Homocysteine induces metalloproteinase and shedding of beta-1 integrin in microvessel endothelial cells.
Although studies have suggested microvessel endothelial cells (MVEC) activation and induction of matrix metalloproteinases (MMPs) by homocysteine (Hcy), the transduction mechanism leading to endothelial activation was unclear. We hypothesized that Hcy induced metalloproteinase and altered the levels of integrin in MVEC. MVEC from mouse brain were isolated and characterized by CD-31 (PECAM-1) FITC labeling. The MVEC were activated with different doses (6-40 microM) of Hcy. The cultured-conditioned-medium was analyzed for MMP activity by gelatin gel-zymography. TIMP-1, -4, beta-1 integrin, and a disintegrin and metalloproteinase-12 (ADAM-12) were quantified by Western blot analysis. We used MVEC in cell culture to study the effect of increasing concentrations of Hcy upon the secretion of various proteins into the culture medium. MMP-9, beta-1 integrin, ADAM-12, and TIMP-1 were found in increased concentrations in the culture medium of Hcy-treated cells whereas TIMP-4 was decreased. We have shown that purified TIMP-4 blocked the increase of beta-1 integrin shedding in Hcy-treated cells. Interestingly, our results suggest that TIMP-1 and TIMP-4 function antagonistically in Hcy-induced signaling pathways.
Shastry
Suresh
S
Department of Physiology and Biophysics, University of Louisville School of Medicine, Louisville, Kentucky 40202, USA.
Tyagi
Suresh C
SC
D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002196:Capillaries; D019483:Disintegrins / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism; D006710:Homocysteine / Q000494:pharmacology*; D019012:Integrin beta1 / Q000378:metabolism*; D053508:Matrix Metalloproteinase 12; D008666:Metalloendopeptidases / Q000378:metabolism*; D051379:Mice; D015398:Signal Transduction; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism*; D019714:Tissue Inhibitor of Metalloproteinases / Q000378:metabolism*; D000097792:Tissue Inhibitor of Metalloproteinase-4
D019483:Disintegrins; D006710:Homocysteine; D019012:Integrin beta1; D053508:Matrix Metalloproteinase 12; D008666:Metalloendopeptidases; D019715:Tissue Inhibitor of Metalloproteinase-1; D019714:Tissue Inhibitor of Metalloproteinases; D000097792:Tissue Inhibitor of Metalloproteinase-4