Global gene expression analysis reveals a role for the Rho/ROCK pathway in intestinal smooth muscle cells fibrogenic differentiation.
MC Vozenin-Brotons, C Bourgier V Haydont, F Milliat, A François, V Holler, P Lasser, J Bourhis, D Mathé, ELSO 2004 4-8/09/2004, Nice, France.
Background & Aims: Radiation enteritis is associated with activation of intestinal smooth muscle cells and increased CTGF expression (1). cDNA array analysis revealed alteration of the genes coding for the Rho family of proteins (2). As Rho proteins are involved in the regulation of stress fiber formation and may regulate CTGF expression, we postulated that alteration of the Rho Pathway is involved in maintenance of radiation-induced smooth muscle cells fibrogenic differentiation. Methods: Radiation enteritis and normal ileum biopsies were dissected muscularis layers were enzymatically digested to establish primary culture of smooth muscle cells. Cytoskeleton structure and composition, as well as CTGF and type I collagen secretion were studied. The involvement of the Rho/ROCK pathway in the fibrosis-activated smooth muscle cells was assessed using Y-27632, a specific inhibitor of Rho kinase. Results: Intestinal smooth muscle cells retained their fibrogenic differentiation in vitro and exhibited a typical cytoskeletal network. CTGF and pro-Collagen I secretion was enhanced in cells derived from radiation enteritis as compared with their normal counterparts and was concomitant to alteration of the Rho pathway. Specific inhibition of the Rho kinase with Y-27632, induced a simultaneous decrease in the number of actin stress fibers, in the a-sm actin and HSP27 protein level. Y-27632 also decreased CTGF level probably through NF-κB inhibition as well as type I collagen gene. Conclusions: The present study suggests that the Rho/ROCK pathway is involved in the control of actin network and in regulation of CTGF expression associated with radiation-induced fibrogenic differentiation in intestinal smooth muscle cells. Furthermore, inhibition of the Rho/ROCK pathway seemed to reverse the radiation-induced fibrogenic differentiation in vitro and suggests that specific inhibition of Rho kinase may be a promising approach for the development of antifibrotic therapies. 1) Vozenin-Brotons et al. Fibrogenic signals in patients with radiation enteritis are associated with increased connective tissue growth factor expression. Int J Radiat Oncol Biol Phys, 56, 561-72, 2003. Vozenin-Brotons et al. cDNA array reveals genes differentially expressed in human radiation enteritis. Rad. Res. 3:299-311, 20