Induction of human-fetal-membrane remodeling in-vitro by the alpha hemolysin of Escherichia coli

Abstract

IntroductionAlmost 80% of urinary tract infections during pregnancy are caused by uropathogenic strains of Escherichia coli. Alpha-hemolysin, toxin secreted by them, has a fundamental role in this pathology development. Considering that urinary tract infections are related with premature rupture of fetal membranes, we proposed to evaluate the effects that alpha-hemolysin induces on human-fetal-membranes. MethodsThirteen fetal membranes obtained from elective cesarean sections (>37 weeks) were mounted in a transwell-device generating two independent chambers. To mimic an ascendant-urinary-tract infection, membranes were incubated with different concentrations of pure alpha-hemolysin from the choriodecidual side during 24h. Extensive histological analyses were performed and transepithelial electrical-resistance were determined. Cell viability, metalloproteinase activity and cyclooxygenase-2- gene expression was estimated by lactate-dehydrogenase–release assay, zymography and RT-qPCR, respectively. Finally, four fetal membranes were treated with hemolysin preincubated with polyclonal anti-hemolysin antibodies. Cell viability and metalloproteinase activity were monitored. ResultsAfter 24 h of treatment, fetal membranes evidenced a structural damage and a decrease in membrane resistance that progressed as the concentration of alpha hemolysin increased. While the amniotic-epithelial-layer remained practically unaffected, the chorion cells manifested an increase in vacuolization and necrosis. In addition, the extracellular matrix exhibited collagen-fiber disorganization, a marked decrease in fiber content, and became thicker in presence of the toxin. Cyclooxigenase-2 expression and metalloproteinase activity were also higher in the treated groups than in untreated ones. Finally, a preincubation of hemolysin with specific antibodies prevented the cytotoxicity on the chorion cells and the increase in metalloproteinase activity. DiscussionHemolysin induces structural and molecular changes associated with the remodeling of human-fetal-membranes in-vitro.