Effect of lymphocyte activation gene 3(LAG3)on immune function of CD8+T cells in a mouse model of Echinococcus multilocularis infection
Objective To investigate the regulatory role of lymphocyte activation gene 3(LAG3)on the im-mune function of CD8+T cells in mice chronically infected with Echinococcus multilocularis(E.multilocularis).Methods 10 C57BL/6 wild-type(WT)and LAG3 knock-out(KO)mice were used,with each mouse receiving an intraportal injection of 3 000 protoscoleces of E.multilocularis to establish an infection model.12 weeks post-infection,liver and spleen tissues were collected from both groups.He-matoxylin-eosin(HE)staining was performed to observe inflammatory cell infiltration and pathological changes around liver lesions,while immunohistochemistry was utilized to assess the ratio of CD8+T cells in the"inflammatory microenvironment"surrounding liver lesions and within the spleen.Liver and spleen lymphocytes were isolated from both groups,and flow cytometry was employed to identify different CD8+T cell subsets,measuring the proportions and absolute numbers of CD8+T cells,effector memory CD8+T cells(CD8+Tem),central memory CD8+T cells(CD8+Tcm),and naive CD8+T cells(CD8+Tn),as well as their secretion of cytokines interferon-gamma(IFN-γ),tumor necrosis factor-alpha(TNF-α),in-terleukin-10(IL-10)and interleukin-17A(IL-17A).Additionally,liver lymphocytes from both groups were stimulated ex vivo withE.multilocularis antigen,and flow cytometry was used to assess the ratios of IFN-γ,L-10 and IL-17A secreted by CD8+T cells.Results HE staining showed that LAG3 KO mice had significantly more inflammatory foci in their livers compared to WT mice(P＜0.05).Immunohisto-chemical analysis revealed that CD8+T cell recruitment around liver lesions was significantly higher in LAG3 KO mice than in WT mice(P＜0.05),while there was a non-significant trend toward reduced CD8+T cell recruitment in the spleens of LAG3 KO mice(P＞0.05).Flow cytometry results indicated that the proportion of effector memory CD8+T cells in the livers of LAG3 KO mice showed a non-signifi-cant increasing trend compared to WT mice(P＞0.05);however,the proportion of CD8+T cells in the spleen weredecreased,with a significantly increased proportion of CD8+Tem phenotype(P＜0.01).Both liver and spleen CD8+T cells from LAG3 KO mice displayed enhanced TNF-α and IL-10 secretion abili-ties,with statistically significant differences(P＜0.05).In the ex vivo stimulation experiments using E.multilocularis antigens,CD8+T cells from LAG3 KO mice secreted significantly higher levels of IFN-γand IL-10 compared to WT mice(P＜0.05),and there was a non-significant trend toward increased IL-17A secretion(P＞0.05).Conclusion In chronic E.multilocularis infection in mice,LAG3 suppresses the im-mune response capability of CD8+T cells in both liver and spleen,modulating the inflammatory reaction.
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