Objective To detect the uric acid effect on the expression and function of 11 β-hydroxy steroid dehy-drogenase type Ⅰ(11β-HSD1) and the osteogenic ability of hBMSCs in the course of human bone marrow mesenchymal stem cells (hBMSCs) into osteoblasts in vitro. Methods Healthy adult hBMSCs were isolated and cultured using the whole bone marrow cultured method in vitro. The nature of the cells was identified by the cell morphology and cell surface markers. After culturing hBMSCs to the passage 3 , we respectively used complete medium ( containing the volume fraction of 10% FBS, 1% double-antibiotics, 89% low glucose DMEM) for blank control group, osteogenic induction medium (containing 10 mol/L dexamethasone, 50 mg/L vitamin C, 10 mol/L p-glycerol phosphatesodium in complete medi- um) , uric acid intervention osteogenic induction mediums ( respectively containing 0. 2, 0. 4, 0. 8 mmol/L uric acid in osteogenic induction medium) for condition control groups to culture and induce hBMSCs. After 14 days, cell morphology was observed through an inverted microscope, alkaline phosphatase ( ALP) staining and alizarin red staining were performed to identify osteoblasts, the expression of 11 β-HSD1 mRNA was detected by immunocytochemistry staining and RT-PCR technology. Results The isolated and cultured cells was positive and CD34 was negative for the surface expression of CD44. High pure hBMSCs were obtained by the methods. The ALP staining and Alizarin red staining of all induced cells were positive, and with the increased of uric acid concentration, the calcium nodules of induced cells increased gradually (P<0. 05). Immunocytochemical stain indicated brown positive staining granules in the cytoplasm in all cells, pathology image analysis indicated cells of complete medium had the highest optical density value, and with the increased of uric acid concentration and osteogenic ability, the optical density values reduced gradually. ( P < 0. 05 ) . RT-PCR indicated cells of the complete medium had the highest expression of 11 β-HSDl mRNA, and with the increased of uric acid concentration and osteogenic ability, the expressions of llfl-HSDl mRNA reduced gradually ( P < 0. 05). Conclusion Uric acid can promote hBMSCs to proliferate and differentiate into osteoblasts in vitro, with a characteristic of concentration-dependent and time-dependent. Uric acid can promote hBMSCs to proliferate and differentiate into osteoblasts in vitro, by down-regulating the expression of 11 β-HSDl.%目的 探讨体外诱导人骨髓间充质干细胞(hBMSCs)向成骨细胞分化过程中,尿酸对其成骨能力以及对11β-羟化类固醇脱氢酶1(11β-HSD1)表达及功能的影响.方法 全骨髓体外分离培养健康成年人骨髓间充质干细胞,用细胞形态学及细胞表面标志物对其进行鉴定,培养hBMSCs至第3代后分别以完全培养基(含体积分数10％ FBS、1％双抗、89％低糖DMEM)为空白对照组、以成骨培养基(含10-8 mol/L地塞米松、50 mg/L维生素C、10-2 mol/L β-甘油磷酸钠的完全培养基)和尿酸干预成骨培养基(分别含0.2、0.4、0.8 mmol/L尿酸的成骨培养基)为条件对照组进行培养和诱导.培养诱导14 d后,倒置显微镜下观察细胞形态,用碱性磷酸酶染色和茜素红染色进行成骨细胞鉴定,用11β-HSD1免疫细胞化学染色、RT-PCR技术检测各组11β-HSD1 mRNA的表达.结果 分离培养的细胞表面标志物CD44表达阳性,CD34表达阴性.成骨培养基和各尿酸干预成骨培养基诱导的细胞碱性磷酸酶染色和茜素红染色均为阳性,钙结节数量随尿酸浓度增高逐渐增多(P＜0.05).免疫细胞化学染色结果显示各组细胞胞质均可见棕黄色阳性染色颗粒.病理图象分析软件测定11β-HSD1含量结果显示随尿酸浓度增高,成骨能力增加,光密度值逐渐减少(P＜0.05).RT-PCR结果显示各组细胞均有11β-HSD1 mRNA表达,随尿酸浓度增高和成骨能力的增加,11β-HSD1 mRNA表达逐渐减少(P＜0.05).结论 尿酸能促进hBMSCs向成骨细胞增生和分化,具有浓度依赖性和时间依赖性,尿酸通过下调11β-HSD1 mRNA的表达促进hBMSCs向成骨细胞分化.
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