接骨木对酒精性骨重构骨保护、m6A甲基化及骨髓间充质干细胞成骨分化的作用  

Effects of elderberry on bone protection,m6A methylation and osteogenic differentiation of bone marrow mesenchymal stem cells in alcoholic bone remodeling

作　　者：申意伟 李雪 刘颜 李佐 刘鑫[6] SHEN Yiwei;LI Xue;LIU Yan;LI Zuo;LIU Xin(Binhai New Area Hospital of TCM.Tianjin(Fourth Teaching Hospital of Tanjin University of TCM),Tianjin 300000,China;Ningbo Hospital of Traditional Chinese Medicine(Ningbo Hospital of Traditional Chinese Medicine,Zhejiang Chinese Medical University),Ningbo 315010,China;Heilongjiang University of Chinese Medicine,Harbin 150040,China;Shenzhen Hospital,Beijing University of Chinese Medicine,Shenzhen 518172,China;School of Life Science,Northeast Normal University,Changchun 130024,China;Harbin Medical University,Harbin 150000,China)

机构地区：[1]天津市滨海新区中医医院(天津中医药大学第四附属医院),天津300000 [2]宁波市中医院(浙江中医药大学附属宁波中医院),宁波315010 [3]黑龙江中医药大学,哈尔滨150040 [4]北京中医药大学深圳医院,深圳518172 [5]东北师范大学生命科学院,长春130024 [6]哈尔滨医科大学,哈尔滨150000

出　　处：《中华中医药杂志》2024年第12期6740-6744,共5页China Journal of Traditional Chinese Medicine and Pharmacy

基　　金：国家自然科学基金项目(No.81904222);黑龙江中医药大学国家级项目配套(No.2019PT08);黑龙江省普通本科高等学校青年创新人才项目(No.UNPYSCT-2020228);黑龙江省博士后资助项目(No.LBH-Z20034);北京中医药大学深圳医院“三龙计划”项目(No.2022-BUCMSZYLRC08)。

摘　　要：目的:探讨接骨木对酒精性骨重构(ABR)骨保护、m6A甲基化及骨髓间充质干细胞成骨分化的作用。方法:雄性SD大鼠36只随机分为空白组、模型组、接骨木低剂量组和接骨木高剂量组,每组9只。模型组、接骨木低剂量组和接骨木高剂量组大鼠给予20%乙醇腹腔注射,日1次,共12周;空白组腹腔注射等量0.9%氯化钠溶液。造模开始第1天,接骨木低、高剂量组在模型组基础上分别给予340、680 mg/kg接骨木灌胃;空白组和模型组给予等量0.9%氯化钠溶液灌胃,均日1次。HE、Masson染色检测骨小梁、胶原纤维等变化;qRT-PCR检测骨代谢标记基因runt相关转录因子2(Runx2)、Ⅰ型胶原Alpha 2链(COL1A2)、骨形态发生蛋白2(BMP2)和骨保护素(OPG)表达情况;甲基化RIP-qPCR检测Runx2 m6A甲基化水平;Western Blot检测Runx2蛋白水平;DCFHDA染色检测活性氧含量;茜素红染色检测钙化结节形成情况。结果:与模型组比较,接骨木低、高剂量组骨小梁数目、胶原纤维含量增加,弹性模量、弹性载荷、最大应力显著增加(P<0.05),Runx2甲基化mRNA及蛋白质水平显著升高(P<0.05),活性氧含量明显减少,均趋近于空白组水平,骨形成相关基因BMP2、Runx2、COL1A2和OPG mRNA表达显著增加(P<0.05)。结论:接骨木可通过调节m6A RNA甲基化修饰和骨髓间充质干细胞成骨分化,增加骨小梁数目和胶原纤维含量,来恢复骨稳态,从而拮抗ABR的发生发展。Objective:To investigate the effects of elderberry on bone protection,m6A methylation and osteogenic differentiation of bone marrow mesenchymal stem cells(BMSCs)after alcoholic bone remodeling(ABR).Methods:A total of 36 male SD rats were randomly divided into blank group,model group,elderberry low-dose group and elderberry high-dose group,9 rats in each group.Rats in model group,elderberry low-dose group and elderberry high-dose group were given intraperitoneal injection of 20%ethanol once a day for 12 weeks.The blank group was intraperitoneally injected with the same amount of normal saline.On the first day of modeling,elderberry low-dose and high-dose group was given 340,680 mg/kg elderberry on the basis of model group,blank group and model group were given the same amount of normal saline intragastric administration,once a day.The changes of bone trabecula and collagen fibers were detected by HE and Masson staining.The expression of bone metabolic marker genes Runx2,COL1A2,BMP2 and OPG were detected by qRT-PCR.The methylation level of Runx2 m6A was detected by methylated RIP-qPCR.Runx2 protein levels were detected by Western Blot.The content of reactive oxygen species was detected by DCFH-DA staining.Alizarin red staining was used to detect the formation of calcified nodules.Results:Compared with model group,the number of bone trabeculae,collagen fiber content increased,elastic modulus,elastic load and maximum stress significantly increased(P<0.05),Runx2 methylation and protein levels significantly increased(P<0.05),while the content of reactive oxygen species was decreased,all of which were close to the level of blank group,bone formation related genes BMP2,Runx2,COL1A2,OPG mRNA expression significantly increased(P<0.05)in elderberry low-dose group and elderberry highdose group.Conclusion:By regulating m6A RNA methylation modification and osteogenic differentiation of BMSCs,elderberry can increase the number of bone trabeculae and the content of collagen fibers to restore bone homeostasis,thus antagonizing th

关 键 词：接骨木 酒精性骨重构 m6A甲基化 骨髓间充质干细胞 成骨分化 独神散 runt相关转录因子2 骨保护 

分 类 号：R73[医药卫生—肿瘤]