Interleukin‐12 (IL‐12, p70) a heterodimeric cytokine of p40 and p35 subunits, important for Th1‐type immune responses, has been attributed a prominent role in multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). Recently, the related heterodimeric cytokine, IL‐23, composed of the same p40 subunit as IL‐12 and a unique p19 subunit, was shown to be involved in Th1 responses and EAE. We investigated whether astrocytes and microglia, CNS cells with antigen‐presenting cell (APC) function can present antigen to myelin basic protein (MBP)‐reactive T cells, and whether this presentation is blocked with antibodies against IL‐12/IL‐23p40. Interferon (IFN)‐γ‐treated APC induced proliferation of MBP‐reactive T cells. Anti‐IL‐12/IL‐23p40 antibodies blocked this proliferation. These results support and extend our previous observation that astrocytes and microglia produce IL‐12/IL‐23p40. Moreover, we show that stimulated astrocytes and microglia produce biologically active IL‐12p70. Because IL‐12 and IL‐23 share p40, we wanted to determine whether astrocytes also express IL‐12p35 and IL‐23p19, as microglia were already shown to express them. Astrocytes expressed IL‐12p35 mRNA constitutively, and IL‐23 p19 after stimulation. Thus, astrocytes, under inflammatory conditions, express all subunits of IL‐12/IL‐23. Their ability to present antigen to encephalitogenic T cells can be blocked by neutralizing anti‐IL‐12/IL‐23p40 antibodies.

Summary: Using a high throughput gene microarray technology that detects ∼22 000 genes, we found that arginase I was the most significantly up‐regulated gene in the murine spinal cord during experimental autoimmune encephalomyelitis (EAE). By Northern blot and arginase enzyme assay, we detected high levels of arginase I mRNA and protein, respectively, in the spinal cord of EAE mice, but not in the spinal cord of normal mice or mice that had recovered from EAE. In vitro, both microglia and astrocytes produced arginase and nitric oxide synthase, two enzymes that are involved in arginine metabolism. To explore the roles of arginase in EAE, we injected the arginase inhibitor amino‐6‐boronohexanoic acid (ABH) into mice during the inductive and effector phases of the disease. Compared with mice that received vehicle control, mice treated with ABH developed milder EAE with delayed onset, reduced disease score and expedited recovery. Spleen mononuclear cells from ABH‐treated mice produced more nitric oxide and secreted less interferon‐γ and tumour necrosis factor‐α as compared to control mice. These results indicate that arginase plays important roles in autoimmune inflammation in the central nervous system.

Introduction: The TAM (tyro 3, axl, mer) kinases are key regulators of innate immunity and are important in the phagocytosis of apoptotic cells. Gas6 and protein S are ligands for these TAM kinases and bind to phosphatidyl serine residues exposed during apoptosis. In animal models, absence of TAM kinases is associated with lupus-like disease. To test whether human systemic lupus erythematosus (SLE) patients might have deficient levels of TAM ligands, we measured Gas 6 and protein S levels in SLE. Methods: 107 SLE patients were recruited. Of these, 45 SLE patients were matched age, gender and ethnicity with normal controls (NC). Gas6 and free protein S were measured with sandwich enzyme linked immunosorbent assays (ELISAs). Results: Overall, the plasma concentrations of Gas6 and free protein S were not different between 45 SLE patients and 45 NC. In SLE patients, the levels of free protein S were positively correlated with age (r = 0.2405, P = 0.0126), however those of Gas6 were not. There was no correlation between the concentrations of Gas6 and free protein S in individuals. Levels of free protein S were significantly lower in SLE patients with a history of serositis, neurologic disorder, hematologic disorder and immunologic disorder. Gas6 levels were elevated in patients with a history of neurologic disorder. The SLE patients with anti-Sm or anti-cardiolipin IgG showed lower free protein S levels. Circulating free protein S was positively correlated with complement component 3 (C3) (r = 0.3858, P < 0.0001) and complement component 4 (C4) (r = 0.4275, P < 0.0001). In the patients with active BILAG hematologic involvement, the levels of free protein S were lower and those of Gas6 were higher. Conclusions: In SLE, free protein S was decreased in patients with certain types of clinical history and disease activity. Levels of free protein S were strongly correlated with C3 and C4 levels. Gas6 levels in SLE patients differed little from levels in NC, but they were elevated in the small numbers of patients with a history of neurological disease. The correlation of decreased protein S levels with lupus disease activity is consistent with a role for the TAM receptors in scavenging apoptotic cells and controlling inflammation. Protein S appears more important functionally in SLE patients than Gas6 in this regard.

Objective: Increased levels of type I interferon (IFN) and type I IFN–regulated genes are found in patients with systemic lupus erythematosus (SLE) and may be central to its pathogenesis. Mitochondrial antiviral signaling protein (MAVS) is a key regulator of type I IFN that undergoes a dramatic prion‐like aggregation and self propagates the activation signal from viral RNA to amplify downstream IFN production. We undertook this study to determine whether such MAVS aggregates might play a role in the sustained increased production of type I IFN in SLE.Methods: Peripheral blood mononuclear cells were isolated and mitochondrial extracts were prepared. MAVS aggregation was detected by semidenatured agarose gel electrophoresis and confirmed by immunofluorescence staining. MAVS‐associated signaling proteins were analyzed by Western blotting. MAVS aggregation–associated gene expression signature was analyzed by microarray.Results: In blood cells from 22 of 67 SLE patients, essentially all MAVS was in a high molecular weight aggregated form. None of 6 rheumatoid arthritis patients and only 3 of 33 healthy controls had abnormal MAVS. Compared to MAVS aggregate–negative patients, MAVS aggregate–positive SLE patients had significantly higher serum levels of IFNβ and significantly increased levels of autoantibodies against Sm and U1 RNP. Gene array data revealed a characteristic gene expression pattern in these patients, with altered expression of genes involved in IFN signaling and membrane trafficking.Conclusion: Persistent MAVS aggregates may lead to increased type I IFN production and result in unmitigated signals leading to autoimmunity.

Angiotensin converting enzyme (ACE)¹ mediates inflammation, participates in T cell stimulation by certain antigenic peptides, and influences the permeability of the blood brain barrier (BBB). ACE is elevated in multiple sclerosis (MS), an autoimmune disease of the central nervous system (CNS), characterized by increased BBB permeability. ACE inhibitor captopril suppresses certain immune functions and inhibits inflammatory or autoimmune diseases. We studied the effect of captopril on Lewis rat EAE, an animal model of MS. Fourteen rats with EAE were treated with captopril 30 mg/kg daily from immunization to day 21 post-immunization, and compared with 14 untreated rats. Severity scores and lymphocyte reactivity to myelin basic protein and mitogen were measured. There was a statistically significant (p< 0.05) difference between the mean and cumulative clinical scores of captopril-treated and untreated animals. Lymphocytes from captopril treated EAE rats at the peak of disease severity had diminished responses to MBP and concanavalin A. The data suggest a significant beneficial effect of captopril in Lewis rat EAE. Further studies including other inhibitors of ACE or of other peptidases with immune, inflammatory or BBB role, may identify potentially valuable immunopharmacologic agents.