An Immunosuppressive Polypeptide From Myotis Btandtii To Treat Mouse Psoriasis

LTA4H and LTB4 play important roles in the inflammatory response and immune process.It may cause serious inflammatory reactions and even mediate autoimmune diseases if LTB4 content exceeds the threshold.LTB4 can participate in the inflammatory process by affecting NF-κB,MAPK and JAK-STAT pathways,at the same time,it can participate in the activation and differentiation of Th 17 cell or induce the release of various factors such as IL-6 and TNF-α.The experiment proved that the plasma LTA4H content of psoriasis patients was significantly higher than that of normal people,indicating that LTA4H and LTB4 were involved in the pathogenesis of psoriasis.LTB4 and LTA4H may become new targets for the treatment of psoriasis since inflammatory response and psoriasis can be inhibitted by inhibiting theirs activity.Variety of active polypeptide have been authenticated in the saliva of blood-sucking bats.These active polypeptides have many functions such as analgesia,anticoagulation,and anti-inflammatory,that are helpful for blood-sucking.We find a 117-residue peptide named MTX from the saliva of B.cerevisiae,which have serine protease inhibitory activity and anti-inflammatory activity in vitro.It’s a Kazal type protein.We acquired His-r-MTX by PET-32a expression system of BL21(DE3).Then we purified the His-r-MTX by Ni-NTA column and formic acid cutting.We got the His-r-MTX with Trypsin inhibitory activity through RP-HPLC.The molecular weight of MTX was 13 KDa by MALDI-TOF-MS,about 100 Da more than the theoretical molecular weight because formic acid cutting will leave a proline.R-MTX has twelve cysteines,which have formed six pairs of disulfide bond,enduing high stability of r-MTX with a half-time of 16 h in plasma.We performed a safety test and results showed that MTX has no cytotoxicity and hemolytic activity,so,the potential use in clinical practice may be not influenced.MTX itself is a serine protease inhibitor,but at the same time it binds to LTA4H and inhibits its epoxide hydrolase activity,which is an important factor in the production of LTB4.Therefore,we judge that MTX may be a potential immunosuppr-essive agent,and its LTA4H inhibitory activity may be used in the autoimmune diseases’ treatment such as psoriasis.We injected MTX with imiquimod mice and rusults showed that MTX can reduce the levels of LTB4,inflammatory factors and the phosphorylation level of inflammatory signaling pathway,at the same time,it can alleviate the inflammatory symptoms of mouse skin.The lesion status was significantly reduced in the 5 mg/kg group compared with the model group.In summary,MTX may become a new potential drug for psoriasis.It has obvious anti-inflammatory effects,no cytotoxicity and hemolytic activity,and can achieve mass production through prokaryotic expression technology,in line with industrial production requirements.