Steven E. Carsons

TNF in combination with GM‐CSF enhances the differentiation of neonatal cord blood stem cells into dendritic cells and macrophages

We describe dendritic cell progenitors within the CD34+ stem cell compartment in neonatal cord blood and identify growth factors contributing to their differentiation. Granulocyte‐macrophage colony‐stimulating factor (GM‐CSF), although mainly promoting the growth and differentiation of monocyte‐macrophages (mono‐mφs), also induced the differentiation of cells with the distinctiv morphological features of dendritic cells (DCs). Tumo necrosis factor (TNF) in combination with GM‐CSF promoted further growth of both cell types but most notably increased the DC content. In situ analysis revealed that the cells exhibiting DC morphology were positive for class II major histocompatibility complex antigens but were CD14 negative, did not exhibit nonspecific esterase activity, and were nonphagocytic. Moreover, the mixed leukocyte reaction stimulatory capacity of cultures with the higher DC content was greater. TNF, interleukin‐1 (IL‐1), IL‐6, or platelet‐derived growth factor (PDGF) was inactive in promoting stem cell proliferation or DC morphology. IL‐1 or PDGF synergized with GM‐CSF to increase mono‐mφ–associated cell proliferation but did not increase the DC content. The development of a common DC‐monocyte precursor was suggested by the presence of colony‐forming unit–like clusters containing mono‐mφ8 and DCs and one sharp proliferative peak. The loss of DC morphology after 21 days, coupled with increases in mono‐mφ–associated markers and a constant number of viable cells, further suggests that DC morphology may fluctuate in culture or is a transient feature acquired by certain cells of the mono‐mφ lineage.

Fibronectin fragments and their role in inflammatory arthritis

OBJECTIVES To identify potential immunopathogenic links between fibronectin (Fn) fragmentation and the inflammatory response in chronic joint disease. METHODS Scientific papers involving studies of Fn fragments and inflammatory processes important in the pathogenesis of arthritis, including chondrolysis, synoviocyte growth and adhesion, polymorphonuclear leukocyte (PMN) and monocyte function, proteolysis, and immune complex activation were reviewed. In addition, reports identifying Fn fragments in synovial fluid (SF) were assessed. RESULTS A series of Fn fragments have been identified in arthritic SF by several investigators. Fn and fragments ranging from 30 to 200 kd are present in elevated concentrations in inflammatory SF. SF Fn fragments display reduced affinity for fibrin and collagen. The 29- and 50-kd amino terminal fragments mediate release of proteoglycan from articular cartilage by RGD-independent mechanisms. Fn fragments can induce fibroblast gene expression of metalloproteinases or can act as proteinases themselves. A 90-kd plasmin generated fragment possesses homology with streptokinase. Fragments mediate PMN chemotaxis and enhance proliferation of CD4+ lymphocytes as well as binding to the C1q component of complement and influencing the behavior of immune complexes. CONCLUSIONS Fn fragments can be functionally and biochemically characterized in diseased SF. Modification of fragment formation and inhibition of fragment function may have potential therapeutic value in the interruption of chronic synovial inflammation.

Atheroprotective effects of methotrexate on reverse cholesterol transport proteins and foam cell transformation in human THP-1 monocyte/macrophages.

OBJECTIVE To determine whether methotrexate (MTX) can overcome the atherogenic effects of cyclooxygenase 2 (COX-2) inhibitors and interferon-gamma (IFNgamma), both of which suppress cholesterol efflux protein and promote foam cell transformation in human THP-1 monocyte/macrophages. METHODS Message and protein levels of the reverse cholesterol transport proteins cholesterol 27-hydroxylase and ATP-binding cassette transporter A1 (ABCA1) in THP-1 cells were evaluated by real-time polymerase chain reaction and immunoblot, respectively. Expression was evaluated in cells incubated in the presence or absence of the COX-2 inhibitor NS398 or IFNgamma, with and without MTX. Foam cell transformation of lipid-laden THP-1 macrophages was detected with oil red O staining and light microscopy. RESULTS MTX increased 27-hydroxylase message and completely blocked NS398-induced down-regulation of 27-hydroxylase (mean +/- SEM 112.8 +/- 13.1% for NS398 plus MTX versus 71.1 +/- 4.3% for NS398 alone; P < 0.01). MTX also negated COX-2 inhibitor-mediated down-regulation of ABCA1. The ability of MTX to reverse inhibitory effects on 27-hydroxylase and ABCA1 was blocked by the adenosine A2A receptor-specific antagonist ZM241385. MTX also prevented NS398 and IFNgamma from increasing transformation of lipid-laden THP-1 macrophages into foam cells. CONCLUSION This study provides evidence supporting the notion of an atheroprotective effect of MTX. Through adenosine A2A receptor activation, MTX promotes reverse cholesterol transport and limits foam cell formation in THP-1 macrophages. This is the first reported evidence that any commonly used medication can increase expression of antiatherogenic reverse cholesterol transport proteins and can counteract the effects of COX-2 inhibition. Our results suggest that one mechanism by which MTX protects against cardiovascular disease in rheumatoid arthritis patients is through facilitation of cholesterol outflow from cells of the artery wall.

Aberrant Extracellular and Dendritic Cell (DC) Surface Expression of Heat Shock Protein (hsp)70 in the Rheumatoid Joint: Possible Mechanisms of hsp/DC-Mediated Cross-Priming

We describe, in rheumatoid arthritis (RA), abnormalities in the expression and distribution of heat shock protein (hsp) and dendritic cells (DCs) that are conducive to cross-priming and DC cross-talk. As detected by ELISA, inducible (i)hsp70 was dramatically increased in RA synovial fluid (RASF) vs normal human and RA sera and osteoarthritis and gout synovial fluid. Immunoblot analysis of fresh RASF cells revealed marked increases in ihsp70 and activation of its transcription factor heat shock factor-1, compared with fresh normal peripheral blood cells. Flow cytometry and microscopy demonstrated high levels of ihsp70 on the surface of RASF myeloid DCs (but not normal myeloid DCs) that occurred concurrently with hspRs (CD91/CD14). ihsp70 present in RASF exhibited chaperoning potential, as indicated by the capture of ihsp70 present in RASF on the surface of normal DCs. Binding was partially competitively inhibited by excess α2-macroglobulin, indicating that hspRs in addition to CD91 participate in the capture process. These data indicate that ihsp70 may chaperone autologous Ags into immature RASF DCs via hspRs, and that cross-talk between DCs coexpressing hsp/hspRs reflects a disease process in RA. The induction of surface ihsp70 on normal cells after sublethal heat stress and the release of ihsp70 from normal DCs after inflammatory stress also suggest that the pattern of ihsp70 expression in RASF occurs in response to sustained stress.

2015 Recommendations for the management of polymyalgia rheumatica: a European League Against Rheumatism/American College of Rheumatology collaborative initiative

Therapy for polymyalgia rheumatica (PMR) varies widely in clinical practice as international recommendations for PMR treatment are not currently available. In this paper, we report the 2015 European League Against Rheumatism (EULAR)/American College of Rheumatology (ACR) recommendations for the management of PMR. We used the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) methodology as a framework for the project. Accordingly, the direction and strength of the recommendations are based on the quality of evidence, the balance between desirable and undesirable effects, patients’ and clinicians’ values and preferences, and resource use. Eight overarching principles and nine specific recommendations were developed covering several aspects of PMR, including basic and follow-up investigations of patients under treatment, risk factor assessment, medical access for patients and specialist referral, treatment strategies such as initial glucocorticoid (GC) doses and subsequent tapering regimens, use of intramuscular GCs and disease modifying anti-rheumatic drugs (DMARDs), as well as the roles of non-steroidal anti-rheumatic drugs and non-pharmacological interventions. These recommendations will inform primary, secondary and tertiary care physicians about an international consensus on the management of PMR. These recommendations should serve to inform clinicians about best practices in the care of patients with PMR.

Resveratrol mediates anti-atherogenic effects on cholesterol flux in human macrophages and endothelium via PPARγ and adenosine.

Resveratrol is a bioactive molecule used in dietary supplements and herbal medicines and consumed worldwide. Known cardioprotective and anti-inflammatory properties of resveratrol have spurred investigation of the mechanisms involved. The present study explored potential atheroprotective actions of resveratrol on cholesterol metabolism in cells of the arterial wall, including human macrophages and arterial endothelium. Using QRT-PCR and Western blotting techniques, we measured expression of the proteins involved in reverse cholesterol transport (ABCA1, ABCG1 and SR-B1) and the scavenger receptors responsible for uptake of modified cholesterol (CD36, SR-A1 and LOX-1). We analyzed the effect of resveratrol on apoA-1-and HDL-mediated cholesterol efflux in human THP-1 macrophages. The effect of resveratrol on oxLDL internalization and foam cell formation were evaluated using confocal and light microscopy. Our data indicate that resveratrol regulates expression of major proteins involved in cholesterol transport, promotes apoA-1 and HDL-mediated efflux, downregulates oxLDL uptake and diminishes foam cell formation. Mechanistically, resveratrol effects were dependent upon PPAR-γ and adenosine 2A receptor pathways. For the first time we demonstrate that resveratrol regulates expression of the cholesterol metabolizing enzyme cytochrome P450 27-hydroxylase, providing efficient cholesterol elimination via formation of oxysterols. This study establishes that resveratrol attenuates lipid accumulation in cultured human macrophages via effects on cholesterol transport. Further in vivo studies are needed to determine whether resveratrol may be an additional resource available to reduce lipid deposition and atherosclerosis in humans.

EULAR Sjögren's syndrome disease activity index (ESSDAI): a user guide

The EULAR Sjögrens syndrome (SS) disease activity index (ESSDAI) is a systemic disease activity index that was designed to measure disease activity in patients with primary SS. With the growing use of the ESSDAI, some domains appear to be more challenging to rate than others. The ESSDAI is now in use as a gold standard to measure disease activity in clinical studies, and as an outcome measure, even a primary outcome measure, in current randomised clinical trials. Therefore, ensuring an accurate and reproducible rating of each domain, by providing a more detailed definition of each domain, has emerged as an urgent need. The purpose of the present article is to provide a user guide for the ESSDAI. This guide provides definitions and precisions on the rating of each domain. It also includes some minor improvement of the score to integrate advance in knowledge of disease manifestations. This user guide may help clinicians to use the ESSDAI, and increase the reliability of rating and consequently of the ability to detect true changes over time. This better appraisal of ESSDAI items, along with the recent definition of disease activity levels and minimal clinically important change, will improve the assessment of patients with primary SS and facilitate the demonstration of effectiveness of treatment for patients with primary SS.

Effect of cyclooxygenase inhibition on cholesterol efflux proteins and atheromatous foam cell transformation in THP-1 human macrophages: a possible mechanism for increased cardiovascular risk

Both selective cyclooxygenase (COX)-2 inhibitors and non-steroidal anti-inflammatory drugs (NSAIDs) have been beneficial pharmacological agents for many patients suffering from arthritis pain and inflammation. However, selective COX-2 inhibitors and traditional NSAIDs are both associated with heightened risk of myocardial infarction. Possible pro-atherogenic mechanisms of these inhibitors have been suggested, including an imbalance in prostanoid production leaving the pro-aggregatory prostaglandins unopposed, but the precise mechanisms involved have not been elucidated. We explored the possibility that downregulation of proteins involved in reverse cholesterol transport away from atheromatous plaques contributes to increased atherogenesis associated with COX inhibition. The reverse cholesterol transport proteins cholesterol 27-hydroxylase and ATP-binding cassette transporter A1 (ABCA1) export cholesterol from macrophages. When mechanisms to process lipid load are inadequate, uncontrolled cholesterol deposition in macrophages transforms them into foam cells, a key element of atheromatous plaques. We showed that in cultured THP-1 human monocytes/macrophages, inhibition of COX-1, COX-2, or both reduced expression of 27-hydroxylase and ABCA1 message (real-time reverse transcription-polymerase chain reaction) and protein (immunoblot). The selective COX-2 inhibitor N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide (NS398) significantly reduced 27-hydroxylase and ABCA1 message (to 62.4% ± 2.2% and 71.1% ± 3.9% of control, respectively). Incubation with prostaglandin (PG) E2 or PGD2 reversed reductions in both of these cholesterol transport proteins induced by NS398. Cholesterol-loaded THP-1 macrophages showed significantly increased foam cell transformation in the presence of NS398 versus control (42.7% ± 6.6% versus 20.1% ± 3.4%, p = 0.04) as determined by oil red O staining. Pharmacological inhibition of COX in monocytes is involved in downregulation of two proteins that mediate cholesterol efflux: cholesterol 27-hydroxylase and ABCA1. Because these proteins are anti-atherogenic, their downregulation may contribute to increased incidence of cardiac events in patients treated with COX inhibitors. Reversal of inhibitory effects on 27-hydroxylase and ABCA1 expression by PGD2 and PGE2 suggests involvement of their respective signaling pathways. NS398-treated THP-1 macrophages show greater vulnerability to form foam cells. Increased cardiovascular risk with COX inhibition may be ascribed at least in part to altered cholesterol metabolism.

2015 recommendations for the management of polymyalgia rheumatica: a European League Against Rheumatism/American College of Rheumatology collaborative initiative.

Therapy for polymyalgia rheumatica (PMR) varies widely in clinical practice as international recommendations for PMR treatment are not currently available. In this paper, we report the 2015 European League Against Rheumatism (EULAR)/American College of Rheumatology (ACR) recommendations for the management of PMR. We used the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) methodology as a framework for the project. Accordingly, the direction and strength of the recommendations are based on the quality of evidence, the balance between desirable and undesirable effects, patients’ and clinicians’ values and preferences, and resource use. Eight overarching principles and nine specific recommendations were developed covering several aspects of PMR, including basic and follow‐up investigations of patients under treatment, risk factor assessment, medical access for patients and specialist referral, treatment strategies such as initial glucocorticoid (GC) doses and subsequent tapering regimens, use of intramuscular GCs and disease modifying anti‐rheumatic drugs (DMARDs), as well as the roles of non‐steroidal anti‐rheumatic drugs and non‐pharmacological interventions. These recommendations will inform primary, secondary and tertiary care physicians about an international consensus on the management of PMR. These recommendations should serve to inform clinicians about best practices in the care of patients with PMR.

Altered Development of NKT Cells, γδ T Cells, CD8 T Cells and NK Cells in a PLZF Deficient Patient

In mice, the transcription factor, PLZF, controls the development of effector functions in invariant NKT cells and a subset of NKT cell-like, γδ T cells. Here, we show that in human lymphocytes, in addition to invariant NKT cells, PLZF was also expressed in a large percentage of CD8+ and CD4+ T cells. Furthermore, PLZF was also found to be expressed in all γδ T cells and in all NK cells. Importantly, we show that in a donor lacking functional PLZF, all of these various lymphocyte populations were altered. Therefore, in contrast to mice, PLZF appears to control the development and/or function of a wide variety of human lymphocytes that represent more than 10% of the total PBMCs. Interestingly, the PLZF-expressing CD8+ T cell population was found to be expanded in the peripheral blood of patients with metastatic melanoma but was greatly diminished in patients with autoimmune disease.