Cao Zhen, Liu Chuan, Bai Yun, Dou Ce, Li Jian-Mei, Shi Duo-Wei, Dong Shi-Wu, Xiang Qiang
Department of Anatomy, Third Military Medical UniversityChongqing, China.
Department of Biomedical Materials Science, Third Military Medical UniversityChongqing, China.
Am J Transl Res. 2017 Jun 15;9(6):2748-2759. eCollection 2017.
Chondrocytes located in hyaline cartilage may maintain phenotype while the chondrocytes situated in calcified cartilage differentiate into hypertrophy. Chondrogenic and hypertrophic differentiation of mesenchymal stem cells (MSCs) are two subsequent processes during endochondral ossification. However, it is necessary for chondrocytes to hold homeostasis and to inhibit hypertrophic differentiation in stem cell-based regenerated cartilage. Dihydroartemisinin (DHA) is derived from artemisia apiacea which has many biological functions such as anti-malarial and anti-tumor. Whereas the effects of DHA on chondrogenic and hypertrophic differentiation are poorly understand. In this study, the cytotoxicity of DHA was determined by CCK8 assay and the cell apoptosis was analyzed by flow cytometry. Additionally, the effects of DHA on chondrogenic and hypertrophic differentiation of MSCs are explored by RT-PCR, western blotting and immunohistochemistry. The results showed that DHA inhibited expression of chondrogenic markers including Sox9 and Col2a1 by activating Nrf2 and Notch signaling. After induced to chondrogenesis, cells were treated with hypertrophic induced medium with DHA. The results revealed that hypertrophic markers including Runx2 and Col10a1 were down-regulated following DHA treatment through Pax6/HOXA2 and Gli transcription factors. These findings indicate that DHA is negative to chondrogenesis and is protective against chondrocyte hypertrophy to improve chondrocytes stability. Therefore, DHA might be not suited for chondogenesis but be potential as a new therapeutic candidate to maintain the biological function of regenerated cartilage.
位于透明软骨中的软骨细胞可维持其表型，而位于钙化软骨中的软骨细胞则分化为肥大细胞。间充质干细胞（MSC）的软骨生成和肥大分化是软骨内骨化过程中的两个后续过程。然而，在基于干细胞的再生软骨中，软骨细胞保持内环境稳定并抑制肥大分化是必要的。双氢青蒿素（DHA）源自青蒿，具有许多生物学功能，如抗疟疾和抗肿瘤。然而，DHA对软骨生成和肥大分化的影响尚不清楚。在本研究中，通过CCK8法测定DHA的细胞毒性，并通过流式细胞术分析细胞凋亡。此外，通过RT-PCR、蛋白质印迹和免疫组织化学探讨了DHA对MSC软骨生成和肥大分化的影响。结果表明，DHA通过激活Nrf2和Notch信号通路抑制包括Sox9和Col2a1在内的软骨生成标志物的表达。诱导细胞软骨生成后，用含DHA的肥大诱导培养基处理细胞。结果显示，通过Pax6/HOXA2和Gli转录因子，DHA处理后包括Runx2和Col10a1在内的肥大标志物表达下调。这些发现表明，DHA对软骨生成具有负性作用，对软骨细胞肥大具有保护作用，可提高软骨细胞的稳定性。因此，DHA可能不适合用于软骨生成，但作为维持再生软骨生物学功能的新治疗候选药物具有潜力。
