Maternal immune activation elicits rapid and sex-dependent changes in gene expression and vascular dysfunction in the rat placenta

Introduction: Maternal immune activation (MIA), characterized by increased circulating inflammatory mediators during pregnancy, is associated with adverse pregnancy outcomes and neurodevelopmental deficits in offspring. These health outcomes often manifest differently depending on fetal-placental sex. A well-established model of MIA involves administration of a viral mimetic, polyinosinic:polycytidilic acid (PolyI:C), to pregnant rodents. Placental responses to PolyI:C contribute to the detrimental effects of MIA on offspring, but these responses have not yet been well characterized. In the present study, we profiled acute gene expression changes in male and female placentas following PolyI:C administration to pregnant rats. Methods: Pregnant rats received 4 mg/kg PolyI:C or saline intravenously on gestational day 18.5, and tissues were harvested 4-5 hours later. Gene expression profiling on placental tissue was performed. Enzyme immunoassays and immunohistochemistry were conducted to determine levels of select proteins in maternal blood and placental tissue, respectively. Results: Maternal PolyI:C exposure caused a robust increase in levels of inflammatory markers in maternal blood and placental tissue. There were more genes differentially expressed in female placentas after PolyI:C exposure (765) than male placentas (221), including reduced expression of genes associated with maternal-fetal communication. Placentas also had increased expression of genes linked with vascular dysfunction after PolyI:C-induced MIA. Discussion: PolyI:C elicited a powerful inflammatory response in the placenta along with vascular dysfunction, likely contributing to the adverse pregnancy outcomes triggered by MIA. Female placentas responded to PolyI:C more vigorously than male placentas, which could underlie the differential outcomes of MIA depending on sex. Pregnant Sprague-Dawley rats (4 rats per group) were injected with sterile saline or 4 mg/kg PolyI:C sodium salts on gestational day 18.5. Placentas were collected 5 h later. Placentas were collected 5 h after injections. Based on genotyping fetuses, one male and one female placenta were used for downstream analyses (therefore 8 placentas per treatment group). RNA was extracted by homogenizing in TRIzol. After adding chloroform, the aqueous phase was collected and diluted 1:1 with 70% ethanol, placed on RNeasy columns (Qiagen Inc), treated with DNase I, and RNA purified. Libraries were generated using the NEBNext directional library prep kit for Illumina (New England Biolabs). Indexed libraries were sequenced on a NovaSeq 6000 system (Illumina) at Génome Québec (Montréal, QC, Canada) using a 100 bp, paired-end read format.