Lu Tao, Liu Zi-Yi, Ge Yang-Shuo, Jiang Shuai-Yu, Zhao Qing-Ao, Ding Dao-Fang
School of Traditional Chinese Medicine, Shanxi Datong University, Datong 037009, China.
Datong Key Laboratory of Smart Medicine and Health Care for Elderly Chronic Diseases, Shanxi Datong University, Datong 037009, China.
Int J Mol Sci. 2025 Jun 9;26(12):5519. doi: 10.3390/ijms26125519.
Osteoarthritis (OA) is a debilitating joint disorder characterized by cartilage degradation and disruption of chondrocyte homeostasis. Although retinoic acid (RA) has been used in OA models, its precise targets are not clear. A translational framework was employed, integrating RNA-sequencing results, network pharmacology prediction, computational ligand-receptor molecular docking, and biological experimental validation, to systematically elucidate RA's disease-modifying targets in OA pathogenesis. RNA-sequencing of RA-treated chondrocytes revealed 656 differentially expressed genes (DEGs). Protein-protein interaction (PPI) network analysis and functional enrichment [Gene Ontology (GO)/Kyoto Encyclopedia of Genes and Genomes (KEGG)] highlighted key pathways, including extracellular matrix (ECM) reorganization and PI3K-Akt-mediated mechanotransduction and others. Network pharmacology analysis identified 42 shared targets between RA and OA. PPI analysis and functional enrichment (GO/KEGG) highlighted pathways including the renin-angiotensin system and the neuroactive ligand-receptor interaction, among others. Molecular docking ranked candidate targets by binding affinity of RA in descending order as MAPK14 (p38α), PTGER3 (PGE2 receptor), CA2 (CA2), and others. Five intersecting targets CA2, ACE, PTGS1 (COX-1), PGR, and EDNRA (ETAR) were identified by integrating RNA-sequencing (RNA-seq) results and network pharmacology predictions. These interactions were experimentally validated via western blot, RT-qPCR and immunofluorescence. RA increased the expression of MMP13, CA2 and ACE, and decreased the expression of COL2A1 in chondrocytes. siRNA-mediated knockdown of both CA2 (human CA2 homolog) and ACE (human ACE homolog) inhibit cartilage degradation through downregulating MMP13 and upregulating COL2A1. This study not only elucidates potential molecular mechanisms by which RA modulates chondrocyte catabolism but also offers a valuable reference for the development of novel OA therapeutics.
骨关节炎（OA）是一种使人衰弱的关节疾病，其特征是软骨降解和软骨细胞稳态破坏。尽管视黄酸（RA）已用于OA模型，但其确切靶点尚不清楚。采用了一个转化框架，整合RNA测序结果、网络药理学预测、计算配体-受体分子对接和生物学实验验证，以系统地阐明RA在OA发病机制中的疾病修饰靶点。对经RA处理的软骨细胞进行RNA测序，发现了656个差异表达基因（DEG）。蛋白质-蛋白质相互作用（PPI）网络分析和功能富集[基因本体论（GO）/京都基因与基因组百科全书（KEGG）]突出了关键途径，包括细胞外基质（ECM）重组和PI3K-Akt介导的机械转导等。网络药理学分析确定了RA和OA之间的42个共同靶点。PPI分析和功能富集（GO/KEGG）突出了包括肾素-血管紧张素系统和神经活性配体-受体相互作用等途径。分子对接根据RA的结合亲和力将候选靶点按降序排列为MAPK14（p38α）、PTGER3（PGE2受体）、CA2（CA2）等。通过整合RNA测序（RNA-seq）结果和网络药理学预测，确定了五个相交靶点CA2、ACE、PTGS1（COX-1）、PGR和EDNRA（ETAR）。通过蛋白质免疫印迹、RT-qPCR和免疫荧光对这些相互作用进行了实验验证。RA增加了软骨细胞中MMP13、CA2和ACE的表达，并降低了COL2A1的表达。siRNA介导的CA2（人CA2同源物）和ACE（人ACE同源物）敲低通过下调MMP13和上调COL2A1抑制软骨降解。本研究不仅阐明了RA调节软骨细胞分解代谢的潜在分子机制，也为新型OA治疗药物的开发提供了有价值的参考。
