Effect of mild stress in mice latently infected pseudorabies virus.

Stress is one of the important factors that induces reactivation of pseudorabies virus (PrV) in latently infected pigs. We established a murine model of latent PrV infection and examined the effects of mild stress treatment in order to demonstrate that this model simulates natural infection in the pig. Latently infected mice excreted PrV from the nasal cavity under stress treatments consisting of restraint, exposure to cold or transport. Similar reactions have been observed upon treatment with acetylcholine and dexamethasone. The present findings demonstrate that these kinds of mild stress reactivate the virus in murine latent infection models in a manner similar to the induction of latent infection in pigs in the field.     

Lack of Interleukin-6 (IL-6) Enhances Susceptibility to Infection but Does Not Alter Latency or Reactivation of Herpes Simplex Virus Type 1 in IL-6 Knockout Mice

The ability of the pleotropic, proinflammatory cytokine interleukin-6 (IL-6) to affect the replication, latency, and reactivation of herpes simplex virus type 1 (HSV-1) in cell culture and in IL-6 knockout (KO) mice was studied. In initial studies, we found no effect of exogenous IL-6, monoclonal antibodies to IL-6, or monoclonal antibody to the IL-6 coreceptor, gp130, on HSV-1 replication in vitro by plaque assay or reactivation ex vivo by explant cocultivation of latently infected murine trigeminal ganglia (TG). Compared with the wild-type (WT) mice, the IL-6 KO mice were less able to survive an ocular challenge with 10(5) PFU of HSV-1 (McKrae) (40% survival of WT and 7% survival KO mice; P = 0.01). There was a sixfold higher 50% lethal dose of HSV-1 in WT than IL-6 KO mice (1.7 x 10(4) and 2.7 x 10(3) PFU, respectively). No differences were observed in titers of virus recovered from the eyes, TG, or brains or in the rates of virus reactivation by explant cocultivation of TG from latently infected WT or KO mice. Exposure of latently infected mice to UV light resulted in comparable rates of reactivation and in the proportions of WT and KO animals experiencing reactivation. Moreover, quantitative PCR assays showed nearly identical numbers of HSV-1 genomes in latently infected WT and IL-6 KO mice. These studies indicate that while IL-6 plays a role in the protection of mice from lethal HSV infection, it does not substantively influence HSV replication, spread to the nervous system, establishment of latency, or reactivation.     

Activation of latent pseudorabies virus infection in mice treated with acetylcholine.

Pseudorabies virus (PrV) YS-81 strain latently infected in 6-week-old BALB/c mice was detectable by nasal swabbing, and serum antibody was shown to increase in titer after intraperitoneal injection for 3 days with acetylcholine or dexamethasone.     

Lipopolysaccharide, tumor necrosis factor alpha, or interleukin-1beta triggers reactivation of latent cytomegalovirus in immunocompetent mice

We have previously shown that cytomegalovirus (CMV) can reactivate in lungs of nonimmunosuppressed patients during critical illness. Our recent work has shown that polymicrobial bacterial sepsis can trigger reactivation of latent murine CMV (MCMV). We hypothesize that MCMV reactivation following bacterial sepsis may be caused by inflammatory mediators. To test this hypothesis, BALB/c mice latently infected with Smith strain MCMV received sublethal intraperitoneal doses of lipopolysaccharide (LPS), tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), or saline. Lung tissue homogenates were evaluated for viral reactivation 3 weeks after mediator injection. Because LPS is known to signal via Toll-like receptor 4 (TLR-4) in mice, further studies blocking this signaling mechanism were performed using monoclonal MTS510. Finally, mice were tested with intravenous TNF-alpha to determine whether this would cause reactivation. All mice receiving sublethal intraperitoneal doses of LPS, TNF-alpha, or IL-1beta had pulmonary reactivation of latent MCMV 3 weeks following injection, and LPS caused MCMV reactivation with kinetics similar to those for sepsis. When TLR-4 signaling was blocked, exogenous LPS did not reactivate latent MCMV. Intravenous TNF-alpha administration at near-lethal doses did not reactivate MCMV. Exogenous intraperitoneal LPS, TNF-alpha, and IL-1beta are all capable of reactivating CMV from latency in lungs of previously healthy mice. LPS reactivation of MCMV appears dependent on TLR-4 signaling. Interestingly, intravenous TNF-alpha did not trigger reactivation, suggesting possible mechanistic differences that are discussed. We conclude that inflammatory disease states besides sepsis may be capable of reactivating CMV from latency.     

Establishment of HSV1 Latency in Immunodeficient Mice Facilitates Efficient In Vivo Reactivation

The establishment of latent infections in sensory neurons is a remarkably effective immune evasion strategy that accounts for the widespread dissemination of life long Herpes Simplex Virus type 1 (HSV1) infections in humans. Periodic reactivation of latent virus results in asymptomatic shedding and transmission of HSV1 or recurrent disease that is usually mild but can be severe. An in-depth understanding of the mechanisms regulating the maintenance of latency and reactivation are essential for developing new approaches to block reactivation. However, the lack of a reliable mouse model that supports efficient in vivo reactivation (IVR) resulting in production of infectious HSV1 and/or disease has hampered progress. Since HSV1 reactivation is enhanced in immunosuppressed hosts, we exploited the antiviral and immunomodulatory activities of IVIG (intravenous immunoglobulins) to promote survival of latently infected immunodeficient Rag mice. Latently infected Rag mice derived by high dose (HD), but not low dose (LD), HSV1 inoculation exhibited spontaneous reactivation. Following hyperthermia stress (HS), the majority of HD inoculated mice developed HSV1 encephalitis (HSE) rapidly and synchronously, whereas for LD inoculated mice reactivated HSV1 persisted only transiently in trigeminal ganglia (Tg). T cells, but not B cells, were required to suppress spontaneous reactivation in HD inoculated latently infected mice. Transfer of HSV1 memory but not OVA specific or naïve T cells prior to HS blocked IVR, revealing the utility of this powerful Rag latency model for studying immune mechanisms involved in control of reactivation. Crossing Rag mice to various knockout strains and infecting them with wild type or mutant HSV1 strains is expected to provide novel insights into the role of specific cellular and viral genes in reactivation, thereby facilitating identification of new targets with the potential to block reactivation.     

Alpha/beta interferons regulate murine gammaherpesvirus latent gene expression and reactivation from latency

Alpha/beta interferon (IFN-alpha/beta) protects the host from virus infection by inhibition of lytic virus replication in infected cells and modulation of the antiviral cell-mediated immune response. To determine whether IFN-alpha/beta also modulates the virus-host interaction during latent virus infection, we infected mice lacking the IFN-alpha/beta receptor (IFN-alpha/betaR(-/-)) and wild-type (wt; 129S2/SvPas) mice with murine gammaherpesvirus 68 (gammaHV68), a lymphotropic gamma-2-herpesvirus that establishes latent infection in B cells, macrophages, and dendritic cells. IFN-alpha/betaR(-/-) mice cleared low-dose intranasal gammaHV68 infection with wt kinetics and harbored essentially wt frequencies of latently infected cells in both peritoneum and spleen by 28 days postinfection. However, latent virus in peritoneal cells and splenocytes from IFN-alpha/betaR(-/-) mice reactivated ex vivo with >40-fold- and 5-fold-enhanced efficiency, respectively, compared to wt cells. Depletion of IFN-alpha/beta from wt mice during viral latency also significantly increased viral reactivation, demonstrating an antiviral function of IFN-alpha/beta during latency. Viral reactivation efficiency was temporally regulated in both wt and IFN-alpha/betaR(-/-) mice. The mechanism of IFN-alpha/betaR action was distinct from that of IFN-gammaR, since IFN-alpha/betaR(-/-) mice did not display persistent virus replication in vivo. Analysis of viral latent gene expression in vivo demonstrated specific upregulation of the latency-associated gene M2, which is required for efficient reactivation from latency, in IFN-alpha/betaR(-/-) splenocytes. These data demonstrate that an IFN-alpha/beta-induced pathway regulates gammaHV68 gene expression patterns during latent viral infection in vivo and that IFN-alpha/beta plays a critical role in inhibiting viral reactivation during latency.     

Mechanisms for macrophage-mediated HIV-1 induction

Viral latency is a long-term pathogenic condition in patients infected with HIV-1. Low but sustained virus replication in chronically infected cells can be activated by stimulation with proinflammatory cytokines such as TNF-alpha, IL-1 beta, or other host factors. However, the precise mechanism by which cellular activation induces latently infected cells to produce virions has remained unclear. In the present report, we present evidence that activation of HIV-1 replication in latently infected U1 or ACH2 cells by human macrophages is mediated by a rapid nuclear localization of NF-kappaB p50/p65 dimer with concomitant increased expression of proinflammatory cytokines. Multiplexed RT-PCR amplification of mRNA isolated from cocultures of macrophages and U1 and ACH2 cells showed significant induction of IL-1beta, IL-6, IL-8, TNF-alpha, and TGF-beta expression within 3 h of coincubation. Fixation of macrophages, U-1, or ACH2 cells with paraformaldehyde before coculture completely abrogated the induction of NF-kappaB subunits and HIV-1 replication, suggesting that cooperative interaction between the two cell types is an essential process for cellular activation. Pretreatment of macrophage-U1 or macrophage-ACH2 cocultures with neutralizing anti-TNF-alpha Ab down-regulated the replication of HIV-1. In addition, pretreatment of macrophage-U1 or macrophage-ACH2 cocultures with the NF-kappaB inhibitor (E)3-[(4-methylphenyl)sulfonyl]-2-propenenitrile (BAY 11-7082) prevented the induction of cytokine expression, indicating a pivotal role of NF-kappaB-mediated signaling in the reactivation of HIV-1 in latently infected cells by macrophages. These results provide a mechanism by which macrophages induce HIV-1 replication in latently infected cells.     

Ex vivo stimulation of B cells latently infected with gammaherpesvirus 68 triggers reactivation from latency

Murine gammaherpesvirus 68 (gammaHV68) infection of mice results in the establishment of a chronic infection, which is largely maintained through latent infection of B lymphocytes. Acute virus replication is almost entirely cleared by 2 weeks postinfection. Spontaneous reactivation of gammaHV68 from latently infected splenocytes upon ex vivo culture can readily be detected at the early stages of infection (e.g., day 16). However, by 6 weeks postinfection, very little spontaneous reactivation is detected upon explant into tissue culture. Here we report that stimulation of latently infected splenic B cells harvested at late times postinfection with cross-linking surface immunoglobulin (Ig), in conjunction with anti-CD40 antibody treatment, triggers virus reactivation. As expected, this treatment resulted in B-cell activation, as assessed by upregulation of CD69 on B cells, and ultimately B-cell proliferation. Since anti-Ig/anti-CD40 stimulation resulted in splenic B-cell proliferation, we assessed whether this reactivation stimulus could overcome the previously characterized defect in virus reactivation of a v-cyclin null gammaHV68 mutant. This analysis demonstrated that anti-Ig/anti-CD40 stimulation could drive reactivation of the v-cyclin null mutant virus in latently infected splenocytes, but not to the levels observed with wild-type gammaHV68. Thus, there appears to be a role for the v-cyclin in B cells following anti-Ig/anti-CD40 stimulation independent of the induction of the cell cycle. Finally, to assess signals that are not mediated through the B-cell receptor, we demonstrate that addition of lipopolysaccharide to explanted splenocyte cultures also enhanced virus reactivation. These studies complement and extend previous analyses of Epstein-Barr virus and Kaposi's sarcoma-associated virus reactivation from latently infected cell lines by investigating reactivation of gammaHV68 from latently infected primary B cells recovered from infected hosts.     

The Probability of In Vivo Reactivation of Herpes Simplex Virus Type 1 Increases with the Number of Latently Infected Neurons in the Ganglia

The purpose of this study was to define the relationship between herpes simplex virus (HSV) latency and in vivo ganglionic reactivation. Groups of mice with numbers of latently infected neurons ranging from 1.9 to 24% were generated by varying the input titer of wild-type HSV type 1 strain 17syn+. Reactivation of the virus in mice from each group was induced by hyperthermic stress. The number of animals that exhibited virus reactivation was positively correlated with the number of latently infected neurons in the ganglia over the entire range examined (r = 0.9852, P < 0.0001 [Pearson correlation]).     

ICP0 is required for efficient reactivation of herpes simplex virus type 1 from neuronal latency

Relative to wild-type herpes simplex virus type 1 (HSV-1), ICP0-null mutant viruses reactivate inefficiently from explanted, latently infected mouse trigeminal ganglia (TG), indicating that ICP0 is not essential for reactivation but plays a central role in enhancing the efficiency of reactivation. The validity of these findings has been questioned, however, because the replication of ICP0-null mutants is impaired in animal models during the establishment of latency, such that fewer mutant genomes than wild-type genomes are present in latently infected mouse TG. Therefore, the reduced number of mutant viral genomes available to reactivate, rather than mutations in the ICP0 gene per se, may be responsible for the reduced reactivation efficiency of ICP0-null mutants. We have recently demonstrated that optimization of the size of the ICP0 mutant virus inoculum and transient immunosuppression of mutant-infected mice with cyclophosphamide can be used to establish wild-type levels of ICP0-null mutant genomes in latently infected TG (W. P. Halford and P. A. Schaffer, J. Virol. 74:5957-5967, 2000). Using this procedure to equalize mutant and wild-type genome numbers, the goal of the present study was to determine if, relative to wild-type virus, the absence of ICP0 function in two ICP0-null mutants, n212 and 7134, affects reactivation efficiency from (i) explants of latently infected TG and (ii) primary cultures of latently infected TG cells. Although equivalent numbers of viral genomes were present in TG of mice latently infected with either wild-type or mutant viruses, reactivation of n212 and 7134 from heat-stressed TG explants was inefficient (31 and 37% reactivation, respectively) relative to reactivation of wild-type virus (KOS) (95%). Similarly, n212 and 7134 reactivated inefficiently from primary cultures of dissociated TG cells plated directly after removal from the mouse (7 and 4% reactivation, respectively), relative to KOS (60% reactivation). The efficiency and kinetics of reactivation of KOS, n212, and 7134 from cultured TG cells (treated with acyclovir to facilitate the establishment of latency) in response to heat stress or superinfection with a nonreplicating HSV-1 ICP4(-) mutant, n12, were compared. Whereas heat stress induced reactivation of KOS from 69% of latently infected TG cell cultures, reactivation of n212 and 7134 was detected in only 1 and 7% of cultures, respectively. In contrast, superinfection with the ICP4(-) virus, which expresses high levels of ICP0, resulted in the production of infectious virus in nearly 100% of cultures latently infected with KOS, n212, or 7134 within 72 h. Thus, although latent mutant viral genome loads were equivalent to that of wild-type virus, in the absence of ICP0, n212 and 7134 reactivated inefficiently from latently infected TG cells during culture establishment and following heat stress. Collectively, these findings demonstrate that ICP0 is required to induce efficient reactivation of HSV-1 from neuronal latency.     

Autonomic and glucocorticoid associations with the steady-state expression of latent Epstein-Barr virus

Previous studies have demonstrated the impact of psychological stress on the steady-state expression/reactivation of latent Epstein-Barr virus (EBV). Stress-induced decrements in the cellular immune response result in less control over the expression of the latent virus, resulting in increases in antibody to the virus. In Study 1, we investigated whether the steady-state expression of latent EBV in vivo differed between high and low stress reactors, as defined by sympathetic cardiac reactivity. Autonomic activity and antibody titers to Epstein-Barr virus capsid antigen (VCA) were measured in 50 elderly women latently infected with EBV. Results revealed that women who were high stress reactors were characterized by higher antibody titers to the latent virus than low stress reactors. High reactors tended to show larger stress-related increases in cortisol than low reactors, but the differences were not significant. Daily stressors can activate the autonomic nervous system and promote the release of pituitary and adrenal hormones, especially in high reactors. Glucocorticoid hormones have been shown to reactivate EBV in vitro from cells latently infected with the virus. We hypothesized that absolute levels of plasma cortisol may not be the only explanation for stress-induced reactivation of latent EBV and that the diurnal changes in the production of cortisol may be an important factor in these interactions. To examine the feasibility of this hypothesis, an in vitro study was conducted (Study 2) to determine whether changing glucocorticoid concentrations in the medium, in which EBV latently infected cells were cultured, to mimic diurnal changes in plasma cortisol concentrations would enhance the reactivation of the latent virus. Cells latently infected with EBV were exposed to either constant or varying concentrations of the synthetic glucocorticoid hormone dexamethasone (Dex), for 72 h. Results revealed a three- to eightfold enhancement of reactivation of latent EBV in cells pulsed with varying Dex concentrations when compared with cells exposed to a constant and/or a higher mean level of one Dex concentration. Together, these studies raise the possibility that differences in the kinetics of glucocorticoid concentrations may contribute to differences in the reactivation of latent EBV.     

Murine gamma-herpesvirus-68-induced IL-12 contributes to the control of latent viral burden, but also contributes to viral-mediated leukocytosis

Early IFN-alpha/beta production, followed by the development of a viral-specific CTL response, are critical factors in limiting the level of murine gamma-herpesvirus-68 (gammaHV-68) infection. Development of a long-lived CTL response requires T cell help, and these CTLs most likely function to limit the extent of infection following reactivation. The importance of IL-12 in the development and/or activity of Th1 cells and CTLs is well documented, and we investigated the kinetics and magnitude of gammaHV-68-induced IL-12 production. Following intranasal infection, IL-12 and IL-23 mRNA expression was up-regulated in lung and spleen and lung, respectively, followed by increased levels of IL-12p40 in lung homogenates and sera. Exposure of cultured macrophages or dendritic cells to gammaHV-68 induced secretion of IL-12, suggesting that these cells might be responsible for IL-12 production in vivo. gammaHV-68 infection of mice made genetically deficient in IL-12p40 expression (IL-12p40(-/-)) resulted in a leukocytosis and splenomegaly that was significantly less than that observed in syngeneic C57BL/6 mice. IL-12p40(-/-) mice showed increased levels of infectious virus in the lung, but only at day 9 postinfection. Increased levels of latent virus in the spleen at day 15 postinfection were also observed in IL-12p40(-/-) mice when compared with syngeneic C57BL/6 mice. An overall reduction in gammaHV-68-induced IFN-gamma production was observed in IL-12p40(-/-) mice, suggesting that most of the viral-induced IFN-gamma in C57BL/6 mice was IL-12 dependent. Taken together, these results suggest that gammaHV-68-induced IL-12 contributes to the pathophysiology of viral infection while also functioning to limit viral burden.     

The importance of IL-1 beta and TNF-alpha,and the noninvolvement of IL-6, in the development of monoclonal antibody-induced arthritis

Injection of anti-type II collagen Ab and LPS induces arthritis in mice. The levels of IL-1 beta, IL-6, and chemokines (macrophage inflammatory protein (MIP)-1 alpha, MIP-2, and monocyte chemoattractant protein-1) in the hind paws increased with the onset of arthritis and correlated highly with arthritis scores. The level of TNF-alpha was also elevated, but only transiently. Quantitative real-time PCR analysis revealed increases in cytokine and chemokine mRNA. To elucidate the contribution of inflammatory cytokines and chemokines in arthritis development more directly, recombinant proteins, neutralizing Abs, and knockout mice were used. The injection of rIL-1 beta or TNF-alpha, but not IL-6 or chemokines, induced arthritis when mice were i.v. preinjected with anti-type II collagen Ab. However, a single injection of recombinant cytokines or chemokines into the hind paws did not induce swelling. Arthritis development was inhibited by neutralizing Ab against IL-1 beta, TNF-alpha, or MIP-1 alpha. In contrast, the inhibitory effect by anti-MIP-2 Ab was partial and, surprisingly, Abs to IL-6 and monocyte chemoattractant protein-1 showed no inhibitory effect. Furthermore, arthritis development in IL-1R(-/-) mice and TNFR(-/-) mice was not observed at all, but severe arthritis was developed in IL-6(-/-) mice. These results suggest that IL-1 beta and TNF-alpha play more crucial roles than IL-6 or chemokines in this model. Because arthritis was also developed in SCID mice, the development of arthritis in the Ab-induced mice model is due to a mechanism that does not involve T or B cells.     

The Influence of Stress Factors on the Reactivation of Latent Herpes Simplex Virus Type 1 in Infected Mice

This study shows that the influence of different stress factors impacts the reactivation of latent herpes simplex virus type 1 (HSV-1) specifically in the trigeminal ganglion of infected mice. Different stress factors including hyperthermia, hypothermia, fatigue, and immunosuppression were exerted on mice infected with HSV-1. These viral antigens were then detected in the trigeminal ganglion region of infected mice under the influence of each stress factor, with hyperthermia having the most influence on reactivation. Interestingly, an increase in IL-6 was also detected in mice subjected to hyperthermia. These studies therefore suggest that stress can induce the reactivation of latent HSV-1, possibly through the induction of IL-6, in the trigeminal ganglion region of infected mice. This reveals a new insight on the pathogenesis of relapse infection of HSV-1.     

Coxsackievirus B3-induced production of tumor necrosis factor-alpha, IL-1 beta, and IL-6 in human monocytes.

Infections by coxsackievirus B3 (CVB3) have previously been shown to cause acute and chronic myocarditis characterized by a heavy mononuclear leukocyte infiltration and myocyte necrosis. Because clinical and experimental evidence suggested that cardiac damage may result from immunologic rather than viral mechanisms, we examined in this study the in vitro interaction of CVB3 with human monocytes. CVB3 was capable of infecting freshly harvested monocytes as revealed by immunofluorescence and release of infectious virus particles. Virus infection did not reduce monocyte viability but, on the contrary, enhanced spreading and adherence. In a dose-dependent manner, CVB3 stimulated the release of cytokines from monocytes. Whereas a potent production of TNF-alpha, IL-1 beta, and IL-6 was dependent on exposure to infectious CVB3, IFN release was also induced by UV-inactivated virus. On a molecular level, CVB3 stimulated cytokine gene expression as shown by a marked TNF-alpha, IL-1 beta, and IL-6 mRNA accumulation. Supernatants of CVB3-infected monocytes displayed cytotoxic activity against Girardi heart cells which could be abrogated by an anti-TNF-alpha antiserum. These data suggest that CVB3-induced cytokine release from monocytes may participate in virus-induced organ damage such as myocarditis, which may either occur by a direct cytotoxicity of cytokines or by activation of cytotoxic lymphocytes.     

IL-1 beta-deficient mice are resistant to induction of experimental SLE

IL-1 is one of the most pleiotropic pro-inflammatory and immunostimulatory cytokines. Overproduction of IL-1 has been shown to be involved in the pathogenicity of various autoimmune inflammatory diseases, including systemic lupus erythematosus (SLE). However, the different contributions that the IL-1 agonistic molecules make in their in vivo native milieu, IL-1beta which is mainly secreted against IL-1alpha which is mainly cell-associated, have not been established. Experimental SLE can be induced in mice by injection with monoclonal anti-DNA antibodies bearing a major idiotype designated, 16/6Id. In the present study, experimental SLE was induced in mice deficient in specific IL-1 molecules, i.e. IL-1alpha(-/-), IL-1beta(-/-), IL-1alpha/beta(-/-) (double KO) and in control BALB/c mice. Mice deficient in IL-1beta , i.e. IL-1beta(-/-) and IL-1alpha/beta(-/-) mice, developed lower levels of anti-dsDNA antibodies after immunization with 16/6Id, as compared to IL-1alpha(-/-) or control BALB/c mice. Disease manifestations were milder in mice deficient in IL-1beta expression. The representative cytokine cascade that is characteristic of overt experimental SLE was also shown to be reduced in groups of mice that lacked IL-1beta as compared to mice deficient in IL-1alpha, which is mainly cell-associated. Altogether, our results point to the importance of secretable IL-1beta, rather than cell-associated IL-1alpha, in the immunostimulatory and inflammatory phenomena that mediate the pathogenesis of experimental SLE.     

IL-1 receptor antagonist-mediated therapeutic effect in murine myasthenia gravis is associated with suppressed serum proinflammatory cytokines, C3, and anti-acetylcholine receptor IgG1

In myasthenia gravis (MG), TNF and IL-1beta polymorphisms and high serum levels of these proinflammatory cytokines have been observed. Likewise, TNF and IL-1beta are critical for the activation of acetylcholine receptor (AChR)-specific T and B cells and for the development of experimental autoimmune myasthenia gravis (EAMG) induced by AChR immunization. We tested the therapeutic effect of human recombinant IL-1 receptor antagonist (IL-1ra) in C57BL/6 mice with EAMG. Multiple daily injections of 0.01 mg of IL-1ra administered for 2 wk following two AChR immunizations decreased the incidence and severity of clinical EAMG. Furthermore, IL-1ra treatment of mice with ongoing clinical EAMG reduced the clinical symptoms of disease. The IL-1ra-mediated suppression of clinical disease was associated with suppressed serum IFN-gamma, TNF-alpha, IL-1beta, IL-2, IL-6, C3, and anti-AChR IgG1 without influencing total serum IgG. Therefore, IL-1ra could be used as a nonsteroidal drug for the treatment of MG.     

Mechanism of acute toxicity of IL-1 beta in mice

Human recombinant IL-1 beta was able to kill C3H/HeJ mice only when inoculated intravenously at very high doses. IL-1 beta, inoculated at 100 mg/kg i.v. as a bolus, induced a shock-like state characterized by anorexia, severe hypothermia and hypoglycemia and persistent neutrophilia, leading to death in 55% of animals generally between 24 and 48 h. In contrast, the noninflammatory adjuvant IL-1 beta peptide VQGEESNDK (position 163-171) did not induce any toxic effect in vivo, when administered following the same schedule. At variance with what was previously observed in endotoxin induced shock, IL-1 beta induced death was not preceded by appearance of circulating TNF. On the other hand, very high and persistent levels of circulating IL-6 could be detected after lethal IL-1 beta administration. Treatment of mice with ibuprofen or with chlorpromazine, both known to counteract some of the toxic effects of IL-1 in vivo, could protect from IL-1 beta induced mortality. Both drugs, at doses protecting from IL-1 beta induced death, were able to abolish IL-1 beta-induced rise of circulating phospholipase A2 (PLA2) activity, and the subsequent generation of toxic PLA2-derived metabolites.