Differential Expression And The Biological Effect Of TIM-3 On Immune Cells Of Mice Infected With Toxoplasma Gondii Or Plasmodium Parasites

T-cell immunoglobulin and mucin domain 3(TIM-3),which is an immunoglobulin(Ig)and mucin domain family cell-surface molecule that is often referred to as a checkpoint receptor and exhaustion marker,it plays an important role in the immune escape process of tumor and some parasites such as Schistosoma japonicum and Plasmodium.In the process of parasite infection,the immune state of the body is closely related to the virulence of the infected parasite,some studies also associated the virulence of parasites with its ability to active the immune system by different mechanisms.Previous studies of our group showed that the expression of TIM-3 on the surface of key immune cells of mice increased during Schistosoma japonicum infection and induced Th2 biased immune response of the host;our study also found that the expression of TIM-3 on the surface of key immune cells also increased in patients infected with Plasmodium falciparum and in mice infected with Plasmodium berghei.However,the differences of hosts’ immune response and expression level of TIM-3 on the surface of host immune cells induced by different virulent parasites are still unclear,further,the biological effect of Tim-3 in parasite-infected mice remains to be further studied.Thus,in this study,based on our previous research,two kinds of parasites,Plasmodium and Toxoplasma gondii,two kinds of Intracellular parasitism apicomplexan protozoa were selected,the TIM-3 inducing abilities in host’s immune responses of different virulent strains and its significance in disease resistance and pathogenesis were studied.Plasmodium mainly parasitized in erythrocytes and Toxoplasma gondii mainly parasitized in karyocytes.They would trigger an array of signals and responses in both the innate and adaptive immune systems of the host to control the parasite,which can prevent parasite infection under certain circumstances.Plasmodium infection often causes periodic cold and heat attack,accompanied by headache,nausea and other symptoms,which can be lifethreatening.Different from Plasmodium,T.gondii will gradually transform from tachyzoites to bradyzoites in hosts with normal immune system,then forming cysts to escape the killing of the immune system,and eventually parasitize in the host for a long time,even with its whole life.Only when the host immune system is low or damaged can it cause fatal damage.These may be related to host’s immune exhaustion caused by TIM-3 expression.Previous studies show that the anti-Parasite immune response and the vaccines and drugs designed based on hosts’ immune response can effectively control the infection of parasite,and even prevent the parasite infection in some cases.Thus,our research could provide a theoretical basis for the selection of antimalarial drug targets,the treatment of chronic Toxoplasmosis and revealing the long-term parasitic mechanism of Toxoplasma gondii in the host.Firstly,TIM-3 expression on critical lymphocyte populations and the proportion of these cells were detected by flow cytometry.In P.berghei infection experiment: The expression levels of TIM-3 on splenic and circulatory T cells in P.b ANKA infected mice were the higher than that in P.b NK65 or ΔP.b Tat D infected mice.Meanwhile,increased TIM-3 expression on splenic and circulatory T cells was associated with a reduction in the proportion of these cells.In T.gondii infection experiment: TIM-3 expression on the splenic and circulatory T cells of mice infected with T.gondii(RH strain)was higher than that in mice infected with T.gondii(ME49 strain).Meanwhile,T.gondii infection reduced the proportion of splenic T cells and increased TIM-3 expression.Further,blocking TIM-3 signaling using anti-TIM-3 antibodies in vitro inhibited early apoptosis of lymphocytes.Thus,higher level of Tim-3 was induced by more virulent parasites and TIM-3 on lymphocytes negatively regulates cell-mediated immunity against parasite infection.In order to analysis the levels of cytokines in the sera of mice infected with parasites of different virulence,cytometric bead array(CBA)technique was chosen.The results showed that in Plasmodium infected mice,interleukin(IL)-2,IL-4,IL-6,IL-22,interferon(IFN)-γ,and tumor necrosis factor alpha(TNF-α)levels increased to their highest levels and these cytokines,apart from IL-22,were the most secreted by the P.b NK65 group at 4 or 7 days post infection.In T.gondii infected mice,interleukin(IL)-2,interferon γ,tumor necrosis factor(TNF)-α,IL-12p70,IL-22,IL-17 A,and IL-5 increased significantly after infection.Mice infected with T.gondii(ME49 strain)showed elicited responses of TNF-α,IL-17 A,IL-12p70 and IL-22 than that infected by the RH strain.That is to say,the secretion level of some cytokines in the sera of mice infected with more virulent parasite is higher.In order to further study the biological effect of Tim-3,in vivo TIM-3 / Gal-9 signaling blockade assays using α-lactose(an antagonist of Gal-9)were conducted.Results showed that in Plasmodium infected mice,α-lactose did not significantly protect the mice but induced the expression of another immunosuppressive molecule,T cell immunoreceptor with Ig and ITIM domains(TIGIT).In T.gondii infected mice,α-lactose also has no significant protective effect,which may be related to the irreversible lymphocyte exhaustion in mice.In the immunotherapy progress of tumor,single-target treatment also cannot achieve better results,so the above results suggest that in the treatment of parasites,the multi-target treatment may achieve better results.