| Malaria remains a serious infectious disease caused by Plasmodium parasites,and poses a serious threat to human health and safety.In 2019,a global number of 229 million new cases of malaria and 409000 deaths were reported,involving 87 countries and regions around the world.Malaria caused by Plasmodium vivax infection was defined as vivax malaria.In recent years,severe cases of severe anemia,liver insufficiency,acute lung injury,acute renal failure,and hypersplenism caused by vivax malaria have continued to appear.The mechanism is not yet clear,and it is urgent to strengthen research on it.The interaction between the Plasmodium parasite and the host is an important cause of malaria,in which how to escape the monitoring and clearance of host immune system is a key problem.A hypothesis of the immune evasion of P.vivax is that after entering the spleen,the parasites could interact with spleen fibroblasts and change the morphology or function of fibroblasts,so that the parasites could avoid the killing of antibodies and immune cells under the cover of spleen fibroblasts.It implies that human spleen fibroblasts(HSF)play an important role in the immune regulation of the spleen.Infection of erythrocytes by the P.vivax results in the export of several parasite proteins onto the erythrocyte membrane surface establishing novel interactions between human spleen fibroblasts and parasite proteins,which changes the structure and function of fibroblasts,thus protecting the malaria parasites from been recognized and cleared by human spleen immune system..However,how the export protein interacts with HSF cells and how to change its structure and function have not been clarified.P.vivax tryptophan rich antigens(Pv TRAgs)protein family,which being exported to the erythrocyte membrane surface after parasites infection,have high immunogenicity and antigenicity,and can effectively recognize serum of vivax malaria patients.On the other hand,67 spleen-related genes were identified by full transcription analysis of intact spleen and splenectomized monkey infected by P.vivax,including 5 members of Pv TRAgs protein family.These genes were highly expressed in the spleen of monkeys infected with P.vivax,but these genes could not be detected after splenectomy,suggesting that they play an important role in the immune regulation of spleen.Moreover,HSF cells located in the spleen can bind to the infected red blood cells and protect them from the killing and elimination of the spleen immune system.So in our previous experiments,we have verified and screened the Pv TRAgs protein that can interact with HSF cells.Combined with the spleen-dependent proteins screened from the whole transcriptome analysis of P.vivax infected monkeys,this study targeted PvTRAg23 to clarify the interaction between Pv TRAgs and human spleen fibroblasts,and the changes in the structure and function of fibroblasts,so as to provide a theoretical basis for revealing the immune escape mechanism of P.vivax in human hosts.The main results are as follows:(1)Expression and purification of Pv TRAgs.The sequence of Pv TRAgs were found from the Plasm DB URL.The E.coli expression system was used for the protein expressions.(2)Subcellular localization of PvTRAg23 and PvTRAg26.Indirect immunofluorescence assay confirmed that PvTRAg23 and PvTRAg26 proteins were localized on the surface of erythrocyte membrane infected with P.vivax.(3)Interaction between PvTRAg23 and human spleen fibroblasts.The binding of PvTRAg23 to human spleen fibroblasts was verified by flow cytometry.A PvTRAg23 receptor molecule Vimentin on the surface of HSF cell membrane was screened by mass spectrometry.(4)Effects of PvTRAg23 on human spleen fibroblasts.PvTRAg23 could significantly promote the migration of human spleen fibroblasts.PvTRAg23 could inhibit the expression of collagen Ⅰ in human spleen fibroblasts.(5)The signal transduction mechanism of of PvTRAg23 down-regulating the expression of collagen Ⅰ in HSF cells.PvTRAg23 specifically reduces the expression and secretion of collagen Ⅰ in HSF cells by activating NF-κB p65 signal pathway,and the increase of IL-1β,IL-6 and TNF-α cytokines in HSF cells induced by PvTRAg23 plays a positive feedback role in the decrease of collagen Ⅰ.To sum up,this study provides a reference value for perfecting the immune escape mechanism of P.vivax in human by analyzing the structural and functional changes of HSF cells under the stimulation of PvTRAg23,and provides a scientific theoretical basis for the key role of HSF cells and P.vivax export proteins,especially Pv TRAgs protein family,in the process of malaria parasites immune escape. |
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