脂肪基质细胞脂向分化过程中MicroRNA-134的表达变化

The differential expression of miRNA-134 during adipogenic differentiation of ADSCs

目的了解脂肪基质细胞(ADSCs)脂向分化过程中MicroRNA-134(miR-134)表达变化情况,深入了解ADSCs脂向分化的分子调控机制,为脂肪组织工程的研究和发展奠定基础。方法(1)采取出生30d的雄性体健SD(Sprague-Dawley)幼鼠的腹股沟脂肪组织,用密度梯度离心结合贴壁培养法分离、纯化后诱导培养大鼠脂肪基质细胞(ADSCs)7d,设置未诱导组以对照。(2)应用miRNA芯片技术比较诱导组和未诱导组的miR-134表达差异。(3)用实时定量PCR定量验证ADSCs脂向分化前后miR-134表达差异。结果(1)成功分离、纯化、培养ADSCs,诱导组培养7d后观察到典型脂肪细胞特征,细胞形态学上表现为细胞相互融合,成片生长,呈细长纺锤形,大量脂滴形成;PPARγ免疫细胞化学染色呈阳性;油红O染色脂滴被染成橙红色。(2)成脂诱导前后miR-134表达明显下调,其中诱导修正值2054.5,无诱导修正值2855.25。(3)实时定量PCR结果亦显示miR-134表达显著下调,与miRNA芯片结果一致。结论miR-134在ADSCs脂向分化过程中miR-134表达显著下调,可能参与调控ADSCs的脂向分化。

Objective For better understanding of the molecular mechanisms of adipogenic differentiation of ADSCs, we explored the differential expression of miR - 134 during adipogenic differentiation of ADSCs. Methods The experiment was performed at the State Key Laboratory of Oral Diseases. （ 1 ） Adipose was obtained from eight healthy male young SD rat. ADSCs were isolated and purified with density gradient centrifugation and adherent culture, The third generation cells were induced into adipogenic differentiation in the inductive medium for 7 days, and were compared with the uninduced cells. （2） We used the miRNA chip technology to explore the differential expression of miR - 134 in experimental group （7 days after adipogenic -differentiated） and control group （not induced）. （3）We validated the differential expression of miR - 134 during adipogenic differentiation of ADSCs with reahime quantitative PCR. Results （ 1 ） We got high -purity ADSCs and successfully induced them into osteoplastic differentiation. Character of typical adipocytes was displayed after 7 days induction： the ceils turned to slender fusiform and mixed together, with plenty fat drops; positive PPARγ immunohistochemical staining; fat drops were stained positively with Oil red O. （2）The expression of miR- 134 showed significant down regulation during adipogenic differentiation of ADSCs. The induce allowance 2 054. 5 and non - induce allowance 2 855.25 both exceeded 50. （3） Realtime quantitative PCR results met the results of miRNA chip tech- nology exactly. Conclusion Significant down - regulated expression of miR - 134 during adipogenic differentiation of ADSCs suggests an important role in adipogenic differentiation of ADSCs.