title: Matrix metalloproteinase-9 in homocysteine-induced intestinal microvascular endothelial paracellular and transcellular permeability.
abstract: Although elevated levels of homocysteine (Hcy), known as hyperhomocysteinemia (HHcy), is associated with inflammatory bowel disease (IBD), the mechanism of Hcy action is unclear. In the present study, we tested the hypothesis that HHcy activates matrix metalloproteinase-9 (MMP-9), which in turn enhances permeability of human intestinal microvascular endothelial cell (HIMEC) layer by decreasing expression of endothelial junction proteins and increasing caveolae formation. HIMECs were grown in Transwells and treated with 500 µM Hcy in the presence or absence of MMP-9 activity inhibitor. Hcy-induced permeability to FITC-conjugated bovine serum albumin (FITC-BSA) was assessed by measuring fluorescence intensity of solutes in the Transwells' lower chambers. The cell-cell interaction and cell barrier function was estimated by measuring trans-endothelial electrical impedance. Confocal microscopy and flow cytometry were used to study cell junction protein expressions. Hcy-induced changes in transcellular transport of HIMECs were estimated by observing formation of functional caveolae defined as caveolae labeled by cholera toxin and antibody against caveolin-1 and one that have taken up FITC-BSA. Hcy instigated HIMEC monolayer permeability through activation of MMP-9. The increased paracellular permeability was associated with degradation of vascular endothelial cadherin and zona occludin-1 and transcellular permeability through increased caveolae formation in HIMECs. Elevation of Hcy content increases permeability of HIMEC layer affecting both paracellular and transcellular transport pathways, and this increased permeability was alleviated by inhibition of MMP-9 activity. These findings contribute to clarification of mechanisms of IBD development.
authors.affiliation: Department of Physiology and Biophysics, University of Louisville School of Medicine, Louisville, KY 40202, USA.
authors.forename: Charu
authors.initials: C
authors.lastname: Munjal
authors.forename: Neetu
authors.initials: N
authors.lastname: Tyagi
authors.forename: David
authors.initials: D
authors.lastname: Lominadze
authors.forename: Suresh C
authors.initials: SC
authors.lastname: Tyagi
mesh_terms: D000199:Actins / Q000096:biosynthesis; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000201:enzymology / Q000378:metabolism*; D004789:Enzyme Activation; D005434:Flow Cytometry; D006710:Homocysteine / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D007422:Intestines / Q000098:blood supply* / Q000201:enzymology; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D055806:Microvessels / Q000201:enzymology / Q000378:metabolism*; D010539:Permeability; D010750:Phosphoproteins / Q000378:metabolism; D019108:Tight Junctions; D062826:Zonula Occludens-1 Protein
chemical_list: D000199:Actins; D008565:Membrane Proteins; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D006710:Homocysteine; D020780:Matrix Metalloproteinase 9