不同氧浓度对骨髓基质细胞向成骨细胞分化的影响

Effects of different oxygen concentrations on differentiation of marrow stroma cells into osteoblasts

目的:观察不同低氧环境对在向成骨细胞分化培养系中的骨髓基质细胞核心结合因子α1(Cbfα1/Runx2)、骨形态发生蛋白2(BMP2)和过氧化物酶体增殖物激活受体γ2(PPAR-γ2)表达的影响,探讨低氧环境对成骨细胞分化的影响。方法:取4月龄雌性SD大鼠骨髓基质细胞在生长培养基中培养传代后,随机分成4组,每组样本数为8,加入分化培养基,分别将细胞放入4个不同氧浓度中继续培养,3d后进行下面的实验:用Trizol提取细胞总RNA,用半定量逆转录PCR(RT-PCR)检测(Cbfα1/Runx2)、BMP2和PPARγ2mRNA的表达;用Western blotting检测(Cbfα1/Runx2)、BMP2蛋白的表达。结果:1.与常氧组(20%)相比,各低氧组Runx2mRNA和Runx2蛋白的表达明显增加。2.与常氧组(20%)相比,各低氧组BMP2mRNA的表达明显增加,低氧可促进BMP2蛋白的表达。3.与常氧组(20%)相比,各低氧组PPARγ2mRNA的表达明显降低。结论:低氧能明显促进Runx2mRNA、BMP2mRNA和Runx2蛋白的表达,且氧浓度越低,Runx2mRNA、BMP2mRNA和Runx2、BMP2蛋白表达越多,相反,低氧能明显抑制骨髓基质细胞PPARγ2mRNA的表达,且氧浓度越低,PPARγ2mRNA的表达越低,表明低氧明显抑制骨髓基质细胞向脂肪细胞分化而促进其向成骨细胞分化。

AIM：To investigate the effect of different oxygen concentrations on the differentiation of marrow stroma cells into osteoblasts and to evaluate the expression of Cbfα1 /Runx2,bone-morphogenesis protein 2 （BMP2） and peroxisome proliferator-activated receptor γ2 （PPAR-γ2） in bone marrow stromal cells. METHODS：The bone marrow stomal cells obtained from 4-month-old female SD rats were cultured in growth medium and were used between passages 3 to 5. The cells were divided randomly into 4 groups,each group has 8 samples. The cells in all 4 groups were used for the following experiments after cultured with different oxygen concentrations for 3 d in osteoblastic differentiation medium：total cellular RNA was isolated using total RNA kit; RT -PCR was performed to detect the mRNA expression of Cbfα1 /Runx2,BMP2 and PPARγ2. The protein expression of Cbfα1 /Runx2 and BMP2 was assayed by Western blotting. RESULTS：Compared to the cells in normoxia condition （20% ）,the mRNA and protein expressions of Runx2 were enhanced significantly,the mRNA expression of BMP2 was also enhanced significantly,the protein expression of BMP2 increased and the mRNA expression of PPARγ2 decreased significantly in the cells cultured with lower oxygen concentrations. The lower oxygen con-centration was in the culture,the more Runx2 mRNA,BMP2 mRNA,BMP2 and Runx2 protein were expressed. On the contrary,hypoxia significantly decreased the expression of PPARγ2 mRNA in bone marrow stronmal cells and the lower the oxy-gen concentration was used,the less expression of PPARγ2 mRNA was achieved. CONCLUSION：Hypoxia promotes the mRNA and protein expressions of Runx2 and BMP2,also significantly decreases the expression of PPARγ2 mRNA in bone marrow stronmal cells in an oxygen concentration dependent manner,indicating that hypoxia significantly stimulates the differentiation of bone marrow stromal cells into osteoblasts instead of lipocytes.