Intravenous transplantation of GDNF gene-modified CD34~+ cells from human umbilical cord blood for cerebral ischemie-reperfusion inury in rats
10.3760/cma.j.issn.0254-1785.2010.03.008
- VernacularTitle:静脉移植转染GDNF基因的人脐血CD34~+细胞改善局灶性脑缺血大鼠的神经功能
- Author:
Yali OU
;
Guolong YU
- Publication Type:Journal Article
- Keywords:
Nerve growth factor;
Cord blood stem cell transplantation;
Genes;
Transfection;
Recovery of function;
Brain ischemia
- From:
Chinese Journal of Organ Transplantation
2010;31(3):162-166
- CountryChina
- Language:Chinese
-
Abstract:
Objective To observe the neurologicaI functional recovery after intravenous transplantation of human umbilical cord blood(HUCB)CD34~+ cells transfected with glial cell-derived neurotrophic factor (GDNF) and green fluorescent protein (GFP) in SD rats with middle cerebral artery occlusion (MCAO).and tO investigate the survival,migration and neural differentiation of the graft cells.Methods (1)CD34~+ cells were isolated from HUCB using centrifuge combined with immune beads and then identified by flow cytometry,and transfected by the recombinant plasmid of GDNF.GFP or GFP plasmid by liposome method.(2) Sixty aduh male SD rats with MCAO were randomly divided into three groups (n=20 in each group):GDNF-GFP-CD34~+ cells group,in which the GDNF-GFP-CD34~+ cells were transplanted intravenously at the 24th h after the establishment ofmodels of MCAO;GFP-CD34~+ cells group:in which the GFP-CD34~+ cells were transplanted intravenously at the Same time;Normal saline group,in which normal saline was injected at the same time.Fifteen SD rats served as sham-operated group.(3) Neurological functional measurements were performed using the modified neurolc'gical severity score.Quantitative histological determinations of infarct volume were performed using standard TTC staining and quantitative image analysis. The GDNF level in the cell culture or the cerebral tissue was measured by ELISA. Meanwhile, the survival and migration of GFP-labeled CD34~+ cells and the expression of astrocytie marker-GFAP and the neuron marker-neuronal nuclei (NeuN) were detected by immunohistochemical and fluorescent staining. Results (1) The GDNF level in the cell culture was significantly higher in GDNF-GFP-CD34~+ cells than in GFP-CD34~+ cells (P<0. 05). (2) No significant difference was found in the modified neurological severity score at the day 7 after transplantation among the three groups with MACO, but at day the 28 after transplantation, the neurological function in GFP-GDNF-CD34~+ group was improved significantly (5.0±1. O) as compared with the GFP-CD34~+ cells (5. 9 + 1.4) or saline groups (7. 0±1.7) (P<0. 05). The cerebral infarct volume in GFP-GDNF-CD34~+ group (142±44mm~3) was significantly decreased as compared with the GFP-CD34~+ group (196±58 mm~3) (P<0. 05) or saline group (233<50 mm~3 ) (P<0. 01 ). The GDNF level in cerebral tissue in GFP-GDNF-CD34~+ group was significantly increased as compared with the GFP-CD34~+ group (P<0. 05) at the 28th day after treatment. At the 28th day after treatment, the NeuN positive cells (6.7±2.0), and GFAP positive ceils (14.1±3.3) in GFP-GDNF-CD34~+ group were significantly increased as compared with the GFP-CD34~+ group (P<0. 05), but there were no positive cells in sham-operationgroup. Conclusion Intravenous transplantation of GDNF gene-modified CD34~+ cells from human umbilical cord blood could improve the neurological function in rats with MCAO. The increased GDNF level in cerebral tissue was one of possible mechanisms responsible for the different improvements.
