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Heat shock protein 90 inhibition abrogates TLR4-mediated NF-?B activity and reduces renal ischemia-reperfusion injury.

ABSTRACT: Renal ischemia-reperfusion injury (IRI) is a common cause of acute kidney injury. Toll-like receptor 4 (TLR4) mediates sterile inflammation following renal IRI. Heat shock protein 90 (Hsp90) inhibition is a potential strategy to reduce IRI, and AT13387 is a novel Hsp90 inhibitor with low toxicity. This study assessed if pre-treatment with AT13387 could reduce renal IRI and established if the mechanism of protection involved a reduction in inflammatory signalling. Mice were pre-treated with AT13387 prior to renal IRI. 24?h later, renal function was determined by serum creatinine, kidney damage by tubular necrosis score, renal TLR4 expression by PCR and inflammation by cytokine array. In vitro, human embryonic kidney cells were co-transfected to express TLR4 and a secreted alkaline phosphatase NF-?B reporter. Cells were pre-treated with AT13387 and exposed to endotoxin-free hyaluronan to stimulate sterile TLR4-specific NF-?B inflammatory activation. Following renal IRI, AT13387 significantly reduced serum creatinine, tubular necrosis, TLR4 expression and NF-?B-dependent chemokines. In vitro, AT13387-treatment resulted in breakdown of I?B kinase, which abolished TLR4-mediated NF-?B activation by hyaluronan. AT13387 is a new agent with translational potential that reduces renal IRI. The mechanism of protection may involve breakdown of I?B kinase and repression of TLR4-mediated NF-?B inflammatory activity.

SUBMITTER: O'Neill S

PROVIDER: S-EPMC4528191 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature

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Heat shock protein 90 inhibition abrogates TLR4-mediated NF-κB activity and reduces renal ischemia-reperfusion injury.

O'Neill Stephen S Humphries Duncan D Tse George G Marson Lorna P LP Dhaliwal Kevin K Hughes Jeremy J Ross James A JA Wigmore Stephen J SJ Harrison Ewen M EM

Scientific reports 20150807

Renal ischemia-reperfusion injury (IRI) is a common cause of acute kidney injury. Toll-like receptor 4 (TLR4) mediates sterile inflammation following renal IRI. Heat shock protein 90 (Hsp90) inhibition is a potential strategy to reduce IRI, and AT13387 is a novel Hsp90 inhibitor with low toxicity. This study assessed if pre-treatment with AT13387 could reduce renal IRI and established if the mechanism of protection involved a reduction in inflammatory signalling. Mice were pre-treated with AT133 ...[more]

PMID: 26248657

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Project description:Ischemic stroke is a devastating disease with long-term disability. However, the pathogenesis is unclear and treatments are limited. Meisoindigo, a second-generation derivative of indirubin, has general water solubility and is well-tolerated. Previous studies have shown that meisoindigo reduces inflammation by inhibiting leukocyte chemotaxis and migration. In the present study, we investigated the hypothesis that meisoindigo was also protective against ischemic stroke, then evaluated its underlying mechanisms. In vivo, adult male C57BL/6J wild-type mice were used to produce a middle cerebral artery occlusion (MCAO) stroke model. On day three after reperfusion, obvious improvement in neurological scores, infarct volume reduction and cerebral edema amelioration were observed in meisoindigo treatment. Moreover, immunofluorescence staining and western-blot showed that the expression of NLRP3 inflammasome and its associated proteins in neurons and microglia was inhibited by meisoindigo. The effects of Meisoindigo on NLRP3 inflammasome inactivation and increased the M2 phenotype of microglia/macrophage through shifting from a M1 phenotype, which was possibly mediated by inhibition of TLR4/NF-κB. Furthermore, we verified the inhibitory effect of meisoindigo on TLR4/NF-κB signaling pathway, and found that meisoindigo treatment could significantly suppressed the expression of TLR4/NF-κB pathway-associated proteins in a dose-dependent manner, meanwhile, which resulted in downregulation of HMGB1 and IL-1β. Next, we established an in vitro oxygen glucose deprivation/Reperfusion (OGD/R) model in HT-22 and BV2 cells to simulate ischemic conditions. Cytotoxicity assay showed that meisoindigo substantially improved relative cell vitality and in HT-22 and BV2 cells following OGD/R in vitro. After suffering OGD/R, the TLR4/NF-κB pathway was activated, the expression of NLRP3 inflammasome-associated proteins and M1 microglia/macrophage were increased, but meisoindigo could inhibit above changes in both HT-22 and BV2 cells. Additionally, though lipopolysaccharide stimulated the activation of TLR4 signaling in OGD/R models, meisoindigo co-treatment markedly reversed the upregulation of TLR4 and following activation of NLRP3 inflammasome and polarization of M1 microglia/macrophages mediated by TLR4. Overall, we demonstrate for the first time that meisoindigo post-treatment alleviates brain damage induced by ischemic stroke in vivo and in vitro experiments through blocking activation of the NLRP3 inflammasome and regulating the polarization of microglia/macrophages via inhibition of the TLR4/NF-κB signaling pathway.

Dataset Information

Heat shock protein 90 inhibition abrogates TLR4-mediated NF-?B activity and reduces renal ischemia-reperfusion injury.

Publications

Heat shock protein 90 inhibition abrogates TLR4-mediated NF-κB activity and reduces renal ischemia-reperfusion injury.

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