Role of alpha-hemolysin in the extracellular matrix remodeling of human chorioamniotic membranes

ACCIALINI PAULA; FARINA MARIANA; PUCCI MOLINERIS MELISA; PELINSKI PABLO; HERLAX VANESA; LIMA MS; BARBERO HUGO

Virtual

Congreso; SCHCF + ALACF Joint Meeting 2020; 2020

Introduction: α-hemolysin (HlyA), toxin secreted by uropathogenic strains of Escherichia coli, has a fundamental role in urinary tract infections (UTIs). In pregnancy UTIs are very frequent, being E. coli the etiological agent of almost the 80% of the cases. Considering that UTIs are related with premature rupture of fetal membranes, we proposed to analyze changes of human chorioamniotic membranes treated with HlyA in vitro.Methods: Chorioamniotic membranes (n=8) were obtained from deliveries by elective cesarean section (>37 weeks). All included women had normal pregnancies without evidence of active labor or infection. This protocol was approved by the Hospital Español Review Board (La Plata-EI001/19). Membrane explants were mounted and tied to a Transwell device generating two independent chambers. To simulate an ascending infection, explants were incubated in the chorion-side with 5nM/50nM HlyA during 24h. HlyA was detected by immunohistochemistry and histological signs of damage (like edema, vacuolization, and apoptosis) were evaluated from paraffin-embedded tissue sections stained with hematoxylin/eosin. Transepithelial electrical resistance (TEER) was measured using a Millicell-ERS unit, necrosis was evaluated by LDH release, (n=3), metalloprotease activity by zimography, and cyclooxygenase-2 (COX2) expression by RT-qPCR. Groups were compared using t-, U Mann-Whitney or Chi-squared test as correspond and values are shown as media ± SEM.Results: HlyA interaction with chorioamniotic membranes caused structural alterations and a slight diminish of TEER after 24hs of incubation. The main tissue alterations were observed for the highest toxin concentration tested (50nM HlyA). Epithelial layer remained practically unaltered, while chorion cells showed an increment of vacuolization and necrosis. Extracellular matrix thickness, COX-2 expression and metalloproteinase activity were higher and fibroblast number lower in treated groups compared to control ones.Conclusion: HlyA by itself is capable to introduce structural and molecular modifications in human chorioamniotic membranes, suggesting a role of this toxin in chorioamniotic extracellular matrix remodeling.