超负荷运动诱导小鼠阴虚证模型的构建与评价

Construction and evaluation of a mouse model of yin-deficiency syndrome induced by overloading exercise

关键词:  阴虚证  动物模型  超负荷运动

Key words:  yin-deficiency syndrome  animal model  overload exercise

摘要:

摘要：目的 构建与评价超负荷强迫游泳诱导的阴虚证小鼠模型。方法 通过持续超负荷强迫游泳的方法构建小鼠阴虚证模型，模型小鼠每日游泳时长从第一周的20min逐步增加至第六周的120min，末次游泳次日取其新鲜粪便进行菌群检测，测量肛温后通过负重游泳观察小鼠体力变化，结束后断尾取少量血液进行血糖含量检测；次日，眼内眦取血后检测血常规，剩余血液离心后得到血浆，检测外周血中cAMP、cGMP、ALT、AST、LDH以及BUN水平，剩余血浆进行代谢组学检测；脱颈椎处死小鼠，取胸腺、脾、肝脏、肾脏及部分大肠称重并进行病理学检测。结果 持续超负荷游泳对小鼠有较大的影响，模型小鼠体重显著下降，脏器指数中胸腺指数显著降低（P＜0.01），肝指数与肾指数提高（P＜0.01），脾指数也有所增加（P＜0.05）；造模结束后模型小鼠血糖值较对照组降低明显（P＜0.01），而肛温较对照小鼠略有升高（P＜0.05）；外周血血细胞检测结果中模型小鼠HCT值较对照有显著提高（P＜0.01），LYMPH%上调（P＜0.05），NEUT%下调（P＜0.05）；模型小鼠血浆中ALT、AST活力、BUN与cGMP值较对照组无显著差异（P＞0.05），LDH活性与cAMP水平较对照组显著上升（P＜0.01），从而cAMP/cGMP比值较对照组有显著提高（P＜0.05）；高通量血浆代谢物检测结果说明模型小鼠血浆代谢物与对照有较大变化，其中有807种代谢物下调，202种上升，通过富集分析发现模型组类固醇激素生物合成以及亚油酸代谢相关代谢物的下降明显；模型小鼠粪便菌群特征较对照组也发生了较大的改变，其中模型组菌群丰度下降，Alpha多样性中Shannon、Simpson、pielou-e值均显著下降；Beta多样性分析结果也存在较大的变化，如属水平中乳杆菌、大肠埃希菌等有益菌群丰度下降。结论 超负荷强迫游泳可建立可靠的小鼠阴虚证动物模型。

Abstract:

ABSTRACT: OBJECTIVE Construction and evaluation of a mouse model of yin-deficiency syndrome induced by overloading forced swimming. METHODS The mouse model of yin-deficiency syndrome was constructed by continuous overload forced swimming. The daily swimming time of model mice gradually increased from 20min in the first week to 120min in the sixth week. On the day after the last swim, fresh feces were collected for flora detection. After the anal temperature was measured, the physical changes of mice were detected by weight-bearing swimming, and a small amount of blood was taken from the tail to detect the blood glucose content. The next day, blood was collected from the inner canthus for routine blood testing, and the remaining blood was centrifuged to obtain its plasma. Detect the levels of cAMP, cGMP, ALT, AST, LDH, and BUN in plasma, and perform metabolomics analysis on the remaining plasma. The mice were euthanized after cervical spondylosis, and the thymus, spleen, liver, kidney, and large intestine were weighed and subjected to pathological examination. RESULTS Continuous overload swimming has a significant impact on mice, the model mice showed significant weight loss. In the organ index, the thymus index significantly decreased (P<0.01), the liver index and kidney index significantly increased (P<0.01), and the spleen index slightly increased (P<0.05); After modeling, the blood glucose levels of the model mice decreased significantly compared to the control group (P<0.01), and the anal temperature slightly increased (P<0.05); The results of peripheral blood cell testing showed that the HCT value of the model mice was significantly increased compared to the control group (P<0.01), LYMPH% was upregulated (P<0.05), and NEUT% was downregulated (P<0.05); The activities of ALT, AST, BUN, and cGMP in the plasma of model mice showed no significant differences compared to the control group (P>0.05), while the activities of LDH and cAMP levels significantly increased compared to the control group (P<0.01), resulting in a significant increase in the cAMP/cGMP ratio of the model group (P<0.05). The high-throughput plasma metabolite detection results indicate that there are significant changes in the plasma metabolites of the model mice compared to the control group, with 807 metabolites decreasing and 202 metabolites increasing. Through enrichment analysis, it was found that the model group showed a significant decrease in steroid hormone biosynthesis and linoleic acid metabolism related metabolites. Modeling has a significant impact on the characteristics of mouse fecal microbiota, with a decrease in the abundance of the model group's microbiota and a significant decrease in Shannon, Simpson, and pielou-e values in Alpha diversity; The results of Beta diversity analysis also showed significant changes, such as a decrease in the abundance of beneficial bacterial groups such as Lactobacillus and Escherichia coli at the genus level. CONCLUSION Overloading forced swimming can establish a reliable mouse model of yin-deficiency syndrome.