Role of endoplasmic reticulum calcium disequilibria in the mechanism of homocysteine-induced ER stress.
Our laboratory demonstrated that hyperhomocysteinemia accelerates atherosclerosis in mouse models through ER stress and activation of the unfolded protein response (UPR). In this study, we tested the hypothesis that homocysteine-induced ER stress may arise from ER-Ca(2+) disequilibria. We found that homocysteine-induced cytosolic Ca(2+) transients in T24/83 cells and human aortic smooth muscle cells (HASMCs). These calcium effects occurred at concentrations of homocysteine in the external medium (1-5 mM) that increase intracellular homocysteine in these cell types. Prolonged homocysteine treatment (5 h) at these exogenous concentrations reduced ER-Ca(2+) emptying evoked by thapsigargin. However, these homocysteine-induced effects on ER-Ca(2+) emptying were of a much smaller magnitude than those evoked by A23187 or thapsigargin (ER stressors known to induce ER stress through ER-Ca(2+) depletion). T24/83 cells stably overexpressing the Ca(2+)-binding ER chaperone GRP78 showed diminished cytosolic Ca(2+) transients induced by homocysteine and reduced ER-Ca(2+) emptying evoked by thapsigargin. Prevention of the homocysteine-induced UPR by cycloheximide pretreatment normalized GRP78 expression and ER-Ca(2+) emptying evoked by thapsigargin. These results are inconsistent with a mechanism of ER stress induction by homocysteine through ER-Ca(2+) depletion.
Dickhout
Jeffrey G
JG
Department of Pathology and Molecular Medicine, McMaster University, and the Henderson Research Centre, Hamilton, Ontario, Canada.
Sood
Sudesh K
SK
Austin
Richard C
RC
D001011:Aorta / Q000166:cytology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000032:analysis / Q000172:deficiency* / Q000378:metabolism; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes; D016257:Fura-2; D006360:Heat-Shock Proteins / Q000378:metabolism; D006710:Homocysteine / Q000494:pharmacology*; D006801:Humans; D007476:Ionophores / Q000494:pharmacology; D018832:Molecular Chaperones / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects*; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors; D001749:Urinary Bladder Neoplasms / Q000473:pathology
D000091342:Endoplasmic Reticulum Chaperone BiP; D005456:Fluorescent Dyes; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; C000717248:Hspa5 protein, mouse; D007476:Ionophores; D018832:Molecular Chaperones; D006710:Homocysteine; D000001:Calcimycin; D019284:Thapsigargin; D002118:Calcium; D016257:Fura-2