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 the occurrence and development of ABR.
作者
申意伟
李雪
刘颜
李佐
刘鑫
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)
出处
《中华中医药杂志》
CSCD
北大核心
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)。
