色素上皮衍生因子对高浓度地塞米松刺激下成骨细胞迁移、分化的影响

Effects of pigment epithelium- derived factor on migration and differentiation of osteoblasts after stimulation by high concentrations of dexamethasone

[目的]研究色素上皮衍生因子(pigment epithelium-derived factor,PEDF)在高浓度地塞米松刺激下,体外培养的成骨细胞迁移及分化的影响,探讨PEDF对成骨细胞功能的保护作用。[方法]将体外培养的MC3T3-E1细胞分为六组,分别为对照组、高浓度地塞米松(dexamethasone,DEX 10-6mol/L)组,PEDF干预地塞米松组,PEDF干预地塞米松组又按照PEDF的浓度分为3组(2、10、50 nmol/L)及成骨诱导培养组。采用Transwell小室及划痕实验于48 h后观察细胞的迁移情况。于14 d时采用倒置相差显微镜观察细胞形态的变化,并进行细胞化学染色(茜素红染色、油红O染色),运用Western Blot检测细胞内PPARγ-2、Runx2蛋白的表达。[结果]在高浓度地塞米松作用下,细胞迁移能力受到抑制,细胞分化能力减弱,胞浆中出现脂滴,Western Blot结果显示脂肪细胞标志蛋白PPARγ-2表达增高,成骨细胞标志蛋白Runx2表达减少。而PEDF干预后可明显逆转高浓度地塞米松抑制细胞迁移及分化的作用。[结论]大剂量应用糖皮质激素可能抑制成骨细胞的迁移及成骨分化能力,促进成骨细胞转分化为脂肪细胞,而PEDF可以有效地逆转这一过程,起到保护细胞功能的作用。

[ Objective]To explore the effects of pigment epithelium - derived factor （PEDF） on the migration and differen- tiation of osteoblasts cultured in vitro after stimulation by high concentrations of dexamethasone （ DEX）, and to explore the pro- tective effects of PEDF on osteoblastic function. [ Method] MC3T3 -E1 cells cultured in vitro were randomly divided into con- trol group, DEX （10-6 mol/L） group, DEX ＋ 2 nmol/L PEDF group, DEX ＋ 10 nmol/L PEDF group, DEX ＋50 nmol/L PEDF group, and induced osteogenesis group. The transwell chamber assay and scratch test were used to evaluate the migration of cells after 48 hours. At 14 days after treatment, inverted phase - contrast microscopy and cell staining （ alizarin red staining and oil red O staining） were used to evaluate the morphological changes and differentiation of cells, respectively. Western blot was used to determine the intraeellular expression of PPAR~/- 2 and Runx2. [ Result] After stimulation by high concentrations of DEX, cell migration and differentiation were inhibited, lipid droplets were observed in cytoplasm. Western blot results revealed elevated expression of the adipocyte marker PPART - 2 and reduced expression of the osteoblast marker Runx2. Inter- vention with PEDF substantially reversed the inhibitory effects of high concentrations of DEX on cell migration and differentiation. [ Conclusion ] High - dose glucocorticoids may inhibit the migration and osteogenic differentiation of osteoblasts and promote transdifferentiation of osteoblasts into adipocytes. PEDF can effectively reverse the process and protect osteo- blastic function.