BACKGROUND： Previous studies have shown that transplantation of vascular endothelial growth factor （VEGF）-modified neural stem cells （NSC） provides better outcomes, compared with neural stem cells, in the treatme...BACKGROUND： Previous studies have shown that transplantation of vascular endothelial growth factor （VEGF）-modified neural stem cells （NSC） provides better outcomes, compared with neural stem cells, in the treatment of brain damage. OBJECTIVE： To compare the effects of VEGF-modified NSC transplantation and NSC transplantation on radiation-induced brain injury, and to determine neuron-specific enolase （NSE） expression in the brain. DESIGN, TIME, AND SETTING： The randomized, controlled study was performed at the Linbaixin Experimental Center, Second Affiliated Hospital, Sun Yat-sen University, China from November 2007 to October 2008. MATERIALS： VEGF-modified C17.2 NSCs were supplied by Harvard Medical School, USA. Streptavidin-biotin-peroxidase-complex kit （Boster, China） and 5, 6-carboxyfluorescein diacetate succinimidyl ester （Fluka, USA） were used in this study. METHODS： A total of 84 Sprague Dawley rats were randomly assigned to a blank control group （n = 20）, model group （n = 20）, NSC group （n = 20）, and a VEGF-modified NSC group （n = 24）. Rat models of radiation-induced brain injury were established in the model, NSC, and VEGF-modified NSC groups. At 1 week following model induction, 10 pL （5 ×10^4 cells/μL） VEGF-modified NSCs or NSCs were respectively infused into the striatum and cerebral cortex of rats from the VEGF-modified NSC and NSC groups. A total of 10μL saline was injected into rats from the blank control and model groups. MAIN OUTCOME MEASURES： NSE expression in the brain was detected by immunohistochemistry following VEGF-modified NSC transplantation. RESULTS： NSE expression was significantly decreased in the brains of radiation-induced brain injury rats （P 〈 0.05）. The number of NSE-positive neurons significantly increased in the NSC and VEGF-modified NSC groups, compared with the model group （P 〈 0.05）. NSE expression significantly increased in the VEGF-modified NSC group, compared with the NSC group, at 6 weeks following transplantation （P 〈 0.05）. CONCLUSION： VEGF-modified NSC transplantation increased NSE expression in rats with radiation-induced brain injury, and the outcomes were superior to NSC transplantation.展开更多
To develop a rabbit corneal endothelial (RCE) cell line, in vitro culture of RCE cells was initiated from Oryctolagus curiculus corneas and a novel RCE cell line was established in this study. To initiate the primary ...To develop a rabbit corneal endothelial (RCE) cell line, in vitro culture of RCE cells was initiated from Oryctolagus curiculus corneas and a novel RCE cell line was established in this study. To initiate the primary culture of RCE cells, corneas from rabbit eyes were sliced and attached into glutin-coated wells with endothelial cell surface down. After being cultured at a time-gradient interval from 48 to 6 h, the corneal slices were detached and reattached into new wells, respectively. Cells in the wells containing only a pure population of RCE cells were collected and cultured in 20% FBS-DMEM/F12 medium con- taining chondroitin sulfate, ocular extract, epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), carboxymethyl-chitosan, N-acetylglucosamine hydrochloride, glucosamine hydrochloride, culture medium of rabbit corneal stromal cells and oxidation-degradation products of chondroitin sul- fate at 37℃, 5% CO2. The cultured RCE cells, in quadrangle and polygonal shapes, proliferated to con- fluence 3 weeks later. During the subsequent subculture, the shape of RCE cells changed gradually from polygonal to more fibroblastic. A novel RCE cell line, growing at a steady rate, with a population doubling time of 53.8 h, has been established and subcultured to passage 67. Chromosome analysis showed that the RCE cells exhibited chromosomal aneuploidy with the modal chromosome number of 44. The results of immuno-cytochemical staining with neuron specific enolase (NSE) confirmed that the RCE cells were in neuroectodermal origin. Combined with the results of vascular endothelial growth factor (VEGF) treatment and endothelial cell morphology recovery, it can be concluded that the cell line established here is an RCE cell line. This RCE cell line may serve as a useful tool in theoretical re- searches of mammalian corneal endothelial cells, and may also have potential application in artificial corneal endothelium development.展开更多
基金Supported by:the National Natural Science Foundation of China,No.30870750the Doctor Priming Program of Natural Foundation of Guangdong Province,No. 8451008901000672+1 种基金the Medical Scientific Research Foundation Program of Guangdong Province,No. B2008044the Youth Teacher Foundation Program of Sun Yat-sen University, No,3177915
文摘BACKGROUND： Previous studies have shown that transplantation of vascular endothelial growth factor （VEGF）-modified neural stem cells （NSC） provides better outcomes, compared with neural stem cells, in the treatment of brain damage. OBJECTIVE： To compare the effects of VEGF-modified NSC transplantation and NSC transplantation on radiation-induced brain injury, and to determine neuron-specific enolase （NSE） expression in the brain. DESIGN, TIME, AND SETTING： The randomized, controlled study was performed at the Linbaixin Experimental Center, Second Affiliated Hospital, Sun Yat-sen University, China from November 2007 to October 2008. MATERIALS： VEGF-modified C17.2 NSCs were supplied by Harvard Medical School, USA. Streptavidin-biotin-peroxidase-complex kit （Boster, China） and 5, 6-carboxyfluorescein diacetate succinimidyl ester （Fluka, USA） were used in this study. METHODS： A total of 84 Sprague Dawley rats were randomly assigned to a blank control group （n = 20）, model group （n = 20）, NSC group （n = 20）, and a VEGF-modified NSC group （n = 24）. Rat models of radiation-induced brain injury were established in the model, NSC, and VEGF-modified NSC groups. At 1 week following model induction, 10 pL （5 ×10^4 cells/μL） VEGF-modified NSCs or NSCs were respectively infused into the striatum and cerebral cortex of rats from the VEGF-modified NSC and NSC groups. A total of 10μL saline was injected into rats from the blank control and model groups. MAIN OUTCOME MEASURES： NSE expression in the brain was detected by immunohistochemistry following VEGF-modified NSC transplantation. RESULTS： NSE expression was significantly decreased in the brains of radiation-induced brain injury rats （P 〈 0.05）. The number of NSE-positive neurons significantly increased in the NSC and VEGF-modified NSC groups, compared with the model group （P 〈 0.05）. NSE expression significantly increased in the VEGF-modified NSC group, compared with the NSC group, at 6 weeks following transplantation （P 〈 0.05）. CONCLUSION： VEGF-modified NSC transplantation increased NSE expression in rats with radiation-induced brain injury, and the outcomes were superior to NSC transplantation.
基金Supported by the Grant from the Chinese Ministry of Agriculture [No.NYCYTX-43] the Zhejiang Provincial Natural Science Foundation of China [No.R3090332]
基金863 High-Tech Research and Development Program of China (Grant No. 2001AA625050)
文摘To develop a rabbit corneal endothelial (RCE) cell line, in vitro culture of RCE cells was initiated from Oryctolagus curiculus corneas and a novel RCE cell line was established in this study. To initiate the primary culture of RCE cells, corneas from rabbit eyes were sliced and attached into glutin-coated wells with endothelial cell surface down. After being cultured at a time-gradient interval from 48 to 6 h, the corneal slices were detached and reattached into new wells, respectively. Cells in the wells containing only a pure population of RCE cells were collected and cultured in 20% FBS-DMEM/F12 medium con- taining chondroitin sulfate, ocular extract, epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), carboxymethyl-chitosan, N-acetylglucosamine hydrochloride, glucosamine hydrochloride, culture medium of rabbit corneal stromal cells and oxidation-degradation products of chondroitin sul- fate at 37℃, 5% CO2. The cultured RCE cells, in quadrangle and polygonal shapes, proliferated to con- fluence 3 weeks later. During the subsequent subculture, the shape of RCE cells changed gradually from polygonal to more fibroblastic. A novel RCE cell line, growing at a steady rate, with a population doubling time of 53.8 h, has been established and subcultured to passage 67. Chromosome analysis showed that the RCE cells exhibited chromosomal aneuploidy with the modal chromosome number of 44. The results of immuno-cytochemical staining with neuron specific enolase (NSE) confirmed that the RCE cells were in neuroectodermal origin. Combined with the results of vascular endothelial growth factor (VEGF) treatment and endothelial cell morphology recovery, it can be concluded that the cell line established here is an RCE cell line. This RCE cell line may serve as a useful tool in theoretical re- searches of mammalian corneal endothelial cells, and may also have potential application in artificial corneal endothelium development.
