Modulation Of The Activity Of Wuchangbream(Megalobrama Amblycephala) Phagocytes And Research On Function Mechanism By Chitosan

In this work, on the basis of the dissection structure of Wuchang bream, the separation and morphological and functionally identification of the head kidney phagocytes, we aim to investigate the effects of Oligochitosan(OCS), water soluble chitosan(WSC) and acid soluble chitosan(ASC) on the activity of the Wuchang bream (Megalobrama amblycephala) head-kidney phagocytes and the mechanism of action. In this paper we preliminary studied combining capacity between macrophages and different ligand. Foundations for the further study. The results were as follows:(1) The dissection structure indicated that Wuchang bream head-kidney was located above the sides of musculus retractor in the chest, the head-kidney was tabular, divided into two parts, and the two parts were connected at base. The base of head kidney was connected to the forefront of the kidney through the base of the forth processus transversus vertebrarum and the base sides hole stitch of tabula osteocomma. In front of dissepiment between cardiac chambers and abdomen was head kidney, and the kidney was at posterior side. Microstructure showed that the head kidney was lymphoid tissue, there was lymphocyte zones and granulocyte zones. Ultrastructural features showed the cell types of head kidney.(2) Microstructure showed the separated cells contained phagocytes and a small amount of lymphocytes. Ultrastructural features showed that there were two major types of cells. There were many dense lysosomes of various shape and size in the cytoplasm of macrophage, also contained mitochondria, large vacuoles, oval nuclei locates at the side of cells. There were large vacuoles and rod granule of uneven size in polymorphonuclear(PMN). After3h phagocytes in culture were adherence and had a rounded appearance. After18h anchorage-dependent phagocytes had spread on the bottom of the petri dish. Cells were stretched longer and in shapes.The phagocytosis experiment results indicated that the separated cells and cultured cells for3h all had the ability to phagocytose the formalin killed Staphylococcus aureus(F-SA), the Phagocytic Percentage (PP) of the separated cells was25.50±0.58, the Phagocytic Index (PI) was90.80±0.84. There were two types cells which had phagocytic activity respectively macrophage and polymorphonuclear(PMN) under transmission electron microscope. There were vacuoles in macrophages, the F-SA in vacuoles was out of shaped and degradation. And the same phenomenon observed in polymorphonuclear (PMN). The results of the respiratory burst activity indicated that the fluorescence intensity of control group was only111.07and fluoresce cells were accounted for4.64%in the phagocytes. But the fluorescence intensity of experimental group was up to450.72, fluoresce cells were accounted for78.45%in the phagocytes. The results showed that there was extremely significant difference between experimental group and control group (P<0.01).(4) The results of the effects of Oligochitosan(OCS), water soluble chitosan(WSC) and acid soluble chitosan(ASC) on the activity of the Wuchang bream (Megalobrama amblycephala) head-kidney phagocytes indicated that OCS (100μg/ml), WSC (100μg/mL) and ASC (100μg/mL) were all able to improve the phagocytic activity of the obtained phagocytes and stimulate the respiratory burst activity of head-kidney phagocytes after incubation for30min. In comparison with control group, the Phagocytic Percentage (PP) and Phagocytic Index (PI) in experiment group showed significantly difference (P<0.01). But there was no significant difference between experimental group (P>0.05). The experiments group also showed extremely significantly difference(P<0.01) with control group respective in the percentage of fluoresce cell and the fluorescence intensity of fluoresce cell (two parameters of respiratory burst activity). However, on the percentage of fluoresce cell, the phagocytes stimulated by WSC showed strongest respiratory burst activity. But on the fluorescence intensity of fluoresce cell, the phagocytes stimulated by ASC showed the strongest respiratory burst activity.(5) The results of the binding competition studies indicated that all three chitosan had inbibitional effect on the binding of M-FITC-BSA and macrophages. We also found that compared with respective positive control group, the percentage of fluoresce cell and the fluorescence intensity of fluoresce cell in Oligochitosan+mannose group, Water-soluble chitosan+mannose group, Acid soluble chitosan+mannose group were all decreased. And Oligochitosan+galactose group, Water-soluble chitosan+galactose group, Acid soluble chitosan+galactose group were no obvious change...