METTL3 promotes experimental osteoarthritis development by regulating inflammatory response and apoptosis in chondrocyte
METTL3 promotes experimental osteoarthritis development by regulating inflammatory response and apoptosis in chondrocyte
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DOI：
10.1016/j.bbrc.2019.05.168
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发表时间：
2019-08-13
影响因子：
3.1
通讯作者：
Fu, Bin
中科院分区：
文献类型：
--
作者：
Liu, Qingbai;Li, Meng;Fu, Bin
目的：研究RNA甲基转移酶METTL3（一种催化n6 -甲基腺苷（m6A）在靶mRNA上形成的酶）在骨关节炎（OA）发生中的功能作用及其机制。方法：采用细胞因子IL-1 β刺激软骨祖细胞ATDC5细胞，模拟体外炎症反应。采用qRT-PCR检测METTL3 mRNA、m6A及炎症因子水平。CCK-8检测细胞活性。流式细胞术检测细胞凋亡率。Western blot检测细胞内nf - κ B信号分子和胶原蛋白水平。采用甲基化抑制剂环亮氨酸和甲基供体甜菜碱治疗胶原酶诱导的OA小鼠。结果：在IL-1 β处理的ATDC5细胞中，METTL3 mRNA水平和m6A甲基化mRNA占总mRNA的百分比呈剂量依赖性增加。shRNA沉默METTL3降低了IL-1 β诱导的细胞凋亡百分比，抑制了IL-1 β诱导的炎症细胞因子水平升高和软骨细胞中nf - κ B信号的激活。此外，METTL3的沉默通过降低MMP-13和Coll X的表达，提高Aggrecan和Coll II的表达，促进细胞外基质（ECM）的降解。在胶原酶诱导的OA小鼠模型中，注射甲基化抑制剂环油氨酸或甲基供体甜菜碱不影响METTL3 mRNA的表达，但分别显著抑制或促进m6A总水平以及炎症状况和ECM降解。结论：METTL3通过调节NF-kappa B信号和软骨细胞中ECM的合成，在骨关节炎的进展中具有功能作用，这有助于通过关注METTL3和mRNA甲基化来制定OA的预防和治疗策略。(C) 2019 Elsevier Inc.版权所有。
Objective: This study was to investigate the functional role of RNA methyltransferase METTL3, an enzyme catalyzes the formation of N6-methyladenosine (m6A) on the target mRNA, in the development of osteoarthritis (OA) and the underlying mechanism.Methods: Cytokine IL-1 beta was used to stimulate the chondroprogenitor cell line ATDC5 cells to mimic the inflammatory condition in vitro. The level of METTL3 mRNA and m6A as well as inflammatory cytokines were detected by qRT-PCR. Cell activity was detected by CCK-8. The rate of apoptotic cell was measured by flow cytometry. Western blot was used to detect the levels of NF-kappa B signaling molecules and collagen in cells. Methylation inhibitor cycloleucine and methyl donor betaine were used to treat collagenase-induced OA mice.Results: In IL-1 beta-treated ATDC5 cells, the METTL3 mRNA levels and the percentage of m6A methylated mRNA of total mRNA were increased in a dose-dependent manner. Silencing of METTL3 by shRNA reduced the percentage of IL-1 beta-induced apoptosis, suppressed IL-1 beta-induced increased inflammatory cytokines levels and activation of NF-kappa B signaling in chondrocytes. Moreover, silencing of METTL3 promotes degradation of extracellular matrix (ECM) by reducing the expression of MMP-13 and Coll X, elevating the expression of Aggrecan and Coll II. In a OA mouse model induced by collagenase, injection of methylation inhibitor cycloleucine or methyl donor betaine does not affects METTL3 mRNA expression, but significantly inhibits or promotes the total level of m6A as well as inflammatory condition and ECM degradation, respectively.Conclusion: METTL3 has a functional role in mediates osteoarthritis progression by regulating NF-kappa B signaling and ECM synthesis in chondrocytes that shed insight on developing preventive and curative strategies for OA by focusing on METTL3 and mRNA methylation. (C) 2019 Elsevier Inc. All rights reserved.