| Myocardial fibrosis refers to the excessive accumulation of collagen fibers in the normal tissues and structures, marked elevation of collagen concentration, or the changes of collagen constituents. This pathological change exists in many cardiovascular diseases. Now, it is believed that this change is associated tightly with cardiac arrhythmia, cardiac dysfunction and even cardiogenic sudden death.There are many reasons that can cause myocardial fibrosis. Cardiac interstitium has the possibility of being involved primarily, but in most already known cases, the starting point of myocardial fibrosis is the damage to cardiac parenchymal cells due to various reasons, that is to say, degeneration, necrosis and apoptosis of cardiac cells or only the changes in subcellular structure, biochemistry and metabolism, activate corresponding macrophages and other cells which produce and secrete many kinds of active factors(cytokines, growth factors, etc ) , which in turn, activate ECM-producing cells(various cardiac interstitial cells) in the cardiac tissues. Recently, research of mechanism about the formation of myocardial fibrosis has manifested that myocardial fibrosis is a process in which occurs the mutual facilitation and restriction of lots of factors, among which the imbalance of cytokines and growth factors, mainly the TGF-β1 and CTGF, play a very important role. In this experiment, we replicated a model of myocardium necrosis caused by ischemia and hypoxia in rats by using isoprenaline. Then we studied the expression of cytokines TGF-β1 and CTGF in the early stage of cardiac necrosis caused by ischemia and hypoxia, by means of immunohistochemistry and RT-PCT, as well as the changes in expression of these cytokines as the time of ischemia and hypoxia went on. As a result, we can make clear the functions of TGF-β1 and CTGF in the occurrence and development of myocardial fibrosis. Besides, we can offer a new therapeutic target for the reversion of myocardial fibrosis and alleviation of myocardium reconstruction. The experimental results are as follows:1 Replication of myocardium necrosis caused by ischemia and hypoxia in rats.In the serum-enzyme testing, it was found that 6h after isoprenaline injection, CK and CK-MB had the tendency of elevation, and in 24h, their activities reached the maximum. Later, AST and LDH also started to increase from 6h after injection until 48h. Morphological results further testified that small amounts of spotty necrosis could be seen under the endocardium 6h after the injection, obvious small necrotic focuses appeared in 12h (as the time of ischemia elongated, the degree of necrosis aggravated ) , and sharply-defined, diffuse, sporatic, multiple necrotic focuses appeared in 1w. The results showed that models of myocardium necrosis caused by ischemia and hypoxia could be successfully replicated in this way.2 Relation between TGF-β1 , CTGF and myocardial fibrosis.We studied the expression of TGF-β1 and CTGF in the occurrence and development of myocardial fibrosis, by means of immunohistochemistry. The results were as follows:Results from immunohistochemistry revealed that myocardium of rats in the normal control group did not have obvious positive expression of TGF-β1. 6h after injection of isoprenaline, expression of TGF-β1 in myocardium of rats in the experimental group was similar to that in the normal control group. 12h after the injection, small amounts of TGF-β1 was expressed under the endocardium. As the time after injection became longer, the degree of fibrosis became severer, the expression of TGF-β1 increased, and the extent enlarged apparently. 3w after the injection, expression of TGF-β1 reached the maximum. Results from RT-PCT revealed that expression of TGF-β1 mRAN in the experimental group began to increase 6h after the injection of isoprenaline. Compared to normal control group, the elevation was obvious, with p<0.05. Furthermore, expression of TGF-β1 mRNA reached the maximum 3w after injection of isoprenaline, apparently higher than that of the normal control group, with p<0.05.Results of CTGF from immunohistochemistry were similar to those of TGF-β1. In normal control group, the expression of CTGF was negative, even 6h after the injection of isoprenaline. Small amounts of CTGF were expressed under the endocardium 12h after the injection. The expression increased as the time after injection became longer, and reached the maximum in 1w. Results of CTGF from RT-PCR were also similar to those of TGF-β1. It started to express in 6h, and reached the maximum in 3w.Comparing the results of TGF-β1 and CTGR, the distribution locations of these two cytokines were close to each other, both started the expression from the endocardium, then strengthened the positive expression, and finally extended to the mesocardium and epicardium. Results from RT-PCR showed that expression in molecular lever is consistent with change in the histology, indicating that expression increase of these two cytokines was duo to the increase of transcription of genes. At the same time, expression of these two cytokines had the same tendency, indicating that both of them are coupled to express in muscle fibrosis, possibly with synergy effect.From the above mentioned, we can see that myocardial fibrosis caused by ischemia and hypoxia starts in the early stage after the injection of isoprenaline. As the time elongates, the degree of myocardial fibrosis aggravates. Expressions of TGF-β1 and CTGF changes from negative to positive, increases as time goes on, and have similarity in the expression location and degree, through out the whole process of myocardium necrosis caused by ischemia and hypoxia. It is manifested that positive expressions of TGF-β1 and CTGF are closely related to the occurrence and development of myocardial fibrosis, and the fibrosis is mediated by them.
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