Intra-articular nuclear factor-κB blockade ameliorates collagen-induced arthritis in mice by eliciting regulatory T cells and macrophages.

Nuclear factor (NF)-κB is a transcription factor implicated in the pathogenesis of autoimmune disorders such as rheumatoid arthritis (RA). Here we have examined the effect of intra-articular administration of the IKK inhibitor, NEMO-binding domain peptide (NBD), on the severity of collagen-induced arthritis (CIA). NBD peptides were injected intra-articularly into the knee joints of DBA/1J mice after the onset of disease. Collagen-injected mice given a scrambled peptide served as controls. Arthritis severity was determined by visual examination of paws. Intra-articular NBD injection reduced the arthritis score and ameliorated morphological signs of bone destruction compared to the controls. Serum levels of type-II collagen-specific immunoglobulin (Ig)G2a antibodies were lower in NBD-treated mice versus the control mice, whereas the levels of type-II collagen-specific IgG1 antibodies were increased by NBD treatment. NBD treatment diminished the proinflammatory cytokines interleukin (IL)-17 and interferon (IFN)-γ in serum, but increased the regulatory cytokine IL-10. NBD-treated CIA mice exhibited significantly higher percentages and numbers of forkhead box protein 3 (FoxP3(+)) CD4(+) CD25(+) regulatory T cells than controls. Immunofluorescence analysis of NBD-treated mice revealed that FoxP3 and Ym1, a marker of alternatively activated macrophages, were juxtaposed to each other within draining inguinal lymph nodes. Intra-articular administration of NBD peptide is effective as an experimental therapy in a murine model of RA. Nevertheless, the intra-articular treatment modality is still associated with systemic effects on the immune system.

Haemophilus influenzae serotype B (Hib) seroprevalence in England and Wales in 2009.

A national seroprevalence study was performed to determine the prevalence of Haemophilus influenzae type b (Hib) antibodies in England and Wales in 2009, when Hib disease incidence was the lowest ever recorded. A total of 2,693 anonymised residual sera from routine diagnostic testing submitted by participating National Health Service hospital laboratories were tested for Hib anti-polyribosyl-ribitol phosphate (PRP) IgG antibodies using a fluorescent bead assay. Median anti-PRP IgG concentrations were highest in toddlers aged 1­4 years (2.65 µg/ml), followed by children aged 5­9 years (1.95 µg/ml). Antibody concentrations were significantly lower after this age, but were still significantly higher among 10­19 year-olds (0.54 µg/ml) compared with adults aged >20 years (0.16 µg/ ml; p<0.0001). Half of the adults (51%) did not have Hib antibody concentrations ≥0.15 µg/ml, the level considered to confer short-term protection. Thus, the current excellent Hib control appears to be the result of high anti-PRP antibody concentrations in children aged up to 10 years, achieved through the various childhood vaccination campaigns offering booster immunisation. The lack of seroprotection in adults emphasises the importance of maintaining control of the disease and, most probably carriage, in children, therefore raising the question as to whether long-term routine boosting of either pre-school children or adolescents may be required.

Effect of oestrogen on T cell apoptosis in patients with systemic lupus erythematosus.

Defective control of T cell apoptosis is considered to be one of the pathogenetic mechanisms in systemic lupus erythematosus (SLE). Oestrogen has been known to predispose women to SLE and also to exacerbate activity of SLE; however, the role of oestrogen in the apoptosis of SLE T cells has not yet been documented. In this study, we investigated the direct effect of oestrogen on the activation-induced cell death of T cells in SLE patients. The results demonstrated that oestradiol decreased the apoptosis of SLE T cells stimulated with phorbol 12-myristate 13-acetate (PMA) plus ionomycin in a dose-dependent manner. In addition, oestradiol down-regulated the expression of Fas ligand (FasL) in activated SLE T cells at the both protein and mRNA levels. In contrast, testosterone increased FasL expression dose-dependently in SLE T cells stimulated with PMA plus ionomycin. The inhibitory effect of oestradiol on FasL expression was mediated through binding to its receptor, as co-treatment of tamoxifen, an oestrogen receptor inhibitor, completely nullified the oestradiol-induced decrease in FasL mRNA expression. Moreover, pre-treatment of FasL-transfected L5178Y cells with either oestradiol or anti-FasL antibody inhibited significantly the apoptosis of Fas-sensitive Hela cells when two types of cells were co-cultured. These data suggest that oestrogen inhibits activation-induced apoptosis of SLE T cells by down-regulating the expression of FasL. Oestrogen inhibition of T cell apoptosis may allow for the persistence of autoreactive T cells, thereby exhibiting the detrimental action of oestrogen on SLE activity.

Effect of interleukin-4 on vascular endothelial growth factor production in rheumatoid synovial fibroblasts.

Interleukin (IL)-4 has been demonstrated to have anti-inflammatory and anti-tumour activity. Because aberrant angiogenesis is a significant pathogenic component of tumour growth and chronic inflammation, we investigated the effect of IL-4 on the production of vascular endothelial growth factor (VEGF) by synovial fibroblasts derived from patients with rheumatoid arthritis (RA). Fibroblast-like synoviocytes (FLS) were prepared from synovial tissues of RA and incubated with different concentrations of IL-4 in the presence or absence of transforming growth factor (TGF)-beta. VEGF level was measured by enzyme-linked immunosorbent assay and semiquantitative reverse transcription--polymerase chain reaction. Treatment of FLS with IL-4 alone caused a dose-dependent increase in VEGF levels. In contrast, IL-4 exhibited the inhibitory effect on VEGF production when FLS were stimulated with TGF-beta. Combined treatment of IL-4 and IL-10 inhibited TGF-beta-induced VEGF production in an additive fashion. TGF-beta increased the induction of cyclooxygenase-2 mRNA, which was inhibited significantly by the treatment of IL-4. NS-398, a COX-2 inhibitor, inhibited TGF-beta-induced VEGF production in a dose-dependent manner. Furthermore, exogenous addition of prostaglandin E2 (PGE2) restored IL-4 inhibition on TGF-beta induced VEGF production. Collectively, our results suggest that IL-4 have an anti-angiogenic effect, especially in the inflammatory milieu of RA by inhibiting the VEGF production in synovial fibroblasts.

Hydroxychloroquine potentiates Fas-mediated apoptosis of rheumatoid synoviocytes.

Inadequate apoptosis may contribute to the synovial hyperplasia associated with rheumatoid arthritis (RA). The Fas-associated death domain protein (FADD)-like interleukin (IL)-1beta-converting enzyme (FLICE)-inhibitory protein (FLIP), which is an apoptotic inhibitor, has been implicated in the resistance to Fas-mediated apoptosis of synoviocytes. This study investigated whether hydroxychloroquine (HCQ), an anti-rheumatic drug, induces the apoptosis of rheumatoid synoviocytes, and modulates the expression of FLIP. Fibroblast-like synoviocytes (FLS) were prepared from the synovial tissues of RA patients, and were cultured with various concentrations of HCQ in the presence or absence of the IgM anti-Fas monoclonal antibodies (mAb) (CH11). Treatment with HCQ, ranging from 1 to 100 microM, induced the apoptosis of FLS in a dose- and time-dependent manner. The increase in synoviocytes apoptosis by HCQ was associated with caspase-3 activation. A combined treatment of HCQ and anti-Fas mAb increased FLS apoptosis and caspase-3 activity synergistically, compared with either anti-Fas mAb or HCQ alone. The Fas expression level in the FLS was not increased by the HCQ treatment, while the FLIP mRNA and protein levels were decreased rapidly by the HCQ treatment. Moreover, time kinetics analysis revealed that the decreased expression of FLIP by HCQ preceded the apoptotic event that was triggered by HCQ plus anti-Fas mAb. Taken together, HCQ increases the apoptosis of rheumatoid synoviocytes by activating caspase-3, and also sensitizes rheumatoid synoviocytes to Fas-mediated apoptosis. Our data suggest that HCQ may exert its anti-rheumatic effect in rheumatoid joints through these mechanisms.