促红细胞生成素及受体信号通路调控牙周膜干细胞成骨分化的机制  

Mechanism of erythropoietin/erythropoietin receptor signaling pathway in regulating osteogenic differentiation of periodontal ligament stem cells

作　　者：孙峥[1] 赵华[1] Sun Zheng;Zhao Hua(Department of Stomatology,Second Hospital of Shanxi Medical University,Taiyuan 030000,Shanxi Province,China)

机构地区：[1]山西医科大学第二医院口腔科,山西省太原市030000

出　　处：《中国组织工程研究》2025年第36期7762-7768,共7页Chinese Journal of Tissue Engineering Research

基　　金：山西省重点研发计划项目(201903D321141),项目负责人:孙峥;山西省卫生健康委科研课题项目,项目负责人:孙峥。

摘　　要：背景:促红细胞生成素/促红细胞生成素受体信号通路除了参与骨髓造血,也参与调节非造血组织,如脑、心脏、骨骼肌、脂肪组织等的代谢反应。同时,激活此信号通路能加速牙周膜干细胞的生物矿化过程,减轻细胞氧化应激损伤,但促进牙周膜干细胞成骨分化的机制尚不清楚。目的:探究促红细胞生成素/促红细胞生成素受体信号通路对牙周膜干细胞成骨分化的影响及作用机制。方法:采用酶消化法分离、培养牙周疾病患者及健康人群的牙周膜干细胞,采用qRT-PCR和免疫印迹法检测两种牙周膜干细胞中促红细胞生成素受体的mRNA和蛋白表达水平。使用小干扰RNA沉默促红细胞生成素受体,或使用促红细胞生成素诱导促红细胞生成素受体表达激活,然后采用qRT-PCR、免疫印迹法检测促红细胞生成素受体、成骨标志基因Runt相关转录因子2、骨钙素、骨桥素、骨唾液蛋白的表达水平,采用碱性磷酸酶染色、茜素红染色检测牙周膜干细胞成骨分化能力,采用免疫印迹法检测信号转导及转录激活因子5的磷酸化水平。结果与结论:(1)qRT-PCR和免疫印迹法结果显示,疾病组牙周膜干细胞中促红细胞生成素受体的mRNA和蛋白水平显著低于健康组牙周膜干细胞;(2)碱性磷酸酶染色以及茜素红染色显示,敲低促红细胞生成素受体能抑制牙周膜干细胞的成骨分化能力;qRT-PCR结果显示,与对照组相比,敲低促红细胞生成素受体组Runt相关转录因子2、骨钙素、骨桥素和骨唾液蛋白的表达水平显著降低(P <0.05);(3)qRT-PCR结果显示,促红细胞生成素处理后,牙周膜干细胞中促红细胞生成素受体表达恢复,沉默促红细胞生成素受体后再给予促红细胞生成素处理,逆转了促红细胞生成素受体的表达水平;促红细胞生成素处理后提高了疾病组牙周膜干细胞的成骨分化能力以及成骨标志基因Runt相关转录因子2的表�BACKGROUND:Erythropoietin/erythropoietin receptor signaling pathway not only participates in bone marrow hematopoiesis,but also regulates the metabolic response of non-hematopoietic tissues,such as brain,heart,skeletal muscle,and adipose tissue.Simultaneously,it can accelerate the mineralization process of periodontal ligament stem cells and reduce oxidative stress damage.However,the mechanism of action on osteogenic differentiation of periodontal ligament stem cells is still unclear.OBJECTIVE:To investigate the effect and action mechanism of erythropoietin/erythropoietin receptor signaling pathway on osteogenic differentiation of periodontal ligament stem cells.METHODS:Enzyme digestion method was used to isolate and culture periodontal ligament stem cells from periodontal disease patients and healthy people.The mRNA and protein levels of erythropoietin receptor in two kinds of periodontal ligament stem cells were detected by qRT-PCR and western blot assay.Erythropoietin receptor expression was silenced by small interfering RNA(siRNA)or activated by erythropoietin.qRT-PCR and western blot assay were used to detect the expression of erythropoietin receptor,levels of osteogenic marker genes Runt-related transcription factor 2(Runx2),osteocalcin,osteopontin,and bone sialoprotein.Alkaline phosphatase staining and alizarin red staining were applied to measure osteogenic differentiation ability of periodontal ligament stem cells.The phosphorylation of signal transducer and activator of transcription 5(STAT5)was detected by western blot assay.RESULTS AND CONCLUSION:(1)The results of qRT-PCR and western blot assay showed that the mRNA and protein levels of erythropoietin receptor in periodontal ligament stem cells in the disease group were significantly lower than those in periodontal ligament stem cells in the healthy group.(2)Alkaline phosphatase staining and alizarin red staining showed that knocking down the erythropoietin receptor can inhibit the osteogenic differentiation ability of periodontal ligament stem cells.

关 键 词：牙周膜干细胞 促红细胞生成素 促红细胞生成素受体 信号转导及转录激活因子5 成骨分化 牙周炎 工程化干细胞 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学] R68