Hydrogen sulfide modulates gene networks in hypoxia/reoxygenation-stressed trophoblasts: insights from transcriptome...

Hydrogen sulfide is an endogenous gaseous signalling molecule with recognized roles in vascular regulation, redox homeostasis, and inflammation. In the placenta, H₂S is essential for maintaining trophoblast function and promoting healthy vascular remodelling. Impaired H₂S signalling has been implicated in placental disorders characterized by oxidative stress, particularly in preeclampsia. One of the principal drivers of oxidative stress in the placenta is H/R injury, which mimics the intermittent perfusion patterns seen in early placental maldevelopment. Although the protective roles of H₂S have been described in several ischemia-reperfusion models, its genome-wide transcriptional effects on trophoblasts under hypoxia/reoxygenation-induced oxidative stress remain unknown.

HTR-8/SVneo trophoblasts were subjected to H/R injury induced by varying oxygen concentrations to model the fluctuating oxygen environments of early placental development, followed by treatment with an exogenous H₂S donor (NaHS). A CSE inhibitor (PAG) treatment was also given. RNA sequencing was performed to characterize global gene expression changes. Differentially expressed genes were analyzed using KEGG and Gene Ontology enrichment, protein-protein interaction network mapping, and transcription factor prediction.

H/R induced extensive transcriptional remodelling, with robust activation of HIF-1, PI3K-Akt, MAPK, Rap1/Ras, NF-κB, and focal adhesion pathways. H/R [2/10% O₂] triggered pronounced glycolytic, hypoxia-adaptive, anti-apoptotic, and pro-invasive signatures. NaHS modulated these responses in a context-dependent manner: it attenuated early chemokine-driven inflammation, enhanced angiogenic and ECM-remodelling programs, and strengthened metabolic adaptation under a higher hypoxic burden 2/10% H/R paradigm. PAG induced a chronic inflammatory angiogenic signature, indicating endogenous H₂S restrains basal inflammatory activation. Integrated regulation of proliferation, migration, apoptosis, morphogenesis, and angiogenesis was observed through biological process analysis, with major changes noticed in NaHS-treated 2/10% H/R conditions. JUN, PTGS2, MAP3K5, DUSP1, SFN, NCF2, THBS2, and GADD45A emerged as the central interconnected hub-gene module through PPI analysis. Among these, JUN and PTGS2 appeared as potential integrators of trophoblast remodelling, redox stress, and inflammatory signalling.

Our study provides the first evidence of transcriptomic analysis showing that H₂S alters gene networks in trophoblast cells subjected to H/R-induced oxidative stress. The results highlight coordinated regulation of metabolic, angiogenic, and inflammatory pathways, providing fundamental understanding into how H₂S may influence trophoblast adaptation to stress.