罗汉果皂苷V刺激成骨细胞增殖与分化

Mogroside V promotes bone formation by stimulating osteoblast proliferation and differentiation

背景:骨在一生中不断地进行重塑,骨稳态是成骨细胞的骨形成和破骨细胞的骨吸收之间的动态平衡。在细胞水平严格控制骨重建,对于维持骨稳态以响应结构和代谢需求是至关重要的。目的:探讨罗汉果皂苷V对成骨细胞增殖、分化的影响。方法:采用两步酶消化法提取大鼠成骨细胞,通过MTT实验检测罗汉果皂苷V的细胞毒性;不同质量浓度(0,6.25×10^-3,1.25×10^-2,2.5×10^-2g/L)罗汉果皂苷V对成骨细胞干预2,4,6d后通过FDA/PI荧光染色观察成骨细胞活性,干预6d后通过碱性磷酸酶染色、茜素红染色及qRT-PCR检测观察罗汉果皂苷V对成骨细胞分化的影响。结果与结论:①MTT实验表明罗汉果皂苷V在6.25×10^-3-2.5×10^-2g/L时显著促进成骨细胞增殖;②FDA/PI荧光染色显示6.25×10^-3,1.25×10^-2,2.5×10^-2g/L罗汉果皂苷V可提高成骨细胞活力、促进细胞增殖,尤其以1.25×10^-2g/L效果显著;③罗汉果皂苷V干预6d后,碱性磷酸酶染色(定量)及茜素红染色实验均显示罗汉果皂苷V促进成骨细胞成骨矿化,特别是在1.25×10^-2g/L时最为明显;④qRT-PCR结果显示罗汉果皂苷V组的成骨分化相关基因RUNX2、BSP、OCN和COL1A1表达高于对照组,特别是罗汉果皂苷V在1.25×10^-2g/L时表达最高;⑤结果表明,罗汉果皂苷V可刺激成骨细胞增殖、分化,在1.25×10^-2g/L时作用最为明显。

BACKGROUND:Bone remodeling is going on throughout the lifetime.Bone homeostasis is a dynamic balance between bone formation of osteoblasts and bone resorption of osteoclasts.Controlling of bone remodeling at the cellular level is critical to maintaining bone homeostasis in response to structural and metabolic demands.OBJECTIVE:To explore the effect of mogroside V on the proliferation and differentiation of osteoblasts.METHODS:Rat osteoblasts were extracted by two-step enzymatic digestion,followed by MTT assay to detect the toxicity of mogroside V.The proliferation of osteoblasts was observed by FDA/PI fluorescence staining at 2,4 and 6 days after the treatment with mogroside V(0,6.25×10^-3,1.25×10^-2,2.5×10^-2 g/L).After 6 days of treatment on osteoblasts,the effect of mogroside V on osteoblast differentiation was studied by alkaline phosphatase staining,alizarin red staining and qRT-PCR.RESULTS AND CONCLUSION:MTT assay showed that the mogroside V at 6.25×10^-3 to 2.5×10^-2 g/L promoted osteoblast proliferation.FDA/PI fluorescence staining showed that mogroside V(6.25×10^-3,1.25×10^-2,2.5×10^-2 g/L)could improve osteoblast activity and promote cell proliferation,especially when the concentration of mogroside V was 1.25×10^-2 g/L.After 6 days of treatment,alkaline phosphatase staining and alizarin red staining showed that the mogroside V,especially at 1.25×10^-2 g/L,promoted osteogenesis and mineralization.qRT-PCR results showed that the expressions of runt related transcription factor 2,bone sialoprotein,osteocalcin and alpha-1 type I collagen genes were higher in the mogroside V group than the blank group,especially when the concentration of mogroside V was 1.25×10^-2 g/L.Therefore,mogroside V,especially at 1.25×10^-2 g/L,promotes bone formation by stimulating osteoblast proliferation and differentiation.