Competitive metabolism of L-arginine:arginase as a therapeutic target in asthma

Exhaled breath nitric oxide （NO） is an accepted asthma biomarker. Lung concentrations of NO and its amino acid precursor, L-arginine, are regulated by the relative expressions of the NO synthase （NOS） and arginase isoforms. Increased expression of arginase I and NOS2 occurs in murine models of allergic asthma and in biopsies of asthmatic airways. Although clinical trials involving the inhibition of NO-producing enzymes have shown mixed results, small molecule arginase inhibitors have shown potential as a therapeutic intervention in animal and cell culture models. Their transition to clinical trials is hampered by concerns regarding their safety and potential tox- icity. In this review, we discuss the paradigm of arginase and NOS competition for their substrate L-arginine in the asthmatic airway. We address the functional role of L-arginine in inflammation and the potential role of arginase inhibitors as therapeutics.

Arginase as a Potential Target in the Treatment of Alzheimer’s Disease

Alzheimer’s disease (AD) is a slowly progressive, neurodegenerative disorder with an insidious onset that is characterized by severe decline in memory, thinking and reasoning skills. Advanced age is a prominent risk factor for AD and other metabolic diseases, such as type II diabetes and atherosclerosis. Their causal mechanisms are multifaceted and not fully understood. The precise pathophysiology of AD remains a mystery despite decades of intensive investigation. Thus far, there is no truly successful AD therapy. Arginase is the central enzyme of the urea cycle. Recent studies have identified arginase function in the brain and associated this enzyme with the development of neurodegenerative diseases. Upregulation of arginase has been shown to contribute to endothelial dysfunction, ischemia-reperfusion, atherosclerosis, diabetes, and neurodegeneration. Other state-of-the-art discoveries of the precise molecular machinery of neurodegeneration have provided new directions for the rational development of innovative therapeutic strategies in the treatment of common neurodegenerative diseases. In this context, the regulation of arginase activity appears to be a universal approach in interfering with the pathogenesis of AD and providing relief for it and other metabolic disorders. Therefore, the enzyme represents a novel therapeutic target. Arginase inhibition has been shown to reverse amyloid-driven neuronal dysfunction and microgliosis and prevent the development of other AD symptoms in rodent models of AD. Consequently, the methodology represents a promising direction for clinical development.

C1q/TNF-related protein 1 promotes vasodilatory dysfunctions by increasing arginase 1 activity and uncoupling of endothelial nitric oxide synthase

Objective C1q/TNF-related protein(CTRP)1 was initiallyidentified as a paralog of adiponectin based on the similarity in C1q domain of these two proteins.Previously,we showed that CTRP1promotes the development of atherosclerosis by increasing endothelial adhesiveness.Here,we sought to investigate whether CTRP1 also influences vascular dilatory functions.

Mycobacterium Lrp/AsnC family transcriptional factor modulates the arginase pathway as both a sensor and a transcriptional repressor

L-Arginine is the precursor of nitric oxide(NO),a host immune effector against intracellular pathogens including Mycobacterium tuberculosis(M.tb).Pathogens including M.tb have evolved various strategies targeting arginine to block the production of NO for better survival and proliferation.However,L-arginine metabolism and regulation in Mycobacterium are poorly understood.Here,we report the identification of M.smegmatis MSMEG_1415(homolog of M.tb Rv2324)as an arginine-responsive transcriptional factor regulating the arginase pathway.In the absence of L-arginine,MSMEG_1415 acts as a repressor to inhibit the transcription of the roc(for arginine,ornithine catabolism)gene cluster,thereby switching off the arginase pathway.Treatment with L-arginine relieves the transcriptional inhibition of MSMEG_1415 on the roc gene cluster to activate the arginase pathway.Moreover,the L-arginine-MSMEG_1415 complex activates the transcription of the roc gene cluster by recognizing and binding a 15-bp palindrome motif,thereby preventing the excess accumulation of L-arginine in M.smegmatis.Physiologically,MSMEG_1415 confers mycobacteria resistance to starvation and fluoroquinolones exposure,suggestive of its important role in M.smegmatis persistence.The results uncover a unique regulatory mechanism of arginine metabolism in mycobacteria and identify M.tb Rv2324 as an attractive candidate target for the design of drugs against tuberculosis.

Thermokinetic Study on the Reversible Competitive Inhibition of Bovine Liver Arginase

A new thermokinetic reduced extent method for studying of the reversible competitive inhibition of single sub-strate enzyme-catalyzed reactions was proposed in this paper. The reaction that arginase-catalyzed hydrolysis of L-arginine to L-ornithine and urea and the inhibition of this reaction by the product, L-ornithine, and exogenous L-lysine were studied at 37 ℃ in 40 mmolL-1 sodium barbiturate-HCl buffer solution (pH=9.4). Michealis con-stant Km for arginine and maximum velocity Vm of the reaction were determined to be 5.14 mmolL-1 and 1.13× 10-2 mmolL-1s-1, respectively. The product inhibition constant KP and inhibitory constant KI of L-lysine were de-termined to be 1.18 and 5.6 mmolL-1, respectively. All the results have better repeatability and self-consistency and are in agreement with literature values. This new method using more direct thermal information from the proc-ess would give more reliable kinetic information than the traditional initial rate method.

MicroRNA-155 mediates endogenous angiotensin II type 1 receptor regulation:implications for innovative type 2 diabetes mellitus management

Type 2 diabetes mellitus(T2DM)is a lifelong condition and a threat to human health.Thorough understanding of its pathogenesis is acutely needed in order to devise innovative,preventative,and potentially curative pharmacological interventions.MicroRNAs(miRNA),are small,non-coding,one-stranded RNA molecules,that can target and silence around 60%of all human genes through translational repression.MiR-155 is an ancient,evolutionarily well-conserved miRNA,with distinct expression profiles and multifunctionality,and a target repertoire of over 241 genes involved in numerous physiological and pathological processes including hematopoietic lineage differentiation,immunity,inflammation,viral infections,cancer,cardiovascular conditions,and particularly diabetes mellitus.MiR-155 Levels are progressively reduced in aging,obesity,sarcopenia,and T2DM.Thus,the loss of coordinated repression of multiple miR-155 targets acting as negative regulators,such as C/EBPβ,HDAC4,and SOCS1 impacts insulin signaling,deteriorating glucose homeostasis,and causing insulin resistance(IR).Moreover,deranged regulation of the renin angiotensin aldosterone system(RAAS)through loss of Angiotensin II Type 1 receptor downregulation,and negated repression of ETS-1,results in unopposed detrimental Angiotensin II effects,further promoting IR.Finally,loss of BACH1 and SOCS1 repression abolishes cytoprotective,anti-oxidant,anti-apoptotic,and anti-inflam matory cellular pathways,and promotesβ-cell loss.In contrast to RAAS inhibitor treatments that further decrease already reduced miR-155 Levels,strategies to increase an ailing miR-155 production in T2DM,e.g.,the use of metformin,mineralocorticoid receptor blockers(spironolactone,eplerenone,finerenone),and verapamil,alone or in various combinations,represent current treatment options.In the future,direct tissue delivery of miRNA analogs is likely.

益肺抗瘤饮对癌细胞凝集素受体表达的影响

凝集素受体(Lectin)是一类能与特异性糖蛋白结合的蛋白质,具有凝集细胞和沉淀糖连接物的作用。它的生物学作用涉及多种生理功能。近年来作为研究的工具,已涉及多个领域,如刺激淋巴细胞分裂,细胞分离,含高糖分子的分离和纯化,细胞膜结构和生物功能研究等。肿瘤细胞膜表面有多种凝集素受体,这种受体与癌细胞的恶性程度及癌细胞的侵袭转移能力密切相关。为此,本实验采用益肺抗瘤饮制剂,观察对人肺腺癌细胞(SPC-A-1)的细胞膜表面12种凝集素受体表达的影响。

Role of the metabolism of branched-chain amino acids in the development of Alzheimer’s disease and other metabolic disorders

Alzheimer’s disease is an incurable chronic neurodegenerative disorder and the leading cause of dementia,imposing a growing economic burden upon society.The disease progression is associated with gradual deposition of amyloid plaques and the formation of neurofibrillary tangles within the brain parenchyma,yet severe dementia is the culminating phase of the enduring pathology.Converging evidence suggests that Alzheimer’s disease-related cognitive decline is the outcome of an extremely complex and persistent pathophysiological process.The disease is characterized by distinctive abnormalities apparent at systemic,histological,macromolecular,and biochemical levels.Moreover,besides the well-defined and self-evident characteristic profuse neurofibrillary tangles,dystrophic neurites,and amyloid-beta deposits,the Alzheimer’s disease-associated pathology includes neuroinflammation,substantial neuronal loss,apoptosis,extensive DNA damage,considerable mitochondrial malfunction,compromised energy metabolism,and chronic oxidative stress.Likewise,distinctive metabolic dysfunction has been named a leading cause and a hallmark of Alzheimer’s disease that is apparent decades prior to disease manifestation.State-of-theart metabolomics studies demonstrate that altered branched-chain amino acids(BCAAs)metabolism accompanies Alzheimer’s disease development.Lower plasma valine levels are correlated with accelerated cognitive decline,and,conversely,an increase in valine concentration is associated with reduced risk of Alzheimer’s disease.Additionally,a clear BCAAs-related metabolic signature has been identified in subjects with obesity,diabetes,and atherosclerosis.Also,arginine metabolism is dramatically altered in Alzheimer’s disease human brains and animal models.Accordingly,a potential role of the urea cycle in the Alzheimer’s disease development has been hypothesized,and preclinical studies utilizing intervention in the urea cycle and/or BCAAs metabolism have demonstrated clinical potential.Continual failures to offer a competent treatment strategy directed against amyloid-beta or Tau proteins-related lesions,which could face all challenges of the multifaceted Alzheimer’s disease pathology,led to the hypothesis that hyperphosphorylated Tau and deposited amyloid-beta proteins are just hallmarks or epiphenomena,but not the ultimate causes of Alzheimer’s disease.Therefore,approaches targeting amyloid-beta or Tau are not adequate to cure the disease.Accordingly,the modern scientific vision of Alzheimer’s disease etiology and pathogenesis must reach beyond the hallmarks,and look for alternative strategies and areas of research.

Circulating myeloid-derived suppressor cells in patients with pancreatic cancer

BACKGROUND： Myeloid-derived suppressor cells （MDSCs） are heterogeneous cell types that suppress T-cell responses in cancer patients and animal models, some MDSC subpopulations are increased in patients with pancreatic cancer. The present study was to investigate a specific subset of MDSCs in patients with pancreatic cancer and the mechanism of MDSCs increase in these patients. METHODS： Myeloid cells from whole blood were collected from 37 patients with pancreatic cancer, 17 with cholangiocarcinoma, and 47 healthy controls. Four pancreatic cancer cell lines were co- cultured with normal peripheral blood mononudear cells （PBMCs） to test the effect of tumor cells on the conversion of PBMCs to MDSCs. Levels of granulocyte-macrophage colony-stimulating factor （GM-CSF） and arginase activity in the plasma of cancer patients were analyzed by enzyme-linked immunosorbent assay. RESULTS： CD14＋/CD11b＋/HLA-DR MDSCs were increased in patients with pancreatic or bile duct cancer compared with those in healthy controls, and this increase was correlated with clinical cancer stage. Pancreatic cancer cell lines induced PBMCs to MDSCs in a dose-dependent manner. GM-CSF and arginase activity levels were significantly increased in the se rum of patients with pancreatic cancer. CONCLUSIONS： MDSCs were tumor related： tumor cells induced PBMCs to MDSCs in a dose-dependent manner and circulating CD14＋/CD11b＋/HLA-DR- MDSCs in pancreatic cancer patients were positively correlated with tumor burden. MDSCs might be useful markers for pancreatic cancer detection and progression.

Proteomic analysis of differentially expressed proteins involving in liver metastasis of human colorectal carcinoma

BACKGROUND: Early diagnosis of liver metastasis of colorectal carcinoma is very important for the appropriate treatment of such patients. However, there has been no effective approach available for clinical application. The present study aimed to investigate the differential expression of proteins in patients with liver metastasis of colorectal carcinomas using proteomic analysis and evaluate its potentiality in clinical diagnosis. METHODS: Fluorescence two-dimensional differential in-gel electrophoresis (2-D DIGE) was used to analyze and compare the protein expression between normal mucosa, the primary focus, and liver metastases. Proteomic analysis was made to identify the differentially expressed proteins. Immunohistological staining was used to confirm the expression of differentially expressed proteins in colorectal carcinomas and areas of liver metastasis. RESULTS: A 1.5-fold difference was found with 46 differentially expressed proteins. In 20 differentially expressed proteins, 3 were down-regulated and 17 up-regulated in liver metastases. Proteomic analysis showed that the S-adenosylmethionine transgelin variant was down-regulated in liver metastasis tissues. Zinc finger protein 64 homolog (Zfp64), guanine nucleotide exchange factor 4 (GEF4), human arginase, glutathione S-transferases (GSTs) A3, and tumor necrosis factor a (TNF-alpha)-induced protein 9 were up-regulated in liver metastasis tissues. Immunohistochemical staining confirmed that human arginase expression was higher in liver metastases than in the primary focus. CONCLUSIONS: There was a significant difference in protein expression between the primary focus of colorectal carcinoma and liver metastases. The differentially regulated proteins were closely related to liver metastasis of colorectal carcinoma. Elevated human arginase may be an important molecular marker for liver metastasis from colorectal carcinoma. (Hepatobiliary Pancreat Dis Jut 2010; 9: 149-153)

L-Norvaline, a new therapeutic agent against Alzheimer’s disease

Growing evidence highlights the role of arginase activity in the manifestation of Alzheimer’s disease(AD).Upregulation of arginase was shown to contribute to neurodegeneration.Regulation of arginase activity appears to be a promising approach for interfering with the pathogenesis of AD.Therefore,the enzyme represents a novel therapeutic target.In this study,we administered an arginase inhibitor,L-norvaline(250 mg/L),for 2.5 months to a triple-transgenic model(3×Tg-AD)harboring PS1M146V,APPSwe,and tauP301L transgenes.Then,the neuroprotective effects of L-norvaline were evaluated using immunohistochemistry,proteomics,and quantitative polymerase chain reaction assays.Finally,we identified the biological pathways activated by the treatment.Remarkably,L-norvaline treatment reverses the cognitive decline in AD mice.The treatment is neuroprotective as indicated by reduced beta-amyloidosis,alleviated microgliosis,and reduced tumor necrosis factor transcription levels.Moreover,elevated levels of neuroplasticity related postsynaptic density protein 95 were detected in the hippocampi of mice treated with L-norvaline.Furthermore,we disclosed several biological pathways,which were involved in cell survival and neuroplasticity and were activated by the treatment.Through these modes of action,L-norvaline has the potential to improve the symptoms of AD and even interferes with its pathogenesis.As such,L-norvaline is a promising neuroprotective molecule that might be tailored for the treatment of a range of neurodegenerative disorders.The study was approved by the Bar-Ilan University Animal Care and Use Committee(approval No.82-10-2017)on October 1,2017.

Common and small molecules as the ultimate regulatory and effector mediators of antigen-specific transplantation reactions

In spite of intensive research, the molecular basis of allograft and xenograft rejection still remains not fully understood. The acute rejection of an allograft is associated with the intragraft Th1 cytokine response, while tolerance of an allograft or xenograft rejection is accompanied by a higher production of the Th2 cytokines interleukin(IL)-4 and IL-10. Nevertheless, these cytokines are not the final regulatory and effector molecules mediating transplantation reactions. Data indicate that the functioning of common molecules with enzymatic activities, such are inducible nitric oxide synthase(i NOS), arginase, heme oxygenase-1(HO-1) or indoleamine-2,3-dioxygenase(IDO), the bioavailability of their substrates(L-arginine, tryptophan, heme) and the cytotoxic and regulatory actions of their small gaseous products(NO, CO) can be the ultimate mechanisms responsible for effector or regulatory reactions. Using models of transplantation immunity and tolerance we show that T cell receptor-mediated recognition of allogeneic or xenogeneic antigens as well as the balance between immunity/tolerance induces distinct cytokine production profiles. The ratio between Th1 and Th2 cytokines efficiently regulates the expression of genes for common enzymes, such as i NOS, arginase, HO-1 and IDO. These enzymes may compete for substrates, such as L-arginine or tryptophan, and the final product of their activity are small molecules(NO, CO) displaying effector or regulatory functions of the immune system. Thus, it is suggested that in spite of the high immunological specificity of transplatation reaction, the ultimate players in regulatory and effector functions could be small and common molecules.

TAK1 Deficiency in Macrophages Increases Host Susceptibility to Leishmania Infection

Leishmania parasites mainly infect macrophages and may cause severe immunopathologies in their host,which are called leishman-iases.In the current work,we infected human and mouse macrophages in vitro with Leishmania major,an etiological agent of cu-taneous leishmaniasis,and found that inhibition or deletion of the transforming growth factorβ–activated kinase 1(TAK1)gene re-sulted in increased parasite loads.In vivo,following a challenge with L.major,mice with a macrophage-specific deletion of TAK1 showed increased clinical signs and higher parasite loads compared with wild-type controls.TAK1 deficiency in mouse macro-phages led to biased Th2 cell responses during the acute stage of infection,characterized by a decrease in interferon-γexpression,and increased expression of IL-4,IL-5 and IL-10.Finally,we found that,in the late stage of L.major infection,excessive Th2-related cytokines led to high arginase 1 expression in mouse tissues and a significant reduction of NO production both locally and system-ically,resulting in compromised control of Leishmania.These findings suggest that TAK1 plays a vital role in host resistance to Leish-mania infection.

Arginine metabolism and its functions in growth, nutrient utilization, and immunonutrition of fish

Fish have limited ability in endogenous biosynthesis of arginine.Arginine is an indispensable amino acid for fish,and the arginine requirement varies with fish species and fish size.Recent studies on fish have demonstrated that arginine influences nutrient metabolism,stimulates insulin release,is involved in nonspecific immune responses and antioxidant responses,and elevates disease resistance.Specifically,arginine can regulate energy homeostasis via modulating the adenosine 5'-monophosphate(AMP)-activated protein kinase(AMPK)pathway,and also regulate protein synthesis via activating the target of rapamycin(TOR)signaling pathway.The present article reviews pertinent knowledge of arginine in fish,including dietary quantitative requirements,endogenous anabolism and catabolism,regulation of the endocrine and metabolic systems,and immune-regulatory functions under pathogenic challenge.Our findings showed that further data about the distribution of arginine after intake into specific cells,its sub-cellular sensor to initiate downstream signaling pathways,and its effects on fish mucosal immunity,especially the adaptive immune response against pathogenic infection in different species,are urgently needed.

nduction of M2-1ike macrophages in recipient NOD-scid mice by allogeneic donor CD4＋CD25＋ regulatory T cells

CD4＋CD25＋ regulatory T cells （Tregs） play an important role in maintaining host immune tolerance via regulation of the phenotype and function of the innate and adaptive immune cells. Whether allogeneic CD4＋CD25＋ Tregs can regulate recipient mouse macrophages is unknown. The effect of allogeneic donor CD4＋CD25＋ Tregs on recipient mouse resident F4/80＋macrophages was investigated using a mouse model in which allogeneic donor CD4＋CD25＋ Tregs were adoptively transferred into the peritoneal cavity of host NOD-scid mice. The phenotype and function of the recipient macrophages were then assayed. The peritoneal F4/80＋ macrophages in the recipient mice that received the allogeneic CD4＋CD25＋ Tregs expressed significantly higher levels of CD23 and programmed cell death-ligand I（PD-L1） and lower levels of CD80, CD86, CD40 and MHC II molecules compared to the mice that received either allogeneic CD4＋CD25- T cells （Teffs） or no cells. The resident F4/80＋ macrophages of the recipient mice injected with the allogeneic donor CD4＋CD25＋ Tregs displayed significantly increased phagocytosis of chicken red blood cells （cRBCs） and arginase activity together with increased IL-IO production, whereas these macrophages also showed decreased immunogenicity and nitric oxide （NO） production. Blocking arginase partially but significantly reversed the effects of CD4＋CD25＋ Tregs with regard to the induction of the M2 macrophages in vivo. Therefore, the allogeneic donor CD4＋CD25＋ Tregs can induce the M2 macrophages in recipient mice at least in part via an arginase pathway. We have provided in vivo evidence to support the unknown pathways by which allogeneic donor CD4＋CD25＋ Tregs regulate innate immunity in recipient mice by promoting the differentiation of M2 macrophages.

RBP-J is required for M2 macrophage polarization in response to chitin and mediates expression of a subset of M2 genes

Development of alternatively activated （M2） macrophage phenotypes is a complex process that is coordinately regulated by a plethora of pathways and factors. Here, we report that RBP-J, a DNA-binding protein that integrates signals from multiple pathways including the Notch pathway, is critically involved in polarization of M2 macrophages. Mice deficient in RBP-J in the myeloid compartment exhibited impaired M2 phenotypes in vivo in a chitin-induced model of M2 polarization. Consistent with the in vivo findings, M2 polarization was partially compromised in vitro in Rbpj-deficient macrophages as demonstrated by reduced expression of a subset of M2 effector molecules including arginase 1. Functionally, myeloid Rbpj deficiency impaired M2 effector functions including recruitment of eosinophils and suppression of T cell proliferation. Collectively, we have identified RBP- Jas an essential regulator of differentiation and function of alternatively activated macrophages.

Defective arginine metabolism impairs mitochondrial homeostasis in Caenorhabditis elegans

Arginine catabolism involves enzyme-dependent reactions in both mitochondria and the cytosol,defects in which may lead to hyperargininemia,a devastating developmental disorder.It is largely unknown if defective arginine catabolism has any effects on mitochondria.Here we report that normal arginine catabolism is essential for mitochondrial homeostasis in Caenorhabditis elegans.Mutations of the arginase gene argn-1 lead to abnormal mitochondrial enlargement and reduced adenosine triphosphate(ATP)production in C elegans hypodermal cells.ARGN-1 localizes to mitochondria and its loss causes arginine accumulation,which disrupts mitochondrial dynamics.Heterologous expression of human ARGl or ARG2 rescued the mitochondrial defects of argn-1 mutants.Importantly,genetic inactivation of the mitochondrial basic amino acid transporter SLC-25A29 or the mitochondrial glutamate transporter SLC-25A18.1 fully suppressed the mitochondrial defects caused by argn-1 mutations.These findings suggest that mitochondrial damage probably contributes to the pathogenesis of hyperargininemia and provide clues for developing therapeutic treatments for hyperargininemia.