Effect of LAG3 deficiency on natural killer cell function and hepatic fibrosis in mice infected with Echinococcus multilocularis
Objective To investigate the effect of LAG-3 deficiency(LAG3-/-)on natural killer(NK)cell function and hepatic fibrosis in mice infected with Echinococcus multilocularis.Methods C57BL/6 mice,each weighing(20±2)g,were divided into the LAG3-/-and wild type(WT)groups,and each mouse in both groups was inoculated with 3 000 E.multilocularis protoscoleces via the hepatic portal vein.Mouse liver and spleen specimens were collected 12 weeks post-infection,sectioned and stained with sirius red,and the hepatic lesions and fibrosis were observed.Mouse hepatic and splenic lymphocytes were isolated,and flow cy-tometry was performed to detect the proportions of hepatic and splenic NK cells,the expression of CD44,CD25 and CD69 mole-cules on NK cell surface,and the secretion of interferon γ(IFN-γ),tumor necrosis factor α(TNF-α),interleukin(IL)-4,IL-10 and IL-17A.Results Sirius red staining showed widening of inflammatory cell bands and hyperplasia of fibrotic connective tissues around mouse hepatic lesions,as well as increased deposition of collagen fibers in the LAG3-/-group relative to the WT group.Flow cytometry revealed lower proportions of mouse hepatic(6.29%±1.06%vs.11.91%±1.85%,P<0.000 1)and splenic NK cells(4.44%±1.22%vs.5.85%±1.10%,P>0.05)in the LAG3-/-group than in the WT group,and the mean fluorescence inten-sity of CD44 was higher on the surface of mouse hepatic NK cells in the LAG3-/-group than in the WT group(t =-3.234,P<0.01),while no significant differences were found in the mean fluorescence intensity of CD25 or CD69 on the surface of mouse he-patic NK cells between the LAG3-/-and WT groups(both P values>0.05).There were significant differences between the LAG3-/-and WT groups in terms of the percentages of IFN-γ(t =-0.723,P>0.05),TNF-α(t =-0.659,P>0.05),IL-4(t =-0.263,P>0.05),IL-10(t =-0.455,P>0.05)or IL-17A secreted by mouse hepatic NK cells(t = 0.091,P>0.05),and the percentage of IFN-γ secreted by mouse splenic NK cells was higher in the LAG3-/-group than in the WT group(58.40%±1.64%vs.50.40%±4.13%;t =-4.042,P<0.01);however,there were no significant differences between the two groups in terms of the proportions of TNF-α(t =-1.902,P>0.05),IL-4(t =-1.333,P>0.05),IL-10(t =-1.356,P>0.05)or IL-17A secreted by mouse splenic NK cells(t = 0.529,P>0.05).Conclusions During the course of E.multilocularis infections,LAG3-/-promotes high-level secretion of IFN-γ by splenic NK cells,which may participate in the reversal the immune function of NK cells,resulting in aggravation of hepatic fibrosis.
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