Investigating the Pathogenesis of Hypertensive Disorders of Pregnancy: Role of Placental Inflammation and Mitochondria

Objectives: We hypothesize that inflammatory associated signaling promotes metabolic imbalance and induces mitochondrial dysfunction that alters the fate of human placental trophoblasts. Methods: RNA-sequencing was performed on human placental cross-sections from HDP women, and the data were analyzed using Gene Set Enrichment Analysis with the hallmark gene sets from the human molecular signatures database. Human trophoblasts were subjected to TNFɑ (10-100 ng/mL) in vitro. RvD2 (10-100nM) was applied as a treatment strategy to mitigate inflammation and mitochondrial dysfunction. Migration assays were performed to measure trophoblast function. qPCR and immunoblots were used to measure NRF2 and TFAM mRNA and protein expression. Seahorse® was used to measure oxygen consumption rates (OCAR). LDH and CCK8 colorimetric assays were used to measure cytotoxicity and cell viability. One-way ANOVA or student t-test was performed. Results: RNA-sequencing data identified notable changes in inflammatory and mitochondrial pathways in the human placenta, particularly showing elevated TNFɑ levels in HDP placenta in vivo. In vitro experiments demonstrated that RvD2 significantly enhanced trophoblast cell migration while TNFɑ inhibited it (RvD2: 42% vs. TNFɑ: 32%; p=0.019). TNFɑ exposure increased TFAM mRNA and protein levels in trophoblasts, which were mitigated by RvD2 treatment (TFAM relative expression to β-Actin: TNFɑ: 0.002 vs. TNFɑ+RvD2: 0.001; p=0.026). Nuclear NRF2 protein expression increased with the treatment of TNFɑ. Additionally, treatment of RvD2 modulated OCAR for maximal and basal respiratory rates and ATP production (ATP OCAR: RvD2: 119.7 vs. TNFɑ: 295 vs. RvD2+TNFa: 173; p=0.010; pmol/min/mg/mL). TNFɑ significantly decreased migration but did not affect cellular cytotoxicity or viability. Conclusions: RNA-sequencing showed a change in mitochondrial and inflammatory pathways. RvD2 exhibits a protective role through modulation of the NRF2-TFAM pathway in the mitochondria of trophoblasts during TNFɑ-driven inflammatory insults and can functionally alter trophoblast migration. Funding Sources: Child Health Research Institute.