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TNF induces caspase-dependent inflammation in renal endothelial cells through a Rho- and myosin light chain kinase-dependent mechanism

2009; American Physical Society; Volume: 297; Issue: 2 Linguagem: Inglês

10.1152/ajprenal.00089.2009

1931-857X

Xiaoyan Wu, Rongqing Guo, Peili Chen, Quan Wang, Patrick N. Cunningham,

Inflammasome and immune disorders

The pathogenesis of LPS-induced acute kidney injury (AKI) requires signaling through tumor necrosis factor-alpha (TNF) receptor 1 (TNFR1), which within the kidney is primarily located in the endothelium. We showed previously that caspase inhibition protected mice against LPS-induced AKI and in parallel significantly inhibited LPS-induced renal inflammation. Therefore we hypothesized that caspase activation amplifies TNF-induced inflammation in renal endothelial cells (ECs). In cultured renal ECs, TNF induced apoptosis through a caspase-8-dependent pathway. TNF caused translocation of the p65 subunit of NF-kappaB to the nucleus, resulting in upregulation of inflammatory markers such as adhesion molecules ICAM-1 and VCAM-1. However, the broad-spectrum caspase inhibitor Boc-d-fmk reduced NF-kB activation as assessed by gel shift assay, reduced phosphorylation of subunit IkappaBalpha, and significantly inhibited TNF-induced expression of ICAM-1 and VCAM-1 as assessed by both real-time PCR and flow cytometry. Broad-spectrum caspase inhibition markedly inhibited neutrophil adherence to the TNF-activated endothelial monolayer, supporting the functional significance of this effect. Specific inhibitors of caspases-8 and -3, but not of caspase-1, reduced TNF-induced NF-kappaB activation. Caspase inhibition also reduced TNF-induced myosin light chain (MLC)-2 phosphorylation, and activation of upstream regulator RhoA. Consistent with this, MLC kinase (MLCK) inhibitor ML-7 reduced TNF-induced NF-kappaB activation. Thus caspase activation influences NF-kappaB signaling via its affect on cytoskeletal changes occurring through RhoA and MLCK pathways. These cell culture experiments support a role for caspase activation in TNF-induced inflammation in the renal endothelium, a key event in LPS-induced AKI.