| OBJECTIVE:In this study,a rat model of myocardial ischemia reperfusion injury was established to observe the protective effect of carminative and expectorant traditional Chinese medicine on myocardial ischemia reperfusion injury in rats from the perspective of endothelial function,inflammatory response and cell apoptosis,and to explore the mechanism of its action.This study provides the theoretical basis for the clinical application of carminative and expectorant traditional Chinese medicine.MATERIALS AND METHODS:140 SD rats were fed adaptively for 7 days,and randomly divided into the sham operation group,the model group,the perindopril group,and the carminative and expectorant traditional Chinese medicine group for 35 rats per group.According to the commonly used dose recommended by adults for daily use,the weight conversion coefficient of adults and rats was 6.3,the dosage for rats was calculated,and then the drug was given once a day,and the gavage was continued for 7 days.Rats in the sham operation group and the model group were given normal saline of 10 mL·kg-1 by gavage.Rats in the carminative and expectorant traditional Chinese medicine group were given carminative and expectorant traditional Chinese medicine of 8.1g·kg-1 by gavage.Rats in the perindopril group were given perindopril tablets of 0.4mg·kg-1 by gavage.The myocardial ischemia reperfusion model was established in one hour after the end of the gavage on the 7th day.The procedure of the sham operation group was the same as that of the other three groups,but no ligation was performed.After successfully modeling,10 rats in each group were randomly selected,which are fasting on the eve of selection.4 mL of blood was taken from the abdominal aorta of the rats.After standing for 30 minutes,the cells were centrifuged,and the supernatant was taken and stored at-20?for use.Rats were sacrificed immediately after blood collection,and the hearts of the rats were taken out under aseptic conditions.After washing with pre-cooled sterile saline,the hearts were fixed with 10%formaldehyde,placed in liquid nitrogen,and stored at-20?for use.The contents of VEGF,TXA2,PGI2,sTM,sEPCR,IL-8,IL-6 and TNF-a in serum were detected by ELISA.The protein expression levels of P-Akt,Bax,Bcl-2 were detected by Western Blot.The gene expression levels of Akt mRNA,Bax mRNA and Bcl-2 mRNA were detected by RT-PCR.RESULTS:1.ELISA detection:1.1 VEGF content in each group of rats'serum:Compared with the sham operation group,the VEGF content in the model group was significantly decreased,P<0.05;compared with the model group,the VEGF content in the perindopril group was increased,P<0.05;compared with the model group,the VEGF content in the carminative and expectorant traditional Chinese medicine group was increased,P<0.05;compared with the perindopril group,the VEGF content in the carminative and expectorant traditional Chinese medicine group was significantly increased,P<0.05.1.2 TXA2 and PGI2 contents in each group of rats'serum,and the TXA2/PGI2 specific value:Compared with the sham operation group,the TXA2 content in the model group was significantly increased,the PGI2 content was significantly decreased,the TXA2/PGI2 was significantly increased,P<0.05;compared with the model group,the TXA2 content in the perindopril group was decreased,the PGI2 content was increased,the TXA2/PGI2 was decreased,P<0.05;compared with the model group,the TXA2 content in the carminative and expectorant traditional Chinese medicine group was decreased,the PGI2 content was increased,the TXA2/PGI2 was decreased,P<0.05;compared with the perindopril group,the TXA2 content in the carminative and expectorant traditional Chinese medicine group was significantly decreased,the PGI2 content was not significantly increased,the TXA2/PGI2was significantly decreased,P<0.05.1.3 sTM and sEPCR contents in each group of rats'serum:Compared with the sham operation group,the sTM and sEPCR contents in the model group were increased,P<0.05;compared with the model group,the sTM and sEPCR contents in the perindopril group and the carminative and expectorant traditional Chinese medicine group were decreased,P<0.05;compared with the perindopril group,the sTM and sEPCR contents in the carminative and expectorant traditional Chinese medicine group were increased,P>0.05.1.4 IL-6 and IL-8 contents in each group of rats'serum:Compared with the sham operation group,the IL-6 and IL-8 contents in the model group were increased,P<0.05;compared with the model group,the IL-6 and IL-8 contents in the perindopril group and the carminative and expectorant traditional Chinese medicine group were decreased,P<0.05;compared with the perindopril group,the IL-8 content in the carminative and expectorant traditional Chinese medicine group was decreased,the IL-6 content was increased,P<0.05.1.5 TNF-a content in each group of rats:Compared with the sham operation group,the TNF-a content in the model group was increased,P<0.05;compared with the model group,the TNF-a content in the perindopril group and the carminative and expectorant traditional Chinese medicine group was decreased,P<0.05;compared with the perindopril group,the TNF-a content in the carminative and expectorant traditional Chinese medicine group was increased,P>0.05.2.PCR detection2.1 The expression level of Akt mRNA in each group of rats:Compared with the sham operation group,the expression level of Akt mRNA in the model group was significantly decreased,P<0.05;compared with the model group,the expression level of Akt mRNA in the perindopril group was increased,P<0.05;compared with the model group,the expression level of Akt mRNA in the carminative and expectorant traditional Chinese medicine group was increased,P<0.05;compared with the perindopril group,the expression level of Akt mRNA in the carminative and expectorant traditional Chinese medicine group was not significantly increased,P>0.05.2.2 The expression level of Bax mRNA and Bcl-2 mRNA in each group of rats:Compared with the sham operation group,the expression level of Bax mRNA in the model group was significantly increased,the expression level of Bcl-2 mRNA was significantly increased,P<0.05;compared with the model group,the expression level of Bax mRNA in the perindopril group was decreased,the expression level of Bcl-2 mRNA was increased,P<0.05;compared with the model group,the expression level of Bax mRNA in the carminative and expectorant traditional Chinese medicine group was decreased,the expression level of Bc-l2mRNA was increased,P<0.05;compared with the perindopril group,the expression level of Bax mRNA in the carminative and expectorant traditional Chinese medicine group was not significantly decreased,the expression level of Bcl-2 mRNA was not significantly increased,P>0.05.3.Western Blot detection?Protein grayscale statistics?3.1 The protein expression level of P-Akt in each group of rats:Compared with the sham operation group,the protein expression of P-Akt in the model group was decreased,P<0.05;compared with the model group,the protein expression of P-Akt in the perindopril group was increased,P<0.05;compared with the model group,the protein expression of P-Akt in the carminative and expectorant traditional Chinese medicine group was increased,P<0.05;compared with the perindopril group,the protein expression of P-Akt in the carminative and expectorant traditional Chinese medicine group was not significantly decreased,P>0.05.3.2 The protein expression level of Bax,Bcl-2 in each group of rats:Compared with the sham operation group,the protein expression of Bax and Bcl-2 in the model group was increased,and the protein expression of Bcl-2/Bax was decreased,P<0.05;compared with the model group,the protein expression of Bcl-2 in the perindopril group and carminative and expectorant traditional Chinese medicine was increased,the protein expression of Bax was decreased,the protein expression of Bcl-2/Bax was increased,P<0.05;compared with the perindopril group,the protein expression of Bcl-2 and Bax in the carminative and expectorant traditional Chinese medicine group was increased,the protein expression of Bcl-2/Bax was decreased,P<0.05.CONCLUSION:1.The carminative and expectorant traditional Chinese medicine can increase the VEGF content in the serum,increase PGI2 content,decrease TXA2 content,keep balance of TXA2/PGI2 proportion,decrease the levels of sTM and sEPCR in the serum,and then make the protective effect on myocardial ischemia reperfusion vascular endothelial function in rats.2.The carminative and expectorant traditional Chinese medicine can decrease the contents of IL-8,IL-6 and TNF-?,it proves that the protective effect of the carminative and expectorant traditional Chinese medicine on myocardial ischemia reperfusion injury in rats may be related to the inhibition of inflammatory response.3.The carminative and expectorant traditional Chinese medicine can increase the protein expression level of P-Akt,decrease the protein expression level of Bax,increase the protein expression level of Bcl-2;the carminative and expectorant traditional Chinese medicine can increase the relative gene expression level of cell apoptosis of Akt mRNA and Bcl-2 mRNA,decrease the relative gene expression level of cell apoptosis of Bax mRNA;the protective effect mechanism of the carminative and expectorant traditional Chinese medicine on myocardial ischemia reperfusion injury in rats may be related to the activation of Bax/Akt pathway and the inhibition of cell apoptosis. |
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