Arginase Isoenzymes Research Articles

Determination of Serum Arginase-1 Concentrations and Serum Arginase Activity for the Non-Invasive Diagnosis of Endometriosis.

Backgroud: Endometriosis remains a diagnostic challenge, both clinically and economically, affecting 6% to 15% of women of child-bearing potential. We have attempted to determine whether testing serum concentrations and activity of arginase isoenzymes could be useful for the non-invasive diagnosis of endometriosis. Methods: This study involved 180 women (105 endometriosis subjects-study group B; 22 subjects with other benign gynaecological conditions-control group 1-K1, both undergoing surgery; and 53 healthy subjects without features of endometriosis-control group 2-K2). Results: Preoperative and postoperative arginase-1 (Arg-1) concentrations were significantly higher in patients, as compared with the control groups K1 (p < 0.0001 and p = 0.0005, respectively) and K2 (both p < 0.0001). Similarly, arginase activity was significantly higher in patients than in the control group K1 before surgery and higher than in both control groups after surgery. No significant differences in either Arg-1 concentrations or arginase activity were noted between the operated control group K1 and the non-operated control group K2. A significant postoperative decrease in Arg-1 concentration was observed within both patient (p < 0.0001) and control group K1 (p = 0.0043). Diagnostic performance was assessed using the receiver operating characteristic (ROC) method. The threshold for differentiation between endometriosis patients and healthy non-operated controls was 42.3 ng/mL, with a sensitivity of 90% and specificity of 81%. For differentiation of patients and operated controls with benign gynaecological conditions, the threshold was 78.4 ng/mL, with a sensitivity of 61% and specificity of 95%. Conclusions: We, therefore, conclude that Arg-1 serum concentrations and arginase activity could be considered potential biomarkers for endometriosis but require further studies on larger cohorts of patients.

Modulation of Arginase-2 mRNA Levels by ω-3 PUFAs and Aspirin in Asthmatic Human Lung Fibroblasts.

Airway remodeling (AR) increases disease severity, and morbidity of asthmatic patients by contributing to irreversible airflow obstruction and progressive declines in lung function. Arginase isoenzymes and the downstream enzymes ornithine decarboxylase (ODC) and ornithine aminotransferase (OAT) have been implicated in the hyperplastic and fibrotic changes of AR, respectively. Omega-3 polyunsaturated fatty acids (ω-3 PUFAs) and resolvin metabolites have anti-AR effects, but whether they are mediated through the arginase pathway is unclear. Our study intended to determine the effects of the ω-3 PUFAs eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), resolvin D1 (RvD1), TH1 cytokines, acetylsalicylic acid (ASA), cAMP, and dexamethasone (DEX) on the expression of arginase isoenzymes arginase 1 (ARG1) and arginase 2 (ARG2), ODC, and OAT in human lung fibroblasts (HLF) from normal (NHLF) and diseased (DHLF) asthmatic donors using reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR). Our data showed that EPA and EPA+DHA downregulated ARG2 mRNA 2-fold in both types of HLF. DHA, RvD1, and DEX did not alter mRNA levels for any of the genes studied. EPA lowered the ARG2 protein levels in DHLF, but did not affect those levels in NHLF. ASA upregulated ARG2 mRNA 5-fold and 7-fold in NHLF and DHLF, respectively, TH1 cytokines downregulated ARG2, ODC, and OAT mRNA in DHLF 10-fold, 2-fold, and 2.5-fold, respectively, and cAMP downregulated ARG2 mRNA 2-fold in DHLF. These results are the first to show a direct effect of ω-3 PUFAs on ARG2 mRNA levels and provide further evidence for a role of ω-3 PUFAs in AR.

A Study of Arginase Expression in Chronic Non-healing Wounds.

Arginase expression has been recently shown to increase in numerous disease states like neurodegeneration, inflammation, and malignancies. Although it has been found to be functionally important in various disease pathologies, little is known about its role in wound healing. Here, we look at the expression of arginase and its isoforms in chronic non-healing wounds and also study the expression of nitric oxide synthase (NOS) and oxidative stress enzymes in them. Wound tissues and blood samples were collected at the time of index presentation and follow-up from 61 chronic non-healing wound cases. The expression patterns of arginase isoenzymes, NOS, superoxide dismutases (SOD), lactic acid dehydrogenase (LDH), and catalase were examined by using enzyme-linked immunosorbent assay, immunohistochemistry, and western blot analysis at the transcript and protein level. We reported a significant decrease of serum arginase levels in chronic nonhealing wounds in the progress of wound healing. Interestingly, tissue arginase levels were found to be increased with improved wound condition at follow-up. Tissue NOS, LDH, and catalase activity were also found to be increased with the progress of healing, whereas SOD levels were downregulated. Our findings reported increased expression at the transcript level of arginase-I and arginase-II in chronic non-healing wounds for the first time. In conclusion, we observed decreased serum arginase levels in completely healed patients as compared to non-healed cases. Our study findings support the hypothesis that inhibition of the activity of arginase delays wound healing. Arginase and iNOS may also find their place in the future as possible biomarkers for wound healing.

Significance of arginase determination in body fluids of patients with hepatocellular carcinoma and liver cirrhosis before and after surgical treatment

ObjectiveTo assess the utility of arginase activity and expression in diagnosis of liver diseases. Design and methodsArginase activity, sensitivity and specificity were determined in serum of 140 patients including 50 with HCC, 60 with LC, 30 with choledocholithiasis (CDL) and 90 healthy controls. In HCC and LC arginase activity in serum was studied before and after tumor resection or liver transplantation. Arginase sensitivity in HCC was compared to that of alpha-fetoprotein (AFP) and aminotransferases (AST, ALT). In LC the activity was determined also in bile before and after transplantation. The expression of arginase isoenzymes in serum was studied by Western blotting. ResultsIn HCC and LC the preoperative arginase activity was significantly higher compared to controls, and it decreased after surgery. The sensitivity of arginase in HCC was much higher than that of AFP, AST and ALT (96, 40, 20 and 18%, respectively). In HCC it was higher than in LC (93%) and CDL (33%). The specificity of arginase was above 80%. In bile of cirrhotic patients the highest activity was immediately after liver transplantation. It decreased with time but increased dramatically at the time of the graft rejection. Arginase AII was present in serum of HCC and LC but not the control cases. ConclusionsThe increase of arginase activity in serum accompanied by the presence of isoenzyme AII can be useful in HCC and LC diagnosis. The determination of arginase activity in bile may be helpful in monitoring liver graft recipients.

Arginase enzymes in the human prostate: expression of arginase isoenzymes and effects of arginase inhibitors on isolated human prostate tissue

What's known on the subject? and What does the study add? A previous study by Lexander et al. in 2005, using two-dimensional gel electrophoresis, demonstrated the expression of arginase type II in the different anatomical regions of the prostate; however, to date, no study has addressed, using an in vitro approach, the role of arginase isoenzymes in the human prostate. The results of the present study demonstrate that: both arginase isoenzymes, Arg I and Arg II, are expressed in the transition zone of the human prostate; the inhibition of arginase antagonized, to a certain degree, the tension brought about by noradrenaline in isolated human prostate tissue; exposure of human prostate tissue to arginase inhibitors enhanced the local production of cyclic GMP; and inhibition of arginase enzymes in the human prostate may augment the activity of the nitric oxide/cyclicGMP pathway. • To investigate the expression of arginase isoenzymes type I (Arg I) and type II (Arg II) in the transition zone of the human prostate and the functional significance of arginase enzymes in the control of prostate smooth muscle. • Human prostate tissue was obtained from male patients who had undergone pelvic surgery. • The expression of Arg I and Arg II was investigated using Western blot analysis. • Using the organ bath technique, the effects of cumulative administration of difluoromethylornithine (DFMO), H-Orn-OH × HCl, H-Ile-OH and N-ω-hydroxy-nor-L-arginine (nor-NOHA; 1 nM-10 µM) on the tension induced by noradrenaline in isolated prostate tissue were assessed. • Tissue strips were also exposed to arginase inhibitors and the production of cyclic GMP was determined. • Western blot analysis showed the expression of Arg I and Arg II in the transition zone of the prostate. • The tension induced by noradrenaline was antagonized by the drugs in the following rank order of efficacy: H-Orn-OH × HCl ≥ H-Ile-OH ≥ DFMO > nor-NOHA; however, the maximum reversion of tension recorded ranged from only -25 to -13%. • The enhancement in cyclic GMP production registered in the presence of the arginase inhibitors ranged from four- to 14-fold. • Arg I and Arg II are expressed in the transition zone of the human prostate. • Isometric tension studies and measurement of cyclic GMP showed that inhibition of arginase can reverse, to a certain degree, the tension of human prostate tissue induced by the activation of α-adrenoceptors and enhance the accumulation of cyclic GMP. • Future studies should explore further the role of arginase enzymes in the relaxation mediated by nitric oxide in prostate smooth muscle.

Species differences in expression pattern of arginase isoenzymes and differential effects of arginase inhibition on collagen synthesis in human and rat pulmonary fibroblasts

Arginase was shown to be up-regulated in different animal models of inflammatory and fibrotic airway diseases. Since arginase provides L-ornithine, one precursor for L-proline, an essential substrate for collagen synthesis, it has been suggested that arginase might be a key enzyme in airway remodelling. The present study aimed to characterize expression of arginase isoenzymes in rat and human pulmonary fibroblasts, and to test whether arginase inhibition affects collagen synthesis. In primary rat tracheal and lung fibroblasts, mRNA for arginase I and II could be detected, with arginase I as predominant isoenzyme. In contrast, in human lung fibroblasts (primary cells and different cells lines) mRNA levels for arginase I were at or below detection limit whereas arginase II mRNA was markedly higher than in rat pulmonary fibroblasts. Arginase activity in rat tracheal and lung fibroblasts was between 20 and 30 mU/mg protein, but was below detection limit (2.5 mU/mg) in human lung fibroblasts. In rat tracheal and lung fibroblasts cultured in proline-free medium, arginase inhibition by N(omega)-hydroxy-nor-L-arginine caused a reduction by about one-third of basal collagen I accumulation (determined by western blot analysis) and largely attenuated transforming growth factor beta 1 (TGF-beta(1))-induced increase in collagen accumulation, whereas basal and TGF-beta(1)-induced collagen accumulation by human lung fibroblasts was not affected by arginase inhibition. In conclusion, arginase isoenzymes reveal a species specific expression pattern. Arginase contributes significantly to L-proline supply for collagen synthesis in rat fibroblasts, in which arginase I is the predominant isoenzyme, but not in human fibroblasts, in which arginase II is the only isoenzyme expressed.

Arginase isoenzymes in human cirrhotic liver.

Cirrhosis leads to an inability of the liver to perform its biochemical functions. It can also lead to hepatocellular carcinoma in which, as we showed lately, arginase isoenzyme pattern changes. The present work presents our results on arginase isoenzymes and their possible role in liver cirrhosis. The study was performed on tissues obtained during liver transplantation from 60 patients with liver cirrhosis, and on samples of histologically normal liver (control) from 40 patients with benign or colorectal cancer liver metastases removed during surgery, 6-7 cm from the tumor border. Arginase isoenzymes AI (so-called liver-type arginase) and AII (called extrahepatic arginase) were identified by Western blotting and isolated by ion-exchange chromatography. Their expression on mRNA level was studied by RT-PCR. A significant decrease in arginase activity, dependent of the liver clinical stage, was observed in cirrhotic tissue. Arginase AI activity and its mRNA level were significantly decreased in cirrhotic liver, whereas the activity and expression of arginase AII were concurrently raised, as compared to normal liver. Since arginase AI is a key enzyme of the urea cycle, whereas arginase AII most probably takes part in the biosynthesis of ornithine and polyamines, the defective ammonia inactivation and increased collagen biosynthesis observed in cirrhotic liver may be related to the changes in arginase AI and AII levels, respectively.

Changes in arginase isoenzymes pattern in human hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide affecting preferentially patients with liver cirrhosis. The studies were performed on tissues obtained during surgery from 50 patients with HCC, 40 with liver cirrhosis and 40 control livers. It was found that arginase activity in HCC was nearly 5- and 15-fold lower than in cirrhotic and normal livers, respectively. Isoenzymes AI (so-called liver-type arginase) and AII (extrahepatic arginase) were identified by Western blotting in all studied tissues, however the amount of AI, as well as the expression of AI-mRNA were lower in HCC, in comparison with normal liver, and those of AII were significantly higher. Since HCC is arginine-dependent, and arginine is essential for cells growth, the decrease of AI may preserve this amino acid within tumor cells. Concurrently, the rise of AII can increase the level of polyamines, compounds crucial for cells proliferation. Thus, both arginase isoenzymes seem to participate in liver cancerogenesis.

Common ligands of G‐protein‐coupled receptors and arginine‐utilizing enzymes

A new family of G-protein-coupled receptors (GPRC6A) has recently been described and characterized with unknown physiological role. Three isoforms of GPRC6A are expressed from the same gene by alternative splicing. Agonists for GPRC6A are basic amino acids, particularly the analogues and derivatives of L-arginine and L-ornithine. These compounds are known ligands of nitric oxide synthase and arginase isoenzymes, but amino-acid sequences of these enzymes have not shown significant homologies. Various hypotheses for physiological roles of these receptors have been proposed but not yet substantiated. In order to define the most potent agonists, and elucidate further the biological significance of the receptor family, a detailed ligand-based quantitative structure-activity relationship study may be helpful.

Altered expression of l-arginine metabolism pathway genes in chronic wounds in recessive dystrophic epidermolysis bullosa

Individuals with the severe, mutilating Hallopeau-Siemens form of recessive dystrophic epidermolysis bullosa (HS-RDEB) have trauma-induced blisters and skin erosions which often progress to wounds that are slow to heal. These chronic wounds cause considerable morbidity and there is an increased risk of squamous cell carcinoma arising in the wound margins. Currently, little is known about the keratinocyte cell biology in these wounds. Therefore, we compared the gene expression profiles of wound edge with nonwounded skin from two individuals with HS-RDEB. Trauma-induced wound sites had been present in both patients for more than 3 months. Hybridizations using DermArray gene expression filters showed relative differences in gene expression between wounded and unwounded skin. Notably, there was a fivefold increase in expression of arginase-1 (ARG1) in the chronic wound samples. Expression of seven other genes relevant to L-arginine metabolism also showed differences greater than twofold. L-arginine is known to have a critical role in the synthesis of nitric oxide as part of normal tissue repair. Although alterations in arginase isoenzymes have been detected previously in other chronic wounds (human and animal models), this is the first study to demonstrate differences in several components of the L-arginine metabolism pathway in chronic wounds, and the first to examine chronic wounds in HS-RDEB. The data show that the cascade of L-arginine metabolites is altered in HS-RDEB and the findings may provide new insight into the pathology of chronic wounds in this genodermatosis.

Expression and activity of arginase isoenzymes during normal and diabetes-impaired skin repair.

Within the past years, an important role for nitric oxide (NO) in skin repair has been well defined. As NO is synthesized from L-arginine by NO synthases (NOS), the availability of L-arginine might be one rate-limiting factor of NO production at the wound site. Upon injury, arginase-1 and -2 mRNA, protein, and activity were strongly induced reaching a maximum between day 3 and day 7 postwounding. Immunohistochemistry colocalized both arginases and the inducible NOS (iNOS) at epithelial sites at the margins of the wound. Notably, diabetes-impaired skin repair in leptin-deficient mice (diabetes/diabetes, db/db; and obese/obese, ob/ob) was characterized by an abnormally elevated arginase activity in wound tissue in the absence of an expression of iNOS. Expression analyses demonstrated that arginase-1 contributed to increased arginase activities in impaired repair. Interestingly, an improved healing of chronic wound situations in leptin-supplemented ob/ob mice was strongly associated with an adjustment of the dysregulated expression of L-arginine-converting enzymes: an attenuated iNOS expression was upregulated early in repair and an augmented arginase-1 expression and activity was downregulated in the presence of markedly elevated numbers of macrophages during late repair. These data suggest a coordinated consumption of L-arginine by the NOS and arginase enzymatic pathways at the wound site as a prerequisite for a balanced NO (via iNOS) and polyamine (via arginases) synthesis that drives a normal skin repair.

Glucocorticoid inhibition of interleukin-4 (IL-4) and interleukin-13 (IL-13) induced up-regulation of arginase in rat airway fibroblasts.

Arginase appears to play a significant role in the pathogenesis of inflammatory and obstructive airway diseases by interfering with NO synthesis (hyperreactivity) and by providing substrate for collagen synthesis (remodelling). IL-4 and IL-13 are key proinflammatory cytokines in asthma, and their effects on arginase in rat primary airway fibroblasts in culture were studied. Airway fibroblasts showed significant arginase activity, which was higher when the culture medium contained 10% fetal calf serum (FCS) (20 mU/mg protein) compared to 5% FCS (6 mU/mg protein). In presence of 10% FCS addition of IL-4 or IL-13 (10 ng/ml each) for 20 h or 40 h caused an increase in arginase activity by 76% and 160% (IL-4) and by 134% and 213% (IL-13), respectively. Using RT-PCR mRNA for arginase I was clearly detectable with 30 PCR cycles, whereas mRNA for arginase II was hardly detectable with 35 PCR cycles. IL-4 and IL-13 caused a clear increase in the mRNA of both arginase isoenzymes. Dexamethasone (1 microM) did not affect basal arginase activity, but largely opposed the stimulatory effects of IL-4 and IL-13. In conclusion, IL-4 and IL-13 can cause a marked up-regulation of arginase activity in rat airway fibroblasts and these effects can be inhibited by glucocorticoids.

Utilization of lacrimal urea assay in the monitoring of hemodialysis: conditions, limitations and lacrimal arginase characterization

The lacrimal urea content was found to be proportional to that of blood, which suggested its possible utilization in the monitoring of hemodialysis as a less invasive method. On the other hand, however, arginase activity was detected in tears, which may influence the urea content independently of blood urea concentration. The feasibility of using lacrimal urea measurement to replace blood urea measurement in the monitoring hemodialysis was also investigated. Blood and tear samples of 35 healthy persons and 43 renal patients undergoing hemodialysis were tested. Tear samples were collected on Schirmer paper strips. After elution the lacrimal urea content was measured by a colorimetric method. The determination of arginase activity was based on the release of urea and ornithine. The correlation between blood and lacrimal urea and arginase was studied by multivariate regression analysis. The lacrimal arginase isoenzyme pattern was investigated by native polyacrylamide gel electrophoresis and Western blotting. The effect of partially isoform-specific inhibitors was also studied. Blood urea levels in blood were significantly higher in the renal patients before dialysis than in the control patients (12·86±0·59 vs. 6·45±0·41 m m, p<0·0001). Blood sera arginase activity was very low. Lacrimal arginase activity was significantly higher in tears than in sera ( p<0·0001 for each group). The tear/serum ratio of urea content was significantly different between controls and renal patients, particularly in postdialytic samples (1·89±0·07 vs. 3·49±0·31, p<0·0001). The correlation between lacrimal and blood sera urea was best in controls ( r=0·89) and was better in predialytic ( r=0·75) than in postdialytic ( r=0·52) samples, depending on the level of arginase activity. In postdialytic samples a stronger correlation ( r=0·77) between tear urea and arginase was observed. Both arginase isoforms were detected in tears, but the extrahepatic (arginase II) isoenzyme was present in higher concentration. In conclusion, the determination of lacrimal urea level as a possible less invasive replacement for blood urea determination could only be utilized in the monitoring of hemodialysis if lacrimal arginase is also measured. Blood urea levels can be correctly determined by using equations, which take into account arginase activity. The accuracy of these equations was checked on a new patient population. Both arginase isoenzymes were observed in lacrimal samples.

Identification of two arginase isoenzyme activities along the nephron of Meriones shawi.

Conflicting theories on the existence of several renal arginase isoenzymes remain in debate. Because the activity of arginase is high in two embryologically different nephron segments of the Meriones shawi kidney, namely the cortical (CPST) and medullary (OSPST) proximal straight tubule and the outer medullary collecting duct (OMCD), we postulate that these nephron segments may contain different isoforms. Isolated nephron segments were dissected from collagenase-treated kidneys. Tubules were permeabilized with Triton X-100 (0.25%) and incubated with increasing Arg concentrations to characterize the arginase activity. The results were as follows: (1) in OMCD, one arginase isoform (E1), characterized by a high Arg affinity (1.160 mM), was present; (2) in CPST, two arginase isoforms were discovered - one, E1, had a similar Km (1.407 mM) to that found in OMCD whereas the other (E2) had a low affinity for Arg (Km =18.8 mM); and (3) in OSPST, two isoenzymes were present - E1 which had a Km of 1.478 mM and the second isoform that we named E2 which had a Km of 9.07 mM. In addition, arginase located in CPST and OMCD was strongly inhibited by Orn and Lys. The Ki value for Lys varied between 1.635 and 2.288 mM. Therefore, this work demonstrates that two arginase isoforms are present in the kidney of Meriones shawi. Isoform E1 is present in the proximal tubule and the collecting duct whereas isoform E2 is restricted to the proximal tubule.

Arginine metabolism: nitric oxide and beyond.

Arginine is one of the most versatile amino acids in animal cells, serving as a precursor for the synthesis not only of proteins but also of nitric oxide, urea, polyamines, proline, glutamate, creatine and agmatine. Of the enzymes that catalyse rate-controlling steps in arginine synthesis and catabolism, argininosuccinate synthase, the two arginase isoenzymes, the three nitric oxide synthase isoenzymes and arginine decarboxylase have been recognized in recent years as key factors in regulating newly identified aspects of arginine metabolism. In particular, changes in the activities of argininosuccinate synthase, the arginases, the inducible isoenzyme of nitric oxide synthase and also cationic amino acid transporters play major roles in determining the metabolic fates of arginine in health and disease, and recent studies have identified complex patterns of interaction among these enzymes. There is growing interest in the potential roles of the arginase isoenzymes as regulators of the synthesis of nitric oxide, polyamines, proline and glutamate. Physiological roles and relationships between the pathways of arginine synthesis and catabolism in vivo are complex and difficult to analyse, owing to compartmentalized expression of various enzymes at both organ (e.g. liver, small intestine and kidney) and subcellular (cytosol and mitochondria) levels, as well as to changes in expression during development and in response to diet, hormones and cytokines. The ongoing development of new cell lines and animal models using cDNA clones and genes for key arginine metabolic enzymes will provide new approaches more clearly elucidating the physiological roles of these enzymes.

Purification and Properties of Human Erythrocyte Arginase

An efficient method for purification of human erythrocyte arginase was developed. This method included two new procedures, hydrophobic chromatography and immunoaffinity chromatography, and yielded 0.7 mg of homogeneous arginase protein from 2.1 L of haemolysate. The molecular weight of native arginase was estimated to be 105,000 by gel filtration on a Sephadex G-150 column, and that of its subunit 35,000 by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. This indicates that the native enzyme is composed of three homologous subunits. Amino acid composition of human erythrocyte arginase was found to be very similar to that of liver arginase of several other mammals. After dialysis against distilled water, the purified arginase still retained its enzymatic activity which was decreased by EDTA and reversibly restored by Mn(II) ion. A specific polyclonal antibody for use in an immunoassay was also produced. This antibody revealed one single band on immunoelectrophoretic analysis of the acetone powder extract, suggesting absence of arginase isoenzymes in human erythrocytes.

Studies on l-arginase in developing rat small intestine, brain, and kidney. I. Ontogenic evolution of arginase isoenzymes : L. Konarska and L. Tomaszewski, Department of Clinical Biochemistry, Medical School of Warsaw, ul. Banacha 1, 02-097 Warsaw, Poland

Studies on l-arginase in developing rat small intestine, brain, and kidney. I. Ontogenic evolution of arginase isoenzymes : L. Konarska and L. Tomaszewski, Department of Clinical Biochemistry, Medical School of Warsaw, ul. Banacha 1, 02-097 Warsaw, Poland

Effect of steroid hormones on age dependent changes in rat arginase isoenzymes

Age dependent changes in rat liver and kidney arginase isoenzymes from day 1 to 73 weeks have been studied. It has been observed that total liver arginase activity (activity/liver) increases with age, but there is no appreciable change in the proportions of isoenzymes, their pH optima, electrophoretic mobilities and inhibition characteristics, although K m increases with age. Corticosterone induces liver arginase activity and the degree of induction decreases with age. The hormone has no effect on kidney arginase. Testosterone treatment, on the other hand, induces kidney arginase without any effect on liver enzyme.

Isoenzyme pattern and immunological properties of arginase in normal and hyperargininemia fibroblasts.

Arginase deficiency is an inborn error of the last step in the urea cycle and leads to profound hyperargininemia. The enzyme deficiency has been demonstrated in the liver and red blood cells. In cultured patient fibroblasts, the activity is normal. Arginase exists in multiple molecular forms only one of which is missing in hyperargininemic patients. In fibroblasts, three arginase isoenzymes can be demonstrated by DEAE-cellulose column chromatography, two by electrophoresis and by immunoprecipitation methods. From the present data, it is improbable that part of the A1 isoenzyme in fibroblasts originates from fetal calf serum arginase which supplements the culture media. None of the techniques for the separation and analyses of arginase isoenzyme allows to differentiate between the normal and the arginase-deficient phenotype. A possible explanation would be that the defect in A1 arginase observed in the liver is the result of a regulatory defect.

Separation of Arginase Isoenzymes from Human Tissues by Agar Gel Electrophoresis

Arginase (EC 3.5.3.1) from human liver. kidney, mammary gland, and erythrocytes, was separated by agar-gel electrophoresis using barbital buffer pH 8.6. Three isoenzymes were separated. Two of these, A2 and A3, occur in liver and erythrocytes. The same two isoenzymes were found in the kidney, but in reversed proportions. In addition to the A3 isoenzyme, the mammary gland contains a fast anodically moving A1 isoenzyme. The three isoenzymes differ in their degree of sensitivity to ornithine.