Investigating The Protective Mechanism On Cryopreservation Of Blood Vessels With Fuzi-polysaccharies Based On Mitochondrial Energy Metabolism

Objective: by establish a rat abdominal aortic freezing injury model in cryopreservation,to observe the effects of different concentrations of Fuzi-polysaccharides(FPS)on mitochondrial energy metabolism in cryopreserved vascular tissues,and to explore the protective mechanism of FPS on blood vessels.Methods: In this paper,48 SD male rats were divided into a control group,cryopreserved model group(sterile water for injection),and aconite polysaccharide intervention group(FPS 0.1 mg/ml group,1 mg/ml group,10 mg /ml group,20 mg/ml group).The abdominal aorta was obtained under a microscope.The control group entered the testing process immediately after obtaining the material.In contrast,the lyophilized model and epimedium polysaccharide intervention groups obtained the material after programmed cooling-liquid nitrogen preservation(7 days)-rapid rewarming for index testing.The internal ultrastructure of the mitochondria was observed by transmission electron microscopy;the content of ATP,ADP,and AMP in vascular tissues was measured by high-performance liquid chromatography(HPLC),and the energy charge(EC)was calculated.Na+-K+-ATPase,Ca2+-ATPase),superoxide dismutase(SOD),glutathione peroxidase(GSH-PX),catalase(CAT)activity,and malondialdehyde(MDA)content;UV-Vis spectrophotometer to detect mitochondrial respiratory chain complex I,II,III,and IV activities.Results: 1.The ultrastructural changes in mitochondria were observed by transmission electron microscope: the mitochondria in the control group were regular oval with clear outlines.The outer membrane is smooth and intact.The inner crest was dense and clearly visible.The matrix was evenly distributed;compared with the control group,the mitochondrial structure of the vascular tissue in the cryopreserved model group was seriously damaged.The inner and outer membranes of mitochondria are ruptured and dissolved.Flowed outward,the mitochondrial matrix is transparent.After pretreatment with different concentrations of FPS,the mitochondrial ultrastructure damage was reduced,and the mitochondrial damage was reduced with the increase of FPS concentration,which was concentration-dependent.The group with high FPS concentration was more evident than those with low FPS concentration.2.Vascular tissue mitochondrial protein concentration:Compared with the control group,mitochondrial proteins were heavily degraded in the the cryopreservation model group(P< 0.01).Compared with the cryopreservation model group,the degradation of mitochondrial protein was significantly reduced in the fuzi-Polysaccharides intervention group,the higher the FPS concentration,the higher the protein concentration,the solution reached saturation when the concentration reached FPS 10 mg/ml,at which time the protein concentration was highest(P< 0.01).Compared with the FPS 10 mg/ml group,the FPS 20 mg/ml granulosa protein concentration showed a decreasing trend,and the difference was not statistically significant(P > 0.1).3.ATP、ADP、AMP content and energy charge in vascular tissue: Compared with the control group,the ATP,ADP,AMP content and EC in vascular tissues of the cryopreservation model group and the fuzi-Polysaccharides intervention group(FPS 0.1 mg/ml,1 mg/ml)showed a significant decrease(P<0.01).The differences of ATP,ADP,AMP and EC in vascular tissues of FPS 0.1 mg/ml and FPS 1 mg/ml groups were not statistically significant compared with the cryopreservation model group(P>0.05).The difference was statistically significant(P<0.01)from 10 mg/ml onwards compared with the cryopreservation model group,but the difference between the FPS 10 mg/ml and FPS 20 mg/ml groups was not significant.(P>0.05).4.The activities of mitochondrial respiratory chain complex,ATPase,antioxidant enzyme(SOD,GSH-Px,CAT),and MDA content in vascular tissue: Compared with the control group,,the activities of mitochondrial respiratory chain complex,ATPase and antioxidant enzyme in the vascular tissues of the cryopreservation model group and the fuziPolysaccharides intervention group were significantly decreased(P < 0.05),and MDA content significantly increased(P < 0.05).Except for the FPS 0.1 mg/ml group,the activities of complex,ATPpase and antioxidant enzyme in the fuzi-Polysaccharides intervention group were significantly higher than those in the cryopreservation model group(P < 0.01),and MDA content decreased significantly(P < 0.01).Compared with FPS 1 mg/ m L group,the complex(I-IV)activity,ATPpase activity and antioxidant activity of FPS 10 mg/ m L group and FPS 20 mg/ m L group were significantly increased(P < 0.01)(P < 0.05),and MDA content decreased significantly(P < 0.01)(P < 0.05).When the concentration was increased to 10 mg/ m L,the complex activity,ATPpase activity and antioxidant activity reached the peak,when the concentration was further increased the Enzyme activity showed a downward trend,but the difference between the FPS 10 mg/ml and FPS 20 mg/ml groups was not significant(P > 0.05).Conclusions:1.Mitochondrial structure and function are impaired during cryogenic storage,and energy metabolism of vascular tissue is affected;2.FPS has a protective effect on blood vessels by protecting mitochondrial structure,improving mitochondrial dysfunction and regulating energy metabolism in cryopreservation.