novobiocin and Colitis

Inflammatory bowel diseases are chronic intestinal inflammatory diseases thought to reflect a dysregulated immune response. Although antibody-based inhibition of tumor necrosis factor-α (TNF-α) has provided relief to many inflammatory bowel diseases patients, these therapies are either ineffective in a patient subset or lose their efficacy over time, leaving an unmet need for alternatives. Given the critical role of the heat shock response in regulating inflammation, this study proposed to define the impact of selective inhibition of heat shock protein 90 (HSP90) on intestinal inflammation. Using multiple preclinical mouse models of inflammatory bowel diseases, we demonstrate a potent anti-inflammatory effect of selective inhibition of the HSP90 C-terminal ATPase using the compound novobiocin. Novobiocin-attenuated dextran sulfate sodium-induced colitis and CD45RB adoptive-transfer colitis through the suppression of inflammatory cytokine secretion, including TNF-α. In vitro assays demonstrate that CD4 T cells treated with novobiocin produced significantly less TNF-α measured by intracellular cytokine staining and by enzyme-linked immunosorbent assay. This corresponded to significantly decreased nuclear p65 translocation by Western blot and a decrease in nuclear factor-κB luciferase activity in Jurkat T cells. Finally, to verify the anti-TNF action of novobiocin, 20-week-old TNFΔ mice were treated for 2 weeks with subcutaneous administration of novobiocin. This model has high levels of circulating TNF-α and exhibits spontaneous transmural segmental ileitis. Novobiocin treatment significantly reduced inflammatory cell infiltrate in the ileal lamina propria. HSP90 inhibition with novobiocin offers a novel method of inflammatory cytokine suppression without potential for the development of tolerance that limits current antibody-based methods.

Topics: Adenosine Triphosphatases; Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Cell Nucleus; Cell Proliferation; Colitis; Cytokines; Dextran Sulfate; Enzyme Activation; Enzyme Inhibitors; HSP90 Heat-Shock Proteins; Humans; Ileitis; Intestinal Mucosa; Jurkat Cells; Mice; Mice, Inbred C57BL; Novobiocin; Transcription Factor RelA; Tumor Necrosis Factor-alpha

Several enterohemorrhagic Escherichia coli (EHEC) strains of serotype O157:H7 isolated from patients with hemorrhagic colitis, ischemic colitis, or hemolytic uremic syndrome were all found to be able to invade certain human epithelial cell lines in vitro. Their ability to gain entry into epithelial cells was compared with those of known invasive Shigella flexneri and Salmonella typhi strains and the noninvasive E. coli strain HB101 in invasion assays utilizing gentamicin to kill extracellular bacteria. All EHEC strains under investigation were efficiently internalized into T24 bladder and HCT-8 ileocecal cells. In striking contrast to shigellae, the same EHEC strains were not taken up into human embryonic intestinal INT407 cells or HEp-2 cells any more than the noninvasive E. coli strain HB101. The mechanism(s) of EHEC internalization was characterized by comparing the invasion efficiencies in the absence and presence of a variety of inhibitors acting on structures and processes of prokaryotic or eukaryotic cells. Also, wild-type, plasmid-containing EHEC strains were compared with their plasmid-cured isogenic derivative strains to determine if plasmid genes affect invasion ability. Plasmid-cured EHEC invaded as well as wild-type EHEC, indicating that invasion ability is chromosomally encoded. Inhibition of bacterial protein synthesis by simultaneous addition of bacteria and chloramphenicol to the monolayer blocked EHEC uptake dramatically, suggesting the presence of an invasion protein(s) with a short half-life. Studies utilizing inhibitors which act on eukaryotic cells demonstrated a strong dependence on microfilaments in the process of uptake of all EHEC strains into both T24 and HCT-8 cells. In general, depolymerization of microtubules as well as inhibition of receptor-mediated endocytosis reduced the efficiency of EHEC invasion of T24 cells, whereas interference with endosome acidification reduced EHEC entry into only HCT-8 cells. Taxol-induced stabilization of microtubules did not inhibit internalization into T24 cells or into the HCT-8 cell line. In marked contrast, the ability of S. typhi Ty2 to invade either cell line was inhibited only by depolymerization of microfilaments. In addition to the cell line specificity of EHEC invasion, not all EHEC strains displayed uniform behavior in the presence of inhibitors, suggesting the existence of variant uptake pathways in different strains. Most importantly, previous reports of the inability of EHEC to in

Topics: Actin Cytoskeleton; Bacterial Proteins; Cadaverine; Cell Line; Chloramphenicol; Clathrin; Colitis; Cytoplasm; DNA, Bacterial; Endocytosis; Epithelium; Escherichia coli; Escherichia coli Infections; Hemolytic-Uremic Syndrome; Hemorrhage; Humans; Hydrogen-Ion Concentration; In Vitro Techniques; Microtubules; Novobiocin; Receptors, Cell Surface; Rifamycins; RNA, Bacterial; Salmonella typhi; Shigella flexneri

Topics: Anti-Bacterial Agents; Colitis; Humans; Novobiocin; Tetracycline

Topics: Anti-Bacterial Agents; Candida albicans; Candidiasis; Colitis; Enterocolitis; Micrococcus; Novobiocin