RESUMEN
Although the functions of programmed death-1 (PD-1) on αß T cells have been extensively reported, a role for PD-1 in regulating γδT cell function is only beginning to emerge. Here, we investigated the phenotypic and functional characteristics of PD-1-expressing γδT cells, and the molecular mechanism was also explored in the Plasmodium yoelii nigeriensis (P. yoelii NSM)-infected mice. Flow cytometry and single-cell RNA sequencing (scRNA-seq) were performed. An inverse agonist of RORα, SR3335, was used to investigate the role of RORα in regulating PD-1+ γδT cells. The results indicated that γδT cells continuously upregulated PD-1 expression during the infection period. Higher levels of CD94, IL-10, CX3CR1, and CD107a; and lower levels of CD25, CD69, and CD127 were found in PD-1+ γδT cells from infected mice than in PD-1- γδT cells. Furthermore, GO enrichment analysis revealed that the marker genes in PD-1+ γδT cells were involved in autophagy and processes utilizing autophagic mechanisms. ScRNA-seq results showed that RORα was increased significantly in PD-1+ γδT cells. GSEA identified that RORα was mainly involved in the regulation of I-kappaB kinase/NF-κB signaling and the positive regulation of cytokine production. Consistent with this, PD-1-expressing γδT cells upregulated RORα following Plasmodium yoelii infection. Additionally, in vitro studies revealed that higher levels of p-p65 were found in PD-1+ γδT cells after treatment with a RORα selective synthetic inhibitor. Collectively, these data suggest that RORα-mediated attenuation of NF-κB signaling may be fundamental for PD-1-expressing γδT cells to modulate host immune responses in the spleen of Plasmodium yoelii nigeriensis-infected C57BL/6 mice, and it requires further investigation.
Asunto(s)
Malaria , Plasmodium yoelii , Receptor de Muerte Celular Programada 1 , Bazo , Animales , Plasmodium yoelii/inmunología , Receptor de Muerte Celular Programada 1/metabolismo , Malaria/inmunología , Malaria/parasitología , Ratones , Bazo/inmunología , Bazo/parasitología , Femenino , Transducción de Señal/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Receptores de Antígenos de Linfocitos T gamma-delta/inmunologíaRESUMEN
BACKGROUND: Despite the functions of TLRs in the parasitic infections have been extensively reported, few studies have addressed the role of TLR3 in the immune response to Schistosoma japonicum infections. The aim of this study was to investigate the properties of TLR3 in the liver of C57BL/6 mice infected by S. japonicum. METHODS: The production of TLR3+ cells in CD4+T cells (CD4+CD3+), CD8+T cells (CD8+CD3+), γδT cells (γδTCR+CD3+), NKT cells (NK1.1+CD3+), B cells (CD19+CD3-), NK (NK1.1-CD3+) cells, MDSC (CD11b+Gr1+), macrophages (CD11b+F4/80+), DCs (CD11c+CD11b+) and neutrophils (CD11b+ Ly6g+) were assessed by flow cytometry. Sections of the liver were examined by haematoxylin and eosin staining in order to measure the area of granulomas. Hematological parameters including white blood cell (WBC), red blood cell (RBC), platelet (PLT) and hemoglobin (HGB) were analyzed. The levels of ALT and AST in the serum were measured using biochemical kits. The relative titers of anti-SEA IgG and anti-SEA IgM in the serum were measured by enzyme-linked immunosorbent assay (ELISA). CD25, CD69, CD314 and CD94 molecules were detected by flow cytometry. RESULTS: Flow cytometry results showed that the expression of TLR3 increased significantly after S. japonicum infection (P < 0.05). Hepatic myeloid and lymphoid cells could express TLR3, and the percentages of TLR3-expressing MDSC, macrophages and neutrophils were increased after infection. Knocking out TLR3 ameliorated the damage and decreased infiltration of inflammatory cells in infected C57BL/6 mouse livers.,The number of WBC was significantly reduced in TLR3 KO-infected mice compared to WT-infected mice (P < 0.01), but the levels of RBC, platelet and HGB were significantly increased in KO infected mice. Moreover, the relative titers of anti-SEA IgG and anti-SEA IgM in the serum of infected KO mice were statistically decreased compared with the infected WT mice. We also compared the activation-associated molecules expression between S.japonicum-infected WT and TLR3 KO mice. CONCLUSIONS: Taken together, our data indicated that TLR3 played potential roles in the context of S. japonicum infection and it may accelerate the progression of S. japonicum-associated liver pathology.
Asunto(s)
Schistosoma japonicum , Animales , Ratones , Schistosoma japonicum/metabolismo , Receptor Toll-Like 3/metabolismo , Ratones Endogámicos C57BL , Inmunoglobulina G , Inmunoglobulina MRESUMEN
CD103 is an important marker of tissue-resident memory T cells (TRM) which play important roles in fighting against infection. However, the immunological characteristics of CD103+ T cells are not thoroughly elucidated in the liver of mouse infected with Plasmodium. Six- to eight-week-old C57BL/6 mice were infected with Plasmodium yoelii nigeriensis NSM. Mice were sacrificed on 12-16 days after infection and the livers were picked out. Sections of the livers were stained, and serum aspartate aminotransferase (AST) and alanine transaminase (ALT) levels were measured. Moreover, lymphocytes in the liver were isolated, and the expression of CD103 was determined by using qPCR. The percentage of CD103 on different immune cell populations was dynamically observed by using flow cytometry (FCM). In addition, the phenotype and cytokine production characteristics of CD103+CD8+ Tc cell were analyzed by using flow cytometry, respectively. Erythrocyte stage plasmodium infection could result in severe hepatic damage, a widespread inflammatory response and the decrease of CD103 expression on hepatic immune cells. Only CD8+ Tc and γδT cells expressed higher levels of CD103 in the uninfected state.CD103 expression in CD8+ Tc cells significantly decreased after infection. Compared to that of CD103- CD8+ Tc cells, CD103+ CD8+ Tc cells from the infected mice expressed lower level of CD69, higher level of CD62L, and secreted more IL-4, IL-10, IL-17, and secreted less IFN-γ. CD103+CD8+ Tc cells might mediate the hepatic immune response by secreting IL-4, IL-10, and IL-17 except IFN-γ in the mice infected with the erythrocytic phase plasmodium, which could be involved in the pathogenesis of severe liver damage resulted from the erythrocytic phase plasmodium yoelii nigeriensis NSM infection.
Asunto(s)
Malaria , Plasmodium yoelii , Animales , Ratones , Linfocitos T CD8-positivos/metabolismo , Interleucina-10/metabolismo , Interleucina-17 , Interleucina-4 , Hígado , Malaria/inmunología , Malaria/metabolismo , Ratones Endogámicos C57BLRESUMEN
Splenomegaly is a prominent clinical manifestation of malaria and the causes remain incompletely clear. Anemia is induced in malaria and extramedullary splenic erythropoiesis is compensation for the loss of erythrocytes. However, the regulation of extramedullary splenic erythropoiesis in malaria is unknown. An inflammatory response could facilitate extramedullary splenic erythropoiesis in the settings of infection and inflammation. Here, when mice were infected with rodent parasites, Plasmodium yoelii NSM, TLR7 expression in splenocytes was increased. To explore the roles of TLR7 in splenic erythropoiesis, we infected wild-type and TLR7 -/- C57BL/6 mice with P. yoelii NSM and found that the development of splenic erythroid progenitor cells was impeded in TLR7 -/- mice. Contrarily, the treatment of the TLR7 agonist, R848, promoted extramedullary splenic erythropoiesis in wild-type infected mice, which highlights the implication of TLR7 on splenic erythropoiesis. Then, we found that TLR7 promoted the production of IFN-γ that could enhance phagocytosis of infected erythrocytes by RAW264.7. After phagocytosis of infected erythrocytes, the iron metabolism of RAW264.7 was upregulated, evidenced by higher iron content and expression of Hmox1 and Slc40a1. Additionally, the neutralization of IFN-γ impeded the extramedullary splenic erythropoiesis modestly and reduced the iron accumulation in the spleen of infected mice. In conclusion, TLR7 promoted extramedullary splenic erythropoiesis in P. yoelii NSM-infected mice. TLR7 enhanced the production of IFN-γ, and IFN-γ promoted phagocytosis of infected erythrocytes and the iron metabolism of macrophages in vitro, which may be related to the regulation of extramedullary splenic erythropoiesis by TLR7.
Asunto(s)
Malaria , Bazo , Ratones , Animales , Bazo/metabolismo , Eritropoyesis , Receptor Toll-Like 7 , Ratones Endogámicos C57BL , Interferón gamma/uso terapéutico , Macrófagos/metabolismo , Hierro/metabolismoRESUMEN
The immune system of vertebrates includes innate immunity and adaptive immunity, and the network between them enables the host to fight against invasions of various pathogens. Recently, studies discovered that immune memory is one of the features of innate immunity, breaking the previous opinion that immune memory exists only in adaptive immunity. Immune memory supports innate immune cells to respond efficiently upon reinfection or restimulation. During the Plasmodium infection, the innate immune system is the first to be triggered, and innate immune cells are activated by components from Plasmodium or Plasmodium-infected red blood cells. Innate immune cells could be induced to develop memory after the activation and may play an important role in the subsequent infection of Plasmodium or other pathogens and stimulation. This review will discuss the recent findings relevant to trained immunity and Plasmodium infection, facilitating the understanding of the role of trained immunity in malaria and other diseases and the development of therapeutic strategies based on trained immunity.
Asunto(s)
Malaria , Plasmodium , Animales , Inmunidad Entrenada , Inmunidad Adaptativa , Inmunidad Innata , Memoria InmunológicaRESUMEN
Recently, there is a paucity of studies focus on the characteristics of myeloid cells which expressed γδTCR. The aim of this study was to observe the properties of γδTCR-expressing myeloid cells in the spleen of C57BL/6 mice infected by P. yoelii nigeriensis NSM. Haematoxylin-eosin (HE) staining was used to observe pathological changes in the spleens from infected mice. The differentially expressed genes (DEGs) between the infection and control groups were analyzed by RNA sequencing (RNA -seq). Flow cytometry (FCM) was used to evaluate the frequency of γδTCR+ cells and the characteristics of γδTCR+ cells in P. yoelii nigeriensis NSM-infected mice. Obvious infiltration of inflammatory were observed in the spleens from infected C57BL/6 mouse. The proportions of γδTCR+ cells and CD11b+ γδTCR+ cells from infected group were higher than that from normal group. CD11b+ γδTCR+ cells expressed high levels of activated-mediated genes and inflammatory-mediated genes. The heterogeneous pathway activities among CD11b+ γδTCR+ cells from normal and infected group were characterized. The oxidative phosphorylation, respiratory electron transport chain and leukocyte activation involved in immune response pathways were up-regulated, while the alpha-beta T cell activation and myeloid leukocyte migration pathways were down-regulated in infected mice. Importantly, Ly6c2 was higher expressed in CD11b+ γδTCR+ cells than Ly6g. Consistent with it, flow cytometry results revealed that a subset of Ly6C+ cells was higher than Ly6G+ cells in the spleen. Taken together, our data suggest the existence of a population of γδTCR-expressing myeloid cells and they might be multifunctional cells, which play a role in couse of Plasmodium infection.
Asunto(s)
Malaria , Células Mieloides , Plasmodium yoelii , Receptores de Antígenos de Linfocitos T gamma-delta , Animales , Ratones , Citometría de Flujo , Ratones Endogámicos C57BL , Células Mieloides/metabolismo , Plasmodium yoelii/fisiologíaRESUMEN
The morbidity and mortality of malaria are still high. Programmed cell death-1(PD-1) is an important co-inhibitory factor and CD8 T cells with PD-1 were reported to be exhausted cells. It remains unknown what the role of CD4 T cells expressing PD-1 is and what the upstream regulating molecules of PD-1 in CD4 T cells are. The C57BL/6 mice were injected with Plasmodium yoelii (P. yoelii) in this study. Expressions of PD-1, activation markers, and cytokines were tested. The differentially expressed genes between PD-1+/- CD4 T cells were detected by microarray sequencing. Western blot, chromatin immunoprecipitation (ChIP), siRNA, hypoxia inducible factor-1α (HIF-1α) inducer and inhibitor were used to explore PD-1's upstream molecules, respectively. The proportions of PD-1+ CD4 T cells increased post P. yoelii infection. PD-1+ CD4 T cells expressed more activated surface markers and could produce more cytokines. Nuclear factor of activated T cells 1 (NFATc1) was found to be a key transcription factor to induce PD-1 expression after infection. Both the inducer and the inhibitor of HIF-1α could change the expressions of NFATc1 and PD-1 in vivo and in vitro, respectively. Taken together, P. yoelii infection induced NFATc1 expression by HIF-1α. The highly expressed NFATc1 entered the nucleus and initiated PD-1 expression. PD-1+ CD4 T cells appeared to be more activated and could secrete more cytokines to regulate the host's immune responses against malaria.
Asunto(s)
Linfocitos T CD4-Positivos , Subunidad alfa del Factor 1 Inducible por Hipoxia , Malaria , Factores de Transcripción NFATC , Plasmodium yoelii , Receptor de Muerte Celular Programada 1 , Animales , Linfocitos T CD4-Positivos/inmunología , Citocinas/inmunología , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/inmunología , Malaria/genética , Malaria/inmunología , Malaria/parasitología , Ratones , Ratones Endogámicos C57BL , Factores de Transcripción NFATC/genética , Factores de Transcripción NFATC/inmunología , Receptor de Muerte Celular Programada 1/genética , Receptor de Muerte Celular Programada 1/inmunología , Transducción de SeñalRESUMEN
S. japonicum infection can induce granulomatous inflammation in the liver of the host. Granulomatous inflammation limits the spread of infection and plays a role in host protection. Toll-like receptor 7 (TLR7) is an endosomal TLR that recognizes single-stranded RNA (ssRNA). In this study, the role of TLR7 in S. japonicum infection-induced hepatitis was investigated in both normal and TLR7 knockout (KO) C57BL/6 mice. The results indicated that TLR7 KO could aggravate S. japonicum infection-induced damage in the body, with less granuloma formation in the tissue, lower WBCs in blood, and decreased ALT and AST in the serum. Then, the expression of TLR7 was detected in isolated hepatic lymphocytes. The results indicated that the percentage of TLR7+ cells was increased in the infected mice. Hepatic macrophages, DCs, and B cells could express TLR7, and most of the TLR7-expressing cells in the liver of infected mice were macrophages. The percentage of TLR7-expressing macrophages was also increased after infection. Moreover, macrophages, T cells, and B cells showed significant changes in the counts, activation-associated molecule expression, and cytokine secretion between S. japonicum-infected WT and TLR7 KO mice. Altogether, this study indicated that TLR7 could delay the progression of S. japonicum infection-induced hepatitis mainly through macrophages. DCs, B cells, and T cells were involved in the TLR7-mediated immune response.
Asunto(s)
Hígado/parasitología , Esquistosomiasis Japónica , Receptor Toll-Like 7 , Animales , Glicoproteínas de Membrana , Ratones , Ratones Endogámicos C57BL , Schistosoma japonicum , Esquistosomiasis Japónica/inmunologíaRESUMEN
Despite the paramount role of TLRs in the induction of innate immune and inflammatory responses, there is a paucity of studies on the role of TLRs in Schistosoma japonicum infection. Here, we observed obvious infiltration of inflammatory cells in S. japonicum-infected C57BL/6 mouse lungs. Expression and release of IFN-γ, IL-4, and IL-17 were significantly higher in pulmonary lymphocytes from infected mice compared with control mice in response to anti-CD3 plus anti-CD28 mAbs. Higher percentages of TLR2, TLR3, TLR4, and TLR7 were expressed on such lymphocytes, and the TLR agonists PGN, Poly I:C, LPS, and R848 induced a higher level of IFN-γ. However, a higher level of IL-4 was found in the supernatant of pulmonary lymphocytes from infected mice stimulated by these TLR agonists plus CD3 Ab. Only R848 plus anti-CD3 mAb could induce a higher level of IFN-γ in such lymphocytes. TLR expressions were then compared on different pulmonary lymphocytes after infection, including T cells, B cells, NK cells, NKT cells, and γδT cells. The expression levels of TLR3 on T cells, B cells, NK cells, and γδT cells were increased in the lungs after infection. NK cells also expressed higher levels of TLR4 after infection of control mice. Collectively, these findings highlight the potential role of TLR expression in the context of S. japonicum infection.
Asunto(s)
Pulmón/inmunología , Linfocitos/inmunología , Schistosoma japonicum/fisiología , Esquistosomiasis Japónica/inmunología , Receptor Toll-Like 3/metabolismo , Animales , Femenino , Regulación de la Expresión Génica , Humanos , Interferón gamma/metabolismo , Interleucina-4/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Recuento de Huevos de Parásitos , Receptor Toll-Like 2/genética , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 3/genética , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismo , Receptor Toll-Like 7/genética , Receptor Toll-Like 7/metabolismo , TranscriptomaRESUMEN
Schistosoma japonicum infection can induce granulomatous inflammation and cause tissue damage in the mouse liver. The cytokine secretion profile of T helper (Th) cells depends on both the nature of the activating stimulus and the local microenvironment (e.g. cytokines and other soluble factors). In the present study, we found an accumulation of large numbers of IFN-γ(+) IL-4(+) CD4(+) T cells in mouse livers. This IFN-γ(+) IL-4(+) cell population increased from 0·68 ± 0·57% in uninfected mice to 7·05 ± 3·0% by week 4 following infection and to 9·6 ± 5·28% by week 6, before decreasing to 6·3 ± 5·9% by week 8 in CD4 T cells. Moreover, IFN-γ(+) IL-4(+) Th cells were also found in mouse spleen and mesenteric lymph nodes 6 weeks after infection. The majority of the IFN-γ(+) IL-4(+) Th cells were thought to be related to a state of immune activation, and some were memory T cells. Moreover, we found that these S. japonicum infection-induced IFN-γ(+) IL-4(+) cells could express interleukin-2 (IL-2), IL-9, IL-17 and high IL-10 levels at 6 weeks after S. japonicum infection. Taken together, our data suggest the existence of a population of IFN-γ(+) IL-4(+) plasticity effector/memory Th cells following S. japonicum infection in C57BL/6 mice.
Asunto(s)
Hígado/inmunología , Schistosoma japonicum/inmunología , Esquistosomiasis Japónica/inmunología , Subgrupos de Linfocitos T/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Diferenciación Celular , Células Cultivadas , Femenino , Memoria Inmunológica , Interferón gamma/metabolismo , Interleucina-4/metabolismo , Hígado/parasitología , Ratones , Ratones Endogámicos C57BL , Balance Th1 - Th2RESUMEN
Gamma delta (γδ) T cells are mainly present in mucosa-associated lymphoid tissues, which play an important role in mucosal immunity. In this study, C57BL/6 mice were infected by Schistosoma japonicum and lymphocytes were isolated from the mesenteric lymph node (MLN) to identify changes in the phenotype and function of γδ T cells using flow cytometry. Our results indicated that the absolute number of γδ T cells from the MLNs of infected mice was significantly higher compared with normal mice (P < 0.05). In addition, the infected γδ T cells expressed a high level of the activated molecule CD69 (P < 0.01) and demonstrated an increasing population of CD4(+) γδ T cells (P < 0.05). MLN γδ T cells secrete interferon-γ (IFN-γ), interleukin (IL)-4, IL-9, and IL-17 in response to propylene glycol monomethyl acetate (PMA) plus ionomycin simulation, and the levels of IL-4, IL-9, and IL-17 increased significantly after S. japonicum infection (P < 0.05). Taken together, these findings indicated that S. japonicum infection could induce γδ T cell activation, proliferation, and differentiation in the MLN. Moreover, our results indicated that the expression of NKG2D (CD314) was not increased in γδ T cells after infection, suggesting that other mechanisms are involved in activating γδ T cells. Furthermore, higher expression of programmed death-1 (CD279) but not IL-10 was detected in the γδ T cells isolated from infected mice (P < 0.05), suggesting that the function of γδ T cells is inhibited gradually over the course of S. japonicum infection.
Asunto(s)
Ganglios Linfáticos/citología , Activación de Linfocitos/inmunología , Schistosoma japonicum , Esquistosomiasis Japónica/inmunología , Subgrupos de Linfocitos T/clasificación , Animales , Femenino , Interferón gamma/metabolismo , Interleucinas/metabolismo , Ganglios Linfáticos/patología , Mesenterio/patología , Ratones , Ratones Endogámicos C57BL , Subgrupos de Linfocitos T/metabolismoRESUMEN
The mesenteric lymph node (MLN) is the main draining lymph node in mouse enterocoelia, which contains many types of immune cells. Among these cells, natural killer (NK) and natural killer T (NKT) cells belong to innate lymphoid cells (ILCs), which have potent activities for controlling a variety of pathogenic infections. In this study, C57BL/6 mice were infected with Schistosoma japonicum for 5-7 weeks. Lymphocytes were isolated from the MLN to detect changes in the phenotype and function of NK and NKT cells using a fluorescence activating cell sorter (FACS). These results demonstrated that a S. japonicum infection could significantly increase the percentage of NK cells in the mouse MLN, (P < 0.05). We found an increase in the cell number of both NK and NKT cells. In addition, we found that NK and NKT cells from infected mice expressed higher levels of CD69 compared to normal mice (P < 0.05). These results demonstrated that a S. japonicum infection could induce MLN NK and NKT cell activation. Moreover, we found that the expression of CD4 was increased in infected MLN NK cells (P < 0.05). Furthermore, intracellular cytokine staining revealed that expression of IL-4 and IL-17 were significantly enhanced in both the NK and NKT cells of infected mice after phorbol 12-myristate 13-acetate (PMA) and ionomycin stimulation (P < 0.05). Taken together, these results indicated that infection-induced MLN NK and NKT cells might play roles in modulating the classical T cell response. Finally, our results indicated that the expression of CD94 was decreased in NK cells, suggesting that the downregulation of CD94 expression might served as a mechanism in NK cell activation.
Asunto(s)
Células Asesinas Naturales/citología , Ganglios Linfáticos/inmunología , Células T Asesinas Naturales/citología , Esquistosomiasis Japónica/patología , Animales , Femenino , Interleucina-17/inmunología , Interleucina-4/inmunología , Células Asesinas Naturales/inmunología , Ganglios Linfáticos/citología , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Subfamília D de Receptores Similares a Lectina de las Células NK/metabolismo , Células T Asesinas Naturales/inmunología , Schistosoma japonicum , Esquistosomiasis Japónica/inmunologíaRESUMEN
In schistosomiasis, limited information is available about the role of interleukin-17 (IL-17) in lung, despite the fact that this cytokine plays a crucial role during pro-inflammatory immune responses. In our study, we observed CD4(+)T cells changed after the infection. Furthermore, ELISA and FACS results revealed that Schistosomajaponicum infection could induce a large amount of IL-17 in mouse pulmonary lymphocytes. IL-17-producing cells, including Th17 cells, CD8(+)T (Tc) cells, γδT cells and natural killer T cells, was also associated with the development of lung inflammatory diseases. FACS results indicated that Th17 cell was the main source of IL-17 in the infected pulmonary lymphocytes after phorbol-12-myristate-13-acetate (PMA) and Ionomycin stimulation. Moreover, FACS results revealed that the percentage of Th17 cells continued to increase as over the course of S. japonicum infection. Additionally, cytokines co-expression results demonstrated that Th17 cells could express more IL-4 and IL-5 than IFN-γ. Reducing IL-17 activity by using anti-IL-17 ameliorated the damage and decreased infiltration of inflammatory cells in infected C57BL/6 mouse lungs. Collectively, these results suggest Th17 cells is the major IL-17-producing cells population and IL-17 contributes to pulmonary granulomatous inflammatory during the S. japonicum infection.
Asunto(s)
Granuloma del Sistema Respiratorio/inmunología , Interleucina-17/metabolismo , Schistosoma japonicum/inmunología , Esquistosomiasis Japónica/inmunología , Células Th17/inmunología , Animales , Linfocitos T CD8-positivos/inmunología , Femenino , Inflamación/inmunología , Interferón gamma/metabolismo , Interleucina-4/metabolismo , Interleucina-5/metabolismo , Ionomicina/metabolismo , Pulmón/inmunología , Pulmón/patología , Ratones , Ratones Endogámicos C57BL , Células T Asesinas Naturales/inmunología , Ésteres del Forbol/metabolismo , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Esquistosomiasis Japónica/patologíaRESUMEN
OBJECTIVE: To compare the percentages of γδT cells in the lymphocytes and CD3âº; T cells isolated from the livers, lungs, spleen and mesenteric lymph nodes of C57BL/6 mice and the phonotype and function of γδT cells in different tissues or organs. METHODS: Lymphocytes were isolated from livers, lungs, spleen and mesenteric lymph nodes of normal C57BL/6 mice, respectively. Then, percentages of γδT cells in lymphocytes and CD3âº; T cells, and phenotypic characteristics of γδT cells were examined by flow cytometry. Moreover, IFN-γ, IL-4, IL-9 and IL-17 secreted by γδT cells were detected by means of intracellular cytokine staining after stimulation with PMA plus ionomycin. RESULTS: The percentage of γδT in lymphocyte cells of the livers was significantly higher than that of the lungs, spleen and mesenteric lymph nodes (P<0.05), and the percentage in CD3âº; T cells of the mesenteric lymph nodes was the lowest (P<0.05). CD4â»; CD8â»; γδT cells were the main subpopulation in these tissues and organs and there was also a small proportion of CD8âº; γδT cells. The proportion of CD4âº; γδT cells in the mesenteric lymph nodes was significantly higher than that in the others (P<0.05), and a group of CD4âº; CD8âº; γδT cells existed obviously. The percentage of IL-17âº; cells in γδT cells was significantly higher than IFN-γâº; and IL-4âº; cells, and almost no IL-9 γδT cells were found. The ability of secreting cytokines of γδT cells in the lungs was the strongest and the percentage of IL-17âº; γδT was (26.6 ± 12.1) %. In addition, the proportion of IFN-γâºâ»; γδT cells in the livers and lungs were (1.36 ± 0.37)% and (1.6 ± 0.7)%, respectively. CONCLUSION: The proportion, phenotype and function of γδT cells had significant difference in the livers, lungs, spleen, mesenteric lymph nodes of C57BL/6 mice.
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Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Subgrupos de Linfocitos T/metabolismo , Animales , Citocinas/biosíntesis , Femenino , Citometría de Flujo , Ratones , Ratones Endogámicos C57BL , Especificidad de Órganos/inmunología , Subgrupos de Linfocitos T/inmunologíaRESUMEN
Schistosomiasis japonica is a severe tropical disease caused by the parasitic worm Schistosoma japonicum. Among the most serious pathological effects of S. japonicum infection are hepatic lesions (cirrhosis and fibrosis) and portal hypertension. Interleukin-17 (IL-17) is a pro-inflammatory cytokine involved in the pathogenesis of many inflammatory and infectious conditions, including schistosomiasis. We infected C57BL/6 mice with S. japonicum and isolated lymphocytes from the liver to identify cell subsets with high IL-17 expression and release using flow cytometry and ELISA. Expression and release of IL-17 was significantly higher in hepatic lymphocytes from infected mice compared with control mice in response to both non-specific stimulation with anti-CD3 monoclonal antibody plus/anti-CD28 monoclonal antibody and PMA plus ionomycin. We then compared IL-17 expression in three hepatic T-cell subsets, T helper, natural killer T and γδT cells, to determine the major source of IL-17 during infection. Interleukin-17 was induced in all three subsets by PMA + ionomycin, but γδT lymphocytes exhibited the largest increase in expression. We then established a mouse model to further investigate the role of IL-17 in granulomatous and fibrosing inflammation against parasite eggs. Reducing IL-17 activity using anti-IL-17A antibodies decreased infiltration of inflammatory cells and collagen deposition in the livers of infected C57BL/6 mice. The serum levels of soluble egg antigen (IL)-specific IgGs were enhanced by anti-IL-17A monoclonal antibody blockade, suggesting that IL-17 normally serves to suppress this humoral response. These findings suggest that γδT cells are the most IL-17-producing cells and that IL-17 contributes to granulomatous inflammatory and fibrosing reactions in S. japonicum-infected C57BL/6 mouse liver.
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Mediadores de Inflamación/metabolismo , Interleucina-17/metabolismo , Hígado/inmunología , Schistosoma japonicum/inmunología , Esquistosomiasis Japónica/inmunología , Subgrupos de Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Neutralizantes/farmacología , Anticuerpos Antiprotozoarios/sangre , Células Cultivadas , Colágeno Tipo III/metabolismo , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Citometría de Flujo , Granuloma/tratamiento farmacológico , Granuloma/inmunología , Granuloma/parasitología , Ionomicina/farmacología , Hígado/efectos de los fármacos , Hígado/parasitología , Hígado/patología , Cirrosis Hepática/tratamiento farmacológico , Cirrosis Hepática/inmunología , Cirrosis Hepática/parasitología , Ratones , Ratones Endogámicos C57BL , Células T Asesinas Naturales/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Schistosoma japonicum/patogenicidad , Esquistosomiasis Japónica/tratamiento farmacológico , Esquistosomiasis Japónica/parasitología , Esquistosomiasis Japónica/patología , Subgrupos de Linfocitos T/efectos de los fármacos , Subgrupos de Linfocitos T/parasitología , Células Th17/inmunología , Regulación hacia ArribaRESUMEN
Schistosome infection could cause significant liver damage in animal; Th2 cells play an important role in the progress of this disease. In our study, C57BL/6 mice were infected by Schistosoma japonicum and lymphocytes were isolated from the liver to detect some characteristics of interleukin-5 (IL-5)-producing T cells by different methods. The results revealed that S. japonicum infection could induce a large amount of IL-5 in mouse liver T cells by the means of fluorescent bead immunoassay and RT-PCR. Although, mouse liver contained many T cell subsets, such as Th cells, Tc cells, NKT cells, and γδ T cells. Fluorescence activated cell sorting results indicated that Th cells were the main source of IL-5 in the T cell population after phorbol 12-myristate 13-acetate and ionomycin stimulation. Moreover, the percentage of IL-5-producing Th cells continued to increase from 4 to 8 weeks after S. japonicum infection, which differed from the changes of IFN-γ(+) Th1 cells, IL-4(+) Th2 cells, and IL-17A(+) Th17 cells during S. japonicum infection. Additionally, cytokines co-expression results demonstrated that 36.2 % of IL-5(+) Th cells could express IL-4, and 10 % of it could produce IFN-γ or IL-17A. Collectively, these findings implied that IL-5-producing Th cells posses some properties which differ from other cytokines secreting Th cells.
Asunto(s)
Interleucina-5/biosíntesis , Hígado/inmunología , Schistosoma japonicum/patogenicidad , Esquistosomiasis Japónica/inmunología , Linfocitos T/inmunología , Animales , Interferón gamma/biosíntesis , Interleucina-17/metabolismo , Interleucina-4/biosíntesis , Hígado/parasitología , Ratones , Ratones Endogámicos C57BL , Schistosoma japonicum/inmunología , Esquistosomiasis Japónica/parasitología , Linfocitos T/citología , Linfocitos T/metabolismo , Células TH1/citología , Células TH1/inmunología , Células TH1/metabolismo , Células Th2/citología , Células Th2/inmunología , Células Th2/metabolismoRESUMEN
OBJECTIVE: To observe the immune response of Th17 cells in mesenteric lymph node (MLN) of C57BL/6 mice infected by Schistosoma japonicum. METHODS: Twenty C57BL/6 mice were randomly divided into infected group and control group each with ten mice. The mice in infected group were infected each with 40 +/- 5 S. japonicum cercariae. Five to six weeks later, MLN lymphocytes were separated and stimulated for 4 h by anti-CD3 (1 microg/ml) and anti-CD28 (1 microg/ml) before examination of IL-17 and retinoic acid receptor-related orphan receptor gammat (ROR-gammat) mRNA by reverse transcription PCR. The level of IL-17 and IFN-gamma was detected by ELISA after culturing with supernatant for 72h. MLN lymphocytes were stimulated for 5h by 10 ng/ml phorbol myristoyl acetate (PMA) and 1 microg/ml ionomycin. The intracellular cytokines were stained and the content of Th17 and other cytokines was examined by flow cytometry. RESULTS: The level of IFN-gamma [(214.3 +/- 62.6) pg/ml] and IL-17 [(176.8 +/- 62.1) pg/ml] in the supernatant of cultured MLN cells from the infected mice was significantly higher than that of normal mice [(467 +/- 13.9) and 0 pg/ml) (P < 0.05). The expression level of IL-17 and ROR-gammat mRNA was also considerably higher than that of normal mice. IL-17+ IL-4+, IL-17+ IFN-gamma+, IL-17+ IL-5+ and IL-17+ IL-9 cells accounted for 0.06%, 0.02%, 0.02%, and 0.01% of the mesenteric lymph node CD4+ T cells of the infected mice, respectively. However, IL-17+ IL-10+ and IL-17+ Foxp3+ cells were undetected. CONCLUSION: The MLN of S. japonicum-infected C57BL/6 mice can induce the production of Th17 cells, and these cells can secrete IL-4, less IFN-gamma, IL-5 and IL-9, but not IL-10, and can not express Foxp3 in the infected mice.