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| 组蛋白甲基转移酶2（SMYD2）抑制剂LLY-507对成骨分化的影响 |
| SET and MYND domain containing protein 2 inhibitor LLY-507 antagonizes osteogenic differentiation by MC3T3-E1 cells and human bone marrow mesenchymal stem cells |
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| DOI：10.3969/j.issn.1006.7108.2022.03.012 |
| 中文关键词: SMYD2 LLY-507 MC3T3-E1 hMSCs 成骨分化 |
| 英文关键词:SMYD2 LLY-507 MC3T3-E1 hMSCs osteogenic differentiation |
| 基金项目:国家自然科学基金面上项目（基于ChIP-Seq技术探讨组蛋白修饰在补肾法防治骨质疏松症中的表观遗传机制）（81774339）；国家自然科学基金面上项目（从肾主骨生髓探讨骨质疏松症中m6A去甲基化转移酶FTO介导成骨的分子机制研究）（82074462）；国家自然科学基金青年科学基金项目（LncRNA MALAT1介导组蛋白甲基化修饰在补肾法调控成骨-破骨细胞耦联维持骨稳态中表观遗传机制）（82004395） |
| 作者 | 单位 | | 李建良1,2 张濛3 麦嘉乐1,2 何琪1,2 张罡瑜1,2 陈伟坚1,2 肖嘉聪1,2 潘兆丰1,2 宫大伟1,2,4 王海彬1,2,5* | 1. 广州中医药大学，广东 广州 510405
2. 广州中医药大学岭南医学研究中心中医骨伤科实验室，广东 广州 510405
3. 河南省人民医院，郑州大学人民医院，河南大学人民医院骨科, 河南 郑州 450003
4.山东省文登整骨医院，山东 威海 264400
5. 广州中医药大学第一附属医院，广东 广州 510405 |
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| 中文摘要: |
| 目的 研究SMYD2抑制剂LLY-507对小鼠颅顶前骨细胞亚克隆14（MC3T3-E1）及人骨髓间充质干细胞（hMSCs）成骨分化的影响。方法 通过CCK-8法检测LLY-507对MC3T3-E1和hMSCs增殖的影响；采用碱性磷酸酶( ALP) 染色和活性测定检测不同浓度LLY-507干预下MC3T3-E1和hMSCs早期成骨分化标志物ALP的活性情况；通过茜素红染色及半定量分析检测LLY-507对MC3T3-E1和hMSCs矿化能力的影响；运用荧光定量PCR检测hMSCs的成骨相关基因Osterix、Runx2、OPG、OPN mRNA的表达；Western blot检测hMSCs的Runx2和SMYD2蛋白表达量。结果 CCK-8结果表明LLY-507在0.8 μmol/L的剂量下细胞活性无明显影响，不会明显抑制细胞增殖。ALP染色及活性测定表明，加入50 nmol/L及以上剂量的LLY-507干预7 d后能够明显抑制MC3T3-E1及hMSCs成骨分化以及碱性磷酸酶的表达（P<0.05）；茜素红染色及半定量分析表明100 nmol/L剂量的LLY-507能够明显抑制MC3T3-E1及hMSCs的矿化能力（P<0.05）。通过qPCR，加入25 nmol/L及以上剂量的LLY-507分别干预7 d和14 d后能够明显抑制hMSCs成骨分化相关基因Runx2、Osterix、OPG、OPN的mRNA表达（P<0.05）。通过Western blot, 加入50 nmol/L及以上剂量的LLY-507干预7 d和14 d后均能够明显抑制hMSCs的Runx2蛋白和SMYD2蛋白的表达。结论 抑制SMYD2的表达能够抑制成骨相关基因的表达，说明SMYD2在成骨分化过程中起到促进成骨的作用。 |
| 英文摘要: |
| Objective To observe the effect of SET and MYND domain containing protein 2 (SMYD2) inhibitor LLY-507 on osteogenic differentiation of MC3T3-E1 cells and human bone marrow mesenchymal stem cells (hMSCs). Methods The effect of LLY-507 on cell survival of MC3T3-E1 and hMSCs was detected with CCK-8 assay. The early osteogenic differentiation marker alkaline phosphatase (ALP) was detected with ALP staining and ALP activity in MC3T3-E1 and hMSCs intervened with LLY-507 at different concentrations. The effect of LLY-507 on mineralization ability of MC3T3-E1 and hMSCs was detected with Alizarin red staining and semi-quantitative analysis. The mRNA expression of Osterix, Runx2, OPG, and OPN in hMSCs was detected with real-time PCR. Runx2 and SMYD2 protein expression was detected with Western blotting. Results LLY-507 had no significant effect on cell activity at the dose of 0.8 μmol/L and did not significantly inhibit cell proliferation as shown in CCK-8 assay. The results of ALP staining and activity examination showed that LLY-507 significantly inhibit MC3T3-E1 and hMSCs osteogenic differentiation at 50nmol/L and above dose for 7 days (P<0.05). The mineralization ability of MC3T3-E1 and hMSCs was inhibited at 100nmol/L dose of LLY-507 as shown in Alizarin red staining and semi-quantitative analysis. mRNA expressions of Runx2, Osterix, OPG, and OPN in hMSCs were significantly inhibited when 25nmol/L and above dose of LLY-507 was additioned for 7 days and 14 days as shown with qPCR (P<0.05). The expressions of Runx2 and SMYD2 protein in hMSCs were significantly inhibited when LLY-507 at doses of 50nmol/L and above was additioned for 7 days and 14 days. Conclusion Inhibition of SMYD2 expression inhibits the level of osteogenic related genes expression, indicating that SMYD2 plays a role in promoting osteogenic differentiation. |
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