Male factor infertility and placental pathology in singleton live births conceived with in vitro fertilization.

Purpose We sought to determine whether pregnancies conceived in those with male factor infertility have unique placental pathology profiles compared to those undergoing infertility treatments for other indications.

This was a retrospective cohort study of placental pathology from 464 live births conceived from autologous fresh IVF cycles at an academic fertility center from 2004 to 2017. Placental pathology was compared between live births arising from patients with male factor infertility alone and those with another infertility diagnosis. Placental outcomes were compared with parametric or non-parametric tests; logistic regression was performed to account for potential confounders.

Compared to cycles performed for a non-male factor diagnosis, male factor infertility cycles had a higher mean paternal age (38.2 years vs. 36.5 years, p < 0.001), a higher female mean BMI (24.3 vs. 23.3 kg/m2, p = 0.01), and a lower day 3 follicle stimulating hormone (FSH) level (6.8 vs. 7.3 IU/mL, p = 0.02). The mean numbers of embryos transferred, and day of transfer were similar between groups, and more cycles used ICSI in the male factor infertility group (90.6% vs. 22.5%, p < 0.001). Placental pathology in our adjusted model was similar between the male factor and non-male factor groups. In our unadjusted subgroup analysis, cycles for male factor using ICSI appeared to lead to more small placentas by weight compared to cycles performed with conventional insemination (45.8% < 10th percentile vs. 18.8%, p = 0.04).

Male factor infertility is not associated with significantly different placental pathology compared to other infertility diagnoses.

IVF, ICSI, Placenta, Pathology, Male factor infertility

The unique paternal contribution to placental makeup has long been known but has become of greater interest over the past decade. It was initially explored in the molar pregnancy literature in humans, which highlights their androgenetic makeup [1, 2]. Subsequent advances in epigenetics led to the discovery of imprinted genes, in which genes are only expressed from either the paternal or maternally inherited allele [3]. Imprinted genes in the placenta have demonstrated a careful balance between maternally and paternally expressed genes. Any alteration in this balance could have significant clinical consequences, with associations between alterations in imprinting and fetal growth, neonatal outcomes and childhood neurodevelopment and cancer risk [4–6]. Male infertility could affect this balance. Up to 50% of couples presenting for infertility are found to have male factor infertility [7]. Currently, semen analysis remains the most prevalent method of assessing infertility [8] but serves primarily as a quantitative assessment. Oligospermia is the most common male infertility diagnosis [9] and affected individuals have been shown to have higher concentrations of reactive oxidative species (ROS) present in seminal fluid [10]. In turn, elevated seminal ROS has been associated with recurrent miscarriage and adverse effects on offspring in both animal studies and in humans [7, 11–14]. Men with oligospermia and other spermatic abnormalities including asthenozoospermia have also been demonstrated to have higher rates of alterations in DNA methylation [15–17], including at paternally imprinted gene loci, including IGF2 and PEG3, both genes which are implicated in placental and fetal growth [18]. These epigenetic changes have been postulated to have a “late paternal effect” which has been associated with failure of implantation and early morphological abnormalities [11]. Paternal age and paternal obesity have also been linked with both male factor infertility and epigenetic changes which may impact placental functioning [19, 20]. The prevalence of male factor infertility, including oligospermia, has been increasing in developed nations [21]. The reasons for this remain unclear, but epidemiological studies have linked oligospermia with environmental factors, increasing paternal age and an increasing burden of chronic conditions (e.g., diabetes and obesity) [15]. Thus, it is of interest and clinically relevant that we establish its consequences for maternal and neonatal outcomes. At present, the most common intervention for couples diagnosed with male factor infertility and desiring pregnancy is assisted reproductive technology (ART), often utilizing in vitro fertilization with intracytoplasmic cytoplasmic sperm injection (ICSI) [22]. ART is independently associated with placental pathology, which in turn may be associated with maternal and neonatal morbidity [4, 5, 23, 24]. While there is still no consensus as to whether the ART procedures or the underlying infertility diagnosis lead to this increased risk, it is likely that both play a role [6, 25, 26]. There has been little research to date on the impact of male factor infertility on placental outcomes. Given that infertility is associated with spermatic DNA damage by ROS and altered epigenetic patterns which may be expressed in the placenta, we hypothesized that male factor infertility may be associated with unique placental pathology compared to pregnancies conceived by IVF/ICSI for other diagnoses.

Study design We conducted a retrospective cohort study of singleton live births resulting from autologous IVF pregnancies at an academic institution between 2004 and 2017 with placental pathology available. The study was approved by the Institutional Review Board (#2015P002672) of the Massachusetts General Hospital. Data was collated from the hospital electronic medical record system, the internal IVF database, and the internal pathology database. Exclusion criteria were pregnancies resulting in a multiple gestation, women with uterine factor infertility, and pregnancies conceived with frozen embryo transfers, as these have been linked to placental abnormalities [26]. Donor oocyte and gestational carrier cycles were also excluded. The primary outcomes were placental pathology, subdivided into anatomic, inflammatory, infectious or vascular/thrombotic subgroups, as we have described previously [26]. Infertility diagnosis All patients underwent a standard fertility evaluation. Male factor infertility was defined per SART criteria as an alteration in sperm concentration and/or motility and/or morphology in at least one sample of two sperm analyzes, collected on average 4 weeks apart [27]. Male factor infertility was further classified as mild if the total motile sperm count (TMC, volume x concentration x % motile) was 5–15 million and severe if the TMC was less than 5 million. Non-male factor infertility was classified as patients without any known diagnosis of male factor and with a recorded TMC over 15 million. Ovarian stimulation and transfer protocols Patients underwent controlled ovarian hyperstimulation, as previously described, by luteal-phase gonadotropin-releasing hormone (GnRH) agonist, GnRH-antagonist downregulation, Patch antagonist, or GnRH agonist flare protocol, as clinically indicated and described previously [26, 28]. Follicular measurements, endometrial thickness, and serum estradiol were monitored during the cycles. Intramuscular human chorionic gonadotropin (hCG) (10,000 IU, Novarel, Ferring Pharmaceuticals or 10,000 IU, Pregnyl, Merck) was administered to induce final oocyte maturation and 35–37 h later the patients underwent a transvaginal ultrasound-guided oocyte retrieval [26, 28]. Peak estradiol was measured on the day of hCG trigger. Oocytes then underwent conventional insemination or intracytoplasmic sperm injection (ICSI) as clinically indicated. Our laboratory’s criteria for ICSI includes concentration less than 1 million/mL, motility less than 2%, forward progression score less than 3, or Kruger strict morphology less than 4% normal forms. Couples with prior failed fertilization also undergo ICSI. For those undergoing ICSI, the cumulus cells were stripped from the oocytes 2–3 h after retrieval. Embryos meant for fresh transfer were cultured until post-retrieval day two, three or five, and the decision for number of embryos transferred was made based on institutional and ASRM guidelines [29–32]. Placental pathology assessment All placentas from deliveries at MGH were received at the MGH pathology department and triaged either to undergo complete (gross and histologic) pathologic examination or storage for two weeks before incineration based on provided clinical history, as previously described [5]. A provided clinical history of ART and/or age ≥ 40 years is an indication for full gross and histopathologic examination. The pathology reports of the study patients were reviewed by an experienced placental pathologist (DJR), and the pathologic findings, if present, were categorized as follows: anatomic (e.g., small 90%ile by weight, marginal cord insertion; membranous cord insertion; single umbilical artery; membranous vessels; circummarginate or circumvallate membranes; accessory placental lobe), inflammatory (e.g., villitis of unknown etiology, both low and high grade; chronic deciduitis; chronic histocytic intervillositis), infectious (e.g., acute chorioamnionitis with maternal or fetal inflammatory response,), or vascular (e.g., maternal vascular malperfusion; fetal vascular malperfusion, both low and high grade; intervillous thrombi; subchorionic thrombi; septal thrombi) using accepted diagnostic criteria from the Amsterdam Placental Workshop [26, 33]. Outcomes of interest We compared placental pathology findings in couples with a single infertility diagnosis of male factor infertility to those undergoing IVF/ICSI for other indications and with a documented TMC > 15 million. Statistical analysis Demographic and cycle outcomes were compared with parametric (chi-square, Student’s t-test) or non-parametric tests as appropriate. Multivariate logistic regression models were fitted to estimate odds ratios (OR) and 95% confidence intervals (CIs) of placental pathology findings, adjusted for maternal age, paternal age, race, BMI, gestational age at delivery, number of embryos transferred, and fertilization method. All analyses were conducted using the Stata/IC program (StataCorp, College Station, TX). A two-sided significance level less than 0.05 was considered to be statistically significant, along with a 95% CI that did not contain 1.

Patient demographics and cycle characteristics Autologous IVF cycles from 2004 to 2017 were reviewed for live births. Of 6534 fresh autologous cycles with a transfer, 2648 cycles led to a live birth (40.5%), of which 464 met our inclusion criteria with placental pathology available for review from delivery within the Partners Healthcare system (17.5%). There were 293 cycles for patients with a non-male factor diagnosis and TMC greater than 15 million, and there were 171 cycles performed for couples with male factor infertility alone. Demographic and cycle characteristics per cycle leading to live birth are presented in Table 1. Paternal age was significantly higher in the male factor infertility group compared to pregnancies with a non-male factor ART diagnosis: mean 38 years vs. 36 years, p < 0.001). Maternal BMI was significantly higher in the male factor infertility group (24 vs 23, p = 0.01). Maternal age was similar between the groups, and the majority of patients were Caucasian. 75–78% of women were nulliparous. Non-male factor ART diagnosis group had higher mean FSH level compared to the male factor infertility group, but similar AMH and estradiol levels. There were differences between IVF stimulation protocols between groups, with 80% of the male factor infertility group undergoing a GnRH agonist protocol compared to 63% non-male factor infertility group (p < 0.001). There were no significant differences between numbers of embryos transferred per cycle or the day of transfer. There was a significant difference in fertilization method between groups, with 90% of male factor pregnancies utilizing ICSI, compared to 23% of non-male factor pregnancies. For obstetric outcomes, shown in Table 2, there were no differences between those with and without male factor infertility. Table 1. | Non-male factor ART diagnosish (N = 293) | Male factor infertility (N = 171) | p-valuei | | |---|---|---|---| | Paternal factors | ||| | Paternal age (years) | 36.5 (4.8) | 38.2 (5.9) | 0.001 | | Maternal factors | ||| | Maternal age (years) | 35.0 (3.9) | 34.6 (3.9) | 0.32 | | Race | 0.15 | || | White (1) | 224 (76.5%) | 136 (79.5%) | | | Black (2) | 13 (4.4%) | 5 (2.9%) | | | Asian (3) | 44 (15.0%) | 17 (9.9%) | | | Other | 12 (4.1%) | 13 (7.6%) | | | Ethnicity | 0.30 | || | Hispanic (4) | 11 (3.8%) | 10 (5.8%) | | | BMI (median, IQR)b | 23.3 (21.2, 26.65) | 24.3 (22, 27.75) | 0.01 | | Nulliparous | 237 (80.9%) | 129 (75.4%) | 0.17 | | Day 3 FSHc | 7.3 (2.5) | 6.8 (1.7) | 0.02 | | AMH (2014–2017)d | 3.6 (4.1) | 4.0 (2.9) | 0.50 | | Peak estradiole | 43.5 (32.2) | 41.1 (17.9) | 0.36 | | IVF stimulation protocol | 0.001 | || | GnRH agonist | 187 (65.5%) | 141 (82.5%) | | | Flare | 53 (18.1%) | 16 (9.4%) | | | GnRH antagonist | 42 (14.3%) | 14 (8.2%) | | | Patch | 6 (2.0%) | 0 (0.0%) | | | Number of embryos transferred | 1.9 (0.78) | 1.9 (0.79) | 0.99 | | Stage of transfer | 0.63 | || | Cleavage stage | 146 (49.8%) | 85 (49.7%) | | | Blastocyst stage | 147 (50.2%) | 86 (50.3%) | | | Fertilization method | < 0.001 | || | IVFf | 227 (77.5%) | 16 (9.4%) | | | ICSIg | 66 (22.5%) | 155 (90.6%) | aMale infertility categorized by SART diagnosis bBMI body mass index, (kg/m2) cFSH follicle stimulating hormone, iu/L dAMH anti-Mullerian hormone, ng/ml epg/ml fGnRH gonadotropin releasing hormone gIVF in vitro fertilization hICSI intracytoplasmic sperm injection iOther diagnosis of infertility includes tubal or ovarian causes of infertility; uterine factors excluded. Cases included in the “non-male factor” group all have TMC > 15 million jp < 0.05 is significant Table 2. | Non-male factor ART diagnosis (N = 293) | Male factor infertility (N = 171) | p-valueb | | |---|---|---|---| | Gestational age at delivery (weeks) | 38.4(2.7) | 38.2 (2.9) | 0.64 | | Preterm delivery (% < 37 weeks) | 52 (17.7%) | 37 (21.6%) | 0.30 | | Median singleton birth weight (g) | 3195 (2730, 3547) | 3125 (2725, 3530) | 0.56 | | SGA infant | 30 (10.2%) | 24 (14.0%) | 0.32 | | LGA infant | 12 (4.1%) | 12 (7.0%) | 0.23 | | Mode of delivery | 0.27 | || | Vaginal | 168 (57.3%) | 88 (51.5%) | | | Cesarean | 115 (39.2%) | 75 (43.9%) | | | Unknown | 10 (3.4%) | 8 (4.7%) | | | Preeclampsia | 40 (13.7%) | 22 (12.9%) | 0.81 | | Postpartum hemorrhage | 11 (3.8%) | 7 (4.1%) | 0.82 | aMale infertility categorized by SART diagnosis. All non-male factor ART patients have a TMC > 15 million [1] bp < 0.05 is significant Placental pathology in those with male factor infertility The rates of placental pathologies are shown in Table 3. Compared to placentas of couples that underwent ART but did not have a diagnosis of male factor infertility, placentas in the male factor infertility group had a lower number of membranous cord insertions (2.9% vs 7.5%, p = 0.05). This association was stronger when a subgroup of couples with severe male factor infertility was looked at specifically. Placentas delivered from patients in the severe male factor infertility group had a significantly lower number of membranous cord insertions compared to placentas of couples with other diagnoses of infertility (0% vs 7.2%, p = 0.03, Table 3). Otherwise, there were no significant differences in placental weight or infectious, inflammatory, or vascular pathology between couples with a diagnosis of male factor infertility or another diagnosis of infertility. Table 3. | Non-male factor ART diagnosisc (N = 293) | Male factor infertilityd (N = 171) | Severe male factor infertilitye (N = 59) | p-valuef Other vs male | p-valuef Other vs severe male | | |---|---|---|---|---|---| | Mean placental weight (g) | 450.7(114.2) | 440.8 (114.8) | 443.4 (123.0) | 0.37 | 0.66 | | Mean placental weight percentile | 32.6 (31.0) | 30.5 (30.4) | 31.3 (31.6) | 0.48 | 0.78 | | Anatomic | 197 (67.2%) | 117 (68.4%) | 35 (59.3%) | 0.79 | 0.24 | | Small ( 90%ile) | 26 (8.9%) | 14 (8.2%) | 5 (8.5%) | 0.80 | 0.92 | | Marginal cord insertion | 24 (8.2%) | 19 (11.1%) | 7 (11.9%) | 0.30 | 0.36 | | Membranous cord insertion | 22 (7.5%) | 5 (2.9%) | 0 (0.0%) | 0.04 | 0.03 | | Single umbilical artery | 4 (1.4%) | 3 (1.8%) | 2 (3.4%) | 0.74 | 0.27 | | Membranous vessels | 2 (0.7%) | 6 (3.5%) | 1 (1.7%) | 0.02 | 0.44 | | Circummarginate or circumvallate membranes | 17 (5.8%) | 11 (6.4%) | 3 (5.1%) | 0.78 | 0.83 | | Accessory placental lobe | 20 (6.8%) | 6 (3.5%) | 2 (3.4%) | 0.13 | 0.32 | | Infectious | 98 (33.4%) | 49 (28.7%) | 22 (37.3%) | 0.28 | 0.57 | | Acute chorioamnionitis with maternal inflammatory response, moderate to severe | 33 (11.3%) | 23 (13.5%) | 8 (13.6%) | 0.49 | 0.62 | | Acute chorioamnionitis with fetal inflammatory response, moderate to severe | 13 (4.4%) | 10 (5.8%) | 2 (3.4%) | 0.50 | 0.72 | | Inflammatory | 54 (18.4%) | 29 (17.0%) | 14 (23.7%) | 0.69 | 0.35 | | Villitis of unknown etiology, low-grade | 25 (8.5%) | 17 (9.9%) | 8 (13.6%) | 0.61 | 0.23 | | Villitis of unknown etiology, high-grade | 8 (2.7%) | 5 (2.9%) | 3 (5.1%) | 0.90 | 0.34 | | Chronic deciduitis | 16 (5.5%) | 10 (5.8%) | 5 (8.5%) | 0.86 | 0.37 | | Chronic histiocytic intervillositis | 1 (0.3%) | 1 (0.6%) | 0 (0.0%) | 0.70 | 0.65 | | Vascular | 195 (66.6%) | 113 (66.1%) | 41 (69.5%) | 0.92 | 0.66 | | MVMa | 38 (13.0%) | 29 (17.0%) | 11 (18.6%) | 0.24 | 0.25 | | FVMb, low grade | 27 (9.2%) | 9 (5.3%) | 3 (5.1%) | 0.12 | 0.30 | | FVMb, high grade | 8 (2.7%) | 3 (1.8%) | 2 (3.4%) | 0.51 | 0.78 | | Intervillous thrombi | 48 (16.4%) | 30 (17.5%) | 11 (18.6%) | 0.75 | 0.67 | | Subchorionic thrombi | 5 (1.7%) | 4 (2.3%) | 2 (3.4%) | 0.63 | 0.40 | | Septal thrombi | 1 (0.3%) | 0 (0.0%) | 0 (0.0%) | 0.44 | 0.65 | aMVM refers to maternal vascular malperfusion bFVM refers to fetal vascular malperfusion cOther diagnosis of infertility includes tubal or ovarian causes of infertility; uterine factors excluded. Cases included in the “non-male factor” group all have TMC > 15 million d,eMale infertility categorized by total motile sperm count (TMC), with TMC < 5million characterized as severe male factor infertility, TMC 5–15 million characterized as mild male factor infertility [1] fp < 0.05 is significant Placental pathology by fertilization method in those with male factor infertility To better understand the role of fertilization method, given the high percentage of those with male factor undergoing ICSI, we analyzed only those patients in each group who underwent ICSI, and found that there were no significant differences in placental pathology between couples with male factor compared to those with female factor (Table 4). However, when placental pathology was compared between couples with male factor infertility who underwent conventional insemination (N = 16) and those who underwent ICSI (N = 155, Table 5), there were significantly more placentas < 10%ile in the ICSI group (45% vs 18%, p = 0.04; mean placental weight 436 g vs. 484 g, p = 0.11). There was also a trend towards an increase in maternal vascular malperfusion in the ICSI group (18% vs 0%, p = 0.06. Placentas in the ICSI group had lower rates of low-grade villitis of unknown etiology (VUE) (8.4% vs 25%, p = 0.03), but similar rates of high-grade VUE and overall inflammatory pathology (including chronic chorioamnionitis and plasma cell deciduitis). There were no other significant differences in placental pathology between groups. Table 4. | Non-male factor ART diagnosis, using ICSIc (N = 66) | Male factor infertility, using ICSId (N = 155) | p-valuee | | |---|---|---|---| | Mean singleton placental weight (g) | 435.9 (107.7) | 436.3 (113.7) | 0.98 | | Mean placental weight percentile | 27.2 (28.8) | 28.9 (29.8) | 0.70 | | Anatomic | 43 (65.2%) | 108 (69.7%) | 0.51 | | Small ( 90%ile) | 4 (6.1%) | 13 (8.4%) | 0.55 | | Marginal cord insertion | 10 (15.2%) | 17 (11.0%) | 0.38 | | Membranous cord insertion | 3 (4.5%) | 5 (3.2%) | 0.63 | | Single umbilical artery | 1 (1.5%) | 2 (1.3%) | 0.89 | | Membranous vessels | 0 (0.0%) | 5 (3.2%) | 0.14 | | Circummarginate or circumvallate membranes | 3 (4.5%) | 10 (6.5%) | 0.58 | | Accessory placental lobe | 4 (6.1%) | 6 (3.9%) | 0.47 | | Infectious | 20 (30.3%) | 44 (28.4%) | 0.77 | | Acute chorioamnionitis with maternal inflammatory response, moderate to severe | 6 (9.1%) | 20 (12.9%) | 0.42 | | Acute chorioamnionitis with fetal inflammatory response, moderate to severe | 2 (3.0%) | 9 (5.8%) | 0.39 | | Inflammatory | 15 (22.7%) | 25 (16.1%) | 0.24 | | Villitis of unknown etiology, low-grade | 6 (9.1%) | 13 (8.4%) | 0.86 | | Villitis of unknown etiology, high-grade | 2 (3.0%) | 5 (3.2%) | 0.94 | | Chronic deciduitis | 5 (7.6%) | 10 (6.5%) | 0.76 | | Chronic histiocytic intervillositis | 0 (0.0%) | 1 (0.6%) | 0.51 | | Vascular | 46 (69.7%) | 103 (66.5%) | 0.64 | | MVMa | 7 (10.6%) | 29 (18.7%) | 0.14 | | FVMb, low grade | 7 (10.6%) | 8 (5.2%) | 0.14 | | FVMb, high grade | 3 (4.5%) | 3 (1.9%) | 0.27 | | Intervillous thrombi | 10 (15.2%) | 28 (18.1%) | 0.60 | | Subchorionic thrombi | 1 (1.5%) | 4 (2.6%) | 0.63 | | Septal thrombi | 0 (0.0%) | 0 (0.0%) | – | aMVM refers to maternal vascular malperfusion bFVM refers to fetal vascular malperfusion cOther diagnosis of infertility includes tubal or ovarian causes of infertility; uterine factors excluded. Cases included in the “non-male factor” group all have TMC > 15 million dMale infertility categorized by total motile sperm count (TMC), with TMC < 5million characterized as severe male factor infertility, TMC 5–15 million characterized as mild male factor infertility [1] ep < 0.05 is significant Table 5. | Male factor infertilityc using conventional insemination (N = 16) | Male factor infertility,c using ICSI (N = 155) | p-valued | | |---|---|---|---| | Mean singleton placental weight (g) | 484.1 (120.3) | 436.3 (113.7) | 0.11 | | Mean placental weight percentile | 47.1 (32.8) | 28.9 (29.8) | 0.03 | | Anatomic | 9 (56.2%) | 108 (69.7%) | 0.27 | | Small ( 90%ile) | 1 (6.2%) | 13 (8.4%) | 0.77 | | Marginal cord insertion | 2 (12.5%) | 17 (11.0%) | 0.85 | | Membranous cord insertion | 0 (0.0%) | 5 (3.2%) | 0.47 | | Single umbilical artery | 1 (6.2%) | 2 (1.3%) | 0.15 | | Membranous vessels | 1 (6.2%) | 5 (3.2%) | 0.53 | | Circummarginate or circumvallate membranes | 1 (6.2%) | 10 (6.5%) | 0.98 | | Accessory placental lobe | 0 (0.0%) | 6 (3.9%) | 0.42 | | Infectious | 5 (31.2%) | 44 (28.4%) | 0.81 | | Acute chorioamnionitis with maternal inflammatory response, moderate to severe | 3 (18.8%) | 20 (12.9%) | 0.51 | | Acute chorioamnionitis with fetal inflammatory response, moderate to severe | 1 (6.2%) | 9 (5.8%) | 0.94 | | Inflammatory | 4 (25.0%) | 25 (16.1%) | 0.37 | | Villitis of unknown etiology, low-grade | 4 (25.0%) | 13 (8.4%) | 0.03 | | Villitis of unknown etiology, high-grade | 0 (0.0%) | 5 (3.2%) | 0.47 | | Chronic deciduitis | 0 (0.0%) | 10 (6.5%) | 0.30 | | Chronic histiocytic intervillositis | 0 (0.0%) | 1 (0.6%) | 0.75 | | Vascular | 10 (62.5%) | 103 (66.5%) | 0.75 | | MVMa | 0 (0.0%) | 29 (18.7%) | 0.06 | | FVMb, low grade | 1 (6.2%) | 8 (5.2%) | 0.85 | | FVMb, high grade | 0 (0.0%) | 3 (1.9%) | 0.57 | | Intervillous thrombi | 2 (12.5%) | 28 (18.1%) | 0.58 | | Subchorionic thrombi | 0 (0.0%) | 4 (2.6%) | 0.52 | aMVM refers to maternal vascular malperfusion bFVM refers to fetal vascular malperfusion cMale factor diagnosis defined as oligospermia, categorized by total motile sperm count (TMC), with TMC < 5million characterized as severe male factor infertility, TMC 5–15 million characterized as mild male factor infertility dp < 0.05 is significant Table 6 presents the unadjusted and adjusted odds of clinically significant placental pathologies in all singleton pregnancies resulting from couples with male factor infertility compared to other causes of infertility. There were no differences in the adjusted odds of having any clinically significant placental pathologies. Table 6. | Unadjusted OR (95% CI), p-value | Adjusted ORb,c (aOR) (95% CI), p-value | | |---|---|---| | Anatomic | 1.06 (0.69, 1.63); 0.79 | 0.82 (0.49, 1.36); 0.44 | | Marginal cord insertion | 1.4 (0.74, 2.64); 0.3 | 0.7 (0.31, 1.55); 0.37 | | Membranous cord insertion | 0.21 (0.04, 1.1); 0.06 | 0.4 (0.07, 2.28); 0.3 | | Accessory lobe | 0.5 (0.2, 1.26); 0.14 | 0.62 (0.25, 1.59); 0.32 | | Infectious | 0.8 (0.53, 1.21); 0.28 | 1.04 (0.62, 1.74); 0.88 | | Inflammatory | 0.88 (0.46, 1.67); 0.7 | 1.09 (0.59, 2.03); 0.77 | | Villitis of unknown etiology, high-grade | 1.12 (0.18, 7); 0.9 | 1.06 (0.22, 5.08); 0.94 | | Vascular/thrombotic | 0.96 (0.51, 1.8); 0.9 | 0.83 (0.44, 1.57); 0.57 | | MVM | 1.64 (0.66, 4.09); 0.29 | 0.95 (0.47, 1.89); 0.87 | | FVM, high grade | 0.64 (0.17, 2.43); 0.51 | 1.42 (0.28, 7.11); 0.67 | | Subchorionic thrombi | 1.38 (0.37, 5.21); 0.63 | 0.65 (0.19, 2.2); 0.49 | aSART diagnosis bAdjusted for maternal age, paternal age, race, BMI, gestational age at delivery, number of embryos transferred, fertilization method cPresented as odds ratio (95% CI)

The use of ART and the incidence of male factor infertility are both increasing, and so it is important to understand the potential impact of these variables on placentation. The maternal hormonal and uterine environment play a critical role in placental development [6], but given the increased understanding of the important role for paternal DNA in placental and fetal growth [5], male factor infertility should not be discounted as a potential contributor to placental development. Increased DNA fragmentation, a common finding in men with male factor infertility, reflects modifications to sperm DNA or chromatin structure, often by methylation or covalent modifications to histones and protamines [34, 35]. Methylation errors are associated with oxidative stress and the creation of reactive oxidative species (ROS) [36]. There is ongoing work to establish whether hypomethylation is associated with effects in the blastocyst, morula, and subsequently the placenta and fetus [37]. Our study is the first to assess whether male factor infertility is associated with any unique placental pathology for couples who conceive a singleton after fresh embryo transfer. Compared to those without male factor infertility, couples with male factor infertility did not have significantly different placental pathology once adjusted for relevant confounders. These results remained true when examining placentas arising from ICSI cycles alone and those from couples with severe male factor infertility. Importantly, when couples with male factor undergoing conventional insemination and ICSI were compared, placentas in the ICSI group tended to be smaller, a finding accompanied by a trend towards increased maternal vascular malformation in the ICSI group as well. However, the number of patients in the conventional insemination group was significantly smaller and results should be interpreted with caution. Our findings are interesting for several reasons. First, it is notable that male factor infertility itself is not associated with any clinically significant adverse placental or obstetric outcomes in our cohort. This should be reassuring to couples with a sole diagnosis of male factor. Second, it is interesting that the placentas arising from couples with female factor infertility did not have significantly increased rates of placental pathology compared to couples with only male factor infertility, i.e., no clinically identified female factor. We observed a significantly increased rate of membranous cord insertion in the non-male factor group in our unadjusted data, but this was no longer significant when adjusted for confounders. On the one hand, we excluded a major cause of abnormal placentation: uterine factor. On the other hand, diagnoses such as endometriosis and polycystic ovarian syndrome are known to be associated with abnormal placentation and adverse pregnancy outcomes [25, 38, 39]. However, there were no significant differences between the two groups in this study. Although our findings are limited by power and small sample size, it does raise the question of whether the previously observed adverse placental outcomes in ART pregnancies could be more attributed to the ART procedures themselves rather than a specific non-uterine factor infertility diagnosis [26, 40]. Finally, a subgroup comparison of couples with male factor undergoing IVF vs. ICSI revealed that there might be increased pathology associated with ICSI in particular. Although limited by low power, placentas of couples with male factor infertility who undergo ICSI were more likely than those who underwent conventional insemination to be less than the 10%ile for weight (placental hypoplasia). Importantly, our male factor cohort had no other infertility diagnoses that might contribute to placental outcomes. This finding is of particular interest as elective ICSI is increasing in use, for indications such as pre-implantation genetic testing for aneuploidy (PGT-A) [41]. Lower birth weight: placental weight ratios have been associated with neonatal morbidity in the literature [42], and so the noted trend that ICSI may contribute to adverse placental outcomes should be studied further in a larger cohort. Sperm DNA oxidative stress and subsequent hypomethylation could also explain the putative differences noted in our study between IVF and ICSI cycles. Theoretically, ROS production in situ is more relevant during the IVF process [43, 44], and this can be impacted by DNA hypomethylation [45], whereas the use of ICSI may bypass this step, leading to a higher pregnancy rate but potentially also an increase in placental pathology. There has been no investigation of the role of sperm epigenetic changes in placental function, but further study with a larger cohort could elucidate this process further. There are many strengths of this study, the first of which is that it was performed at a single center, which decreases inter-observer bias. The placental pathology was reviewed by an expert placental pathologist using the Amsterdam Criteria [33], again reducing inter-observer variation and outcome misclassification. At this center, placental evaluation is routine for women with a noted history of ART, reducing the risk of selection bias. Finally, although this is not a randomized study, baseline maternal and cycle characteristics were comparable across groups, reducing selection bias. The limitations of this study include firstly the fact that the comparison group itself has risk factors for abnormal placentation, as discussed above. A better control group would consist of couples undergoing fertility treatment for a diagnosis such as tubal factor (i.e., minimal female risk factor for abnormal placentation). Unfortunately, we were limited in power to restrict our study population to such a group. Therefore, we opted to exclude women with the strongest pre-existing risk factor for abnormal placentation, uterine factor. While this study focused on assessing paternal contribution to placental functioning and outcomes, the maternal contribution to DNA methylation and placental development is also significant, and this could not be fully adjusted for in this study [46]. An interesting future analysis could compare placental and obstetric outcomes for couples with male factor infertility undergoing intrauterine insemination versus IVF/ICSI. Another limitation to the current study was our low number of couples with severe male factor and those with male factor undergoing conventional insemination. Additionally, pathological placental outcomes of clinical significance are also rare, and so the study may be underpowered for these outcomes. Although we were able to assess severe male factor by oligospermia criteria, other markers of semen quality such as morphology may be more significantly associated with placental dysfunction [11]. Perhaps using the usual semen quality parameters of concentration, motility and morphology that are currently available do not correlate with epigenetic alterations within the genome that can lead to adverse perinatal outcomes. For example, murine studies have demonstrated an association between paternal exposure to toxins that is epigenetically transmitted through generations leading to impaired placentation and preterm birth [47, 48]. These epigenetic changes are detectable at a genomic level, but are not necessarily associated with the usual markers of semen quality that we currently use, i.e., concentration, morphology, or motility. As more refined markers of semen quality are developed, these associations can be assessed more robustly.

Male factor infertility status was not significantly associated with placental outcomes amongst patients in this cohort. Our findings suggest a possible association between ICSI and lower placental weight but numbers were too small to draw valid conclusions. In the setting of an increasing prevalence of both male factor infertility and ICSI, further prospective research is needed to reproduce and validate these findings.

The authors wish to thank the MGH IVF laboratory and Department of Pathology for their contributions to data collection. Footnotes Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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