STAT6 and STAT1 Pathway Activation in Circulating Lymphocytes and Monocytes as Predictor of Treatment Response in Rheumatoid Arthritis

Objective To find novel predictors of treatment response to disease-modifying antirheumatic drugs (DMARDs), we studied activation of STAT (signal transducers and activators of transcription) 6 and 1 in circulating leukocytes of patients with rheumatoid arthritis (RA). Methods 19 patients with untreated recent-onset RA, 16 patients with chronic RA irresponsive to synthetic DMARDs and 37 healthy volunteers provided blood samples for whole blood flow cytometric determination of intracellular STAT6 and STAT1 phosphorylation, expressed as relative fluorescence units, in response to IL-4 and IFN-gamma, respectively. Phosphorylation was restudied and treatment response (according to European League Against Rheumatism) determined after 1-year treatment with synthetic DMARDs in recent-onset RA and with biological DMARD in synthetic DMARD-irresponsive RA. Estimation-based exact logistic regression was used to investigate relation of baseline variables to treatment response. 95% confidence intervals of means were estimated by bias-corrected bootstrapping and the significance between baseline and follow-up values was calculated by permutation test. Results At baseline, levels of phosphorylated STAT6 (pSTAT6) induced by IL-4 in monocytes were higher in those who achieved good treatment response to synthetic DMARDs than in those who did not among patients with untreated RA (OR 2.74, 95% CI 1.05 to 9.47), and IFN-gamma-stimulated lymphocyte pSTAT1 levels were higher in those who achieved good treatment response to a biological drug than in those who did not among patients with chronic RA (OR 3.91, 95% CI 1.12 to 20.68). During follow-up, in recent-onset RA patients with good treatment response to synthetic DMARDS, the lymphocyte pSTAT6 levels decreased (p = 0.011), and, consequently, the ratio of pSTAT1/pSTAT6 in lymphocytes increased (p = 0.042). Conclusion Cytokine-stimulated STAT6 and STAT1 phosphorylation in circulating leukocytes was associated with treatment response to DMARDs in this pilot study. The result, if confirmed in larger studies, may aid in developing personalized medicine in RA.Peer reviewe

The constrained median : a way to incorporate side information in the assessment of food samples

A classical problem in the field of food science concerns the consensus evaluation of food samples. Typically, several panelists are asked to provide scores describing the perceived quality of the samples, and subsequently, the overall (consensus) scores are determined. Unfortunately, gathering a large number of panelists is a challenging and very expensive way of collecting information. Interestingly, side information about the samples is often available. This paper describes a method that exploits such information with the aim of improving the assessment of the quality of multiple samples. The proposed method is illustrated by discussing an experiment on raw Atlantic salmon (Salmo salar), where the evolution of the overall score of each salmon sample is studied. The influence of incorporating knowledge of storage days, results of a clustering analysis, and information from additionally performed sensory evaluation tests is discussed. We provide guidelines for incorporating different types of information and discuss their benefits and potential risks

Multivariate statistical analysis for the identification of potential seafood spoilage indicators

Volatile organic compounds (VOCs) characterize the spoilage of seafood packaged under modified atmospheres (MAs) and could thus be used for quality monitoring. However, the VOC profile typically contains numerous multicollinear compounds and depends on the product and storage conditions. Identification of potential spoilage indicators thus calls for multivariate statistics. The aim of the present study was to define suitable statistical methods for this purpose (exploratory analysis) and to consequently characterize the spoilage of brown shrimp (Crangon crangon) and Atlantic cod (Gadus morhua) stored under different conditions (selective analysis). Hierarchical cluster analysis (HCA), principal components analysis (PCA) and partial least squares regression analysis (PLS) were applied as exploratory techniques (brown shrimp, 4 °C, 50%CO2/50%N2) and PLS was further selected for spoilage marker identification. Evolution of acetic acid, 2,3-butanediol, isobutyl alcohol, 3-methyl-1-butanol, dimethyl sulfide, ethyl acetate and trimethylamine was frequently in correspondence with changes in the microbiological quality or sensory rejection. Analysis of these VOCs could thus enhance the detection of seafood spoilage and the development of intelligent packaging technologies.acceptedVersionPeer reviewe

Periodontitis in early and chronic rheumatoid arthritis : a prospective follow-up study in Finnish population

Objectives To investigate the association between rheumatoid arthritis (RA) and periodontitis with special emphasis on the role of antirheumatic drugs in periodontal health. Design Prospective follow-up study. Patients with early untreated RA and chronic active RA were examined at baseline and 16months later. Controls were examined once. Settings and participants The study was conducted in Finland from September 2005 to May 2014 at the Helsinki University Hospital. Overall, 124 participants were recruited for dental and medical examinations: 53 were patients with early disease-modifying antirheumatic drug (DMARD) na1ve RA (ERA), 28 were patients with chronic RA (CRA) with insufficient response to conventional DMARDs. After baseline examination, patients with ERA started treatment with synthetic DMARDs and patients with CRA with biological DMARDs. Controls were 43 age-matched, gender-matched and community-matched participants. Outcome measures Degree of periodontitis (defined according to the Center for Disease Control and Prevention and the American Academy of Periodontology). Prevalence of periodontal bacteria (analysed from plaque samples), clinical rheumatological status by Disease Activity Score, 28-joint count (DAS28), function by Health Assessment Questionnaire (HAQ) and treatment response by European League Against Rheumatism (EULAR) criteria. Results Moderate periodontitis was present in 67.3% of patients with ERA, 64.3% of patients with CRA and 39.5% of control participants (p=0.001). Further, patients with RA had significantly more periodontal findings compared with controls, recorded with common periodontal indexes. In the re-examination, patients with RA still showed poor periodontal health in spite of treatment with DMARDs after baseline examination. The prevalence of Porphyromonas gingivalis was higher in patients with ERA with periodontal probing depth 4mm compared with patients with CRA and controls. Antirheumatic medication did not seem to affect the results. Conclusions Moderate periodontitis was more frequent in patients with RA than in controls. Patients with ERA and CRA exhibited poorer periodontal health parameters when compared with controls. There was no association between antirheumatic treatment and periodontal parameters.Peer reviewe

IL-6 Mediated Transcriptional Programming of Naïve CD4+ T Cells in Early Rheumatoid Arthritis Drives Dysregulated Effector Function.

Objective: We have previously shown that increased circulating interleukin-6 (IL-6) results in enhanced CD4+ T cell signaling via signal transduction and activator of transcription-3 (STAT3) in early rheumatoid arthritis (RA). We tested the hypothesis that transcriptional "imprinting" of T-cells by this mechanism skews downstream effector responses, reinforcing immune dysregulation at a critical, but targetable, disease phase. Methods: We modeled naïve CD4+ T cell exposure to pathophysiological concentrations of IL-6 in vitro, assessing the dynamic transcriptional and functional consequences for downstream effector cells utilizing microarray and flow cytometry. Fresh blood from treatment-naïve early arthritis patients was phenotyped in parallel for comparison. Results: T cell sensitivity to IL-6 was most marked in the naïve subset, and related to gp130 rather than IL-6R expression. Exposure of healthy naïve CD4+ T cells to IL-6 induced the same STAT3 target genes as previously seen to discriminate RA patients from disease controls. After TCR stimulation IL-6 pre-exposed cells exhibited enhanced proliferative capacity, activation, and a propensity toward Th1 differentiation, compared to non-exposed cells. An entirely analogous phenotype was observed in early RA compared to control CD4+ T cells. Conclusions: Sustained IL-6 exposure at a critical point in the natural history of RA "primes" the adaptive immune system to respond aberrantly to TCR stimulation, potentiating disease induction with implications for the optimal timing of targeted therapy

Baseline JAK phosphorylation profile of peripheral blood leukocytes, studied by whole blood phosphospecific flow cytometry, is associated with 1-year treatment response in early rheumatoid arthritis

Background: We found recently that baseline signal transducer and activator of transcription 3 phosphorylation in peripheral blood CD4(+) T cells of patients with early rheumatoid arthritis (RA) is associated with treatment response to synthetic disease-modifying antirheumatic drugs (DMARDs). This prompted us to study the baseline phosphorylation profiles of Janus kinases (JAKs) in blood leukocytes with respect to treatment response in early RA. Methods: Thirty-five DMARD-naive patients with early RA provided blood samples for whole blood flow cytometric determination of phosphorylation of JAKs in CD4(+) and CD8(+) T cells, CD19(+) B cells, and CD14(+) monocytes. Treatment response was determined after 1 year of treatment with synthetic DMARDs, with remission defined as absence of tender and swollen joints and normal erythrocyte sedimentation rate. Exact logistic regression was used to investigate the association of baseline variables with treatment response. Ninety-five percent CIs of means were estimated by bias-corrected bootstrapping. Results: High JAK3 phosphorylation in CD4(+) and CD8(+) T cells, CD19(+) B cells, and CD14(+) monocytes and low JAK2 phosphorylation in CD14(+) monocytes were significantly associated with remission following treatment with synthetic DMARDs. Conclusions: Baseline JAK phosphorylation profile in peripheral blood leukocytes may provide a means to predict treatment response achieved by synthetic DMARDs among patients with early RA.Peer reviewe

Pancreatic cancer is associated with aberrant monocyte function and successive differentiation into macrophages with inferior anti-tumour characteristics

Background/objectives: Inflammation is related to the development and progression of pancreatic cancer (PC). Locally, anti-inflammatory macrophages (M2), and systemically, high levels of certain inflammation-modulating cytokines associate with poor prognosis in PC. The detailed effects of systemic inflammation on circulating monocytes and macrophage polarisation remain unknown. We aimed to find out how intracellular signalling of peripheral blood monocytes is affected by the systemic inflammatory state in PC patients and how it affects their differentiation into macrophages. Methods: Monocytes were isolated from 50 consenting PC patients and 20 healthy controls (HC). The phosphorylation status of the signalling molecules was assessed by flow cytometry both from unstimulated and appropriately stimulated monocytes. Monocytes derived from HC and PC patients were cocultured with cancer cells (MIA PaCa-2 and HPAF-II) in media supplemented with autologous serum, and the CD marker expression of the obtained macrophages was assessed by flow cytometry. Results: Phosphorylation levels of unstimulated STAT2, STAT3 and STAT6 were higher (p <0.05) and those of stimulated NF-kB (p = 0.004) and STAT5 (p = 0.006) were lower in patients than in controls. The expression of CD86, a proinflammatory (Ml) marker, was higher in control- than patient-derived cocultured macrophages (p = 0.029). Conclusions: Circulating monocytes from PC patients showed constitutive phosphorylation and weaker response to stimuli, indicating aberrant activation and immune suppression. When co-culturing the patient-derived monocytes with cancer cells, they differentiated into macrophages with reduced levels of M1 macrophage marker CD86, suggesting compromised anti-tumour features. The results highlight the need for global management of tumour-associated immune aberrations in PC treatment. (C) 2021 Published by Elsevier B.V. on behalf of IAP and EPC.Peer reviewe

Immuunijärjestelmän aktivaatiosta nivelreumassa ja reaktiivisessa artriitissa

Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovitis, progressive joint destruction, and disability. Reactive arthritis (ReA) is a sterile joint inflammation following a distant mucosal infection. The clinical course of these diseases is variable and cannot be predicted with reasonable accuracy by clinical and laboratory markers. The predictive value of circulating soluble interleukin-2 receptor (sIL-2R), a marker of lymphocyte activation, measured by Immulite® automated immunoassay analyzer, was evaluated in two cohorts of RA patients. In 175 patients with active early RA randomized to treatment with either on disease-modifying antirheumatic drug (DMARD) or a combination of 3 DMARDs and prednisolone, low baseline sIL-2R level predicted remission after 6 months in patients treated with a single DMARD. In 24 patients with active RA refractory to DMARDs, low baseline sIL-2R level predicted rapid clinical response to treatment with infliximab, an anti-tumour necrosis factor antibody. Furthermore, in a cohort of 26 patients with acute ReA, high baseline sIL-2R level predicted remission after 6 months. Levels of circulating soluble E-selectin (sE-selectin), a marker of endothelial activation, were measured annually by enzyme-linked immunosorbent assay (ELISA) in a cohort of 85 patients with early RA. During a five-year follow-up, sE-selectin levels were associated with activity and outcome of RA. The levels of neutrophil and monocyte CD11b/CD18 expression measured by flow cytometry, and circulating levels of sE-selectin measured by ELISA, and procalcitonin by immunoluminometric assay, were compared in 28 patients with acute ReA and 16 patients with early RA. The levels of the markers were comparable in ReA, RA, and healthy control subjects. In conlusion, sIL-2R may provide a new predictive marker in early RA treated with a single DMARD and refractory RA treated with infliximab. In addition, sIL-2R level predicts remission in acute ReA.Nivelreuma on krooninen tulehduksellinen nivelsairaus jota Suomessa sairastaa noin 1 % väestöstä. Taudinkulku vaihtelee ohimenevästä muutaman nivelen tulehduksesta vakavaan niveliä vaurioittavaan monen nivelen tulehdukseen. Nivelreuman lääkehoito aloitetaan kaikilla potilailla heti diagnoosin varmistuttua, koska nopeasti aloitetun tehokkaan hoidon on osoitettu johtavan parhaaseen hoitotulokseen eikä taudinkulkua voida nykymenetelmillä ennustaa. Mikäli tyydyttävää hoitovastetta ei saavuteta perinteisillä reumalääkkeillä, voidaan hoitoon lisätä niin sanottu biologinen reumalääke. Niillä saadaan hyvä hoitotulos noin kolmasosalla potilaista, mutta ne ovat hyvin kalliita ja niihin liittyy vakavienkin sivuvaikutusten riski. Hoitotulosta ei nykymenetelmillä osata ennustaa. Väitöskirjatutkimuksessani selvitettiin verinäytteistä mitatun lymfosyyttien aktivaatiota kuvastavan liukoisen interleukiini-2 reseptorin (sIL-2R) ennustemerkitystä nivelreumassa. Kun 175 tuoretta aktiivista nivelreumaa sairastavaa potilasta hoidettiin satunnaistetusti joko yhdellä reumalääkkeellä tai usean reumalääkkeen yhdistelmällä, matala sIL-2R-pitoisuus ennen hoidon aloitusta ennakoi hyvää hoitotulosta yhdellä lääkkeellä hoidettujen ryhmässä. Tutkittaessa 24 kroonisesti aktiivista nivelreumaa sairastavaa potilasta joille aloitettiin biologinen reumalääke (infliksimabi), ennusti matala sIL-2R-pitoisuus nopeaa hoitovastetta. Väitöskirjassani tutkittiin myös verinäytteistä mitatun verisuonten sisäkalvon aktivaatiota kuvaavan liukoisen E-selektiinin (sE-selektiini) merkitystä 85 tuoretta nivelreumaa sairastavalla potilaalla. Viiden vuoden seurannan aikana sE-selektiinipitoisuudet olivat yhteydessä taudin aktiivisuuteen ja nivelten vaurioitumiseen. Reaktiivinen niveltulehdus on virtsateiden, hengitysteiden tai suoliston bakteeri-infektion jälkeen alkanut niveltulehdus. Suurin osa potilaista paranee kuukausien kuluessa mutta joillakin potilailla tauti kroonistuu krooniseksi spondyloartropatiaksi. Väitöskirjatutkimuksessani selvitettiin sIL-2R-pitoisuuden merkitystä akuutissa reaktiivisessa niveltulehduksessa. Korkea sIL-2R-pitoisuus ennusti niveltulehduksen paranemista. Väitöskirjatutkimuksessani vertailtiin myös virtaussytometrisesti mitattuja neutrofiilien ja monosyyttien CD11b/CD18-ekspressiota ja verinäytteistä mitattuja sE-selektiini- ja prokalsitoniinipitoisuuksia nivelreumassa ja reaktiivisessa niveltulehduksessa. Sairauksien välillä ei havaittu eroja eivätkä potilaiden tulokset poikenneet terveiden verrokkien tuloksista. Väitöskirjatutkimuksieni perusteella verinäytteestä mitattu sIL-2R-pitoisuus ennustaa hoitovastetta hoidettaessa tuoretta nivelreumaa yhdellä reumalääkkeellä ja kroonista nivelreumaa infliksimabilla. Lisäksi sIL-2R-pitoisuudella on ennustemerkitystä reaktiivisessa niveltulehduksessa. Mikäli löydösten kliininen merkitys voidaan lisätutkimuksissa varmistaa, on mahdollista että tätä verinäytteestä mitattavaa merkkiainetta voidaan tulevaisuudessa käyttää apuna nivelreuman yksilöllisen hoidon suunnittelussa

Activated matrix metalloproteinase 8 in serum predicts severity of acute pancreatitis

Objectives: Severe acute pancreatitis (SAP) has high morbidity and mortality but there are no widely accepted predictive biomarkers in clinical use. Matrix metalloproteinases (MMPs) are active in tissue destruction and inflammatory responses. We studied whether serum levels of activated MMP-8 (aMMP8), MMP-9 and their regulators tissue inhibitor of matrix metalloproteinases (TIMP)-1, myeloperoxidase (MPO) and human neutrophil elastase (HNE) could predict the development of SAP. Methods: The study comprised 214 AP patients (revised Atlanta classification: 142 mild, MAP; 54 moderately severe, MSAP; 18 SAP) referred to Helsinki University Hospital. A venous blood sample was taken within 72 h from the onset of symptoms. Serum levels of aMMP-8 were determined using immunofluorometric assay, and those of MMP-9, TIMP-1, MPO and HNE using enzyme-linked immunosorbent assay. AP groups were compared using Jonckheere-Terpstra test and predictive value for SAP was analyzed using receiver operating characteristics (ROC) analysis. Results: Serum aMMP-8 levels were higher in SAP (median 657 ng/ml, interquartile range 542-738 ng/ ml) compared to MSAP (358 ng/ml, 175-564 ng/ml; p < 0.001) and MAP (231 ng/ml, 128-507 ng/ml; p < 0.001). Similar trend was seen with TIMP-1 and MPO. In ROC analysis aMMP-8, MPO and TIMP-1 emerged as potential markers for the development of SAP (areas under ROC curves 0.83, 0.71 and 0.69, respectively). Conclusions: Serum aMMP-8 measured early in the course of AP (within 72 h of symptom onset) predicted the development of SAP. (c) 2021 The Authors. Published by Elsevier B.V. on behalf of IAP and EPC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).Peer reviewe