Effects Of Arginase Inhibition Research Articles

Effects of arginase inhibition on myocardial Ca2+ and contractile responses.

Nitric oxide (NO) is thought to increase cardiac contractility by increasing cytosolic Ca2+ concentration ([Ca2+]cyt) during excitation. Alternatively, NO could increase the sensitivity of the contractile response to [Ca2+]cyt (Ca2+ sensitivity). Arginase regulates NO production by competing with NO synthase (NOS), and thus, arginase inhibition should increase cardiac contractility by increasing NO production. We hypothesized that arginase inhibition increases cardiac contractility by increasing both [Ca2+]cyt and Ca2+ sensitivity. [Ca2+]cyt and contractile (sarcomere length [SL] shortening) responses to electrical stimulation were measured simultaneously in isolated rat cardiomyocytes using an IonOptix system. In the same cardiomyocytes, measurements were obtained at baseline, following 3‐min exposure to an arginase inhibitor (S‐[2‐boronoethyl]‐l‐cysteine; BEC) and following 3‐min exposure to BEC plus a NOS inhibitor (N G‐nitro‐l‐arginine‐methyl ester; l‐NAME). These responses were compared to time‐matched control cardiomyocytes that were untreated. Compared to baseline, BEC increased the amplitude and the total amount of evoked [Ca2+]cyt, and the extent and velocity of SL shortening in cardiomyocytes, whereas addition of l‐NAME mitigated these effects. The [Ca2+]cyt at 50% contraction and relaxation were not different across treatment groups indicating no effect of BEC on Ca2+ sensitivity. The [Ca2+]cyt and SL shortening responses in time‐matched controls did not vary with time. Arginase inhibition by BEC significantly increased the amplitude and the total amount of evoked [Ca2+]cyt, and the extent and velocity of SL shortening in cardiomyocytes, but did not affect Ca2+ sensitivity. These effects of BEC were mitigated by l‐NAME. Together, these results indicate an effect of NO on [Ca2+]cyt responses that then increase the contractile response of cardiomyocytes.

Effects of arginase inhibition via jugular infusion of Nω-hydroxy-nor-l-arginine on metabolic and immune indices in lactating dairy cows

Transformation of Arg to nitric oxide and ornithine (Arg-Orn) constitutes the main route of Arg metabolism in mammals. The primary objective of this work was to determine the effects of inhibiting the Arg-Orn pathway via Nω-hydroxy-nor-l-arginine (nor-NOHA) on health of lactating cows. Furthermore, we also explored the effect of Arg-Orn inhibition on the efficiency of nitrogen utilization to find support for previous research that showed the inhibition of Arg-Orn inhibited milk protein synthesis. Six healthy Chinese Holstein cows of similar body weight (550.0 ± 20 kg), parity (4.0 ± 0), body condition score (3.0 ± 0), milk yield (21.0 ± 1.0 kg), and days in milk (80 ± 2 d) were selected and randomly assigned to 3 treatments in a replicated 3 × 3 Latin square design with 22 d for each period (7 d for infusion and 15 d for washout). The treatments were (1) saline infusion (control); (2) infusion of 125 mg/L of nor-NOHA; and (3) infusion of 125 mg/L of nor-NOHA with 9.42 g/L of Arg. Dry matter intake, apparent digestibility of nutrients, urinary N, N in milk, and blood indices of metabolism and immune function were determined. Compared with the control, the infusion of nor-NOHA had no effect on the concentrations of cholesterol, high-density lipoproteins, IgA, IL-1β, tumor necrosis factor-α, and alanine transaminase. In addition, the dry matter intake, apparent digestibility of N, and the concentration of milk protein N in the Nor-NOHA did not differ from the control; however, the infusion of nor-NOHA and Arg resulted in greater concentrations of high-density lipoprotein, IgA, IL-1β, and tumor necrosis factor-α, and lower concentrations of cholesterol in serum compared with the control. Moreover, the addition of Arg to cows infused with nor-NOHA increased the concentration of nitrate (the indicator of nitric oxide) in serum and was associated with greater milk protein N production due to greater milk yield compared with those infused with nor-NOHA. Overall, the results indicated important roles of Arg in immunity and mammary N utilization, whereas a minor role of the Arg-Orn pathway in these physiologic processes was found.

Arginase inhibition: a new treatment for preventing progression of established diabetic nephropathy

Our previous publication showed that inhibition of arginase prevents the development of diabetic nephropathy (DN). However, identification of targets that retard the progression of established DN-which is more clinically relevant-is lacking. Therefore, we tested the hypothesis that arginase inhibition would prevent the progression of established DN. Effects of arginase inhibition were compared with treatment with the angiotensin-converting enzyme inhibitor captopril, a current standard of care in DN. Experiments were conducted in Ins2(Akita) mice treated with the arginase inhibitor S-(2-boronoethyl)-l-cysteine (BEC) or captopril starting at 6 wk of age for 12 wk (early treatment) or starting at 12 wk of age for 6 wk (late treatment). Early and late treatment with BEC resulted in protection from DN as indicated by reduced albuminuria, histological changes, kidney macrophage infiltration, urinary thiobarbituric acid-reactive substances, and restored nephrin expression, kidney nitrate/nitrite, kidney endothelial nitric oxide synthase phosphorylation, and renal medullary blood flow compared with vehicle-treated Ins2(Akita) mice at 18 wk of age. Interestingly, early treatment with captopril reduced albuminuria, histological changes, and kidney macrophage infiltration without affecting the other parameters, but late treatment with captopril was ineffective. These findings highlight the importance of arginase inhibition as a new potential therapeutic intervention in both early and late stages of diabetic renal injury.

Effects of Arginase Inhibition in Hypertensive Hyperthyroid Rats

This study analyzed the effects of chronic administration of N[omega]-hydroxy-nor-l-arginine (nor-NOHA), an inhibitor of arginase, on the hemodynamic, oxidative stress, morphologic, metabolic, and renal manifestations of hyperthyroidism in rats. Four groups of male Wistar rats were used: control, nor-NOHA-treated (10 mg/kg/day), thyroxine (T4)-treated (75 μg/rat/day), and thyroxine- plus nor-NOHA-treated rats. All treatments were maintained for 4 weeks. Body weight, tail systolic blood pressure (SBP), and heart rate (HR) were recorded weekly. Finally, morphologic, metabolic, plasma, and renal variables were measured. Arginase I and II protein abundance and arginase activity were measured in aorta, heart, and kidney. The T4 group showed increased arginase I and II protein abundance, arginase activity, SBP, HR, plasma nitrates/nitrites (NOx), brainstem and urinary isoprostanes, proteinuria and cardiac and renal hypertrophy in comparison to control rats. In hyperthyroid rats, chronic nor-NOHA prevented the increase in SBP and HR and decreased proteinuria in association with an increase in plasma NOx and a decrease in brainstem and urinary isoprostanes. In normal rats, nor-NOHA treatment did not significantly change any hemodynamic, morphologic, or renal variables. Acute nor-NOHA administration did not affect renal or systemic hemodynamic variables in normal or T4-treated rats. Hyperthyroidism in rats is associated with the increased expression and activity of arginase in aorta, heart, and kidney. Chronic arginase inhibition with nor-NOHA suppresses the characteristic hemodynamic manifestations of hyperthyroidism in association with a reduced oxidative stress. These results indicate an important role for arginase pathway alterations in the cardiovascular and renal abnormalities of hyperthyroidism.

Correction: Effect of Arginase Inhibition on Pulmonary L-Arginine Metabolism in Murine Pseudomonas Pneumonia

Correction: Effect of Arginase Inhibition on Pulmonary L-Arginine Metabolism in Murine Pseudomonas Pneumonia

Effect of Arginase Inhibition on Pulmonary L-Arginine Metabolism in Murine Pseudomonas Pneumonia

RationaleInfection of the lung with Pseudomonas aeruginosa results in upregulation of nitric oxide synthases (NOS) and arginase expression, and both enzymes compete for L-arginine as substrate. Nitric oxide (NO) production may be regulated by arginase as it controls L-arginine availability for NOS. We here studied the effect of systemic arginase inhibition on pulmonary L-arginine metabolism in Pseudomonas pneumonia in the mouse.MethodsMice (C57BL/6, 8–10 weeks old, female) underwent direct tracheal instillation of Pseudomonas (PAO-1)-coated agar beads and were treated by repeated intra-peritoneal injections of the arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH) or PBS until lungs were harvested on day 3 of the infection. L-arginine metabolites were quantified using liquid chromatography-tandem mass spectrometry, NO metabolites nitrate and nitrite by Griess reagent and cytokines by ELISA.ResultsNO metabolite concentrations (48.5±2.9 vs. 10.9±2.3 µM, p<0.0001), as well as L-ornithine (29.6±1.7 vs 2.3±0.4 µM, p<0.0001), the product of arginase activity, were increased in Pseudomonas infected lungs compared to naïve controls. Concentrations of the NOS inhibitor asymmetric dimethylarginine (ADMA) were also increased (0.44±0.02 vs. 0.16±0.01 µM, p<0.0001). Arginase inhibition in the infected animals resulted in a significant decrease in L-ornithine (14.6±1.6 µM, p<0.0001) but increase in L-arginine concentration (p<0.001), L-arginine/ADMA ratio (p<0.001), L-arginine availability for NOS (p<0.001), and NO metabolite concentrations (67.3±5.7 µM, p<0.05). Arginase inhibitor treatment also resulted in an increase in NO metabolite levels in animals following intratracheal injection of LPS (p = 0.015). Arginase inhibition was not associated with an increase in inflammatory markers (IFN-γ, IL-1β, IL-6, MIP-2, KC or TNF-α) in lung. Concentrations of the L-ornithine-dependent polyamines putrescine, spermidine and spermine were increased in Pseudomonas infected lungs (p<0.001, respectively) but were unaffected by ABH treatment.ConclusionsSystemic arginase inhibition with ABH during Pseudomonas pneumonia in mice results in an increase in pulmonary NO formation but no pro-inflammatory effect.

51 Effect of arginase inhibition on NO formation in cystic fibrosis sputum

51 Effect of arginase inhibition on NO formation in cystic fibrosis sputum

52 Evaluation of marker proteins for quantitation of CFTR

Objectives: P. aeruginosa, a common pathogen in airways of patients with CF is sensitive to NOand nitrite-mediated killing. L-arginine is a substrate for nitric oxide (NO) synthase and arginase and both substitution of L-arginine and/or inhibition of arginase could have positive effects on Pseudomonas infection. We investigated the effect of L-arginine and of arginase inhibitor ABH on NOmetabolite formation in a novel CF sputum assay. Methods: Expectorated sputum was homogenized with the use of sputulysin and DPBS following standardized protocols before incubation with different concentrations of L-arginine or ABH. Nitrate and nitrite were measured using Griess reagent after 1 hour of incubation time at 37oC. Results: Total NO-metabolites (NOx) and nitrite concentrations were unaffected by incubation with buffer alone. L-arginine (10, 30, 100 and 300mmol) resulted in a concentration-dependent increase in both total NOx (1.7, 2.9, 3.9 and 3.9fold, p< 0.01, ANOVA) and nitrite (2.0, 3.4, 4.9 and 5.4-fold, p< 0.01). ABH (10, 30, and 100mmol) also resulted in a significant increase in NOx and nitrite (1.5fold, p< 0.01). The combination of ABH 1mmol with increasing concentrations of L-arginine (0, 0.33, 1, 3, and 10mmol) (n = 21) resulted in additive effects with significant increase in sputum nitrite observed for L-arginine of 3mmol and higher (p< 0.001) compared to ABH 1 mmol, respectively. Conclusions: Arginase inhibitor added to CF sputum alone or in combination with L-arginine significantly increases NO production. Studies to assess the effects of arginase inhibition on NO-mediated killing of bacteria in CF sputum are currently under way. 52 Evaluation of marker proteins for quantitation of CFTR

Arginase Modulates NF-κB Activity via a Nitric Oxide–Dependent Mechanism

NF-kappaB is a versatile transcription factor that regulates a wide array of processes, including inflammation and survival, and plays a critical role in the etiology of inflammatory lung diseases. Nitric oxide (NO) has been suggested to play an antiinflammatory role through S-nitrosation of components of NF-kappaB pathway. NO production can be modulated by changing the availability of its substrate, L-arginine. Arginases compete with NO synthases (NOSs) for their common substrate, L-arginine, and thereby have the potential to alter the signaling function of NO. The goal of the present study was to determine the impact of arginase manipulation on NO, and subsequent effects on NF-kappaB activation, in lung epithelial cells. Our results demonstrate that reduction of arginase activity enhanced cellular content of NO and S-nitrosated proteins, and resulted in decreases in TNF-alpha- or LPS-stimulated NF-kappaB DNA binding and transcriptional activity, in association with enhanced S-nitrosation of p50. The effects of arginase inhibition on NF-kappaB were reversed by the generic NOS inhibitor, N-omega-nitro-L-arginine methyl ester (L-NAME), suggesting a causal role for NO in the attenuation of NF-kappaB induced by arginase suppression. Conversely, overexpression of arginase I decreased cellular S-nitrosothiol content and enhanced IkappaB kinase activity and NF-kappaB DNA binding, and decreased S-nitrosation of p50. Collectively, our data point to a regulatory mechanism wherein NF-kappaB is controlled through arginase-dependent regulation of NO levels, which may impact on chronic inflammatory diseases that are accompanied by NF-kappaB activation and upregulation of arginases.

Beneficial effects of arginase inhibition in spontaneously hypertensive rats

Beneficial effects of arginase inhibition in spontaneously hypertensive rats