| BackgroundStem cells are a kind of cells which possess self-renewing abilities during the whole life activity. The dult stem cells abilities of differentiation, self-renewing and self-maintenance tended to be limited so that it, which is characterized of short life time, could differentiate by means of mono-direction or double direction. What is the most important is that it can not differentiate all cells from the three embryonic layer. Many investigation have confirmed that there existed hepatic stem cells in adult liver, which not only possesed self-renewing ability, but also could differentiate into bile duct cells, hepatic parenchymal cells and intestinal tract epithelial cells. It not only possessed self-renewing ability but also produced all characterized cells which can developed tissue itself. Nowadays, the researches about hepatic stem cells have become a heated topic in the world. Hepatic stem cells are a indispensible key role in the physiological and pathophysiological process of liver cell development, growth, proliferation, regeneration, fibrosis and carcinomatous change. In the same time, it plays an important role involving in liver diseases and stem cell transplantation, bioartificia liver in vitro and gene therapy. However, there exist some disputes and difficulties about the localization, separation and culture as well as regulation in vitro of hepatic stem cells. And how to locate and detect the origin of hepatic stem cells? How to separate and culture them simple and easy? What are the conditions of their differentiation and proliferation in vitro? In order to clarify the location of hepatic stem cells, DAB was adopted to construct the proliferation model of cell in rats, and its origin and location of hepatic stem cell were studied by methods of pathology and inimunohistochemistry, and hepatic stem cells were isolated from adult rats liver by portal vein filling digestion and gradient centrifugation and cultured in vitro, then the biological properts of the cells were observed by phase contrast microscope, cell transmission electon microscope and scanning electron microscope and by measuring cellgrowth curve. Hepatic cell growth factor and epithelial cell growth factor were applied to research the mechanism of stem cell proliferation as interventional factors primarily.ObjectiveThe study aimed at exploring the origin and location of hepatic stem cells and studying a simple, cheap and credible method of separating and culturing stem cells in vitro. Meantime, the cultured stem cells were regulated with cytokines to explore a new way for whose clinical applications.Methods1.Constructing the model of hepatic stem cell and locating the stem cells in rats: DAB was used to constructe the model of hepatic stem cells and pathological and immunohistochemistry method were used to study the distribution of hepatic stem cells of SD rats.2.1solating culturing and ultrastructure observation of hepatic stem cells of rats:The methods of portal vein filling digestion and gradient centrifugation were used to isolate stem cells from adult rats liver to culture in vitro and biological properts of the cells were observed by cell phase contrast microscope, cell transmission electron microscope, scanning electron microscope and measuring cell growth curve.3.Effect of HGF and EGF on culturing hepatic stem cell in vitro: separated hepatic stem cells were cultured with serum-free medium, different concentration EGF, HGF and EGF associated with HGF were added respectively and MTT chromatometry was used to analyse the proliferation effects of hepatic stem cells intervened by different cytokines.Resultsl.The CK18 CK19 and CD34 positive cells were hepatic stem cells with inimunohistocheniistry . Hepatic stem cells located in portal area and It's quantity increased with aggravation of liver injury, which clarified that the bile duct epithelia cells in portal area were potential, effective hepatic stem cell pool.2.The multiple potential hepatic stem cells could be isolated from adultrat liver f...
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