Ramipril attenuates lipid peroxidation and cardiac fibrosis in an experimental model of rheumatoid arthritis

Introduction

Recent studies revealed that co-morbidity and mortality due to cardiovascular disease are increased in patients with rheumatoid arthritis (RA) but little is known about factors involved in these manifestations. This study aimed at characterizing the impact of arthritis on oxidative stress status and tissue fibrosis in the heart of rats with adjuvant-induced arthritis (AIA).

Methods

AIA was induced with complete Freund's adjuvant in female Lewis rats. Animals were treated by oral administration of vehicle or angiotensin-converting enzyme inhibitor ramipril (10 mg/kg/day) for 28 days, beginning 1 day after arthritis induction. Isolated adult cardiomyocytes were exposed to 10 μM 4-hydroxynonenal (HNE) for 24 hours in the presence or absence of 10 μM ramipril.

Results

Compared to controls, AIA rats showed significant 55 and 30% increase of 4-HNE/protein adducts in serum and left ventricular (LV) tissues, respectively. Cardiac mitochondrial NADP+-isocitrate dehydrogenase (mNADP-ICDH) activity decreased by 25% in AIA rats without any changes in its protein and mRNA expression. The loss of mNADP-ICDH activity was correlated with enhanced accumulation of HNE/mNADP-ICDH adducts as well as with decrease of glutathione and NADPH. Angiotensin II type 1 receptor (AT₁R) expression and tissue fibrosis were induced in LV tissues from AIA rats. In isolated cardiomyocytes, HNE significantly decreased mNADP-ICDH activity and enhanced type I collagen and connective tissue growth factor expression. The oral administration of ramipril significantly reduced HNE and AT₁R levels and restored mNADP-ICDH activity and redox status in LV tissues of AIA rats. The protective effects of this drug were also evident from the decrease in arthritis scoring and inflammatory markers.

Conclusion

Collectively, our findings disclosed that AIA induced oxidative stress and fibrosis in the heart. The fact that ramipril attenuates inflammation, oxidative stress and tissue fibrosis may provide a novel strategy to prevent heart diseases in RA.     

Arginase attenuates inhibitory nonadrenergic noncholinergic nerve-induced nitric oxide generation and airway smooth muscle relaxation

Background

Recent evidence suggests that endogenous arginase activity potentiates airway responsiveness to methacholine by attenuation of agonist-induced nitric oxide (NO) production, presumably by competition with epithelial constitutive NO synthase for the common substrate, L-arginine. Using guinea pig tracheal open-ring preparations, we now investigated the involvement of arginase in the modulation of neuronal nitric oxide synthase (nNOS)-mediated relaxation induced by inhibitory nonadrenergic noncholinergic (iNANC) nerve stimulation.

Methods

Electrical field stimulation (EFS; 150 mA, 4 ms, 4 s, 0.5 – 16 Hz)-induced relaxation was measured in tracheal preparations precontracted to 30% with histamine, in the presence of 1 μM atropine and 3 μM indomethacin. The contribution of NO to the EFS-induced relaxation was assessed by the nonselective NOS inhibitor L-NNA (0.1 mM), while the involvement of arginase activity in the regulation of EFS-induced NO production and relaxation was investigated by the effect of the specific arginase inhibitor nor-NOHA (10 μM). Furthermore, the role of substrate availability to nNOS in EFS-induced relaxation was measured in the presence of various concentrations of exogenous L-arginine.

Results

EFS induced a frequency-dependent relaxation, ranging from 6.6 ± 0.8% at 0.5 Hz to 74.6 ± 1.2% at 16 Hz, which was inhibited with the NOS inhibitor L-NNA by 78.0 ± 10.5% at 0.5 Hz to 26.7 ± 7.7% at 8 Hz (P < 0.01 all). In contrast, the arginase inhibitor nor-NOHA increased EFS-induced relaxation by 3.3 ± 1.2-fold at 0.5 Hz to 1.2 ± 0.1-fold at 4 Hz (P < 0.05 all), which was reversed by L-NNA to the level of control airways in the presence of L-NNA (P < 0.01 all). Similar to nor-NOHA, exogenous L-arginine increased EFS-induced airway relaxation (P < 0.05 all).

Conclusion

The results indicate that endogenous arginase activity attenuates iNANC nerve-mediated airway relaxation by inhibition of NO generation, presumably by limiting L-arginine availability to nNOS.     

RANKL inhibition by osteoprotegerin prevents bone loss without affecting local or systemic inflammation parameters in two rat arthritis models: comparison with anti-TNFα or anti-IL-1 therapies

Introduction

Rat adjuvant-induced arthritis (AIA) and collagen-induced arthritis (CIA) feature bone loss and systemic increases in TNFα, IL-1β, and receptor activator of NF-κB ligand (RANKL). Anti-IL-1 or anti-TNFα therapies consistently reduce inflammation in these models, but systemic bone loss often persists. RANKL inhibition consistently prevents bone loss in both models without reducing joint inflammation. Effects of these therapies on systemic markers of bone turnover and inflammation have not been directly compared.

Methods

Lewis rats with established AIA or CIA were treated for 10 days (from day 4 post onset) with either PBS (Veh), TNFα inhibitor (pegsunercept), IL-1 inhibitor (anakinra), or RANKL inhibitor (osteoprotegerin (OPG)-Fc). Local inflammation was evaluated by monitoring hind paw swelling. Bone mineral density (BMD) of paws and lumbar vertebrae was assessed by dual X-ray absorptiometry. Markers and mediators of bone resorption (RANKL, tartrate-resistant acid phosphatase 5b (TRACP 5B)) and inflammation (prostaglandin E₂ (PGE₂), acute-phase protein alpha-1-acid glycoprotein (α₁AGP), multiple cytokines) were measured in serum (day 14 post onset).

Results

Arthritis progression significantly increased paw swelling and ankle and vertebral BMD loss. Anti-TNFα reduced paw swelling in both models, and reduced ankle BMD loss in AIA rats. Anti-IL-1 decreased paw swelling in CIA rats, and reduced ankle BMD loss in both models. Anti-TNFα and anti-IL-1 failed to prevent vertebral BMD loss in either model. OPG-Fc reduced BMD loss in ankles and vertebrae in both models, but had no effect on paw swelling. Serum RANKL was elevated in AIA-Veh and CIA-Veh rats. While antiTNFα and anti-IL-1 partially normalized serum RANKL without any changes in serum TRACP 5B, OPG-Fc treatment reduced serum TRACP 5B by over 90% in both CIA and AIA rats. CIA-Veh and AIA-Veh rats had increased serum α₁AGP, IL-1β, IL-8 and chemokine (C-C motif) ligand 2 (CCL2), and AIA-Veh rats also had significantly greater serum PGE₂, TNFα and IL-17. Anti-TNFα reduced systemic α₁AGP, CCL2 and PGE₂ in AIA rats, while anti-IL-1 decreased systemic α₁AGP, IL-8 and PGE₂. In contrast, RANKL inhibition by OPG-Fc did not lessen systemic cytokine levels in either model.

Conclusions

Anti-TNFα or anti-IL-1 therapy inhibited parameters of local and systemic inflammation, and partially reduced local but not systemic bone loss in AIA and CIA rats. RANKL inhibition prevented local and systemic bone loss without significantly inhibiting local or systemic inflammatory parameters.     

Les inhibiteurs de la phosphodiestérase de type 5 : une révolution dans le traitement des sympt?mes du bas appareil urinaire?

Context

The incidence of lower urinary tract symptoms (LUTS) related to benign prostatic hyperplasia (BPH) increases with age, affecting 50% of patients aged over 50 years and 90% of those aged over 80 years. The prevalence and severity of erectile dysfunction (ED) also increase with age. Its prevalence is estimated to be 31.6% in men over 40 years. LUTS as well as ED significantly affect the quality of life of patients and their partners. Several studies have shown that LUTS represent an independent risk factor for ED. The severity of LUTS is correlated with that of ED. The pathophysiological hypothesis linking LUTS to ED are an increase in sympathetic tone, alteration of NO/cGMP system, alteration of rho kinase system, and pelvic atherosclerosis.

Goal

Some treatments of LUTS have adverse effects on the erectile function. The phosphodiesterase type 5 inhibitors (IPDE 5) revolutionized the treatment of ED.

Material and methods

Several recent clinical studies evaluated the effect of daily treatment by IPDE 5 on LUTS secondary to BPH among patients with or without ED.

Results

This review shows that IPDE 5 administration improves LUTS significantly in 12 randomized clinical trials, as well as in both storage and voiding parts of the international prostate symptom score (IPSS) and in quality of life questionnaire. No adverse events were observed, and ED, which has a high prevalence among this population, was also improved.

Conclusion

The treatment of LUTS by IPDE 5 looks very promising, even though they are not yet approved for this indication in France.     

Functional claudication distance: a reliable and valid measurement to assess functional limitation in patients with intermittent claudication

Background

Pharmacological inhibition of endothelial arginase-II has been shown to improve endothelial nitric oxide synthase (eNOS) function and reduce atherogenesis in animal models. We investigated whether the endothelial arginase II is involved in inflammatory responses in endothelial cells.

Methods

Human endothelial cells were isolated from umbilical veins and stimulated with TNFα (10 ng/ml) for 4 hours. Endothelial expression of the inflammatory molecules i.e. vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin were assessed by immunoblotting.

Results

The induction of the expression of endothelial VCAM-1, ICAM-1 and E-selectin by TNFα was concentration-dependently reduced by incubation of the endothelial cells with the arginase inhibitor L-norvaline. However, inhibition of arginase by another arginase inhibitor S-(2-boronoethyl)-L-cysteine (BEC) had no effects. To confirm the role of arginase-II (the prominent isoform expressed in HUVECs) in the inflammatory responses, adenoviral mediated siRNA silencing of arginase-II knocked down the arginase II protein level, but did not inhibit the up-regulation of the adhesion molecules. Moreover, the inhibitory effect of L-norvaline was not reversed by the NOS inhibitor L-NAME and L-norvaline did not interfere with TNFα-induced activation of NF-κB, JNK, p38mapk, while it inhibited p70s6k (S6K1) activity. Silencing S6K1 prevented up-regulation of E-selectin, but not that of VCAM-1 or ICAM-1 induced by TNFα.

Conclusion

The arginase inhibitor L-norvaline exhibits anti-inflammatory effects independently of inhibition of arginase in human endothelial cells. The anti-inflammatory properties of L-norvaline are partially attributable to its ability to inhibit S6K1.     

The severity of experimental arthritis is independent of IL-36 receptor signaling

Introduction

Interleukin (IL)-36 refers to three related IL-1 family cytokines, IL-36α, IL-36β, and IL-36γ, that bind to the IL-36 receptor (IL-36R). IL-36 exerts proinflammatory effects in skin and lung and stimulates T cell responses. In the present study, we examined the expression and function of IL-36R and its ligands in experimental arthritis.

Methods

Collagen-induced arthritis (CIA), antigen-induced arthritis (AIA), and K/BxN serum transfer-induced arthritis were induced according to standard protocols. Messenger RNA levels for IL-36R and its ligands in the joints of mice with CIA were determined by RT-qPCR. Mice with CIA were injected with a blocking monoclonal anti-IL-36R, a blocking anti-IL-1RI, or their isotype-matched control antibodies at the time of arthritis onset. Anti-IL-36R or control antibodies were also injected at the time of AIA induction. Finally, IL-36R-deficient mice were examined in AIA and serum transfer-induced arthritis. The development and severity of arthritis were assessed by clinical and histological scoring.

Results

IL-36R, IL-36Ra and IL-36γ mRNA were detected in the joints of mice with CIA, but their levels did not correlate with arthritis severity. As opposed to anti-IL-1RI antibody treatment, the injection of an anti-IL-36R antibody was devoid of effect on the development and severity of CIA. The severity of joint inflammation and structural damage in AIA was also unaltered by anti-IL-36R antibody treatment. Finally, the severity of AIA and K/BxN serum transfer-induced arthritis was similar in IL-36R-deficient and wild-type mice.

Conclusions

The development and severity of experimental arthritis are independent of IL-36R signaling.     

Arginase inhibition restores in vivo coronary microvascular function in type 2 diabetic rats

Nitric oxide (NO) is crucial for maintaining normal endothelial function and vascular integrity. Increased arginase activity in diabetes might compete with NO synthase (NOS) for their common substrate arginine, resulting in diminished production of NO. The aim of this study was to evaluate coronary microvascular function in type 2 diabetic Goto-Kakizaki (GK) rats using in vivo coronary flow velocity reserve (CFVR) and the effect of arginase inhibition to restore vascular function. Different groups of GK and Wistar rats were given vehicle, the arginase inhibitor N(ω)-hydroxy-nor-l-arginine (nor-NOHA), l-arginine, and the NOS inhibitor N(G)-monomethyl -l-arginine (l-NMMA). GK rats had impaired CFVR compared with Wistar rats (1.31 ± 0.09 vs. 1.87 ± 0.05, P < 0.001). CFVR was restored by nor-NOHA treatment compared with vehicle in GK rats (1.71 ± 0.13 vs. 1.23 ± 0.12, P < 0.05) but remained unchanged in Wistar rats (1.88 ± 0.10 vs. 1.79 ± 0.16). The beneficial effect of nor-NOHA in GK rats was abolished after NOS inhibition. CFVR was not affected by arginine compared with vehicle. Arginase II expression was increased in the aorta and myocardium from GK rats compared with Wistar rats. Citrulline-to-ornithine and citrulline-to-arginine ratios measured in plasma increased significantly more in GK rats than in Wistar rats after nor-NOHA treatment, suggesting a shift of arginine utilization from arginase to NOS. In conclusion, coronary artery microvascular function is impaired in the type 2 diabetic GK rat. Treatment with nor-NOHA restores the microvascular function by a mechanism related to increased utilization of arginine by NOS and increased NO availability.     

RANKL synthesized by articular chondrocytes contributes to juxta-articular bone loss in chronic arthritis

Introduction

The receptor activator nuclear factor-kappaB ligand (RANKL) diffuses from articular cartilage to subchondral bone. However, the role of chondrocyte-synthesized RANKL in rheumatoid arthritis-associated juxta-articular bone loss has not yet been explored. This study aimed to determine whether RANKL produced by chondrocytes induces osteoclastogenesis and juxta-articular bone loss associated with chronic arthritis.

Methods

Chronic antigen-induced arthritis (AIA) was induced in New Zealand (NZ) rabbits. Osteoarthritis (OA) and control groups were simultaneously studied. Dual X-ray absorptiometry of subchondral knee bone was performed before sacrifice. Histological analysis and protein expression of RANKL and osteoprotegerin (OPG) were evaluated in joint tissues. Co-cultures of human OA articular chondrocytes with peripheral blood mononuclear cells (PBMCs) from healthy donors were stimulated with macrophage-colony stimulating factor (M-CSF) and prostaglandin E₂ (PGE₂), then further stained with tartrate-resistant acid phosphatase.

Results

Subchondral bone loss was confirmed in AIA rabbits when compared with controls. The expression of RANKL, OPG and RANKL/OPG ratio in cartilage were increased in AIA compared to control animals, although this pattern was not seen in synovium. Furthermore, RANKL expression and RANKL/OPG ratio were inversely related to subchondral bone mineral density. RANKL expression was observed throughout all cartilage zones of rabbits and was specially increased in the calcified cartilage of AIA animals. Co-cultures demonstrated that PGE₂-stimulated human chondrocytes, which produce RANKL, also induce osteoclasts differentiation from PBMCs.

Conclusions

Chondrocyte-synthesized RANKL may contribute to the development of juxta-articular osteoporosis associated with chronic arthritis, by enhancing osteoclastogenesis. These results point out a new mechanism of bone loss in patients with rheumatoid arthritis.     

Arginase Inhibition Ameliorates Hepatic Metabolic Abnormalities in Obese Mice

Objectives

We examined whether arginase inhibition influences hepatic metabolic pathways and whole body adiposity in diet-induced obesity.

Methods and Results

After obesity induction by a high fat diet (HFD), mice were fed either the HFD or the HFD with an arginase inhibitor, N^ω-hydroxy-nor-L-arginine (nor-NOHA). Nor-NOHA significantly prevented HFD-induced increases in body, liver, and visceral fat tissue weight, and ameliorated abnormal lipid profiles. Furthermore, nor-NOHA treatment reduced lipid accumulation in oleic acid-induced hepatic steatosis in vitro. Arginase inhibition increased hepatic nitric oxide (NO) in HFD-fed mice and HepG2 cells, and reversed the elevated mRNA expression of hepatic genes in lipid metabolism. Expression of phosphorylated 5′ AMPK-activated protein kinase α was increased by arginase inhibition in the mouse livers and HepG2 cells.

Conclusions

Arginase inhibition ameliorated obesity-induced hepatic lipid abnormalities and whole body adiposity, possibly as a result of increased hepatic NO production and subsequent activation of metabolic pathways involved in hepatic triglyceride metabolism and mitochondrial function.     

La dysfonction érectile associée à une hypertrophie bénigne de prostate (HBP) symptomatique: son lien avec le stade évolutif de l’HBP, et son évolution sous différentes thérapeutiques

Introduction

Although the relationship between the lower urinary tract symptoms (LUTS) and the erectile dysfunction (ED) is no more debated, its underlying mechanism remains obscure so far. Several studies emphasized the correlation between the severity of LUTS and the sexual function, and the impact of the different medications used. This study is the first to highlight the association between the stage of evolution of BPH (complicated and noncomplicated BPH) and the severity of the ED.

Objectives

To assess the correlation between ED and the stage of evolution of BPH, and to evaluate the impact of different medications on ED.

Patients and methods

This is a prospective trial relating of 100 patients admitted for urologic consultation, in the Universitary Hospital Center of Fez in Morocco, in a period of 12 months. To evaluate the severity of ED, we used International Index of Erectile Function (IIEF). In our patients, it was not possible to use the International Prostate Symptom Score (IPSS) to assess the severity of urinary symptoms, and it was not possible to date exactly the beginning of LUTS. Hence, we studied patients’ age, the stage of evolution of BPH (complicated or noncomplicated BPH) and the response of the ED to different treatments.

Results

The average man age was 64.3 years. Forty patients had complicated BPH and 60 patients had noncomplicated BPH. Thirty patients (75%) among 40 with complicated BPH had severe ED, whereas an ED rate of 33% (20 patients) was noticed in patients presenting with noncomplicated BPH. Alpha-blockers (tamsulosin) improved erectile function in 12 patients (20%) among those with noncomplicated BPH. Patients presenting with complicated BPH underwent surgical procedure (either transurethral resection of the prostate or open prostatectomy). Erectile function was not statistically improved in this group of patients.

Conclusion

ED showed a correlation with the stage of evolution of symptomatic BPH. Indeed, the risk of ED is higher in patients with complicated BPH. The alphablockers improved the erectile function in the group of noncomplicated BPH, contrary to the surgical approach.     

Effects of the New Arginase Inhibitor N-Hydroxy-nor- -Arginine on NO Synthase Activity in Murine Macrophages

In stimulated murine macrophage, arginase and nitric oxide synthase (NOS) compete for their common substrate, -arginine. The objectives of this study were (i) to test the new α-amino acid N^ω-hydroxy-nor- -arginine (nor-NOHA) as a new selective arginase inhibitor and (ii) to elucidate the effects of arginase inhibition on -arginine utilization by an inducible NOS. Nor-NOHA is about 40-fold more potent than N^ω-hydroxy- -arginine (NOHA), an intermediate in the -arginine/NO pathway, to inhibit the hydrolysis of -arginine to -ornithine catalyzed by unstimulated murine macrophages (IC₅₀ values 12 ± 5 and 400 ± 50 μM, respectively). Stimulation of murine macrophages with interferon-γ and lipopolysaccharide (IFN-γ + LPS) results in clear expression of an inducible NOS (iNOS) and to an increase in arginase activity. Nor-NOHA is also a potent inhibitor of arginase in IFN-γ + LPS-stimulated macrophage (IC₅₀ value 10 ± 3 μM). In contrast to NOHA, nor-NOHA is neither a substrate nor an inhibitor for iNOS and it appears as a useful tool to study the interplays between arginase and NOS. Inhibition of arginase by nor-NOHA increases nitrite and -citrulline accumulation for incubation times higher than 12 h, under our conditions. Our results allow the determination of the kinetic parameters of the two competitive pathways and the proposal of a simple model which readily explains the differences observed between experiments. This model readily accounts for the observed effects and should be useful to predict the consequences of arginase inhibition in the presence of an active NOS on -arginine availability.     

Study of the effects of oral zinc supplementation on peroxynitrite levels, arginase activity and NO synthase activity in seminal plasma of Iraqi asthenospermic patients

Background

Low concentrations of nitric oxide (NO) are necessary for the biology and physiology of spermatozoa, but high levels of NO are toxic and have negative effects on sperm functions. Although several studies have considered the relationship between infertility and semen NO concentrations, no study on the effects of asthenospermia treatments such as oral zinc supplementation on concentrations of NO, which are important in fertility, has been reported. Studies have shown that oral zinc supplementation develops sperm count, motility and the physical characteristics of sperm in animals and in some groups of infertile men. The present study was conducted to study the effect of zinc supplementation on the quantitative and qualitative characteristics of semen, along with enzymes of the NO pathway in the seminal plasma of asthenospermic patients.

Methods

Semen samples were obtained from 60 fertile and 60 asthenozoospermic infertile men of matched age. The subfertile group was treated with zinc sulfate; each participant took two capsules (220 mg per capsule) per day for 3 months. Semen samples were obtained (before and after zinc sulfate supplementation). After liquefaction of the seminal fluid at room temperature, routine semen analyses were performed. The stable metabolites of NO (nitrite) in seminal plasma were measured by nitrophenol assay. Arginase activity and NO synthase activity were measured spectrophotometrically.

Results

Peroxynitrite levels, arginase activity, NO synthase activity and various sperm parameters were compared among fertile controls and infertile patients (before and after treatment with zinc sulfate). Peroxynitrite levels and NO synthase activity were significantly higher in the infertile patients compared to the fertile group. Conversely, arginase activity was significantly higher in the fertile group than the infertile patients. Peroxynitrite levels, arginase activity and NO synthase activity of the infertile patient were restored to normal values after treatment with zinc sulfate. Volume of semen, progressive sperm motility percentage and total normal sperm count were increased after zinc supplementation.

Conclusions

Treatment of asthenospermic patients with zinc supplementation leads to restored peroxynitrite levels, arginase activity and NO synthase activity to normal values and gives a statistically significant improvement of semen parameters compared with controls.

Trial registration

ClinicalTrials.gov identifier: NCT01684059     

Erectile dysfunction is frequent in systemic sclerosis and associated with severe disease: a study of the EULAR Scleroderma Trial and Research group

Introduction

Erectile dysfunction (ED) is common in men with systemic sclerosis (SSc) but the demographics, risk factors and treatment coverage for ED are not well known.

Method

This study was carried out prospectively in the multinational EULAR Scleroderma Trial and Research database by amending the electronic data-entry system with the International Index of Erectile Function-5 and items related to ED risk factors and treatment. Centres participating in this EULAR Scleroderma Trial and Research substudy were asked to recruit patients consecutively.

Results

Of the 130 men studied, only 23 (17.7%) had a normal International Index of Erectile Function-5 score. Thirty-eight per cent of all participants had severe ED (International Index of Erectile Function-5 score ≤ 7). Men with ED were significantly older than subjects without ED (54.8 years vs. 43.3 years, P < 0.001) and more frequently had simultaneous non-SSc-related risk factors such as alcohol consumption. In 82% of SSc patients, the onset of ED was after the manifestation of the first non-Raynaud's symptom (median delay 4.1 years). ED was associated with severe cutaneous, muscular or renal involvement of SSc, elevated pulmonary pressures and restrictive lung disease. ED was treated in only 27.8% of men. The most common treatment was sildenafil, whose efficacy is not established in ED of SSc patients.

Conclusions

Severe ED is a common and early problem in men with SSc. Physicians should address modifiable risk factors actively. More research into the pathophysiology, longitudinal development, treatment and psychosocial impact of ED is needed.     

Human articular chondrocytes express 15-lipoxygenase-1 and -2: potential role in osteoarthritis

Introduction

Although cardiovascular morbidity and mortality are increased in rheumatoid arthritis, little is known about the burden of subclinical coronary atherosclerosis in these patients.

Methods

Using computed tomography, coronary artery calcification was measured in 195 men and women with rheumatoid arthritis aged 45 to 84 years without clinical cardiovascular disease and compared with 1,073 controls without rheumatoid arthritis enrolled in the Baltimore cohort of the Multi-Ethnic Study of Atherosclerosis.

Results

The prevalence of coronary calcification (Agatston score > 0) was significantly higher in men, but not women, with rheumatoid arthritis after adjusting for sociodemographic and cardiovascular risk factors (prevalence ratio = 1.19; P = 0.012). Among participants with prevalent calcification, those with rheumatoid arthritis had adjusted mean Agatston scores 53 units higher than controls (P = 0.002); a difference greater for men than women (P for interaction = 0.017). In all analyses, serum IL-6 attenuated the association between rheumatoid arthritis and coronary calcification, suggesting its role as a potential mediator of enhanced atherosclerosis. Notably, increasing severity of rheumatoid arthritis was associated with a higher prevalence and extent of coronary calcification among both men and women with rheumatoid arthritis, and for all age categories. The largest percentage difference in coronary arterial calcification between rheumatoid arthritis patients and their nonrheumatoid arthritis counterparts was observed in the youngest age category.

Conclusions

Increasing rheumatoid arthritis disease severity was associated with a higher prevalence and greater extent of coronary artery calcification, potentially mediated through an atherogenic effect of chronic systemic inflammation. Gender and age differences in association with coronary calcification suggest that preventive measures should be emphasized in men with rheumatoid arthritis, and considered even in younger rheumatoid arthritis patients with low levels of traditional cardiovascular risk factors.     

Toll-like receptor homolog RP105 modulates the antigen-presenting cell function and regulates the development of collagen-induced arthritis

Introduction

RP105 is a Toll-like receptor homolog expressed on B cells, dendritic cells (DCs), and macrophages. We investigated the role of RP105 in the development of collagen-induced arthritis (CIA).

Methods

CIA was induced in RP105-deficient DBA/1 mice and the incidence and arthritis index were analyzed. The cytokine production by spleen cells was determined. The functions of the DCs and regulatory T cells (Tregs) from RP105-deficient or control mice were determined by adding these cells to the lymph node cell culture. Arthritis was also induced by incomplete Freund's adjuvant (IFA) plus collagen or by injecting anti-collagen antibody and lipopolysaccharide.

Results

RP105-deficient mice showed accelerated onset of arthritis and increased severity. Interferon-gamma (IFN-γ) and tumor necrosis factor-alpha production by spleen cells from RP105-deficient mice was increased in comparison with that from wild-type mice. The DCs from RP105-deficient mice induced more IFN-γ production, whereas Tregs from those mice showed less inhibitory effect against IFN-γ production. RP105-deficient mice also showed more severe arthritis induced by collagen with IFA.

Conclusions

These results indicate that RP105 regulates the antigen-presenting cell function and Treg development, which induced the attenuation of the cell-mediated immune responses and, as a result, suppressed the development of CIA.     

Is arginase a potential drug target in tobacco-induced pulmonary endothelial dysfunction?

Background

Tobacco-induced pulmonary vascular disease is partly driven by endothelial dysfunction. The bioavailability of the potent vasodilator nitric oxide (NO) depends on competition between NO synthase-3 (NOS3) and arginases for their common substrate (L-arginine). We tested the hypothesis whereby tobacco smoking impairs pulmonary endothelial function via upregulation of the arginase pathway.

Methods

Endothelium-dependent vasodilation in response to acetylcholine (Ach) was compared ex vivo for pulmonary vascular rings from 29 smokers and 10 never-smokers. The results were expressed as a percentage of the contraction with phenylephrine. We tested the effects of L-arginine supplementation, arginase inhibition (by N(omega)-hydroxy-nor-l-arginine, NorNOHA) and NOS3 induction (by genistein) on vasodilation. Protein levels of NOS3 and arginases I and II in the pulmonary arteries were quantified by Western blotting.

Results

Overall, vasodilation was impaired in smokers (relative to controls; p < 0.01). Eleven of the 29 smokers (the ED⁺ subgroup) displayed endothelial dysfunction (defined as the absence of a relaxant response to Ach), whereas 18 (the ED⁻ subgroup) had normal vasodilation. The mean responses to 10⁻⁴ M Ach were −23 ± 10% and 31 ± 4% in the ED⁺ and ED⁻ subgroups, respectively (p < 0.01). Supplementation with L- arginine improved endothelial function in the ED⁺ subgroup (−4 ± 10% vs. -32 ± 10% in the presence and absence of L- arginine, respectively; p = 0.006), as did arginase inhibition (18 ± 9% vs. -1 ± 9%, respectively; p = 0.0002). Arginase I protein was overexpressed in ED⁺ samples, whereas ED⁺ and ED⁻ samples did not differ significantly in terms of NOS3 expression. Treatment with genistein did not significantly improve endothelial function in ED⁺ samples.

Conclusion

Overexpression and elevated activity of arginase I are involved in tobacco-induced pulmonary endothelial dysfunction.     

PAD4 is not essential for disease in the K/BxN murine autoantibody-mediated model of arthritis

Introduction

Both murine and human genome-wide association studies have implicated peptidyl arginine deiminase (PAD4) as a susceptibility gene in rheumatoid arthritis (RA). In addition, patients with RA commonly have autoantibodies which recognize PAD4 or and/or citrullinated peptides. This study aims to evaluate the role of PAD4 in the effector phase of arthritis.

Methods

PAD4 knock out (KO) and wild type (WT) C57BL/6J mice were injected with K/BxN sera to induce disease. Progression of disease was monitored by measuring paw and ankle swelling and clinical indexes of disease, and pathogenesis was assessed by indexing of clinical progression on paws collected from WT and PAD4 KO mice injected with K/BxN serum. PAD4 activity was determined by visualization of neutrophil extracellular traps (NETs) and immunohistological analysis of histone citrullination.

Results

PAD4 activity is readily detectable in the inflamed synovium of WT but not PAD4 deficient animals, as demonstrated by histone citrullination and NET formation. However, PAD4 WT and KO animals develop K/BxN serum transfer disease with comparable severity and kinetics, with no statistically significant differences noted in clinical scores, swelling, joint erosion or joint invasion.

Conclusions

PAD4 WT and KO mice develop disease in the K/BxN serum transfer model of arthritis with similar severity and kinetics, indicating that PAD4 is dispensable in this effector phase model of disease.     

Nitric oxide synthase activity and the level of nitrates/nitrites in brain regions during spontaneous morphine withdrawal in rats

Activity of nitric oxide synthase (NOS) and concentrations of nitrate/nitrites (NO _x ^? ) were measured in brain regions of rats during spontaneous morphine withdrawal, which was modeled in male Wistar rats. The animals were injected with the increasing intraperitoneal doses (10–100 mg/kg, twice a day) of morphine hydrochloride for 6 days. Thirty six hours after the last injection the severity of the spontaneous morphine withdrawal syndrome was determined by specific autonomic and locomotor indices The withdrawal was accompanied by the increase of both NOS activity and NO _x ^? levels in the midbrain and hippocampus, the decrease of these parameters in striatum and hypothalamus, and lack of changes in cerebral cortex and brain stem. In cerebellum NOS activity decreased whereas NO _x ^? concentrations remained unchanged. In the cerebral cortex, striatum, midbrain, and cerebellum activity of NOS and NO _x ^? concentrations correlated with the withdrawal syndrome severity and also with the specific signs of abstinence.     

Estradiol ameliorates arthritis and protects against systemic bone loss in Staphylococcus aureus infection in mice

Introduction

Staphylococcus aureus is a common cause of bacterial arthritis, which is associated with progressive bone loss in affected joints. We recently showed that S. aureus infection also induces a significant systemic bone loss in mice. This study was performed to assess the effect of estradiol treatment on the clinical course and outcome of S. aureus arthritis and on infection-induced bone loss in experimental S. aureus infection.

Methods

Mice were ovariectomized, treated with estradiol or placebo, and S. aureus infection was established by intravenous inoculation of bacteria.

Results

Estradiol treatment was found to decrease significantly the frequency and clinical severity of S. aureus arthritis, a finding that was accompanied with significantly higher serum levels of interleukin-10 in estradiol-treated mice. Estradiol was also highly protective against S. aureus-induced systemic trabecular, and cortical bone loss. Lack of endogenous estrogens and S. aureus infection had additive effects on trabecular bone loss. The S. aureus-infected, ovariectomized mice lost as much as 76% of their trabecular bone mass.

Conclusions

Treatment with estradiol ameliorates S. aureus arthritis and is protective against infection-induced systemic bone loss in experimental S. aureus infection.     

Angiotensin II type 2 receptor correlates with therapeutic effects of losartan in rats with adjuvant‐induced arthritis

The angiotensin II type 1 receptor (AT1R) blocker losartan ameliorates rheumatoid arthritis (RA) in an experimental model. In RA, AT2R mainly opposes AT1R, but the mechanism by which this occurs still remains obscure. In the present study, we investigated the role of AT2R in the treatment of rats with adjuvant-induced arthritis (AIA) by losartan. Adjuvant-induced arthritis rats were treated with losartan (5, 10 and 15 mg/kg) and methotrexate (MTX; 0.5 mg/kg) in vivo from day 14 to day 28. Arthritis was evaluated by the arthritis index and histological examination. Angiotensin II, tumour necrosis factor-α, and VEGF levels were examined by ELISA. The expression of AT1R and AT2R was detected by western blot and immunohistochemistry analysis. After stimulation with interleukin-1β in vitro, the effects of the AT2R agonist {"type":"entrez-protein","attrs":{"text":"CGP42112","term_id":"874777115","term_text":"CGP42112"}}CGP42112 (10⁻⁸–10⁻⁵ M) on the chemotaxis of monocytes induced by 10% foetal calf serum (FCS) were analysed by using Transwell assay. Subsequently, the therapeutic effects of {"type":"entrez-protein","attrs":{"text":"CGP42112","term_id":"874777115","term_text":"CGP42112"}}CGP42112 (5, 10 and 20 μg/kg) were evaluated in vivo by intra-articular injection in AIA rats. After treatment with losartan, the down-regulation of AT1R expression and up-regulation of AT2R expression in the spleen and synovium of AIA rats correlated positively with reduction in the polyarthritis index. Treatment with {"type":"entrez-protein","attrs":{"text":"CGP42112","term_id":"874777115","term_text":"CGP42112"}}CGP42112 inhibited the chemotaxis of AIA monocytes in vitro, possibly because of the up-regulation of AT2R expression. Intra-articular injection with {"type":"entrez-protein","attrs":{"text":"CGP42112","term_id":"874777115","term_text":"CGP42112"}}CGP42112 (10 and 20 μg/kg) ameliorated the arthritis index and histological signs of arthritis. In summary, the present study strongly suggests that the up-regulation of AT2R might be an additional mechanism by which losartan exerts its therapeutic effects in AIA rats.