Department of Orthopaedics, The Wangjing Hospital, Chinese Academy of Traditional Chinese Medicine, Beijing, China.
Department of Pediatrics, Dongzhimen Hospital, Beijing University of Traditional Chinese Medicine, Beijing, China.
Orthop Surg. 2021 Feb;13(1):276-284. doi: 10.1111/os.12845. Epub 2020 Dec 6.
To confirm the role of long noncoding RNA differentiation antagonizing non-protein coding RNA (DANCR) in chondrocyte inflammatory injury in osteoarthritis (OA) in vitro, as well as its molecular mechanism.
Human primary chondrocytes were treated with lipopolysaccharide (LPS) to construct a chondrocyte inflammatory injury in human OA cell model. Gene expression was detected using real-time quantitative polymerase chain reaction. Cell inflammatory injury was evaluated by Cell Counting Kit-8 assay, flow cytometry, and enzyme-linked immunosorbent assay. The interplay between miRNA-19a-3p (miR-19a) and DANCR was validated by dual-luciferase reporter assay and RNA immunoprecipitation.
Expression of DANCR was upregulated, and miR-19a was downregulated in human OA cartilage and LPS-treated primary chondrocytes in vitro. Moreover, DANCR expression was inversely correlated with miR-19a in OA patients. LPS reduced cell viability and increased the apoptotic rate and secretion of interleukin (IL)-1β, IL-6, IL-8, as well as tumor necrosis factor (TNF)-α in primary chondrocyte cells in vitro, suggesting an inflammatory injury model of OA. Functionally, knockdown of DANCR could attenuate LPS-induced apoptosis and inflammatory response, as evidenced by improved cell viability, and reduced apoptotic rate and products of IL-1β, IL-6, IL-8, and TNF-α. Notably, DANCR negatively regulated miR-19a expression, presumably via sponging. Furthermore, miR-19a deletion eliminated the effect of DANCR knockdown on apoptosis and the inflammatory response of primary chondrocytes under LPS stress.
Differentiation antagonizing non-protein coding RNA silencing could protect human chondrocyte cells against LPS-induced inflammatory injury and apoptosis through targeting miR-19a, suggesting a vital role of the DANCR/miR-19a axis in OA.
在体外确认长链非编码 RNA 分化拮抗非编码 RNA（DANCR）在骨关节炎（OA）软骨细胞炎症损伤中的作用及其分子机制。
用人脂多糖（LPS）处理原代软骨细胞，构建人 OA 细胞模型中的软骨细胞炎症损伤。采用实时定量聚合酶链反应检测基因表达。通过细胞计数试剂盒-8 检测、流式细胞术和酶联免疫吸附试验评估细胞炎症损伤。通过双荧光素酶报告基因检测和 RNA 免疫沉淀验证 miRNA-19a-3p（miR-19a）与 DANCR 之间的相互作用。
在人 OA 软骨和体外 LPS 处理的原代软骨细胞中，DANCR 表达上调，miR-19a 表达下调。此外，OA 患者中 DANCR 表达与 miR-19a 呈负相关。LPS 降低原代软骨细胞的活力，增加细胞凋亡率和白细胞介素（IL）-1β、IL-6、IL-8 和肿瘤坏死因子（TNF）-α的分泌，提示建立了 OA 的炎症损伤模型。功能上，DANCR 敲低可减轻 LPS 诱导的凋亡和炎症反应，表现为细胞活力提高，细胞凋亡率降低，IL-1β、IL-6、IL-8 和 TNF-α产物减少。值得注意的是，DANCR 通过海绵作用负调控 miR-19a 表达。此外，miR-19a 缺失消除了 DANCR 敲低对 LPS 应激下原代软骨细胞凋亡和炎症反应的影响。
分化拮抗非编码 RNA 沉默通过靶向 miR-19a 可保护人软骨细胞免受 LPS 诱导的炎症损伤和凋亡，表明 DANCR/miR-19a 轴在 OA 中具有重要作用。
