1Department of Orthopedics, First Affiliated Hospital of Kunming Medical University, Kunming, 650032 Yunnan Province People's Republic of China.
Department of Orthopedics, Dalian University Affiliated Xinhua Hospital, No. 156 Xinhua Street, Shahekou District, Dalian, 116021 Liaoning Province People's Republic of China.
Cell Mol Biol Lett. 2018 Feb 22;23:7. doi: 10.1186/s11658-018-0072-6. eCollection 2018.
Osteoarthritis is characterized by the continuous degradation of the articular cartilage. The microRNA miR-448 has been found to be broadly involved in cellular processes, including proliferation, apoptosis, invasion and EMT. While aberrant expression of miR-448 has been found in multiple cancers, its level in osteoarthritis cartilage and its role in the progression of this disease are still unknown. Here, we examined the functional roles of miR-448 and its expression in osteoarthritis tissues, including IL-1β-stimulated osteoarthritis chondrocytes.
Chondrocytes were isolated from human articular cartilage and stimulated with IL-1β. The expression levels of miR-448 in the cartilage and chondrocytes were both determined. After transfection with an miR-448 mimic or inhibitor, the mRNA levels of aggrecan, type II collagen and MMP-13 were determined. Luciferase reporter assay, qRT-PCR and western blot were performed to explore whether matrilin-3 was a target of miR-448. Furthermore, we co-transfected chondrocytes with miR-448 inhibitor and siRNA for matrilin-3 and then stimulated them with IL-1β to determine whether miR-448-mediated IL-1β-induced cartilage matrix degradation resulted from directly targeting matrilin-3.
The level of miR-448 was significantly higher and matrilin-3 expression was significantly lower in osteoarthritis cartilage and IL-1β-induced chondrocytes than in normal tissues and cells. Furthermore, matrilin-3 expression was reduced by miR-448 overexpression. MiR-448 downregulation significantly alleviated the IL-1β-induced downregulation of aggrecan and type II collagen expression, and upregulation of MMP-13 expression. MiR-448 overexpression had the opposite effects. Knockdown of matrilin-3 reversed the effects of the miR-448 inhibitor on the expressions of aggrecan, type II collagen and MMP-13.
The findings showed that miR-448 contributed to the progression of osteoarthritis by directly targeting matrilin-3. This indicates that it has potential as a therapeutic target for the treatment of osteoarthritis.
骨关节炎的特征是关节软骨的持续降解。已经发现 microRNA miR-448 广泛参与细胞过程，包括增殖、凋亡、侵袭和 EMT。虽然 miR-448 的异常表达已在多种癌症中发现，但它在骨关节炎软骨中的水平及其在疾病进展中的作用尚不清楚。在这里，我们研究了 miR-448 的功能作用及其在包括 IL-1β 刺激的骨关节炎软骨细胞在内的骨关节炎组织中的表达。
从人关节软骨中分离软骨细胞，并用 IL-1β 刺激。确定软骨和软骨细胞中 miR-448 的表达水平。转染 miR-448 模拟物或抑制剂后，测定聚集蛋白聚糖、II 型胶原和 MMP-13 的 mRNA 水平。进行荧光素酶报告基因检测、qRT-PCR 和 Western blot 以探索 matrilin-3 是否是 miR-448 的靶标。此外，我们将 miR-448 抑制剂和 matrilin-3 的 siRNA 共转染软骨细胞，然后用 IL-1β 刺激它们，以确定 miR-448 介导的 IL-1β 诱导的软骨基质降解是否是由于直接靶向 matrilin-3。
与正常组织和细胞相比，骨关节炎软骨和 IL-1β 诱导的软骨细胞中的 miR-448 水平显著升高，而 matrilin-3 的表达水平显著降低。此外，miR-448 的过表达降低了 matrilin-3 的表达。miR-448 的下调显著缓解了 IL-1β 诱导的聚集蛋白聚糖和 II 型胶原表达下调以及 MMP-13 表达上调。miR-448 的过表达则产生相反的效果。matrilin-3 的敲低逆转了 miR-448 抑制剂对 aggrecan、II 型胶原和 MMP-13 表达的影响。
研究结果表明，miR-448 通过直接靶向 matrilin-3 促进骨关节炎的进展。这表明它具有作为骨关节炎治疗的治疗靶点的潜力。
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