Background and aims

Myofibroblast hyperplasia contributes to muscularis mucosae thickening and stricture formation in Crohn's disease (CD). Protease-activated receptor-2 (PAR-2) and cytosolic phospholipase A₂ (cPLA₂) are known regulators of cell growth, but their significance in intestinal myofibroblast proliferation remain to be elucidated. The principle aims of the present study were to investigate if PAR-2 is expressed in the expanded muscularis mucosa in ileal CD specimens, if inflammatory cytokines may stimulate PAR-2 expression in intestinal myofibroblasts, and if PAR-2 and cPLA₂ may regulate intestinal myofibroblast growth.

Methods

Immunohistochemistry was used for detection of PAR-2 in ileal CD specimens. Studies on PAR-2 expression, PLA₂ activation and cell growth were performed in a human intestinal myofibroblast cell line, CCD-18Co. PAR-2 expression was investigated by RT-PCR and immunocytochemistry. PLA₂ activity was analyzed by quantification of released ¹⁴C-arachidonic acid (¹⁴C-AA). Cell growth was examined by ³H-thymidine incorporation and cell counting.

Results

The thickened muscularis mucosae of the CD specimens showed strong PAR-2 expression. In cultured myofibroblasts, tumor necrosis factor-α (TNF-α) up-regulated PAR-2 mRNA and protein, and potentiated PAR-2-stimulated ¹⁴C-AA release by two known PAR-2 activators, trypsin and SLIGRL-NH₂. The release of ¹⁴C-AA was dependent on cPLA₂. Trypsin stimulated the proliferation of serum-starved cells, and inhibition of cPLA₂ reduced normal cell growth and abolished the growth-promoting effect of trypsin.

Conclusions

The results suggest that PAR-2-mediated cPLA₂ activation might be of importance in intestinal myofibroblast proliferation. The results also point to the possibility that PAR-2 up-regulation by inflammatory cytokines, like TNF-α, may modulate this effect.