Effects Of High Glucose, Insulin And AGEs On Number And Activity Of Vascular Endothelial Cells Differentiated By Bone Marrow Stem Cells Of Mouse

Part I:Effects of glucose and insulin on number and activity of vascular endothelial cells differentiated by bone marrow stem cells of mouseObjective To investigate the effects of glucose and insulin of different levels on number and activity of vascular endothelial cells(VEC) from bone marrow stem cells of mouse. Methods Total mononuclear cells (MNCs) were isolated from bone marrow by Percoll density gradient centrifugation; attached cells were collected and cultured. After 8 days, attached cells were incubated with glucose and insulin in a series of concentrations (glucose: 5.6 mmol/L, 16.7mmol/L, 25mmol/L; insulin: 0 mU/L, 15mU/L, 200mU/L) for 72h. Vascular endothelial cells and endothelial progenitor cells were characterized as adherent cells which were documented by demonstrating the expression of Flk-1, VEGFR-2 with flow cytometry. Cells positive for Flk-1 were further identified under a laser scanning confocal microscope and counted. Results On the same lever of insulin, the amount of cells positive for Flk-1 decreased with the raise of glucose lever, showed significant differences (p<0.01); on the same lever of glucose, compared with 0mU/L insulin, 15mU/L insulin elevate the amount of cells positive for Flk-1 significantly (p<0.01), but the differences in the contents between 200mU/L insulin and 15mU/L insulin were little (p>0.05). Conclusion High glucose might inhibit BMSC's differentiation into EPCs, insulin would promote the differentiation, but high glucose might impair the effects of insulin. Part II: Effects of advanced glycation end-products on number and activity of vascular endothelial cells differentiated by bone marrow stem cells of mouseObjective To investigate the effects of AGE on number and activity of vascular endothelial cells(VEC)from bone marrow stem cells of mouse. Methods Total mononuclear cells (MNCs) were isolated from bone marrow by Percoll density gradient centrifugation; attached cells were collected and cultured. After 8 days, attached cells were incubated with AGE or BSA for 72h, compared with the controlled group. VEC and endothelial progenitor cells were characterized as adherent cells which were documented by demonstrating the expression of Flk-1, VEGFR-2 with flow cytometry. Cells positive for Flk-1 were further identified under a laser scanning confocal microscope and counted. Results Compared with the control group, cells positive for Flk-1 in 100μg /ml AGE group have sharply decreased, showed significant differences (p<0.01), but within 20μg /ml AGE group and BSA group the differences in the contents were little(p>0.05). Conclusion 100μg /ml AGE might inhibit BMSCs'differentiation into EPCs, 20μg/ml AGE and BSA would have little effects on the differentiation.