PLoS neglected tropical diseases · 2025
Distinct strategies of epithelial cell barrier disruption by Leptospira interrogans isolated from human patients in Okinawa, Japan.
СтатьяПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод в очереди
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Kakita Tetsuya, Toma Claudia, Sebastián Isabel, Barbee Hunter, Yamashiro Tetsu
Аннотация
BACKGROUND: Leptospirosis is a bacterial infection common in tropical and subtropical regions, which causes feverish conditions. Although approximately half of human leptospirosis cases in Japan are reported in Okinawa, a subtropical area, the pathogenic mechanisms of clinical isolates from this region remain unknown. This study aimed to identify the infection mechanisms of L. interrogans isolates from Okinawa (Oki-strains) in human renal proximal tubule epithelial cells (RPTECs).
METHODOLOGY/PRINCIPAL FINDINGS: The transepithelial electrical resistance (TEER) measurements of 11 Oki-strains in infected renal proximal tubule epithelial cells (RPTECs) showed that all strains caused a decrease in TEER by 48 hours post-infection. Imaging analysis of RPTECs infected with two selected strains (Oki53 and Oki65) revealed that both strains induced the displacement of adherens junction (AJ) proteins E-cadherin, α-catenin, afadin, and nectin-2 and cytoskeletal F-actin disorganization. However, western blotting analysis revealed that AJ protein levels were not reduced, except for afadin-an important protein for linking F-actin to AJs. Chemical inhibition revealed that the proteosome inhibitors MG132 and bortezomib and pan-caspase inhibitor Z-VAD-FMK prevented the Oki53-induced TEER decrease, AJ protein mislocalization, afadin degradation, and F-actin disorganization. However, in Oki65-infected RPTECs, the inhibitors partially prevented these effects. Thus, the AJ-F-actin link and epithelial barrier were fully preserved in Oki53-infected RPTECs pretreated with these inhibitors at 24 h post-infection (~TEER 130% of initial TEER), whereas in Oki65-infected cells, the AJ-F-actin link was maintained only partially (~TEER 70%).
CONCLUSIONS/SIGNIFICANCE: Our findings suggest that the maintenance of epithelial barrier integrity requires both the afadin-F-actin and α-catenin-F-actin links, and that Oki53 and Oki65 employ distinct strategies to disrupt the AJ-F-actin link and, thus, the epithelial barrier.
METHODOLOGY/PRINCIPAL FINDINGS: The transepithelial electrical resistance (TEER) measurements of 11 Oki-strains in infected renal proximal tubule epithelial cells (RPTECs) showed that all strains caused a decrease in TEER by 48 hours post-infection. Imaging analysis of RPTECs infected with two selected strains (Oki53 and Oki65) revealed that both strains induced the displacement of adherens junction (AJ) proteins E-cadherin, α-catenin, afadin, and nectin-2 and cytoskeletal F-actin disorganization. However, western blotting analysis revealed that AJ protein levels were not reduced, except for afadin-an important protein for linking F-actin to AJs. Chemical inhibition revealed that the proteosome inhibitors MG132 and bortezomib and pan-caspase inhibitor Z-VAD-FMK prevented the Oki53-induced TEER decrease, AJ protein mislocalization, afadin degradation, and F-actin disorganization. However, in Oki65-infected RPTECs, the inhibitors partially prevented these effects. Thus, the AJ-F-actin link and epithelial barrier were fully preserved in Oki53-infected RPTECs pretreated with these inhibitors at 24 h post-infection (~TEER 130% of initial TEER), whereas in Oki65-infected cells, the AJ-F-actin link was maintained only partially (~TEER 70%).
CONCLUSIONS/SIGNIFICANCE: Our findings suggest that the maintenance of epithelial barrier integrity requires both the afadin-F-actin and α-catenin-F-actin links, and that Oki53 and Oki65 employ distinct strategies to disrupt the AJ-F-actin link and, thus, the epithelial barrier.