Department of Orthopedics, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, People's Republic of China.
Department of Orthopedics, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an 710061, People's Republic of China.
Bone. 2020 Oct;139:115522. doi: 10.1016/j.bone.2020.115522. Epub 2020 Jul 3.
To explore the role of cellular communication network factor 1 (CCN1) in cartilage inflammaging and osteoarthritis (OA) pathogenesis in the isolated primary human chondrocytes in vitro, cartilage explants ex vivo, and a pre-clinical mice model.
Recombinant human CCN1 stimulation and small interfering RNA inhibition were conducted in human chondrocytes. The RNA was extracted to quantify catabolic targets and pro-inflammatory genes and the proteins were probed with specific antibodies. IL-1β and IL-6 were monitored by ELISA. IHC was performed to evaluate important hypertrophic hallmarks and catabolic markers. The effects of Tanshinone IIA on chondrocytes were investigated in both time-dependent and dose-dependent processes. Cartilage explants were cultured in growth medium and further treated with Tanshinone IIA. The intra-articular injection was performed in 13 months old C57BL/6J mice. Safranin O and fast green staining were performed to evaluate the histological change of cartilage followed by a semi-quantitative analysis using the OARSI scoring system.
RNA and protein levels of CCN1 increased in an age-dependent manner compared to young donors. Increased CCN1 expression was also found in the damaged area compared to the non-lesion area which correlated with the advanced pathological change in human OA. The overexpression of CCN1 promoted chondrocytes senescence, while the down-regulation of CCN1 by small interfering RNA reduced CCN1 production and limited inflammation secretion suggesting that CCN1 was a possible novel target to intervene OA. Inhibition of CCN1 by using Tanshinone IIA could reduce SASP components in a dose- and time-dependent manner. Additionally, our data showed that Tanshinone IIA was able to preserve articular cartilage integrity, suppress CCN1 production, and inhibit SASP factors in human cartilage explants and in aged mice model.
This study showed that CCN1 signaling aggravated cartilage inflammaing and matrix degradation. Collectively, our findings showed new insight into repurposing Tanshinone IIA for slowing down OA advancement in human and mice by inhibiting the CCN1 axis.
在体外原代人软骨细胞、离体软骨外植体和临床前小鼠模型中，探索细胞通讯网络因子 1 (CCN1) 在软骨炎症和骨关节炎 (OA) 发病机制中的作用。
在人软骨细胞中进行重组人 CCN1 刺激和小干扰 RNA 抑制。提取 RNA 以定量代谢目标和促炎基因，并用特异性抗体探测蛋白质。通过 ELISA 监测 IL-1β 和 IL-6。免疫组化评估重要的肥大特征和代谢标志物。在时间和剂量依赖性过程中研究丹参酮 IIA 对软骨细胞的影响。在生长培养基中培养软骨外植体，并进一步用丹参酮 IIA 处理。在 13 个月大的 C57BL/6J 小鼠中进行关节内注射。用番红 O 和固绿染色评估软骨的组织学变化，然后使用 OARSI 评分系统进行半定量分析。
与年轻供体相比，CCN1 的 RNA 和蛋白水平随年龄增长呈依赖性增加。在受损区域与非病变区域相比，CCN1 的表达也增加，这与人类 OA 的进展性病理变化相关。CCN1 的过表达促进软骨细胞衰老，而小干扰 RNA 下调 CCN1 可减少 CCN1 的产生并限制炎症分泌，这表明 CCN1 可能是干预 OA 的一个新的潜在靶点。丹参酮 IIA 抑制 CCN1 可呈剂量和时间依赖性减少 SASP 成分。此外，我们的数据表明，丹参酮 IIA 能够在人软骨外植体和老年小鼠模型中维持关节软骨完整性，抑制 CCN1 产生并抑制 SASP 因子。
本研究表明 CCN1 信号加重了软骨炎症和基质降解。总之，我们的研究结果表明，丹参酮 IIA 通过抑制 CCN1 轴在人类和小鼠中为减缓 OA 进展提供了新的见解。
