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目的:探讨在人骨髓间充质干细胞(h BMSCs)成骨分化过程中,不同浓度尿酸(UA)对骨形态形成蛋白-2(BMP-2)表达的影响。方法:以全骨髓贴壁培养法分离h BMSCs,将生长状态良好的第3代h BMSCs分为5组,分别为空白对照组(加入完全培养基)和成骨诱导组(加入成骨诱导液及含0 mmol/L、0.2 mmol/L、0.4 mmol/L、0.8 mmol/L尿酸的完全培养基)。连续干预诱导14d后,用倒置显微镜观察细胞形态的变化,通过观察茜素红染色情况及检测碱性磷酸酶(ALP)活性进行成骨情况的检测。RT-PCR技术检测各组细胞BMP-2 mR NA的表达情况。结果:第3代h BMSCs大多为形态单一的长梭形,呈旋涡状生长;干预诱导后的细胞逐渐变成不规则的立方形,局部形成团块状结节,以含尿酸浓度为0.8 mmol/L的成骨诱导培养基最为显著。连续干预14d后,空白对照组茜素红染色为阴性,而各成骨诱导组细胞茜素红染色结果为阳性,提示干预诱导后的细胞为成骨细胞。碱性磷酸酶活性随尿酸浓度的增加和干预时间的延长而增强(P<0.05)。RT-PCR检测结果显示,空白对照组无BMP-2 mR NA的表达。成骨诱导组随培养基中尿酸浓度的增加,BMP-2 mR NA表达逐渐增强,呈浓度依赖性(P<0.05)。结论:尿酸上调h BMSCs向成骨细胞分化过程中BMP-2 mR NA的表达。
OBJECTIVE: To investigate the effects of different concentrations of uric acid (UA) on the expression of bone morphogenetic protein-2 (BMP-2) during the osteogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs). Methods: The hBMSCs were isolated by whole bone marrow adherent culture. The third generation of hBMSCs with good growth status were divided into 5 groups, which were blank control group (complete medium addition) and osteogenic induction group (osteogenic induction group And complete medium containing 0 mmol / L, 0.2 mmol / L, 0.4 mmol / L, 0.8 mmol / L uric acid). After 14 days of continuous intervention, the changes of cell morphology were observed by inverted microscope. The osteogenesis was detected by observing the alizarin red staining and detecting alkaline phosphatase (ALP) activity. The expression of BMP-2 mRNA in each group was detected by RT-PCR. Results: The third generation of hBMSCs were mostly long fusiform and swirling in shape. After induction, hBMSCs gradually became irregular cubes and formed clumps nodules. The concentration of uric acid was 0.8 mmol / L osteogenic induction medium is the most significant. After 14 days of continuous intervention, alizarin red staining of the blank control group was negative, while the alizarin red staining of the osteogenic induction group was positive, suggesting that the cells after the intervention were osteoblasts. Alkaline phosphatase activity increased with the increase of uric acid concentration and prolonged intervention (P <0.05). RT-PCR results showed that there was no BMP-2 mR NA expression in the blank control group. With osteogenic induction group, the expression of BMP-2 mR NA gradually increased in a concentration-dependent manner with the increase of uric acid concentration in culture medium (P <0.05). CONCLUSION: Uric acid up-regulates the expression of BMP-2 mRNA in hBMSCs during osteoblast differentiation.