals maintained on a pumpless AP circuit exhibited subphysiologic flows. On the pumped AP circuit, there was a progressive decline in UV flow and oxygen delivery. We conclude that the addition of a centrifugal pump to the AP circuit improves survival of preterm pigs by augmenting UV flow through the reduction of right ventricular afterload. However, we continued to observe the development of heart failure within a matter of days.\u003c/p\u003e","manuscriptTitle":"Advantages of a Centrifugal Pump in the Development of a Swine Model of an Artificial Placenta","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2022-01-24 18:37:46","doi":"10.21203/rs.3.rs-1251735/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"67df32a5-6d4c-41ed-b728-767129085807","owner":[],"postedDate":"January 24th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":9953460,"name":"General Cell Biology \u0026 Physiology"}],"tags":[],"updatedAt":"2022-03-16T06:59:23+00:00","versionOfRecord":[],"versionCreatedAt":"2022-01-24 18:37:46","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-1251735","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1251735","identity":"rs-1251735","version":["v1"]},"buildId":"wLkW0s4AflPzk-lpfg-fK","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}