| [Background and Objective] The worldwide research and development of chitin and chitosan are booming since the international symposium on chitin and chitosan was held in America in 1977. But chitin and chitosan's applications were limited for their insolubility in water. So the utilization of oligochitosan or oligochitin becomes important. Much domestic research work has been carried out and the oligosaccharide products have been industrialized.As the advances of research ,oligochitosan has been found with many unique physiological activities and functions. E.g.:l. It's low m.w. and it can be soluble in water and biodegradable by human body.2. It can enhance the phagocytosis function of macrophage and inhibit proliferation and metastasis of tumor cells.3. It can stimulate the proliferation of bifidobacterium and lactobacillus of human intestine and keep the homestasis of microorgnism in human intestine. 4. It has strong antimicrobial effect in low PH environment in vitro, and an excellent moisture retention effect.5. It can act as plant function regulator , stimulate plant growth and resist to insect .So many aspects of oligochitosan 's functions have been discovered by researchers, but the mechanism is still unclear.Macrophage is a kind of immune cell as well as accessory cell. It can function as antiinfection, anticancer,immune response and immunoregulation by phagocytosis, antigen presentation, cytokines secretion, lymphocyte activation ,reactive oxygen and NO production. It's an important part of body immune system.Our experiment observed the combinding of fluorescence labeled oligochitosan and macrophage from mouse peritoneum and explored the specific combinding activity of oligochitosan with the component of macrophage. And the experiment trys to elucidate the signal transduction pathway after the combinding of oligochitosan and macrophage. This work provides a foundation for further research on separation of combinding oligochitosan component of macrophage and elucidation on the immunoregulation mechanism of oligochitosan with macrophage. From this work we endeavor to find the theoretical ground for development of antitumor drugs and functional food from oligochitosan. It's a brandnew work and no report was found in recent literature.[Methods] 1. Preparation of macrophage from NIH mouse peritoneum: Macrophage cells were prepared from NIH mouse peritoneal fluids, adjust the cell density to 1 X 106cells/ml. 2. label oligochitosan with dansyl hydrazine:160mg oligochitosan solved in 1ml H2O was mixed with 2ml freshly prepared dansyl hydrazine and 1ml 3%TCA, then dry the mixture in vacuum centrifuge. 3. The combinding between macrophage and oligochitosan labeled with dansyl hydrazine: Macrophage cells were co-cultured with dansyl hydrazine labeled oligochitosan for Ih, 2h, 24h; The control group was treated with dansyl hydrazine only. After flushed the fluorescence intensity difference between treatment group and control group was observed under fluorescence microscope.4. The effect of D (+)-mannose > invertase, 3-dextran on the combinding of dansyl hydrazine labeled oligochitosan and macrophage: Macrophage cells were treated with D(+)-mannosex invertase or 3 - dextran for 20min, followed with a further co-culture with 3.75mM dansyl hydrazine labeled oligochitosan for another 2h. The control group was treated with dansyl hydrazine labeled oligochitosan directly without D(+)-mannose > invertase > 0 - dextran treatment. After flushing the fluorescence intensity difference between treatment group and control group was observed under fluorescence microscope. 5. label oligochitosan with AMAC: 400 u 1 of a 5mM aqueous solution of oligochitosan was lyophilized. A 30 u 1 sample of a 34mM solution of AMAC and lOOu 1 of a l.OM solution of NaBH3CN were added. Then the mixture was incubated at 45 癈 for 5h, then lyophilized. 6. The combination between macrophage and oligochitosan labeled with AMAC: Macrophage cells were co-cultured with AMAC labeled oligochitosan for 2min, 5min, 30min: The control group...
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