Gene Expression
Nebulas
Gene Expression NebulasSummary: Pregnant women appear to be at increased risk for severe outcomes associated with COVID-19, but the pathophysiology underlying this increased morbidity and its potential impact on the developing fetus is not well understood. In this study of pregnant women with and without COVID-19, we assessed viral and immune dynamics at the placenta during maternal SARS-CoV-2 infection. Amongst uninfected women, ACE2 was detected by immunohistochemistry in syncytiotrophoblast cells of the normal placenta during early pregnancy but was rarely seen in healthy placentas at full term. Term placentas from women infected with SARS-CoV-2, however, displayed a significant increase in ACE2 levels. Using immortalized cell lines and primary isolated placental cells, we determined the vulnerability of various placental cell types to direct infection by SARS-CoV-2 in vitro. Yet, despite the susceptibility of placental cells to SARS-CoV-2 infection, viral RNA was detected in the placentas of only a subset (∼13%) of women in this cohort. Through single cell transcriptomic analyses, we found that the maternal-fetal interface of SARS-CoV-2-infected women exhibited markers associated with pregnancy complications, such as preeclampsia, and robust immune responses, including increased activation of placental NK and T cells and increased expression of interferon-related genes. Overall, this study suggests that SARS-CoV-2 is associated with immune activation at the maternal-fetal interface even in the absence of detectable local viral invasion. While this likely represents a protective mechanism shielding the placenta from infection, inflammatory changes in the placenta may also contribute to poor pregnancy outcomes and thus warrant further investigation.
Overall Design: Analysis of single-cell RNA-seq data from decidua and placental villi of control and SARS-CoV-2-positive samples.
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| Treatment Protocol: | - |
| Extract Protocol: | Dissected tissues were rinsed thoroughly in PBS and minced with scissors in a tissue digestion buffer of Liberase TM (0.28 WU/ml) and DNase I (30 μg/ml) in HBSS with Ca2+ and Mg2+. Finely minced tissue was enzymatically digested at 37°C for 1 hour with agitation, pipetting, and further mincing every 10 minutes until disaggregated. The suspension was passed through sterile gauze, centrifuged at 1000 x g for 5 minutes at 4°C to pellet cells, and washed with fresh digestion buffer. After centrifugation, the supernatant was aspirated and the cell pellet was resuspended in ACK lysing buffer for 5 minutes. Cells were centrifuged and resuspended in RPMI media before filtering through a 70-μm mesh cell strainer. |
| Library Construction Protocol: | scRNA-seq libraries were generated using the 10x Chromium Single Cell 3’ Reagent Kit. |
| Molecule Type: | polyA(+) RNA |
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| Library Layout: | PAIRED |
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| Platform: | ILLUMINA |
| Instrument Model: | Illumina NovaSeq 6000 |
| Strand-Specific: | - |
| Data Resource | GEN Sample ID | GEN Dataset ID | Project ID | BioProject ID | Sample ID | Sample Name | BioSample ID | Sample Accession | Experiment Accession | Release Date | Submission Date | Update Date | Species | Race | Ethnicity | Age | Age Unit | Gender | Source Name | Tissue | Cell Type | Cell Subtype | Cell Line | Disease | Disease State | Development Stage | Mutation | Phenotype | Case Detail | Control Detail | Growth Protocol | Treatment Protocol | Extract Protocol | Library Construction Protocol | Molecule Type | Library Layout | Strand-Specific | Library Strand | Spike-In | Strategy | Platform | Instrument Model | Cell Number | Reads Number | Gbases | AvgSpotLen1 | AvgSpotLen2 | Uniq Mapping Rate | Multiple Mapping Rate | Coverage Rate |
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