ABSTRACT
Objective:To investigate the phenotype and function of the intestinal γδT lymphocytes in post-infectious irritable bowel syndrome mouse model. Methods:The mouse model for post-infectious irritable bowel syndrome was established by the infection with trichinella spiralis. The intestinal inflammation,abdominal withdrawal reflex( AWR) and colon transportation test were observed. 2 and 8 weeks later,the animals were sacrificed and the lymphocytes in the intestinal lymph nodes and spleen were collected,from which the γδT lymphocytes were isolated and purified by monoclonal antibody-immuno-microbeads method. The functions of the purified γδT lymphocytes were evaluated,including proliferation by 3 HTdR;CD69,CD62L molecule staining by flow cytometry. Furthermore,the con-centration of cytokine IL-17 and IFN-γ in the supernatant of the cultured γδT lymphocytes were detected by ELISA. Results: At 2nd weeks after infection,significant intestinal inflammation was observed,with increasingγδT lymphocytes,proliferating and activating with increasing production of IL-17. At 8th weeks after infection, the intestinal inflammation disappeared, whereas the number of γδT lymphocytes remained increasing,also with proliferating and activating with increasing production of IL-17. Meanwhile,the mice show higher AWR score and Bristol score. Conclusion: γδT lymphocytes could participate in the pathogenesis of PI-IBS via their proliferation,activation and production of IL-17.
ABSTRACT
Background C57BL/6andB10R Ⅲareroutinemurinespeciesusedinexperimental autoimmune uveitis (EAU).The inflammation is light for mouse after immunization whereas it is prominent for B10R Ⅲ.ObjectiveThis study was to observe the killing effect of interphotoreceptor retinoid binding protein (IRBP) 1-20-specific T cells on mouse retinal astrocyte.Th1 and Th17 cells effect in the EAU mechanism was discussed.MethodsB10RllⅢ mice and C57BL/6 mice were immunized with IRBP 161-180 and IRBP 1-20 in complete Freund adjuvant (CFA).The infiltrating cells of diseased B10R Ⅲ eyes were analyzed by flow cytometry.IRBP 1-20-specific T cells were isolated from the drainage lymph node and spleen and cultured in IL-2 or IL-23 for Th1 and Th17 cells polarization,respectively.Th1 and Th17 cells cultured for 5 days were seeded on the mouse retinal astrocyte monolayer pretreated with gamma interferon.Cell interaction was observed and the quantity of TNF-α was tested by ELISA.Every test was repeated 6 times and the mean was calculated.The maintenance of experimental animals complied with the Statement of ARVO.ResultsThere were lots of infiltrating cells in the eyes of B10Rm mice after immunization,including 9.5％ IFNγ+ cells,5.1％ IL-17+cells and 41.4％ CD45+ cells.Six days after IRBP1-20 stimulation and cultured by IL-2 and IL-23,44.0％ and 8.0％ cells were IFNγ+,and 1.0％ and 26.0％ cells were IL17+.Twentyfour hours after the interaction between Th1 or Th17 and retinal astrocyte,retinal astrocyte died and detached.The killing effect of Th17 was stronger than Th1.48 hours after co-culture of Th1 or Th17T cells with astrocytes,the concentrations of TNF-α were ( 500± 10 ) and ( 801 ±24 μg/L) μg/L,respectively,with a significant statistical difference (t =-20.36,P =0.00).ConclusionsBoth Th1 and Th17 can kill retinal astrocyte,but Th17 plays a key role in the EAU pathogenesis process.The killing effect is caused by intercellular contact and interaction under the induction of cytokines.