Effect Of JM No.1 WYCF On The Intracellular Calcium Concentration In Vascular Cells Of ASO Rabbits

Objective: This study is a part of the National Natural Science Foundation of China.The purpose of this project is to explore the prevention and treatment of JM No.1 WYCF,JM No.1 and JKSQ on rabbit ASO model through animal experiments.Further studies were conducted to investigate the effects of Wenyang Recipe on serum calcium ion,intracellular calcium concentration and calmodulin in rabbits with ASO model.Preliminary verification of Professor Deng Baiyang's hypothesis related to the calcium overload of senile kidney yang blood cells based on the principle of heterosexual disease and the existing literature research and its own previous research.Methods: Thirty-five New Zealand white rabbits were randomly divided into normal control group,model group,JM No.1 WYCF Group,JM No.1 group,JKSQ group,with 7 groups in each group.The rabbits were replicated for 12 weeks using pure high-fat-fed experimental rabbits and administered simultaneously during model replication.After 12 weeks,the preparation of the model was judged by color Doppler abdominal aortic angiography and abdominal aortic HE staining pathological section.Blood samples were taken at the beginning and end of the experiment to detect blood lipids,blood rheology and serum calcium ions.After the model was successfully prepared,the 2-3 cm abdominal aorta was cut under anesthesia and divided into two parts.One section was used for paraffin section HE staining to observe the morphological changes of vascular tissue.A liquid nitrogen rapid freezing qPCR was used to detect the relative expression of cam(calmodulin)mRNA in vascular tissues of each group.Two rabbits were randomly selected from each group and washed rapidly with pre-cooled PBS solution.The vascular tissue was cut into meat emulsions,and the cell suspension was added to the flo-4-am fluorescent probe for flow cytometry to detect fluorescence intensity and cells.The proportion of positive calcium ions.Results: After 12 weeks,the results of vascular ultrasound showed that there was no clear plaque echo in the abdominal aorta of the normal control group through the normal wall.In the model group,the intima of the abdominal aorta of the rabbits were partially smooth,and multiple strong echo plates were observed.Pathological section examination of abdominal aorta showed that the model group had obvious plaque formation and lumen narrowing;the blood index test results of each group: 1.Results of each group before and after the experiment 1 normal control group,plasma total cholesterol(TC),triglyceride(TG),low-density lipoprotein cholesterol(LDL-C)and high-density lipoprotein cholesterol(HDL-C)concentrations did not differ(P>0.05);blood rheology index whole blood low-cut viscosity,high-cut viscosity,plasma viscosity And hematocrit,the difference was not statistically significant(P>0.05);model group,JM No.1 WYCF group,JM No.1 group and JKSQ group were TC,LDL-C concentration was significantly increased(P<0.05),there was no significant difference in TG and HDL-C concentrations(P>0.05);blood rheology index whole blood low-cut viscosity,high-cut viscosity,plasma viscosity and hematocrit were all different degrees.The difference was statistically significant(P<0.05);There was no difference in serum calcium concentration before and after the experiment(P>0.05);2.After the model was successfully prepared,the blood indexes of each group were compared with the model group and each drug.Compared with the normal control group,the treatment group was TC and LDL-C.There was no significant difference in the concentration of TG and HDL-C(P>0.05);The blood flow index showed low blood viscosity,high viscosity,plasma viscosity and hematocrit.The difference was statistically significant(P<0.05);there was no difference in serum calcium ion concentration(P>0.05);There was no difference in concentration of TC,TG,LDL-C and HDL-C between the drug-treated group and the model group.P>0.05);blood rheology index whole blood low-cut viscosity,high-cut viscosity,plasma viscosity and hematocrit,the difference was not statistically significant(P>0.05);there was no difference in serum calcium ion concentration(P>0.05);3.HE staining results of pathological sections of abdominal aorta in each group showed that the surface of the vascular tissue of the normal control group was smooth and intact,the endothelial cells were arranged neatly without bulging,and the smooth muscle cells of the middle membrane were arranged in a ring,neat;model group and each drug treatment There were different degrees of endothelial plaque formation in the group.The vascular tissue morphology of the model group was the worst,and the JM No.1 group was basically normal.The vascular tissue morphology of the JM No.1 WYCF group and the JKSQ group was poor but better than the model group;4.qPCR Detection of cams in various groups of vascular tissue The relative expression of mRNA of calmodulin gene showed no significant difference between the groups(P>0.05);5.The results of cell flow assay showed that the difference of intracellular calcium concentration in the model group compared with the normal control group was different.Statistical significance(P<0.05);Compared with the drug treatment group,the calcium concentration in the model group was lower than that in each drug treatment group(P<0.05).Conclusion:1.The ASO animal model was successfully prepared by feeding the rabbits with high fat for 12 weeks.2.After the experiment,except for the normal control group,the model group and the drug treatment group all suffered from hyperlipidemia and the blood was in a highly viscous state.The drug has no obvious effect on the blood lipid and blood viscosity of rabbits in the prevention and treatment of ASO disease,which may be due to the high-fat feeding during the feeding process.3,through the morphological changes of vascular tissue,Wenyang split can be split to reduce the risk of plaque formation and protect the integrity of the vascular structure to prevent lesions.The effect may be that the Wenyang method can promote the influx of calcium ions and increase the concentration of calcium ions in vascular tissue cells.The intracellular calcium concentration in the vascular tissue of the model group is low,indicating that the pathogenesis of ASO may be the "no-load" rather than "overload" of calcium ions in vascular tissue cells.4,the use of classic Wenshen Yangyang Recipe JKSQ to prevent certain symptoms,but the effect is not as good as the JM No.1 WYCF,the high-fat induced ASO animal model may be yang deficiency syndrome.