Browsing by Subject "Tight junction"

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    Occludin Content Modulates Hydrogen Peroxide–Induced Increase in Renal Epithelial Paracellular Permeability
    (Wiley, 2016-03) Janosevic, Danielle; Axis, Josephine; Bacallao, Robert L.; Amsler, Kurt; Department of Medicine, IU School of Medicine
    The ability of hydrogen peroxide (H2O2) to increase paracellular permeability of renal epithelial cell monolayers was examined and the role of occludin in this regulation was investigated. H2O2 treatment increased the paracellular movement of calcein, a marker for the leak pathway permeability, across monolayers of two renal epithelial cell lines, MDCK and LLC-PK1, in a concentration-dependent manner. At the same concentrations, H2O2 did not alter transepithelial resistance (TER) nor increase cell death. The magnitude of the H2O2-induced increase in leak pathway permeability was inversely related to cellular occludin protein content. H2O2 treatment did not produce any major change in total cellular content or Triton X-100-soluble or -insoluble fraction content of occludin protein. Occludin protein staining at the tight junction region was diminished following H2O2 treatment. The most dramatic effect of H2O2 was on the dynamic mobility of GFP-occludin into the tight junction region. H2O2 treatment slowed lateral movement of GFP-occludin into the tight junction region but not on the apical membrane. Further, removal of the cytoplasmic C-terminal region of occludin protein eliminated the effect of H2O2 on GFP-occludin lateral movement into the tight junction region. An increase in the mobile fraction of GFP-occludin was associated with a loss of response to H2O2. These data indicate that the H2O2-induced increase in renal epithelial cell paracellular permeability is mediated, at least in part, through occludin protein, possibly through a slowing of the rate of occludin movement into the tight junction region.
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    UCHL1, a deubiquitinating enzyme, regulates lung endothelial cell permeability in vitro and in vivo
    (American Physiological Society, 2021) Mitra, Sumegha; Epshtein, Yulia; Sammani, Saad; Quijada, Hector; Chen, Weiguo; Bandela, Mounica; Desai, Ankit A.; Garcia, Joe G.N.; Jacobson, Jeffrey R.; Biochemistry and Molecular Biology, School of Medicine
    Increasing evidence suggests an important role for deubiquitinating enzymes (DUBs) in modulating a variety of biological functions and diseases. We previously identified the upregulation of the DUB ubiquitin carboxyl terminal hydrolase 1 (UCHL1) in murine ventilator-induced lung injury (VILI). However, the role of UCHL1 in modulating vascular permeability, a cardinal feature of acute lung injury (ALI) in general, remains unclear. We investigated the role of UCHL1 in pulmonary endothelial cell (EC) barrier function in vitro and in vivo and examined the effects of UCHL1 on VE-cadherin and claudin-5 regulation, important adherens and tight junctional components, respectively. Measurements of transendothelial electrical resistance confirmed decreased barrier enhancement induced by hepatocyte growth factor (HGF) and increased thrombin-induced permeability in both UCHL1-silenced ECs and in ECs pretreated with LDN-57444 (LDN), a pharmacological UCHL1 inhibitor. In addition, UCHL1 knockdown (siRNA) was associated with decreased expression of VE-cadherin and claudin-5, whereas silencing of the transcription factor FoxO1 restored claudin-5 levels. Finally, UCHL1 inhibition in vivo via LDN was associated with increased VILI in a murine model. These findings support a prominent functional role of UCHL1 in regulating lung vascular permeability via alterations in adherens and tight junctions and implicate UCHL1 as an important mediator of ALI.