ABSTRACT
BACKGROUND:Currently,there is not a standard method for in vitro culture and large scale amplification of umbilical cord blood mesenchymal stem cells(UCB-MSCs).OBJECTIVE:To investigate the isolation,purification and culture of UCB-MSCs in vitro,and to detect its surface marker variation.METHODS:The monocytes were harvested from UCB using 1.077 g/cm3 lymphocytes separating solution and density gradient centrifugation,followed by incubation in an incubator containing 5%CO2 at 37℃.The cell morphological changes were observed at different time points and the expression of surface marker was detected using flow cytometry.RESULTS AND CONCLUSION:The monocytes isolated from the UCB grew initially into numerous hematopoietic cell clones,most of which were granulocyte/macrophage colony-forming units and burst forming unit-erithroid,increasing by(37.1±2.3)and (10.4±1.7),respectively.Switzerland staining showed most of them were granulocyte clones(80,1±85.2)%,next was erythroid clones(14.2±1.8)%.At 7 days after culture,some shuttle fibroblast-like cells and fiat osteogenic-like cell spread the whole plastic well.At 14 days after culture,flow cytometry showed CD38+ cells accounted for 1.64%,and CD34+/CD38+ cells accounted for 1,71%,and CD34+/CD38- were 0.55%.PI+ and Annexin-V+ cells accounted for 0.05% and 0.18% respectively.At 21 days after culture,CD38+,CD34+/CD38+ and CD34+/CD38- cells were 74.32%,1.61%,and 0.24%.The results reveled that UCB-MSCs can be isolated and cultured in vitro.
ABSTRACT
In order to investigate the protective effect of hypoxic preconditioning on the cerebral ischemia-reperfusion injury, the expression of Bcl-2 and Bax was detected by using immunohistochemical staining after 3 h cerebral ischemia followed by 1, 6, 12, 24 and 48 h reperfusion respectively in rats treated with or without hypoxic preconditioning before cerebral ischemia. In addition,the apoptosis of neural cells and the behavioral scores for neurological functions recovery were evaluated by TUNEL staining and "crawvling method", respectively. Compared with control group (cerebral ischemia-reperfusion without hypoxic preconditioning), the expression of Bcl-2 was significantly increased, but that of Bax decreased in the hypoxic preconditioning group (cerebral ischemiareperfusion with hypoxic preconditioning), both P＜0. 05. The pre-treatment with hypoxic preconditioning could reduce the apoptosis of neural cells and promote the neurological function recovery as compared to control group. It was suggested that hypoxic preconditioning may have protective effects on the cerebral ischemia-reperfusion injury by inhibiting the apoptosis of neural cells, increase the expression of Bcl-2 and decrease the expression of Bax.
ABSTRACT
Objective To investigate the involvement of Bcl-2 and Bax mRNA expressions in mitochondrial toxin 3-nitropropionic acid(3-NPA)induced ischemic tolerance to focal cerebral ischemia in rats.Methods Rats were administrated 3-NPA intraperitoneally at dose of 20 mg/kg 3 days prior to a 2 h middle cerebral artery occlusion followed by 1 h,6 h,12 h,24 h and 48 h of reperfusion.The Bcl-2 and Bax mRNA expressions were measured by reverse transcriptase polymerase chain reaction and compared to the sham operation group and ischemic reperfusion group.Results Compared to the sham operation group,the ischemic reperfusion and 3-NPA pretreated groups exhibited an increase in Bcl-2 and Bax mRNA after reperfusion every time point of focal cerebral ischemia(all(P