Study On The Regulation Mechanism Of Fuzi-polysaccharide On The Mitochondrial Apoptosis Pathway In Blood Vessels Cryopreservation

Objective:By establishing a cryopreserved rat abdominal aorta model,to explore the regulation effect of Fuzi-polysaccharide(FPS)on genes expression of mitochondrial apoptosis pathway in vascular tissue cryopreservation.Methods:The abdominal aortas of 36 SD rats were randomly divided into 6 groups with 6 rats in each group,namely the control group,the cryoprcserved model group and the FPS 0.1 mg/ml group,FPS 1 mg/ml group,FPS 10 mg/ml group,FPS 20 mg/ml group.The cryopreservation control group and the FPS concentration groups were stored in liquid nitrogen after programmed cooling,and quickly rewarmed after 1 week.The expression of cytochrome c(Cyt-c)in vascular tissue was detected by immunohistochemistry;Western blotting(WB)was used to detect the expression of Cyt-c,the second Expression of mitochondrial-derived cystinase activator(Smac),cysteinyl aspartate specific proteinase-9(Caspase-9),Bcl-2-associated X protein gene(Bax)and B-lymphoma-2 gene(Bcl-2).The expression levels of Bax and Bcl-2 mRNA in vascular tissues were detected by reverse transcription real-time PCR(qRT-PCR).The above indexes were detected immediately after sampling in the control group.Results:1.The expression of Cyt-c in vascular tissues:Cyt-c is rarely expressed in fresh vascular tissues.The expression of Cyt-c in the vascular tissues of the cryopreserved model group was the highest,which has a large amount of brown in the cytoplasm of the cells.The expression of FPS 10 mg/ml and FPS 20 mg/ml group was significantly lower than that of the control group(P<0.05).2.Cyt-c,Smac,Caspase-9,Bax,Bcl-2 protein expression levels:Compared with the control group,the expressions of Cyt-c,Smac,Caspase-9 and Bax proteins in the cryopreserved model group and each concentration group of FPS were all up-regulated,and the expression of Bcl-2 protein was down-regulated;Compared with the cryopreserved model group,the relative protein expressions of Cyt-c,Smac,Caspase-9 and Bax in the FPS 1,10 and 20 mg/ml groups were decreased,The expression level of FPS 10 mg/ml group was significantly lower than that of other concentration groups(P<0.05).Compared with the cryopreserved model group,the relative expression of Bcl-2 in each FPS concentration group increased significantly(P<0.05),but the difference between the FPS 10 mg/ml and FPS 20 mg/ml groups was not significant(P>0.05).3.The expression levels of Bax and Bcl-2 mRNA in vascular tissues:Compared with the control group,the expression levels of Bax mRNA in the cryopreserved model group and each concentration group of FPS increased(P<0.05),while the expression levels of Bcl-2 mRNA decreased(P<0.05);Compared with the cryopreserved model group,Bax mRNA expression in all FPS groups was decreased(P<0.05);Except for the FPS 0.1 mg/ml group,the expression of Bcl-2 mRNA was increased in other FPS concentration groups(P<0.05);Compared with the FPS group with different concentrations,the FPS 10 mg/ml group could significantly up-regulate the expression level of Bcl-2 mRNA,while down-regulating the expression level of Bax mRNA(P<0.05).Conclusions:1.Hypothermic damage can cause apoptosis in vascular tissue.2.Fuzi-polysaccharide can protect vascular tissues cryopreservation through the regulation of factors involved in the mitochondrial apoptosis pathway..