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P024. Regional differences in effect of TGF-beta1 and PDGF on the early onset of intestinal fibrosis

B.T. Pham, W.T. van Haaften, R. Iswandana, D. Oosterhuis, P. Olinga, University of Groningen, Pharmaceutical Technology and Biopharmacy, Groningen, Netherlands

Background

Intestinal fibrosis (IF) is one of the major complications in inflammatory bowel disease patients. IF can cause narrowing of the intestinal lumen, which may lead to stricture formation. The mechanism of IF is still unknown and adequate models are lacking. By using precision-cut intestinal slice (PCIS) from different regions of the bowel, we studied the early onset of fibrosis in mouse jejunum, ileum and colon PCIS, in the presence of transforming growth factor (TGF)-beta1 and platelet-derived growth factor (PDGF).

Methods

Mouse jejunum, ileum and colon were excised and prepared as a segment embedded in agarose. PCIS (estimated 300–400 µm) was prepared and incubated up to 48 hr with or without the presence of TGF-beta1 and PDGF. ATP content of the PCIS was used to assess the general viability. The gene expression of different fibrosis markers including Pro-Collagen 1 A1 (COL1A1), heat shock protein 47 (HSP47), alpha-smooth muscle actin (SMA), connective tissue growth factor (CTGF), synaptophysin (SYN) and fibronectin (FN2) were determined.

Results

Mouse PCIS from different segments were viable up to 48 hr. After 48 hr of incubation, HSP47 and FN2 gene expression were significantly up-regulated, compared to PCIS directly after slicing, in jejunum (3.6 and 4.9 fold, respectively) and in ileum (4.9 and 5.5 fold, respectively). When incubated with 5 ng/mL TGF-beta1, in jejunum PCIS, COL1A1, HSP47, CTGF and FN2 were significantly up-regulated compared to 48 hr control. In ileum PCIS the gene expression of HSP47 (1.9 fold) and FN2 (3.9 fold) were also significantly increased.

In the presence of 50 ng/mL PDGF, only in ileum PCIS, CTGF (1.4 fold) and SYN (1.9 fold) were significantly increased compared to 48 hr control. Interestingly, in PCIS from the colon, 5 ng/ml TGF-beta1 did not affect the gene expression of the fibrosis markers. However, HSP47 (1.4 fold) and FN2 (1.7 fold) were significantly increased when colon PCIS were incubated with 50 ng/mL PDGF.

Conclusion

TGF-beta1, but not PDGF, was able to induce HSP47 and FN2 in mouse jejunum and ileum PCIS. This is in contrast to the result in colon PCIS, where only PDGF was able to induce these fibrosis markers. Moreover, PDGF increased CTGF and SYN only in ileum PCIS. These results indicate differences in the effect of TGF-beta1 and PDGF on the early onset of fibrosis in different regions of the intestine.