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We hypothesized that hypoxia would activate epidermal growth factor receptor (EGFR) tyrosine kinase, leading to increased arginase expression and resulting in proliferation of human pulmonary microvascular endothelial cell (hPMVEC). To test this hypothesis, hPMVEC were incubated in normoxia (20% 02, 5% C[O.sub.2]) or hypoxia (1% [O.sub.2], 5% C[O.sub.2]). Immunoblotting for EGFR and proliferating cell nuclear antigen was done, and protein levels of both total EGFR and proliferating cell nuclear antigen were greater in hypoxic hPMVEC than in normoxic hPMVEC. Furthermore, hypoxic hPMVEC had greater levels of EGFR activity than did normoxic hPMVEC. Hypoxic hPMVEC had a twofold greater level of proliferation compared with normoxic controls, and this increase in proliferation was prevented by the addition of AG-1478 (a pharmacological inhibitor of EGFR). Immunoblotting for arginase I and arginase II demonstrated a threefold induction in arginase II protein levels in hypoxia, with little change in arginase I protein levels. The hypoxic induction of arginase II protein was prevented by treatment with AG-1478. Proliferation assays were performed in the presence of arginase inhibitors, and hypoxia-induced proliferation was also prevented by arginase inhibition. Finally, treatment with an EGFR small interfering RNA prevented hypoxia-induced profiferation and urea production. These findings demonstrate that hypoxia activates EGFR tyrosine kinase, leading to arginase expression and thereby promoting proliferation in hPMVEC. vascular remodeling; L-arginine; pulmonary hypertension doi: 10.1152/ajplung.00122.2009.

Chen B, Calvert AE, Cui H, Nelin LD. Hypoxia promotes human pulmonary artery smooth muscle cell proliferation through induction of arginase. Am J Physiol Lung Cell Mol Physiol 297 : L1151-L1159, 2009. First published October 2, 2009; doi: 10.1152/ajplung.00183.2009.--Vascular remodeling and smooth muscle cell proliferation are hallmark pathogenic features of pulmonary artery hypertension (PAH). Alterations in the metabolism of L-arginine via arginase and nitric oxide synthase play a critical role in the endothelial dysfunction seen in PAH. L-arginine metabolism by arginase produces L-ornithine and urea. L-ornithine is a precursor for polyamine and proline synthesis, ultimately leading to an increase in cellular proliferation. Given the integral role of the smooth muscle layer in the pathogenesis of hypoxia-induced PAH, we hypothesized that hypoxia would increase cellular proliferation via arginase induction in human pulmonary artery smooth muscle cells (hPASMC). We found that arginase II mRNA and protein expression were significantly increased in cultured hPASMC exposed to 1% [O.sub.2] for 24 and 48 h, which coincided with an increase in arginase activity at 48 h. There were no hypoxia-induced changes in levels of arginase I mRNA or protein in cultured hPASMC. Exposure to hypoxia resulted in more than one and a half times as many viable ceils after 120 h than normoxic exposure. The addition of the arginase inhibitor, S-(2-boronoethyl)-L-cysteine, completely prevented both the hypoxia-induced increase in arginase activity and proliferation in hPASMC. Furthermore, transfection of small interfering RNA (siRNA) targeting arginase II in hPASMC resulted in knockdown of arginase II protein levels and complete prevention of the hypoxia-induced cellular proliferation. These data support our hypothesis that hypoxia increases proliferation of hPASMC through the induction of arginase II. pulmonary hypertension; L-arginine; vascular remodeling doi: 10.1152/ajplung.00183.2009.

Liver dysfunction secondary to severe inflammation is associated with the release of enzymes normally sequestered within hepatocytes. The purpose of these studies was to test the hypothesis that these enzymes are released, at least in part, to modulate potentially deleterious inflammatory processes in distant tissues like the gut. Human [Caco-2.sub.BBe] enterocyte-like cells were exposed to cytomix (IFN-[gamma] TNF-[alpha], and IL-1[beta]) in the absence or presence of human liver cytosol (LC). Nitric oxide ([NO.sup..]) and inducible nitric oxide synthase (iNOS) protein production were measured by the Griess assay and Western analysis, respectively. Cytomix induced the expression of iNOS and release of [NO.sup..]. LC protein (400 [micro]g/ml) added to the basal compartment but not apical compartment completely blocked the release of [NO.sup..] but only slightly decreased the magnitude of iNOS protein induction. Ultrafiltration and ultracentrifugation studies demonstrated that microsome-associated arginase-1 activity was the iNOS-suppressing activity in LC. Liver arginase required activation by a < 10-kDa factor that was present in supernatants of cytomix-stimulated cells. The selective iNOS inhibitor L-[N.sup.6]-(1-iminoethyl)-lysine x 2HCl prevented production of this factor. The biotin switch assay detected increased S-nitrosylation of arginase-1 after incubation with supernatants from immunostimulated Caco-2 ceils. Serum from endotoxemic mice contained significantly greater arginase activity compared with serum from control mice. Furthermore, the ratio of mucosal monomeric to dimeric iNOS increased in endotoxemic mice compared with controls. Thus reciprocal activation of arginase-1 and modulation of mucosal iNOS activity may be protective because it would be expected to decrease [NO.sup..]-dependent intestinal barrier dysfunction on that basis. shock; systemic inflammatory response syndrome; liver damage doi: 10.1152/ajpgi.90716.2008

Asthma is a dynamic disorder of airway inflammation and airway remodeling with an imbalance in T helper type 1 ([Th.sub.l])/[Th.sub.2] immune response. Increased [Th.sub.2] cytokines such as IL-4 and IL-13 induce arginase either directly or indirectly through transforming growth factor-[[beta].sub.1] (TGF-[[beta].sub.1]) and lead to subepithelial fibrosis, which is a crucial component of airway remodeling. Synthetic antimalarials have been reported to have immunomodulatory properties. Mepacrine is known for its reduction of airway inflammation in short-term allergen challenge model by reducing The cytokines and cysteinyl leukotrienes, which has an important role in the development of airway remodeling features. Therefore, we hypothesized that mepacrine may reduce airway remodeling. For this, extended subacute ovalbumin mice model of asthma was developed; these mice showed an increased expression of profibrotic mediators, subepithelial fibrosis, and goblet cell metaplasia along with airway inflammation, increased [Th.sub.2] cytokines, allergen-specific IgE, IgG1, increased cytosolic PLA2 (cPLA2), and airway hyperresponsiveness. Presence of intraepithelial eosinophils and significant TGF-[[beta].sub.1] expression in subepithelial mesenchymal regions by repeated allergen exposures indicate that asthmatic mice of this study have developed human mimicking as well as late stages of asthma. However, mepacrine treatment decreased [Th.sub.2] cytokines and subepithelial fibrosis and alleviated asthma features. These reductions by mepacrine were associated with a decrease in levels and expression of TGF-[[beta].sub.1] and the reduction in activity, expression of arginase in lung cytosol, and immunolocalization in inflammatory cells present in perivascular and peribronchial regions. These results suggest that mepacrine might reduce the development of subepithelial fibrosis by reducing the arginase and TGF-[[beta].sub.1]. These effects of mepacrine likely underlie its antiairway remodeling action in asthma. transforming growth factor-[[beta].sub.1]; lung

Arginases compete with nitric oxide (NO) synthases for L-arginine as common substrate. Pulmonary vascular and airway diseases in which arginase activity is increased are associated with decreased NO production and reduced smooth muscle relaxation. The developmental patterns of arginase activity and type I and II isoforms expression in the lung have not been previously evaluated. Hypothesizing that lung arginase activity is developmentally regulated and highest in the fetus, we measured the expression of both arginase isoforms and total arginase activity in fetal, newborn, and adult rat lung, pulmonary artery, and bronchial tissue. In addition, intrapulmonary arterial muscle force generation was evaluated in the absence and presence of the arginase inhibitor [N.sup.[omega]]-hydroxy-nor-L-arginine (nor-NOHA). Arginase II content, as well as total arginase activity, was highest in fetal rat lung, bronchi, and pulmonary arterial tissue and decreased with age (P < 0.05), and its lung cell expression was developmentally regulated. In the presence of nor-NOHA, pulmonary arterial force generation was significantly reduced in fetus and newborn (P < 0.01). No significant change in force generation was noted in bronchial tissue following arginase inhibition. In conclusion, arginase II is regulated developmentally, and both expression and activity are maximal during fetal life. We speculate that the maintenance of a high pulmonary vascular resistance and decreased lung NO production prenatally may. in part, be dependent on increased arginase expression and/or activity. pulmonary vascular resistance; airway resistance; nitric oxide

Vascular tissues express arginase that metabolizes L-arginine to L-ornithine and urea and thus reduces substrate availability for nitric oxide formation. Dahl salt-sensitive (Dahl-S) rats with salt-induced hypertension show endothelial dysfunction, including decreased vascular nitric oxide formation. This study tests the hypothesis that increased vascular arginase activity contributes to endothelial dysfunction in hypertensive Dahl-S rats. Male Dahl-S rats (5-6 wk) were placed on high (8%) or low (0.3%) NaCl diets for 4 wk. With respect to the low-salt group, mean arterial blood pressure was increased in the high-salt animals. Immunohistochemical stainings for arginase I and II were enhanced in arterioles isolated from high-salt Dahl-S rats. Experiments used isolated Krebs buffer-superfused first-order gracilis muscle arterioles with constant pressure (80 mmHg) and no luminal flow or constant midpoint but altered endpoint pressures to establish graded levels of luminal flow (0-50 [micro]l/min). In high-salt arterioles, responses to an endothelium-dependent vasodilator acetylcholine (1 nmol/l to 3 [micro]mol/l) and flow-induced dilation were decreased. Acute in vitro treatment with an inhibitor of arginase, 100 [micro]mol/l (S)-(2-boronoethyl)-L- cystine, or the nitric oxide precursor, 1 mmol/l L-arginine, similarly enhanced acetylcholine and flow-induced maximal dilations and abolished the differences between high- and low-salt arterioles. These data show that arteriolar arginase expression is increased and that endothelium-dependent vasodilation is decreased in high-salt Dahl-S rats. Acute pretreatment with an arginase inhibitor or with L-arginine restores endothelium-dependent vasodilation and abolishes the differences between high- and low-salt groups. These results suggest that enhanced vascular arginase activity contributes to endothelial dysfunction in Dahl-S rats with salt-induced hypertension and identifies arginase as a potential therapeutic target to prevent endothelial dysfunction. salt-sensitive hypertension; vascular tone; arterioles

The arginases catalyze the divalent cation dependent hydrolysis of L-arginine to produce L-ornithine and urea. Although traditionally considered in terms of its role as the final enzyme of the urea cycle, the enzyme is found in a variety of nonhepatic tissues. These findings suggest that the enzyme may have other functions in addition to its role in nitrogen metabolism. High-resolution crystal structures have been determined for recombinant rat liver (type I) arginase and for recombinant human kidney (type II) arginase, their variants, and complexes with products and inhibitors. Each identical subunit of the trimeric enzyme contains an active site that lies at the bottom of a 15 [Angstrom] deep cleft. The 2 essential Mn(II) ions are located at the bottom of this cleft, separated by ~3.3 [Angstrom] and bridged by oxygens derived from 2 aspartic acid residues and a solvent-derived hydroxide. This metal bridging hydroxide is proposed to be the nucleophile that attacks the guanidinium carbon of substrate arginine. On the basis of this proposed mechanism, boronic acid inhibitors of the enzyme have been synthesized and characterized kinetically and structurally. These inhibitors display slow-onset inhibition at the pH optimum of the enzyme, and are found as tetrahedral species at the active site, as determined by X-ray diffraction. The potent inhibition of arginases I and II by these compounds has not only delineated key enzyme-substrate interactions, but has also led to a greater understanding of the role of arginase in nonhepatic tissues. J. Nutr. 134: 2760S-2764S, 2004. KEY WORDS: * arginase * manganese * protein structure * enzyme inhibitors * boronic acids

The NO-cGMP pathway plays a key role in the male and female genital sexual arousal response. Nitric oxide synthase (NQS) utilizes L-arginine and oxygen as substrates to produce nitric oxide (NO) and citrulline. Arginase is a metalloenzyme that catalyzes the hydrolysis of L-arginine to produce L-ornithine and urea. It is proposed that arginase competes for L-arginine and reduces NOS activity in genital tissues, thus modulating sexual function. Using 2 transition state analogue inhibitors of arginase, 2(S)-Amino-6-boronohexanoic acid (ABH) and S-(2-boronoethyl)-L-cysteine (BEC), we have characterized arginase activity in penile and vaginal tissue. Neither of these inhibitors has activity against NOS. Thus, ABH and BEC are useful compounds for examining the role of arginase in genital tissue physiology, without directly influencing NOS activity. We present data to suggest that arginase may regulate NO production by competing for endogenous pools of L-arginine. In this fashion, arginase is an indirect regulator of penile and vaginal blood flow and specific arginase inhibitors may improve genital blood flow during sexual arousal. As evidenced by the upregulation of arginase in specific disease states, its distribution in the vagina, and its modulation by sex steroid hormones, this enzyme may also participate in numerous other physiological and pathophysiological processes, such as tissue growth, fibrosis, and immune function. KEY WORDS: * arginase * arginine * nitric oxide synthase * erectile function * genital hemodynamics

In this review the current knowledge about the arginine-degrading enzyme arginase and its unexpected roles in survival and regeneration in the central nervous system will be discussed. Recent data suggest the neuroprotective effects of extracellularly applied arginase can be attributed to an activation of the endoplasmic reticulum stress response with a consequent change of the pro-survival gene expression profile. However, the activation of neural regeneration pathways caused by an upregulation of endogenous arginase I is mediated by polyamines, a group of arginase downstream products with widespread biological effects. In light of these new discoveries, there is heightened interest in the regulation of arginase I gene expression within the central nervous system. A number of transcription factors such as Sp1, C/EBP (CCATT/enhancer-binding protein), and CREB seem to be involved in the transcriptional control of arginase I and may contribute to the complex expression pattern of arginase I in distinct brain regions and during development. Beyond molecular mechanisms, this review will also include relevant clinical findings in patients with neurodegenerative diseases. J. Nutr. 134: 2812S-2817S, 2004. KEY WORDS: * arginase * arginine * neurodegeneration

For many years it has been supposed that the production of an excess of nitric oxide (NO) by inducible NO synthase (iNOS) plays a major role in inflammatory diseases, including asthma. However, recent studies indicate that a deficiency of beneficial, bronchodilating constitutive NOS (cNOS)-derived NO is important in allergen-induced airway hyperresponsiveness. Although several mechanisms are proposed to explain the reduction of cNOS activity, reduced substrate availability, caused by a combination of increased arginase activity and decreased cellular uptake of L-arginine, appears to play a key role. Recent evidence also indicates that iNOS-induced pathophysiological effects involve substrate deficiency. Thus, at low concentrations of L-arginine iNOS produces both NO and superoxide anions, which results in the increased synthesis of the highly reactive, detrimental oxidant peroxynitrite. Based on these observations, we propose that a relative deficiency of NO caused by increased arginase activity and altered L-arginine homeostasis is a major factor in the pathology of asthma.

Activities of arginase I and II are limiting for endothelial cell proliferation. Am J Physiol Regulatory Integrative Comp Physiol 282: R64-R69, 2002.--Polyamines are essential for cell proliferation; therefore, we hypothesized that arginase I or arginase II activities, via production of ornithine for polyamine synthesis, may be limiting for proliferation of endothelial cells (EC). Bovine coronary venular EC stably transfected with a lacZ gene (lacZ-EC, control), rat arginase I cDNA (AI-EC), or mouse arginase II cDNA (AII-EC) were utilized to test this hypothesis. Cell-proliferation assays showed that EC proliferation was markedly increased in AI-EC and AII-EC compared with lacZ-EC. Expression of proliferating cell nuclear antigen was also enhanced in AI-EC and AII-EC. DL-[alpha]-difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase, was used to establish that increased polyamine synthesis was involved in mediating the enhanced growth of AI-EC and AII-EC. Addition of 5 mM DFMO to the culture medium completely abolished the differences in cellular putrescine concentrations and reduced the differences in spermidine concentrations among AI-EC, AII-EC, and lacZ-EC. The DFMO treatment also prevented an increase in AI-EC and AII-EC proliferation compared with lacZ-EC. Addition of 10 and 50 [micro]M putrescine dose-dependently increased AI-EC, AII-EC, and lacZ-EC growth to the same extent. These results demonstrate that either arginase isoform can potentially play a role in modulating EC proliferation by regulating polyamine synthesis. ornithine; polyamines; cell transfection

Research has been conducted on the mitochondrial human type II arginase. The inhibition of this arginase by N (super)omega hydroxy-L-arginine and its analogue N (super)omega hydroxy-nor-L-arginine has been investigated and the results are discussed.

Research has been conducted on the arginase, a binuclear Mn (super)2+ metalloinsyme. The x-ray crystal structutes of arginase have been determined.

Research has been conducted on the boronic acid-based arginine analogue S-(2-boronoethyl)-L-cysteine. The synthesis of this analogue has been carried out demonstrating that it is a slow-binding competitive inhibitor of the binuclear manganese metalloenzyme arginase.

The early induction of inducible nitric oxide synthase (iNOS) and arginase II in the lung and spleen of mice treated with bacterial lipopolysaccharides has been studied. RNA blot and immunoblot analyses as well as enzyme assay were used in examining the expression of arginase isoforms and related enzymes in mouse tissues and in peritoneal macrophages. Results indicate that iNOS and arginase II were induced early in the lung and spleen of mice but arginase I was induced later in the lung. There was also an early induction of iNOS, arginase II, argininosuccinate synthase and ornithine decarboxylase in peritoneal macrophages.

A study was conducted to determine the specific arginase isoforms induced by interleukin (IL)-4, a Th2 cytokine shown by others to enhance arginase activity in macrophages, and serum. Findings showed that IL-4, in combination with serum, enhances arginase I, but not arginase II, mRNA in cultured murine macrophages. Also, serum induces both arginase isoforms and is needed for maximal arginase I induction by IL-4.

A study was conducted to investigate the role and regulation of arginase and inducible nitric oxide synthase (iNOS) in the macrophage cells of a mouse. Results show that increases in arginase mRNAs increase arginase activity. Moreover, complex patterns of iNOS and arginase expressions suggest that arginase isoforms may play distinct but partially overlapping functional roles in macrophage arginine metabolism.

The mechanisms by which N(supra G)-hydroxy-L-arginine (NOHA) and nitric oxide (NO) suppress the proliferation of a cultured human colon carcinoma cell line (Caco-2) was examined. Results showed that both NOHA and NO were able to inhibit Caco-2 in a concentration-dependent manner. It was observed that NOHA inhibits the Caco-2 cell arginase activity both in cell-free extracts and in intact cells. On the other hand, NO inhibits Caco-2 by inhibiting the ornithine decarboxylase present in Caco-2 cells.

A study was conducted to examine the induction of arginase isoforms in the rat lung during 100% O2 exposure. A polytron homogenizer was utilized to homogenized harvested lung tissues while perfused rat lung were inflation fixed with an intratracheal instillation of paraformaldehyde in a phosphate buffer. Results indicated that the function of pulmonary arginase is correlated with connective tissue elements during lung injury and recovery.

Research was conducted to test whether nitric oxide (NO) production in lipopolysaccharide-activated macrophages is influenced by arginase and to examine whether arginase-regulated NO production is modulated by the extracellular level of L-arginine. NO synthase was assayed for cultured cells while cell lysates were utilized for enzyme assays in the presence and absence of L-norvaline. Results indicated that inducible NO synthase activity was not influenced by L-norvaline.

The biochemical function of cortisol in the regulation of glutamine and arginine metabolism was analyzed in the enterocytes isolated from 21-day-old suckling pigs. Analysis of jejunal enterocytes isolated from suckling pigs after intramuscular injections of hydrocortisone 21-acetate and mifepristone indicated the role of cortisol in increasing the activities of arginase, argininosuccinate lyase and pyrroline-5-carboxylate (P5C) synthase. Furthermore, the induction by cortisol of P5C synthase and arginase in mammalian cells increased the rates of glutamine and arginine metabolism.

Previous studies have shown that pretreatment with lipopolysaccharide (LPS) induces arginase and nitric oxide synthase (NOS) formation in murine macrophage cells. The induction of arginase and NOS is accompanied by NO, citrulline and urea. Herein, immunoprecipitation experiments using rat aortic endothelial cells revealed that N(super G)-hydroxy-L-arginine (NOHA) and not urea is produced in association with the production of arginase and NOS. In contrast to previous results, urea is produced during the inhibition of NOS activity and is accompanied by decreased NO and NOHA production.

L-Canavanine [2-amino-4-(guanidinooxy) butyric acid], a non-protein amino acid that is structurally analogous to arginine, has been proposed as a major antinutritional factor responsible for the toxic effects induced by raw Canavalia ensiformis (L.) seeds in chicks. We investigated the effects of L-canavanine on performance and select metabolic responses of growing chicks. Canavanine was added to a control diet, in an amount equivalent to that provided by 300 g raw canavalia seeds/kg diet (10 g free base canavanine/kg diet). Growth, plasma basic amino acids and kidney arginase activity were measured. The incorporation of canavanine into a nutritionally balanced diet for growing chicks depressed feed intake and growth by [approximately] 25% (P < 0.01) compared with the control diet. Performance was unaffected by equimolar amounts of arginine. Canavanine exerted its growth-depressing effect exclusively by reducing feed intake, because this effect was not observed in a pair-feeding experiment. Chicks fed a diet containing 473 mmol canavanine sulfate/kg for 11 d were given an intracrop dose of 946 mmol of canavanine sulfate or arginine hydrochloride. In both cases, plasma histidine and lysine concentrations were significantly decreased compared with a placebo group dosed with water. Plasma arginine concentration was unaffected by the canavanine sulfate dose but, as expected, was significantly increased by the arginine hydrochloride dose. Free base canavanine significantly (P < 0.05) reduced kidney arginase activity. No overt toxic effects were observed at any point during the study. These data indicate that, although canavanine is not the principal antinutritional factor in Canavalia ensiformis seeds, its presence in the diet precludes optimum performance of chicks.

A 4-year-old girl presented at our clinic with autistic-like symptoms, aggressivity and occasional hyperactivity. She had no history of neurologic or physical symptoms. Her condition was diagnosed as pervasive developmental [...]

Research has been conducted on binuclear manganese metalloenzyme arginase which catalyzes L-arginine hydrolysis forming L-ornithine and urea. Results demonstrate that arginase II is responsible for male and female sexual arousal disorders.

X-ray crystal structures of the arginase I elucidate structure-stability relationships of the first-shell metal ligands in the binuclear manganese cluster of arginase I as well as structure-mechanism of action of the intact cluster in catalysis. Data indicate that metal ligand substitutions compromise the catalytic activity either by the loss or disruption of the metal cluster or metal-bridging hydroxide ion.

Arginase, which exists as the isoforms arginase I and II, catalyzes the hydrolysis of arginine to ornithine and urea. Ornithine is the principal precursor for production of polyamines, which are required for cell proliferation. Rat aortic smooth muscle cells (RASMC) contain constitutive arginase I, and arginase inhibitors cause inhibition of cell proliferation. The objective of this study was to determine whether the elevated expression of arginase I in RASMC causes increased cell proliferation. RASMC were stably transfected with either rat arginase I cDNA or a [Beta]-galactosidase control expression plasmid. Western blots and arginase enzymatic assays revealed high-level expression of cytosolic arginase I in arginase I-transfected RASMC. Moreover, this observation was associated with the increased production of urea and polyamines and higher rates of RASMC proliferation. The two selective inhibitors of arginase, [N.sup.G]-hydroxy-L-arginine and S-(2-boronoethyl)-L-cysteine, inhibited arginase and decreased the production of urea and polyamines in arginase I-transfected RASMC, all of which were associated with the inhibition of cell proliferation. This study demonstrates that elevated arginase I expression increases RASMC proliferation by mechanisms involving increased production of polyamines. These observations suggest that arginase I plays a potentially important role in controlling RASMC proliferation. nitric oxide | vascular smooth muscle growth | polyamines | atherosclerosis

Han, Xiaoli, Michael N. Kazarinoff, Nikolaus Seiler, and Bruce A. Stanley. Rat colon ornithine and arginine metabolism: coordinated effects after proliferative stimuli. Am J Physiol Gastrointest Liver Physiol 280: G389-G399, 2001.--Ornithine decarboxylase (ODC) catalyzes the first step in the polyamine biosynthetic pathway, a highly regulated pathway in which activity increases during rapid growth. Other enzymes also metabolize ornithine, and in hepatomas, rate of growth correlates with decreased activity of these other enzymes, which thus channels more ornithine to polyamine biosynthesis. Ornithine is produced from arginase cleavage of arginine, which also serves as the precursor for nitric oxide production. To study whether short-term coordination of ornithine and arginine metabolism exists in rat colon, ODC, ornithine aminotransferase (OAT), arginase, ornithine, arginine, and polyamine levels were measured after two stimuli (refeeding and/or deoxycholate exposure) known to synergistically induce ODC activity. Increased ODC activity was accompanied by increased putrescine levels, whereas OAT and arginase activity were reduced by either treatment, accompanied by an increase in both arginine and ornithine levels. These results indicate a rapid reciprocal change in ODC, OAT, and arginase activity in response to refeeding or deoxycholate. The accompanying increases in ornithine and arginine concentration are likely to contribute to increased flux through the polyamine and nitric oxide biosynthetic pathways in vivo. ornithine decarboxylase; arginase; ornithine aminotransferase; tumor promoters; polyamines

Kepka-Lenhart, Diane, Sanjay K. Mistry, Guoyao Wu, and Sidney M. Morris, Jr. Arginase I: a limiting factor for nitric oxide and polyamine synthesis by activated macrophages? Am J Physiol Regulatory Integrative Comp Physiol 279: R2237-R2242, 2000.--Because arginase hydrolyzes arginine to produce ornithine and urea, it has the potential to regulate nitric oxide (NO) and polyamine synthesis. We tested whether expression of the cytosolic isoform of arginase (arginase I) was limiting for NO or polyamine production by activated RAW 264.7 macrophage cells. RAW 264.7 cells, stably transfected to overexpress arginase I or [Beta]-galactosidase, were treated with interferon-[Gamma] to induce type 2 NO synthase or with lipopolysaccharide or 8-bromo-cAMP (8-BrcAMP) to induce ornithine decarboxylase. Overexpression of arginase I had no effect on NO synthesis. In contrast, cells overexpressing arginase I produced twice as much putrescine after activation than did cells expressing [Beta]-galactosidase. Cells overexpressing arginase I also produced more spermidine after treatment with 8-BrcAMP than did cells expressing [Beta]-galactosidase. Thus endogenous levels of arginase I are limiting for polyamine synthesis, but not for NO synthesis, by activated macrophage cells. This study also demonstrates that it is possible to alter arginase I levels sufficiently to affect polyamine synthesis without affecting induced NO synthesis. nitric oxide synthase; ornithine decarboxylase; putrescine; RAW 264.7

Argininemia may be misdiagnosed as cerebral palsy in some children. Argininemia is a metabolic disorder caused by reduced levels of arginase, the enzyme that metabolizes arginine. This leads to elevated blood levels of arginine. An African-American boy was diagnosed with cerebral palsy at the age of five. When he was nine, he had a seizure and his symptoms worsened. He was hospitalized, where his blood levels of arginine were found to be three times the normal concentration. A five-year-old Hispanic boy was also misdiagnosed with cerebral palsy. He was diagnosed with argininemia after a laboratory test revealed increased blood levels of arginine. Both children were put on a protein-restricted diet and treated with oral sodium phenylbutyrate.

Arginase is a multifunctional enzyme in nitrogen metabolism. There are two forms of arginase in terrestrial vertebrates (ureotelic) encoded by separate genes, ARG01 (cytosolic, hepatic) and ARG02 (mitochondrial, non-hepatic). In ammoniotelic fish (eg., rainbow trout, Oncorhynchus mykiss) without a hepatic urea cycle, arginase catalyses the conversion of dietary arginine to urea and ornithine. Trout arginase activity is largely mitochondrial. We predicted, therefore, that the arginase gene(s) in trout would be more similar to the mammalian ARG02 gene. Using PCR cloning techniques, we characterized the full (1221 bp) and partial (196 bp) coding region of two rainbow trout arginase genes (Onmy-ARG01 and ARG02). In contrast to our prediction, the nucleic acid sequences for Onmy-ARG01 and Onmy-ARG02 aligned with the mammalian ARG01 and ARG02 genes, respectively. Using Northern analysis, we determined the pattern of mRNA expression between various trout tissues. Onmy-ARG01 was expressed in liver and intestine. Onmy-ARG02 was expressed in liver, kidney, gill, heart, intestine, white muscle and red muscle. There was a 3-fold increase in liver arginase activity and a 2-fold increase in Onmy-ARG01:beta-actin mRNA levels, but no change in Onmy-ARG02 mRNA levels in juvenile fish fasted 6 weeks relative to fed fish. These findings indicate that trout arginase genes are differentially expressed and in response to dietary stress, differentially regulated.