Ornithine Decarboxylase Activity Research Articles

Putrescine independent wound response phenotype is produced by ODC-like RNAi in planarians

Despite increasing evidence indicates polyamines as a convergence point for signaling pathways, including cell growth and differentiation, a unifying concept to interpret their role is still missing. The activity of ornithine decarboxylase (ODC), the rate-limiting enzyme in polyamine biosynthesis, is tightly regulated by a complex molecular machinery, and the demonstration of the existence of multiple ODC paralogs, lacking decarboxylation activity, suggests additional layers of complexity to the intricate ODC regulatory pathway. Because of their extraordinary regenerative abilities and abundance of stem cells, planarians have potential to contribute to our understanding of polyamine function in an in vivo context. We undertook a study on ODC function in planarians and we found six planarian ODCs (ODC1-6). Five out of six ODC homologs carry substitutions of key aminoacids for enzymatic activity, which makes them theoretically unable to decarboxylate ornithine. Silencing of ODC5 and 6 produced a complex phenotype, by prompting animals to an aberrant response, following chronic injury without tissue removal. Phenotype is neither rescued by putrescine, nor mimicked by difluoromethylornithine treatment. Moreover, the co-silencing of other genes of the ODC regulatory pathway did not modulate phenotype outcome or severity, thus suggesting that the function/s of these ODC-like proteins might be unrelated to decarboxylase activity and putrescine production.

A Label-Free Continuous Fluorescence-Based Assay for Monitoring Ornithine Decarboxylase Activity with a Synthetic Putrescine Receptor

Polyamines play an important role in cell growth, differentiation, and cancer development, and the biosynthetic pathway of polyamines is established as a drug target for the treatment of parasitic diseases, neoplasia, and cancer chemoprevention. The key enzyme in polyamine biosynthesis is ornithine decarboxylase (ODC). We report herein an analytical method for the continuous fluorescence monitoring of ODC activity based on the supramolecular receptor cucurbit[6]uril (CB6) and the fluorescent dye trans-4-[4-(dimethylamino)styryl]-1-methylpyridinium iodide (DSMI). CB6 has a significantly higher binding constant to the ODC product putrescine (>107 M–1) than to the substrate L-ornithine (340 M–1). This enables real-time monitoring of the enzymatic reaction through a continuous fluorescence change caused by dye displacement from the macrocycle by the formed product, which allowed a straightforward determination of enzyme kinetic parameters (kcat = 0.12 s–1 and KM = 24 µM) and inhibition constants of the two ODC inhibitors α-difluoromethylornithine (DFMO) and epigallocatechin gallate (EGCG). The potential for high-throughput screening (HTS) was demonstrated by excellent Z′ factors (>0.9) in a microplate reader format, and the sensitivity of the assay is comparable to or better than most established complementary methods, which invariably have the disadvantage of not being compatible with direct implementation and upscaling to HTS format in the drug discovery process.

Characterization of an androgen-responsive, ornithine decarboxylase-related protein in mouse kidney.

We have investigated and characterized a novel ornithine decarboxylase (ODC) related protein (ODCrp) also annotated as gm853. ODCrp shows 41% amino acid sequence identity with ODC and 38% with ODC antizyme inhibitor 1 (AZIN1). The Odcrp gene is selectively expressed in the epithelium of proximal tubuli of mouse kidney with higher expression in males than in females. Like Odc in mouse kidney, Odcrp is also androgen responsive with androgen receptor (AR)-binding loci within its regulatory region. ODCrp forms homodimers but does not heterodimerize with ODC. Although ODCrp contains 20 amino acid residues known to be necessary for the catalytic activity of ODC, no decarboxylase activity could be found with ornithine, lysine or arginine as substrates. ODCrp does not function as an AZIN, as it neither binds ODC antizyme 1 (OAZ1) nor prevents OAZ-mediated inactivation and degradation of ODC. ODCrp itself is degraded via ubiquination and mutation of Cys363 (corresponding to Cys360 of ODC) appears to destabilize the protein. Evidence for a function of ODCrp was found in ODC assays on lysates from transfected Cos-7 cells where ODCrp repressed the activity of endogenous ODC while Cys363Ala mutated ODCrp increased the enzymatic activity of endogenous ODC.

Abstract 2474: The mechanistic role of polyamines in DNA double-strand break repair

Abstract Polyamines are essential organic cations for cell growth, proliferation and tissue remodeling. The rate-limiting step in the biosynthesis of polyamines is catalyzed by ornithine decarboxylase (ODC). The activity of ODC and the level of intracellular polyamines significantly increase in many cancers, indicating the critical role of polyamines in tumorigenesis. Depletion of polyamines sensitizes cells to DNA damage agents that lead to DNA double-strand breaks (DSBs), suggesting that endogenous polyamines may participate in DNA repair process. However, the mechanistic role of polyamines in DSB repair remains largely unknown. Here, we wish to ascertain the role of polyamines in DSBs repair pathway and to delineate the underlying mechanism by combining biochemical analyses, cell-based and animal approaches. Our cell-based experiments demonstrated that polyamines indeed participate in DNA double-strand break repair. Specifically, the level of intracellular polyamines regulates homologous recombination (HR) repair pathway rather than non-homologous end joining (NHEJ). Furthermore, our in vitro reconstitution system and functional analyses demonstrated that polyamines significantly enhance RAD51-mediated DNA strand exchange reaction. Importantly, we reveal that the stimulatory effect of polyamines by RAD51 stems from the enhancement of duplex DNA captures in the DNA exchange reaction. Finally, the physiological role of polyamines in DSBs repair was examined by using mouse hair follicle as a model system. The animal model clearly highlights the significant contribution of polyamines to the DNA repair in vivo. Our findings thus furnish valuable insight into the mechanistic basis of polyamines in DNA repair. Citation Format: Chih-Ying Lee, Guan-Chin Su, Min-Yu Ko, Wen-Yen Huang, Geen-Dong Chang, Sung-Jan Lin, Peter Chi. The mechanistic role of polyamines in DNA double-strand break repair [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2474. doi:10.1158/1538-7445.AM2017-2474

ROLES OF ORNITHINE DECARBOXYLASE (ODC) IN REGULATION OF AMYLOID β-INDUCED MICROGLIAL NEUROINFLAMMATION

Alzheimer's disease (AD) is the most common neurodegenerative disorder characterized by amyloid β (Aβ) deposition. Increasing evidence supports neuroinflammation is one of primarily concerns in AD. Particularly, microglia are the primary immune cells contribute to neuroinflammation by releasing various proinflammatory cytokines and neurotoxins in the brain. Therefore, inhibition of microglia shows potential value of therapeutic or preventive intervention. Interestingly, it has been demonstrated that arginine metabolism pathway may contribute to the balance between M2 and M1 phenotypes of microglia. In particular, ornithine decarboxylase (ODC) is a rate-limiting enzyme to the regulation of arginine catabolism. As a result, blocking ODC activity may result in an alternative metabolic pathway of arginine in stimulation of inducible NO synthase (iNOS). This is believed to drive neuroinflammation in AD pathogenesis. However, the detailed mechanism remains unclear. In this study, we investigated whether Aβ disrupts ODC homeostasis and contribute to microglial over-activation and neurotoxicity by BV2 and EOC13.31 microglial cell lines. Our results indicated that Aβ interferes endogenous ODC homeostasis and its downstream effects including altered arginine metabolism pathway resulting in microglial over-activation and neurotoxicity. On the contrary, restoration of ODC activity was associated with an increase in M2/M1 microglia ratio, indicating that Aβ-inhibited ODC may contribute to microglial inflammation. Our results demonstrated the modulation of ODC activities in microglia may be a workable strategy for therapeutics against Aβ-induced microglial neuroinflammation.

Abstract 2006: AMX-513 polyamine depletion therapy inhibits tumor growth and reverses immunosuppression in cancers including MYC-driven neuroblastoma and pancreatic cancer

Abstract Tumorigenesis is associated with increased polyamine levels and involves the induction of ornithine decarboxylase (ODC), the initial rate-limiting enzyme in polyamine biosynthesis, and increased uptake of polyamines from the blood and diet. As well as contributing to proliferation, polyamines are reported to exert an immunosuppressive effect. Amplification of the MYC/MYCN oncogenes has been shown to directly induce ODC activity and inhibition of this enzyme by α-difluoromethylornithine (DFMO) markedly delays tumor development. Aminex Therapeutics is developing a polyamine depletion approach targeting both biosynthesis and transport of polyamines with AMX-513, a combination of the approved ODC inhibitor, DFMO, together with AMXT 1501, an alkylated polyamine mimetic which blocks polyamine uptake. In the syngeneic CT26.CL25 mouse model of colorectal cancer, AMX-513, dosed daily for four weeks, reduced tumor growth > 75% compared to vehicle-treated control in immunocompetent Balb/C mice. There was no effect in athymic nude mice indicating that tumor growth inhibition by AMX-513 is T-cell-dependent. In the induced transgenic K6/ODC squamous tumor mouse model, stable regression was sustained 10 weeks after treatment ended and was accompanied by tumor infiltrate increases in IFNγ and in CD3+ and CD8+ T-cells. Tumor infiltrates from AMX-513-treated KPC pancreatic cancer transgenic mice with tumor regressions showed >90% reductions in myeloid-derived suppressor cells (MDSCs; CD11b+ Gr-1+) but no changes in mature myeloid cells (CD11+Gr-1neg) by FACS analysis. AMX-513 treatment did not impact the percentage or number of CD4+CD25+FoxP3+ Tregs, but did significantly increase the percentage of activated CD8+ T cells in tumors. Neuroblastoma is an aggressive childhood cancer frequently associated with MYCN and ODC deregulation. In neuroblastoma cell lines, the AMX-513 combination was highly synergistic (CI<0.5). Prophylactic treatment of neuroblastoma-prone TH-MYCN transgenic mice with AMX-513 significantly extended survival compared to either agent alone (median survival time = 81.0±11.8 days versus DFMO alone = 57.1±7.1 days; P<0.0001). Treatment of mice with small palpable tumors with AMX-513 in combination with cyclophosphamide/topotecan significantly improved survival compared with either AMX-513 or cyclo/topo alone (5/9 long term survivors compared to 0/10 and 0/9 for cyclo/topo and AMT-513, respectively; P<0.001 in each case). Polyamine levels were significantly decreased in mice undergoing AMX-513 treatment compared to DFMO or AMXT 1501 alone. In conclusion, AMX-513 treatment alone or in combination with other cancer therapies results in significant tumor growth reduction in multiple cancer models and demonstrates novel immunotherapeutic potential. Clinical evaluation of AMX-513 is planned in 2017. Citation Format: Mark R. Burns, Kathy Fosnaugh, Michael G. Palfreyman, Laura Gamble, Jayne Murray, Sophie Allan, Georgina Eden, Sara Sarraf, Murray Norris, David Ziegler, Michelle Haber. AMX-513 polyamine depletion therapy inhibits tumor growth and reverses immunosuppression in cancers including MYC-driven neuroblastoma and pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2006. doi:10.1158/1538-7445.AM2017-2006

Antiproliferative and antimicrobial efficacy of the compounds isolated from the roots of Oenothera biennis L.

Oenothera biennis L., commonly known as evening primrose, harbours the flavonoids, steroids, tannins, fatty acids and terpenoids responsible for a diverse range of biological activity, such as antitumour, anti-arthritic and anti-inflammatory effects. In addition to the previous reports from aerial parts of this plant, studies related to antiproliferative or antimicrobial activity from the roots are warranted. To investigate antiproliferative and antimicrobial activity of compounds/mixture (1-8) isolated and characterized from the roots of O.biennis L. A possible mechanism of antiproliferative activity was also studied by targeting ornithine decarboxylase (ODC) and cathepsin D (CATD). Antiproliferative efficacy of the compounds/mixture was examined in selected cancer cell lines along with their probable mechanism of action. The antimicrobial activity was also studied against selected microbes (bacteria and fungi). Antiproliferative potential was evaluated by MTT assay against selected cell lines. The mechanism of action was studied spectrophotometrically by targeting ODC and CATD using both an in-vitro and an in-silico approach. The antimicrobial efficiency was analysed using the disc diffusion and broth dilution methods. Oenotheralanosterol B (3) and the mixture of oenotheralanosterol A and oenotheralanosterol B (4) exhibited antiproliferative activity against breast, hepatic, prostate and leukaemia cancer cell lines as well as in mouse macrophages (IC50 8.35-49.69μg/ml). Oenotheralanosterol B (3) and the mixture of oenotheralanosterol A and oenotheralanosterol B (4) displayed a strong molecular interaction with succinate dehydrogenase (binding energy -6.23 and -6.84kcal/mol and Ki 27.03 and 9.6μm, respectively). Oenotheralanosterol A (1), oenotheralanosterol B (3) and mixture of oenotheralanosterol A and oenotheralanosterol B (4) potently inhibited the ODC activity with IC50 ranging from 4.65±0.35 to 19.06±4.16μg/ml and also showed a strong interaction with ODC (BE -4.17 to -4.46kcal/mol). Oenotheralanosterol A (1), cetoleilyl diglucoside (2), oenotheralanosterol B (3), dihydroxyprenylxanthone acetylated (6) and dihydroxyprenylxanthone (7) inhibited CATD activity (IC50 3.95±0.49 to 24.35±2.89μg/ml). The in-silico molecular interaction analysis of compounds with CATD revealed the non-specific interaction. A moderate antimicrobial activity was observed against selected microbes with a growth inhibition ranging from 6 to 14mm and minimum inhibitory concentration between 125 and 500μg/ml. Oenotheralanosterol B (3) and dihydroxyprenylxanthone acetylated (6) exhibited better antimicrobial activity with an MIC range from 62.50 to 500μg/ml. Oenotheralanosterol B (3) exhibited stronger antiproliferative and antimicrobial potential with respect to the other compounds tested, whereas oenotheralanosterol A (1) was a potent inhibitor of ODC and CATD. Hence, it is suggested that these in-vitro findings could be studied further invivo for biological activity, safety evaluation and derivatization to enhance potency and efficacy.

The effect of Tetraneura ulmi L. galling process on the activity of amino acid decarboxylases and the content of biogenic amines in Siberian elm tissues.

Tetraneura ulmi (L.), a member of Eriosomatinae subfamily, is one of the gall-forming aphids occurring on elms. Sap-sucking behaviour of founding mothers results in the formation of new plant organs. This study documents the changes in the content of plant biogenic amines (putrescine, cadaverine, spermidine, tryptamine, spermine and histamine) and key enzymes of their biosynthesis: lysine decarboxylase (LDC), tyrosine decarboxylase and ornithine decarboxylase (ODC) in galls and other parts of Siberian elm (Ulmus pumila L.) leaves during the galling process. The direction and intensity of these changes for particular amines and enzymes were dependent on the stage of gall development and part of the galling leaf. Generally, the amine content tended to increase in gall tissues during the 1st and 2nd period of the galling process and decreased in later phases. LDC and ODC activities were markedly enhanced, especially in gall tissues at the initial stage of the galling process.

Biochemical evaluation of the anticancer potential of the polyamine-based nanocarrier Nano11047.

Synthesizing polycationic polymers directly from existing drugs overcomes the drug-loading limitations often associated with pharmacologically inert nanocarriers. We recently described nanocarriers formed from a first-generation polyamine analogue, bis(ethyl)norspermine (BENSpm), that could simultaneously target polyamine metabolism while delivering therapeutic nucleic acids. In the current study, we describe the synthesis and evaluation of self-immolative nanocarriers derived from the second-generation polyamine analogue PG-11047. Polyamines are absolutely essential for proliferation and their metabolism is frequently dysregulated in cancer. Through its effects on polyamine metabolism, PG-11047 effectively inhibits tumor growth in cancer cell lines of multiple origins as well as in human tumor mouse xenografts. Promising clinical trials have been completed verifying the safety and tolerance of this rotationally restricted polyamine analogue. We therefore used PG-11047 as the basis for Nano11047, a biodegradable, prodrug nanocarrier capable of targeting polyamine metabolism. Following exposure of lung cancer cell lines to Nano11047, uptake and intracellular degradation into the parent compound PG-11047 was observed. The release of PG-11047 highly induced the polyamine catabolic enzyme activities of spermidine/spermine N1-acetyltransferase (SSAT) and spermine oxidase (SMOX). By contrast, the activity of ornithine decarboxylase (ODC), a rate-limiting enzyme in polyamine biosynthesis and a putative oncogene, was decreased. Consequently, intracellular levels of the natural polyamines were depleted concurrent with tumor cell growth inhibition. This availability of Nano11047 as a novel drug form and potential nucleic acid delivery vector will potentially benefit and encourage future clinical studies.

The Variation Tendency of Polyamines Forms and Components of Polyamine Metabolism in Zoysiagrass (Zoysia japonica Steud.) to Salt Stress with Exogenous Spermidine Application.

To understand dynamic changes in polyamines (PAs) forms and components of polyamine metabolism in zoysiagrass (Zoysia japonica Steud.) response to salt stress with exogenous spermidine (Spd) application, two Chinese zoysia cultivars, z081 and z057, were exposed to sodium chloride stress for 2, 4, 6, and 8 days. The z057 cultivar possesses higher salinity tolerance than the z081 cultivar. Salt stress decreased the zoysiagrass fresh weight (FW) and increased free Spd and spermine (Spm) levels and soluble and insoluble putrescine (Put), Spd and Spm levels in both cultivars. Moreover, salt stress enhanced the activities of arginine decarboxylase (ADC), ornithine decarboxylase (ODC), S-adenosylmethionine decarboxylase (SAMDC), and diamine oxidase (DAO). Exogenous Spd increased PA metabolism and ADC, SAMDC, and DAO activities and decreased free Put levels under salt stress conditions in both cultivars. In addition, structural equation modeling (SEM) showed that ODC, SAMDC, and DAO contributed to PA metabolism, and endogenous Spd levels also contributed to endogenous Spm levels. Free PAs may be the primary factor influencing the variation of other PA forms. SEM also indicated that ADC and polyamine oxidase (PAO) play a limited role in enhancing zoysia salt tolerance via PA metabolism under salt stress.

Protective Effect of Lactobacillus casei on DMH-Induced Colon Carcinogenesis in Mice.

The administration of probiotics is a promising approach to reduce the prevalence of colon cancer, a multifactorial disease, with hereditary factors, as well as environmental lifestyle-related risk factors. Biogenic polyamines, putrescine, spermidine, and spermine are small cationic molecules with great roles in cell proliferation and differentiation as well as regulation of gene expression. Ornithine decarboxylase is the first rate-limiting enzyme for polyamine synthesis, and upregulation of ornithine decarboxylase activity and polyamine metabolism has been associated with abnormal cell proliferation. This paper is focused on studying the protective role of Lactobacillus casei ATCC 393 in a chemically induced mouse model of colon carcinogenesis, directing our attention on aberrant crypt foci as preneoplastic markers, and on polyamine metabolism as a possible key player in carcinogenesis. BALB/c mice were administered 1,2-dimethylhydrazine dihydrochloride (DMH) to induce colon cancer (20mg/kg body weight, subcutaneous, twice a week for 24weeks). L. casei ATCC 393 was given orally (106CFU, twice a week), 2weeks before DMH administration. Hematoxylin and eosin staining, high-performance liquid chromatography, and Western blotting were used to evaluate aberrant crypt foci, urinary polyamines, and ornithine decarboxylase expression in the colon. The experimental data showed that the preventive administration of L. casei ATCC 393 may delay the onset of cancer as it significantly reduced the number of DMH-induced aberrant crypt foci, the levels of putrescine, and the expression of ornithine decarboxylase. Hence, this probiotic strain has a prospective role in protection against colon carcinogenesis, and its antimutagenic activity may be associated with the maintenance of polyamine metabolism.

Short communication: Arginase inhibition reduces the synthesis of casein in bovine mammary epithelial cells

The main purpose of this work was to determine the effect of arginase inhibition via Nω-hydroxy-nor-l-arginine (nor-NOHA) on casein synthesis in bovine mammary epithelial cells (BMEC). Passage 2 BMEC isolated from dairy cows were seeded to 6-well plates and randomly divided into 4 treatments: (1) control [Dulbecco's modified Eagle medium:Nutrient Mixture F-12 medium (DMEM/F12)]; (2) nor-NOHA (DMEM/F12 + 1 mmol/L nor-NOHA); (3) nor-NOHA + arginine (DMEM/F12 + 1 mmol/L nor-NOHA + 3.2 mmol/L Arg); and (4) nor-NOHA + ornithine (DMEM/F12+ 1 mmol/L nor-NOHA + 1 mmol/L Orn). Then, we determined the activity of enzymes related to Arg metabolism and casein synthesis in BMEC and the proliferation of cells. The addition of nor-NOHA reduced the activity of arginase and ornithine decarboxylase but had no effect on the activity of nitric oxide synthase, and these responses were the same at the gene expression level. In addition, supplementation of nor-NOHA in BMEC reduced cellular proliferation and casein synthesis. Addition of Arg to nor-NOHA resulted in cellular proliferation and casein synthesis similar to that of nor-NOHA alone. In contrast, addition of Orn to the medium with nor-NOHA increased the synthesis of casein and cellular proliferation compared with Nor-NOHA. In conclusion, suppression of the Arg-arginase-Orn pathway reduced casein synthesis and cellular proliferation, which indicated that this pathway is an important regulator of the synthesis of casein in BMEC.

Ornithine decarboxylase or gamma-glutamylcysteine synthetase overexpression protects Leishmania (Vianna) guyanensis against antimony

Trypanosomatids present a unique mechanism for detoxification of peroxides that is dependent on trypanothione (bisglutathionylspermidine). Ornithine decarboxylase (ODC) and γ-glutamylcysteine synthetase (GSH1) produce molecules that are direct precursors of trypanothione. In this study, Leishmania guyanensis odc and gsh1 overexpressor cell lines were generated to investigate the contribution of these genes to the trivalent antimony (SbIII)-resistance phenotype. The ODC- or GSH1-overexpressors parasites presented an increase of two and four-fold in SbIII-resistance index, respectively, when compared with the wild-type line. Pharmacological inhibition of ODC and GSH1 with the specific inhibitors α-difluoromethylornithine (DFMO) and buthionine sulfoximine (BSO), respectively, increased the antileishmanial effect of SbIII in all cell lines. However, the ODC- and GSH1-overexpressor were still more resistant to SbIII than the parental cell line. Together, our data shows that modulation of ODC and GSH1 levels and activity is sufficient to affect L. guyanensis susceptibility to SbIII, and confirms a role of these genes in the SbIII-resistance phenotype.

Antizyme inhibitor 1: a potential carcinogenic molecule.

Polyamines are multivalent and organic cations essential for cellular growth, proliferation, differentiation, and apoptosis. Increased levels of polyamines are closely associated with numerous forms of cancer. An autoregulatory circuit composed of ornithine decarboxylase (ODC), antizyme (AZ) and antizyme inhibitor (AZI) govern the intracellular level of polyamines. Antizyme binds with ODC to inhibit ODC activity and to promote the ubiquitin‐independent degradation of ODC. Antizyme inhibitor binds to AZ with a higher affinity than ODC. Consequently, ODC is released from the ODC–AZ complex to rescue its activity. Antizyme inhibitor increases the ODC activity to accelerate the formation of intracellular polyamines, triggering gastric and breast carcinogenesis as well as hepatocellular carcinoma and esophageal squamous cell carcinoma development. Antizyme inhibitor 1 (AZIN1), a primary member of the AZI family, has aroused more attention because of its contribution to cancer. Even though its conformation is changed by adenosine‐to‐inosine (A→I) RNA editing, it plays an important role in tumorigenesis through regulating intracellular polyamines. Encouragingly, AZIN1 has been revealed to have an additional function outside the polyamine pathway so as to bypass the deficiency of targeting the polyamine biosynthetic pathway, promising to become a critical target for cancer therapy. Here, we review the latest research advances into AZIN1 and its potential contribution to carcinogenesis.

IN VITRO ANALYSIS OF ORNITHINE DECARBOXYLASE INHIBITORY ACTIVITY OF THE EXTRACTS OF EXCOECARIA AGALLOCHA L.

Objective: Ornithine decarboxylase (ODC) is one of the key enzymes involved in polyamine synthesis that has a prominent regulatory role in cell growth in prokaryotes and eukaryotes. Its regulation/inhibition is a potential means of controlling cell growth. The purpose of the study was to check the effect of the methanol extracts of leaves and seeds of the mangrove plant Excoecaria agallocha in controlling/inhibiting ODC enzyme produced by Escherichia coli.Methods: The methanol extracts of leaves and seeds of the mangrove plant E. agallocha were made. The inhibitory effect of the extracts on ODC enzyme produced by E. coli was tested by a modified spectrophotometric assay (measured in terms of µM of putrescine produced), and the antibacterial effect of the extract was determined by well diffusion assay.Results: It was found that both the leaf and seed extracts inhibited ODC activity thereby preventing the conversion of ornithine to putrescine (the prominent by-product of ODC activity) in a concentration-dependent manner. The leaf extract exhibited 50% inhibition of ODC activity at a concentration of 17.5 µg/ml whereas seed extract inhibited 50% of the activity at a concentration of 16.1 µg/ml. In the growth inhibition assay by well diffusion method, the effect of the extracts on the growth of E. coli was studied and a concentration-dependent growth inhibition was found - 100 µl of leaf extract showed a zone of inhibition of 12.31 mm (diameter) and seed extract showed 12.64 mm, confirming the antibacterial activity of the extracts.Conclusion: The ODC inhibition-mediated growth inhibition of E. coli is a novel finding as far as the extracts of E. agallocha are concerned. The extract was potent enough to be suggested as an antibacterial agent.Keywords: Ornithine decarboxylase, Excoecaria agallocha, Escherichia coli, Ornithine decarboxylase inhibition, Putrescine, Ornithine, Antibacterial effect.

Elucidation of the polyamine biosynthesis pathway during Brazilian pine (Araucaria angustifolia) seed development.

Polyamines (PAs), such as spermidine and spermine, as well as amino acids that are substrates for their biosynthesis, are known to be essential for plant development. However, little is known about the gene expression and metabolic switches associated with the ornithine/arginine and PA biosynthetic pathway during seed development in conifers. To understand these metabolic switches, the enzyme activity of arginine decarboxylase and ornithine decarboxylase, as well as the contents of PAs and amino acids were evaluated in three Araucaria angustifolia (Bertol. Kuntze) seed developmental stages in combination with expression profile analyses of genes associated with the ornithine/arginine and PA biosynthetic pathway. Twelve genes were selected for further analysis and it was shown that the expression profiles of AaADC and AaSAMDC were up-regulated during zygotic embryo development. Polyamines and amino acids were found to accumulate differently in embryos and megagametophytes, and the transition from the globular to the cotyledonary stage was marked by an increase in free and conjugated spermidine and spermine contents. Putrescine is made from arginine, which was present at low content at the late embryogenesis stage, when high content of citrulline was observed. Differences in amino acids, PAs and gene expression profiles of biosynthetic genes at specific seed stages and at each seed transition stage were investigated, providing insights into molecular and physiological aspects of conifer embryogenesis for use in future both basic and applied studies.

Thyronamines and Derivatives: Physiological Relevance, Pharmacological Actions, and Future Research Directions.

Thyronamines (3-T1AM, T0AM) are endogenous compounds probably derived from L-thyroxine or its intermediate metabolites. Combined activities of intestinal deiodinases and ornithine decarboxylase generate 3-T1AM in vitro. Alternatively, 3-T1AM might be formed by the thyroid gland and secreted into the blood. 3-T1AM and T0AM concentrations have been determined by liquid chromatography-tandem mass spectrometry analysis (LC-MS/MS) in tissues, serum, and cell lines. However, large variations of 3-T1AM concentrations in human serum were reported by LC-MS/MS compared with a monoclonal antibody-based immunoassay. These differences might be caused by strong binding of the highly hydrophobic 3-T1AM to apolipoprotein B100. Pharmacological administration of 3-T1AM results in dose-dependent reversible effects on body temperature, cardiac function, energy metabolism, and neurological functions. The physiological relevance of these actions is unclear, but may occur at tissue concentrations close to the estimated endogenous concentrations of 3-T1AM or its metabolites T0AM or thyroacetic acid (TA1). A number of putative receptors, binding sites, and cellular target molecules mediating actions of the multi-target ligand 3-T1AM have been proposed. Among those are members of the trace amine associated receptor family, the adrenergic receptor ADRα2a, and the thermosensitive transient receptor potential melastatin 8 channel. Preclinical studies employing various animal experimental models are in progress, and more stable receptor-selective agonistic and antagonistic analogues of 3-T1AM are now available for testing. The potent endogenous thyroid hormone-derived biogenic amine 3-T1AM exerts marked cryogenic, metabolic, cardiac and central actions and represents a valuable lead compound linking endocrine, metabolic, and neuroscience research to advance development of new drugs.

Ornithine decarboxylase gene expression and activity in lung cancer

Lung cancer is the malignant tumor of highest mortality rate in men worldwide, and its rate is increasing in women. Ornithine decarboxylase (ODC) is the first enzyme in the pathway of polyamines synthesis and its expression was found increased in many types of tumor tissues. This investigation aimed at measuring ODC activity and gene expression in patients with non-small cell lung cancer (NSCLC). 3 different histopathologic types were included; adenocarcinoma (AC), squamous lung carcinoma (SCC) and large cell lung carcinoma (LCC). The patients were at stages I or II and undergone curative tumor resection. The study revealed that, ODC gene expression and activity were significantly elevated in tumor tissue compared to benign tissue (P<0.0001). In tumor tissues from patients with adenocarcinoma and large cell carcinoma ODC had a higher genetic expression level and activity than in tumor tissues from patients with squamous cell carcinoma (P<0.05). We concluded that ODC may have a clinical and prognostic significance regarding the type and severity of tumor in patients with NSCLC.

Chemomodulatory effect Melastoma Malabathricum Linn against chemically induced renal carcinogenesis rats via attenuation of inflammation, oxidative stress, and early markers of tumor expansion.

Melastoma malabathricum Linn (MM) has high valued for its commercial significance. Indian market (northeast) has great demand for the plants, which extended, its use as a traditional home remedy due to its anti-inflammatory effects. In this study, we scrutinize the therapeutic and protective effect of MM against diethylnitrosamine (DEN) and ferric nitrilotriacetate (Fe-NTA)-induced renal carcinogenesis, renal hyperproliferation, and oxidative stress in rats. Liquid chromatography mass spectroscopy (LC-MS) was used for identification of phytoconstituents. Administration of DEN confirmed the initiation the renal carcinogenesis via enhancing the expansion of tumor incidence. Intraperitoneally, administration of Fe-NTA boost the antioxidant enzymes (phase I), viz., superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx) and phase II, viz., quinone reductase (QR) and glutathione-S-transferase (GST). It also increased the content of renal lipid peroxidation (LPO), hydrogen peroxidase (H2O2) with decrease content in glutathione content (GSH). It also increased the renal biochemical and non-biochemical parameter. It also confirmed the augment the level of thymidine [3H] incorporation into renal DNA, ornithine decarboxylase (ODC) activity and increased the generation of proinflammatory (TNF-α, IL-6 and IL-β) and inflammatory mediator (PGE2). We also analyzed the macroscopic and histologic of renal tissue. In addition, the effect of phytoconstituent of MM extract was evaluated in silico and free radical scavenging activity against the DPPH and ABTS free radicals. LC-MS confirmed the presence of quercetin >gallic acid in MM extract. Renal carcinogenesis rats treated with MM (100, 250, and 500mg/kg) confirmed the significantly (P<0.001) protective effect via reduction the antioxidant (phase I and phase II) enzymes, biochemical parameter and restore the proinflammatory and inflammatory mediator at dose dependent manner. MM altered the ODC and thymidine activity in renal DNA. The chemoprotective effect of MM was confirmed via decreased the renal tumor incidence, which was confirmed by the macroscopic and histopathological observation. Consequently, our result suggests that MM is a potent chemoprotective agent and suppresses DEN+ Fe-NTA-induced renal carcinogenesis, inflammatory reaction, and oxidative stress injury in Wister rats.

Amino acid decarboxylase activities and biogenic amine formation abilities of lactic acid bacteria isolated from shalgam

ABSTRACT56 lactic acid bacteria were isolated during shalgam fermentation and identified as Lactobacillus spp. (51 isolates), Lactococcus spp. (3 isolates), Streptococcus sp. (one isolate), and Leuconostoc sp. (one isolate). 53 of all isolates decarboxylated both arginine and tyrosine, while others decarboxylated one of arginine or tyrosine. None of the isolates could decarboxylate histidine, ornithine, lysine, phenylalanine, or tryptophan. All isolates produced both agmatine (105.8–867.5 mg L–1) and tyramine (24.5–649.7 mg L–1). Although none of the isolates displayed ornithine decarboxylase activity, putrescine was produced (2.1–33.3 mg L–1) by all isolates, except one Lactobacillus strain. Therefore, lactic acid bacteria seem to be responsible mainly for tyramine and agmatine formation during shalgam fermentation, as well as a small amount of putrescine.