Inhibition of Rho kinase modulates radiation induced fibrogenic phenotype in intestinal smooth muscle cells through alteration of the cytoskeleton and connective tissue growth factor expression. Issue 3 (11th February 2005)

• Record Type: Journal Article
• Authors: Bourgier, C; Haydont, V; Milliat, F; François, A; Holler, V; Lasser, P; Bourhis, J; Mathé, D; Vozenin-Brotons, M-C
• Is Part Of: Gut. Volume 54:Issue 3(2005)
• Page Start: 336
• Page End: 343
• Publication Date: 2005-02-11
• Subjects: CTGF, connective tissue growth factor -- α/γ-sm actin, α/γ smooth muscle actin -- HSP, heat shock protein -- ROCK, Rho kinase -- N/RE SMC, normal/radiation enteritis smooth muscle cells -- COL1A1, type I collagen alpha 1 -- MLCK, myosin light chain kinase -- RT-PCR, reverse transcription-polymerase chain reaction -- EMSA, electrophoretic mobility shift assay -- PAGE, polyacrylamide gel electrophoresis -- NFκB, nuclear factor κB -- TNF-α, tumour necrosis factor α -- TGF-β1, transforming growth factor β1; connective tissue growth factor -- intestinal smooth muscle cells -- Rho pathway -- fibrosis -- radiotherapy; Gastroenterology -- Periodicals; 616.33
• DOI: 10.1136/gut.2004.051169
• Languages: English
• ISSNs: 0017-5749

Abstract

Abstract : Background: Late radiation enteritis in humans is associated with accumulation of extracellular matrix and increased connective tissue growth factor (CTGF) expression that may involve intestinal muscular layers. Aims: We investigated the molecular pathways involved in maintenance of radiation induced fibrosis by gene profiling and postulated that alteration of the Rho pathway could be associated with radiation induced fibrogenic signals and CTGF sustained expression. Patients and methods: Ileal biopsies from individuals with and without radiation enteritis were analysed by cDNA array, and primary cultures of intestinal smooth muscle cells were established. Then, the effect of pharmacological inhibition of p160 Rho kinase, using Y-27632, was studied by real time reverse transcription-polymerase chain reaction, western blot, and electrophoretic mobility shift assay. Results: Molecular profile analysis of late radiation enteritis showed alterations in expression of genes coding for the Rho proteins. To investigate further the involvement of the Rho pathway in intestinal radiation induced fibrosis, primary intestinal smooth muscle cells were isolated from radiation enteritis. They retained their fibrogenic differentiation in vitro, exhibited a typical cytoskeletal network, a high constitutive CTGF level, increased collagen secretory capacity, and altered expression of genes coding for the Rho family. Rho kinase blockade induced a simultaneous decrease in the number of actin stress fibres, α smooth muscle actin, and heat shock protein 27 levels. It also decreased CTGF levels, probably through nuclear factor κB inhibition, and caused decreased expression of the type I collagen gene. Conclusion: This study is the first showing involvement of the Rho/Rho kinase pathway in radiation fibrosis and intestinal smooth muscle cell fibrogenic differentiation. It suggests that specific inhibition of Rho kinase may be a promising approach for the development of antifibrotic therapies.