| Acute coronary syndromes such as myocardial infarctions are usually caused by the eruption of the instable plaque of the atherosclerosis. However, the increasing combination of matrix metalloptoteinases( MMPs) in the plaque of atherosclerosis and /or the decrease of MMP inhibitors can serve as one main mechanism in deciding the decline of stability of the atherosclerotic plaque. By degrading the extracelluar matrix MMPs can promote the monocytes in the blood and the smooth muscle cells in the mediuthelium migrating towards the endothe-lium as well as the process of atherosclerosis. Besides, MMPs could also promote degradation of the fibrous cap of the atherosclerotic plaque and the formation of angiogenesis, leading the rupture of the plaque and hemorrhage, and ultimately the occurrence of acute coronary syndrome. Recently, the role that the inflammatory factors play in the atherosclerosis begins to catch people's attention, and put forward the atherosclerosis inflammation theory. Lots of research showed that many inflammatory factors such as IL - 1, TNF - a etc can promote the secretion of MMPs in the plaque.Currently, researches about the influence brought by granular - macrophage colony stimulating factor ( GM - CSF) on the MMPs expression and activity are quite rare. Thus this experiment intended to discuss the relation between inflammation factor GM - CSF and the formation and stability of the atherosclerosis by observing the influence the GM - CSF brought on the expression and activity of MMP - 2 in vitro cultured human vascular endothelial cells.MethodFirstly, HUVECs were cultured by modified Jaffe method and adding the stimulating factors when the cells were sub - confluent. Group one, dividing HUVECs into four groups, normally control( serum - free DMEM ) and GM -CSF stimulating groups in varying concentrations ( 5 25 50ng/ml ) then collecting the culture supematants after 24 hours. Group two, dividing HUVECs into four groups, normally control ( serum - free DMEM ) and the same concentration GM - CSF in varying stimulating time. Then culture supematants were collected , cleared of debris by centrifugation. Gelatin zymography was used to determine the activity of MMP - 2 in the supematants of each group; Western Blot was used to determine the protein amounts of MMP - 2 in the supematants of each group.ResultsThe activity of MMP - 2 was determined by SDS - PAGE containing gelatin had shown that, after treatment with GM - CSF in varying concentrations ( 5 25 50ng/ml ), the activity of MMP - 2 in the culture media of the ECs were higher compared with untreated controls in a dose - dependent manner; and after treatment with GM - CSF of the same concentration ( 50ng/ml ) but at varying time ( 6 12 24h ) , the activity of MMP -2 gradually increased showing a time - dependent manner. The analysis of Western Blot had shown that the 24 hour - attendance of varying concentration GM - CSF ( 5 25 50ng/ml ) on HUVECs, the amount of MMP - 2 in culture media is higher than untreated control in a dose - dependent manner; and after treatment with GM - CSF of the same concentration ( 50ng/ml ) but at varying time ( 6 12 24h ) , the protein amounts of MMP - 2 gradually increased showing a time - dependent manner. According to molecular weight of the MMP - 2, the MMP - 2 tested in this experiment is mainly its proMMP - 2 form.DiscussionThe MMPs are a family of zinc - dependent endopeptidases which are capable of degrading all know extracellular matrix (ECM ) components. This family is mainly composed of the collagenases, the stromelysins, the gelatinases and the membrane - type MMPs ( MT - MMPs ). They were synthesized and secreted in the form of proenzyme by lots of cells such as monocytes vascular smooth muscle cells vascular endothelial cells and fibroblast etc. MMP -2, know as gelatinase A, is the most important MMPs for vascular cells expression and secretion. Once activated, it can degrade extracelluar matrix components such as gelatin collagens and basement membrane components.The ruptu...
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