BACKGROUND Inflammatory bowel disease（IBD） is characterized by chronic and non-specific inflammation of the intestinal mucosa and mainly includes ulcerative colitis and Crohn’s disease.AIM To explore the beneficial...BACKGROUND Inflammatory bowel disease（IBD） is characterized by chronic and non-specific inflammation of the intestinal mucosa and mainly includes ulcerative colitis and Crohn’s disease.AIM To explore the beneficial effect of Toxo ROP16Ⅰ/Ⅲ-induced M2 phynotype macrophages in homeostasis of IBDs through downregulation of M1 inflammatory cells.METHODS RAW264.7 macrophages stimulated by lipopolysaccharide（LPS）（M1 cells） were co-cultured with Caco-2 cells as an inflammatory model of IBD in vitro.The expression of Toxo ROP16Ⅰ/Ⅲ was observed in RAW264.7 macrophages that were transfected with p EGFP-rop16Ⅰ/Ⅲ.The phenotypes of M2 and M1 macrophage cells were assessed by quantitative real-time reverse transcriptase polymerase chain reaction and the expression of tumor necrosis factor（TNF）-α,interleukin（IL）-1β,IL-6,transforming growth factor（TGF）-β1,IL-10,inducible nitric oxide synthase（i NOS）,and arginase-1（Arg-1） was detected.The expression of i NOS,Arg-1,signal transducer and activator of transcription 3（Stat3）,p-Stat3,Stat6,pStat6,programmed death ligand-2（PD-L2）,caspase-3,-8,and-9 was analyzed by Western blotting,and Griess assays were performed to detect nitric oxide（NO）.TNF-α,IL-1β,IL-6,TGF-β1,and IL-10 expression in the supernatants was detected by enzyme-linked immunosorbent assay,and Caco-2 cell apoptosis was determined by flow cytometry after mixing M1 cells with M2 cells in a Caco-2 cell co-culture system.RESULTS M1 cells exhibited significantly increased production of i NOS,NO,TNF-α,IL-1β,and IL-6,while Toxo ROP16Ⅰ/Ⅲ induced macrophage bias to M2 cells in vitro,showing increased expression of Arg-1,IL-10 and TGF-β1 and elevated production of p-Stat3 and p-Stat6.The mixed M1 and M2 cell culture induced by Toxo ROP16 Ⅰ/Ⅲ exhibited decreased production of NO and i NOS and upregulated expression of Arg-1 and PD-L2.Accordingly,Caco-2 cells became apoptotic,and apoptosis-associated proteins such as caspase-3,-8 and-9 were dampened during co-cu展开更多
Recent studies indicated that type II Toxoplasma gondii(Tg)GRA15II favored the generation of classically activated macrophages(M1),whereas type I/III TgROP16I/III promoted the polarization of alternatively activated m...Recent studies indicated that type II Toxoplasma gondii(Tg)GRA15II favored the generation of classically activated macrophages(M1),whereas type I/III TgROP16I/III promoted the polarization of alternatively activated macrophages(M2).A number of studies have demonstrated that M2 cells are involved in the pathogenesis of the liver fibrogenesis caused by Schistosoma japonicum.The purpose of the present study was to explore the inhibitory effect of Toxoplasma-derived TgGRA15II on mouse hepatic fibrosis with schistosomiasis.The gra15II and rop16I/III genes were amplified from strains T.gondii PRU and Chinese 1 Wh3,respectively.Lentiviral vectors containing the gra15II or rop16I/III plasmid were constructed and used to infect the RAW264.7 cell line.The polarization of the transfected cells was evaluated,followed by co-culture of the biased macrophages with mouse hepatic stellate JS1 cells.Then,mice were injected with GRA15II-driven macrophages via the tail vein and infected with S.japonicum cercariae.TgGRA15II induced a M1-biased response,whereas TgROP16I/III drove the macrophages to a M2-like phenotype.The in vitro experiments indicated that JS1 cell proliferation and collagen synthesis were decreased following co-culture with TgGRA15II-activated macrophages.Furthermore,mice inoculated with TgGRA15II-biased macrophages displayed a notable alleviation of collagen deposition and granuloma formation in their liver tissues.Our results suggest that TgGRA15II-induced M1 cells may dampen the M2 dominant pathogenesis of hepatic fibrosis and granulomatosis.These results provide insights into the use of parasite-derived immunomodulators as potential anti-fibrosis agents and to re-balance the schistosomiasis-induced immune response.展开更多
基金Supported by the National Natural Science Foundation of China,No.81471983the Key Research and Development Plan Project of Anhui Province,Department of Science and Technology 2019,No.201904a07020043+1 种基金the Key Project of Natural Science Research in the Universities of Anhui Provence,No.KJ2017A202the Research Fund Project of Anhui Institute of Transforming Medicine,No.2017zhyx04
文摘BACKGROUND Inflammatory bowel disease（IBD） is characterized by chronic and non-specific inflammation of the intestinal mucosa and mainly includes ulcerative colitis and Crohn’s disease.AIM To explore the beneficial effect of Toxo ROP16Ⅰ/Ⅲ-induced M2 phynotype macrophages in homeostasis of IBDs through downregulation of M1 inflammatory cells.METHODS RAW264.7 macrophages stimulated by lipopolysaccharide（LPS）（M1 cells） were co-cultured with Caco-2 cells as an inflammatory model of IBD in vitro.The expression of Toxo ROP16Ⅰ/Ⅲ was observed in RAW264.7 macrophages that were transfected with p EGFP-rop16Ⅰ/Ⅲ.The phenotypes of M2 and M1 macrophage cells were assessed by quantitative real-time reverse transcriptase polymerase chain reaction and the expression of tumor necrosis factor（TNF）-α,interleukin（IL）-1β,IL-6,transforming growth factor（TGF）-β1,IL-10,inducible nitric oxide synthase（i NOS）,and arginase-1（Arg-1） was detected.The expression of i NOS,Arg-1,signal transducer and activator of transcription 3（Stat3）,p-Stat3,Stat6,pStat6,programmed death ligand-2（PD-L2）,caspase-3,-8,and-9 was analyzed by Western blotting,and Griess assays were performed to detect nitric oxide（NO）.TNF-α,IL-1β,IL-6,TGF-β1,and IL-10 expression in the supernatants was detected by enzyme-linked immunosorbent assay,and Caco-2 cell apoptosis was determined by flow cytometry after mixing M1 cells with M2 cells in a Caco-2 cell co-culture system.RESULTS M1 cells exhibited significantly increased production of i NOS,NO,TNF-α,IL-1β,and IL-6,while Toxo ROP16Ⅰ/Ⅲ induced macrophage bias to M2 cells in vitro,showing increased expression of Arg-1,IL-10 and TGF-β1 and elevated production of p-Stat3 and p-Stat6.The mixed M1 and M2 cell culture induced by Toxo ROP16 Ⅰ/Ⅲ exhibited decreased production of NO and i NOS and upregulated expression of Arg-1 and PD-L2.Accordingly,Caco-2 cells became apoptotic,and apoptosis-associated proteins such as caspase-3,-8 and-9 were dampened during co-cu
基金We thank Dr Jinsheng Guo at Shanghai Zhongshan Hospital for providing the murine hepatic stellate cell line JS1The work was funded by the National Science Foundation of China(No.81471983 and No.81171606)+1 种基金the National Basic Research Program of China(No.2010CB530001)the Science Foundation of Anhui Province(No.KJ2014A106 and No.1308085MH124).
文摘Recent studies indicated that type II Toxoplasma gondii(Tg)GRA15II favored the generation of classically activated macrophages(M1),whereas type I/III TgROP16I/III promoted the polarization of alternatively activated macrophages(M2).A number of studies have demonstrated that M2 cells are involved in the pathogenesis of the liver fibrogenesis caused by Schistosoma japonicum.The purpose of the present study was to explore the inhibitory effect of Toxoplasma-derived TgGRA15II on mouse hepatic fibrosis with schistosomiasis.The gra15II and rop16I/III genes were amplified from strains T.gondii PRU and Chinese 1 Wh3,respectively.Lentiviral vectors containing the gra15II or rop16I/III plasmid were constructed and used to infect the RAW264.7 cell line.The polarization of the transfected cells was evaluated,followed by co-culture of the biased macrophages with mouse hepatic stellate JS1 cells.Then,mice were injected with GRA15II-driven macrophages via the tail vein and infected with S.japonicum cercariae.TgGRA15II induced a M1-biased response,whereas TgROP16I/III drove the macrophages to a M2-like phenotype.The in vitro experiments indicated that JS1 cell proliferation and collagen synthesis were decreased following co-culture with TgGRA15II-activated macrophages.Furthermore,mice inoculated with TgGRA15II-biased macrophages displayed a notable alleviation of collagen deposition and granuloma formation in their liver tissues.Our results suggest that TgGRA15II-induced M1 cells may dampen the M2 dominant pathogenesis of hepatic fibrosis and granulomatosis.These results provide insights into the use of parasite-derived immunomodulators as potential anti-fibrosis agents and to re-balance the schistosomiasis-induced immune response.
