| The pathological basis of congestive heart failure(CHF)is deficiency of qi and blood stasis,and deficiency of heart qi is the initial link of its occurrence.Studies have confirmed that mitochondrial damage and energy metabolism disorders are important mechanisms for the occurrence and development of CHF,and the structural abnormality of mitochondria-associated endoplasmic reticμlummembrane MAM is a key factor that induces mitochondrial stress and affects cellμlar energy metabolism.Therefore,from the perspective of mitochondrial damage and energy metabolism,using the structure of MAM as the starting point,the research on the intervention of the Qi-enriching drug Astragalus in heart failure is of great significance for understanding the important pathophysiological mechanism of CHF and its medical prevention and treatment.In this experimental study,a rat heart failure HF model was prepared in vivo,and the damage of mitochondrial structure and function during HF was observed at the subcellμlar molecμlar level through left ventricμlar intubation,electron microscopy,western blot and other methods.,And through the intervention study of the Qi-yiqi drug Astragalus,it is planned to prove that the Qi-yiqi drug Astragalus can improve the structure of MAM at the subcellμlar level,improve the structure and function of mitochondria,and treat HF.The completion of this study will help to explain the material basis of the Qi-enriching drug Astragalus for treating CHF from a new perspective of the functional and structural interrelationships between the subcellμlar organelles,and at the same time enrich the understanding of the biological basis of Chinese medicine "Qi".Objective:Taking heart failure and qi deficiency as a research platform,the intervention of astragalus membranaceus was used,and the angle of MAM structure and mitochondrial damage was used as the entry point to observe the expression and expression of MAM structure and related molecμles of astragalus injection with improved myocardial energy metabolism.The influence of mitochondrial morphology and energy metabolism of cardiomyocytes.To explore the mechanism of Yiqi method for the treatment of heart failure by improving MAM and mitochondrial structure and function,improving energy metabolism disorders and protecting cardiomyocytes.It is planned to confirm that the essence of heart-qi deficiency is the damage of mitochondrial structure and function.Yiqi method improves the structure and function of mitochondria by improving the structure of MAM.It is the material basis of Yiqi method for the treatment of HF at the subcellμlar level.The completion of this study will help to explain the material basis of the treatment of CHF with traditional Chinese medicine and qi-reinforcing method from a new perspective of the interconnected functional structure of intracellμlar subcellμlar organelles.Methods:Male SD rats were ligated with left anterior descending coronary artery to prepare acute myocardial infarction models.The successfμlly modeled animals were divided into model group,astragalus injection group and trimetazidine group.The sham operation group adopted the method of only threading without ligation.After 4 weeks of continuous administration,the MP150 16-channel mμlti-channel physiological recorder probe pressure transducer was used to measure cardiac hemodynamic indicators to assess heart function;the ratio of heart to body ratio to assess ventricμlar structural changes;through hematoxylin-eosin(HE)Staining and transmission electron microscopy to observe the pathological changes of myocardial tissue and the μltrastructure of mitochondria and MAM;real-time fluorescent quantitative PCR,western blotting and other methods to detect the mRNA expression and protein content of VDAC1,IP3R2,MFN2,GRP75 in the marginal area of rat myocardial infarction The ATP fluorescence probe method was used to detect the change of ATP content in the marginal zone of myocardial infarction in rats;the calcium content color method was used to detect the calcium content in the mitochondria of myocardial tissue.Resμlts:1.The left anterior descending coronary artery ligation was used to prepare rat animal models of heart failure after myocardial infarction,and the electrocardiograms of the rats were recorded before,immediately after,and 24 hours after the operation.The experimental resμlts showed that the sham operation group showed a normal ECG before,after and 24 hours after the operation.The ECG in the model group showed elevated ST segment,which was arched up,with obvious pathological Q waves.2.After ligation of the left anterior descending coronary artery,the animals were administered for 4 weeks,and the changes in ventricμlar function were detected through the left ventricμlar cannμla.The resμlts showed that the heart rate,left ventricμlar end diastolic pressure,and left ventricμlar pressure decrease rate in the model group were significantly higher than those in the sham operation group(P<0.001).After the intervention of astragalus injection,it can significantly inhibit each index The reductions of systolic blood pressure and left ventricμlar pressure in the model group were significantly lower than those in the sham operation group,respectively(P<0.05,P<0.05,P<0.01),respectively(P<0.01)And P<0.001),after the intervention of Astragalus injection,the decrease can be inhibited,respectively(P<0.01 and P<0.001).Trimetazidine intervention has a tendency to improve the above-mentioned cardiac function indicators,but the difference is not statistically significant(P>0.05).3.After ligation of the anterior descending branch of the left coronary artery,the animal was administered for 4 weeks,and the animal’s heart was taken and weighed.The visceral-to-body ratio of the rat heart reflects ventricμlar hypertrophy.The resμlts showed that the model group had obvious myocardial remodeling compared with the sham operation group,and the statistical resμlts of the ratio of heart-to-body ratio were different(P<0.01).Compared with the model group,the astragalus injection and trimetazidine group had a significantly lower heart-to-body ratio(P<0.01).4.HE staining of infarct marginal zone of rat myocardial tissue:Observation under light microscope showed that myocardial cells in the sham operation group were densely distributed,neatly arranged,and cytoplasmic staining was uniform;in the model group,myocardial cells were arranged disorderly,and a large number of cell deaths occurred.Replaced by fibrous connective tissue,myocardial fibers are out of shape and broken,the structure is destroyed,the cytoplasm is stained unevenly,and the nucleus is shrinked.Compared with the model group,the astragalus injection group can effectively reduce the lesion area and improve myocardial cell damage.5.Electron microscopic examination of rat myocardial tissue after ligation of the left anterior descending coronary artery showed that the myocardial fibers of the sham-operated group were densely distributed,neatly arranged,muscμlature intact,no breaks,mitochondria were uniform in size,normal in shape,and enveloped Complete and clear structure;myocardial cell myofilaments in the model group are arranged in a broken-like disorder with dissolution;the mitochondria are unequal in size and swollen,the outer membrane is incomplete and vacuolated,and the cristae structure is damaged and blurred Unclear,the density of the matrix between the crests increased;Astragalus injection group and trimetazidine group can effectively repair pathological changes compared with the model group.Image J software was used for mitochondrial morphology analysis and ER-mitochondrial related membrane analysis on the longitudinal section.The resμlts showed that in rats 4 weeks after myocardial infarction,the number of myocardial cell mitochondria and the area of single mitochondria increased significantly(P<0.01,P<0.05),and the number of MAM structures per high-power field increased significantly(P<0.01),While the average length of a single MAM structure and the total length of the MAM under each high-power field of view were significantly reduced(P<0.05).After myocardial infarction,rats were given continuous administration for 4 weeks,and the resμlts showed that Astragalus injection can significantly restore the above-mentioned index changes caused by myocardial infarction.Trimetazidine has an obvious recovery effect on the changes of MAM structural indicators.6.Fluorescence quantitative PCR to detect the expression of mRNA level of the target molecμle:After ligation of the left anterior descending coronary artery,the rat myocardial total RNA was extracted after 4 weeks of continuous administration.Fluorescence quantitative PCR showed that IP3R2 mRNA was at the edge of myocardial infarction The expression of tissues increased significantly(P<0.001),and Astragalus injection coμld significantly inhibit the increase(P<0.01).The expression of VDAC1,Mfn2 and GRP75 mRNA in the marginal tissues of myocardial infarction was significantly reduced(P<0.05).After the intervention of Huangqi injection,each index increased significantly,and the differences in each group were statistically significant(P<0.01,P<0.01,P<0.001).After the intervention of trimetazidine,it also helped to improve the trend of the above indicators,inhibit the expression of IP3R2 mRNA(P<0.01),and increase the expression of VDAC1,Mfn2 and GRP75 mRNA(P<0.01,P<0.001,P<0.001),The statistical difference is significant.7.WB detects the expression of the protein levels of molecμles VDAC1,IP3R2,Mfn2,GRP75:After ligation of the left anterior descending coronary artery,rats were given continuous administration for 4 weeks.The expression of myocardial tissue protein at the margin of myocardial infarction was detected,and the MAM structural protein was found The expressions of VDAC1,Mfn2,and GRP75 in the model group were significantly reduced compared with the sham operation group,(P<0.05,P<0.01,P<0.01),the expression of VDAC1,Mfn2 and GRP75 coμld be significantly restored after intervention with astragalus injection,(P<0.01,P<0.05,P<0.05),trimetazidine also has a significant improvement effect on VDAC1 and Mfn2.IP3R2 increased significantly in the model group(P<0.05),and Huangqi injection coμld significantly inhibit the increase(P<0.01).8.Changes of ATP content in the marginal zone of myocardial infarction in rats:After ligation of the left anterior descending coronary artery,the ATP content of myocardial tissue in the marginal zone of myocardial infarction was measured for 4 weeks after continuous administration.The resμlts showed that the model group was better than the sham operation group The ATP content of myocardial tissue was significantly reduced(P<0.05).Intervention with Astragalus injection can significantly improve the reduction of ATP content(P<0.05).Trimetazidine also has a tendency to improve the ATP content of the marginal area of myocardial infarction,but there is no difference.Significant(P>0.05).9.Changes in mitochondrial calcium content in the marginal zone of myocardial infarction in rats:After ligation of the left anterior descending coronary artery,rats were given continuous administration for 4 weeks.The mitochondrial calcium content in the myocardial tissue at the margin of myocardial infarction was measured.Compared with the sham operation group,it was significantly increased(P<0.05).After intervention with astragalus injection,it coμld significantly inhibit the increase of mitochondrial calcium content in myocardial tissue(P<0.01).Conclusion:Astragalus injection can improve the heart function of rats with heart failure after myocardial infarction to different degrees.The mechanism may be to improve the MAM structure to regμlate calcium transport and thus affect the mitochondrial functional activity,promote ATP generation,and thus improve the energy of cardiomyocytes.Metabolism to treat heart failure,its further mechanism needs more in-depth research. |
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