摘要
目的探讨黄芪多糖通过调控微小RNA-375(microRNA-375,miR-375)对人牙周膜干细胞生物学特性的影响及其相关机制的研究。方法体外分离培养人牙周膜干细胞,采用黄芪多糖浓度为0、40、100、400μg/mL进行处理48 h,记为不同浓度的黄芪多糖组;将anti-miR-NC和anti-miR-375转染至人牙周膜干细胞,选取400μg/mL的黄芪多糖进行处理,记为黄芪多糖+anti-miR-NC组和黄芪多糖+anti-miR375组。通过噻唑蓝(methylthiazolyldiphenyl-tetrazolium bromide,MTT)检测细胞增殖能力和碱性磷酸酶(alkaline phosphatase,ALP)活性;蛋白免疫印迹法(Western blot)检测Runt相关转录因子2(runt-related transcription factor 2,Runx-2)、骨桥蛋白(osteopontin,OPN)和骨钙素(osteocalcin,OCN)的蛋白表达;实时荧光定量聚合酶链式反应(real-time polymerase chain reaction,qRT-PCR)检测miR-375的表达。结果不同浓度的黄芪多糖组可以促进miR-375表达(1.54±0.16、1.92±0.20、2.54±0.22)(0.92±0.07)、细胞A值[(0.46±0.05、0.58±0.07、0.72±0.06)比(0.35±0.04),(0.53±0.05、0.69±0.07、0.84±0.09)(0.40±0.03)]、ALP活性(0.73±0.08、0.84±0.07、1.03±0.09)比(0.32±0.03),Runx-2(1.32±0.11、1.51±0.14、1.83±0.17)(0.92±0.07)、OCN(1.47±0.13、1.65±0.16、1.89±0.19)(0.99±0.06)、OPN(1.53±0.15、1.72±0.16、1.99±0.20)(1.00±0.04)蛋白表达上调。黄芪多糖+anti-miR-375组的miR-375的表达、细胞A值、ALP活性,Runx-2、OCN、OPN蛋白表达明显高于黄芪多糖+anti-miR-NC组。结论黄芪多糖可以促进人牙周膜干细胞的增殖和成骨分化能力,其作用机制可能与miR-375有关。
Objective To investigate the effect of astragalus polysaccharides on human periodontal ligament stem cells(PDLSCs)and the related mechanisms by regulating microRNA-375(miR-375).Methods In this study,human PDLSCs were isolated and cultured in vitro.Cells were treated with 0,40,100,400μg/mL astragalus polysaccharide for 48 h.Cells in the astragalus polysaccharides+anti-miR-NC group and astragalus polysaccharides+anti-miR-375 group were transfected with anti-miR-NC and anti-miR-375 respectively,and treated with 400μg/mL astragalus polysaccharides for 48 h.Alkaline phosphatase(ALP)activity were measured and the proliferation of PDLSCs was assayed by using methylthiazolyldiphenyl-tetrazolium bromide(MTT).The expression levels of Runt-related transcription factor 2(Runx-2),osteopontin(OPN)and osteocalcin(OCN)were detected by Western blotting.The expression level of miR-375 was detected by real-time quantitative polymerase chain reaction(qRT-PCR).Results Astragalus polysaccharide can promote the expression of miR-375[(1.54±0.16,1.92±0.20,2.54±0.22)of the astragalus polysaccharide groups vs.(0.92±0.07)of the control group].Astragalus polysaccharide treatment promoted the proliferation of PDLSCs,the A values were(0.46±0.05,0.58±0.07,0.72±0.06)of the astragalus polysaccharide groups vs.(0.35±0.04)of the control group for 48 h treatment,and(0.53±0.05,0.69±0.07,0.84±0.09)of the astragalus polysaccharide groups vs.(0.40±0.03)of the control group for 72 h.The values for ALP activity were(0.73±0.08,0.84±0.07,1.03±0.09)of the astragalus polysaccharide groupsvs.(0.32±0.03)of the control group.Treatment with various concentrations of astragalus polysaccharide also up-regulated the expression of Runx-2[(1.32±0.11,1.51±0.14,1.83±0.17)vs.(0.92±0.07)],OCN[(1.47±0.13,1.65±0.16,1.89±0.19)vs.(0.99±0.06)],OPN[(1.53±0.15,1.72±0.16,1.99±0.20)vs.(1.00±0.04)].The expression level of miR-375,cell proliferation,ALP activity,and the expression levels of Runx-2,OCN,OPN of astragalus polysaccharide+antimiR-375 group w
作者
李鲲
谢陆莉
邓琳
LI Kun;XIE Luli;DENG Lin(Department of Stomatology,First People’s Hospital of Longquanyi District,Chengdu,610100,China)
出处
《医学分子生物学杂志》
CAS
2022年第6期496-500,共5页
Journal of Medical Molecular Biology
关键词
黄芪多糖
miR-375
人牙周膜干细胞
增殖
成骨分化
astragalus polysaccharide
miR-375
human periodontal ligament stem cells
proliferation
osteogenic differentiation
