The Effects Of Advanced Glycation End Products In Human Mesenchymal Stem Cells And Its Mechanisms

Objective To isolate and culcure human bone marrow derived Mesenchymal stem cells (hMSCs), investigate the immuno phenotype of cell surface. To investigate the effects of advanced glycation end products (AGE-BSA) on oxidative stress and apoptosis in hMSC, provide new evidences for a higher survival of donor MSCs in the heart of diabetes cardiomyopathy after clinical stem cell transplantation.Methods The hMSCs were isolated from adult bone marrow, cultured in LG-DMEM with 10% FCS, were suspended by trypsin and passaged for subsequent passages.the morphologial characteristics of hMSCs were observed under inverted phase-contrast microscope. At the third passage, the expressions of immunophenotype were analyse by Flow cytometry (CD34, CD44, CD105). the absence or presence of AGE-BSA for 24 hours.CCK8 were used to evaluate the proliferation capability, The DNA ladder was examined by agarose gel electrophoresis, and the cell apoptosis was determined by Annexin V/PI flow cytometry. Reactivated oxygen species (ROS) were analyzed using the ROS assay kit, the content of MDA, the activity of SOD was measured.Results (1)The hMSCs exhibited adherent long spindle-shaped cells. (2)The immuno phenotype of hMSCs show that both stromal cell-associated markers (CD44,) and the stem cell-associated marker (CD105) are positive, they are 97.8% and 98.9%, while blood cell, endothelial cell-associated marker(CD34) is negative, it is only 0.8%.(3)Compared with the control group, co-culturing with 20 ug/ml,50 ug/ml,100 ug/ml,200 ug/ml AGE-BSA increased the rate of apoptosis. Simultaneously, AGE-BSA decreased proliferation of the cells. Both of them are in a dose dependent manner.(4)Compared with the control group, co-culturing with 20 ug/ml,50 ug/ml,100 ug/ml,200 ug/ml AGE-BSA increased ROS production, the content of MDA in cells, Simultaneously, AGE-BSA decreased antioxidase of the cells. Both of them are in a dose dependent manner.Conclusions AGEs increases the production of ROS in hMSCs, while weakens the production of antioxidases, it increases the oxidative stress, breaks the homeostasis of cells, accordingly, AGEs inhibits the proliferation of human Mesenchymal stem cells and increases the apoptosis of these cells, all of them are in a dose dependent manner.