Jun Akaogi

Induction of lupus autoantibodies by adjuvants

Exposure to the hydrocarbon oil pristane induces lupus specific autoantibodies in non-autoimmune mice. We investigated whether the capacity to induce lupus-like autoimmunity is a unique property of pristane or is shared by other adjuvant oils. Seven groups of 3-month-old female BALB/cJ mice received a single intraperitoneal injection of pristane, squalene (used in the adjuvant MF59), incomplete Freunds adjuvant (IFA), three different medicinal mineral oils, or saline, respectively. Serum autoantibodies and peritoneal cytokine production were measured. In addition to pristane, the mineral oil Bayol F (IFA) and the endogenous hydrocarbon squalene both induced anti-nRNP/Sm and -Su autoantibodies (20% and 25% of mice, respectively). All of these hydrocarbons had prolonged effects on cytokine production by peritoneal APCs. However, high levels of IL-6, IL-12, and TNFalpha production 2-3 months after intraperitoneal injection appeared to be associated with the ability to induce lupus autoantibodies. The ability to induce lupus autoantibodies is shared by several hydrocarbons and is not unique to pristane. It correlates with stimulation of the production of IL-12 and other cytokines, suggesting a relationship with a hydrocarbons adjuvanticity. The potential to induce autoimmunity may complicate the use of oil adjuvants in human and veterinary vaccines.

Role of PGE2 and EP Receptors in the Pathogenesis of Rheumatoid Arthritis and as a Novel Therapeutic Strategy

Recent progress in understanding the pathogenesis of rheumatoid arthritis (RA) in parallel with elucidation of the functional role of the prostaglandin receptor subfamily has revealed an important regulatory role of PGE2, in addition to its well-known proinflammatory role in the progression of RA. Characteristic features of RA are synovial proliferation and pannus formation, which result in the destruction of cartilage and bone. Pannus tissue is mainly composed of macrophages and fibroblast-like synoviocytes. Both T cell-derived IL-17 and macrophage-derived TNF-alpha seem to play a central role in the progression of proinflammatory cascades in RA. PGE2 is also produced in response to proinflammatory cytokines, which in turn negatively regulates both IL-17 and TNF-alpha expression and TNF/IL-1-induced activation of fibroblast-like synoviocytes through EP2/EP4 receptors, resulting in the modulation of proinflammatory cascades. IL-17- and TNF-activated macrophages differentiate into osteoclasts in the presence of M-CSF and RANKL expressed by fibroblast-like synoviocytes. PGE2 binding to EP4 stimulates osteoclastogenesis through enhancing RANKL expression. At the same time, PGE2 suppresses osteoclastogenesis by inhibiting M-CSF expression of fibroblast-like synoviocytes as well as both IL-17 and IL-17-induced TNF-alpha expression of macrophages. PGE2-EP4 also activates osteoblastogenesis through increasing cbfa1 and osterix, two essential transcription factors required for bone formation. The net effect of PGE2 may direct toward repair of eroding bone through the suppression of inflammation and enhancement of bone remodeling. Here, we discuss a diverse action of PGE2/EP receptors and their important regulatory roles in the pathogenesis of RA, which may lead to a novel therapeutic strategy.

Prostaglandin E2 receptors EP2 and EP4 are up-regulated in peritoneal macrophages and joints of pristane-treated mice and modulate TNF-α and IL-6 production

Prostaglandin E2 (PGE2) can have pro‐ or anti‐inflammatory effects, depending on engagement of different PGE2 receptor (EP) subtypes. The role of EPs in regulating autoimmune inflammation was studied in the murine arthritis/lupus model induced by pristane. Peritoneal macrophages were isolated (biomagnetic beads) from BALB/c, DBA/1, or C57BL/6 mice treated with pristane (intraperitoneally, 3 months earlier) or thioglycolate (3 days earlier) or with untreated controls. EPs, inducible nitric oxide synthase (iNOS), and cyclooxygenase‐2 (COX‐2) mRNA expression was examined by reverse transcriptase‐polymerase chain reaction (RT‐PCR). Cells were cultured unstimulated or stimulated with lipopolysaccharide (LPS) or LPS + interferon‐γ in combination with EP subtype‐specific agonists. Tumor necrosis factor α (TNF‐α) and interleukin (IL)‐6 production was tested by enzyme‐linked immunosorbent assay (culture supernatant) and flow cytometry. TNF‐α mRNA levels also were examined. High levels of EPs (EP4/2>EP1>EP3), iNOS, and COX‐2 mRNA were expressed in peritoneal macrophages from pristane‐treated but not untreated or thioglycolate‐treated mice (RT‐PCR). TNF‐α production was inhibited 50–70% at 2–24 h by EP4/2 agonists, whereas IL‐6 was enhanced up to ∼220%. TNF‐α inhibition is mediated partly via the protein kinase A pathway and partly via IL‐6. Intracellular TNF‐α staining was inhibited 20% by EP4/2 agonists. TNF‐α mRNA levels were inhibited 50–70% at 2–24 h, indicating that TNF‐α inhibition was partly at the level of transcription. EP1/3 agonists had little effect. Synovial cells from mice with pristane‐induced arthritis (DBA/1) also expressed EP2/4, and the EP2/4 agonist inhibited TNF‐α production. PGE2 can modulate inflammatory reactions via the EP2/4 receptor through its regulation of TNF‐α and IL‐6. Modification of EP signaling may be a new therapeutic strategy in inflammatory/autoimmune diseases.

Citrulline Dependence of Anti-Cyclic Citrullinated Peptide Antibodies in Systemic Lupus Erythematosus as a Marker of Deforming/Erosive Arthritis

Objective. Anti-cyclic citrullinated peptide (CCP) antibodies are a serological marker for rheumatoid arthritis (RA); up to 10%–15% of patients with systemic lupus erythematosus (SLE) are also positive. While anti-CCP in RA is citrulline-dependent, anti-CCP in some other diseases is citrulline-independent and reacts with both CCP and the unmodified (arginine-containing) cyclic arginine peptide (CAP). We investigated the citrulline dependence of anti-CCP and its significance in the arthritis of SLE. Methods. IgG anti-CCP was compared by ELISA to anti-CAP in sera from patients with SLE (n = 335) and RA (n = 47) and healthy controls (n = 35). SLE patients were divided into 5 groups based on their joint involvement: subset I: deforming/erosive arthritis (n = 20); II: arthritis fulfilling (or likely fulfilling) American College of Rheumatology criteria for RA but without erosions (n = 18); III: joint swelling but not fulfilling RA criteria (n = 39); IV: arthritis without documented joint swelling (n = 194); and V: no arthritis (n = 58). Results. Anti-CCP (> 1.7 units) was found in 68% (32/47) of patients with RA and 17% (55/329) of those with SLE. It was more common in SLE patients with deforming/erosive arthritis (38%). High anti-CCP (> 10 units) was found in RA (26%) and deforming/erosive SLE (12%). High anti-CCP/CAP ratios (> 2, indicating a selectivity to CCP) were found in 91% of anti-CCP-positive RA and 50% of anti-CCP-positive SLE patients with deforming/erosive arthritis. Patients from subset II did not have high anti-CCP/CAP. Conclusion. Citrulline dependence or high levels (> 10) of anti-CCP were common in SLE patients with deforming/erosive arthritis, while most anti-CCP in SLE patients was citrulline-independent. This may be useful in identifying a subset of SLE patients with high risk for development of deforming/erosive arthritis.

8 - Induction of Autoimmunity by Adjuvant Hydrocarbons

Although susceptibility to systemic lupus erythematosus (SLE) is genetically determined, environmental factors such as ultraviolet radiation, chemicals, or infections are likely to play an important role in triggering the disease in susceptible individuals. It is reported that a lupus-like disease with disease-specific autoantibodies and nephritis develops in non-autoimmune prone mice treated with pristane, a hydrocarbon derived from the metabolism of chlorophyll. More recently, it has become clear that other hydrocarbons, notably the mineral oil Bayol F and the endogenous hydrocarbon squalene, also can induce lupus-like disease in mice. These substances share the capacity to serve as immunological adjuvants, defined as “substances used in combination with a specific antigen that produce more immunity than the antigen alone”. The induction of murine lupus by immunological adjuvants is significant for two reasons. First, it provides a model for the interaction of environmental triggers with the genetic background in systemic autoimmunity, and secondly, it raises the possibility that adjuvant hydrocarbons might trigger autoimmune disease in susceptible humans. This chapter elaborates on the current state of understanding about the immunological effects of adjuvants and presents a recent work on the induction of autoimmunity by these materials.