RAB3B regulates vesicle uptake and tight junction integrity in amniotic epithelial cells and is downregulated in premature rupture of membranes

BACKGROUND: Premature rupture of membranes (PROM) is a major cause of preterm birth and neonatal mortality. Tight junctions are critical for maintaining fetal membrane barrier integrity. Ras-related protein Rab-3B (RAB3B), a key regulator of vesicle trafficking, may influence barrier homeostasis; however, its role in PROM remains unclear. METHODS: Whole-transcriptome RNA sequencing was performed on fetal membrane tissues from 31 PROM patients and 10 full-term controls to identify differentially expressed genes. A PROM mouse model was established to assess RAB3B expression and related pathological changes. In vitro, RAB3B was overexpressed or silenced in human amniotic epithelial cells (hAECs) to evaluate its effect on cell migration, epithelial-mesenchymal transition (EMT), epithelial permeability, and tight junction integrity. Extracellular vesicles (EVs) were isolated to explore the role of RAB3B in EV internalization and EV-mediated regulation of tight junctions. RESULTS: RAB3B expression was significantly downregulated in fetal membranes from PROM patients and PROM mice. PROM mice exhibited increased membrane permeability, enhanced apoptosis, elevated pro-inflammatory cytokine levels, and reduced expression of tight junction proteins. In hAECs, RAB3B overexpression promoted cell migration, suppressed EMT, reduced epithelial permeability, and upregulated tight junction proteins, whereas RAB3B knockdown produced opposite effects. Furthermore, RAB3B enhanced EV internalization and potentiated the beneficial effects of EVs on tight junction integrity. CONCLUSION: RAB3B is markedly downregulated in PROM and plays a critical role in maintaining fetal membrane barrier integrity. By enhancing tight junctions and promoting EV-mediated material transport, RAB3B contributes to fetal membrane repair, highlighting its potential as a therapeutic target for PROM prevention and treatment.