| Chitin is one of the most abundant biological natrue resources, have many kinds of physiological functions. But high insolubility restrict its application, so we need to carry on chemical milling to it , carry on the structure and decorate or degrade its strand partly. Chitin's partially deacetylated product is chitosan. Unlike these polysaccharides, chitooligosaccharide and oligochitosan, which are the oligomer homologs of chitin or chitosan, are soluble and can be satisfactorily utilized.Previous investments have shown that chitin, chitosan or their oligosaccharides were able to stimulate immune system. Either intravenously or intraperitoneal administered to mouse,these polysacca-rides or oligosaccharides were able to increase polymorphonuclear cells in blood or peritoneal exudate, and activated macrophages, T lymphocytes and NK cells of mouse. These cells released numbers of immune-functional molecules, such as interleukin, TNF, IFN, H2O2 and NO, which protected animals from pathogen infection or tumor invasion. Chitin and its derivative can strengthen the function of resisting on the microorganism of many kinds of etiologies of organism through regulating the function of immune system of organism , have effect of suppressing to tumour cells, such as mouse MethA fibrosarcoma , S180 tumour, MM44 entity tumour and Lewis lung cancer, etc.Most of these investiments, however, were carried out through in vivo experiments, and research on stimulating function of oligochitosan to one particular part of immune system was very few. Here we managed to make cultured mice peritoneal macrophages be directly influenced by oligochitosan,and be stimulated by IFN-r before oligochitosan added,then measured the changes of gene transcriptionand translation level of both IL-1 and IMF- a , respectively by methods of relatively quantitive RT-PCR and ELISA.First, RT-PCR results showed that 18 hours was the most effective time and 40ug/ml was the most effective concentration of oligochitosan ,then by the same method, confirm that 4hours is the most effective time and lOOu/ml is the most effective concentration of IFN-r stimulating. Because IFN-r can enhance IL-1 and TNF-a gene expression of macrophages alone, so add IFN-r to microphages alone for 22 hours, then examined by RT-PCR, the results showed that IL-1 and TNF-a gene expression have no remarkable difference compared with the blank contrast group . this means that joining of IFN-Yonly to stimulate and make the macrophages in sensitive state, help to strengthen the enhancing effect of oligochitosan on IL-1 and TNF-a gene expression of macrophages.After confirming the most effective time and the most effective concentration of oligochitosan and IFN- Y , select three groups of cells, experiment group join lOOU/ml IFN-r , after 4 hours, add 40ug/ml oligochitosan; oligochitosan contrast group add oligochitosan only;blank contrast groupadd neither IFN-Y nor oligochitosan, then the three goups culture 18 hours together. Collect three groups culture supernatant for ELISA experiment, RT-PCR and ELISA results indicated that both transcription and translation of two cytokine genes can be enhanced obviously.RT-PCR results showed that the transcription level of IL-1 and TNF-a genes of oligochitosan contrast group were as 2. 1 times and 1. 9 times as the level of blank contrast group(p < 0.01), and the transcription level of IL-1 and TNF-a genes of experiment groupwere as 3.7 times and 4.3 times as the level of blank contrast group(p <0. 01).ELISA results showed that the translation level of IL-1 and TNF- a detected in culture supernatant of oligochitosan contrast group were as 2.4 times and 2 times as the level of blank contrast group (p < 0.01), and the translation level of IL-1 and TNF-a genes of experiment group were as 4. 2 times and 2. 9 times as the level of blank contrast group(p< 0. 01).Conclusively, our experiments indicated that oligochitosan was an immune stimulant, which was able to enhance IL-1 and TNF-a gene expression of mice macrophages,... |
PDF Full Text Request