Autophagosome activity in macrophage for atherosclerotic plaques in ApoE^-/-mice enhanced by Tiaozhi Tongmai Granules

Objective:Atherosclerosis is the fundamental pathophysiologic component of cardiovascular disease,and Tiaozhi Tongmai Granules show great efficiency in the treatment of the disease.However,the mechanism of Tiaozhi Tongmai Granules is still unclear.In this study,we have combined experiments with network pharmacology to explore the antiatherosclerosis mechanism of Tiaozhi Tongmai Granules.Method:120 male ApoE
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mice were randomly divided into three groups:the model group,Chinese herb group and Atorvastatin group.The model group,Atorvastatin group and Chinese herb group were fed with a high-fat diet,a high-fat diet plus atorvastatin(5.1 mg/kg/d)and a high-fat diet plus Tiaozhi Tongmai Granules(16.5 g/kg/d)for 16 weeks,respectively.Atherogenesis was identified by H&E staining.The colocalization of neutral lipid stain BODIPY and microtubule-associated protein 1 light chain 3(LC3)and the colocalization of BODIPY and lysosomal-associated membrane protein 1(LAMP1)within ApoE
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mice aortic plaques were tested using fluorescence confocal microscopy and the Pearson’s coefficients were calculated.To further explore the anti-atherosclerosis mechanism of Tiaozhi Tongmai Granules,the network pharmacology was used to construct the herb-compound-target network.Results:The size of the aortic lipid plaque in the Chinese herb group and Atorvastatin group were smaller compared with the model group on the 16th week.Compared with the model group on the 16th week,the BODIPY and LC3 colocalization rate,the BODIPY and LAMP1 colocalization rate of the Chinese herb group and Atorvastatin group all presented significant increase in the aortic plaque(P<.001),showing that Tiaozhi Tongmai Granules could enhance autophagosome activity in the macrophage.In the herb-compound-target network,17 active compounds and 27 targets were obtained through literature searching and using LHRI&DAVID Bioinformatics.It was found that 23 targets were correlated with the macrophage.Some of them participated in macrophage inflammatory response,and the other targets could promote/inhibit phagocytosis of the macrophage.It was hypothesized that the active compounds of Tiaozhi Tongmai Granules were acting on these targets and having y the biological effects.Conclusions:In the progression stage of atherosclerosis,Tiaozhi Tongmai Granules can still make the macrophage have higher autophagosome activity,and play a role of anti-atherogenesis.

Yiqi Huoxue prescription can prevent and treat post-MI myocardial remodeling through promoting the expression of AMPK signal pathway

Objective:To investigate how Yiqi Huoxue (YQHX) prescription regulates mitochondrial biosynthesis and ATP synthesis via AMP-activated protein kinase (AMPK) and to reveal its molecular mechanism in preventing and treating post-MI myocardial remodeling.Methods:The MI animal model of myocardial infarction were established by ligating Sprague Dawley (SD) rats' left anterior descending coronary arteries;the animals were randomly divided into MI group,YQHX prescription group and perindopril group,and a sham operation group was set at the same time.Related drug intervention was administered on the 2nd day after surgery,the YQHX group was given Astragalus,angelica,ginseng,Ligusticum wallichii and pseudoginseng,provided by the Dongzhimen Hospital of Beijing University of Chinese Medicine,once per day,at a dose of 21 g/kg body weight/day (the clinical equivalent dose based on a previous study),and the changes in relevant indicators were observed at 1 week and 4 weeks.Echocardiography (ECG) was used to observe the changes in rat cardiac structure and functions;the morphology-based technique was used to observe the changes in myocardial cells and mitochondria;the expression of AMPK signal pathway-related proteins and mRNA was detected using western blotting and real-time fluorescence quantification respectively,while fluorescence enzyme-labeled method was used for detecting ATP synthesis.Results:Cardiac structure and functions:Compared with the MI group at 1 week,the YQHX prescription group and the perindopril group exhibited increased LVEF and LVFS (Pall <.05);their LVEDs and LVEDd were reduced but suggested no statistically significant differences (F =2.258,F =0.3464,Pall >.05).At 4 weeks,both LVEF and LVFS were elevated in the YQHX prescription group (P =.008,.009) and the perindopril group (P =.279,.333),where differences in the later group indicated no statistical significance;the YQHX prescription group and the perindopril group were also featured by reduced LVEDs and LVEDd (Pall <.05).Morphology:Compared with the MI group at two time nodes,the YQHX prescription group and the perindopril group exhibited significant improvement in the pathological changes in myocardial cells and mitochondrial structure.Expression of AMPK signal pathway-related proteins and mRNA:The expression of both pLKB1 and pAMPK proteins followed a rising trend in the YQHX prescription group and the perindopril group at 1 week.The expression of LKB1mRNA and AMPKmRNA was elevated (Pall <.05).The increased expression of PGC-1α,NRF1 and mtTFA proteins demonstrated statistically significant differences (Pall <.05).The expression of mtDNA protein followed an increasing trend.At 4 weeks,the expression of both pLKB1 and pAMPK proteins was elevated (Pall <.05).Heightened expression was also reported in LKB1mRNA and AMPKmR-NA (Pall <.05).The increased expression of PGC-1α,NRF1,mtTFA and mtDNA proteins demonstrated statistically significant differences (Pall <.05).ATP synthesis:ATP synthesis was increased in the YQHX prescription group and the perindopril group at both 1 week and 4 weeks (Pall <.001).Conclusion:The possible mechanism of YQHX prescription for preventing and treating post-MI myocardial remodeling may function through strengthening the activation AMPK signal pathway by LKB1,thus further increasing the expression of downstream transcription factor proteins and initiating mitochondrial replication and transcription;as a result,the YQHX prescription can improve the post-MI damage in mitochondrial morphological structure in the tissue at heart marginal zone as well as enhance mitochondrial biosynthesis and ATP synthesis.