The Studies On Structure Identification,Bioactivity Activity And The Mechanism Of Anti-tumor Activity Of The Morehella Esculenta(L.)pers Polysaccharide(ME-X)

Morehella esculenta(L.)Pers belongs to Morchellaceae,Morchella.its fruiting body is similar to sheep belly,so it is named Morehella esculenta(L.)Pers.The wild Morehella esculenta(L.)Pers is a good edible and medicinal fungus with delicious taste and rich nutrition.At present,the research on Morehella esculenta(L.)Pers focused on artificial cultivation.The structure of Morehella esculenta(L.)Pers has not been elucidated,and the mechanism of Morehella esculenta(L.)Pers polysaccharide inhibiting S180 solid tumors has not been reported.The crude polysaccharides of Morehella esculenta(L.)Pers were extracted by hot water extraction,which were fractionated and purified by DEAE-52 cellulose chromatography and dialysis method.Combined with HPGPC,HPLC,FT-IR,GC-MS and NMR analysis,the primary structure of Morehella esculenta(L.)Pers polysaccharide(ME-X)was analyzed to provide the basis for analysis of its secondary and tertiary structure.CCK-8 method and neutral red method were used to detect the effects of ME-X on proliferation of immune cells and phagocytosis of macrophage to evaluate the immune regulatory activity of ME-X;CCK-8 method was used to detect the results of ME-X on tumor cells to evaluate the anti-tumor activity of ME-X in vitro;The model of S180 tumor in mice was establelished to evaluate the anti-tumor activity in vivo of ME-X.The Illumina HiScq TM 200 technology was used for transcription sequencing of S180 tumor tissues to preliminarily explore anti-tumor mechanism,ELISA method was used to validate it.The main results were as follows:(1)40g crude polysaccharides were obtained from 200 g Morehella esculenta(L.)Pers fruiting bodies collected from Xiaojin County of Sichuan Province by hot water extraction,and the yield of polysaccharide was 20%.(2)ME-X was composed of mannose,glucose and galactose with a ratio of 4:4:1.It had a skeleton structure of(2,4?6)-alpha-D-mannopyranose with branches at 6-O and the annular branches consisted of three(1?6)-alpha-D-glucosepyranose and(1,2 ? 6)-alpha-D-galactopyranose.Residues of branch structures were terminated with ?2)-alpha-D-glucose.(3)The immunoregulatory activities of ME-X showed that: compared with the blank control group,ME-X had a good promoting effect on the proliferation of immune cells in a certain concentration range.When the concentration of ME-X was 10 ug/mL,the proliferative activities were best on T and B lymphocytes,with the proliferation rates of 31.8% and 63.02%respectively.When the concentration of ME-X was 20 ug/mL,the proliferative and phagocytic activity on RAW264.7 cells was best,with the proliferation rate of 63.09% and the phagocytic rate of 22.49%.(4)The anti-tumor activities of ME-X showed that compared with the blank control group,ME-X had a good inhibitory effect on the proliferation of tumor cells in a certain concentration range.When the concentration of ME-X was 20 ug/mL,the inhibitory effect on tumor cells was the best.The inhibition rates were 40.31%(MFC),33.81%(CT26.WT)and 21.19%(S180),respectively.(5)The results showed that ME-X could inhibit the growth in vivo of S180 tumors,with an inhibition rate of 50.32%.And it inhibited the growth of tumor cells through PI3K/AKT signaling pathway and Ras signaling pathway.These studies could not only provide a scientific basis for immune regulation mechanism of ME-X,but also provide theoretical support for ME-X in anti-cancer research.