Zonulin, regulation of tight junctions, and autoimmune diseases

Abstract

Recent studies indicate that besides digestion and absorption of nutrients and water and electrolytes homeostasis, another key function of the intestine is to regulate the trafficking of environmental antigens across the host mucosal barrier. Intestinal tight junctions (TJs) create gradients for the optimal absorption and transport of nutrients and control the balance between tolerance and immunity to nonself antigens. To meet diverse physiological challenges, intestinal epithelial TJs must be modified rapidly and in a coordinated fashion by regulatory systems that orchestrate the state of assembly of the TJ multiprotein network. While considerable knowledge exists about TJ ultrastructure, relatively little is known about their physiological and pathophysiological regulation. Our discovery of zonulin, the only known physiologic modulator of intercellular TJs described so far, has increased our understanding of the intricate mechanisms that regulate the intestinal epithelial paracellular pathway and has led us to appreciate that its upregulation in genetically susceptible individuals leads to autoimmune diseases.

Figure 1

Western blotting using zonulin cross-reacting anti-Zot polyclonal antibodies on CD patient sera showed three main patterns: sera showing a 18 kDa immunoreactive band and a fainter _45 kDa band (lane 1), sera showing only a 9 kDa band (lane 2), and sera showing both the 18 and 9 kDa bands (lane 3). The cartoon shows the structure of both pre-haptoglobin (HP) 1 and pre-HP2 and their mature proteins. HPs evolved from a complement-associated protein (mannose-binding lectin-associated serine protease, MASP), with their α-chain containing a complement control protein (CCP), while the β chain is related to chymotrypsin-like serine proteases (SP domain) containing an epidermal growth factor-like motif. The gene encoding the α2-chain of pre-HP2 originated in India almost 2 million years ago through a chromosomal aberration (unequal crossing over) of HP1. Pre-HPs are translated as single-chain precursor proteins. Pre-HPs may be proteolytically cleaved intracellularly into α - and β-chains that remain disulfide linked, referred to as cleaved, two-chain mature HPs.

Figure 2

Zonulin can activate EGFR through direct binding (1) and/or through PAR2 transactivation (2). This second mechanism can be mediated by either Src signaling (2a) or by the release of MMPs and/or ADAMS that in turn will activate Pro-HB-EGF. When cell tryptase IV cleaves zonulin in its two subunits (so eliminating one of the three required disulfide bridges necessary for EGF activity), the molecule is not able to bind to EGFR (3), while will acquire a different function (Hb binding) and becomes an inflammatory marker.

Figure 3

(A). CD patients showed higher serum zonulin levels compared to both their relatives and controls. (B). Similar results were obtained in T1D patients. (C). Serum zonulin correlated with intestinal permeability evaluated by the LA/MA test. The percentage of CD patients (81%) and their relatives (50%) with elevated serum zonulin levels was higher compared to T1D patients (42%) and their relatives (29%), respectively.