Ja Gracie

Interleukin 18: a pleiotropic participant in chronic inflammation

The wide-ranging effects of interleukin 18 as a regulator of innate and acquired immune responses, and particularly its role in human autoimmune diseases, suggest its potential importance as a therapeutic target.

A novel therapeutic approach targeting articular inflammation using the filarial nematode-derived phosphorylcholine-containing glycoprotein ES-62

Understanding modulation of the host immune system by pathogens offers rich therapeutic potential. Parasitic filarial nematodes are often tolerated in human hosts for decades with little evidence of pathology and this appears to reflect parasite-induced suppression of host proinflammatory immune responses. Consistent with this, we have previously described a filarial nematode-derived, secreted phosphorylcholine-containing glycoprotein, ES-62, with immunomodulatory activities that are broadly anti-inflammatory in nature. We sought to evaluate the therapeutic potential of ES-62 in vitro and in vivo in an autoimmune disease model, namely, collagen-induced arthritis in DBA/1 mice. ES-62 given during collagen priming significantly reduced initiation of inflammatory arthritis. Crucially, ES-62 was also found to suppress collagen-induced arthritis severity and progression when administration was delayed until after clinically evident disease onset. Ex vivo analyses revealed that in both cases, the effects were associated with inhibition of collagen-specific pro-inflammatory/Th1 cytokine (TNF-α, IL-6, and IFN-γ) release. In parallel in vitro human tissue studies, ES-62 was found to significantly suppress macrophage activation via cognate interaction with activated T cells. Finally, ES-62 suppressed LPS-induced rheumatoid arthritis synovial TNF-α and IL-6 production. Evolutionary pressure has promoted the generation by pathogens of diverse mechanisms enabling host immune system evasion and induction of “tolerance.” ES-62 represents one such mechanism. We now provide proof of concept that parasite-derived immunomodulatory strategies offer a novel therapeutic opportunity in inflammatory arthritis.

The phosphorycholine moiety of the filarial nematode immunomodulator ES-62 is responsible for its anti-inflammatory action in arthritis

Objective: In countries where parasitic infections are endemic, autoimmune disease is relatively rare, leading to the hypothesis that parasite-derived immunomodulators may protect against its development. Consistent with this, we have previously demonstrated that ES-62, a 62 kDa phosphorylcholine (PC)-containing glycoprotein that is secreted by filarial nematodes, can exert anti-inflammatory action in the murine collagen-induced arthritis (CIA) model and human rheumatoid arthritis-derived synovial tissue cultures. As a first step to developing ES-62-based drugs, the aim of this study was to determine whether the PC-moiety of ES-62 was responsible for its anti-inflammatory actions. Methods: We compared the anti-inflammatory activity of a PC-free form of recombinant ES-62 (rES-62) and a synthetic PC-ovalbumin conjugate (OVA-PC) with that of native ES-62 in the CIA model and synovial tissues from patients with rheumatoid arthritis. Results: The anti-inflammatory actions of ES-62 in CIA appear to be dependent on the PC moiety as indicated by the reduction in severity of disease and also suppression of collagen-specific T helper 1 cytokine production observed when testing OVA-PC, but not rES-62. Interestingly, the anti-inflammatory activity of PC did not correlate with a reduction in anti-collagen IgG2a levels. Also, the ES-62-mediated suppression of interferon-γ from human patient tissues could be mimicked by OVA-PC but not rES-62 or ovalbumin. Conclusions: In countries where filariasis is endemic the reduced detection of inflammatory diseases, such as rheumatoid arthritis may be because of the anti-inflammatory action of the PC moieties of ES-62. PC may thus provide the starting point for the development of novel, safe immunomodulatory therapies.

Disease association of two distinct interleukin-18 promoter polymorphisms in Caucasian rheumatoid arthritis patients

Interleukin (IL)-18 is an important mediator of innate and adaptive immunity. We searched for an association of IL-18 promoter single-nucleotide polymorphisms (SNP) with rheumatoid arthritis (RA) in Caucasians. The entire study population was composed of two independent cohorts from Germany (n=200) and Scotland (n=410). Presence of IL-18 SNP at positions −607 and −137 was determined by allele-specific PCR in 327 RA patients and 283 healthy donors (HD). Diplotype distributions of both loci were in Hardy–Weinberg equilibrium (HWE) in the German and Scottish HD cohorts. In contrast, locus −607 was in HW disequilibrium in German, and locus −137 in Scottish RA patients. Diplotypic exact χ2 tests suggested that −607CC was overrepresented in German, and −137CC in Scottish RA patients, but conservative χ2 trend analyses could not prove any significant disease association of these single loci. SNP −607 and −137 were in strong linkage disequilibrium. The −607C*−137C haplotype was more prevalent in German RA (3.2 vs 1.2%) and in Scottish RA patients (4.1 vs 0.9%) than in the respective HD cohorts. These observations suggest that SNP of both positions contribute to the genetic background of RA pathogenesis.

New strategies to control inflammatory synovitis: interleukin 15 and beyond

A broad range of strategies to target inflammatory synovitis is currently being explored. Most approaches require the identification of a single molecule/pathway that is tractable to modulation in the clinic. In general, molecular activities with relevant biological effects are sought within the appropriate target lesion—namely, the synovium. Thereafter, such an activity may be targeted in rodent models of arthritis and the effects on inflammation and articular destruction measured. A moiety with plausible biology and bioactivity in model systems may then move to phase I “proof of concept” studies and thereafter to clinical development (fig 1). The successful targeting of tumour necrosis factor α (TNFα) represents an example of such an approach whereby an effective therapeutic agent has been derived.1,2 However, even after TNF blockade, unmet clinical need clearly persists in inflammatory arthritis. Major challenges remain, in particular the identification of therapeutic responders a priori, and the rational choice of additional targets, either as independent primary therapeutic targets or as synergistic biological targets for future combination applications. This short review will test interleukin 15 (IL15) in this model of new biological development to evaluate its therapeutic potential. Thereafter, an alternative approach will be considered for comparative purposes, whereby a model in which multiple pathways within the synovium are targeted simultaneously using a strategy borrowed from the host-parasite relationship arising from evolutionary pressures over millennia (fig 1). Figure 1 Alternative model approaches, resulting in “proof of concept” clinical studies to target synovitis. Our group has recently sought mechanisms whereby innate and acquired immune responses interact during chronic inflammation, with particular emphasis on cytokine biology. Many cytokines present within the synovium derive from macrophages and synovial fibroblasts.3 IL15, a cytokine with structural similarities to IL2,4 is produced primarily by macrophages and as such attracted our attention at an early stage as …

Inhibition of neutrophil function by the Kaposi's sarcoma-associated herpesvirus vOX2 protein

Kaposis sarcoma is multifactorial, involving Kaposis sarcoma-associated herpesvirus (KSHV) infection and immune dysfunction. A KSHV protein (vOX2), fused with the Fc domain of human immunoglobulin G1 to create vOX2:Fc, suppressed neutrophil oxidative burst and inhibited the production of pro-inflammatory chemokines (IL-8 and monocyte chemoattractant protein 1) by monocyte/macrophage cells. vOX2:Fc suppressed the acute inflammatory response in mice in which neutrophil-mediated inflammation was induced by carrageenan. The data suggest that vOX2 can contribute to immune dysfunction and could have anti-inflammatory therapeutic potential.

TLR4 mutations (Asp299Gly and Thr399Ile) are not associated with ankylosing spondylitis

Background: Immunoregulatory genes and Gram negative gut bacteria are thought to be important in disease expression in ankylosing spondylitis (AS). Objective: To compare the frequency of two common and functional TLR4 mutations (Asp299Gly, and Thr399Ile) between patients with AS and HLA-B27 healthy controls. Methods: The TLR4 genotypes of patients and healthy HLA-B27 controls were determined using allele-specific PCR and restriction fragment length polymorphism analysis. Asp299Gly genotype was determined in 193 patients and 125 HLA-B27 positive controls and Thr399Ile genotype in 184 patients and 113 HLA-B27 controls. Allele frequencies were compared using a χ2 test of association. Results: 29/193 (15%) patients with AS had a polymorphism in the Asp299 site compared with 18/125 (14.4%) healthy HLA-B27 controls. Of the patients genotyped for the Thr399Ile allele, 29/184 (15.8%) carried the polymorphism compared with 19/113 (16.8%) HLA-B27 controls. No significant difference between the frequencies of the Asp299Gly genotype or the Thr399Ile genotype between patients with AS and healthy HLA-B27 controls was found. No significant difference in allele frequency was found at either site. Conclusion: Two common TLR4 polymorphisms, which cause a functional deficiency in host immune response to Gram negative bacteria, are not overrepresented in patients with AS.

Inflammatory cytokines in juvenile idiopathic arthritis: effects on physical growth and the insulin-like-growth factor axis

OBJECTIVE To investigate the relationship between markers of inflammation with physical growth and systemic markers of GH secretion in JIA. DESIGN This is a cross sectional prospective study of patients with JIA recruited during therapeutic arthrocentesis of 17 children with JIA (F,10): 8 oligoarticular (OJIA) and 9 polyarticular (PJIA). RESULTS Median adjusted height (AHt) SDS was -0.3 (-2.2 to 1.6). Serum ALS SDS (median -1.3, range -2.7 to -0.6) was reduced compared with serum IGFBP-3 SDS (median -0.5, range -7.7 to 2.3) and IGF-1 SDS (median -0.2, range -0.5 to 0.5). Log serum IL5 (95% CI -3.25, -0.81) and log serum IL15 (95% CI -9.58, -4.10) were independent factors associated with AHt SDS. Inflammatory cytokines individually showed no association with IGF-1, IGFBP-1, -2, -3 and ALS. CONCLUSION Children with JIA and mild degree of growth retardation show decreased ALS and IGFBP-3. Cytokines did not show an association to systemic markers of GH secretion. However, this study reports the novel, preliminary association between serum levels of IL5 and IL15 and the extent of short stature.

Skin allograft rejection in mice lacking inducible nitric oxide synthase.

BACKGROUND During allograft rejection, up-regulation of cytokine-inducible nitric oxide synthase (iNOS) leads to the production of large amounts of nitric oxide (NO). The net effect of NO on the alloimmune response is, however, difficult to predict because of its diverse biological effects, which include potentially opposing roles as an effector and immunoregulatory molecule. METHODS In this study, the role of iNOS on the in vitro and in vivo alloimmune response was defined using mutant mice that lack a functional iNOS gene. The ability of spleen cells obtained from iNOS-deficient mutants to proliferate and to produce cytokines in response to irradiated BALB/c stimulator cells was determined, and the rate at which iNOS-deficient mice were able to reject BALB/c skin allografts was observed. RESULTS Spleen cells from homozygous iNOS-deficient (129xMF1)F1 mice, when compared with cells from heterozygous control mice, showed an increased in vitro proliferative response and produced substantially higher levels of interferon-gamma, and also more interleukin-2 and interleukin-12, in response to allogeneic stimulation. The kinetics of BALB/c skin graft rejection were comparable in heterozygous control animals and iNOS-deficient mice. Moreover, no net effect of iNOS on skin allograft rejection was apparent in mice treated with depleting monoclonal antibodies (mAb) to CD4 or CD8 T cells, either alone or in combination, or in mice treated with both anti-CD8 mAb and a neutralizing anti-tumor necrosis factor mAb. CONCLUSIONS These results show that iNOS has an immunomodulatory effect on the in vitro alloimmune response but lack of iNOS has no net influence on the kinetics of murine skin allograft rejection in either unmodified recipients or recipients in which the early contribution of T-cell subsets and tumor necrosis factor-alpha to graft rejection has been abrogated.

Phosphodiesterase 4 (PDE4) regulation of proinflammatory cytokine and chemokine release from rheumatoid synovial membrane

Background The cAMP-metabolising enzyme, phosphodiesterase 4 (PDE4), has been implicated in a number of immune responses, including tumour necrosis factor α (TNFα) production. To date, few data have directly addressed whether synovial cytokine and chemokine production is modified by PDE4. Objective Using specific PDE4 inhibitors, roflumilast plus two novel inhibitors, INH 0061 and INH 0062, the authors studied the effect of PDE4 inhibition on proinflammatory cytokine and chemokine release from primary rheumatoid arthritis (RA) synovial digest suspensions and in a macrophage T cell co-culture assay system. Results All PDE4 inhibitors dose-dependently reduced the release of TNFα from primary synovial membrane cultures (n=5), half maximal inhibitory concentration (IC50) 300–30 nM, p<0.05. Similarly, a significant suppression in the release the proinflammatory chemokines, monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein (MIP)-1α, MIP-1β (IC50 300–30 nM) and regulated upon activation normal T-cell expressed and secreted (RANTES) (IC50 3 nM) was also observed, p<0.05. While interleukin 1β was also reduced, it did not achieve an IC50. These observations were further confirmed in a macrophage T cell co-culture system, demonstrating the importance of PDE4 pathways in regulating cytokine/chemokine release in a cellular interaction implicated in inflammatory synovitis. Subsequent studies using the human monocytic cell line U937 also demonstrated cytokine regulation with PDE4 knockdown utilising a small interfering RNA approach. Conclusion These data provide direct evidence of PDE4-dependent pathways in human RA synovial inflammatory cytokine and chemokine release and may provide a novel approach in treating chronic autoimmune conditions such as RA.