Mechanism And Effect Of Sulfated Modification On The Immunological Activity Of Mesona Chinensis Benth Polysaccharide

Mesona chinensis Benth polysaccharide(MP)is one of the main active substances of Mesona chinensis Benth,and it is a medicine and food resource with various biological activities such as antidiabetics,antioxidant,and anticancer activities.Due to the structural difference of polysaccharides,it is possible to limit their biological activity.Therefore,the introduction of specific chemical groups through chemical modification can specifically improve the biological activity of polysaccharides to a certain extent.In this paper,the MP was extracted and purified from Mesona chinensis Benth.The chlorosulfonic acid-pyridine method was used to modify the MP to obtain a sulfated derivative named SMP.The main physical and chemical properties of polysaccharides were analyzed by measuring their sugar content,uronic acid content,protein content,monosaccharide composition,molecular weight(Mw),degree of substitution(DS),and surface morphology.By establishing the model of scavenging free radicals in vitro and oxidative damage to RAW264.7 cells induced by H2O2,the antioxidant activity of MP and SMP in vitro was evaluated.Meanwhile,the immunoregulatory effects of MP and SMP on splenic lymphocytes and RAW264.7 cells in vitro were explored.In addition,the effects of MP and SMP on the immune function of immunosuppressed mice were investigated by establishing a cyclophosphamide(Cy)-induced mouse immunosuppressive model.The main results are as follows:1.Sulfated modification changed the sugar content,monosaccharide composition,Mw and apparent structure of MP.Infrared spectrum results showed that the sulfate group was successfully incorporated into the structure of MP,and the DS of SMP was 0.52.MP is a homogeneous polysaccharide containing 39.01±0.25%neutral sugar,29.30±1.68%uronic acid and 27.52±0.61%protein,while SMP containing 31.49±0.13%neutral sugar,33.27±0.10%uronic acid and 20.85±0.15%protein.The Mw of MP increased from 157 kDa to 177 kDa after sulfated modification,and the monosaccharide composition was changed,but this change did not cause degradation of MP skeleton.Scanning electron microscopes(SEM)results showed that SMP has a more dispersed morphology than MP and contains some curled forms,which indicateed that the introduction of sulfate group makes the MP surface have more voids,and more conducive to exert their biological activities.2.Sulfated modification can increase the in vitro antioxidant activity of MP.By establishing a model of oxidative damage to RAW264.7 cells induced by H2O2,both MP and SMP can protect RAW264.7 cells from H2O2-induced damage by enhancing superoxidase dismutase(SOD)activity and reducing malondialdehyde(MDA)content.In vitro free radical scavenging experiments showed that SMP has better DPPH and hydroxyl radicals scavenging abilities than MP.3.Sulfated modification can enhance the therapeutic effect of MP on Cy-induced immunosuppressed mice.MP and SMP can restore body weight,organ index and abnormalities in routine blood indicators in Cy-induced immunosuppressed mice.MP and SMP can increase the proliferation activity of mice spleen T/B lymphocytes,and increase the secretion of interleukin(IL)-2,interleukin(IFN)-? and nitric oxide(NO)in spleen lymphocytes and spleen.While exerting immune function,MP and SMP can treat Cy-induced oxidative damage in mice by increasing the activity of total antioxidant capacity(T-AOC),glutathione peroxidase(GSH-Px),SOD and catalase(CAT),and reducing the content of MDA in spleen.MP can restore Cy-induced mouse immunosuppression by activating the phosphorylation levels of c-Jun N-terminal kinase(JNK),rabbit p38 kinase(p38)and rabbit extracellular regulated protein kinase(ERK)in the mitogen activated protein kinases signaling pathways(MAPKs)signaling pathway,but the signaling pathways through which SMP exerts its immunological activity still need further study.4.Sulfated modification can enhance the immunoregulatory capacity of MP on spleen lymphocytes and RAW264.7 cells.MP and SMP can synergize with ConA to promote T lymphocyte proliferation and further exert immune activity.The introduction of sulfate groups enhanced the regulatory effect of MP on the immune function of spleen lymphocytes.MP and SMP may promote the proliferation of RAW264.7 cells and the ability to secrete cytokines through TLR4 antibodies,and further exert immunoregulatory functions.