| Background:At present,many studies have confirmed that Gli1 plays a key role in the formation and development of fibrosis,mainly manifested as abnormal activation.However,there were few studies on Gli1 in cardiovascular diseases,especially in myocardial infarction.Therefore,we hope to explore the molecular mechanism of GANT61 in reducing myocardial fibrosis after myocardial infarction by detecting the expression of Gli1 in ischemic myocardium in adult mice.Part one Explore the expression of Gli1 in myocardial tissue after myocardial infarctionObjective: Explore the expression of Gli1 in myocardial tissue after myocardial infarction by establishing the animal model of myocardial infarction in mice.Method:In 27 C57BL/6 male mice,a random number method was used to establish the model of myocardial infarction in 15 mice,and the elected mice were divided into myocardial infarction1 week group,myocardial infarction 2 weeks group and myocardial infarction 3 weeks group.9 rats were randomly selected as the sham operation group in theremaining 12 mice,divided into sham operation 1 week group,sham operation 2 weeks group and sham operation 3 weeks group.The last 3were used as normal group.The expression of Gli1 m RNA and protein in myocardial tissue of myocardial infarction and ischemic myocardium at the end of the first week,the second week and the third week after myocardial infarction were detected by real time quantitative PCR and western blot,and compared with normal group and sham operation group.The expression of Gli1 in myocardial infarction and peripheral ischemic myocardium was detected by immunofluorescence.Result: Comparison of myocardial infarction 2 weeks group,the expression of Gli1 m RNA in myocardial infarction group 3 weeks increased significantly(P<0.05),the difference was statistically significant;The expression of Gli1 m RNA in the sham operation group and normal group in the myocardial tissue showed no significant change(P>0.05),the difference was not statistically significant;And compared with 2 weeks after myocardial infarction,the expression of Gli1 protein the heart was increased 3 weeks after myocardial infarction,the results are similar to q RT-PCR;Immunofluorescence results showed the expression of Gli1 2weeks after myocardial infarction is less than 3 weeks after myocardial infarction.Conclusion: There were no Gli1 expression in the normal group and sham operation group,Gli1 was activated in the second week aftermyocardial infarction;With the extension of time,the expression of Gli1 m RNA and protein increased gradually.Part two Explore the molecular mechanism of GANT61 in alleviating myocardial fibrosis after myocardial infarction in adult miceObjective: Explore the molecular mechanism of GANT61 in alleviating myocardial fibrosis after myocardial infarction in adult mice.Method: In 80 C57BL/6 mice,60 mice were randomly selected to create the model of myocardial infarction by ligation of the left anterior descending coronary artery method.18 mice died after operation,only 42 survived.The surviving mice were randomly divided into myocardial infarction group,saline group and intervention group.The intervention group was treated with GANT61 at the dose of 50mg/kg/d by subcutaneous injection on the day of operation,saline group was injected with equal volume of normal saline.In addition,10 mice were randomly selected as sham operation group,not ligated the coronary artery after thoracotomy.The remaining 10 mice were used as normal group.The expression of Gli1,TGF-beta 1,type I and type III collagen protein and m RNA in the ischemia tissues of myocardial infarction and the border zone of myocardial infarction 1week,2weeks and 3weeks after operation were detected by using real-time fluorescence quantitative PCR and Western blot.And compared with the normal group and sham operation group.Masson staining was used to observe the morphology changes ofmyocardial infarction tissue.Result: Masson staining showed that the degree of myocardial fibrosis after myocardial infarction in mice was increased with the prolongation of time,and the myocardial fibrosis could be alleviated by GANT61intervention;The detection results of real-time fluorescent quantitative PCR and Western blotting showed that 3weeks after the GANT61 intervention can decrease the level of Gli1 m RNA and protein in tissues,compared with myocardial infarction 3 weeks group(P<0.05),the difference was statistically significant;Compared with the normal group,the m RNA and protein expression of TGF-beta 1,Collagen I and Collagen III increased slightly 2 weeks after myocardial infarction(P<0.05),the difference was statistically significant;After the GANT61 intervention for3 weeks,the m RNA and protein expression of TGF-beta 1,Collagen I and Collagen III were decreased,compared with the myocardial infarction 3weeks group(P<0.05),the difference was statistically significant.Conclusion: GANT61 can inhibit the expression of Gli1 in the HH pathway to alleviate myocardial fibrosis after myocardial infarction.It can be predicted that Gli1 may be a new molecular target for inhibiting myocardial fibrosis after myocardial infarction. |
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