Mining and verification of inflammation-related genes in skeletal muscle of exhaustive exercise rats undergoing cannabidiol intervention

doi:10.12307/2025.338

Abstract

Abstract: BACKGROUND: Cannabidiol is effective in ameliorating the body’s inflammatory response, but no clear mechanistic studies have been conducted to ameliorate skeletal muscle inflammation induced by exhaustive exercise.
OBJECTIVE: To explore the mechanism by which cannabidiol improves skeletal muscle inflammation during exhaustive exercise by using transcriptome sequencing technology.
METHODS: Thirty-six Sprague-Dawley rats were randomly divided into six groups: blank control group, exercise coconut oil group, exercise control group, 50 mg/kg cannabidiol group, 60 mg/kg cannabidiol group, and 70 mg/kg cannabidiol group, with six rats in each group. Except for rats in the blank control group, rats in each group were subjected to swimming exercise for 9 days to produce the exhaustive exercise model. At the end of each swimming exercise, rats in the cannabidiol groups were given 2 mL of fat-soluble cannabidiol at different concentrations (50, 60, and 70 mg/kg) by gavage; rats in the exercise coconut oil group were given the same volume of coconut oil by gavage until the end of the exercise on the 9th day; and rats in the blank control group and the exercise control group were not given any special treatment. The levels of inflammatory factors and differentially expressed genes in the skeletal muscle of rats in each group were determined using ELISA and transcriptome sequencing techniques. Differentially expressed genes obtained were subjected to KEGG analysis, and the accuracy of the sequencing data was verified by fluorescence quantitative PCR.
RESULTS AND CONCLUSION: The results of ELISA showed that the contents of interleukin-6 (P < 0.05), tumor necrosis factor-α (P < 0.01), interleukin-10 and other inflammatory factors in the exercise group increased significantly compared with the blank control group and the coconut oil group. After cannabidiol intervention, the mass concentrations of interleukin-6 and tumor necrosis factor-α showed a sequential decrease with increasing cannabidiol concentration. By comparing GO and KEGG databases, the functional properties of differentially expressed genes were analyzed, and the results showed that the differentially expressed genes were mainly involved in the tumor necrosis factor signaling pathway and the Toll-like receptor signaling pathway. RT-qPCR results showed that the trends of five randomly selected differentially expressed genes were in agreement with the transcriptome sequencing results. To conclude, cannabidiol can improve skeletal muscle inflammation caused by exhaustive exercise.

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: transcriptome, cannabidiol, exhaustive exercise, skeletal muscle inflammation, differentially expressed gene, inflammatory factor, KEGG analysis, inflammatory pathway

CLC Number:

Zhu Wenning, Sun Lili, Peng Lina, Si Juncheng, Zang Wanli, Yin Weidong, Li Mengqi . Mining and verification of inflammation-related genes in skeletal muscle of exhaustive exercise rats undergoing cannabidiol intervention[J]. Chinese Journal of Tissue Engineering Research, 2025, 29(11): 2347-2356.

Figures/Tables 9

2.5 肿瘤坏死因子信号通路的KEGG富集分析肿瘤坏死因子信号通路上的基因调控机体的炎症反应与免疫应答，使用KEGG的富集通路进行分析，发现该通路上的基因表达变化情况。 2.5.1 空白对照组和运动对照组比较如图6所示。在肿瘤坏死因子信号通路中，共测得8个上调基因，包括AP-1、Cxcl1、Cxcl2、Cxcl3、Il1b、IL15、Fos、Socs3；3个下调基因，包括Ccl5、Cxcl10、Ifi47。相比于空白对照组，运动对照组炎症因子上调数量更高，这在一定程度说明了剧烈运动后机体产生了免疫炎症反应。 2.5.2 运动对照组和大麻二酚组比较如图7所示，在肿瘤坏死因子信号通路中共测得4个上调基因，包括Cxcl10、Ifi47、P13K、IRF1；13个下调基因，包括IkBα、P38、MSK1/2、Ccl2、Cxcl1、Cxcl2、Cxcl3、Jag1、IL1b、Nfkbia、JunB、Sele、Ptgs2。相比于运动对照组，大麻二酚组炎症因子下调数量更多，这从侧面证明，大麻二酚能够改善力竭运动大鼠自身的炎症状态。 2.6 TLR信号通路的KEGG富集分析 TLR信号通路上的基因参与机体的炎症反应与免疫应答，使用KEGG的富集通路进行分析，发现该通路上的基因表达变化情况。 2.6.1 空白对照组与运动对照组比较如图8所示，在TLR信号通路中共测得4个上调基因，包括TLR5、白细胞介素1β、AP-1、TLR7/8，4个下调基因，包括IRF7、RANTES、IP-10、MIG。 2.6.2 运动对照组和大麻二酚组比较如图9所示，在TLR信号通路中测得2个上调基因，包括PI3K、MIG，4个下调基因，包括TLR2、P38、IkBα、白细胞介素1β。相比于运动对照组，大麻二酚组炎症因子下调数量更多，这从侧面证明大麻二酚能够改善力竭运动大鼠自身的炎症状态。 2.7 RT-qPCR验证结果如图10所示，5个基因的变化趋势与转录组测序结果相一致，进一步证实了转录组数据具有较高的可信度。"

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