Transforming growth factor β signalling and matrix metalloproteinases in the mucosa overlying Crohn’s disease strictures

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Abstract

Background and Aims:

In addition to its crucial role in dampening tissue-damaging immune responses in the gut, transforming growth factor β (TGFβ) is a potent profibrogenic agent inducing collagen synthesis and regulating the balance between matrix-degrading matrix metalloproteinases (MMPs) and their inhibitors (TIMPs). TGFβ signalling was investigated by analysis of Smad proteins and MMPs/TIMPs in the mucosa overlying strictures in patients with Crohn’s disease (CD).

Methods:

Specimens were collected from macroscopically normal mucosa overlying strictured and non-strictured gut of patients with fibrostenosing CD. Isolated myofibroblasts were cultured with anti-TGFβ blocking antibody or TGFβ1. TGFβ transcripts were analysed by quantitative reverse transcription-PCR (RT-PCR). Smad proteins and MMPs were determined by immunoblotting. MMP-12 activity was measured by a real-time MMP-12 activity assay. An in vitro wound-healing scratch assay was used to assess myofibroblast migration.

Results:

TGFβ transcripts, phosphorylated Smad2–Smad3 (pSmad2–3) and TIMP-1 proteins were higher in mucosa overlying strictures than in mucosa overlying non-strictured areas. In contrast, mucosa overlying strictured gut had lower expression of Smad7, MMP-12 and MMP-3. Myofibroblasts from mucosa overlying strictured gut showed higher TGFβ transcripts, a greater pSmad2–3 response to TGFβ, increased TIMP-1, lower Smad7, increased collagen production and reduced migration ability compared with myofibroblasts from mucosa overlying non-strictured gut. TGFβ blockade increased myofibroblast MMP-12 production and migration, more obviously in myofibroblasts isolated from mucosa overlying non-strictured compared with strictured gut.

Conclusions:

Changes in TGF-β signalling and MMP production were identified in the mucosa overlying strictures in CD which may give a window into the process of fibrosis.

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