Expression of matrix metalloproteinases-2 and -9 in intestinal tissue of patients with inflammatory bowel diseases

Abstract

Background/aims.

Matrix metalloproteinases are major contributors in the breakdown and reconstitution of basement membranes and extracellular matrix in pathophysiological processes. We assessed the expression of matrix metalloproteinases-2 and -9 in intestinal tissue of patients with inflammatory bowel disease.

Patients/methods.

Resected tissue specimens from patients with Crohn's disease or ulcerative colitis and control tissue from patients with a colorectal carcinoma were used for enzyme-linked immunosorbent assay, zymography, activity assay, reverse transcription polymerase chain reaction and immunohistochemistry to evaluate the expression of these matrix metalloproteinases.

Results.

Matrix metalloproteinase-2 and more strongly matrix metalloproteinase-9 protein and mRNA were markedly increased in inflammatory bowel disease tissues, with the highest levels in severely inflamed tissues. Immunohistochemistry showed that matrix metalloproteinase-2 was present in the extracellular matrix of the submucosa, with a lower but more generalised expression in the severely inflamed regions. Matrix metalloproteinase-9 was most prominent in polymorphonuclear leukocytes and was increased, also in activity, in all inflammatory bowel disease tissues. An increased matrix metalloproteinase-9 expression in the extracellular matrix was observed in relation to the severity of inflammation.

Conclusions.

Matrix metalloproteinases-2 and -9 are enhanced in the intestinal tissue and seem to be actively involved in the inflammatory and remodelling processes in inflammatory bowel disease, without major differences between CD and UC.

Introduction

Inflammatory bowel diseases (IBD), i.e. Crohn's disease (CD) and ulcerative colitis (UC), are characterised by inflammation and ulceration of the gastrointestinal tract, with numerous gastrointestinal tract and systemic complications, but of unknown specific aetiology [1]. In the pathophysiological process of IBD, a variety of inflammatory mediators, such as proteolytic enzymes, cytokines and growth factors, and many kinds of cells, like leukocytes and stromal cells, are implicated in the tissue injury and healing processes [2].

Matrix metalloproteinases (MMPs) are a family of Zn²⁺-containing neutral proteinases and are thought to be major contributors to breakdown and reconstitution of extracellular matrix (ECM) in physiological processes, like tissue remodelling during development, growth and wound repair, and in pathological conditions, including destructive diseases, such as arthritis, atherosclerotic plaque rupture and tumour progression [3], [4], [5]. In general, MMPs are secreted as inactive proenzymes that require proteolytic cleavage for activation. The proteolytic activity of MMPs is precisely regulated by the balance between zymogen activation and enzyme inhibition through endogenous inhibitors, such as α-macroglobulins and tissue inhibitors of metalloproteinases (TIMPs) [6]. To date, over 20 members of the MMPs family have been found in vertebrates. Depending on substrate specificity, amino acid similarity and identifiable sequence modules, MMPs are divided into four major subgroups: collagenases, stromelysins, gelatinases and membrane-type MMPs. Gelatinases are composed of two members: MMP-2 (gelatinase A), a 72 kDa proteinase, and MMP-9 (gelatinase B), a 92 kDa proteinase, which specifically degrade basement membrane (BM) type IV collagen, as well as gelatin, collagen type I, V, VII, X, elastin, laminin and fibronectin [7], [8]. MMP-2, a most commonly expressed enzyme in normal adult tissue, is primarily produced by stromal cells. MMP-9 is mainly synthesised by inflammatory cells, particularly polymorphonuclear leukocytes (PMNL) [3], [9], [10], [11]. The expression of MMP-2 and MMP-9 has been found to change in different situations, such as embryonic development, diverse pathophysiological conditions and during culture [6], [12]. For example, MMPs are proposed to be major factors for intestinal tissue injury mediated by T cells in IBD [13], [14]. Previous studies by other groups and us showed an aberrant expression of MMPs in CD or UC, either on the protein or mRNA level, or immunohistochemically [15], [16], [17], [18], [19], [20], [21], [22]. In the present study, we evaluated the expression of MMP-2 and MMP-9 protein, both in level and localisation, and their mRNA gene products in relation to mucosal inflammation in patients with CD or UC.