| Objectives:1)Construct BDNF（Brain derived neurotrophic factor）cardiomyocyte conditional knockout mice and perform preliminary study on their phenotypes such as myocardial cell size,cardiac function,motor function,behaviors,blood BDNF,blood lipid,blood glucose,insulin,and leptin levels;2)Observe the possible differences on the infarction area and cardiac function between the 2-month-old male BDNF cardiomyocyte conditional knockout mice and the wild-type mice after myocardial infarction;3)Compare the sizes of cardiomyocyte of 2-month-old male BDNF cardiomyocyte conditional knockout mice and wild-type mice.Study the changes of cardiomyocyte size and the numbers and sizes of nuclei in wild-type C57 mice with the increase of age.Methods:1)Construct BDNF myocardial conditional knockout(BDNF-/-)mice by mating of STOCK Bdnf（tm3Jae）/J and B6.FVB-Tg（Myh6-cre）2182Mds/J mice;2)Using Vevo ultra high-resolution small animal ultrasound imaging system to analyze the cardiac function of the experimental mice;3)Detect the BDNF,leptin,insulin,high density lipoprotein,low density protein,triglyceride,total cholesterol,free cholesterol levels in the blood of BDNF cardiomyocyte conditional knockout(BDNF-/-)mice by ELISA;4)Detect the blood glucose level in BDNF cardiomyocyte conditional knockout(BDNF-/-)mice by paper strip method;5)Using the forelimb grasping force measuring instrument,the body upward activity tester,the endurance tester and the mouse experimental treadmill to measure mice forelimb tension,body upward activity,endurance and exercise ability respectively;6)Using the morris water maze,the mouse jumping instrument,the mouse automatic avoidance instrument,the mouse spontaneous activity recorder and the mouse open field test cage to measure mice spatial orientation ability,memory ability,behavioral stress of autonomic exploration,autonomic activity and anxiety state respectively;7)Myocardial infarction was induced by ligation of the left anterior descending coronary artery,and the infarct size and cardiac function were compared with those of the wild-type mice with BDNF cardiomyocyte conditional knockout mice;8)The single myocardial cell was isolated by Langendorff enzyme digestion method,using Hochest33342 stain myocardial cell and the sizes of myocardial cells and nuclei were measured by Image-Pro-Plus 6 software;9)Perform statistic analysis on each index between the experimental group and the control group using SPSS 19.0 and Origin 9.4,GraphPad Prism7 statistical analysis software.Results:1)There was no significant difference between the weights of the 2-month-old male BDNF cardiomyocyte conditional knockout mice and the male wild-type littermate（p>0.05）.However,the 8-month-old male BDNF cardiomyocyte conditional knockout mice were heavier than the male wild-type littermate,and difference between their weights was statistic significant（p<0.05）;2)The ejection fraction and shortening fraction of 2 and 8-month-old male BDNF cardiomyocyte conditional knockout mice were lower than the male wild-type littermate and the difference is statistic significant（p<0.05）,while there was no significant difference between the two groups on the systolic diameter,diastolic diameter,systolic volume and diastolic volume（p>0.05）;3)Among the tested exercise ability index,the forelimb tension,endurance and exercise ability of the 2-month-old male BDNF cardiomyocyte conditional knockout mice were lower than the male wild-type littermate except the body upward activity,and the differences were statistic significant（p<0.05）;4)The blood BDNF level in the 8-month-old male BDNF myocardial conditional knockout mice was higher than the male wild-type littermate and the difference is statistic significant（p<0.05）,while there was no significant difference between the two groups on leptin,insulin,high density lipoprotein,low density protein,triglyceride,total cholesterol,free cholesterol levels in the blood（p>0.05）;5)Using the morris water maze,the mouse jump instrument,the mouse automatic avoidance instrument,the mouse autonomic activity recorder,the mouse open field test cage to measure mice spatial orientation ability of learning and memory,learning and memory ability,autonomic behavior exploration behavior and tension,autonomic activity and anxiety state respectively,there was no significant difference for all the detected paramenters between the 8-month-old male BDNF cardiomyocyte conditional knockout mice and the male wild-type littermate（p>0.05）;6)The infarction area of the 2-month-old male BDNF cardiomyocyte conditional knockout mice was bigger than the male wild-type littermate after myocardial infarction,however the difference was not statistic significant（p>0.05）.The ejection fraction of the BDNF myocardial conditional knockout mice was lower than the wild-type littermate after myocardial infarction and the difference was no statistic significant（p>0.05）;7)The length,width,size,volume and nuclear area of cardiomyocytes of C57 mice increased significantly with the increase of age,while the area per unit nucleus decreased significantly with the increase of age（p<0.05）.The percentage of multinucleated cells decreased with age,while the percentage of monocytes increased significantly（p<0.05）.Conclusions:1)There is no significant difference between the wights of the 2-month-old male BDNF cardiomyocyte conditional knockout mice and the male wild-type littermate;while the weight of the 8-month-old male BDNF cardiomyocyte conditional knockout mice was significantly higher than the male wild-type littermate;2)The ejection fraction and shortening fraction of 2 and 8-month-old male BDNF cardiomyocyte conditional knockout mice are significantly lower than the male wild-type littermate;3)Among the tested exercise ability index,the forelimb tension,endurance and exercise ability of the 2-month-old male BDNF cardiomyocyte conditional knockout mice are significantly lower than the male wild-type littermate except the body upward activity;4)The blood BDNF level in the 8-month-old male BDNF cardiomyocyte conditional knockout mice is significantly higher than the male wild-type littermate,while there is no significant difference between the two groups for leptin,insulin,high density lipoprotein,low density protein,triglyceride,total cholesterol,free cholesterol levels in the blood;5)There is no significant difference for indexs（the morris water maze,the mouse jump,the mouse automatic avoidance,the mouse autonomic activity and the mouse open field test）between the 8-month-old male BDNF cardiomyocyte conditional knockout mice and the male wild-type littermate;6)When the 2-month-old male BDNF cardiomyocyte conditional knock out mice were induced myocardial infarction,the infarct area wasn’t significantly greater than that of the male control littermate,also the cardiac function indicator ejection fraction is not significantly lower than that of the same-sex control littermate;7)The length,width,size,volume and nuclear area of myocardial cells of C57 mice increased significantly with the increase of age,while the area per unit nucleus decreased significantly with the increase of age.With increase of age,the percentage of multinucleated cells decreased,while the percentage of monocytes increased significantly. |
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