Immunomodulatory Effect Of Dioscorea Opposita Thunb Polysaccharide DOTP-B On RAW264.7 Cells

Polysaccharides are biological macromolecules with various functions that are widely distributed in nature.They not only serve as energy substances and cell structure skeletons,but also have various biological activities such as immune regulation,anti-oxidation,and anti-tumor.Plant polysaccharides have attracted much attention due to their wide range of sources,small side effects and remarkable curative effects.In addition,plant polysaccharide can act by mediating the host,without cytotoxicity,and it does not produce drug resistance when it acts on the body,so it is a natural immune regulator or immune enhancer.Yam polysaccharide is an important active ingredient of yam.In recent years,there are many studies on the physiological activity of yam polysaccharide.However,most of the current researches focus on the total polysaccharide or crude polysaccharide of yam,and the research on homogeneous polysaccharide is still lacking.In our laboratory,a water-soluble homogeneous dioscorea opposita thunb polysaccharide DOTP-B with an average molecular weight of 5623 Da was obtained by extraction and purification.It is mainly composed of glucose and galactose,and the molar ratio is 14.6:1.Its immunomodulatory effect has not been studied.Therefore,in this paper,the purified water-soluble dioscorea opposita thunb polysaccharide DOTP-B was used as the material,and RAW264.7 cells were used as the immunomodulatory model to study the immunomodulatory activity of dioscorea opposita thunb polysaccharide DOTP-B on RAW264.7 cells.The specific contents include:1.Taking RAW264.7 cells as an immunomodulatory model,after co-incubating with different concentrations of dioscorea opposita thunb polysaccharide DOTP-B for24 h,the immunomodulatory effects of dioscorea opposita thunb polysaccharide DOTP-B on RAW264.7 cells were evaluated by CCK-8 assay,neutral red phagocytosis assay,Griess assay and enzyme-linked immunosorbent assay(ELISA).The results show that the dioscorea opposita thunb polysaccharide DOTP-B has no cytotoxicity to RAW264.7 cells,and promotes cell proliferation,can significantly enhance the phagocytic ability of neutral red,and dose-dependently promotes RAW264.7 cells to synthesize NO and secrete cytokines TNF-α,IL-6 and IL-10.The above results show that dioscorea opposita thunb polysaccharide DOTP-B has significant immune-enhancing activity on RAW264.7 cells,which can enhance the resistance to pathogens,providing theoretical support for the development and utilization of new immune adjuvants.2.Taking RAW264.7 cells as an immune regulation model,after co-incubating with different concentration gradients of dioscorea opposita thunb polysaccharide DOTP-B for 24 h,these gene expression levels of inducible nitric oxide synthase(i NOS),TNF-α,IL-6 and IL-10 were detected by RT-PCR.The result showed that dioscorea opposita thunb polysaccharide DOTP-B promotes the release of NO,TNF-α,IL-6 and IL-10 from RAW264.7 cells by affecting the gene expression levels of i NOS,TNF-α,IL-6 and IL-10.The result indicated that dioscorea opposita thunb polysaccharide DOTP-B enhanced immune activity by regulating gene transcription level,which may be one of the mechanisms by which DOTP-B enhanced the immune activity of RAW264.7 cells.