| Severely traumatic hemorrhage can be a directly threaten to the life, often accompanied by systemic inflammatory response, in which could be harmful to some important organs and sometimes it could be lethal.Tumor necrosis factor (TNF-a) is a kind of endogenous regulatory factor which is produced by the stimulations. At present, the role of TNF-a in the wound healing process and in apoptosis of myocardium has attracted more and more attention.Notch signaling pathway, having influence on cell fate, is conserved signal transduction pathway, which determines the maintainance of normal structure and function in adult animal tissues and organs and the repairing after injury.ObjectIn this paper, we study the protective effect of Notch signaling pathway on myocardium in the model of traumatic hemorrhage, to screen drugs that can protect cardiac muscle cells from apoptosis caused by traumatic hemorrhage, which is helpful to clarify the mechanism of Notch signaling pathway in the process of injury and repair of myocardium. The purpose is to further understand the mechanism and treatment of trauma hemorrhage, and to explore new ideas, and to lay a foundation for the follow-up study.MethodPart one:Preliminary assessment of the effect of bone fracture injury on rat cardiac muscle cells and drug protection from apoptosis1 To establish a rat model of traumatic hemorrhage, and randomly divided into five groups, with 10 rats in each group. The experimental groups were fed with different drugs, and the control group and the trauma group normally diet.2 Elisa detected the level of 1L-2,IL-6, IL-10 and TNF-a in rats serum at Oh, 1h,2h, 4h,8h,12h,16h,24h,48h, to find the most significant point HE staining pathological cardiac injury in rats.3 To detect the expression of apoptosis gene in the myocardium of rats.4 The expression of Akt/Notch1/Hes1 signaling pathway in myocardium was detected.Part two:Low concentration TNF-a activates Akt/Notchl/Hesl signaling pathway and its role in the apoptosis of cardiac muscle cells1 To construct the model of myocardial apoptosis in vitro, and to detect the apoptosis of myocardial cells by Tunel method.2 To detect the protective effect of low concentration TNF-a on cardiac muscle cell apoptosis in vitro.3 The changes of Akt/Notchl/Hesl signaling pathway in the cells treated by low concentration TNF-a were detected by Western blot assay.4 To detect the effect of Akt/Notchl/Hesl signaling pathway activation on apoptosis while knockdown Notch 1.Part three:The protection and comparison of Rosuvastatin, Spironolactone and Irbesartan in myocardium apoptosis induced by trauma.1 To construct the model of myocardial apoptosis in vitro, and to detect the apoptosis of myocardial cells by Tunel method.2 Rt-PCR and Western blotting were used to detect the apoptosis of the myocardial cells treated with myocardial protective drugs and knocking down Notch 1 expression separately.ResultsPart one:1 At 4h, IL-10 and IL-6 were detected to reach the highest. IL-2 was detected to reach the lowest at 8h and TNF-a was detected at 1h the highest in the serum of rats.2 The results of HE staining showed that the degree of inflammation in the drug treatment group was significantly better than that in the trauma group, the macrophage was detected and cell morphology in the trauma group were significantly changed.3 The expression of Pro-apoptotic protein was decreased and the expression of anti-apoptosis protein was increased in the myocardial protective group.4 The Akt/Notch1/Hes1 signaling pathway was activated in the rat myocardial cells in the myocardial protective drugs given group.Part two:1 In the model of myocardial cell apoptosis in vitro, the trauma serum can lead to apoptosis.2 Low concentration TNF-a can inhibit the apoptosis of cardiac muscle cells.3 Low concentration TNF-a can activate Akt/Notch1/Hes1 signaling pathway.4 Activated Akt/Notchl/Hesl signaling pathway can protect cardiac muscle cells from apoptosis.Part three:1 In the model of myocardial cell apoptosis in vitro, myocardial protective drugs can protect cardiac muscle cells from apoptosis.2 Knocking down Notch1 expression, in groups of spironolactone and rosuvastatin the apoptosis was partially weakened, but in irbesartan did not change.Conclusion1 Compared with traumatic model, the TNF-a in the myocardial protective drugs given groups are elevated and could last for a relatively long time.2 Low concentration TNF-α can activate the Akt/Notchl/Hesl signaling pathway, and protect cardiac muscle cells from apoptosis induced by trauma.3 Myocardial protective drugs (Spironolactone, Rosuvastatin and Irbesartan) can protect myocardial cells from injury, Spironolactone and Rosuvastatin is mainly through upregulating of TNF-a, so as to activate the Akt/Notchl/Hesl signaling pathway, and ultimately protect cardiomyocytes from apoptosis caused by trauma. It seems that the mechanism of Irbesartan’s anti-apoptosis of myocardial cell was not mediated by Akt/Notchl/Hesl signaling pathway activation. And its specific mechanism remains to be further study. |
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