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Differential gene expression during placentation in pregnancies conceived with different fertility treatments compared with spontaneous pregnancies

ABSTRACT: This SMAART cohort study uses bulk total RNA-sequencing to assses the impact of mode of conception on first trimester human placenta gene expression [PMID: 30611556]. Pregnancies were singleton, gestational age 10-14 weeks, normal karyotype, fetal race Caucasian or biracial Caucasian/Asian, and conceived either with fertility treatments (IVF=in vitro fertilization, NIFT=non-IVF fertility treatment) or without fertility treatments (spontaneous, also called unassisted). Pregnancies were balanced for fetal sex. Chorionic villi tissue leftover after clinical genetic testing was collected for research with informed consent and stored in RNAlater RNA Stabilization Reagent (QIAGEN) at -80C until RNA isolation with the AllPrep DNA/RNA Mini Kit (QIAGEN). RNA-seq libraries of >200 nt were constructed with Illumina TruSeq Stranded Total RNA with Ribo-Zero Gold sample prep kits (Illumina) and depleted of cytoplasmic and mitochondrial ribosomal RNAs. After quality check with FastQC, transcript abundances were quantified against the human reference genome (Ensembl build GRCh38) using Kallisto. Differential expression analysis with DESeq2 was performed to compare mode of conception, adjusted for fetal sex and RNA-seq dataset. The dataset adjustment corrected for batch effects from RNA isolation and/or sequencing runs. There were N=141 subjects total, including 74 spontaneous, 33 non-IVF (NIFT), and 34 IVF pregnancies. The subject columns at the end of the DESeq2 files are counts normalized for sequencing depth.

ORGANISM(S): Homo sapiens

PROVIDER: GSE215421 | GEO | 2022/10/18

REPOSITORIES: GEO

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Project description:Human placenta bulk mRNA-sequencing from n=124 first trimester (59 female, 65 male) and n=43 third trimester (18 female, 25 male) tissue samples. A subset of 23 pregnancies has matched placenta tissue collected at both first and third trimester. Tissue was collected at Cedars-Sinai Medical Center in Los Angeles, California, USA. First trimester placenta was collected at gestational ages of 70-100 days from leftover tissue from chorionic villus sampling for prenatal genetic diagnosis, after cleaning to remove maternal decidua. Third trimester placenta was collected after delivery at gestational ages 254-290 days from the fetal side near the umbilical cord insertion site beneath the amnion. Tissue was stored in RNAlater RNA Stabilization Solution (Invitrogen) at -80C until further processing. All pregnancies were conceived without fertility treatments, were normal karyotype, and resulted in live singleton births. Mothers with pre-existing diabetes or hypertension were excluded from the study, but not excluded if complications developed during pregnancy, though most pregnancies were uncomplicated. The average parental age was advanced (over 35 years old) but principal components analysis did not show clustering by either maternal or paternal age. RNA extraction was performed with physical homogenization of tissue followed by the AllPrep DNA/RNA/miRNA Universal Kit (QIAGEN), then 1 ug of the total RNA elution was used for library construction using the Illumina TruSeq Stranded mRNA library preparation kit (Illumina), with polyA mRNA selection then cDNA synthesis using SuperScript II reverse transcriptase (Invitrogen) and random primers. The cDNA was converted into double stranded DNA and PCR-amplified, then purified with Agencourt AMPure XP beads (Beckman Coulter). Sample libraries were multiplexed and sequenced on a NovaSeq 6000 platform (Illumina) using 75bp single-end mRNA-sequencing, with average 30 million reads per sample. Differential expression analysis was performed with DEseq2 to compare first versus third trimester placenta, adjusted for fetal sex [PubMed ID: 38271627]. Subanalyses were also performed to identify sex differences.

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Differential gene expression during placentation in pregnancies conceived with different fertility treatments compared with spontaneous pregnancies

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