Nevertheless, the antimicrobial mechanism of quercetin is not elucidated

Nevertheless, the antimicrobial mechanism of quercetin is not elucidated. integrin 1 and FAK association augmented by O157:H7. Furthermore, quercetin reduced FA development induced by bacterial infection and recovered host cell motility. Taken together, data showed that O157:H7 interacts with integrin 1 to facilitate its adhesion to host cells. Quercetin inhibits bacterial infection possibly by blocking the interaction between O157:H7 and integrin 1. Collectively, these data indicate that quercetin provides an alternative antimicrobial to mitigate and control O157:H7 intestinal infection, and suggest potential broad benefits of quercetin and related polyphenols in fighting other enteric pathogen infections. O157:H7, quercetin, integrin 1, anti-adhesion, focal adhesion Introduction Formation intestinal attaching and effacing (A/E) lesions is of necessary for the pathogenesis of O157:H7 (Kaper, 2005). After attachment to intestinal epithelial cells, O157:H7 induces actin PF 06465469 rearrangement to form pedestals (Knutton et al., 1989). Through this tight association with the host cell surface, O157:H7 utilizes various strategies to manipulate host signaling, leading to enhanced bacterial colonization and persistence, and host tissue damage (Xue et al., 2017). The host extracellular matrix (ECM) is composed of multiple macromolecules, which mediate multiple biological functions including cell to cell adhesion, migration, proliferation, and death (Meredith et al., 1993). Integrin 1, the most abundant cell surface integrin, is a transmembrane glycoprotein receptor that interacts with ECM components such as CYFIP1 fibronectin, laminin, and collagen. Through interactions with ECM components, integrin 1 induces multiple bidirectional signal exchanges (Schwartz et al., 1995; Burridge and Chrzanowska-Wodnicka, 1996). In addition, integrin 1 recruits intracellular proteins such as talin, paxillin, and -actinin, leading to the formation of the focal adhesion (FA) complex. To tightly associate with host cells, pathogens utilize integrin 1 as an adhesion factor. interacts with integrin 1 via adhesin YadA to promote tight binding to the host cells (Eitel et al., 2005). attaches to ECM substrate with the assistance of host integrin 1 (Muenzner et al., 2005). In response to infection, the rapid turnover and exfoliation of epithelial cells are innate defense mechanisms against pathogens (Mulvey et al., 2000). However, many pathogenic bacteria can circumvent host exfoliation and colonize the PF 06465469 epithelium efficiently. reduces adhesion complex turnover and suppresses the detachment of infected cells from the basement membrane to manipulate host exfoliation (Kim et al., 2009). PF 06465469 Integrins transduce extracellular signals into the host cells through association with intracellular adaptor proteins and protein kinases such as focal adhesion kinase (FAK) (Dia and Gonzalez de Mejia, 2011) and integrin-linked kinase (ILK) (Gagne et al., 2010). FAK deficiency increases the recruitment of FAs and reduces cell motility (Ilic et al., 1995), indicating FAK is involved in FA formation during cell migration. Thus, pathogens may manipulate FAK and associated kinases, which stabilize the FAs and ultimately enable them to colonize the host cells. Quercetin is a polyphenol widely found in vegetables and fruits. Our previous study demonstrated that quercetin had anti-inflammatory and anti-oxidative properties that prevented O157:H7-induced inflammasome activation (Xue et al., 2017). However, the antimicrobial mechanism of quercetin has not been elucidated. We hypothesized that O157:H7 attaches to host cells via interacting with host integrin 1 and stabilizing FAs formation; quercetin inhibits integrin 1 expression and FA formation thus preventing O157:H7 infection. Materials and Methods Cell Line, Media and Bacterial Strains The human colonic epithelial cell line Caco-2 was obtained from the American Type Culture Collection (Manassas, VA, United States). Caco-2 cells were cultured in Dulbeccos Modified Eagles medium (DMEM) (Sigma, St. Louis, MO, United States) supplemented with 10% fetal bovine serum (Sigma), 100 units/ml penicillin G, and 100 g/ml of streptomycin (Sigma) at 37C with 5% CO2. The O157:H7 EDL933 wild type (EDL933) strain was obtained from the STEC center at Michigan State University. The O157:H7 EDL933 intimin ((plasmid was a generous gift from Dr. John M Leong at Tufts University (Campellone et al., 2002). EDL933pEHEC strain was derived from O157:H7 EDL933strain transformed with pEHEC plasmid. These strains were routinely grown in LB broth at 37C overnight with aeration. Infection of O157:H7 to Colonic Epithelial Cells Caco-2 cells were seeded in a PF 06465469 24-well plate at 5 105 cells/ml for 12 h..