葡萄糖对小鼠胚胎干细胞生长的影响  

Effect of glucose on the growth of mouse embryonic stem cells

作　　者：李华[1,2] Amy Lam 吴浩佳[1] 郝敏[2] Sookja Kim Chung 

机构地区：[1]香港大学李嘉诚医学院解剖系 [2]大连医科大学药学院药理教研室

出　　处：《中国临床药理学与治疗学》2009年第7期760-765,共6页Chinese Journal of Clinical Pharmacology and Therapeutics

摘　　要：目的:目前小鼠胚胎干细胞(mouse embry-onic stem cells,mESC)常规应用高浓度葡萄糖(25 mmol/L)培养基培养,但是在应用干细胞向糖尿病胰岛beta细胞分化的研究中发现,慢性高糖培养可促进干细胞的凋亡、降低细胞分化的效率及分化后胰岛beta细胞对葡萄糖的反应性,因此本研究拟选择合适的较低浓度的葡萄糖以优化胚胎干细胞的培养基、提高胚胎干细胞的生长、分化效率。方法:mESC传代4或12 h后,将传统的25 mmol/L葡萄糖培养基分别换为5、10、15、25 mmol/L葡萄糖培养基培养,均用碱性磷酸酶(AP)染色计数细胞集落形成情况、台盼兰染色测定细胞数目,MTT法测定细胞活力及用4’,6-联脒-2-苯基吲哚(4’,6-diamidino-2-phenylindole,DAPI)染色检测细胞凋亡情况。结果:mESC传代4 h后即调换葡萄糖浓度:(1)各葡萄糖浓度组(5、10、15 mmol/L)与25 mmol/L组相比,集落形成明显减少。(2)各葡萄糖浓度组与25 mmol/L组相比,mESC增殖及细胞活力均受到不同程度明显影响。mESC传代12 h后调换葡萄糖浓度:(1)各葡萄糖浓度组(5、10、15 mmol/L)与25 mmol/L组相比,集落形成无明显变化,且AP染色呈强阳性。(2)15 mmol/L葡萄糖组对mESC增殖及细胞活力均无明显影响。(3)15 mmol/L葡萄糖组与25 mmol/L葡萄糖组细胞核形态正常,均未见明显凋亡。结论:ESC传代12 h后将培养基中传统的高糖(25 mmol/L)降低为15 mmol/L,不影响mESC细胞活力及多能分化潜能和未分化状态。AIM： Mouse embryonic stem cells （mESC） are routinely cultured in the medium with high concentration of glucose （25 mmol/L）. However, it was found that chronic high glucose culture promoted the apoptosis of stem cells, inhibited the differentiation efficiency and decreased the response of differentiated beta cells on glucose stimulation. The present study was designed to find suitable lower concentration of glucose to optimize the ESC culture medium so as to improve the ESC growth and differentiation. METHODS： The routinely used high glucose （25 mmol/L） medium was changed to 5, 10, 15,25 mmol/L respectively at 4 or 12 hours after mESC passage. The colony formation by alkaline phosphatase （AP） staining, cell counting by Trypan blue, cell viability by 3- [4,5-Dim- ethyl-2-thiazol ]-2, 5-diphenyl-2H-tetrazolium bromide （MTT） method were performed. RESULTS： mESCs were changed culturing with different glucose concentrations at 12 hours after passage： （1） There was no significant difference on colony numbers between 5, 10, 15 mmol/L groups and 25 mmol/L group, and AP staining was strongly positive in all groups. （2） There was no significant difference of cell numbers and cell viability at 15 mmol/L group compared with 25 mmol/L group. CONCLUSION： 15 mmol/L glucose, which was used at 12 hours after mESC passage, did not affect the cell viability, pluripotency and undifferentiated state.

关 键 词：胚胎干细胞 葡萄糖 浓度 

分 类 号：R965.2[医药卫生—药理学]