湖南中医药大学学报2025，Vol.45Issue(9)：1644-1652,9.DOI:10.3969/j.issn.1674-070X.2025.09.008
桃红四物汤通过GLUL/NOTCH通路促进BMMSCs成骨分化及核糖体生成研究
Taohong Siwu Decoction promoting osteogenic differentiation and ribosome biogenesis of BMMSCs through the CLUL/NOTCH pathway
摘要
Abstract
Objective To explore the specific mechanisms of Taohong Siwu Decoction(THSWD)in promoting osteogenic differentiation of bone marrow mesenchymal stem cells(BMMSCs)and affecting fracture healing.Methods BMMSCs from rats were cultured in vitro.Well-growing BMMSCs were selected and divided into the following groups:the control(CONTROL)group,the THSWD intervention(THSWD)group,the THSWD intervention+glutamine synthetase(GLUL)knockdown or overexpression blank control(THSWD+sh-NC or THSWD+OE-NC)group,and the THSWD intervention+GLUL knockdown or overexpression(THSWD+sh-GLUL or THSWD+OE-GLUL)group.Cells in the CONTROL group were cultured in complete BMSC medium.The THSWD group was treated with 100 ng/mL THSWD-containing serum.The THSWD+sh-NC and THSWD+OE-NC groups were incubated with 100 ng/mL THSWD-containing serum and then transfected with sh-NC and OE-NC lentiviruses,respectively.The THSWD+sh-GLUL and THSWD+OE-GLUL groups were also treated with 100 ng/mL THSWD-containing serum followed by transduction with sh-GLUL and OE-GLUL lentiviruses,respectively.The protein expression levels of alkaline phosphatase(ALP),Osterix(Osx),runt-related transcription factor 2(Runx2),GLUL,NOTCH intracellular domain(NICD),neurogenic locus notch homolog protein 1(NOTCH1),and hairy and enhancer of split-1(HES1)were detected by Western blot.Mineralization was assessed using Alizarin Red staining.The expression of 5S rRNA,5.8S rRNA,18S rRNA,and 28S rRNA was measured via qRT-PCR.Network pharmacology analysis was employed to predict and identify GLUL as a key target through which THSWD regulates ribosome biogenesis to affect BMMSCs.Cell viability was evaluated using the CCK-8 assay,and cell migration distance was quantified using the Transwell assay.Results Compared with the CONTROL group,the expression levels of ALP,Osx,and Runx2 significantly increased in the THSWD group(P<0.01).Alizarin Red staining results were positive in BMMSCs of the THSWD group.The expression of key ribosome biogenesis factors,including 5S rRNA,5.8S rRNA,18S rRNA,and 28S rRNA,was also markedly upregulated in the THSWD group(P<0.01).Network pharmacology analysis identified GLUL as a key target of THSWD.Cellular validation experiments showed that,compared with the THSWD+sh-NC group,the THSWD+sh-GLUL group exhibited decreased cell proliferation ability,protein expression levels of ALP,Osx,and Runx2,cell migration capacity and wound healing rate,expression of 5S/5.8S/18S/28S rRNAs,and protein expression levels of NICD,NOTCH1,and HES1(all P<0.01).Conversely,compared with the THSWD+OE-NC group,the THSWD+OE-GLUL group showed increases in all these parameters(all P<0.01).Conclusion THSWD may promote osteogenic differentiation of BMMSCs by upregulating the expression of GLUL,activating the expression of the NOTCH signaling pathway,and thereby enhancing ribosome biogenesis.关键词
骨折愈合/桃红四物汤/骨髓间充质干细胞/核糖体生成/NOTCH通路Key words
fracture healing/Taohong Siwu Decoction/bone marrow mesenchymal stem cells/ribosome biogenesis/NOTCH signaling pathway分类
医药卫生引用本文复制引用
陈培,简功辉,杨雷,陈振华,张天择,叶政显,齐新宇,熊辉,陆小龙..桃红四物汤通过GLUL/NOTCH通路促进BMMSCs成骨分化及核糖体生成研究[J].湖南中医药大学学报,2025,45(9):1644-1652,9.基金项目
湖南中医药大学"刘良院士工作站"指导项目(22YS004) （22YS004）
湖南创新型省份建设重点领域研发计划项目(2023SK2047) （2023SK2047）
中医药防治筋骨疾病基础研究创新平台(kh2201056) （kh2201056）
国家自然科学基金面上项目(81874478) （81874478）
湖南省自然科学基金项目(2024JJ9419) （2024JJ9419）
湖南省教育厅科学研究项目(23A0311) （23A0311）
长沙市自然科学基金项目(kq2208197). （kq2208197）
