Losartan protects endothelium-dependent relaxation in vivo in a murine model of rheumatoid arthritis.

Angiotensin II-type 1 receptor stimulation is recognised to promote inflammation, a state central to the development and maintenance of rheumatoid arthritis. Herein we examined the use of losartan, an angiotensin II-type 1 receptor antagonist, on vascular reactivity, knee joint diameter and behavioural assessment of pain in a Freund's complete adjuvant (FCA) mouse model of joint inflammation. Monoarthritis was induced via FCA in the presence or absence of losartan with naive mice serving as controls. Knee joint swelling, joint pain (assessed by dynamic weight bearing of limb use), knee joint artery reactivity (assessed ex vivo) and blood perfusion of the knee joint (assessed in vivo) were determined. FCA mediated a significant increase in knee joint diameter and reduced weight-bearing (a surrogate for pain sensation) of the affected limb. Notably, these phenomena were substantially reduced when mice were prophylactically treated with losartan. Assessment of arterial relaxation and blood perfusion with acetylcholine stimulation revealed that FCA resulted in significant vascular dysfunction, which was resolved to naïve levels with losartan treatment. Through the actions of losartan, these findings indicate that the angiotensin II-type 1 receptor is a likely therapeutic target of importance in the development of the physical changes, pain sensation and vascular dysfunction found in inflammatory arthritis.

Protease activated receptor 2 and matriptase expression in the joints of cats with and without osteoarthritis.

OBJECTIVES: The aim of this study was to determine the presence of protease-activated receptor 2 (PAR2) and matriptase proteins and quantify PAR2 and matriptase mRNA expression in the articular cartilage and synovial membrane of cats with and without osteoarthritis (OA). METHODS: A total of 28 articular cartilage samples from adult cats (14 OA and 14 normal), 10 synovial membranes from adult cats (five OA and five normal) and three cartilage samples from 9-week-old fetal cats were used. The presence of PAR2 and matriptase in the cartilage and synovial membrane of the adult samples was detected by immunohistochemical (IHC) staining, while real-time PCR was used for mRNA expression analyses in all samples. RESULTS: PAR2 was detected in all OA and normal articular cartilage and synovial membrane samples but confined to only a few superficial chondrocytes in the normal samples. Matriptase was only detected in OA articular cartilage and synovial membrane samples. PAR2 and matriptase mRNA expression were, however, detected in all cartilage and synovial membrane samples. PAR2 and matriptase mRNA expression levels in OA articular cartilage were five (P <0.001) and 3.3 (P <0.001) times higher than that of the healthy group, respectively. There was no significant difference (P = 0.05) in the OA synovial membrane PAR2 and matriptase mRNA expression compared with the normal samples. CONCLUSIONS AND RELEVANCE: Detection of PAR2 and matriptase proteins and gene expression in feline articular tissues is a novel and important finding, and supports the hypothesis that serine proteases are involved in the pathogenesis of feline OA. The consistent presence of PAR2 and matriptase protein in the cytoplasm of OA chondrocytes suggests a possible involvement of proteases in cartilage degradation. Further investigations into the PAR2 and matriptase pathobiology could enhance our understanding of the proteolytic cascades in feline OA, which might lead to the development of novel therapeutic strategies.

Rheumatic Disease: Protease-Activated Receptor-2 in Synovial Joint Pathobiology.

Protease-activated receptor-2 (PAR2) is one member of a small family of transmembrane, G-protein-coupled receptors. These receptors are activated via cleavage of their N terminus by serine proteases (e.g., tryptase), unveiling an N terminus tethered ligand which binds to the second extracellular loop of the receptor. Increasing evidence has emerged identifying key pathophysiological roles for PAR2 in both rheumatoid arthritis (RA) and osteoarthritis (OA). Importantly, this includes both pro-inflammatory and destructive roles. For example, in murine models of RA, the associated synovitis, cartilage degradation, and subsequent bone erosion are all significantly reduced in the absence of PAR2. Similarly, in experimental models of OA, PAR2 disruption confers protection against cartilage degradation, subchondral bone osteosclerosis, and osteophyte formation. This review focuses on the role of PAR2 in rheumatic disease and its potential as an important therapeutic target for treating pain and joint degradation.

Endothelium-derived contraction in a model of rheumatoid arthritis is mediated via angiotensin II type 1 receptors.

A role for endothelium-derived constricting factors (EDCF), and the angiotensin II type 1 receptor (AT1R) pathway, in the vascular impairment found in the rat Freund's complete adjuvant (FCA)-model of rheumatoid arthritis (RA) was examined. FCA arthritis was induced in rats±losartan. Vehicle-treated rats served as controls. Knee-joint swelling and red blood cell (RBC) aggregation were measured as indicators of inflammation and endothelium reactivity assessed by response to acetylcholine (ACh) on aortic rings. Results show that knee-joint swelling and RBC aggregation were elevated in the FCA+vehicle group and restored to control levels in the FCA+losartan-treated animals. ACh-induced relaxation of aortic rings taken from FCA+vehicle animals was significantly impaired compared to vehicle-controls and this vasoreactivity was restored to control levels in the FCA+losartan-treated group. Further examination of aorta from the FCA+vehicle animals revealed an EDCF that was reliant on cyclooxygenase-2 (but not cyclooxygenase-1), generation of superoxide anion generation (but not hydrogen peroxide) and activation of thromboxane-prostanoid receptor. Losartan administration in vivo or ex vivo (to aortic rings) prevented the generation of the EDCF. In summary, this is the first evidence of an EDCF in a model of RA and identifies this mechanism as potentially significant in the cardiovascular disorder associated with the disease.

The serine proteinase hepsin is an activator of pro-matrix metalloproteinases: molecular mechanisms and implications for extracellular matrix turnover.

Increasing evidence implicates serine proteinases in the proteolytic cascades leading to the pathological destruction of extracellular matrices such as cartilage in osteoarthritis (OA). We have previously demonstrated that the type II transmembrane serine proteinase (TTSP) matriptase acts as a novel initiator of cartilage destruction via the induction and activation of matrix metalloproteinases (MMPs). Hepsin is another TTSP expressed in OA cartilage such that we hypothesized this proteinase may also contribute to matrix turnover. Herein, we demonstrate that addition of hepsin to OA cartilage in explant culture induced significant collagen and aggrecan release and activated proMMP-1 and proMMP-3. Furthermore, hepsin directly cleaved the aggrecan core protein at a novel cleavage site within the interglobular domain. Hepsin expression correlated with synovitis as well as tumour necrosis factor α expression, and was induced in cartilage by a pro-inflammatory stimulus. However, a major difference compared to matriptase was that hepsin demonstrated markedly reduced capacity to activate proteinase-activated receptor-2. Overall, our data suggest that hepsin, like matriptase, induces potent destruction of the extracellular matrix whilst displaying distinct efficiencies for the cleavage of specific substrates.

Proteinase-activated receptor 2 modulates OA-related pain, cartilage and bone pathology.

OBJECTIVE: Proteinase-activated receptor 2 (PAR2) deficiency protects against cartilage degradation in experimental osteoarthritis (OA). The wider impact of this pathway upon OA-associated pathologies such as osteophyte formation and pain is unknown. Herein, we investigated early temporal bone and cartilage changes in experimental OA in order to further elucidate the role of PAR2 in OA pathogenesis. METHODS: OA was induced in wild-type (WT) and PAR2-deficient (PAR2-/-) mice by destabilisation of the medial meniscus (DMM). Inflammation, cartilage degradation and bone changes were monitored using histology and microCT. In gene rescue experiments, PAR2-/- mice were intra-articularly injected with human PAR2 (hPAR2)-expressing adenovirus. Dynamic weight bearing was used as a surrogate of OA-related pain. RESULTS: Osteophytes formed within 7 days post-DMM in WT mice but osteosclerosis was only evident from 14 days post induction. Importantly, PAR2 was expressed in the proliferative/hypertrophic chondrocytes present within osteophytes. In PAR2-/- mice, osteophytes developed significantly less frequently but, when present, were smaller and of greater density; no osteosclerosis was observed in these mice up to day 28. The pattern of weight bearing was altered in PAR2-/- mice, suggesting reduced pain perception. The expression of hPAR2 in PAR2-/- mice recapitulated osteophyte formation and cartilage damage similar to that observed in WT mice. However, osteosclerosis was absent, consistent with lack of hPAR2 expression in subchondral bone. CONCLUSIONS: This study clearly demonstrates PAR2 plays a critical role, via chondrocytes, in osteophyte development and subchondral bone changes, which occur prior to PAR2-mediated cartilage damage. The latter likely occurs independently of OA-related bone changes.

Angiotensin II Type 1 receptor blockade protects endothelium-derived hyperpolarising factor-mediated relaxation in a rat model of monoarthritis.

AIMS: Rheumatoid arthritis (RA) is associated with high cardiovascular mortality. Impaired endothelial cell (EC) function and elevated angiotensin II levels may be central to the link between vascular dysfunction and RA. Here we investigated the action of angiotensin type 1 receptor (AT1R) blockade on endothelium-dependent relaxation of the isolated saphenous artery in a rat model of monoarthritis. MAIN METHODS: Adjuvant arthritis was induced in rats with and without prophylactic losartan (AT1R antagonist) treatment. Vehicle-treated rats were used as controls. Wire myography was employed to investigate EC function of isolated rings of saphenous artery. KEY FINDINGS: EC-dependent relaxation in arteries from non-inflamed control rats was mediated by both nitric oxide (NO) and endothelium-derived hyperpolarising factor (EDHF) with the EDHF response dependent principally on functional myoendothelial gap junctions. While NO-dependent relaxation remained unaffected, the EDHF-mediated response was abolished in arteries from arthritic rats (P<0.001), however, substantial protection (approximately 50%) of the EDHF-relaxation was found in arthritic rats treated with losartan (P<0.01). Thus, the attenuated EDHF response found in the saphenous artery of arthritic rats was significantly reversed by AT1R blockade. SIGNIFICANCE: These results suggest a key role for the angiotensin system in the EC dysfunction found in chronic joint inflammation and highlights AT1R as a potential therapeutic target to redress the vascular impairment and mortality associated with RA.

Proteinase-activated receptor-2 modulates human macrophage differentiation and effector function.

Proteinase-activated receptor-2 (PAR-2) was shown to influence immune regulation; however, its role in human macrophage subset development and function has not been addressed. Here, PAR-2 expression and activation was investigated on granulocyte macrophage (GM)-CSF(M1) and macrophage (M)-CSF(M2) macrophages. In both macrophages, the PAR-2-activating peptide, SLIGKV, increased PAR-2 expression and regulated TNF-α and IL-10 secretion in a manner similar to LPS. In addition, HLA-DR on M1 cells also increased. Monocytes matured to an M1 phenotype in the presence of SLIGKV had reduced cell area, and released less TNF-α after LPS challenge compared with vehicle (P < 0.05, n = 3). Cells matured to an M2 phenotype with SLIGKV also had a reduced cell area and made significantly more TNF-α after LPS exposure compared to vehicle (P < 0.05, n = 3) with reduced IL-10 secretion (P < 0.05, n = 3). Thus, PAR-2 activation on macrophage subsets regulates HLA-DR and PAR-2 surface expression, and drives cytokine production. In contrast, PAR-2 activation during M1 or M2 maturation induces altered cell morphology and skewing of phenotype, as evidenced by cytokine secretion. These data suggest a complex role for PAR-2 in macrophage biology and may have implications for macrophage-driven disease in which proteinase-rich environments can influence the immune process directly.

Immunomodulatory role of proteinase-activated receptor-2.

OBJECTIVE: Proteinase-activated receptor-2 (PAR(2)) has been implicated in inflammatory articular pathology. Using the collagen-induced arthritis model (CIA) the authors have explored the capacity of PAR(2) to regulate adaptive immune pathways that could promote autoimmune mediated articular damage. METHODS: Using PAR(2) gene deletion and other approaches to inhibit or prevent PAR(2) activation, the development and progression of CIA were assessed via clinical and histological scores together with ex vivo immune analyses. RESULTS: The progression of CIA, assessed by arthritic score and histological assessment of joint damage, was significantly (p<0.0001) abrogated in PAR(2) deficient mice or in wild-type mice administered either a PAR(2) antagonist (ENMD-1068) or a PAR(2) neutralising antibody (SAM11). Lymph node derived cell suspensions from PAR(2) deficient mice were found to produce significantly less interleukin (IL)-17 and IFNγ in ex vivo recall collagen stimulation assays compared with wild-type littermates. In addition, substantial inhibition of TNFα, IL-6, IL-1ß and IL-12 along with GM-CSF and MIP-1α was observed. However, spleen and lymph node histology did not differ between groups nor was any difference detected in draining lymph node cell subsets. Anticollagen antibody titres were significantly lower in PAR(2) deficient mice. CONCLUSION: These data support an important role for PAR(2) in the pathogenesis of CIA and suggest an immunomodulatory role for this receptor in an adaptive model of inflammatory arthritis. PAR(2) antagonism may offer future potential for the management of inflammatory arthritides in which a proteinase rich environment prevails.

Correlation of protease-activated receptor-2 expression and synovitis in rheumatoid and osteoarthritis.

Protease-activated receptor-2 (PAR-2) is known to be pro-inflammatory and increasing evidence points to an inflammatory component in osteoarthritis. This investigation examined the relationship between synovitis and PAR-2 expression, histological and immunohistochemical analysis being performed on synovial samples obtained from OA and RA patients, along with non-arthritic samples obtained by post mortem (PM). Samples were also analysed for PAR-4 expression, this receptor also having putative pro-inflammatory roles. Analysis involved comparison of inflammatory indices (synovial thickness and monocyte infiltration) with expression of PAR-2 and PAR-4. Synovial explants were also analysed for TNFα generation in the presence of a PAR-2 antagonist (ENMD-1068) or vehicle. OA synovia showed heterogeneity of inflammatory indicators, some samples overlapping with those from the RA cohort whilst others appeared similar to the PM cohort. PAR-2 expression, both in the lining layer and the interstitium, correlated strongly and significantly with synovial thickness (r = 0.91) and monocyte infiltration (r = 0.83), respectively (P < 0.001 in both cases), and this remains significant on individual cohort analysis. PAR-2 was co-localised to CD3 and CD68 cells in RA and OA synovium as well as fibroblasts derived from these synovia. PAR-4 was also expressed, but the relationship with inflammatory indicators was substantially weaker. Inflammatory indicators in OA synovia showed considerable variability, but correlated strongly with PAR-2 expression, suggesting PAR-2 upregulation in synovitis. Heterogeneity of inflammatory indicators was paralleled by wide variation in TNFα generation between samples. Secretion of this cytokine was dose-dependently inhibited by ENMD-1068, providing evidence of a functional role for PAR-2 in promoting synovitis.

Protease-activated receptor 2: a novel pathogenic pathway in a murine model of osteoarthritis.

OBJECTIVE: Osteoarthritis is a global clinical challenge for which no effective disease-modifying agents currently exist. This study identified protease-activated receptor 2 (PAR-2) as a novel pathogenic mechanism and potential therapeutic target in osteoarthritis. METHODS: Experimental osteoarthritis was induced in wild-type and PAR-2-deficient mice by sectioning the medial meniscotibial ligament (MMTL), leading to the development of a mild arthropathy. Cartilage degradation and increased subchondral bone formation were assessed as indicators of osteoarthritis pathology. RESULTS: Four weeks following MMTL section, cartilage erosion and increased subchondral bone formation was evident in wild-type mice but was substantially reduced in PAR-2-deficient mice. Crucially, the therapeutic inhibition of PAR-2 in wild-type mice, using either a PAR-2 antagonist or a monoclonal antibody targeting the protease cleavage site of PAR-2, was also equally effective at reducing osteoarthritis progression in vivo. PAR-2 was upregulated in chondrocytes of wild-type but not sham-operated mice. Wild-type mice showed further joint degradation 8 weeks after the induction of osteoarthritis, but PAR-2-deficient mice were still protected. CONCLUSIONS: The substantial protection from pathology afforded by PAR-2 deficiency following the induction of osteoarthritis provides proof of concept that PAR-2 plays a key role in osteoarthritis and suggests this receptor as a potential therapeutic target.

Matriptase is a novel initiator of cartilage matrix degradation in osteoarthritis.

OBJECTIVE: Increasing evidence implicates serine proteinases in pathologic tissue turnover. The aim of this study was to assess the role of the transmembrane serine proteinase matriptase in cartilage destruction in osteoarthritis (OA). METHODS: Serine proteinase gene expression in femoral head cartilage obtained from either patients with hip OA or patients with fracture to the neck of the femur (NOF) was assessed using a low-density array. The effect of matriptase on collagen breakdown was determined in cartilage degradation models, while the effect on matrix metalloproteinase (MMP) expression was analyzed by real-time polymerase chain reaction. ProMMP processing was determined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis/N-terminal sequencing, while its ability to activate proteinase-activated receptor 2 (PAR-2) was determined using a synovial perfusion assay in mice. RESULTS: Matriptase gene expression was significantly elevated in OA cartilage compared with NOF cartilage, and matriptase was immunolocalized to OA chondrocytes. We showed that matriptase activated proMMP-1 and processed proMMP-3 to its fully active form. Exogenous matriptase significantly enhanced cytokine-stimulated cartilage collagenolysis, while matriptase alone caused significant collagenolysis from OA cartilage, which was metalloproteinase-dependent. Matriptase also induced MMP-1, MMP-3, and MMP-13 gene expression. Synovial perfusion data confirmed that matriptase activates PAR-2, and we demonstrated that matriptase-dependent enhancement of collagenolysis from OA cartilage is blocked by PAR-2 inhibition. CONCLUSION: Elevated matriptase expression in OA and the ability of matriptase to activate selective proMMPs as well as induce collagenase expression make this serine proteinase a key initiator and inducer of cartilage destruction in OA. We propose that the indirect effects of matriptase are mediated by PAR-2, and a more detailed understanding of these mechanisms may highlight important new therapeutic targets for OA treatment.

Getting the most out of Student Selected Components: 12 tips for participating students.

BACKGROUND: Student Selected Components (SSCs) are an established feature of UK undergraduate medical curricula that offer students choice. They represent a large investment in time and resources. Although programmes vary between Schools, the major learning objectives remain broadly similar. Providing students engage fully with the activity, the final learning outcomes should also be comparable. However, engaging effectively and purposefully with such programmes may not be a clear and straightforward process for students. AIM: To present the challenges and solutions to inform students how to derive the greatest benefit from the learning activities in their SSC programmes. METHODS: Synthesis of the accumulated experience over more than 10 years of developing, running and evaluating SSCs by the Directors of SSCs in five Scottish Medical Schools, combined with analysis of course evaluation and student feedback. RESULTS: Consensus defined 12 tips aimed at improving the approach taken by students to their SSCs, and to provide a structure to maximise their final learning outcomes. CONCLUSION: SSC programmes provide diverse opportunities for students to develop and expand their learning. With increasing emphasis being placed upon student assessment to judge a wide range of professional skills and standards into foundation and specialist training, much greater importance is now being given to SSCs as an opportunity for personal, professional and academic developments. However, it is important that this is performed in a purposeful manner to maximise this opportunity. These 12 tips provide guidance to students on how they can maximise the opportunity presented to them by SSCs.

Vascular dysfunction and alteration of novel and classic cardiovascular risk factors in mothers of growth restricted offspring.

OBJECTIVE: Offspring and maternal birthweight are inversely associated with maternal cardiovascular disease. However, whether established or putative novel cardiovascular risk factors including vascular and metabolic function are disrupted in women who delivered small for gestational age (SGA) offspring is unknown. METHODS: Case control study with analysis of inflammatory, lipid, metabolic and haemostatic markers and microvascular function as assessed by laser Doppler iontophoresis 4 years after the index pregnancy in 28 mothers who delivered SGA offspring at term and 29 matched controls. RESULTS: Delivery of a SGA infant was associated with altered lipids [triglyceride median (IQR) mmol/l; control 0.64 (0.49-0.84); SGA 0.95 (0.67-0.95), p=0.012] [cholesterol:HDL ratio: control 2.64 (2.10-3.10); SGA 3.06 (2.65-3.89), p=0.013], systolic blood pressure [control mmHg: 110 (108-118); SGA 120 (110-130), p=0.031], subclinical inflammation [CRP mg/l: control 0.7 (0.3-2.1); SGA 2.2 (1.2-4.0), p=0.002] [IL-6 pg/ml: control 1.2 (0.8-1.4); SGA 1.5 (1.1-2.2), p=0.009] and endothelial activation [ICAM-1 ng/ml: control 237.7 (210.0-279.4); SGA 283.1 (240.5-366.3), p=0.013], with differences robust to confounder adjustment. Endothelium dependent (p=0.003) and independent microvascular function (p<0.001) were also impaired in mothers of SGA offspring. CONCLUSIONS: Mothers of term SGA offspring exhibit perturbation of metabolic and vascular function, which may underlie a lifelong trajectory of impaired health incorporating adverse perinatal and cardiovascular events.

Twelve tips for developing and sustaining a programme of student selected components.

BACKGROUND: Student selected components (SSCs) represent a significant component of medical curricula in the UK and a new approach in medical education. Despite the prominence given to SSCs by the General Medical Council in each of its seminal papers regarding undergraduate medical education, there remains a diverse view of the purpose, outcomes, structure and assessment of SSCs. Many Schools have adopted their own perspective of SSCs and created different but often innovative courses. AIMS: This article brings together the Scottish Medical Schools and their experience in organising SSCs, highlights some of the challenges and offers possible solutions to some of the difficulties encountered. METHOD: The SSC Director from each of the Scottish medical schools each contributed their own '12 Tips'. From these a consensus was achieved. RESULTS: Even though the Scottish medical schools have a wide range of curriculum and timetable formats, there was a great deal of agreement in the challenges and problems encountered in their SSC programmes, as expressed through these 12 Tips. CONCLUSION: There is much diversity in SSC programmes at different medical schools, although there is also much commonality in the challenges that arise. We hope that this paper will promote thought and discussion amongst those involved, and be useful to those involved in curriculum and programme development and also to those new to medical education.

Endothelial function in patients with proteinuric primary glomerulonephritis.

BACKGROUND: Cardiovascular disease is the commonest cause of mortality among patients with end-stage renal disease. Endothelial function and inflammation have previously been shown to be abnormal among such individuals, and are known to be important factors in the progression of atherosclerosis. The aim of this study was to assess endothelial function early in the natural history of renal disease. METHODS: Patients with primary glomerulonephritis, and healthy controls were recruited. In addition to routine laboratory assessment of renal function and proteinuria, assays were undertaken to measure CRP, vWF, VCAM and ICAM. Furthermore, a direct assessment of microvascular endothelial function was undertaken, using laser Doppler imaging to measure perfusion to areas of skin under the influence of transdermally delivered vasodilator agents. RESULTS: Data were collected from 39 patients and 22 controls. No patient was taking anti-platelet agents, statins or angiotensin-converting enzyme inhibitors at the time of endothelial function assessment. All 3 biomarkers of endothelial function were significantly elevated in the patient group compared to controls: ICAM 455 versus 359 ng/ml (p = 0.009), VCAM 1,101 versus 771 ng/ml (p = 0.007) and vWF 184 versus 125 IU/ml (p < 0.001). These differences remained significant after adjusting for blood pressure and body mass index. Endothelium-dependent and endothelium-independent vascular responses were blunted in the patient group, compared to controls (AUC: 2,204 vs. 3,721 PU for dependent and 2,190 vs. 3,555 PU for independent responses). CONCLUSIONS: Microvascular endothelial and vascular smooth muscle function is abnormal in patients with primary glomerulonephritis and moderate proteinuria but well-maintained renal function. We believe these findings to be of particular importance as they compare 2 well-matched groups in the absence of the confounding influence of drugs known to affect endothelial function.

Reproducibility and repeatability of peripheral microvascular assessment using iontophoresis in conjunction with laser Doppler imaging.

Interrogation of peripheral vascular function is increasingly recognized as a noninvasive surrogate marker for coronary vascular function and carries with it important prognostic information regarding future cardiovascular risk. Laser Doppler imaging (LDI) is a completely noninvasive method for looking at peripheral microvascular function. We sought to look at reproducibility and repeatability of LDI-derived assessment of peripheral microvascular function between arms and 8 weeks apart. We used LDI in conjunction with iontophoretic application of ACh and SNP to look at endothelium-dependent and -independent microvascular function, respectively, in a mixture of women with cardiac syndrome X and healthy volunteers. We looked at variation between arms (n = 40) and variation at 8 weeks apart (n = 22). When measurements were corrected for skin resistance, there was nonsignificant variation between arms for ACh (2.7%) and SNP (3.8%) and nonsignificant temporal variation for ACh (3.5%) and SNP (4.7%). Construction of Bland-Altman plots reinforce that measurements have good repeatability. Elimination of the baseline perfusion response had deleterious effects on repeatability. LDI can be used to assess peripheral vascular response with good repeatability as long as measurements are corrected for skin resistance, which affects drug delivery. This has important implications for the future use of LDI.

Differences in arterial compliance, microvascular function and venous capacitance between patients with heart failure and either preserved or reduced left ventricular systolic function.

BACKGROUND: Up to 50% of patients with the clinical syndrome of heart failure have preserved left ventricular systolic function (HF-PSF). These patients may have abnormalities of ventriculo-vascular coupling, due to increased vascular and ventricular stiffness. METHODS: We compared arterial compliance, microvascular vasodilator function and venous capacitance (VC) in 3 groups of patients (n=12 each) matched for the presence of coronary heart disease: 1) HF and preserved systolic function (HF-PSF), 2) HF and reduced systolic function (HF-RSF) and 3) controls (no HF, PSF). Arterial compliance was assessed by measuring aortic pulse wave velocity (PWV) with applanation tonometry. Cutaneous microvascular function was assessed using Laser Doppler imaging (LDI) coupled with iontophoresis of endothelium-dependent (acetylcholine) and -independent (sodium nitroprusside) vasodilators. VC was measured using venous occlusion plethysmography. RESULTS: PWV was significantly higher in HF-PSF subjects than in both HF-RSF and control groups (10.7 [1.1], 8.9 [1.7] and 8.6 [2.1] m/s respectively, p<0.05). Acetylcholine and nitroprusside induced vasodilatation were equally impaired in HF-PSF and HF-RSF, as compared to controls (p<0.01). VC was higher in HF-RSF subjects compared with HF-PSF subjects (1.75 [0.41], 1.34 [0.34] ml/100 ml forearm vol. respectively, p<0.05). CONCLUSIONS: These findings are consistent with a more marked increase in vascular stiffness in HF-PSF than in HF-RSF and suggest that arterial stiffness, dynamic vasodilator function and venous abnormalities may be implicated in the complex pathophysiology of HF-PSF.

Expression and proinflammatory role of proteinase-activated receptor 2 in rheumatoid synovium: ex vivo studies using a novel proteinase-activated receptor 2 antagonist.

OBJECTIVE: Serine proteinases activate the G protein-coupled receptor, proteinase-activated receptor 2 (PAR-2), via cleavage and exposure of a tethered ligand. PAR-2 is known to exert proinflammatory actions in a murine model of arthritis, since PAR-2-deficient mice exhibit strikingly reduced articular inflammation. This study was undertaken to examine synovial PAR-2 expression and to determine the effect of a novel PAR-2 antagonist on synovial cytokine production, in order to investigate the hypothesis that PAR-2 plays a critical role in the pathogenesis of rheumatoid arthritis (RA). METHODS: Using a monoclonal antibody to human PAR-2, expression in RA synovium and cultured synovial fibroblasts was characterized. The novel PAR-2 antagonist, ENMD-1068, was added to primary cultures of RA synovial tissue, from which spontaneous cytokine release was measured. RESULTS: PAR-2 was substantially up-regulated in RA synovium compared with control synovial tissue from patients with osteoarthritis or seronegative inflammatory arthritis, neither of which exhibited significant PAR-2 expression. Importantly, spontaneous release of tumor necrosis factor alpha and interleukin-1beta from RA synovium was substantially inhibited by ENMD-1068, in a dose-dependent manner. CONCLUSION: These findings identify PAR-2 as a novel upstream regulator of proinflammatory cytokine production in RA and indicate its potential as a novel therapeutic target in inflammatory arthritis.