| The discovery of neurogenesis from neural stem cells (NSCs) in the adult brain has raised hope for the development of new regenerative therapies for neurological disease. Relevant type of neurons used for therapies can be obtained by regulate the differentiation of NSCs. NSCs transplanted studies on animal models have shown that the NSCs can proliferation and differentiation in recipient injury brain and functional synapse junctions with ectopic NSCs have been found in injury part. The NSCs transplanting therapy was considered as a promise method for degenerative neurological diseases.Alzheimer's disease (AD) is the most common form of age-related neuro -degenerative disorders. The pathogenesis of this and other neurodegenerative diseases remains unclear. AD is characterized by synaptic loss and neuronal death in the cerebral cortex and the hippocampus, with the presence of extensive extracellular amyloid plaques and intracellular neurofibrillary tangles, and effective treatments are currently lacking. Neurogenesis occured in the adult mammalian brain may play roles in improvement of learning and memory processes and recovery from injury, suggesting that the NSCs can be an ideal cell resource for cell replacement therapy for AD. Whether NSCs can survive, form functional synapses, the proliferation and differentiation of them into relevant neuron types in vitro are the key factor for an successfully transplantation therapy. There were some obstacles for AD therapy by NSCs transplantation. For example, the transplanted NSCs suffered apoptosis after transplanted into APP transgenic AD model rat. The study shows that the β-amyloid protein in model rat has toxic action on transplanted NSCs. In addition, how to regulate the transplanted NSCs to differentiating into neurons is another obstacle in AD therapy.To solve the problem of deficient survive and neuronal differentiation of transplanted NSCs is very important in cell replace therapy for AD. but the methods are currently lacking. Falk(Experimental cell research, 2002) has reported that the single gene can drive NSCs into specific fate. The objective of this study is try to solve the problem by transfecting ectopic Humanin gene targeted to NSCs. Humanin(HN) is a 24 amino acids linear polypeptides. The present study has been shown that Humanin could ameliorates neuronal cells death caused by various types of familial Alzheimer disease(AD) genes[V642I and K595N/M596L (NL) mutants of amyloid precursor protein (APP), M146L-presenilin (PS) 1, and N141I-PS2, A617G-APP, L648P-APP, A246E-PS1, L286V-PS1, C410Y-PS1, and H163R-PS1] and by Aβ(Aβ1-42,Aβ1-43,Aβ25-35)with clear action specificity ineffective on neurotoxicity by polyglutamine repeat Q79 or superoxide dismutase 1 mutants. Although the previous study shows that HN is a promising drug for therapy of AD, the obstacle still exists including that: (1) the effects of HN on NSCs remain unclear; (2) It is difficult for the drug reach the injury part of brain because of blood-brain barrier and metabolize enzyme in blood,and the already lost neuron cannot be compensated by HN. So, if the HN can prevent the NSCs from apoptosis induced by Aβ, there will be provide a functional cell resource for AD therapy if NSCs can express HN by itself.To study the effect of HN on NSCs and the possibility that if HN can be expressed by NSCs, studies were carried out study as below.Our study shows that the [Gly14]-Humanin, one of the most powerful variant of this peptide,can effectively enhances the proliferation and neuronal defferentiaion and inhibits the toxic action of Aβ. The screened cell clone consistently express [Gly14]-Humanin and possesses a character of Aβ-resistant when the recombinant plasmid was transfected into NSCs. 1. Isolation and identification of NSCs and study of the toxic action of AβP1-42 on cultured NSCs①With the presence of mitogen bFGF, serum free medium was used to culture the NSCs, and nestin, MAP2, GFAP, Gla-c were used to identify the NSCs, neuron, astroglia and ol... |
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