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P004. Smooth muscle cells participate in Crohn's disease intestinal fibrosis

A. Scirocco1, S. Rosati1, R. Sferra2, A. Vetusti2, N. Pallotta1, G. Tellan1, A. Pronio3, E. Corazziari1, C. Severi1, 1La Sapienza, Medicina Interna e Specialità Mediche, Rome, 2Università degli Studi dell'Aquila, 3Università Sapienza, Italy

Background

Intestinal fibrosis is a progressive process driven by interaction between inflammatory mediators/growth factors with mesenchimal fibroblasts, myofibroblasts and smooth muscle cells (SMC). Extracellular matrix arises from sub-epithelial myofibroblasts but a contribution of both fibroblasts and SMC has been hypothesized. Among growth factors, PDGF drives SMC switch from a contractile to synthetic phenotype. To asses the potential contribution of smooth muscle to the fibrogenetic process by analyzing molecular and morphofunctional alterations occurring in human smooth muscle in normal and Crohn's disease (CD) affected ileum.

Methods

Morphometric analysis and mRNA expression of inflammatory molecules (IL-1b, TNFa), growth factors (PDGFb, TGFa) and collagen III were performed on circular SMC isolated from human normal and 5 CD pre- and stenotic ileum. Immunohistochemistry (IHC) was carried out on whole mount tissue for PDGFb, TGFb and collagen III. Data are expressed as mean±SE, p < 0.05 considered significant.

Results

SMC expressed constitutively mRNA encoding for IL1b, TNFa, PDGFb, TGFb and collagen III. qPCR analysis showed an increase in constitutive mRNA expression of IL1 b, TNFa and PDGFb from normal to pre-stenotic CD specimen. In parallel a decrease in TGFb expression was observed. The compared analysis between pre- and stenotic CD specimen highlighted a further significant fold increase in IL1b (4.93±0.99), TNFa (2.85±0.91) and PDGFb (4.27±1.10) with a decrease in TGFb (0.75±1.18). Collagen III expression did not differ between normal and pre-stenotic CD but presented a significant fold increase in stenotic ileum (4.83±1.24). In parallel, IHC analysis showed a PDGFb and collagen increase expression within the CD muscle layer with no variation in TGFb. In addition, SMC isolated from stenotic CD presented altered morpho-functional parameters consisting in a decrease in cell length (control: 82.45±16.50 µm; CD: 71.89±3.21) and contractile response (control: 22.89±0.89; CD: 13.43±4.15%).

Conclusion

Smooth muscle alterations occurring in CD highlight a possible myogenic contribution to intestinal fibrosis. These alterations already present in pre-stenotic CD specimen may represent a new possible anti-fibrotic therapeutic target.