芒果苷对同型半胱氨酸诱导骨髓间充质干细胞分化的影响

Effect of mangiferin on differentiation of bone marrow mesenchymal stem cells induced by homocysteine

目的旨在利用同型半胱氨酸(Hcy)构建体外培养原代大鼠骨髓间充质干细胞(rBMSCs)脂向分化模型,剖析Hcy对BMSCs脂向分化与骨向分化的具体作用,对Hcy诱导的BMSCs施加不同浓度的芒果苷进行干预,探究芒果苷对BMSCs脂向分化与骨向分化的影响。方法首先提取分离rBMSCs,将培养至一定代数状态良好的rBMSCs置于含不同浓度Hcy(0.125、0.250、0.5、1、2、4 mmol/L)的培养基中培养,建立rBMSCs脂向分化模型。随后,对实验组细胞施加不同浓度的芒果苷(37.5、75、150μmol/L)进行干预。培养一定时间后,收集细胞进行以下检测:运用油红O染液半定量方法检测细胞内脂质累积情况,以评估脂向分化程度;采用ρNPP法精准测定碱性磷酸酶(ALP)活力;Western blot法检测骨形成蛋白-2(BMP-2)、Ⅰ型胶原(ColⅠ)的表达变化,以评估骨向分化程度。结果芒果苷在体外能显著上调BMP-2、ColⅠ的表达,升高ALP水平,促进rBMSCs的骨向分化。Hcy则通过增加脂质累积,促进rBMSCs的脂向分化;与此同时,Hcy还下调BMP-2与ColⅠ的表达,降低胞内ALP水平,从而抑制rBMSCs的骨向分化。然而,上述与Hcy相关的效应能够被芒果苷成功扭转。结论芒果苷在体外能够显著促进rBMSCs的骨向分化,而Hcy可抑制rBMSCs骨向分化并促进其脂向分化,芒果苷具有扭转该效应的能力,说明芒果苷在治疗骨质疏松症相关疾病方面具有一定潜能。

Objective To establish a lipid differentiation model of primary rat bone marrow mesenchymal stem cells(rBMSCs)in vitro using homocysteine(Hcy),and analyze the specific effects of Hcy on lipid and bone differentiation of BMSCs;to comprehensively explore the effects of mangiferin on lipid and bone differentiation of BMSCs,and further reveal the potential mechanism of mangiferin in the treatment of osteoporosis through the intervention of Hcy-induced BMSCs by different concentrations of mangiferin.Methods First,rBMSCs were extracted and isolated.The rBMSCs that were well-cultured to a certain generation were placed in culture medium containing different concentrations of Hcy(0.125,0.250,0.5,1,2,4 mmol/L)to establish lipid differentiation model of rBMSCs.Then,different concentrations of mangiferin(37.5,75,150μmol/L)were applied to the experimental cells for intervention.After culture for a certain period of time,the cells were collected for the following tests:The accumulation of lipids in the cells was detected by semi-quantitative method of oil red O dye solution to evaluate the degree of lipid differentiation;The activity of alkaline phosphatase(ALP)was measured byρNPP method;The expression of bone morphogenetic protein-2(BMP-2)and type I collagen(ColⅠ)were detected by western blot assay to evaluate the degree of bone differentiation.Results Mangiferin could significantly up-regulate the expression of BMP-2 and ColⅠin vitro,increase the level of ALP,and promote bone differentiation of rBMSCs.Hcy promoted lipid differentiation of rBMSCs by increasing lipid accumulation,and down-regulated the expression of BMP-2 and ColⅠ,reduced the intracellular ALP level,thereby inhibiting bone differentiation of rBMSCs.However,the above Hcy-related effects could be successfully reversed by mangiferin.Conclusion Mangiferin can significantly promote bone differentiation of rBMSCs in vitro,while Hcy can inhibit bone differentiation of rBMSCs and promote its lipid differentiation.Mangiferin has the ability to reverse this effect,indicating that mangiferin has certain potential in the treatment of osteoporosis-related diseases.