Ye Shanyu, Si Wenwen, Qin Wei, Yang Lin, Luo Ziwei, Li Zhen, Xie Yulu, Pan Hao, Li Xinrong, Huang Zifeng, Zhu Meiling, Chen Dongfeng
Department of Anatomy, School of Basic Medical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China.
Traditional Chinese Medicine Innovation Research Center, Shenzhen Hospital of Integrated Traditional Chinese and Western Medicine, Guangzhou University of Chinese Medicine, Shenzhen, China.
Anat Rec (Hoboken). 2023 Dec;306(12):3006-3020. doi: 10.1002/ar.24930. Epub 2022 Apr 21.
Atractylodeslancea Rhizoma (Rhizoma atractylodis [RA]) has long been recommended for the treatment of arthritis in traditional Chinese medicine, but its mechanism of action is still unclear. RA contains a large amount of Atractylodes lancea volatile oils (Atr). In this study, we investigated whether Atr can promote mesenchymal stem cells (MSCs) chondrogenic differentiation. The Atr were extracted from RA by steam distillation method, and the effect of Atr on MSCs was detected by the CCK8 assay. The optimal concentration of Atr for MSCs cultivation was 3 μg/ml. The differentially expressed miR-181a-5p was screened by miRNA microarray assay, and its mimics and inhibitors were transfected into MSCs. It was found that the inhibitor of miR-181a-5p could upregulate cartilage-specific genes such as SOX9, COL2A1, and ACAN. Meanwhile, we also found that the expression of gene editing enzyme ADAR2 was significantly increased in the chondrogenic differentiation of MSCs induced by Atr, and the bases of precursor sequence of miR-181a-5p were changed from A to G. After ADAR2 deletion, the expression of cartilage-specific genes was significantly down-regulated and the precursor sequence bases of miR-181a-5p were not changed. Bioinformatics analysis revealed that the predicted target gene of miR-181a-5p was yingyang1 (YY1), and the targeting relationship was verified by dual-luciferase reporter assay. After deleting YY1, the expression of cartilage-specific genes was significantly down-regulated. In conclusion, our study demonstrated that Atr can promote chondrogenic differentiation of MSC through regulation of the ADAR2-miR-181a-5p signaling pathway. This may provide a new insight into the possible mechanism of traditional Chinese medicine (Atr) in treating inflammatory joint diseases.
苍术根茎（苍术 [RA]）长期以来在中国传统医学中被推荐用于治疗关节炎，但其作用机制仍不清楚。RA 含有大量苍术挥发油（Atr）。在本研究中，我们研究了 Atr 是否能促进间充质干细胞（MSCs）向软骨细胞分化。采用水蒸气蒸馏法从 RA 中提取 Atr，并通过 CCK8 法检测 Atr 对 MSCs 的作用。MSCs 培养的 Atr 最佳浓度为 3 μg/ml。通过 miRNA 微阵列分析筛选出差异表达的 miR-181a-5p，并将其模拟物和抑制剂转染到 MSCs 中。发现 miR-181a-5p 的抑制剂可上调软骨特异性基因如 SOX9、COL2A1 和 ACAN 的表达。同时，我们还发现基因编辑酶 ADAR2 的表达在 Atr 诱导的 MSCs 软骨分化过程中显著增加，且 miR-181a-5p 前体序列的碱基从 A 变为 G。ADAR2 缺失后，软骨特异性基因的表达显著下调，miR-181a-5p 的前体序列碱基未改变。生物信息学分析显示 miR-181a-5p 的预测靶基因是阴阳 1（YY1），并通过双荧光素酶报告基因检测验证了靶向关系。删除 YY1 后，软骨特异性基因的表达显著下调。总之，我们的研究表明 Atr 可通过调节 ADAR2-miR-181a-5p 信号通路促进 MSCs 的软骨分化。这可能为中药（Atr）治疗炎症性关节疾病的潜在机制提供新的见解。
