CXCL12 expression within the CNS contributes to the resistance against experimental autoimmune encephalomyelitis in Albino Oxford rats

Experimental autoimmune encephalomyelitis (EAE) is an animal model of multiple sclerosis, a chronic inflammatory and demyelinating disease of the CNS. Albino Oxford (AO) rats are resistant to the induction of EAE, while the disease can be readily induced in Dark Agouti (DA) rats. Here we investigated a potential contribution of the CNS milieu in the limitation of the encephalitogenic autoimmune response. EAE was induced by immunization of the respective rat strains with spinal cord homogenate emulsified in complete Freund's adjuvant. AO rats did not exhibit clinical signs after immunization while DA rats developed severe neurologic deficits. Infiltration of immune cells into spinal cords (SC) was evident in both strains 12-14 days after the immunization. EAE lesions of AO rats contained substantially lower numbers of CD4+ T cells and CD11b+ cells compared to those in DA rats. This went together with lower levels of interferon (IFN)-γ and interleukin (IL)-17 in the cells isolated from SC. We found a dramatic increase of CXCL12 expression in SC tissue and microvessels of AO rats, whereas DA rats markedly decreased the expression of this chemokine within their CNS. Administration of the CXCL12 antagonist AMD3100 to a substrain of AO rats that developed a weak EAE led to earlier onset and exacerbation of the disease. These results suggest a role of CXCL12 in down-regulating autoimmune processes in AO rats during EAE. Therapeutic modulation of CXCL12 could be a promising strategy for the treatment of CNS autoimmunity.     

Genetic influence on disease course and cytokine response in relapsing experimental allergic encephalomyelitis

A protracted and relapsing form of experimental allergic encephalomyelitis (EAE) develops in the DA rat after immunization with rat spinal cord homogenate (SCH) emulsified in incomplete Freund's adjuvant (IFA). The genetic influence on this model has been analyzed by immunizing MHC congenic strains on both LEW and DA genetic backgrounds, and recombinant inbred strains between DA and E3 rats. An in situ hybridization assay was used to examine the expression of mRNA for IFN-gamma, IL-4, IL-10 and transforming growth factor (TGF)-beta both in sections of spinal cords and the antigen-induced expression for these cytokines by splenocytes after in vitro stimulation with encephalitogenic MBP peptides. The susceptibility of relapsing EAE after immunization with SCH in IFA in the DA strain, but not the E3 strain, was correlated with a lack of expression for TGF-beta in the spinal cord. The recombinant inbred DXEB rats developed a severe EAE while surprisingly no signs of disease were observed in the DXEA strain, which shares the MHC region with the DXEB strain, after immunization with the MBP 63-87 peptide. Resistance to relapsing EAE in the DXEA strain correlated with increased non-MHC controlled expression for TGF-beta and lack of IFN-gamma in the spinal cord. The same pattern of cytokine expression was seen in splenocytes after stimulation in vitro with the MBP 63-87 peptide. A spreading of the immune response to the MBP 87-110 peptide was seen. Non-MHC genes controlled the quality of this response: splenocytes from MBP 63-87 immunized DXEB rats responded in vitro towards the MBP 87-110 peptide by expressing mRNA for IFN-gamma, IL-10 and IL-4, whereas in the DXEA strain the corresponding response involved IL-4 and TGF-beta. Taken together these data show that non-MHC controlled expression of mRNA for TGF-beta is associated with resistance to EAE.      

Elevated interferon gamma expression in the central nervous system of tumour necrosis factor receptor 1-deficient mice with experimental autoimmune encephalomyelitis

Inflammation in the central nervous system (CNS) can be studied in experimental autoimmune encephalomyelitis (EAE). The proinflammatory cytokines interferon-gamma (IFN-gamma) and tumour necrosis factor (TNF) are implicated in EAE pathogenesis. Signals through the type 1 TNF receptor (TNFR1) are required for severe EAE to develop, whereas deficiency in IFN-gamma or its receptor result in more severe EAE. We investigated IFN-gamma expression in TNFR1-deficient (TNFR1-/-) mice. We describe here that there were more IFN-gamma-secreting T cells present in the CNS of TNFR1-/- mice during EAE compared to wild-type (WT) mice, despite that clinical symptoms were mild, with delayed onset. There was greater expression of IL-12/23p40 by antigen-presenting cells in these mice, and in vitro, TNFR1-/- antigen-presenting cells induced greater secretion of IFN-gamma but not interleukin (IL)-17 when cultured with primed T cells than did WT antigen presenting cells. TNFR1-/- mice with EAE had significantly higher expression of CXCL10 mRNA (but not CCL5 mRNA) in the CNS compared to WT mice with EAE. These data demonstrate that IFN-gamma expression is enhanced in the CNS of TNFR1-/- mice with EAE and suggest that IFN-gamma levels do not necessarily correlate with EAE severity.     

Reversal of experimental allergic encephalomyelitis with non-mitogenic, non-depleting anti-CD3 mAb therapy with a preferential effect on T(h)1 cells that is augmented by IL-4

This study examined whether therapy with a non-mitogenic, non-activating anti-CD3 mAb (G4.18) alone, or in combination with the T(h)2 cytokines, could inhibit induction or facilitate recovery from experimental allergic encephalomyelitis (EAE) in Lewis rats. G4.18, but not rIL-4, rIL-5 or anti-IL-4 mAb, reduced the severity and accelerated recovery from active EAE. A combination of rIL-4 with G4.18 was more effective than G4.18 alone. The infiltrate of CD4(+) and CD8(+) T cells, B cells, dendritic cells, and macrophages in the brain stem was less with combined G4.18 and IL-4 than G4.18 therapy or no treatment. Residual cells had preferential sparing of T(r)1 cytokines IL-5 and transforming growth factor-beta with loss of T(h)1 markers IL-2, IFN-gamma and IL-12Rbeta2, and the T(h)2 cytokine IL-4 as well as macrophage cytokines IL-10 and tumor necrosis factor-alpha. Lymph nodes draining the site of immunization had less mRNA for T(h)1 cytokines, but T(h)2 and T(r)1 cytokine expression was spared. Treatment with G4.18, rIL-4 or rIL-5 from the time of immunization had no effect on the course of active EAE. MRC OX-81, a mAb that blocks IL-4, delayed onset by 2 days, but had no effect on severity of active EAE. G4.18 also inhibited the ability of activated T cells from rats with active EAE to transfer passive EAE. This study demonstrated that T cell-mediated inflammation was rapidly reversed by a non-activating anti-CD3 mAb that blocked effector T(h)1 cells, and spared cells expressing T(h)2 and T(r)1 cytokines.     

Suppression of experimental allergic encephalomyelitis by intraventricular administration of interferon-gamma in Lewis rats.

Experimental allergic encephalomyelitis (EAE) is an autoimmune inflammatory disease of the central nervous system (CNS) which causes paralysis. Several studies have reported the involvement of Ia antigen-expressing cells in the pathogenesis of EAE. Interferon-gamma (IFN-gamma) can induce Ia antigen expression on a wide range of cells. We examined the effect of IFN-gamma on EAE in Lewis rats. Systemically administered IFN-gamma did not change the disease course of EAE, whereas IFN-gamma applied locally into the ventricular system of the CNS resulted in complete suppression of clinical signs. Furthermore, we found that systemic administration of anti-IFN-gamma just prior to the onset of clinical symptoms resulted in a more severe disease course. We conclude that IFN-gamma is capable of exerting a suppressive action in EAE, possibly through induction of Ia antigen expression or through the induction of suppressive mechanisms locally in the CNS.     

Cellular mRNA expression of interferon-gamma, IL-4 and transforming growth factor-beta (TGF-beta) by rat mononuclear cells stimulated with peripheral nerve myelin antigens in experimental allergic neuritis.

Experimental allergic neuritis (EAN) serves as a useful model for inflammation in the peripheral nervous system. To study the potential role of important immunoregulatory and effector cytokines in EAN, we examined the expression of mRNA for interferon-gamma (IFN-gamma), IL-4 and TGF-beta by in situ hybridization in lymph node and splenic cells cultured with bovine peripheral nerve myelin (BPM), P2 and P0 during the course of EAN in Lewis rats. Levels of IFN-gamma mRNA-expressing mononuclear cells (MNC) from lymph nodes and spleens roughly correlated with clinical status, consistent with a disease-promoting role for IFN-gamma. BPM, P0 and P2-reactive IFN-gamma mRNA-expressing T cells appeared in lymph nodes and spleen before onset of the disease, whereas a significant TGF-beta response to BPM, P2 and P0 was observed at lower levels than the IFN-gamma response and at onset of recovery, consistent with a disease down-regulating role of TGF-beta. IL-4 mRNA-expressing cells were found at levels similar to TGF-beta mRNA-expressing cells, and with the latest peak of the three cytokines examined. This result suggests that IL-4 may also suppress IFN-gamma expression at late recovery phase of EAN.     

Differential expression of suppressors of cytokine signaling-1 and -3 and related cytokines in central nervous system during remitting versus non-remitting forms of experimental autoimmune encephalomyelitis

SJL mice exhibit a relapsing-remitting course of experimental autoimmune encephalomyelitis (EAE), whereas C57BL/6 (B6) mice display a more chronic course without complete remissions. Suppressor of cytokine signaling (SOCS)-1 and SOCS-3 are members of a family of inducible intracellular proteins that negatively regulate cytokine signaling in cells of hematopoietic origin and may influence the Th1 to Th2 balance. SOCS-1 and SOCS-3 are induced by cytokines that are known to be up-regulated during EAE, including IFN-gamma (IFN-g) and IL-6, respectively. To test the hypothesis that the level of induction of SOCS-1 and SOCS-3 correlates with the course of EAE, mRNA levels were compared in spinal cords of SJL and B6 mice during discrete stages of disease. SOCS-1 and SOCS-3 were elevated throughout active disease in both strains. At peak EAE, SOCS-1 was higher and SOCS-3 was lower in B6 cords compared with SJL cords. This correlated with greater expression of the Th1 cytokine, IFN-g, and less of the Th2 cytokine, IL-10, in B6 cords relative to SJL cords during onset and peak disease. SOCS-3 inducers in the IL-6 family were expressed differentially between the strains. IL-6 and leukemia inhibitory factor were higher at onset in B6 cords whereas ciliary neurotrophic factor was increased in SJL cords during peak disease. Expression of fibroblast growth factor-2, which may be involved in remyelination, was higher in SJL cords at peak. Comparison of these models suggests that cytokine autoregulatory mechanisms involving SOCS may play a role in determining the course of EAE.     

Balance of Th1/Th2 Cytokines Associated with the Preventive Effect of Incomplete Freund's Adjuvant on the Development of Adjuvant Arthritis in LEW Rats

Complete Freund's adjuvant (CFA) could induce adjuvant arthritis (AA) in LEW rats and incomplete Freund's adjuvant (IFA) could induce oil induced arthritis (OIA) in DA but not in LEW rats. Lymph node cells (LNCs) from these AA and OIA rats showed increased mRNA expression of IFN-gamma, IL-2 and TNF-alpha but not IL-4. LNCs from IFA immunized LEW rats showed increased expression of IL-4, reduced expression of IFN-gamma and TNF-alpha and no IL-2, in contrast to IFA immunized DA rats. The pretreatment of IFA before CFA challenge could completely prevent AA in LEW rats and their LNCs showed increased expression of IL-4 and IFN-gamma but not IL-2 and TNF-alpha. In F1 (LEW x DA) rats, IFA could not induce OIA but the pretreatment of IFA before CFA challenge could induce very mild AA with 80% incidence, LNCs showing an elevated expression of all the above cytokines. These findings suggest that increased Th1 cytokine expression is associated with disease development and that increased IL-4 expression or the balance of Th2 over Th1 cytokine expression plays an important regulatory role in disease development.     

Dual-label immunohistochemical study of interleukin-4-and interferon-gamma-expressing cells within the pancreas of the NOD mouse during disease acceleration with cyclophosphamide

Beta cell destruction has been shown to occur when rodent or human islets are exposed in vitro to inflammatory cytokines, such as interleukin-1beta (IL-1beta), tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). Other cytokines such as interleukin-4 (IL-4) or interleukin-10 (IL-10), when given to NOD mice, prevent insulin-dependent diabetes mellitus (IDDM). In this study, we have employed immunofluorescence histochemistry to study the expression of IFN-gamma and IL-4 in the pancreas of female NOD mice at various time-points (days 0, 4, 7, 11 and at onset of diabetes) following disease acceleration with cyclophosphamide (Cy). Dual-label confocal and light microscopy were employed to determine the precise cellular sources of the two cytokines. IL-4 immunolabelling was observed in a few immune cells at days 0, 4, and 7 within the pancreatic islets but in larger numbers at day 11 and at onset of diabetes. The cytokine was co-localized predominantly in CD4 cells, while only a small minority of CD8 cells and macrophages also expressed IL-4. At days 0, 4, 7 and 11, weak to moderate immunolabelling for IL-4 was also observed in beta cells. In contrast, immunolabelling for IFN-gamma within the islets was not observed until day 11 and this labelling persisted at onset of diabetes. It was immunolocalized in macrophages and to a lesser extent in CD4 cells. Only a few CD8 cells were immunopositive for IFN-gamma. At day 11, a proportion of beta cells showed weak immunolabelling for IFN-gamma. During the study period, immunolabelling for IFN-gamma was also observed in a proportion of endothelial cells located in the intra-islet and exocrine regions of Cy and diluent-treated mice. From day 11 onwards, both the cytokines were observed in some of the peri-vascular regions. Our results demonstrate that during Cy-induced diabetes, there is increasing expression of both IL-4 and IFN-gamma in specific immune cells within the inflamed islets in the late prediabetic stage and at onset of diabetes. Further studies are required to correlate our protein immunohistochemical findings with in situ cytokine gene expression and to determine whether there is a clear Th1 cytokine protein bias at clinical onset of diabetes and immediately preceding it.     

Application of dendritic cells stimulated with Trichinella spiralis excretory–secretory antigens alleviates experimental autoimmune encephalomyelitis

The parasitic nematode, Trichinella spiralis (T. spiralis), exerts an immunomodulatory effect on the host immune response through excretory–secretory products (ES L1) released from encysted muscle larvae. Our model of combined T. spiralis infection and experimental autoimmune encephalomyelitis (EAE) in Dark Agouti (DA) rats demonstrated a significant reduction in EAE severity in infected animals. Recently, we have created an immune status characteristic for the live infection by in vivo application of dendritic cells (DCs) stimulated with ES L1 products of T. spiralis muscle larvae. Moreover, these cells were able to ameliorate EAE when applied 7 days before EAE induction. ES L1-stimulated DCs increased production of IL-4, IL-10 and TGF-β, and decreased production of IFN-γ and IL-17, both at the systemic level and in target organs. A significant increase in the proportion of CD4+CD25+Foxp3+ T cells was found among spleen cells, and CNS infiltrates from DA rats treated with ES L1-stimulated DCs before EAE induction, compared to controls injected with unstimulated DCs. Regulatory T cells, together with elevated levels of IL-10 and TGF-β, are most likely involved in restraining the production of Th1 and Th17 cytokines responsible for autoimmunity and thus are responsible for the beneficial effect of ES L1-educated DCs on the course of EAE. Our results show that ES L1 antigen-stimulated DCs are able not only to provoke, but also to sustain anti-inflammatory and regulatory responses regardless of EAE induction, with subsequent amelioration of EAE, or even protection from the disease.     

Regulation of autoimmune arthritis by the pro-inflammatory cytokine interferon-gamma

The pathogenesis of T cell-mediated diseases like rheumatoid arthritis (RA) has typically been explained in the context of the Th1-Th2 paradigm: the initiation/propagation by pro-inflammatory cytokines, and downregulation by Th2 cytokines. However, in our study based on the adjuvant-induced arthritis (AA) model of RA, we observed that Lewis (LEW) (RT.1(l)) rats at the recovery phase of AA showed the highest level of IFN-gamma in recall response to mycobacterial heat-shock protein 65 (Bhsp65), whereas AA-resistant Wistar-Kyoto (WKY) (RT.1(l)) rats secreted high levels of IFN-gamma much earlier following disease induction. However, no significant secretion of IL-10 or TGF-beta was observed in either strain. Furthermore, pre-treatment of LEW rats with a peptide of self (rat) hsp65 (R465), which induced T cells secreting predominantly IFN-gamma, afforded protection against AA and decreased IL-17 expression by the arthritogenic epitope-restimulated T cells. These results provide a novel perspective on the pathogenesis of autoimmune arthritis.     

Cytokine and chemokine alterations in tissue,CSF, and plasma in early presymptomatic phase of experimental allergic encephalomyelitis (EAE), in a rat model of multiple sclerosis

Background

Experimental allergic encephalomyelitis (EAE) is the most commonly used experimental animal model for human multiple sclerosis (MS) that has been used so far to study the acute and remission-relapsing phases of the disease. Despite the vast literature on neuroinflammation onset and progression in EAE, important questions are still open regarding in particular the early asymptomatic phase between immunization and clinical onset.

Methods

In this study, we performed a time-course investigation of neuroinflammation and demyelination biomarkers in the spinal cord (SC), cerebrospinal fluid (CSF), and blood in EAE induced in dark agouti (DA) female rats compared to the controls and adjuvant-injected rats, using high-throughput technologies for gene expression and protein assays and focusing on the time-course between immunization, clinical onset (1, 5, 8 days post-immunization (DPI)), and progression (11 and 18 DPI). The expression profile of 84 genes related to T cell activation/signaling, adaptive immunity, cytokine/chemokine inflammation, demyelination, and cellular stress were analyzed in the tissue; 24 cytokines were measured in the CSF and plasma.

Results

The macrophage colony-stimulating factor (CSF1) was the first up-regulated protein as far as 1 DPI, not only in blood but also in CSF and SC. A treatment with GW2580, a selective CSF1R inhibitor, slowed the disease progression, significantly reduced the severity, and prevented the relapse phase. Moreover, both pro-inflammatory (IL-1β, TNF-α) and anti-inflammatory cytokines (IL-5, IL-10, VEGF) were up-regulated starting from 8 DPI. Myelin genes were down-regulated starting from 8 DPI, especially MAL, MBP, and PMP22 while an opposite expression profile was observed for inflammation-related genes, such as CXCL11 and CXCL10.

Conclusions

This early cytokine and chemokine regulation indicates that novel biomarkers and therapeutic options could be explored in the asymptomatic phase of EAE. Overall, our findings provide clear evidence that CSF1R signaling regulates inflammation in EAE, supporting therapeutic targeting of CSF1R in MS.

     

ICOS Deficiency Results in Exacerbated IL-17 Mediated Experimental Autoimmune Encephalomyelitis

Introduction  Inducible costimulatory molecule (ICOS) is important for the effector function of T cells, especially for Th2 and T cell dependent B cell responses. However, it has been shown that ICOS is required for the differentiation of Th17 cells. Since IL-17 has been identified as a major cytokine involved in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), the enhanced severity of EAE in ICOS-deficient mice (ICOS^−/−) mice is unexpected. Methods  To better understand the role of ICOS and of IL-17 in EAE, we induced EAE in ICOS^−/− by immunization with myelin oligodendrocyte glycoprotein peptide (MOG_35–55) in complete Freund’s adjuvant. Results  As previously reported, we found that ICOS^−/− mice developed more severe EAE. Upon restimulation with MOG_35–55, splenocytes from ICOS^−/− mice with EAE produced higher amounts of IL-17 and ICOS^−/− mice had a higher expression of IL-17, IL-6, and TGF-β mRNA in the spinal cords at the onset of the disease. Finally, the blockade of IL-17 strongly inhibited disease even in ICOS^−/− mice, showing that IL-17 is playing a major role in the pathogenesis of EAE both in WT and ICOS^−/− mice. Conclusion  In conclusion, MOG immunization induces MOG-specific Th17 cells also in ICOS^−/− mice, and a higher expression of IL-17 and of Th17-driving cytokines IL-6 and TGF-β in the central nervous system at the onset of EAE that correlates with their more severe disease.     

Combined nasal administration of encephalitogenic myelin basic protein peptide 68-86 and IL-10 suppressed incipient experimental allergic encephalomyelitis in Lewis rats

Mucosal administration of low doses of myelin basic protein (MBP) peptide 68-86 (MBP 68-86) or anti-inflammatory cytokine IL-10 effectively prevented experimental allergic encephalomyelitis (EAE), but failed to suppress the disease if given after 7 days postimmunization (p.i.), i.e., after T cell priming had occurred. We anticipated that combined administration of autoantigen and IL-10 can treat incipient EAE. Lewis rats with EAE actively induced with MBP 68-86 and complete Freund's adjuvant received 120 microg MBP 68-86 + 200 ng IL-10 per rat per day from day 7 p.i. and for 5 consecutive days. These rats showed later onset, lower clinical scores, less body weight loss, and shorter duration of EAE than rats receiving MBP 68-86 or IL-10 only or PBS. EAE amelioration was associated with decreased infiltration of ED1(+) macrophages and CD4(+) T cells within the central nervous system and with decreased proliferative responses of lymph node cells, indicating that combined administration of MBP 68-86 and IL-10 induced immune hyporesponsiveness. IFN-gamma secretion as well as IFN-gamma, TNF-alpha, IL-4, and IL-10 mRNA expression by lymph node MNC was down-regulated in the treated rats. Immune hyporesponsiveness, rather than immune deviation or regulatory mechanisms, seems to be responsible for the protection of EAE after autoantigen + IL-10 administration by the nasal route.     

Infection of non-encapsulated species of Trichinella ameliorates experimental autoimmune encephalomyelitis involving suppression of Th17 and Th1 response

Epidemiological and experimental studies have indicated that helminth infections can ameliorate autoimmune diseases. The present study investigated the amelioration effect of the Trichinella pseudospiralis infection on experimental autoimmune encephalomyelitis (EAE), a T-cell-mediated autoimmune disease of central nervous system (CNS), and expression kinetics of Th17 and Th1 cytokine which play a crucial role in the pathogenesis of EAE. The results indicated that the infection of helminth T. pseudospiralis obviously ameliorated clinical severity and greatly delayed the onset of EAE induced by myelin oligodendrocyte glycoprotein (MOG) immunization. Infection caused much lesser inflammatory infiltration and demyilination in the CNS of infected EAE mice than uninfected EAE mice. The reduced infiltration was also suggested by the expressions of the inflammation cytokines, IL-17, IL-6, IL-1β, IFN-γ, and TNF-α, which were high in the spinal cords of the uninfected EAE mice, but was nearly normal or low in the infected EAE mice. The increased production of MOG-induced IL-17 and IFN-γ and the expression of IL-6, IL-1β, TGF-β in splenocytes after restimulation with MOG was inhibited in the infected EAE mice. On the other hand, the greatly induced Th2 response was observed in the splenocytes of the infected EAE mice. The present study showed that T. pseudospiralis infection can suppresses EAE by reducing the inflammatory infiltration in CNS, likely associated with the suppression of Th17 and Th1 responses by the infection.     

Essential role of TGF-beta in the natural resistance to experimental allergic encephalomyelitis in rats

Experimental allergic encephalomyelitis (EAE) is a T cell-mediated autoimmune disease induced in susceptible rat strains by a single immunization with myelin basic protein (MBP). The Lewis (LEW) strain is susceptible to disease induction while the Brown Norway (BN) strain is resistant. This resistance involves non-MHC genes since congenic BN-1L rats, with LEW MHC on a BN-derived background, are also resistant. In the present study we show that, upon immunization with MBP, the non-MHC-encoded resistance to develop clinical EAE in BN-1L rats is associated with a decreased production of IFN-gamma. This may be due to a difference between LEW and BN-1L rats in their ability to produce regulatory cytokines such as IL-4, IL-10 and TGF-beta. In comparison to LEW rats, immune lymph node cells from BN-1L rats express an increased amount of IL-4 mRNA but produce less IL-10. Furthermore, the sera from BN-1L rats contain higher amounts of active TGF-beta1. Therefore, we have investigated the involvement of IL-4 and TGF-beta in the resistance of BN-1L rats to develop EAE using neutralizing mAb. Neutralization of TGF-beta, but not IL-4, renders BN-1L rats susceptible to clinical EAE without affecting the proliferation or the cytokine repertoire of immune lymph node cells. With respect to the origin of the endogenous TGF-beta production, we excluded the involvement of CD8 T cells and discuss a possible role of platelets and of CD4 T cells exhibiting the CD45RC(low) phenotype.     

Kinetics of IL-17- and interferon-γ-producing PLPp-specific CD4 T cells in EAE induced by coinjection of PLPp/IFA with pertussis toxin in SJL mice

Systemic administration of Pertussis toxin (PTX) abrogates T cell tolerance mediated by injection of neuroantigens in incomplete Freund's adjuvant (IFA) and causes experimental autoimmune encephalomyelitis (EAE). PTX concomitantly induces high frequencies of neuroantigen-specific IFN-γ- and IL-17-producing T cells. Both IL-17 and IFN-γ have been implicated as a key effector cytokines in the pathogenesis of EAE, possibly with different functions. We therefore investigated potential differences in the temporal and spatial kinetics of the PTX-induced neuroantigen-specific IFN-γ- and IL-17-producing T cell effector populations. IFN-γ- and IL-17-producing PLPp-specific T cells initially arose in comparable frequencies in the local draining lymph nodes (drLN) after immunization as measured by cytokine ELISPOT. High frequencies of both IFN-γ- and IL-17-producing T cells were present in the immune periphery before onset of EAE. The highest frequencies of PTX-induced IFN-γ- and IL-17-producing PLPp-specific cells coincided in the inflamed CNS during acute EAE. During recovery, both IFN-γ- and IL-17-producing PLPp-specific T cells simultaneously disappeared from the CNS, whereas high frequencies of these cells remained present in the immune periphery. The functional affinity of both IFN-γ- and IL-17-producing T cells did not change during EAE. Therefore, autoimmune pathology in this model did not correlate with specific PTX effects either on Th1 or Th17 cells regarding their kinetics and CNS migration.     

Cytokine expression and synovial pathology in the initiation and spontaneous resolution phases of adjuvant arthritis: interleukin-17 expression is upregulated in early disease

The aim of this study was to understand the immune processes controlling the initiation and spontaneous resolution of adjuvant arthritis (AA). We investigated synovial T-cell recruitment and mRNA expression of IL-17 and other important disease related cytokines, IFN-gamma, IL-2, IL-4, TNF and TGF-beta in inguinal lymph node (ILN) and synovial membrane (SM). Arthritis severity was assessed by a numerical rating score and rats were sacrificed every 3--4 days postadjuvant induction. Further assessment involved quantitative radiology and histology of the ankle joints on each day, and the ILN and SM were removed for RNA extraction. Cytokine mRNA expression was measured using RT-PCR and densitometry. Paraffin sections of rat ankle joints were stained for T-cells (CD3) by immunohistochemistry. In the ILN, there was an increase in IL-17, TNF and IFN-gamma expression in the early stages of disease, with a secondary sustained increase in IFN-gamma expression. In the SM, there was expression of T-cell cytokines in early arthritis (day 13), and prolonged TNF and TGF-beta expression, which reflected disease progression. IL-4 mRNA expression increased in the later stages of AA. Synovial T-cell numbers transiently increased at day 6, and remained high from days 13--28. Increased pro-inflammatory cytokine expression, including IL-17, in the ILN reflects the initiating events in the early stage of disease. IL-17 may therefore play an important role in the pathogenesis of AA. The increase in IL-4 (an anti-inflammatory cytokine) in the SM in the later stages of AA suggests that IL-4 is involved in the spontaneous resolution of AA. The initial increase in IFN-gamma in the ILN may reflect a pro-inflammatory response, while the prolonged secondary increase may indicate activation of regulatory T-cells.