摘要
目的探究铁在绝经后大鼠模型中对骨髓间充质干细胞(bone marrow derived mesenchymal stem cell,BMSCs)成骨代谢影响,方法选取赣南医科大学从北京动物实验中心购置的50只大鼠为研究对象,按照随机数字表法分为模型组(40只)和空白组(10只),模型组进行双侧卵巢切除,随后接受铁铵柠檬酸盐(ferric acid,FAC)干预,又分为模型组、低、中、高剂量组,每组10只。通过酶联免疫吸附测定、实时荧光定量逆转录聚合酶链反应检测各组大鼠的骨代谢、成骨分化和铁代谢指标,随后取大鼠股骨培养原代BMSCs细胞并进行铁剂干预,进行CCK-8、免疫印迹检测其机制。结果空白组、模型组、低剂量组、中剂量组、高剂量组碱性磷酸酶(alkaline phosphatase,ALP)水平分别为(28.45±4.15)U/L、(22.49±3.45)U/L、(17.46±2.38)U/L、(13.44±1.79)U/L、(11.56±1.33)U/L,模型组ALP值随铁剂用量增加而下降,高剂量组最低,差异有统计学意义(F=68.300,P<0.05);模型组氨基端中段骨钙素值随着铁剂用量增加而下降,高剂量组最低,差异有统计学意义(P<0.05)。模型组大鼠成骨相关转录因子2、骨桥蛋白表达、骨钙素表达明显下降,且随着铁剂用量增加,表达水平逐渐降低,高剂量组最低,差异均有统计学意义(P均<0.05)。模型组总铁结合力、转铁蛋白饱和度和血清铁蛋白水平随着铁剂用量增加而上升,差异均有统计学意义(P均<0.05)。FAC浓度≥200μmol/L促进细胞凋亡,显示高浓度FAC对BMCSs细胞的潜在毒性,处理48 h后,200μmol/L组细胞凋亡率明显上升(P均<0.05)。铁剂处理增强了BM-MSCs中p38表达,暗示丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路激活,同时抑制p-p38、p53和P细胞外信号调节激酶的表达。结论铁通过MAPK信号通路调节BMSCs的双重作用(铁代谢和骨代谢),FAC的浓度处于0~<200μmol/L范围内时,对细胞的增殖并没有产生显著的影响,当FAC的浓度≥200μmol/L时,其对细胞的影响发生了显著变化,明显促进了细胞的凋亡过程,高浓度FAC对BMCSs细胞的潜在毒性。未来的研究应探索针对铁稳态的治疗策略,以缓解骨质疏松症。
Objective To investigate the effect of iron on osteogenic metabolism of bone marrow derived mesenchymal stem cells(BMSCs)in postmenopausal rat models.Methods A total of fifty rats purchased from Beijing Animal Experimental Center of Gannan Medical University were selected as the research objects.According to the random number table method,they were divided into model group(forty rats)and blank group(ten rats).The model group underwent bilateral ovariectomy and then received ferric acid(FAC)intervention,which was divided into model group,low,medium and high doses group.ten rats in each group.The indexes of bone metabolism,osteogenic differentiation and iron metabolism of rats in each group were detected by enzyme-linked immunosorbent assay and real-time fluorescence quantitative reverse transcription polymerase chain reaction.Subsequently,primary BMSCs cells were cultured from rat femurs and intervened with iron.CCK-8 and immunoblotting were used to detect the apoptosis and mechanism.Results The levels of alkaline phosphatase(ALP)in the blank group,model group,low-dose group,medium-dose group and high-dose group were(28.45±4.15)U/L,(22.49±3.45)U/L,(17.46±2.38)U/L,(13.44±1.79)U/L and(11.56±1.33)U/L,the ALP value of the model group decreased with the increase of iron dosage,and the high-dose group was the lowest,the difference was statistically significant(F=68.300,P<0.05).The osteocalcin value in the middle of the amino end of the model group decreased with the increase of the amount of iron,and the high-dose group was the lowest,the difference was statistically significant(P<0.05).The expression of osteoblast-related transcription factor 2,osteopontin and osteocalcin in the model group decreased significantly,and the expression level decreased gradually with the increase of iron dosage,and the high-dose group was the lowest,with statistically significant differences(all P<0.05).The total iron binding capacity,transferrin saturation and serum ferritin level in the model group increased with the increase of iron dosage,and the differences were statistically significant(all P<0.05).FAC concentration≥200μmol/L promoted cell apoptosis,indicating the potential toxicity of high concentration FAC to BMCSs cells.After 48 h of treatment,the apoptosis rate of 200μmol/L group increased significantly(all P<0.05).Iron treatment enhanced the expression of p38 in BM-MSCs,suggesting the activation of mitogen-activated protein kinase(MAPK)signaling pathway,and inhibited the expression of p-p38,p53 and P-extracellular signal-regulated kinase.Conclusion Iron regulates the dual effects of BMSCs(iron metabolism and bone metabolism)through the MAPK signaling pathway.When the concentration of FAC is in the range of 0~<200μmol/L,it has no significant effect on cell proliferation.When the concentration of FAC≥200μmol/L,its effect on cells has changed significantly,which significantly promotes the apoptosis process of cells,and the potential toxicity of high concentration FAC on BMCSs cells.Future research should explore therapeutic strategies for iron homeostasis to alleviate osteoporosis.
作者
杜永伟
徐房添
赖光松
唐文
袁阁欢
姚飞
刘真
DU Yongwei;XU Fangtian;LAI Guangsong;TANG Wen;YUAN Gehuan;YAO Fei;LIU Zhen(Department of Orthopedics,The First Affiliated Hospital of Gannan Medical University,Ganzhou 341000,Jiangxi,China;Department of Rehabilitation Medicine,The First Affiliated Hospital of Gannan Medical University,Ganzhou 341000,Jiangxi,China)
出处
《系统医学》
2025年第13期38-42,46,共6页
Systems Medicine
基金
2021年赣州市指导性科技计划项目(GZ2021ZSF001)
2020年江西省重点研发计划项目(S2020ZPYFB1863)
2019年江西省教育厅科技计划项目(GJJ190809)。
关键词
铁蓄积
骨髓间充质干细胞
成骨代谢
丝裂原活化蛋白激酶通路
铁代谢
绝经后骨质疏松
Iron accumulation
Bone marrow derived mesenchymal stem cell
Osteogenic metabolism
Mitogen activated protein kinase pathway
Iron metabolism
Postmenopausal osteoporosis
