BACKGROUND： Neurotrophin-4 （NT-4） can promote neuronal growth, development, differentiation, maturation, and survival. NT-4 can also improve recovery and regeneration of injured neurons, but cannot pass through the...BACKGROUND： Neurotrophin-4 （NT-4） can promote neuronal growth, development, differentiation, maturation, and survival. NT-4 can also improve recovery and regeneration of injured neurons, but cannot pass through the blood-brain barrier, which limits its activity in the central nervous system. Delivering NT-4 into the central nervous system v/a cells or vectors may have therapeutic benefit. OBJECTIVE： To construct a recombinant vector with a human embryonic brain-derived NT-4 gene and pEGFP-NI. DESIGN, TIME AND SETTING： Neural genetic engineering experiment. The study was performed at the Neuroscience Institute of Kunming Medical College between October 2007 and March 2008. MATERIALS： The pEGFP-N1 plasmid vector was provided by Kunming Institute of Zoology, Chinese Academy of Sciences; embryonic brain tissues were provided by the First Affiliated Hospital of Kunming Medical College. TRIzol RNA extraction Kit was purchased from Sigma （USA）, One Step RNA PCR Kit （AMV） etc. were from Takara （Dalian, China）. METHODS： Total RNA was extracted from human embryonic brain tissues using Trizol. The agarose gel electrophoresis showed two bands： 18 S and 28 S, which were essential subunits of total RNA. The human NT-4 DNA was obtained via RT-PCR and inserted into the pEGFP-N1 vector using ligation and transformation reaction. MAIN OUTCOME MEASURES： The sequencing results of the DNA in the recombinant of NT-4- pEGFP-NI. RESULTS： The NT-4-pEGFP-N1 vector was sequence-verified and showed the expected molecular weight. CONCLUSION： The recombinant of NT-4-pEGFP-N1 was constructed successfully in vitro.展开更多
Objective Cloning and sequencing of the human neurotrophin 4 gene.Methods With the chromosomal DNA of human blood lymphocytes as template,hNT 4 coding genes were amplified by polymerase chain reaction and recombinated...Objective Cloning and sequencing of the human neurotrophin 4 gene.Methods With the chromosomal DNA of human blood lymphocytes as template,hNT 4 coding genes were amplified by polymerase chain reaction and recombinated into phage vector pGEM T Easy,which were sequenced by using Sanger’s single stranded DNA terminal termination method.Results The sequence of the cloned gene is completely the same as that reported in the literature.Conclusion This study successfully cloning and sequenced the gene of mhNT 4,and it would be convenient for us to study the expression of mhNT 4 in eukaryote,and to continue the research on the gene therapy of Alzheimer’s disease intensively.This study indicate that the hNT 4 is conservative in different races and individuals.展开更多
The p75 neurotrophin receptor, which is a member of the tumor necrosis factor receptor superfamil facilitates apoptosis during development and following central nervous system injury. Previous studies have shown that ...The p75 neurotrophin receptor, which is a member of the tumor necrosis factor receptor superfamil facilitates apoptosis during development and following central nervous system injury. Previous studies have shown that programmed cell death is likely involved in the neurotoxic effects of 3, 4-methylenedioxy-N-methylamphetamine （MDMA）, because MDMA induces apoptosis of immortalized neurons through regulation of proteins belonging to the Bcl-2 family. In the present study, intrapedtoneal injection of different doses of MDMA （20, 50, and 100 mg/kg） induced significant behavioral changes, such as increased excitability, increased activity, and irritability in rats. Moreover, changes exhibited dose-dependent adaptation. Following MDMA injection in rat brain tissue, the number of apoptotic cells dose-dependently increased and p75 neurotrophin receptor expression significantly increased in the prefrontal cortex, cerebellum, and hippocampus. These findings confirmed that MDMA induced neuronal apoptosis, and results suggested that this effect was related by upregulated protein expression of the p75 neurotrophin receptor.展开更多
基金Supported by:the Scientific Research Foundation of Yunnan Provincial Education Academy,No.07C10388
文摘BACKGROUND： Neurotrophin-4 （NT-4） can promote neuronal growth, development, differentiation, maturation, and survival. NT-4 can also improve recovery and regeneration of injured neurons, but cannot pass through the blood-brain barrier, which limits its activity in the central nervous system. Delivering NT-4 into the central nervous system v/a cells or vectors may have therapeutic benefit. OBJECTIVE： To construct a recombinant vector with a human embryonic brain-derived NT-4 gene and pEGFP-NI. DESIGN, TIME AND SETTING： Neural genetic engineering experiment. The study was performed at the Neuroscience Institute of Kunming Medical College between October 2007 and March 2008. MATERIALS： The pEGFP-N1 plasmid vector was provided by Kunming Institute of Zoology, Chinese Academy of Sciences; embryonic brain tissues were provided by the First Affiliated Hospital of Kunming Medical College. TRIzol RNA extraction Kit was purchased from Sigma （USA）, One Step RNA PCR Kit （AMV） etc. were from Takara （Dalian, China）. METHODS： Total RNA was extracted from human embryonic brain tissues using Trizol. The agarose gel electrophoresis showed two bands： 18 S and 28 S, which were essential subunits of total RNA. The human NT-4 DNA was obtained via RT-PCR and inserted into the pEGFP-N1 vector using ligation and transformation reaction. MAIN OUTCOME MEASURES： The sequencing results of the DNA in the recombinant of NT-4- pEGFP-NI. RESULTS： The NT-4-pEGFP-N1 vector was sequence-verified and showed the expected molecular weight. CONCLUSION： The recombinant of NT-4-pEGFP-N1 was constructed successfully in vitro.
文摘Objective Cloning and sequencing of the human neurotrophin 4 gene.Methods With the chromosomal DNA of human blood lymphocytes as template,hNT 4 coding genes were amplified by polymerase chain reaction and recombinated into phage vector pGEM T Easy,which were sequenced by using Sanger’s single stranded DNA terminal termination method.Results The sequence of the cloned gene is completely the same as that reported in the literature.Conclusion This study successfully cloning and sequenced the gene of mhNT 4,and it would be convenient for us to study the expression of mhNT 4 in eukaryote,and to continue the research on the gene therapy of Alzheimer’s disease intensively.This study indicate that the hNT 4 is conservative in different races and individuals.
基金supported by the National Natural Science Foundation of China (MDMA Neurotoxicity and Mechanism),No.C090302
文摘The p75 neurotrophin receptor, which is a member of the tumor necrosis factor receptor superfamil facilitates apoptosis during development and following central nervous system injury. Previous studies have shown that programmed cell death is likely involved in the neurotoxic effects of 3, 4-methylenedioxy-N-methylamphetamine （MDMA）, because MDMA induces apoptosis of immortalized neurons through regulation of proteins belonging to the Bcl-2 family. In the present study, intrapedtoneal injection of different doses of MDMA （20, 50, and 100 mg/kg） induced significant behavioral changes, such as increased excitability, increased activity, and irritability in rats. Moreover, changes exhibited dose-dependent adaptation. Following MDMA injection in rat brain tissue, the number of apoptotic cells dose-dependently increased and p75 neurotrophin receptor expression significantly increased in the prefrontal cortex, cerebellum, and hippocampus. These findings confirmed that MDMA induced neuronal apoptosis, and results suggested that this effect was related by upregulated protein expression of the p75 neurotrophin receptor.
