Serum and salivary inflammatory biomarkers in juvenile idiopathic arthritis-an explorative cross-sectional study.

BACKGROUND: Biomarkers may be useful in monitoring disease activity in juvenile idiopathic arthritis (JIA). With new treatment options and treatment goals in JIA, there is an urgent need for more sensitive and responsive biomarkers. OBJECTIVE: We aimed to investigate the patterns of 92 inflammation-related biomarkers in serum and saliva in a group of Norwegian children and adolescents with JIA and controls and in active and inactive JIA. In addition, we explored whether treatment with tumor necrosis factor inhibitors (TNFi) affected the biomarker levels. METHODS: This explorative, cross-sectional study comprised a subset of children and adolescents with non-systemic JIA and matched controls from the Norwegian juvenile idiopathic arthritis study (NorJIA Study). The JIA group included individuals with clinically active or inactive JIA. Serum and unstimulated saliva were analyzed using a multiplex assay of 92 inflammation-related biomarkers. Welch's t-test and Mann-Whitney U-test were used to analyze the differences in biomarker levels between JIA and controls and between active and inactive disease. RESULTS: We included 42 participants with JIA and 30 controls, predominantly females, with a median age of 14 years. Of the 92 biomarkers, 87 were detected in serum, 73 in saliva, and 71 in both biofluids. A pronounced difference between serum and salivary biomarker patterns was found. Most biomarkers had higher levels in serum and lower levels in saliva in JIA versus controls, and in active versus inactive disease. In serum, TNF and S100A12 levels were notably higher in JIA and active disease. The TNF increase was less pronounced when excluding TNFi-treated individuals. In saliva, several biomarkers from the chemokine family were distinctly lower in the JIA group, and levels were even lower in active disease. CONCLUSION: In this explorative study, the serum and salivary biomarker patterns differed markedly, suggesting that saliva may not be a suitable substitute for serum when assessing systemic inflammation in JIA. Increased TNF levels in serum may not be a reliable biomarker for inflammatory activity in TNFi-treated children and adolescents with JIA. The lower levels of chemokines in saliva in JIA compared to controls and in active compared to inactive disease, warrant further investigation.

The Copenhagen Actinic Keratosis Study (COAKS). A decentralised clinical trial to evaluate tolerability, safety and efficacy of daily field-directed topical treatment with cytosolic phospholipase A2 inhibitor, AVX001, in participants with actinic keratosis: protocol for a randomised controlled phase I/IIa trial.

INTRODUCTION: Actinic keratosis (AK) is the most common precancerous skin condition caused by long-term UV exposure. Given the high recurrence rate of 15%-53%, identifying safe and effective treatment options is warranted. AVX001, a cytosolic phospholipase A2α (cPLA2α) enzyme inhibitor, is a novel anti-inflammatory drug for field-directed, self-administered, topical therapy of AK. METHODS AND ANALYSIS: This study is a single-centre, randomised, vehicle-controlled, double-blind, parallel-group hybrid clinical trial in adults with multiple AK lesions Olsen grade 1 or 2. The hybrid design combines decentralised participant tasks and assessments with conventional in-clinic visits. Recruitment using targeted advertising on social media and eligibility prescreening are conducted via the Studies&Me online recruitment platform. Participants (n=60) are randomly assigned to 1 of 3 treatment arms: AVX001 gel 1%, AVX001 gel 3% or vehicle gel. The trial consists of a 4-week treatment period with daily field-directed topical application of the gel and an 8-week follow-up period. Participants attend in-clinic visits at baseline, week 4 and week 12. The remote participant trial tasks include questionnaires and upload of smartphone-obtained photos of the treated skin area using a study-specific web-based app. Both remote and in-clinic assessments of safety and efficacy will be performed. The primary objective is to evaluate the local tolerability of daily application of AVX001 gel (1% or 3%) compared with vehicle gel. Secondary objectives include safety, efficacy, dose-response efficacy relationship, treatment satisfaction and cosmetic outcome. Exploratory objectives include evaluations of tolerability and efficacy assessed by dermatologists using smartphone photos uploaded by participants, comparisons of in-clinic and remote assessments and assessment of AK-related skin changes by non-invasive optical imaging. ETHICS AND DISSEMINATION: Approved by the Ethics Committee of the Capital Region of Denmark (H-21018064) and the Danish Medicines Agency (2021032485). Results will be submitted for publication in peer-reviewed scientific journals. TRIAL REGISTRATION NUMBERS: 2021-000934-32; NCT05164393.

A study of socio-economic inequalities in self-reported oral and general health in South-East Norway.

This study assesses the association between socioeconomic determinants and self-reported health using data from a regional Norwegian health survey. We included 9,068 participants ≥ 25 years. Survey data were linked to registry data on education and income. Self-reported oral and general health were separately assessed and categorized into 'good'/'poor'. Exposures were educational level, personal income, and economic security. Prevalence ratios (PR) were computed to assess the associations between socioeconomic determinants and self-reported health using Poisson regression models. Participants with low education or income had poorer oral and general health than those with more education or higher income. Comparing the highest and lowest education levels, adjusted PRs for poor oral and general health were 1.27 (95%CI, 1.11-1.46) and 1.43 (95%CI, 1.29-1.59), respectively. Correspondingly, PRs for lowest income quintiles compared to highest quintile were 1.34 (95%CI, 1.17-1.55) and 2.10 (95%CI, 1.82-2.43). Low economic security was also significantly associated with poor oral and general health. There were socioeconomic gradients and positive linear trends between levels of education and income in relation to both outcomes (P-linear trends < 0.001). We found statistical evidence of effect modification by gender on the association between education and oral and general health, and by age group between income and oral health.

Inhibition of Cytosolic Phospholipase A2α Induces Apoptosis in Multiple Myeloma Cells.

Cytosolic phospholipase A2α (cPLA2α) is the rate-limiting enzyme in releasing arachidonic acid and biosynthesis of its derivative eicosanoids. Thus, the catalytic activity of cPLA2α plays an important role in cellular metabolism in healthy as well as cancer cells. There is mounting evidence suggesting that cPLA2α is an interesting target for cancer treatment; however, it is unclear which cancers are most relevant for further investigation. Here we report the relative expression of cPLA2α in a variety of cancers and cancer cell lines using publicly available datasets. The profiling of a panel of cancer cell lines representing different tissue origins suggests that hematological malignancies are particularly sensitive to the growth inhibitory effect of cPLA2α inhibition. Several hematological cancers and cancer cell lines overexpressed cPLA2α, including multiple myeloma. Multiple myeloma is an incurable hematological cancer of plasma cells in the bone marrow with an emerging requirement of therapeutic approaches. We show here that two cPLA2α inhibitors AVX420 and AVX002, significantly and dose-dependently reduced the viability of multiple myeloma cells and induced apoptosis in vitro. Our findings implicate cPLA2α activity in the survival of multiple myeloma cells and support further studies into cPLA2α as a potential target for treating hematological cancers, including multiple myeloma.

Oral health in children and adolescents with juvenile idiopathic arthritis - a systematic review and meta-analysis.

BACKGROUND: Observational studies examining the association between oral health and juvenile idiopathic arthritis (JIA) among children and adolescents have reported inconsistent findings. The aims of this systematic review and meta-analysis were to ascertain a potential difference in oral health and oral health-related quality of life (OHRQoL) among children and adolescents with JIA and healthy peers, and to assess the association of prevalence of oral diseases/conditions, temporomandibular disorders (TMD), including temporomandibular joint (TMJ) diseases, in relation to activity and severity of JIA. METHOD: Medline Ovid, Embase, CINAHL, SweMed+ and Cochrane Library were searched up to 25 November 2018. All articles published in English, German and Scandinavian languages focusing on children and adolescents with JIA and without JIA in relation to oral health measures, were considered. Two authors independently evaluated observational studies for inclusion. The study quality was assessed using modified Newcastle Ottawa Scale. Meta-analysis was performed for studies focusing on dental caries as an outcome. RESULTS: Nineteen articles met the inclusion criteria, covering a range of oral diseases/conditions and OHRQoL. Eighteen studies had cross-sectional design. No mean difference of dmft/DMFT indices (decayed/missed/filled teeth) was observed between the JIA - and healthy group. None of the oral health measures including dental erosive wear, enamel defects, dental maturation and OHRQoL, indicated better oral health among children and adolescents with JIA compared to healthy group. However, periodontal conditions and TMD were more predominant among children and adolescents with JIA compared to healthy peers. CONCLUSIONS: Based on the cross-sectional studies, periodontal diseases and TMD were found to be more frequent in children and adolescents with JIA compared to healthy peers. Furthermore, more high-quality studies with large sample size are needed before we infer any concrete conclusion regarding the association between the prevalence of oral and TMJ diseases or oral conditions in relation to activity and severity of JIA.

Cytosolic phospholipase A2 modulates TLR2 signaling in synoviocytes.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic synovitis leading to destruction of cartilage and bone. PLA2 enzymes are key players in inflammation regulating the release of unsaturated fatty acids such as arachidonic acid (AA), a precursor of pro-inflammatory eicosanoids. Several lines of evidence point to toll-like receptors (TLRs) as drivers of synovitis and joint destruction in RA. However, few studies have addressed the implication of PLA2 activity downstream TLR activation in the synovium. Here, we aimed to characterize PLA2 enzyme involvement in TLR2-induced signaling in synovial fibroblast-like cells. TLRs1-7 and a range of sPLA2, iPLA2 and cPLA2 enzymes were found to be transcriptionally expressed in cultured synoviocytes. Activation of TLR2/1 and TLR2/6 led to phosphorylation of cPLA2α at Ser505, and induced AA release and PGE2 production; effects that were attenuated by cPLA2α inhibitors. In contrast, sPLA2 inhibitors did not affect AA or PGE2 release. cPLA2α inhibitors furthermore attenuated TLR-induced expression of IL-6, IL-8 and COX2. COX1/2 inhibitors attenuated TLR2/6-induced IL-6 transcription and protein production comparable to cPLA2α inhibition. Moreover, exogenously PGE2 added alone induced IL-6 production and completely rescued IL-6 transcription when added simultaneously with FSL-1 in the presence of a cPLA2α inhibitor. Our results demonstrate for the first time that cPLA2α is involved in TLR2/1- and TLR2/6-induced AA release, PGE2 production and pro-inflammatory cytokine expression in synoviocytes, possibly through COX/PGE2-dependent pathways. These findings expand our understanding of cPLA2α as a modulator of inflammatory molecular mechanisms in chronic diseases such as RA.

Inhibition of group IVA cytosolic phospholipase A2 by thiazolyl ketones in vitro, ex vivo, and in vivo.

Group IVA cytosolic phospholipase A2 (GIVA cPLA2) is the rate-limiting provider of pro-inflammatory mediators in many tissues and is thus an attractive target for the development of novel anti-inflammatory agents. In this work, we present the synthesis of new thiazolyl ketones and the study of their activities in vitro, in cells, and in vivo. Within this series of compounds, methyl 2-(2-(4-octylphenoxy)acetyl)thiazole-4-carboxylate (GK470) was found to be the most potent inhibitor of GIVA cPLA2, exhibiting an XI(50) value of 0.011 mole fraction in a mixed micelle assay and an IC50 of 300 nM in a vesicle assay. In a cellular assay using SW982 fibroblast-like synoviocytes, it suppressed the release of arachidonic acid with an IC50 value of 0.6 µM. In a prophylactic collagen-induced arthritis model, it exhibited an anti-inflammatory effect comparable to the reference drug methotrexate, whereas in a therapeutic model, it showed results comparable to those of the reference drug Enbrel. In both models, it significantly reduced plasma PGE2 levels.

Cytosolic phospholipase A2 regulates TNF-induced production of joint destructive effectors in synoviocytes.

INTRODUCTION: Rheumatoid arthritis (RA) is an inflammatory disease of the joint characterized by chronic synovitis causing pain, swelling and loss of function due to destruction of cartilage and bone. The complex series of pathological events occurring in RA is largely regulated via excessive production of pro-inflammatory cytokines, the most prominent being tumor necrosis factor (TNF). The objective of this work was to elucidate possible involvement of group IVA cytosolic phospholipase A2 (cPLA2α) in TNF-induced regulation of synovitis and joint destructive effectors in RA, to evaluate the potential of cPLA2α as a future therapeutic target. METHODS: The involvement of cPLA2α in tumor necrosis factor (TNF)-induced intracellular signaling cascades in synoviocytes (synovial fibroblast-like cells) was analyzed by arachidonic acid (AA) release assay, synoviocyte enzyme activity assay, gene expression analysis by real-time PCR and ELISA immunoassay for the detection of prostaglandin E2 (PGE2), interleukin 8 (IL8) and stromelysin-1 (MMP3), respectively. RESULTS: Inhibitors of cPLA2α enzyme activity (AVX002, ATK) significantly reduced TNF-induced cellular release of AA, PGE2, IL8 and MMP3. This reduction was evident both at transcriptional, protein or metabolite levels. Interestingly, cPLA2α inhibition affected several key points of the arachidonyl cascade; AA-release, cyclooxygenase-2 (COX2) expression and PGE2 production. Furthermore, the results suggest that cPLA2α is subject to transcriptional auto-regulation as inhibition of cPLA2α resulted in reduced PLA2G4A gene expression in TNF-stimulated synoviocytes. CONCLUSIONS: cPLA2α appears to be an important regulator of central effectors of inflammation and joint destruction, namely MMP3, IL8, COX2, and PGE2. Decreased transcription of the PLA2G4A and COX2 genes in response to cPLA2α enzyme inhibition further suggest a self-reinforcing effect of cPLA2α inhibition in response to TNF. Collectively, these results support that cPLA2α is an attractive therapeutic target candidate as its inhibition reduces the production of multiple key pro-inflammatory factors involved in RA pathogenesis.

Platelet-activating factor induces proliferation in differentiated keratinocytes.

Increased levels of platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) are found in several inflammatory dermatoses, but PAF's exact role in epidermis is uncertain. In order to better understand the physiological consequences of excess PAF production in epidermis, we examined the gene regulatory effects of PAF short-term stimulation in differentiated HaCaT keratinocytes by transcriptional profiling. Even though PAF induces COX2 expression, we found that PAF regulates only few genes associated with inflammation in differentiated keratinocytes. Rather, we show that natural PAF rapidly regulates genes involved in proliferation, (anti)-apoptosis and migration, all sub-processes of re-epithelialization and wound healing. Moreover, profiling of phosphorylated kinases, cellular wound-scratch experiments, resazurin assay and flow cytometry cell cycle phase analysis all support a role for PAF in keratinocyte proliferation and epidermal re-epithelialization. In conclusion, these results suggest that PAF acts as an activator of proliferation and may, therefore, function as a connector between inflammation and proliferation in differentiated keratinocytes.

The ω3-polyunsaturated fatty acid derivatives AVX001 and AVX002 directly inhibit cytosolic phospholipase A(2) and suppress PGE(2) formation in mesangial cells.

BACKGROUND AND PURPOSE: ω3-polyunsaturated fatty acids (ω3-PUFAs) are known to exert anti-inflammatory effects in various disease models although their direct targets are only poorly characterized. EXPERIMENTAL APPROACH: Here we report on two new cPLA(2) inhibitors, the ω3-derivatives AVX001 and AVX002, and their effects on inflammatory PGE(2) production in cultures of renal mesangial cells. KEY RESULTS: AVX001 and AVX002 dose-dependently inhibited the group IVA cytosolic phospholipase A(2) (cPLA(2) ) in an in vitro activity assay with similar IC(50) values for AVX001 and AVX002, whereas the known cPLA(2) inhibitor AACOCF(3) was less potent and docosahexaenoic acid (DHA) was inactive. In renal mesangial cells, AVX001 and AVX002 suppressed IL-1ß-induced PGE(2) synthesis. Mechanistically, this effect occurred by a down-regulation of IL-1ß-induced group IIA-sPLA(2) protein expression, mRNA expression and promoter activity. A similar but less potent effect was seen with AACOCF(3) and no effect was seen with DHA. As gene expression of sPLA(2) is known to be regulated by the transcription factor NF-κB, we further investigated NF-κB activation. Both compounds prevented NF-κB activation by blocking degradation of the inhibitor of κB. CONCLUSIONS AND IMPLICATIONS: These data show for the first time that the novel cPLA(2) inhibitors AVX001 and AVX002 exert an anti-inflammatory effect in cultures of renal mesangial cells and reduce the pro-inflammatory mediator PGE(2) through an inhibitory effect on NF-κB activation. Therefore, these compounds may represent promising novel drugs for the treatment of inflammatory disorders.

Presence of secretory group IIa and V phospholipase A2 and cytosolic group IValpha phospholipase A2 in chondrocytes from patients with rheumatoid arthritis.

Both secretory and cytosolic phospholipase A2 enzymes have been implicated in the pathogenesis of arthritis in animal models, but the exact expression patterns of the enzymes in diseased human joint tissue are uncertain. We investigated the messenger RNA expression of group IIa, IValpha and V phospholipase A2 and localized the presence of group IIa and IValpha phospholipase A2 at protein levels in articular cartilage from patients with rheumatoid arthritis, osteoarthritis and patients with non-arthritic joints. Both group IIa phospholipase A2 messenger RNA and protein were detected in all samples independent of diagnosis, but were far more prominent in cartilage from rheumatoid arthritis samples. In cartilage with rheumatoid arthritis, the enzyme was detected both within the chondrocytes and in the extracellular matrix, whereas only few osteoarthritic cartilage samples showed positive staining in the matrix. In the cartilage matrix of non-arthritic controls, group IIa phospholipase A2 was totally absent. Messenger RNA for the group IValpha and V phospholipase A2 was, except for one osteoarthritic cartilage sample, exclusively detected in rheumatoid arthritic cartilage. For group IValpha phospholipase A2 this was also confirmed at the protein level. These results suggest that each phospholipase A2 enzyme has distinct roles in both healthy and diseased joint tissue, and that the diversity and amount of enzyme correlate with the grade of inflammation and disease severity.