Study on the role of aerobic exercise in regulating the CNPY2-mediated AKT/GSK3β pathway for improving non-alcoholic fatty liver

doi:10.12307/2025.780

Abstract

Abstract: BACKGROUND: Non-alcoholic fatty liver disease is one of the common chronic liver diseases in the world. Aerobic exercise is considered to be an important means for the treatment of non-alcoholic fatty liver disease. However, the mechanism of exercise to improve non-alcoholic fatty liver disease has not been fully clarified.
OBJECTIVE: To investigate the effects of aerobic exercise on the protein kinase B/glycogen synthase kinase-3β pathway mediated by Canopy FGF signaling regulator 2 (CNPY2) in the liver canopy of non-alcoholic fatty liver disease mice and its mechanism.
METHODS: Thirty male CNPY2 knockout mice (ko) and thirty their litters of wild-type mice (wt) were fed adaptively for one week and randomly divided into control group, model group, and model exercise group, with 10 mice in each group. The control group was fed with ordinary diet. The model group and the model exercise group were fed with high-fat diet for 17 weeks. The model exercise group received continuous aerobic exercise intervention from week 10 until the end of the experiment at week 18. Liver histopathology was observed by hematoxylin-eosin and oil red O staining. The levels of serum lipids and liver function were detected by automatic biochemical analyzer. The expression levels of CNPY2, protein kinase B/glycogen synthase kinase-3β pathway, and Caspase-3 protein in liver tissues were detected by Western Blotting. The apoptosis rate of hepatocytes was detected by TUNEL staining.
RESULTS AND CONCLUSION: (1) Compared with wt control group, CNPY2 expression in liver tissues of wt model group was increased (P < 0.05), while CNPY2 expression in wt model exercise group was decreased compared with wt model group (P < 0.05). Compared with control group, wt mice and ko mice in model group showed steatosis, increased lipid droplets, abnormal blood lipids and liver function, decreased protein kinase B/glycogen synthase kinase-3β expression
(P < 0.05) and increased Caspase-3 expression (P < 0.05), and increased hepatocyte apoptosis rate in liver tissue (P < 0.05). (2) Compared with the model group, wt mice and ko mice showed improvement in the above indexes in model exercise group. (3) Compared with wt mice, the above indexes of ko mice were improved. (4) These findings indicate that CNPY2 gene deletion and aerobic exercise can effectively improve non-alcoholic fatty liver disease. The mechanism may be related to aerobic exercise reducing CNPY2 expression, activating protein kinase B/glycogen synthase kinase-3β signaling pathway, and thus inhibiting hepatocyte apoptosis.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: fatty liver, high-fat diet, hepatocyte apoptosis, aerobic exercise, CNPY2, AKT, GSK3β, Caspase-3

CLC Number:

Wang Jiaqian, , Jiang Changjun, Peng Yi, Ma Mi, Li Junhan. Study on the role of aerobic exercise in regulating the CNPY2-mediated AKT/GSK3β pathway for improving non-alcoholic fatty liver [J]. Chinese Journal of Tissue Engineering Research, 2025, 29(30): 6441-6448.

Figures/Tables 7

2.2.2 肝指数结果显示：第18周wt模型组较wt对照组小鼠肝指数显著升高(P < 0.05)； wt模型运动组较wt模型组小鼠肝指数显著下降(P < 0.05)；ko小鼠组间肝指数无显著差异。wt小鼠与ko小鼠对应组相比，ko模型组小鼠肝指数降低，但差异无显著性意义(P > 0.05)，见表2。 2.3 各组小鼠血清生化指标结果 2.3.1 血脂代谢指标在wt小鼠和ko小鼠各自组内，模型组较对照组小鼠血清总胆固醇、三酰甘油、低密度脂蛋白均显著升高(P < 0.05)，高密度脂蛋白显著降低(P < 0.05)；模型运动组较模型组小鼠血清总胆固醇、三酰甘油、低密度脂蛋白均显著降低(P < 0.05)，wt模型运动组小鼠高密度脂蛋白表达显著升高(P < 0.05)，ko模型运动组小鼠高密度脂蛋白表达升高，但差异无显著性意义(P > 0.05)。wt小鼠与ko小鼠对应组相比，ko模型组较wt模型组小鼠血清总胆固醇、三酰甘油、低密度脂蛋白均显著降低 (P < 0.05)，高密度脂蛋白显著升高(P < 0.05)；ko模型运动组较wt模型运动组小鼠血清总胆固醇、三酰甘油、低密度脂蛋白均显著降低(P < 0.05)，高密度脂蛋白差异无显著性意义(P > 0.05)，见图2。 2.3.2 肝功能指标在wt小鼠和ko小鼠各自组内，模型组较对照组小鼠血清丙氨酸氨基转氨酶、天门冬氨酸氨基转氨酶均显著升高(P < 0.05)；模型运动组较模型组小鼠血清丙氨酸氨基转氨酶、天门冬氨酸氨基转氨酶均显著降低(P < 0.05)。wt小鼠和ko小鼠对应组相比，ko模型组较wt模型组血清丙氨酸氨基转氨酶显著降低(P < 0.05)；其他对应组比较差异无显著意义(P > 0.05)，见图2。 2.4 各组小鼠肝组织病理形态结果 2.4.1 苏木精-伊红染色结果显示：在wt小鼠和ko小鼠各自组内，对照组肝细胞形态规整，结构清晰无病变；模型组较对照组肝细胞排列紊乱、伴肿胀空泡，炎性浸润及脂肪性变显著(P < 0.05)；模型运动组较模型组肿胀空泡、炎性浸润、脂肪性变情况显著改善(P < 0.05)。wt小鼠和ko小鼠对应组相比，ko组较wt组肝细胞结构空泡化、炎性细胞浸润、脂肪性变情况显著改善(P < 0.05)，见图3。 2.4.2 油红O染色结果显示：在wt小鼠和ko小鼠各自组内，对照组仅有少量脂肪颗粒，模型组较对照组出现严重脂质沉积，且细胞核被脂滴挤至边缘，脂滴密度显著升高(P < 0.05)，模型运动组较模型组脂滴密度显著下降(P < 0.05)。wt小鼠和ko小鼠对应组相比，ko组较wt组脂滴密度均显著下降(P < 0.05)，见图4。 2.5 各组小鼠肝组织蛋白表达结果由图可知，在wt小鼠组内，模型组较对照组CNPY2表达显著升高(P < 0.05)，模型运动组较模型组CNPY2表达显著降低(P < 0.05)；ko组小鼠CNPY2基因敲除几乎不表达。在wt小鼠和ko小鼠各自组内，模型组较对照组p-AKT/AKT、p-GSK3β/GSK3β表达均显著降低(P < 0.05)， Caspase-3表达均显著升高(P < 0.05)；模型运动组较模型组p-AKT/AKT、p-GSK3β/GSK3β表达均显著升高(P < 0.05)，Caspase-3表达均显著降低(P < 0.05)。wt小鼠与ko小鼠对应组相比，ko组较wt组p-AKT/AKT、p-GSK3β/GSK3β表达均显著升高(P < 0.05)， Caspase-3表达均显著降低(P < 0.05)，见图5。 2.6 各组小鼠肝组织TUNEL检测结果结果可见，蓝色为正常细胞核，绿色为凋亡细胞核(箭头标记处为例)。在wt小鼠和ko小鼠各自组内，模型组较对照组呈现较多凋亡细胞核，肝细胞凋亡率显著升高(P < 0.05)；模型运动组细胞凋亡数量有所减少，肝细胞凋亡率显著下降(P < 0.05)。wt小鼠与ko小鼠对应组相比，ko组较wt组凋亡数量减少，肝细胞凋亡率显著下降(P < 0.05)，见图6。"

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