Dysregulated Arginine Metabolism Research Articles

Dysregulated arginine metabolism in precursor B-cell acute lymphoblastic leukemia in children: a metabolomic study

BackgroundPrecursor B-cell acute lymphoblastic leukemia (B-ALL) is the most common cancers in children. Failure of induction chemotherapy is a major factor leading to relapse and death in children with B-ALL. Given the importance of altered metabolites in the carcinogenesis of pediatric B-ALL, studying the metabolic profile of children with B-ALL during induction chemotherapy and in different minimal residual disease (MRD) status may contribute to the management of pediatric B-ALL.MethodsWe collected paired peripheral blood plasma samples from children with B-ALL at pre- and post-induction chemotherapy and analyzed the metabolomic profiling of these samples by ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS). Healthy children were included as controls. We selected metabolites that were depleted in pediatric B-ALL and analyzed the concentrations in pediatric B-ALL samples. In vitro, we study the effects of the selected metabolites on the viability of ALL cell lines and the sensitivity to conventional chemotherapeutic agents in ALL cell lines.ResultsForty-four metabolites were identified with different levels between groups. KEGG pathway enrichment analyses revealed that dysregulated linoleic acid (LA) metabolism and arginine (Arg) biosynthesis were closely associated with pediatric B-ALL. We confirmed that LA and Arg were decreased in pediatric B-ALL samples. The treatment of LA and Arg inhibited the viability of NALM-6 and RS4;11 cells in a dose-dependent manner, respectively. Moreover, Arg increased the sensitivity of B-ALL cells to L-asparaginase and daunorubicin.ConclusionArginine increases the sensitivity of B-ALL cells to the conventional chemotherapeutic drugs L-asparaginase and daunorubicin. This may represent a promising therapeutic approach.

Dysregulated Arginine Metabolism Is Linked to Retinal Degeneration in Cep250 Knockout Mice.

Degeneration of retinal photoreceptors is frequently observed in diverse ciliopathy disorders, and photoreceptor cilium gates the molecular trafficking between the inner and the outer segment (OS). This study aims to generate a homozygous global Cep250 knockout (KO) mouse and study the resulting phenotype. We used Cep250 KO mice and untargeted metabolomics to uncover potential mechanisms underlying retinal degeneration. Long-term follow-up studies using optical coherence tomography (OCT) and electroretinography (ERG) were performed. OCT and ERG results demonstrated gradual thinning of the outer nuclear layer (ONL) and progressive attenuation of the scotopic ERG responses in Cep250-/- mice. More TUNEL signal was observed in the ONL of these mice. Immunostaining of selected OS proteins revealed mislocalization of these proteins in the ONL of Cep250-/- mice. Interestingly, untargeted metabolomics analysis revealed arginine-related metabolic pathways were altered and enriched in Cep250-/- mice. Mis-localization of a key protein in the arginine metabolism pathway, arginase 1 (ARG1), in the ONL of KO mice further supports this model. Moreover, adeno-associated virus (AAV)-based retinal knockdown of Arg1 led to similar architectural and functional alterations in wild-type retinas. Altogether, these results suggest that dysregulated arginine metabolism contributes to retinal degeneration in Cep250-/- mice. Our findings provide novel insights that increase understanding of retinal degeneration in ciliopathy disorders.

Dysregulated Arginine Metabolism in Young Patients with Chronic Persistent Asthma and in Human Bronchial Epithelial Cells.

Background: Recent metabolomics studies have found circulatory metabolism alterations in patients with asthma, indicating that altered metabolites played a significant role in asthma. However, the regulatory mechanisms in asthma, especially in young chronic persistent asthma remain underexplored. Methods: In this study, a prospective cohort of 162 patients diagnosed of asthma admitted to the First Affiliated Hospital of Xi’an Jiaotong University from January 2018 to December 2019 was used to perform a nested case-control study. Among them, we included 30 patients with chronic persistent asthma between 20 to 35 years old; 30 health control with evenly distributed age and sex were then recruited. Nontargeted metabolomics was applied to identify serum metabolic profiles and altered metabolic pathways. Results: In vitro, human bronchial epithelial cells (HBECs) line BEAS-2B with the addition of L-citrulline and/or asymmetric dimethylarginine (ADMA) model was utilized and the concentrations of nitric oxide (NO) metabolites were tested to evaluate the therapeutic potential of L-citrulline. The young patients with chronic persistent asthma displayed dysregulated serum metabolic profiles, especially enriched in arginine metabolism. The ratio of L-citrulline to ornithine is associated with blood eosinophil count. In vitro, adding L-citrulline could reverse ADMA-mediated reduction of NOx at lower L-arginine concentration (25 μM), but was ineffective in the higher L-arginine concentration (100 μM) media. Conclusions: The arginine metabolism balance is of vital importance during the pathogenesis and progression of chronic asthma. L-citrulline could be a powerful approach to restore airway NO production, potentially exhibiting therapeutic benefits among young patients with chronic asthma.

Myeloid Arginase 1 Insufficiency Exacerbates Amyloid-β Associated Neurodegenerative Pathways and Glial Signatures in a Mouse Model of Alzheimer's Disease: A Targeted Transcriptome Analysis.

Brain myeloid cells, include infiltrating macrophages and resident microglia, play an essential role in responding to and inducing neurodegenerative diseases, such as Alzheimer’s disease (AD). Genome-wide association studies (GWAS) implicate many AD casual and risk genes enriched in brain myeloid cells. Coordinated arginine metabolism through arginase 1 (Arg1) is critical for brain myeloid cells to perform biological functions, whereas dysregulated arginine metabolism disrupts them. Altered arginine metabolism is proposed as a new biomarker pathway for AD. We previously reported Arg1 deficiency in myeloid biased cells using lysozyme M (LysM) promoter-driven deletion worsened amyloidosis-related neuropathology and behavioral impairment. However, it remains unclear how Arg1 deficiency in these cells impacts the whole brain to promote amyloidosis. Herein, we aim to determine how Arg1 deficiency driven by LysM restriction during amyloidosis affects fundamental neurodegenerative pathways at the transcriptome level. By applying several bioinformatic tools and analyses, we found that amyloid-β (Aβ) stimulated transcriptomic signatures in autophagy-related pathways and myeloid cells’ inflammatory response. At the same time, myeloid Arg1 deficiency during amyloidosis promoted gene signatures of lipid metabolism, myelination, and migration of myeloid cells. Focusing on Aβ associated glial transcriptomic signatures, we found myeloid Arg1 deficiency up-regulated glial gene transcripts that positively correlated with Aβ plaque burden. We also observed that Aβ preferentially activated disease-associated microglial signatures to increase phagocytic response, whereas myeloid Arg1 deficiency selectively promoted homeostatic microglial signature that is non-phagocytic. These transcriptomic findings suggest a critical role for proper Arg1 function during normal and pathological challenges associated with amyloidosis. Furthermore, understanding pathways that govern Arg1 metabolism may provide new therapeutic opportunities to rebalance immune function and improve microglia/macrophage fitness.

Dysregulated Arginine Metabolism and Cardiopulmonary Dysfunction in Patients with Thalassemia

Pulmonary hypertension (PH) is common in thalassemia and contributes to mortality. Advancing age and a history of splenectomy are major risk factors in this population. The etiology of PH is multifactorial, involving a complex interaction of platelets, the coagulation system, erythrocytes, and endothelial cells along with inflammatory and vascular mediators. The long-term effect of splenectomy, red cell membrane pathology, coagulation abnormalities, low nitric oxide (NO) bioavailability, excess arginase activity, platelet activation, oxidative stress, iron overload, and chronic hemolysis play a role. The process of hemolysis disables the arginine-NO pathway through the simultaneous release of erythrocyte arginase and cell-free hemoglobin. Both NO and its obligate substrate arginine are rapidly consumed. The biological consequences of hemolysis on NO bioavailability ultimately translate into the clinical manifestations of PH. Guidelines for the management of PH in thalassemia have not yet been established; however, clinical trials are ongoing in an effort to guide future therapy.

Dysregulated Arginine Metabolism and Left Ventricular Hypertrophy in Pediatric Chronic Kidney Disease and End Stage Renal Disease

Dysregulated Arginine Metabolism and Left Ventricular Hypertrophy in Pediatric Chronic Kidney Disease and End Stage Renal Disease

Dysregulated Arginine Metabolism and Left Ventricular Hypertrophy in Pediatric Chronic Kidney Disease and End Stage Renal Disease

Background: Cardiovascular disease (CVD) is the leading cause of death in patients with Chronic Kidney Disease (CKD) and End Stage Renal Disease (ESRD). Left ventricular hypertrophy (LVH) is common in pediatric CKD patients and is an independent risk factor for cardiovascular (CV) morbidity/mortality. Nitric oxide (NO), an endothelium derived vasoactive substance critical for vascular homeostasis, is synthesized from arginine via NO synthase enzymes. Arginine is also a substrate for arginase enzymes therefore arginine bioavailability affects NO synthesis. The kidneys play a key role in de novo arginine synthesis as well as …

Dysregulated Urinary Arginine Metabolism in Older Adults With Amnestic Mild Cognitive Impairment.

Background: Urine samples, which capture an individual’s metabolic profile, are ideal for the exploration of non-invasive biomarkers to confirm the amnestic mild cognitive impairment (aMCI) status of patients vs. unimpaired ones.Objective: We aimed to detect differentially metabolized amino acids, which are important objectives in metabolomics, garnering particular attention in biomedical pathogenesis from the urine of aMCI patients, which may give clinicians the possibility to intervene with early treatments that curb Alzheimer’s disease (AD).Methods: The study included 208 subjects, 98 of whom were aMCI patients, and 110 who were control subjects without dementia. Urine samples were taken from each participant and supernatant was obtained for analysis. The concentrations of amino acids were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS).Results: Urinary arginine levels in aMCI patients are obviously lower than in normal controls (q < 0.2 and p < 0.05). Meanwhile, aMCI patients had significant reduced urinary global arginine bioavailability ratio (GABR), and GABR in urine displayed a positive correlation with the score of CMMSE.Conclusion: Urinary dysregulated arginine metabolism that may serve as a helpful clinical diagnostic biomarker for aMCI in older adults.

Partial Least Squares Discriminant Analysis and Bayesian Networks for Metabolomic Prediction of Childhood Asthma.

To explore novel methods for the analysis of metabolomics data, we compared the ability of Partial Least Squares Discriminant Analysis (PLS-DA) and Bayesian networks (BN) to build predictive plasma metabolite models of age three asthma status in 411 three year olds (n = 59 cases and 352 controls) from the Vitamin D Antenatal Asthma Reduction Trial (VDAART) study. The standard PLS-DA approach had impressive accuracy for the prediction of age three asthma with an Area Under the Curve Convex Hull (AUCCH) of 81%. However, a permutation test indicated the possibility of overfitting. In contrast, a predictive Bayesian network including 42 metabolites had a significantly higher AUCCH of 92.1% (p for difference < 0.001), with no evidence that this accuracy was due to overfitting. Both models provided biologically informative insights into asthma; in particular, a role for dysregulated arginine metabolism and several exogenous metabolites that deserve further investigation as potential causative agents. As the BN model outperformed the PLS-DA model in both accuracy and decreased risk of overfitting, it may therefore represent a viable alternative to typical analytical approaches for the investigation of metabolomics data.

Co-ingestion of whole eggs or egg whites with glucose protects against postprandial hyperglycaemia-induced oxidative stress and dysregulated arginine metabolism in association with improved vascular endothelial function in prediabetic men.

Replacing a portion of a glucose challenge with whole eggs (EGG) or egg whites (WHITE) was shown to protect against glucose-induced impairments in vascular function. We hypothesised in the present study that previously observed vasoprotection following co-ingestion of EGG or WHITE with glucose was attributed to limiting postprandial hyperglycaemia-induced oxidative stress that improves NO∙ bioavailability. Prediabetic men completed a randomised, cross-over study in which they ingested isoenergetic meals containing 100 g glucose (GLU), or 75 g glucose with 1·5 EGG, seven WHITE or two egg yolks (YOLK). At 30 min intervals for 3 h, we assessed plasma NO∙ metabolites, the lipid peroxidation biomarker malondialdehyde, antioxidants, arginine and its methylated metabolites (asymmetric dimethylarginine and symmetric dimethylarginine), tetrahydrobiopterin redox status, vasoconstrictors and inflammatory markers. Compared with GLU, malondialdehyde was lower and NO∙ metabolites were greater in EGG and WHITE, but YOLK was not different from GLU. Malondialdehyde was inversely correlated with NO∙ metabolites and vascular function, whereas NO∙ metabolites were positively correlated with vascular function. Compared with GLU, arginine was greater, but asymmetric and symmetric dimethylarginine and angiotensin-II were lower in all egg-based meals. Antioxidants, tetrahydrobiopterin redox status and inflammatory markers did not differ among treatments. Thus, while each egg-based meal improved arginine metabolism, only EGG and WHITE limited lipid peroxidation. This suggests that vasoprotection mediated by EGG and WHITE likely occurs in an NO∙-dependent manner by improving arginine metabolism and attenuating oxidative stress that otherwise limit NO∙ biosynthesis and bioavailability to the vascular endothelium.

Dysregulated arginine metabolism and cardiopulmonary dysfunction in patients with thalassaemia.

Pulmonary hypertension (PH) commonly develops in thalassaemia syndromes, but is poorly characterized. The goal of this study was to provide a comprehensive description of the cardiopulmonary and biological profile of patients with thalassaemia at risk for PH. A case-control study of thalassaemia patients at high versus low PH-risk was performed. A single cross-sectional measurement for variables reflecting cardiopulmonary status and biological pathophysiology were obtained, including Doppler-echocardiography, 6-min-walk-test, Borg Dyspnoea Score, New York Heart Association functional class, cardiac magnetic resonance imaging (MRI), chest-computerized tomography, pulmonary function testing and laboratory analyses targeting mechanisms of coagulation, inflammation, haemolysis, adhesion and the arginine-nitric oxide pathway. Twenty-seven thalassaemia patients were evaluated, 14 with an elevated tricuspid-regurgitant-jet-velocity (TRV)≥2·5m/s. Patients with increased TRV had a higher frequency of splenectomy, and significantly larger right atrial size, left atrial volume and left septal-wall thickness on echocardiography and/or MRI, with elevated biomarkers of abnormal coagulation, lactate dehydrogenase (LDH) levels and arginase concentration, and lower arginine-bioavailability compared to low-risk patients. Arginase concentration correlated significantly to several echocardiography/MRI parameters of cardiovascular function in addition to global-arginine-bioavailability and biomarkers of haemolytic rate, including LDH, haemoglobin and bilirubin. Thalassaemia patients with a TRV≥2·5m/s have additional echocardiography and cardiac-MRI parameters suggestive of right and left-sided cardiac dysfunction. In addition, low arginine bioavailability may contribute to cardiopulmonary dysfunction in β-thalassaemia.

Pulmonary Hypertension Associated With Advanced Systolic Heart Failure: Dysregulated Arginine Metabolism and Importance of Compensatory Dimethylarginine Dimethylaminohydrolase-1

This study sought to examine the hemodynamic determinants of dysregulated arginine metabolism in patients with acute decompensated heart failure and to explore possible mechanisms of arginine dysregulation in human heart failure.Accumulating methylated arginine metabolites and impaired arginine bioavailability have been associated with heart failure, but the underlying pathophysiology remains unclear.This study prospectively determined plasma levels of asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor, and global arginine bioavailability ratio [GABR = arginine/(ornithine + citrulline)] by tandem mass spectrometry in subjects with advanced decompensated heart failure in the intensive care unit (n = 68) and with stable chronic heart failure (n = 57).Compared with chronic heart failure subjects, plasma ADMA was significantly higher (median [interquartile range]: 1.29 [1.04 to 1.77] μmol/l vs. 0.87 [0.72 to 1.05] μmol/l, p < 0.0001), and global arginine bioavailability ratio significantly lower (median [interquartile range]: 0.90 [0.69 to 1.22] vs. 1.13 [0.92 to 1.37], p = 0.002) in advanced decompensated heart failure subjects. Elevated ADMA and diminished global arginine bioavailability ratio were associated with higher systolic pulmonary artery pressure (sPAP) and higher central venous pressure, but not with other clinical or hemodynamic indices. We further observed myocardial levels of dimethylarginine dimethylaminohydrolase-1 were increased in chronic heart failure without elevated sPAP (<50 mm Hg), but diminished with elevated sPAP (≥50 mm Hg, difference with sPAP <50 mm Hg, p = 0.02).Dysregulated arginine metabolism was observed in advanced decompensated heart failure, particularly with pulmonary hypertension and elevated intracardiac filling pressures. Compared with hearts of control subjects, we observed higher amounts of ADMA-degradation enzyme dimethylarginine dimethylaminohydrolase-1 (but similar amounts of ADMA-producing enzyme, protein methyltransferase-1) in the human failing myocardium.

Free Arginase-I in Human Pulmonary Microvascular Endothelial Cell Culture Leads to Endothelial Dysfunction and Increased Reactive Oxygen Species.

Abstract 3041Poster Board II-1017 IntroductionEndothelial dysfunction in chronic hemolytic diseases (e.g., sickle cell disease) is believed to lead to hemolysis-associated pulmonary hypertension. Dysregulated arginine metabolism, via free arginase-I released during hemolysis, is one mechanism proposed in prior studies. However, the effects of free arginase-I on endothelial cell function have not previously been reported. We hypothesized that free arginase-I would decrease hPMVEC nitric oxide (NO) production and therefore limit NO bioavailability. MethodsCultured human pulmonary microvascular endothelial cells (hPMVEC), at passage 6-8 and approximately 80% confluence, with increasing arginase-I concentrations added to the media (CN: 0 ng prot/mL, A30: 30 ng prot/mL, A90: 90 ng prot/mL) were studied. To maximize arginase-I activity, 0.1 mM MnCl2 (an important cofactor for arginase-I) and 3mM of L-arginine were added to each condition. The hPMVEC were incubated for 24h in normoxia (n=8) or hypoxia (1% O2; n=4), then media was harvested for urea/nitrite analyses and cell protein was harvested for Western blot analyses. ANOVA analyses were conducted to determine statistical significance. ResultsUrea production significantly increased with increasing media arginase-I in both normoxia (CN: 1.47±0.22, A30: 1.46±0.15, A90: 2.09±0.20 umol/mgPr; p<0.05) and hypoxia (CN: 0.913±0.22, A30: 1.01±0.04, A90: 1.99±0.14 umol/mgPr; p<0.005); interestingly, hypoxia means were significantly lower than those in normoxia (p<0.01). However, the percent change from controls in hypoxia (CN: 100±24, A30: 110±4, A90: 218±15) was significantly higher (p<0.05) than in normoxia conditions (CN: 100±15, A30: 99±10, A90: 142±13). Nitrite production showed no significant change with increasing media arginase-I in normoxia (CN: 17.6±2.4, A30: 20.8±4.6, A90: 18.8±2.1 nmol/mgPr; p=0.79); however, in hypoxia nitrites trended down with increasing media arginase-I (CN: 5.26±0.42, A30: 4.35±0.27, A90: 3.69±0.51 nmol/mgPr; p=0.068). In addition, means for nitrite production in hypoxia were significantly less than in normoxia conditions (p<0.0001). In normoxia, Western blot analyses showed significantly decreased 3-nitrotyrosine (3-NT) levels with increasing media arginase-I (CN: 1.00±0.03, A30: 0.88±0.07, A90: 0.77±0.07 [fold change]; p<0.05). 3-NT expression in hypoxia showed no differences with increasing media arginase-I (CN: 1.90±0.16, A30: 1.88±0.19, A90: 1.75±0.27 [fold change, normalized to normoxia control]; p=0.87); however, hypoxic means were significantly higher than in normoxia conditions (p<0.0001). In normoxia, NAD(P)H oxidase isoform 4 (Nox4) show a decreasing trend in protein levels with increasing media arginase-I (CN: 1.00±0.03, A30: 0.93±0.10, A90: 0.86±0.06 [fold change]), with an increase in Nox4 protein levels in hypoxia with increasing media arginase-I (CN: 1.18±0.02, A30: 1.33±0.08, A90: 1.47±0.01 [fold change, normalized to normoxia control]). The hypoxic levels of Nox4 were also higher than in normoxia conditions. ConclusionsAs expected, increasing free media arginase-I concentrations in hPMVEC culture resulted in significant increases in urea production in both normoxia and hypoxia. Hypoxia resulted in lower nitrite production by endothelial nitric oxide synthase (eNOS). However, increasing amounts of free arginase-I only affected nitrite production in hypoxia, suggesting that increased arginase-I serum concentrations may also become an important regulator of NO production in clinical cases of hypoxia (e.g., acute chest syndrome and vaso-occlusive crises in sickle cell patients). The concurrent increased expression of 3-NT in hypoxia, despite lower nitrites, implies an increase in reactive oxygen species (ROS). Our preliminary data for Nox4 expression, the predominant source of ROS in endothelial cells, indicates that ROS production via this pathway may increase in hypoxia with increasing free arginase-I. Further studies on how free arginase-I in the media affects endothelial function are warranted, particularly in regards to the mechanism by which free arginase-I leads to greater Nox4 expression and ROS production. We speculate that arginase-I inhibitors may represent a potential therapeutic target in patients with chronic hemolysis and acute hypoxic events. DisclosuresNo relevant conflicts of interest to declare.

Asthma management: Reinventing the wheel in sickle cell disease

Asthma is a common comorbidity in sickle cell disease (SCD) with a reported prevalence of 30-70%. The high frequency of asthma in this population cannot be attributed to genetic predisposition alone, and likely reflects in part, the contribution of overlapping mechanisms shared between these otherwise distinct disorders. There is accumulating evidence that dysregulated arginine metabolism and in particular, elevated arginase activity contributes to pulmonary complications in SCD. Derangements of arginine metabolism are also emerging as newly appreciated mechanism in both asthma and pulmonary hypertension independent of SCD. Patients with SCD may potentially be at risk for an asthma-like condition triggered or worsened by hemolysis-driven release of erythrocyte arginase and low nitric oxide bioavailability, in addition to classic familial asthma. Mechanisms that contributed to asthma are complex and multifactorial, influenced by genetic polymorphisms as well as environmental and infectious triggers. Given the association of asthma with inflammation, oxidative stress and hypoxemia, factors known to contribute to a vasculopathy in SCD, and the consequences of these factors on sickle erythrocytes, comorbid asthma would likely contribute to a vicious cycle of sickling and subsequent complications of SCD. Indeed a growing body of evidence documents what should come as no surprise: Asthma in SCD is associated with acute chest syndrome, stroke, pulmonary hypertension, and early mortality, and should therefore be aggressively managed based on established National Institutes of Health Guidelines for asthma management. Barriers to appropriate asthma management in SCD are discussed as well as strategies to overcome these obstacles in order to provide optimal care.

Pulmonary Hypertension in Thalassemia: Association with Hemolysis, Arginine Metabolism Dysregulation, and a Hypercoagulable State

Pulmonary Hypertension in Thalassemia: Association with Hemolysis, Arginine Metabolism Dysregulation, and a Hypercoagulable State

Dysregulated Arginine Metabolism, Hemolysis-Associated Pulmonary Hypertension, and Mortality in Sickle Cell Disease

Dysregulated Arginine Metabolism, Hemolysis-Associated Pulmonary Hypertension, and Mortality in Sickle Cell Disease

Dysregulated Arginine Metabolism and Elevated Arginase Activity in Thalassemia.

Pulmonary hypertension (PHT) is associated with high mortality rate and is a leading factor in heart failure and death in thalassemia (Thal) intermedia. However, data on PHT and Thal major is limited. While there are several risk factors for PHT in Thal major, ongoing chronic hemolysis, despite transfusion, supports the development of hemolysis-associated PHT. Recently, we reported in sickle cell disease a novel paradigm whereby hemolysis liberates vasoactive hemoglobin and erythrocyte arginase, resulting in impaired NO bioavailability, endothelial dysfunction, PHT and death. Since arginase activity is also elevated in Thal red cells, we hypothesize that elevated arginase contributes to dysregulated arginine (Arg) metabolism in Thal. Pilot data was analyzed from transfused Thal patients (8 Thal-major, 4 E-beta Thal, 2 Hb H alpha Thal). Echocardiogram were performed in steady state in 10 of the patients; 70% (7/10) demonstrated PHT (tricuspid regurgitant jet velocity ≥ 2.5 m/s). Plasma Arg trended lower in patients with Thal (19.5 to 122mM, median 50 mM) compared to controls. Ornithine (Orn) levels were higher, and Arg/Orn ratio was lower in Thal vs. contrls. Plasma arginase activity was significantly elevated (median 0.83 mmol/cc/hr, range 0.06 – 1.17). Proline, a downstream metabolite of arginase causing pulmonary vascular remodeling was also elevated. Exhaled nitric oxide (NO) levels are significantly elevated in Thal vs. controls (49±41 parts per billion vs.18±8 ppb, p=0.02), suggesting upregulation of NO synthase in the lungs in addition to higher plasma arginase activity. Recent findings indicate diverse origins of PHT share similar pathophysiologic abnormalities, particularly decreased Arg bioavailability and a shift of metabolism towards ornithine-dependent pathways. Hemolysis is most likely a trigger for these pathologic pathyways in hemoglobinopathies. Therapies that maximize Arg and NO bioavailability may benefit patients with thalassemia.VariableNL Control (n=36)Thalassemia (n=14)p*Arginine (μM)67±1857±26 (50)0.15Ornithine (μM)62±2285±680.05Arg/Orn ratio1.2±0.50.79±0.4<0.01Proline (μM)161±48258±116<0.001Citrulline (μM)25±1142±17<0.001Arginase (μmol/cc/hr)0.33±0.2 (n=45)0.71±0.3<0.001

Dysregulated Arginine Metabolism, Hemolysis-Associated Pulmonary Hypertension, and Mortality in Sickle Cell Disease

Sickle cell disease is characterized by a state of nitric oxide resistance and limited bioavailability of l-arginine, the substrate for nitric oxide synthesis. We hypothesized that increased arginase activity and dysregulated arginine metabolism contribute to endothelial dysfunction, pulmonary hypertension, and patient outcomes. To explore the role of arginase in sickle cell disease pathogenesis, pulmonary hypertension, and mortality. Plasma amino acid levels, plasma and erythrocyte arginase activities, and pulmonary hypertension status as measured by Doppler echocardiogram were prospectively obtained in outpatients with sickle cell disease. Patients were followed up for survival up to 49 months. Urban tertiary care center and community clinics in the United States between February 2001 and March 2005. Two hundred twenty-eight patients with sickle cell disease, aged 18 to 74 years, and 36 control participants. Plasma amino acid levels, plasma and erythrocyte arginase activities, diagnosis of pulmonary hypertension, and mortality. Plasma arginase activity was significantly elevated in patients with sickle cell disease, with highest activity found in patients with secondary pulmonary hypertension. Arginase activity correlated with the arginine-ornithine ratio, and lower ratios were associated with greater severity of pulmonary hypertension and with mortality in this population (risk ratio, 2.5; 95% confidence interval [CI], 1.2-5.2; P = .006). Global arginine bioavailability, characterized by the ratio of arginine to ornithine plus citrulline, was also strongly associated with mortality (risk ratio, 3.6; 95% CI, 1.5-8.3; P<.001). Increased plasma arginase activity was correlated with increased intravascular hemolytic rate and, to a lesser extent, with markers of inflammation and soluble adhesion molecule levels. These data support a novel mechanism of disease in which hemolysis contributes to reduced nitric oxide bioavailability and endothelial dysfunction via release of erythrocyte arginase, which limits arginine bioavailability, and release of erythrocyte hemoglobin, which scavenges nitric oxide. The ratios of arginine to ornithine and arginine to ornithine plus citrulline are independently associated with pulmonary hypertension and increased mortality in patients with sickle cell disease.