| ObjectiveThe study on nerve stem cell has become a hot spot of Neuroscience due to applying concept of nerve stem cell to nervous system since 90 in 20 century. Studying the proliferation and the differentiation mechanism of nerve stem cell has been a focal point for researchers following the success in isolation cell and culture cell, especially its orientation differentiation. Nerve stem cell is a multi-potential stem cell and can be continue proliferation in serum-free medium including EGF or bFGF, as well as keeping feature of stem cell. Nerve stem cell can be differentiation into astroblast, oligodendrocyte and nerve cell induced by factors. Proliferation and differentiation of nerve stem cell can be affected by various factors, such as many cytokine, growth factors, nutritional factors and external environment at al, which can induce nerve stem cell differentiation to various end cell. The data about mechanism of stem cell differentiation would be provided if cultured nerve stem cell in vitro was affected by promote differentiation factors and induced to a certain definite direction.Cellular skeleton is a fiber reticular formation in cytoplasm of eukaryocyte and is constituted by microtubules, microfilaments and intermediate filament, it is not only correlation with cell shape but also takes part in cell moving, material transport, information convection and cell division, differentiation and gene expression.Microtubule is one kind of the most coarse cellosilk, and is a dynamicstructure in cells, it can get together or depolymerization in a certain condition, therefore, various kinds arrangement model were showed in cells for adaptation moving cytoplasm and finishing their functions respectively. Microtubule was made up of a and tubulin, the difference in structure and function was made by structure different of tubulin. The tubulin and microtubule kept to change in order to maintain cell form which grew and changed during development of cell from immature to maturity, therefore, the change of cell form was depended on structure supporting of microtubule.Lysosome is a organell sacked by mono-limitans in cell, and contains more than of 70 hydrolase, can moving in cell, deformation, takes part in phago of cell self and cell xenoma, it is a digestive apparatus in cell, round in usually condition, having special marker enzyme( ACPase). micro-tube lysosome with ACPase positive was firstly found since 1973 , this kind of structure was observed in many cells, and called line lysosome, these two kinds of lysosome were regarded as manifestation pattern in different period, and was round when it was in relative silence, and line when activating state. Researching believed that lysosome has not only digestive function but also traffic action.Cellular skeleton maintains cell form and takes part in various functions in cells. Some researches believed that tubulin was traffic railway in cells, then, how was the correlation during development of lysosome and tubulin? Our experiment objective was to induce cortex nerve stem cell orientation differentiation to nerve cell, and observed the relationship between lysosome and tubulin during the process of cortex nerve stem cell orientation differentiation, that was the development of nerve cell from immature to maturity. We expected to provide the data about nerve stem cell orientation differentiation and material traffic during the development of nerve cell.Methods1. Primary and secondary cultured methods of cortex nerve stem cell for newly born rat: shaving out a thin layer of cerebral cortex in sterility in ultra clean table using newly born rat of Wistar in 24 hours, shears craniotripsotome,digestion in 0.25% pancreatin, filtration, centrifugation, implantation cell into serum-free medium ( DMEM/F12 (1:1 ) + 10% B27 + 20ng/ml EGF + 20ng/ml bFGF) in density of 1 ~2 105/ml, incubate in 37 degree, 5% C02, and changed medium one times every other day, observed clone formed under phase contrast microscope. Collecting clone which cultured 7 day...
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