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Suppressor of cytokine signaling (SOCS) proteins have emerged as important regulators of cytokine signals in lymphocytes. In this study, we have investigated regulation of SOCS expression and their role in Th cell growth and differentiation. We show that SOCS genes are constitutively expressed in naive Th cells, albeit at low levels, and are differentially induced by Ag and Th-polarizing cytokines. Whereas cytokines up-regulate expression of SOCS1, SOCS2, SOCS3, and cytokine-induced Src homology 2 protein, Ags induce down-regulation of SOCS3 within 48 h of Th cell activation and concomitantly up-regulate SOCS1, SOCS2, and cytokine-induced Src homology 2 protein expression. We further show that STAT1 signals play major roles in inducing SOCS expression in Th cells and that induction of SOCS expression by IL-4, IL-12, or IFN-gamma is compromised in STAT1-deficient primary Th cells. Surprisingly, IL-4 is a potent inducer of STAT1 activation in Th2 but not Th1 cells, and SOCS1 or SOCS3 expression is dramatically reduced in STAT1(-/-) Th2 cells. To our knowledge, this is the first report of IL-4-induced STAT1 activation in Th cells, and suggests that its induction of SOCS, may in part, regulate IL-4 functions in Th2 cells. In fact, overexpression of SOCS1 in Th2 cells represses STAT6 activation and profoundly inhibits IL-4-induced proliferation, while depletion of SOCS1 by an anti-sense SOCS1 cDNA construct enhances cell proliferation and induces constitutive activation of STAT6 in Th2 cells. These results are consistent with a model where IL-4 has dual effects on differentiating T cells: it simulates proliferation/differentiation through STAT6 and autoregulates its effects on Th2 growth and effector functions via STAT1-dependent up-regulation of SOCS proteins.

Positive regulatory factors induced by IL-12/STAT4 and IL-4/STAT6 signaling during T cell development contribute to polarized patterns of cytokine expression manifested by differentiated Th cells. These two critical and antagonistic signaling pathways are under negative feedback regulation by a multimember family of intracellular proteins called suppressor of cytokine signaling (SOCS). However, it is not known whether these negative regulatory factors also modulate Th1/Th2 lineage commitment and maintenance. We show here that CD4(+) naive T cells constitutively express low levels of SOCS1, SOCS2, and SOCS3 mRNAs. These mRNAs and their proteins increase significantly in nonpolarized Th cells after activation by TCR signaling. We further show that differentiation into Th1 or Th2 phenotype is accompanied by preferential expression of distinct SOCS mRNA transcripts and proteins. SOCS1 expression is 5-fold higher in Th1 than in Th2 cells, whereas Th2 cells contain 23-fold higher levels of SOCS3. We also demonstrate that IL-12-induced STAT4 activation is inhibited in Th2 cells that express high levels of SOCS3 whereas IL-4/STAT6 signaling is constitutively activated in Th2 cells, but not Th1 cells, with high SOCS1 expression. These results suggest that mutually exclusive use of STAT4 and STAT6 signaling pathways by differentiated Th cells may derive in part, from SOCS3- or SOCS1-mediated repression of IL-12/STAT4- or IL-4/STAT6 signaling in Th2 and Th1 cells, respectively. Given the strong correlation between distinct patterns of SOCS expression and differentiation into the Th1 or Th2 phenotype, SOCS1 and SOCS3 proteins are therefore Th lineage markers that can serve as therapeutic targets for immune modulation therapy.

Immunological responses to pathogens are stringently regulated in the eye to prevent excessive inflammation that damage ocular tissues and compromise vision. Suppressors of cytokine signaling (SOCS) regulate intensity/duration of inflammatory responses. We have used SOCS1-deficient mice and retina-specific SOCS1 transgenic rats to investigate roles of SOCS1 in ocular herpes simplex virus (HSV-1) infection and non-infectious uveitis. We also genetically engineered cell-penetrating SOCS proteins (membrane-translocating sequence (MTS)-SOCS1, MTS-SOCS3) and examined whether they can be used to inhibit inflammatory cytokines. Overexpression of SOCS1 in transgenic rat eyes attenuated ocular HSV-1 infection while SOCS1-deficient mice developed severe non-infectious anterior uveitis, suggesting that SOCS1 may contribute to mechanism of ocular immune privilege by regulating trafficking of inflammatory cells into ocular tissues. Furthermore, MTS-SOCS1 inhibited IFN-γ-induced signal transducers and activators of transcription 1 (STAT1) activation by macrophages while MTS-SOCS3 suppressed expansion of pathogenic Th17 cells that mediate uveitis, indicating that MTS-SOCS proteins maybe used to treat ocular inflammatory diseases of infectious or autoimmune etiology. [ABSTRACT FROM AUTHOR]

Suppressors of cytokine signaling (SOCS) have been implicated in regulation of T-cell activation and cytokine-mediated differentiation of T-helper cells. In this study we have characterized the pattern of SOCS expression in naive and activated primary T-helper cells, examined whether expression of SOCS genes is regulated by cytokine or T-cell receptor signaling, and analyzed the function of SOCS in differentiated T-cells. We show that SOCS1, SOCS2, SOCS3, CIS (cytokine-induced SH2 protein) genes are constitutively expressed in naive T-helper cells, with SOCS3 being the most abundant. Antigen stimulation of naive T-helper cells down-regulates SOCS3 expression and concomitantly up-regulates SOCS1, SOCS2, and CIS gene transcription, suggesting that SOCS genes are regulated differentially by T-cell activation. Down-regulation of SOCS3 expression is subsequently followed by gradual increase in SOCS3 level and corresponding decline in interleukin 2 (IL-2) secretion. In fact, SOCS3 mRNA levels are inversely correlated with the amount of IL-2 secretion and proliferative responses of differentiating T-helper cells, suggesting mutually antagonistic effects of SOCS3 and IL-2 and feedback regulation of T-cell activation by SOCS3. Furthermore, the degree of SOCS3 inhibition is antigen concentration-dependent and is mediated in part by growth factor independence-1, a T-cell transcription factor that regulates S-phase entry in T-cells. Forced overexpression of SOCS3 inhibits proliferation of T-helper cells, whereas depletion of endogenous SOCS3 by antisense SOCS3 cDNA enhances T-cell receptor- and cytokine-induced proliferation. Taken together, these results suggest a role for SOCS3 in maintaining T-helper cells in a quiescent state. Transient inhibition of SOCS3 by antigen stimulation may therefore be essential in allowing activation of resting T-cells. [ABSTRACT FROM AUTHOR]

Suppressors of cytokine signaling (SOCS) proteins are negative-feedback regulators of the JAK/STAT pathway, and SOCS3 contributes to host immunity by regulating the intensity and duration of cytokine signals and inflammatory responses. Mice with Socs3 deletion in myeloid cells exhibit enhanced STAT3 signaling, expansion of Th1 and Th17 cells, and develop severe experimental autoimmune encephalomyelitis. Interestingly, development of the unique IL-17/IFN-γ double-producing (Th17/IFN-γ and Tc17/IFN-γ) subsets that exhibit strong cytotoxic activities and are associated with pathogenesis of several autoimmune diseases has recently been shown to depend on epigenetic suppression of SOCS3 expression, further suggesting involvement of SOCS3 in autoimmunity and tumor immunity. In this study, we generated mice with Socs3 deletion in the CD4 T cell compartment (CD4-SOCS3 knockout [KO]) to determine in vivo effects of the loss of Socs3 in the T cell-mediated autoimmune disease, experimental autoimmune uveitis (EAU). In contrast to the exacerbation of experimental autoimmune encephalomyelitis in myeloid-specific SOCS3-deleted mice, CD4-SOCS3KO mice were protected from acute and chronic uveitis. Protection from EAU correlated with enhanced expression of CTLA-4 and expansion of IL-10-producing regulatory T cells with augmented suppressive activities. We further show that SOCS3 interacts with CTLA-4 and negatively regulates CTLA-4 levels in T cells, providing a mechanistic explanation for the expansion of regulatory T cells in CD4-SOCS3 during EAU. Contrary to in vitro epigenetic studies, Th17/IFN-γ and Tc17/IFN-γ populations were markedly reduced in CD4-SOCS3KO, suggesting that SOCS3 promotes expansion of the Th17/IFN-γ subset associated with development of severe uveitis. Thus, SOCS3 is a potential therapeutic target in uveitis and other autoinflammatory diseases. [ABSTRACT FROM AUTHOR]

IL-12 family cytokines are important in host immunity. Whereas some members (IL-12, IL-23) play crucial roles in pathogenesis of organ-specific autoimmune diseases by inducing the differentiation of Th1 and Th17 lymphocytes, others (IL-27 and IL-35) suppress inflammatory responses and limit tissue injury induced by these T cell subsets. In this study, we have genetically engineered a novel IL27p28/ IL12p40 heterodimeric cytokine (p28/p40) that antagonizes signaling downstream of the gp130 receptor. We investigated whether p28/p40 can be used to ameliorate uveitis, a CNS inflammatory disease. Experimental autoimmune uveitis (EAU) is the mouse model of human uveitis and is mediated by Th1 and Th17 cells. We show here that p28/p40 suppressed EAU by inhibiting the differentiation and inflammatory responses of Th1 and Th17 cells while promoting expansion of IL-+- and Foxp3+-expressing regulatory T cells. Lymph node cells from mice treated with p28/p40 blocked adoptive transfer of EAU to naïve syngeneic mice by immunopathogenic T cells and suppressive effects of p28/p40 derived in part from antagonizing STAT1 and STAT3 pathways induced by IL-27 and IL-6. Interestingly, IL27p28 also suppressed EAU, but to a lesser extent than p28/p40. The inhibition of uveitogenic lymphocyte proliferation and suppression of EAU by p28/p40 and IL27p28 establish efficacy of single chain and heterodimeric IL-12 family cytokines in treatment of a CNS autoimmune disease. Creation of the biologically active p28/p40 heterodimeric cytokine represents an important proof-of-concept experiment, suggesting that cytokines comprising unique IL-12 α- and β-subunit pairing may exist in nature and may constitute a new class of therapeutic cytokines. [ABSTRACT FROM AUTHOR]

Uveitis is a diverse group of potentially sight-threatening intraocular inflammatory diseases and pathology derives from sustained production of pro-inflammatory cytokines in the optical axis. Although topical or systemic steroids are effective therapies, their adverse effects preclude prolonged usage and are impetus for seeking alternative immunosuppressive agents, particularly for patients with refractory uveitis. In this study, we synthesized a 16 amino acid membrane-penetrating lipophilic suppressor of cytokine signaling 1 peptide (SOCS1-KIR) that inhibits JAK/STAT signaling pathways and show that it suppresses and ameliorates experimental autoimmune uveitis (EAU), the mouse model of human uveitis. Fundus images, histological and optical coherence tomography analysis of eyes showed significant suppression of clinical disease, with average clinical score of 0.5 compared to 2.0 observed in control mice treated with scrambled peptide. We further show that SOCS1-KIR conferred protection from ocular pathology by inhibiting the expansion of pathogenic Th17 cells and inhibiting trafficking of inflammatory cells into the neuroretina during EAU. Dark-adapted scotopic and photopic electroretinograms further reveal that SOCS1-KIR prevented decrement of retinal function, underscoring potential neuroprotective effects of SOCS1-KIR in uveitis. Importantly, SOCS1-KIR is non-toxic, suggesting that topical administration of SOCS1-Mimetics can be exploited as a non-invasive treatment for uveitis and for limiting cytokine-mediated pathology in other ocular inflammatory diseases including scleritis. [ABSTRACT FROM AUTHOR]