葡萄糖和谷氨酰胺浓度对人脐带间充质干细胞体外培养的影响

Effects of Various Concentrations of Glucose and Glutamine on the Cultivation of Human Umbilical Cord-derived Mesenchymal Stem Cells in Vitro

目的探索葡萄糖和谷氨酰胺浓度对人脐带间充质干细胞(hUC-MSCs)体外培养的影响。方法分离、培养hUC-MSCs,光学显微镜观察细胞形态,流式细胞仪检测细胞表面抗原。采用不同浓度葡萄糖和谷氨酰胺对hUC-MSCs进行体外培养,结晶紫计数法记录细胞增殖曲线,并检测培养液中营养物质及其代谢产物。结果传代的hUC-MSCs均一地高表达CD29、CD44、CD105和CD90,不表达CD14、CD45。葡萄糖浓度增加,会促进细胞的增殖,对葡萄糖消耗、乳酸生成和谷氨酰胺消耗没有影响,但增加了氨的生成;谷氨酰胺浓度增加,对细胞增殖的促进作用有限,但可提高细胞的生长速率,从而缩短培养时间,对葡萄糖和谷氨酰胺的消耗影响不大,但增加了乳酸和氨的生成。结论葡萄糖和谷氨酰胺都可促进细胞的增殖。葡萄糖缺失时其他氨基酸充当氮源,谷氨酰胺充足时谷氨酰胺转化成乳酸的通量增大。

Objective To explore the effects of various concentrations of glucose and glutamine on the cultivation of human umbilical cord-derived mesenchymal stem cells （hUC-MSCs） in vitro. Methods The hUC-MSCs were separated and cultured. The morphology and surface antigens were investigated by optical microscopy and flow cytometry. Then hUC-MSCs were cultured with various concentrations of glucose and glutamine in vitro. Cells on microcarriers were counted by crystal violet staining. The nutrients and their by-products in the medium supematant were measured. Results The markers of CD29, CD44, CD105, and CD90 were strongly expressed in hUC-MSCs, but the markers of CD14 and CIM5 were not. With the increase of glucose concentration, the expansion of cells increased, glucose consumption, lactate formation and glutamine consumption had no change, and the ammonium formation improved. The increase of glutamine concentration had a limited effects to improving cell proliferation, but it shorted the culture period as improving growth rate. The increase of glutamine concentration had no effect on the consumption of glucose and glutamine, but improved the formation of lactate and ammonium. Conclusion Glucose and glutamine both enhanced cell proliferation. Lack of glucose results in the larger consumption of amino acids to provide nitrogen, and large supply of glutamine leads to the larger formation of lactate from glutamine.