IntroductionThe study was undertaken to investigate the interrelation of toll-like receptor (TLR) and interleukin (IL)-17 in the salivary glands of patients with primary Sjogren's syndrome (pSS) and to determine the role of TLR and IL-17 in the pathophysiology of pSS.MethodsThe expressions of various TLRs, IL-17 and the cytokines involved in Th17 cell differentiation including IL-6, IL-23, tumor necrosis factor-alpha (TNF-α) and IL-1β were examined by immunohistochemistry in salivary glands of pSS patients. The IL-17 producing CD4+ T cells (Th17 cells) were examined by flow cytometry and confocal staining in peripheral mononuclear blood cells (PMBCs) and salivary glands of pSS patients. After PBMCs were treated with TLR specific ligands, the induction of IL-17 and IL-23 was determined using real-time PCR and ELISA. The signaling pathway that mediates the TLR2 stimulated production of IL-17 and IL-23 was investigated by using treatment with specific signaling inhibitors.ResultsWe showed that TLR2, TLR4, TLR6, IL-17 and the cytokines associated with Th17 cells were highly expressed in salivary glands of pSS patients but not in controls. The expressions of TLR2, TLR4 and TLR6 were observed in the infiltrating mononuclear cells and ductal epithelial cells, whereas IL-17 was mainly observed in infiltrating CD4+ T cells. The number of IL-17 producing CD4+ T cells was significantly higher in pSS patients both in PBMCs and minor salivary glands. The stimulation of TLR2, TLR4 and TLR6 additively induced the production of IL-17 and IL-23 from the PBMCs of pSS patients especially in the presence of TLR2 stimulation. IL-6, signal transducer and activator of transcription 3 (STAT3) and nuclear factor-kappaB (NF-kB) pathways were implicated in the TLR2 stimulated IL-17 and IL-23.ConclusionsOur data demonstrate that TLR2 ligation induces the production of IL-23/IL-17 via IL-6, STAT3 and NF-kB pathway in pSS. Therefore, therapeutic strategies that target TLR/IL-17 pathway might be strong candidates for treatment modalities of pSS.

Abbreviations: GSPE, grape seed proanthocyanidin extract; MIA, monosodium iodoacetate; MMP, matrix metalloproteinase; O2 -, superoxide anion; OA, osteoarthritis; OH, hydroxyl radicals; ONOO -, peroxynitrite; PWL, paw withdrawal latency; PWT, paw withdrawal threshold; ROS, reactive oxygen species; TRP, transient receptor potential; VIP, vasoactive intestinal peptide Abstract Osteoarthritis (OA) is an age-related joint disease that is characterized by degeneration of articular cartilage and chronic pain. Oxidative stress is considered one of the pathophysiological factors in the progression of OA. We investigated the effects of grape seed proanthocyanidin extract (GSPE), which is an antioxidant, on monosodium iodoacetate (MIA)-induced arthritis of the knee joint of rat, which is an animal model of human OA. GSPE (100 mg/kg or 300 mg/kg) or saline was given orally three times per week for 4 weeks after the MIA injection. Pain was measured using the paw withdrawal latency (PWL), the paw withdrawal threshold (PWT) and the hind limb weight bearing ability. Joint damage was assessed using histological and microscopic analysis and microcomputerized tomography. Matrix metalloproteinase-13 (MMP13) and nitrotyrosine were detected using immunohistochemistry. Administration of GSPE to the MIA-treated rats significantly increased the PWL and PWT and this resulted in recovery of hind paw weight distribution (P ＜ 0.05). GSPE reduced the loss of chondrocytes and proteoglycan, the production of MMP13, nitrotyrosine and IL-1β and the formation of osteophytes, and it reduced the number of subchondral bone fractures in the MIA-treated rats. These results indicate that GSPE is antinociceptive and it is protective against joint damage in the MIA-treated rat model of OA. GSPE could open up novel avenues for the treatment of OA.

B cells play an important role in the pathogenesis of rheumatoid arthritis (RA). High levels of B cell activating factor (BAFF) are detected in autoimmune diseases. BAFF and BAFF receptor (BAFF-R) are expressed in B and T cells of RA synovium. The study was undertaken to identify the NF-κB signal pathway involved in the induction of BAFF-R in human B cells. Immunohistochemical staining of NF-κB p65, NF-κB p50, BAFF, and BAFF-R was performed on sections of synovium from severe and mild RA and osteoarthritis (OA) patients. Peripheral blood mononuclear cells (PBMCs) were isolated from control and RA patients and B cells were isolated from controls. BAFF-R was analyzed by flow cytometry, realtime PCR and confocal staining after treatment with NF-κB inhibitors. NF-κB p65, NF-κB p50, BAFF, and BAFF-R were highly expressed in severe RA synovium relative to mild RA synovium or OA synovium. BAFF-R expression was reduced by NF-κB inhibitors in PBMCs and B cells from normal controls. We also showed reduction in expression of BAFF-R via inhibition of the NF-κB pathway in PBMCs of RA patients. BAFF/BAFF-R signaling is an important mechanism of pathogenesis in RA and that BAFF-R reduction by NF-κB blocking therapy is another choice for controlling B cells in autoimmune diseases such as RA.

We have previously demonstrated that semimature dendritic cell- (smDC-) based immunotherapy is effective for the treatment of collagen-induced arthritis (CIA) prior to disease onset. In the present study, we examined the efficacy of combination therapy with smDCs and methotrexate (MTX) in advanced CIA with a score of 2-3. Combination therapy with low-dose MTX and type II collagen- (CII-) pulsed smDCs (CII-smDCs) was more effective in inhibiting disease progression than high or low-dose MTX alone or a combination of high dose MTX and CII-smDCs. The effect of CII-smDCs alone was also comparable to the combination therapy. CD4+Foxp3+ Treg populations and IL-10 secretion markedly increased, and CII-specific autoreactive T cells decreased in mice treated with CII-smDCs alone or in combination with MTX. Combination therapy reduced the secretion of interferon-γ (IFN-γ) and IL-17 with little influence on the IL-4 secretion in the mixed leukocyte reaction. These results imply that the combination therapy with low-dose MTX and smDCs is effective in controlling advanced CIA by enhancing Treg population and suppresses antigen-specific Th1/Th17 immunity, rather than initiating Th1 to Th2 immune deviation. Our findings provide a better understanding of the DC therapy in combination with MTX for the treatment of patients with rheumatoid arthritis (RA).

To examine the effects of NF-κB inhibition on the secretion of macrophage migration inhibitory factor (MIF) in CD4+ T cells. Isolated human CD4+ T cells were cultured with NF-κB inhibitors for 24 hrs. MIF levels were evaluated by intracellular FACs, ELISA, and real time PCR. The levels of intracellular O2-, H2O2, and glutathione were evaluated by intracellular FACs. The amounts of phosphorylated Akt and c-Jun were measured by Western blotting. Treatment of CD4+ T cells with NF-κB inhibitors resulted in a significant increase of MIF concentration in culture supernatants, MIF gene expression and O2- production and decrease of intracellular MIF, H2O2, and GSH. Treatment with LY294002, and SP600125 suppressed NF-κB inhibitors induced MIF mRNA expression and MIF secretion and inhibited parthenolide induced phosphorylation of Akt and c-Jun. Treatment with H2O2 also decreased the amounts of intracellular MIF protein and increased MIF level in the culture supernatant. NAC, an antioxidant precursor of GSH, inhibited parthenolide or H2O2 induced secretion of MIF. These results indicate inhibition of NF-κB leads to release of MIF through de novo synthesis of MIF and secretion of preformed MIF in CD4+ T cells through reactive oxygen species production. .

Objective: Mature B cells in the spleen of mouse can be divide into two main subsets: the follicular (FO) B cells and the marginal zone (MZ) B cells. In this study, we investigated which subtype of B cells is involved in the production of costimulatory molecules, cytokine and antibody during the induction of autoimmune arthritis.Methods: The MZB and FOB cells isolated from DBA/1J induced-and collagen-induced arthritis (CIA) mice were stimulated with LPS or CpG. The costimulatory molecules were measured by flow cytometry (FACs). The cytokines were measured by ELISA. Production of antibodies by the MZB cells or FOB cells was measured by ELISA and the results were observed by confocal microscopy. Results:The expression of co-stimulatory molecules was stronger in the MZB cells than that in the FOB cells. The production of cytokines (IL-10, IL-6) and antibodies was higher in the MZB cells. The IgG expression of the MZB cells, which is known to be associated with the acceleration of autoimmunity, was higher in the CIA mice than that in the DBA/1J mice. Conclusion:We observed that the MZB cells were increased in the CIA mice. The

Introduction This randomized, double-blind, placebo-controlled, phase 2a trial was conducted to evaluate the safety and immunogenicity of the ID93 + glucopyranosyl lipid adjuvant (GLA)-stable emulsion (SE) vaccine in human immunodeficiency virus (HIV)-negative, previously Bacillus Calmette–Guérin (BCG)-vaccinated, and QuantiFERON-TB-negative healthy adults in South Korea. Methods Adults ( n = 107) with no signs or symptoms of tuberculosis were randomly assigned to receive three intramuscular injections of 2 μg ID93 + 5 μg GLA-SE, 10 μg ID93 + 5 μg GLA-SE, or 0.9% normal saline placebo on days 0, 28, and 56. For safety assessment, data on solicited adverse events (AEs), unsolicited AEs, serious AEs (SAEs), and special interest AEs were collected. Antigen-specific antibody responses were measured using serum enzyme-linked immunosorbent assay. T-cell immune responses were measured using enzyme-linked immunospot and intracellular cytokine staining. Results No SAEs, deaths, or AEs leading to treatment discontinuation were found. The solicited local and systemic AEs observed were consistent with those previously reported. Compared with adults administered with the placebo, those administered with three intramuscular vaccine injections exhibited significantly higher antigen-specific antibody levels and Type 1 T-helper cellular immune responses. Conclusion The ID93 + GLA-SE vaccine induced antigen-specific cellular and humoral immune responses, with an acceptable safety profile in previously healthy, BCG-vaccinated, Mycobacterium tuberculosis -uninfected adult healthcare workers. Trial Registration This clinical trial was retrospectively registered on 16 January 2019 at Clinicaltrials.gov (NCT03806686). Supplementary Information The online version contains supplementary material available at 10.1007/s40121-023-00806-0.

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