| Human plasmin is an important enzyme present in blood that degrades many blood plasma proteins,and it plays a role in many normal and pathological conditions in the body.It has been shown that plasmin can not only degrade fibrin,but also directly act on leukocytes and participate in immune regulation.It has been proved that human plasmin and human plasminogen can affect macrophage polarization and promote neutrophil apoptosis and efferocytosis.However,natural plasmin and pasminogen originally came from limited sources and could only be purified and isolated from human plasma.Because of its large molecular mass and complex molecular structure,it is difficult to construct and gene express.However,the human plasminogen serine protease Domain(Plg-SP)can retain the whole human plasminogen activity after being stimulated,and its molecular mass is smaller and its structure is relatively simple.Therefore,in this study,the human plasminogen serine protease domain was constructed and expressed in Pichia pastoris,and its effect on macrophage polarization was preliminarily explored.In this study,the human plasmin Proserine protease domain was constructed and expressed using X33 constitutive Pichia pastoris strain by genetic engineering.The purity of 97% protein samples was obtained after being purified by affinity chromatography.The samples were identified as constructed proteins with a concentration of 0.53 mg/ml,which provided protein samples to meet the needs of subsequent research.In this study,the effects of the human plasminogen serine protease domain on the polarization of macrophages from two sources were studied.It was found that after stimulation with Plg-SP protein,the gene expression levels of Arg-1 and CD206,the surface markers of M2 macrophages in RAW264.7 and MV-4-11 increased significantly,while the expression level of iNOS,the surface marker of M1 macrophages,decreased significantly.Preliminary studies have shown that Plg-SP can affect the polarization of macrophages toward M2. |
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