Impact of SARS-CoV-2 Infection During Various Pregnancy Trimesters on Maternal and Fetal Outcomes

Impact of SARS-CoV-2 Infection During Various Pregnancy Trimesters on Maternal and Fetal Outcomes | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Impact of SARS-CoV-2 Infection During Various Pregnancy Trimesters on Maternal and Fetal Outcomes Mengyue Yin, Lingling Peng, Chunling Zhang, Xueyan Zhang, Mei Han, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4488503/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 Background The specific impact and pathology of long COVID effects on mother and fetus post-infection have not been deeply investigated. Therefore, we explored the inflammatory response in fetal tissue from pregnant women infected at various stages. Methods We collected villi, decidua, and placenta samples from 57 patients at Tai'an Central Hospital, who underwent either induced abortion or delivery between November 2022 and March 2023. HE staining was employed for histological examinations of these tissues. Furthermore, the concentrations of IL-6 in both umbilical cord blood and amniotic fluid were quantitatively analyzed using ELISA kits. Additionally, Western blot analysis was conducted to evaluate the expression levels of TNF-α and IL-1β in the placental tissues. Results Among participants, 53 pregnant women were diagnosed with SARS-CoV-2 infection. No significant placental pathology differences were found between mid-term and late-term stages with the control group (P>0.05). However, Western blot analysis indicated that infected placental tissues showed higher TNF-α and IL-1β levels (P0.05). IL-6 levels in amniotic fluid showed no significant difference, and detected no IL-6 expression in umbilical cord blood. Conclusion Infection with SARS-CoV-2 during different periods of pregnancy can have different effects on the placenta and other tissues, emphasizing the importance of preventing and managing viral infection during pregnancy and providing clinical reference for formulating management strategies for SARS-CoV-2 infection. SARS-CoV-2 Pregnancy Inflammatory reaction Figures Figure 1 Figure 2 Background On March 11, 2020, the World Health Organization (WHO) declared a global pandemic due to coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) [ 1 , 2 ]. SARS-CoV-2 is an RNA virus enveloped with a nucleocapsid protein, primarily transmitted through respiratory droplets and close contact, exhibiting strong infectivity. Pregnant women are considered susceptible to the virus [ 3 ]. To maintain the high metabolic demands required for normal fetal development during pregnancy, the burden of oxidative stress in pregnant women increases significantly. Due to alterations in the physiological and immunosuppressive states during pregnancy, clinical outcomes of SARS-CoV-2 infection in pregnant women may differ from the general population. Diagnosis and treatment of SARS-CoV-2 infection during pregnancy may increase the health risks of both the mother and the newborn [ 4 – 6 ]. The long COVID effect, known as "post-acute sequelae of SARS-CoV-2 infection" or PASC, leads to weakness and affects multiple organ systems [ 7 , 8 ]. Research indicates that SARS-CoV-2 can impact the reproductive system, manifesting as a decline in ovarian reserve and reproductive endocrinological imbalances in COVID-19 patients, affecting the production of ovarian hormones and/or the response of the uterine lining [ 8 – 10 ]. Additionally, babies born to women who had COVID-19 during pregnancy are more likely to undergo neurodevelopmental diagnosis within the first year after birth, with the extent and pathology of the impact still requiring further research [ 11 , 12 ]. Understanding the impact of SARS-CoV-2 infection at different stages of pregnancy on maternal and infant outcomes, as well as the long-term effects of infection during pregnancy, provides a reference for the management of newborns after maternal SARS-CoV-2 infection [ 12 ]. The research employs H&E staining for histological examination, intended to observe pathological alterations in the placenta. Western Blot analysis was used to detect the expression levels of TNF-α and IL-1β in placental tissues. IL-6 levels in umbilical cord blood and amniotic fluid were determined using the ELISA technique. Analyzing the impact of SARS-CoV-2 infection on placental immune response will provide valuable information for developing effective management and prevention strategies to reduce adverse outcomes of SARS-CoV-2 infection during pregnancy [ 13 ]. Methods Participants We admitted 57 pregnant women from the Obstetrics Department of Tai'an Central Hospital as the study population from November 2022 to March 2023. All pregnant women were singleton, and relevant examinations excluded primary diseases and other diseases. The patient group with infections totaled 53 cases, divided based on SARS-CoV-2 infection status and timing into early pregnancy infection group (1st to 12th week of pregnancy, n=20), mid-pregnancy group (13th to 28th week of pregnancy, n=14) and late-pregnancy infection group (29th week of pregnancy to delivery, n=19). There were 4 cases in the non-infection group, serving as the control group. According to the "Diagnosis and Treatment Protocol for Novel Coronavirus Pneumonia (Trial version 6)," all patients were categorized as ordinary cases. All the selected pregnant women gave informed consent and signed the consent form. There were no statistical differences in age and parity among the groups of pregnant women (P > 0.05) ( Table 1 ). Table 1 Participant characteristics Early pregnancy infection (n=20) Mid-pregnancy infection (n=14) Late-pregnancy infection (n=19) Control (n=4) P value Age (years) Mean (SD) 31.8 (5.9) 28.4 (3.6) 31.0 (5.0) 28.0 (4.3) >0.05 Parity Mean (SD) N/A 1.3 (0.5) 1.7 (0.8) 1.3 (0.5) >0.05 Abbreviations: n, number of participants with available data; SD, standard deviation. Instruments and reagents Anti-TNF-α antibody, recombinant rabbit monoclonal antibody against IL-1β, horseradish peroxidase-conjugated sheep anti-rabbit secondary antibody, and β-Actin antibody were purchased from Beijing Boaosen Biotechnology Co., LTD. Pre-stained protein markers and pre-mixed glue kits were purchased from Shanghai Biyantian Biotechnology Co., LTD. RIPA tissue lysate and protein loading buffer were purchased from Beijing Pulley Gene Technology Co., LTD. The human IL-6 ELISA test kit was purchased from Wuhan Doctor De Bioengineering Co., LTD. CKX53 inverted phase contrast microscope (OLYMPUS), microplate reader (Thermo Fisher Scientific). Pathological The chorionic villi and decidua tissues of patients with induced abortion and placenta tissues of pregnant women after cesarean section were collected after infection in early pregnancy, washed with sterile saline, and fixed in 4% paraformaldehyde for 48 hours. The slides were routinely embedded in paraffin and HE stained, and each slide was 4 μm thick. All HE slides were observed and reviewed by an attending physician and an associate chief physician. IL-6 levels in cord blood serum and amniotic fluid Before the rupture of membranes during the cesarean section, amniotic fluid was extracted with a syringe and centrifuged at 3000 r/min for 15 min. After separation, the supernatant was collected in EP tubes and stored in a freezer at -80℃. At delivery, 5 mL of umbilical cord blood was collected into blood collection tubes and centrifuged at 3000 r/min for 15 min. Serum was divided into 300 μL frozen tubes and stored in the refrigerator at -80 ℃. According to the kit instructions, perform the assay and calculate the corresponding concentration based on the standard curve according to the OD value of the sample. TNF-α and IL-1β levels in the placenta We collected two pieces of placental tissue (excluding decidua and amnion) in sterile conditions immediately after delivery. After cleaning with sterile saline, the placenta tissue was quickly frozen in a -180℃ liquid nitrogen tank and then transferred to the -80℃ refrigerator for storage. Added RIPA lysate and ground the tissues at low temperatures. The grinding solution was centrifuged at 12000 r/min for 20 min, the supernatant was collected, and the protein concentration was determined using the BCA method. Prepared a 12% separation gel and concentration gel, loaded 200 μg protein; the electrophoresis conditions were set to 80 V and 120 V. Membrane transfer was performed at 200 mA for 30 minutes. Blocking was done at room temperature for 1 h in blocking solution, followed by incubation with primary antibody (TNF-α antibody 1∶500 dilution, IL-1β antibody 1∶2000 dilution), overninght at 4℃ overnight, and then washed with TBST. The membrane was then incubated with secondary antibodies (diluted 1∶2000) for 2 hours at room temperature before being washed with TBST. Protein expression was calculated based on the gray value of the bands. Statistical analysis Image J was used to quantify the grayscale values in the analysis of Western Blot bands. For sample data analysis, one-way ANOVA was conducted using SPSS 22.0. Statistical plotting was performed with Graph Pad Prism 8, and the data results are presented as mean ± standard deviation (SD), considering a P<0.05 as statistically significant. Categorical data were reported in actual values or percentages, and intergroup differences (control group, mid-pregnancy infection group, late-pregnancy infection group) were analyzed using parametric and non-parametric descriptive statistics. The means were compared using the t-test, while the Wilcoxon rank-sum test was used for median comparison and the chi-square test for categorical variables. Fisher's exact test was applied for any variable with ≤5 cases. P -values were not adjusted for multiple comparisons. Results Pathological outcomes In a comparison between 33 cases in the infection group and 4 in the control group, as shown in Table 2 , 1 out of 33 patients in the infection group exhibited villous edema, with 1 case occurring in mid-pregnancy. This was not significantly different from the control group (0/4, P=0.486). Thrombosis and interstitial fibrin deposition were observed in 3 out of 33 patients during mid-pregnancy, showing no significant difference compared to the control group (0/4, P=0.12). Villous angioma was found in 3 out of 33 patients, with 1 in mid-pregnancy and 2 in late pregnancy, not significantly different from the control group (1/4, P=0.559). Villous space inflammation was present in 3 out of 33 patients during mid-pregnancy, with no significant difference compared to the control group (0/4, P=0.12). One out of 33 patients had chorioamnionitis in mid-pregnancy, not significantly different from the control group (0/4, P=0.486). Decidual vasculopathy was observed in 2 out of 33 patients, one in mid-pregnancy and one in late pregnancy, with no significant difference compared to the control group (1/4, P=0.469). Accelerated villous maturity was noted in 1 out of 33 patients during mid-pregnancy, not significantly different from the control group (0/4, P=0.486). Infarction was present in 2 out of 33 patients, one in mid-pregnancy and one in late pregnancy, with no significant difference compared to the control group (2/4, P=0.087). Meconium staining was observed in 2 out of 33 patients, both in mid-pregnancy, not significantly different from the control group (0/4, P=0.344). In the early pregnancy infection group, 2 cases showed necrosis and inflammation in decidual tissue. Table 2 Results of placental pathology were compared between the infection group and the control group Features Mid-pregnancy infection cases （ n=14 ） Late-pregnancy infection cases （ n=19 ） Control （ n=4 ） P value Villous edema 1 0 0 0.486 Thrombosis and interstitial fibrin deposition 3 0 0 0.12 Villous angioma 1 2 1 0.559 Villous space inflammation 3 0 0 0.12 Chorioamnionitis 1 0 0 0.486 Decidual vasculopathy 1 1 1 0.469 Accelerated villous maturity 1 0 0 0.486 Infarction 1 1 2 0.087 Infarction 2 0 0 0.344 Abbreviations: n, number of participants with available data. IL-6 content in umbilical cord blood and amniotic fluid As shown in Table 3 , the levels of IL-6 in the amniotic fluid were 248.33 ± 109.11 pg/ml and 260.45 ± 78.48 pg/ml in the mid-pregnancy and late-pregnancy infection groups, respectively, compared to 262.72 ± 148.83 pg/ml in the control group. As illustrated in Figure 1 , there were no significant differences in the IL-6 levels between the infection and control groups (P>0.05), nor were there significant differences between the infection groups themselves (P>0.05). IL-6 was not detected in either the infection or control groups in the umbilical cord blood. Table 3 IL-6 content the amniotic fluid (pg/ml, ±SD) Group Cases IL-6 Control 4 262.72±148.83 Mid-pregnancy infection 3 248.33±109.11 Late-pregnancy infection 3 260.45±78.48 P value ＞0.05 TNF-α and IL-1β protein expression in placental tissue Western blot was used to analyze the protein expression levels of TNF-α and IL-1β in placental tissues. Compared to the control group, the levels of TNF-α and IL-1β were significantly lower in the infection group (P0.05) ( Figure 2 ). Discussion This study focuses on the inflammatory response in fetal-related tissues following SARS-CoV-2 infection and investigates the impact of SARS-CoV-2 infection at different stages of pregnancy on maternal-fetal outcomes. By analyzing the pathological changes in the placental tissue, chorionic villi, and decidua of pregnant women infected with the virus during pregnancy, as well as the expression levels of related inflammatory factors, this study aims to provide new insights for the prevention and management of viral infections during pregnancy. SARS-CoV-2 is a pathogenic coronavirus that can affect multiple organs and systems, primarily targeting the respiratory system and causing symptoms ranging from the common cold to severe respiratory distress [7,14–16]. Pregnancy is a physiological state that significantly impacts a woman's body—the immune system changes during pregnancy to tolerate the fetus [17]. Although more studies have assessed the clinical outcomes of pregnant women diagnosed with COVID-19 and their infants, the causes of adverse outcomes remain unclear [18]. In addition to the virus's direct cytopathic effects, virus-mediated hyperinflammatory responses in the human body can cause tissue damage, forming a ''cytokine storm'' [19–22]. The results indicate that in early pregnancy, no unique histopathological features were observed in the decidua following SARS-CoV-2 infection. In mid and late pregnancy, however, the placental tissue showed characteristics such as villous edema, thrombosis and interstitial fibrin deposition, villous angiomas, villitis, chorioamnionitis, decidual vasculopathy, accelerated villous maturity, infarction, and meconium staining. Compared to normal placentae, SARS-CoV-2 infection in the placenta was associated with a higher incidence of decidual arteriopathy and other maternal malperfusion patterns, suggesting that infection could lead to placental inflammation and changes, resulting in long-term multisystem deficits in exposed infants [4,23]. Western blot results showed significantly increased levels of the inflammatory markers TNF-α and IL-1β in the placental tissues of the infection group, which could be associated with activation of the maternal immune system[24]. As critical inflammatory mediators, TNF-α and IL-1β indicate that viral infection may trigger an inflammatory response in placental tissues [25,26]. This inflammatory state may negatively impact the structure and function of the placenta, causing placental dysfunction and thereby affecting fetal growth and development [23]. Maternal SARS-CoV-2 infection may drive the immune activation of the mother, placenta, and fetus, potentially adversely affecting fetal neural development[9]. Although most existing data concern late pregnancy infections or active infections at the time of delivery, the impact of early pregnancy infections on offspring outcomes is crucial for a comprehensive understanding of the risks. Immune activation not only affects fully differentiated cells but can also impact progenitor cells derived from the yolk sac, suggesting that early damage (for example, during early pregnancy) may still affect brain development through microglial progenitor cells [27–29]. The timing of infection, viral strains, fetal sex, other prenatal exposures (such as maternal cardiometabolic diseases, substance use, stress, environmental or drug exposures, other infections during pregnancy), and perinatal/postnatal exposures (such as subsequent SARS-CoV-2 infections in infants or children, breastfeeding status) have a critical impact on offspring neural development and are essential for comprehensively understanding the potential lasting effects of the COVID-19 pandemic on future generations [30]. Viral infections during pregnancy can induce the host to produce inflammatory cytokines such as IL-1β, IL-6, and TNF-α, which not only activate the maternal immune system but can also cross the placental barrier [18,19]. As the placenta is located at the maternal-fetal interface, it not only plays a crucial role in protecting the fetus from infections but can also be influenced by adverse maternal environmental conditions. Infectious agents can cross the placenta, leading to fetal anomalies (e.g., microcephaly caused by Zika virus infection [31–34]); the placenta plays a vital role in maintaining fetal immune tolerance and forms a resistant barrier against pathogen entry. When the placental barrier is breached by infection, the risk of adverse outcomes for the fetus/newborn is high. Or, infectious factors can directly cause placental damage (e.g., villitis caused by cytomegalovirus [35]), leading to fetal growth restriction or death. In both scenarios, it is necessary to collect samples such as villi, decidua, placenta, amniotic fluid, and umbilical cord blood from patients at different stages of pregnancy post-COVID-19 infection to investigate their inflammatory responses and determine the mechanistic links between maternal infection and fetal outcomes. In summary, the pregnant patients in our study remained relatively stable post-infection, with no severe cases. The pregnancy outcomes were good for patients in mid to late pregnancy, with no severe neonatal complications. This study is significant for understanding the impact of viral infection on the health of pregnant women and fetuses, offering valuable insights for improving prevention and treatment strategies for infection-related pregnancy complications. Future research could further explore the molecular mechanisms of placental inflammatory responses caused by viral infection and prevention and treatment strategies to ensure the health of pregnant women and fetuses to the greatest extent. While our study provides critical insights, it also has limitations, including relatively small sample size and the possibility that variations in viral subtypes may affect the consistency of results. Future research needs to address these limitations with larger samples and more in-depth exploration. Further investigations could explore the molecular mechanisms of the inflammatory responses induced by COVID-19 infection during pregnancy and how these inflammatory factors impact placental function and fetal development. Additionally, broader population-based studies and long-term follow-ups will help to understand maternal-fetal outcomes more comprehensively. Abbreviations WHO World Health Organization COVID-19 Coronavirus disease 2019 SARS-CoV-2 Syndrome coronavirus 2 Declarations Acknowledgements The authors thank all the trial participants for making this work possible. Author Contributions MY, LP, CZ, XZ, MH, YW, YH, ZG, HL and LZ acquired the data and contributed to the revision of the final manuscript. Authors HL and LZ designed the study. MY and LP performed the analysis and wrote the first draft of the manuscript. CZ, XZ, MH, YW, YH, ZG interpreted the data. All authors contributed to the intellectual content of the manuscript and approved the manuscript version submitted for publication. Funding There are none funding during the submission process. Ethics approval and consent to participate The study was approved by the the Ethics Committee of the Affiliated Taian City Central Hospital of Qingdao University (accession number MR-37-23-050564) and is in compliance with the principles of the Declaration of Helsinki. Informed consent was obtained from every participant. Consent for publication Consent for publication was obtained from every participant included in the study. Competing interests The authors declare that they have no competing interests. Availability of data and materials The authors confirm that the data supporting the findings of this study are available within the article. References Zhou F, Yu T, Du R, et al. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board 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-4488503","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":312825344,"identity":"d9ae6d5b-0023-4b2f-bf78-3b0292eb8587","order_by":0,"name":"Mengyue Yin","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Mengyue","middleName":"","lastName":"Yin","suffix":""},{"id":312825345,"identity":"df1358f3-df9d-46a0-9048-bbecbfab830b","order_by":1,"name":"Lingling Peng","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Lingling","middleName":"","lastName":"Peng","suffix":""},{"id":312825349,"identity":"5221a809-c3e5-47d0-a5e9-5d7f167e76c9","order_by":2,"name":"Chunling Zhang","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Chunling","middleName":"","lastName":"Zhang","suffix":""},{"id":312825350,"identity":"2340e037-0495-4415-97e4-4306fca1e627","order_by":3,"name":"Xueyan Zhang","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Xueyan","middleName":"","lastName":"Zhang","suffix":""},{"id":312825351,"identity":"ae04cd14-1f45-451d-9ad7-b558d429bdb3","order_by":4,"name":"Mei Han","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Mei","middleName":"","lastName":"Han","suffix":""},{"id":312825352,"identity":"1086aeff-c831-4d55-8780-325c0ba6ae00","order_by":5,"name":"Yongmei Wang","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Yongmei","middleName":"","lastName":"Wang","suffix":""},{"id":312825353,"identity":"d8420cd1-e104-45ba-bbca-aa7521282f76","order_by":6,"name":"Yafei Hao","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Yafei","middleName":"","lastName":"Hao","suffix":""},{"id":312825354,"identity":"9ca11a7f-1dfd-40f2-94eb-70eef30ea018","order_by":7,"name":"Zhaolun Guo","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Zhaolun","middleName":"","lastName":"Guo","suffix":""},{"id":312825355,"identity":"978db862-0da0-4c60-8390-4f11a7da910f","order_by":8,"name":"Hua Li","email":"","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":false,"prefix":"","firstName":"Hua","middleName":"","lastName":"Li","suffix":""},{"id":312825356,"identity":"5808f23c-bc8a-4cf1-b3fd-adddbf9520e2","order_by":9,"name":"Lan Zhang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAxklEQVRIiWNgGAWjYDACdiBOMPgnx8/MfPgBcVqYgfhBxQFjyXa2NAOitTA+OHMg0eA8j4IEUTrkm5mfPUhsu5NgfJiHwYChxiaaoBbGZjZzg8S2Z3lmh3kPPGA4lpbbQNBdzAxmEoltzMVmh/kSDBgbDhPWwsbM/g2kJXFzM4+BBFFaeJh5zCQSzhxO3MBMrBYJZp4yiYSKNGOJw8BATiDGL/Lt7dskfxjYyPH3Hz784EONDWEtqCCBNOWjYBSMglEwCnABAO4nOuMhoU3RAAAAAElFTkSuQmCC","orcid":"","institution":"The Affiliated Taian City Central Hospital of Qingdao University","correspondingAuthor":true,"prefix":"","firstName":"Lan","middleName":"","lastName":"Zhang","suffix":""}],"badges":[],"createdAt":"2024-05-28 06:21:29","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4488503/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4488503/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":58315823,"identity":"a5ea3f5d-a472-496e-937b-6d8d2585e210","added_by":"auto","created_at":"2024-06-13 20:59:52","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":26370,"visible":true,"origin":"","legend":"\u003cp\u003eIL-6 Levels in Amniotic Fluid. The IL-6 content in the amniotic fluid showed no significant differences between the infection and control groups (P\u0026gt;0.05). Additionally, there were no significant differences between the different infection groups themselves (P\u0026gt;0.05). Notably, IL-6 was undetectable in the umbilical cord blood for both the infection and control groups.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-4488503/v1/870bc5dbc259cbeef3f327c9.png"},{"id":58315824,"identity":"5d519777-a246-4914-9a3d-2174cfc90a23","added_by":"auto","created_at":"2024-06-13 20:59:52","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":70414,"visible":true,"origin":"","legend":"\u003cp\u003eProtein Expression Levels of TNF-α and IL-1β in Placental Tissues. Western blot analysis of placental tissues showed that the protein expression levels of TNF-α and IL-1β were significantly lower in the infection group compared to the control group (P\u0026lt;0.05). No statistically significant differences were observed between the mid-pregnancy and late-pregnancy infection groups (P\u0026gt;0.05).\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-4488503/v1/c718be3dc791f59beb5046ef.png"},{"id":72609323,"identity":"94c14272-25fe-4c69-829f-82741fe0f6b1","added_by":"auto","created_at":"2024-12-30 10:09:03","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":578021,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4488503/v1/282d40cd-db37-4abb-ac64-2b2b279e49e5.pdf"},{"id":58315826,"identity":"9b3904bf-21a4-48ea-957a-5950ec939854","added_by":"auto","created_at":"2024-06-13 20:59:53","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":1480321,"visible":true,"origin":"","legend":"","description":"","filename":"supplementaryfile1.docx","url":"https://assets-eu.researchsquare.com/files/rs-4488503/v1/d73ea495cfd9d8e3eb77b64f.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Impact of SARS-CoV-2 Infection During Various Pregnancy Trimesters on Maternal and Fetal Outcomes","fulltext":[{"header":"Background","content":"\u003cp\u003eOn March 11, 2020, the World Health Organization (WHO) declared a global pandemic due to coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. SARS-CoV-2 is an RNA virus enveloped with a nucleocapsid protein, primarily transmitted through respiratory droplets and close contact, exhibiting strong infectivity. Pregnant women are considered susceptible to the virus [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. To maintain the high metabolic demands required for normal fetal development during pregnancy, the burden of oxidative stress in pregnant women increases significantly. Due to alterations in the physiological and immunosuppressive states during pregnancy, clinical outcomes of SARS-CoV-2 infection in pregnant women may differ from the general population. Diagnosis and treatment of SARS-CoV-2 infection during pregnancy may increase the health risks of both the mother and the newborn [\u003cspan additionalcitationids=\"CR5\" citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe long COVID effect, known as \"post-acute sequelae of SARS-CoV-2 infection\" or PASC, leads to weakness and affects multiple organ systems [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Research indicates that SARS-CoV-2 can impact the reproductive system, manifesting as a decline in ovarian reserve and reproductive endocrinological imbalances in COVID-19 patients, affecting the production of ovarian hormones and/or the response of the uterine lining [\u003cspan additionalcitationids=\"CR9\" citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Additionally, babies born to women who had COVID-19 during pregnancy are more likely to undergo neurodevelopmental diagnosis within the first year after birth, with the extent and pathology of the impact still requiring further research [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Understanding the impact of SARS-CoV-2 infection at different stages of pregnancy on maternal and infant outcomes, as well as the long-term effects of infection during pregnancy, provides a reference for the management of newborns after maternal SARS-CoV-2 infection [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe research employs H\u0026amp;E staining for histological examination, intended to observe pathological alterations in the placenta. Western Blot analysis was used to detect the expression levels of TNF-α and IL-1β in placental tissues. IL-6 levels in umbilical cord blood and amniotic fluid were determined using the ELISA technique. Analyzing the impact of SARS-CoV-2 infection on placental immune response will provide valuable information for developing effective management and prevention strategies to reduce adverse outcomes of SARS-CoV-2 infection during pregnancy [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e].\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003e\u003cstrong\u003eParticipants\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe admitted 57 pregnant women from the Obstetrics Department of Tai\u0026apos;an Central Hospital as the study population from November 2022 to March 2023. All pregnant women were singleton, and relevant examinations excluded primary diseases and other diseases. The patient group with infections totaled 53 cases, divided based on SARS-CoV-2 infection status and timing into early pregnancy infection group (1st to 12th week of pregnancy, n=20), mid-pregnancy group (13th to 28th week of pregnancy, n=14) and late-pregnancy infection group (29th week of pregnancy to delivery, n=19). There were 4 cases in the non-infection group, serving as the control group. According to the \u0026quot;Diagnosis and Treatment Protocol for Novel Coronavirus Pneumonia (Trial version 6),\u0026quot; all patients were categorized as ordinary cases. All the selected pregnant women gave informed consent and signed the consent form. There were no statistical differences in age and parity among the groups of pregnant women (P \u0026gt; 0.05) (\u003cstrong\u003eTable 1\u003c/strong\u003e).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1\u0026nbsp;\u003c/strong\u003eParticipant characteristics\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" align=\"\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.02112676056338%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.37323943661972%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eEarly pregnancy infection (n=20)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.781690140845072%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eMid-pregnancy infection (n=14)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.95774647887324%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eLate-pregnancy infection (n=19)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.197183098591548%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(n=4)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.669014084507042%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003evalue\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.02112676056338%\" valign=\"top\"\u003e\n \u003cp\u003eAge (years)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.37323943661972%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.781690140845072%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.95774647887324%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.197183098591548%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.669014084507042%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.02112676056338%\" valign=\"top\"\u003e\n \u003cp\u003eMean (SD)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.37323943661972%\" valign=\"top\"\u003e\n \u003cp\u003e31.8 (5.9)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.781690140845072%\" valign=\"top\"\u003e\n \u003cp\u003e28.4 (3.6)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.95774647887324%\" valign=\"top\"\u003e\n \u003cp\u003e31.0 (5.0)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.197183098591548%\" valign=\"top\"\u003e\n \u003cp\u003e28.0 (4.3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.669014084507042%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026gt;0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.02112676056338%\" valign=\"top\"\u003e\n \u003cp\u003eParity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.37323943661972%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.781690140845072%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.95774647887324%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.197183098591548%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.669014084507042%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.02112676056338%\" valign=\"top\"\u003e\n \u003cp\u003eMean (SD)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.37323943661972%\" valign=\"top\"\u003e\n \u003cp\u003eN/A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.781690140845072%\" valign=\"top\"\u003e\n \u003cp\u003e1.3 (0.5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.95774647887324%\" valign=\"top\"\u003e\n \u003cp\u003e1.7 (0.8)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.197183098591548%\" valign=\"top\"\u003e\n \u003cp\u003e1.3 (0.5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.669014084507042%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026gt;0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eAbbreviations: n, number of participants with available data; SD, standard deviation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eInstruments and reagents\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAnti-TNF-\u0026alpha; antibody, recombinant rabbit monoclonal antibody against IL-1\u0026beta;, horseradish peroxidase-conjugated sheep anti-rabbit secondary antibody, and \u0026beta;-Actin antibody were purchased from Beijing Boaosen Biotechnology Co., LTD. Pre-stained protein markers and pre-mixed glue kits were purchased from Shanghai Biyantian Biotechnology Co., LTD. RIPA tissue lysate and protein loading buffer were purchased from Beijing Pulley Gene Technology Co., LTD. The human IL-6 ELISA test kit was purchased from Wuhan Doctor De Bioengineering Co., LTD. CKX53 inverted phase contrast microscope (OLYMPUS), microplate reader (Thermo Fisher Scientific).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePathological\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe chorionic villi and decidua tissues of patients with induced abortion and placenta tissues of pregnant women after cesarean section were collected after infection in early pregnancy, washed with sterile saline, and fixed in 4% paraformaldehyde for 48 hours. The slides were routinely embedded in paraffin and HE stained, and each slide was 4 \u0026mu;m thick. All HE slides were observed and reviewed by an attending physician and an associate chief physician.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eIL-6 levels in cord blood serum and amniotic fluid\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBefore the rupture of membranes during the cesarean section, amniotic fluid was extracted with a syringe and centrifuged at 3000 r/min for 15 min. After separation, the supernatant was collected in EP tubes and stored in a freezer at -80℃. At delivery, 5 mL of umbilical cord blood was collected into blood collection tubes and centrifuged at 3000 r/min for 15 min. Serum was divided into 300 \u0026mu;L frozen tubes and stored in the refrigerator at -80\u0026nbsp;℃. According to the kit instructions, perform the assay and calculate the corresponding concentration based on the standard curve according to the OD value of the sample.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTNF-\u0026alpha; and IL-1\u0026beta; levels in the placenta\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe collected two pieces of placental tissue (excluding decidua and amnion) in sterile conditions immediately after delivery. After cleaning with sterile saline, the placenta tissue was quickly frozen in a -180℃\u0026nbsp;liquid nitrogen tank and then transferred to the -80℃\u0026nbsp;refrigerator for storage. Added RIPA lysate and ground the tissues at low temperatures. The grinding solution was centrifuged at 12000 r/min for 20 min, the supernatant was collected, and the protein concentration was determined using the BCA method. Prepared a 12% separation gel and concentration gel, loaded 200 \u0026mu;g protein; the electrophoresis conditions were set to 80 V and 120 V. Membrane transfer was performed at 200 mA for 30 minutes. Blocking was done at room temperature for 1 h in blocking solution, followed by incubation with primary antibody (TNF-\u0026alpha; antibody 1∶500 dilution, IL-1\u0026beta; antibody 1∶2000 dilution), overninght at 4℃\u0026nbsp;overnight, and then washed with TBST. The membrane was then incubated with secondary antibodies (diluted 1∶2000) for 2 hours at room temperature before being washed with TBST.\u0026nbsp;Protein expression was calculated based on the gray value of the bands.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStatistical analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eImage J was used to quantify the grayscale values in the analysis of Western Blot bands. For sample data analysis, one-way ANOVA was conducted using SPSS 22.0. Statistical plotting was performed with Graph Pad Prism 8, and the data results are presented as mean \u0026plusmn; standard deviation (SD), considering a P\u0026lt;0.05 as statistically significant. Categorical data were reported in actual values or percentages, and intergroup differences (control group, mid-pregnancy infection group, late-pregnancy infection group) were analyzed using parametric and non-parametric descriptive statistics. The means were compared using the t-test, while the Wilcoxon rank-sum test was used for median comparison and the chi-square test for categorical variables. Fisher\u0026apos;s exact test was applied for any variable with \u0026le;5 cases. \u003cem\u003eP\u003c/em\u003e-values were not adjusted for multiple comparisons.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003ePathological outcomes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn a comparison between 33 cases in the infection group and 4 in the control group, as shown in \u003cstrong\u003eTable 2\u003c/strong\u003e, 1 out of 33 patients in the infection group exhibited\u0026nbsp;villous edema, with 1 case occurring in mid-pregnancy. This was not significantly different from the control group (0/4, P=0.486). Thrombosis and interstitial fibrin deposition were observed in 3 out of 33 patients during mid-pregnancy, showing no significant difference compared to the control group (0/4, P=0.12). Villous angioma was found in 3 out of 33 patients, with 1 in mid-pregnancy and 2 in late pregnancy, not significantly different from the control group (1/4, P=0.559). Villous space inflammation was present in 3 out of 33 patients during mid-pregnancy, with no significant difference compared to the control group (0/4, P=0.12). One out of 33 patients had chorioamnionitis in mid-pregnancy, not significantly different from the control group (0/4, P=0.486). Decidual vasculopathy was observed in 2 out of 33 patients, one in mid-pregnancy and one in late pregnancy, with no significant difference compared to the control group (1/4, P=0.469). Accelerated villous maturity was noted in 1 out of 33 patients during mid-pregnancy, not significantly different from the control group (0/4, P=0.486). Infarction was present in 2 out of 33 patients, one in mid-pregnancy and one in late pregnancy, with no significant difference compared to the control group (2/4, P=0.087). Meconium staining was observed in 2 out of 33 patients, both in mid-pregnancy, not significantly different from the control group (0/4, P=0.344). In the early pregnancy infection group, 2 cases showed necrosis and inflammation in decidual tissue.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2\u0026nbsp;\u003c/strong\u003eResults of placental pathology were compared between the infection group and the control group\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"100%\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eFeatures\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eMid-pregnancy infection cases\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e（\u003c/strong\u003e\u003cstrong\u003en=14\u003c/strong\u003e\u003cstrong\u003e）\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eLate-pregnancy infection cases\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e（\u003c/strong\u003e\u003cstrong\u003en=19\u003c/strong\u003e\u003cstrong\u003e）\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e（\u003c/strong\u003e\u003cstrong\u003en=4\u003c/strong\u003e\u003cstrong\u003e）\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eP\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003evalue\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eVillous edema\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.486\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eThrombosis and interstitial fibrin deposition\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.12\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eVillous angioma\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.559\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eVillous space inflammation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.12\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eChorioamnionitis\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.486\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eDecidual vasculopathy\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.469\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eAccelerated villous maturity\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.486\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eInfarction\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.087\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"27.551020408163264%\"\u003e\n \u003cp\u003eInfarction\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.448979591836736%\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.224489795918368%\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.26530612244898%\"\u003e\n \u003cp\u003e0.344\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eAbbreviations: n, number of participants with available data.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eIL-6 content in umbilical cord blood and amniotic fluid\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAs shown in \u003cstrong\u003eTable 3\u003c/strong\u003e, the levels of IL-6 in the amniotic fluid were 248.33 \u0026plusmn; 109.11 pg/ml and 260.45 \u0026plusmn; 78.48 pg/ml in the mid-pregnancy and late-pregnancy infection groups, respectively, compared to 262.72 \u0026plusmn; 148.83 pg/ml in the control group. As illustrated in \u003cstrong\u003eFigure 1\u003c/strong\u003e, there were no significant differences in the IL-6 levels between the infection and control groups (P\u0026gt;0.05), nor were there significant differences between the infection groups themselves (P\u0026gt;0.05). IL-6 was not detected in either the infection or control groups in the umbilical cord blood.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3\u0026nbsp;\u003c/strong\u003eIL-6 content the amniotic fluid (pg/ml,\u0026nbsp;\u0026plusmn;SD)\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"42.1978021978022%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroup\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"24.395604395604394%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eCases\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.40659340659341%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eIL-6\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"42.1978021978022%\"\u003e\n \u003cp\u003eControl\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"24.395604395604394%\" valign=\"top\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.40659340659341%\" valign=\"top\"\u003e\n \u003cp\u003e262.72\u0026plusmn;148.83\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"42.1978021978022%\"\u003e\n \u003cp\u003eMid-pregnancy infection\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"24.395604395604394%\" valign=\"top\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.40659340659341%\" valign=\"top\"\u003e\n \u003cp\u003e248.33\u0026plusmn;109.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"42.1978021978022%\"\u003e\n \u003cp\u003eLate-pregnancy infection\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"24.395604395604394%\" valign=\"top\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.40659340659341%\" valign=\"top\"\u003e\n \u003cp\u003e260.45\u0026plusmn;78.48\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"42.1978021978022%\"\u003e\n \u003cp\u003e\u003cem\u003eP\u0026nbsp;\u003c/em\u003evalue\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"24.395604395604394%\" valign=\"top\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.40659340659341%\" valign=\"top\"\u003e\n \u003cp\u003e＞0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eTNF-\u0026alpha; and IL-1\u0026beta; protein expression in placental tissue\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWestern blot was used to analyze the protein expression levels of TNF-\u0026alpha; and IL-1\u0026beta; in placental tissues. Compared to the control group, the levels of TNF-\u0026alpha; and IL-1\u0026beta; were significantly lower in the infection group (P\u0026lt;0.05). There was no statistically significant difference between the mid-pregnancy and late-pregnancy infection groups (P\u0026gt;0.05) (\u003cstrong\u003eFigure 2\u003c/strong\u003e).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis study focuses on the inflammatory response in fetal-related tissues following SARS-CoV-2 infection and investigates the impact of SARS-CoV-2 infection at different stages of pregnancy on maternal-fetal outcomes. By analyzing the pathological changes in the placental tissue, chorionic villi, and decidua of pregnant women infected with the virus during pregnancy, as well as the expression levels of related inflammatory factors, this study aims to provide new insights for the prevention and management of viral infections during pregnancy.\u003c/p\u003e\n\u003cp\u003eSARS-CoV-2 is a pathogenic coronavirus that can affect multiple organs and systems, primarily targeting the respiratory system and causing symptoms ranging from the common cold to severe respiratory distress [7,14\u0026ndash;16]. Pregnancy is a physiological state that significantly impacts a woman\u0026apos;s body\u0026mdash;the immune system changes during pregnancy to tolerate the fetus [17]. Although more studies have assessed the clinical outcomes of pregnant women diagnosed with COVID-19 and their infants, the causes of adverse outcomes remain unclear [18]. In addition to the virus\u0026apos;s direct cytopathic effects, virus-mediated hyperinflammatory responses in the human body can cause tissue damage, forming a \u0026apos;\u0026apos;cytokine storm\u0026apos;\u0026apos; [19\u0026ndash;22].\u003c/p\u003e\n\u003cp\u003eThe results indicate that in early pregnancy, no unique histopathological features were observed in the decidua following SARS-CoV-2 infection. In mid and late pregnancy, however, the placental tissue showed characteristics such as villous edema, thrombosis and interstitial fibrin deposition, villous angiomas, villitis, chorioamnionitis, decidual vasculopathy, accelerated villous maturity, infarction, and meconium staining. Compared to normal placentae, SARS-CoV-2 infection in the placenta was associated with a higher incidence of decidual arteriopathy and other maternal malperfusion patterns, suggesting that infection could lead to placental inflammation and changes, resulting in long-term multisystem deficits in exposed infants [4,23]. Western blot results showed significantly increased levels of the inflammatory markers TNF-\u0026alpha; and IL-1\u0026beta; in the placental tissues of the infection group, which could be associated with activation of the maternal immune system[24]. As critical inflammatory mediators, TNF-\u0026alpha; and IL-1\u0026beta; indicate that viral infection may trigger an inflammatory response in placental tissues\u0026nbsp;[25,26]. This inflammatory state may negatively impact the structure and function of the placenta, causing placental dysfunction and thereby affecting fetal growth and development [23].\u0026nbsp;Maternal SARS-CoV-2 infection may drive the immune activation of the mother, placenta, and fetus, potentially adversely affecting fetal neural development[9]. Although most existing data concern late pregnancy infections or active infections at the time of delivery, the impact of early pregnancy infections on offspring outcomes is crucial for a comprehensive understanding of the risks.\u003c/p\u003e\n\u003cp\u003eImmune activation not only affects fully differentiated cells but can also impact progenitor cells derived from the yolk sac, suggesting that early damage (for example, during early pregnancy) may still affect brain development through microglial progenitor cells [27\u0026ndash;29]. The timing of infection, viral strains, fetal sex, other prenatal exposures (such as maternal cardiometabolic diseases, substance use, stress, environmental or drug exposures, other infections during pregnancy), and perinatal/postnatal exposures (such as subsequent SARS-CoV-2 infections in infants or children, breastfeeding status) have a critical impact on offspring neural development and are essential for comprehensively understanding the potential lasting effects of the COVID-19 pandemic on future generations\u0026nbsp;[30].\u003c/p\u003e\n\u003cp\u003eViral infections during pregnancy can induce the host to produce inflammatory cytokines such as IL-1\u0026beta;, IL-6, and TNF-\u0026alpha;, which not only activate the maternal immune system but can also cross the placental barrier [18,19].\u003c/p\u003e\n\u003cp\u003eAs the placenta is located at the maternal-fetal interface, it not only plays a crucial role in protecting the fetus from infections but can also be influenced by adverse maternal environmental conditions. Infectious agents can cross the placenta, leading to fetal anomalies (e.g., microcephaly caused by Zika virus infection [31\u0026ndash;34]); the placenta plays a vital role in maintaining fetal immune tolerance and forms a resistant barrier against pathogen entry. When the placental barrier is breached by infection, the risk of adverse outcomes for the fetus/newborn is high. Or, infectious factors can directly cause placental damage (e.g., villitis caused by cytomegalovirus [35]), leading to fetal growth restriction or death. In both scenarios, it is necessary to collect samples such as villi, decidua, placenta, amniotic fluid, and umbilical cord blood from patients at different stages of pregnancy post-COVID-19 infection to investigate their inflammatory responses and determine the mechanistic links between maternal infection and fetal outcomes.\u003c/p\u003e\n\u003cp\u003eIn summary, the pregnant patients in our study remained relatively stable post-infection, with no severe cases. The pregnancy outcomes were good for patients in mid to late pregnancy, with no severe neonatal complications. This study is significant for understanding the impact of viral infection on the health of pregnant women and fetuses, offering valuable insights for improving prevention and treatment strategies for infection-related pregnancy complications. Future research could further explore the molecular mechanisms of placental inflammatory responses caused by viral infection and prevention and treatment strategies to ensure the health of pregnant women and fetuses to the greatest extent. While our study provides critical insights, it also has limitations, including relatively small sample size and the possibility that variations in viral subtypes may affect the consistency of results. Future research needs to address these limitations with larger samples and more in-depth exploration. Further investigations could explore the molecular mechanisms of the inflammatory responses induced by COVID-19 infection during pregnancy and how these inflammatory factors impact placental function and fetal development. Additionally, broader population-based studies and long-term follow-ups will help to understand maternal-fetal outcomes more comprehensively.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eWHO \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;World Health Organization\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eCOVID-19 \u0026nbsp; \u0026nbsp;Coronavirus disease 2019\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eSARS-CoV-2 \u0026nbsp;Syndrome coronavirus 2\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors thank all the trial participants for making this work possible.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMY, LP, CZ, XZ, MH,\u0026nbsp;YW, YH, ZG, HL and LZ acquired the data and contributed to the revision of the final manuscript. Authors HL and LZ designed the study. MY and LP performed the analysis and wrote the first draft of the manuscript. CZ, XZ, MH, YW, YH, ZG interpreted the data. All authors contributed to the intellectual content of the manuscript and approved the manuscript version submitted for publication.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThere are none funding during the submission process.\u003c/p\u003e\n\u003cp\u003eEthics approval and consent to participate\u003c/p\u003e\n\u003cp\u003eThe study was approved by the the Ethics Committee of the Affiliated Taian City Central Hospital of Qingdao University (accession number MR-37-23-050564) and is in compliance with the principles of the Declaration of Helsinki. Informed consent was obtained from every participant.\u003c/p\u003e\n\u003cp\u003eConsent for publication\u003c/p\u003e\n\u003cp\u003eConsent for publication was obtained from every participant included in the study.\u003c/p\u003e\n\u003cp\u003eCompeting interests\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors confirm that the data supporting the findings of this study are available within the article.\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eZhou F, Yu T, Du R, et al. Clinical course and risk factors for mortality of adult inpatients with COVID-19 in Wuhan, China: a retrospective cohort study. The Lancet. 2020; 395:1054-1062.\u003c/li\u003e\n\u003cli\u003eGorbalenya AE, Baker SC, Baric RS, et al. 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Accessed 21 March 2024.\u003c/li\u003e\n\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":"SARS-CoV-2, Pregnancy, Inflammatory reaction","lastPublishedDoi":"10.21203/rs.3.rs-4488503/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4488503/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground \u003c/strong\u003eThe specific impact and pathology of long COVID effects on mother and fetus post-infection have not been deeply investigated. Therefore, we explored the inflammatory response in fetal tissue from pregnant women infected at various stages.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods \u003c/strong\u003eWe collected villi, decidua, and placenta samples from 57 patients at Tai'an Central Hospital, who underwent either induced abortion or delivery between November 2022 and March 2023. HE staining was employed for histological examinations of these tissues. Furthermore, the concentrations of IL-6 in both umbilical cord blood and amniotic fluid were quantitatively analyzed using ELISA kits. Additionally, Western blot analysis was conducted to evaluate the expression levels of TNF-α and IL-1β in the placental tissues.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003cem\u003e\u003cstrong\u003e \u003c/strong\u003e\u003c/em\u003eAmong participants, 53 pregnant women were diagnosed with SARS-CoV-2 infection. No significant placental pathology differences were found between mid-term and late-term stages with the control group (P\u0026gt;0.05). However, Western blot analysis indicated that infected placental tissues showed higher TNF-α and IL-1β levels (P\u0026lt;0.05), with no notable variance in expression levels between mid-term and late-term pregnancies (P\u0026gt;0.05). IL-6 levels in amniotic fluid showed no significant difference, and detected no IL-6 expression in umbilical cord blood.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion \u003c/strong\u003eInfection with SARS-CoV-2 during different periods of pregnancy can have different effects on the placenta and other tissues, emphasizing the importance of preventing and managing viral infection during pregnancy and providing clinical reference for formulating management strategies for SARS-CoV-2 infection.\u003c/p\u003e","manuscriptTitle":"Impact of SARS-CoV-2 Infection During Various Pregnancy Trimesters on Maternal and Fetal Outcomes","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-06-13 20:59:48","doi":"10.21203/rs.3.rs-4488503/v1","editorialEvents":[{"type":"communityComments","content":1}],"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":"94e22a56-8bbd-4074-89a6-096a84a86fa7","owner":[],"postedDate":"June 13th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-12-30T10:08:15+00:00","versionOfRecord":[],"versionCreatedAt":"2024-06-13 20:59:48","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4488503","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4488503","identity":"rs-4488503","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}