GABA receptors ameliorate Hcy-mediated integrin shedding and constrictive collagen remodeling in microvascular endothelial cells.
Mammalian endothelial cells are deficient in cystathionine beta synthetase (CBS) activity, which is responsible for homocysteine (Hcy) clearance. This deficiency makes the endothelium the prime target for Hcy toxicity. Hcy induces integrin shedding in microvascular endothelial cells (MVEC) by increasing matrix metalloproteinase (MMP). Hcy competes with inhibitory neurotransmitter gamma aminobutyric acid (GABA)-A receptor. We hypothesized that Hcy transduces MVEC remodeling by increasing metalloproteinase activity and shedding beta-1 integrin by inactivating the GABA-A/B receptors, thus behaving as an excitatory neurotransmitter. MVEC were isolated from mouse brain. The presence of GABA-A receptor was determined by immunolabeling. It was induced by muscimol, an agonist of GABA-A receptor as measured by Western blot analysis. Hcy induced MMP-2 activity in a dose- and time-dependent manner, measured by zymography. GABA-A/B receptors ameliorated the Hcymediated MMP-2 activation. Hcy selectively increased the levels of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-3 but decreased the levels of TIMP-4. Treatment with muscimol decreased the levels of TIMP-1 and TIMP-3 and increased the levels of TIMP-4 to control. Hcy caused a robust increase in the levels of a disintegrin and metalloproteinase (ADAM)-12. In the medium of MVEC treated with Hcy, the levels of beta-1 integrin were significantly increased. Treatment with muscimol or baclofen (GABA-B receptor agonist) ameliorated the levels of beta-1 integrin in the medium. These results suggested that Hcy induced ADAM-12. Significantly, Hcy facilitated the beta-1 integrin shedding. Treatment of MVEC with muscimol or baclofen during Hcy administration ameliorated the expression of metalloproteinase, integrin-shedding, and constrictive collagen remodeling, suggesting a role of Hcy in GABA receptor-mediated cerebrovascular remodeling.
Shastry
Suresh
S
Department of Physiology and Biophysics, University of Louisville School of Medicine, Louisville, KY, USA.
Tyagi
Neetu
N
Moshal
Karni S
KS
Lominadze
David
D
Hayden
Melvin R
MR
Tyagi
Suresh C
SC
D051722:ADAM Proteins / Q000378:metabolism; D000072199:ADAM12 Protein; D000818:Animals; D001921:Brain / Q000166:cytology / Q000378:metabolism; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006710:Homocysteine / Q000494:pharmacology*; D016023:Integrins / Q000378:metabolism*; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018377:Neurotransmitter Agents / Q000502:physiology; D018079:Receptors, GABA / Q000378:metabolism*; D011963:Receptors, GABA-A / Q000378:metabolism; D018080:Receptors, GABA-B / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism; D019717:Tissue Inhibitor of Metalloproteinase-3 / Q000378:metabolism; D019714:Tissue Inhibitor of Metalloproteinases / Q000378:metabolism*
D016023:Integrins; D008565:Membrane Proteins; D018377:Neurotransmitter Agents; D018079:Receptors, GABA; D011963:Receptors, GABA-A; D018080:Receptors, GABA-B; D019715:Tissue Inhibitor of Metalloproteinase-1; D019717:Tissue Inhibitor of Metalloproteinase-3; D019714:Tissue Inhibitor of Metalloproteinases; D006710:Homocysteine; D003094:Collagen; D051722:ADAM Proteins; D000072199:ADAM12 Protein; D020782:Matrix Metalloproteinases; D020778:Matrix Metalloproteinase 2