Effect Of Azaserine Research Articles

Effects of a Diet High in Fish Oil (MaxEPA) on the Formation of Micronucleated Erythrocytes in Blood and on the Numberof Atypical Acinar Cell Foci Induced in Rat Pancreas by Azaserine

The present study was performed to investigate the influence of fish oil on the genotoxic effects of azaserine, using the formation of micronucleated erythrocytes as a measure for the degree of initiating potency and the number and size of putative preneoplastic pancreatic atypical acinar cell foci (AACF) as a measure for the actual number of initiated cells. Male Wistar rats were treated twice i.p. with 30 mg azaserine per kg body weight to induce AACF. During the initiation/early promotion phase the rats were maintained on diets containing 5 wt% vegetable oil (safflower and high-oleic sunflower oil), 25 wt% vegetable oil, 25 wt% fat (15% vegetable oil + 10 wt% fish oil), or 25 wt% fat (5% vegetable oil + 20 wt% fish oil), respectively. One day after carcinogen treatment, the numbers of micronucleated polychromatic erythrocytes were determined in blood smears obtained from 10 animals per group. Each high-fat diet resulted in higher percentages of micronucleated polychromatic erythrocytes than the low-fat diet. Dietary fish oil did not significantly influence the number of micronucleated cells. Two weeks after carcinogen treatment, the diets containing fish oil were replaced by the diet containing 25% vegetable oil, and the animals were further maintained for about 14 wk. Pancreatic tissue slides were microscopically evaluated for the number and size of AACF. Dietary fish oil caused an increase in the number and size of AACF, although a clear dose-effect relationship was absent. It was concluded that a high level of dietary fish oil, when given during the induction/early promotion phase, enhances azaserine-induced pancreatic carcinogenesis in rats.

Involvement of purine nucleotide synthetic pathways in gonadotropin-induced meiotic maturation in mouse cumulus cell-enclosed oocytes.

This study was carried out to test the hypothesis that purine nucleotide-generating pathways are required for ligand-stimulated oocyte maturation in meiotically arrested cumulus cell-enclosed oocytes. Oocytes from hormonally primed, immature mice were cultured overnight in Eagle's minimum essential medium containing dibutyryl cyclic AMP (dbcAMP) (to maintain meiotic arrest), plus either mycophenolic acid or alanosine (inhibitors of guanyl and adenyl nucleotide production, respectively). Follicle-stimulating hormone (FSH) was added either at the outset of culture or after a 3-hr preincubation period. Under either of these conditions, the inhibitors suppressed FSH induction of germinal vesicle breakdown (GVB). In addition, the potency of FSH as an inducer of GVB was reduced following the 3-hr preincubation period, but this could be prevented if nucleotide precursors such as hypoxanthine, guanosine, or adenosine were included during the first 3 hr. Furthermore, preincubation had little effect on FSH induction of GVB when hypoxanthine was used to maintain meiotic arrest for the entire culture period. The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine, could not mimic this protective effect of hypoxanthine. Azaserine and aminopterin, inhibitors of purine de novo synthesis, blocked hormone-triggered maturation in dbcAMP-arrested oocytes, but had little effect on hypoxanthine-arrested oocytes. The effect of azaserine on dbcAMP-treated oocytes could be reversed by the inclusion of AICA riboside, a compound that can be taken up by cells and phosphorylated to form AICAR, which can enter the purine de novo pathway at a point distal to the sites of azaserine inhibition. FSH was stimulatory to purine de novo synthesis, while azaserine, aminopterin, hypoxanthine, and AICA riboside all suppressed de novo synthesis in the presence or absence of FSH, with dbcAMP having no effect. HPLC analysis of 14C-hypoxanthine metabolism in oocyte-cumulus cell complexes revealed that changes in the pattern of purine metabolism did not mediate the meiosis-inducing effect of FSH. These data support the conclusion that purine nucleotide-generating pathways are vital participants in the mechanism(s) regulating hormone-induced meiotic maturation, and that either the de novo or salvage pathway can fulfill this nucleotide requirement.

Metabolism of 15N-labelled ammonium by the ectomycorrhizal fungus Pisolithus tinctorim (Pers.) Coker & Couch

Glutamine was the major product accumulated following transfer of nitrogen-limited cultures of the ectomycorrhizal fungus Pisolithus tinctorius to an ammonium medium. Experiments in which mycelium was transferred to [15N]H4+showed glutamine amide was the most heavily labelled product. Assimilation of ammonium into glutamate was markedly inhibited by azaserine. The kinetics of 15N-labelling and the effects of azaserine and methionine sulphoximine on the distribution of 15N-labelled products are entirely consistent with the operation of the glutamate synthase cycle. No evidence was found for ammonium assimilation via glutamate dehydrogenase. The labelling pattern observed in mycelium treated with aminooxyacetate suggests that transamination reactions are an important source of glutamate for the synthesis of glutamine.

Effect of the Glutamine-Analogue "Azaserine" on Embryonic Development of the Horseshoe Crab: (azaserine/inhibition of cell growth/morphogenesis/separation of embryonic area/horseshoe crab).

Embryos of the horseshoe crab were treated with the glutamine-analogue "azaserine". This compound induced malformed embryos at the stage of enlargement of the germ disc. The embryonic areas of these monsters were separated into two pieces. This is the first report of monsters induced with azaserine. The effect of azaserine was counteracted by glutamine. Azaserine repressed cell growth of the embryos, and prevented development of their germ disc. During active morphogenic movement, the embryonic area of treated embryos was separated in the central region of elongation of the embryonic area.

PROCEEDINGS OF THE AUSTRALIAN SOCIETY FOR MEDICAL RESEARCH

PROCEEDINGS OF THE AUSTRALIAN SOCIETY FOR MEDICAL RESEARCH

The effect of azaserine on glutamine uptake by rat renal brush-border membranes.

Azaserine added directly to isolated rat renal brush-border membrane vesicles inhibits uptake of L-glutamine. Azaserine acts as a non-competitive inhibitor of the low-Km system for glutamine transport, but has no effect on the high-Km system. Preincubation of the vesicles with azaserine at 37 degrees C min is not required for transport inhibition to occur, although it is a requirement for gamma-glutamyl transpeptidase inhibition. Removal of azaserine from the vesicle preparation by repeated resuspensions in buffer results in a reversal of the transport inhibition but not in reversal of enzyme inhibition. Azaserine also inhibits vesicle uptake of L-proline and alpha-methyl D-glucoside, indicating a generalized effect on membrane transport systems. The data cast doubt on the postulate that gamma-glutamyl transpeptidase might act as the carrier mechanism for glutamine reabsorption by renal cortical cells.

Production of pancreatic acinar cell carcinoma by combined administration of 4-hydroxyaminoquinoline 1-oxide and azaserine in partial pancreatectomized rats

The effect of azaserine on the pancreatic tumorigenesis of 4-hydroxyaminoquinoline 1-oxide (4-HAQO) after partial pancreatectomy in rats was studied. Pancreatic acinar cell carcinomas were produced in 7 out of 10 rats (70%), which received 7 mg/kg body wt. 4-HAQO 3 days after partial pancreatectomy, followed by 10 weekly injections of 30 mg/kg body wt. azaserine. Partial pancreatectomy enhanced the carcinogenesis of 4-HAQO, which was further promoted by azaserine.

Regulation of ATP pools, rRNA and DNA synthesis in 3T3 cells in response to serum or hypoxanthine.

The serum-induced transition of 3T3 fibroblasts from resting to growing state is characterized by a marked increase in cellular ATP content, the maximal level of which is reached at the onset of DNA replication. This increase in cellular ATP during the G 1 period of the cell cycle is correlated with about 3-fold stimulation of transcription of rRNA measured in permeabilized cell in vitro. Addition of hypoxanthine to serum-depleted quiescent 3T3 cells gives rise to an increase in both the ATP pool and the rate of rRNA synthesis. The expansion of cellular ATP pools after growth induction by serum seems to be a prerequisite for initiation of DNA synthesis since inhibition of purine de novo biosynthesis by azaserine inhibits both ATP pool expansion and DNA replication. This effect of azaserine can be abolished by addition of hypoxanthine to the culture medium. It is concluded that (a) the increase of the rate of rRNA synthesis in 3T3 cells in response to growth factors or serum is controlled by the cellular purine nucleoside triphosphate concentration and (b) an increased ATP level is necessary for initiation of DNA synthesis but is not sufficient to trigger the events that lead to DNA replication.

Effect of azaserine on insulin in release in vitro.

Effect of azaserine on insulin in release in vitro.

Studies on azaserine-induced fatty liver in the rat.

SummaryThe effect of azaserine on liver lipid composition, on lipogenesis in vitro in liver slices and on serum lipoproteins, on liver content of ATP, biotin, and nicotinamide nucleotides in the rat is reported. The effect of ATP or adenine in azaserine-treated rats is also presented. The neutral fats in the liver of azaserine-treated rats significantly increased, while the incorporation in vitro of acetate-14C into total lipids decreased. In the same experimental conditions the liver concentration of biotin and nicotinamide nucleotides fell. A marked decrease of the levels of serum lipoprotein also was observed. The administration of ATP or adenine to azaserine-treated rats did not prevent either the development of fatty liver, the decrease of liver biotin and of serum lipoprotein, or the decrease of acetate-14C incorporation into liver lipids. On the contrary, the administration of ATP or adenine partially reversed the azaserine effects on the liver content of nicotinamide nucleotides and increased the ...

Inhibition of J-128 (Osgood) cell culture by azaserine.

SummaryAdenine, hypoxanthine or AICA will not reverse the inhibitory effect of aza-serine on the reproduction of J-128 cells in tissue cultures. Cells exposed to azaserine increase markedly in size with a parallel increase of nuclei size and DNA content. The drug appears to act quite similarly to certain mustards by stopping cell division between S and G2. A lesion appears in the central sphere of attraction in azaserine-treated cells, increases in size and apparently causes death of the cell.

Nicotinamide adenine dinuleotides in acute liver injury: Effects of azaserine and puromycin in the rat

The effects of single doses of azaserine or puromycin on the nicotinamide adenine dinucleotides (NADs) in rat liver have been studied. Azaserine produced a very rapid decrease in NAD + NADH 2 such that 30 min after dosing the sum was only 36 per cent of the control value. The sum NADP + NADPH 2 was not affected until considerably later in the injury process. Since NADP is synthesised via NAD, the relative rate of decrease of NADP + NADPH 2 compared to NAD + NADH 2 provides some indication of the turnover rate of NADP in the affected liver. The estimate for the “turnover time” of NADP in rat liver was 5 hr. Puromycin had no significant effect on the levels of NAD, NADH 2, NADP and NADPH 2 in rat liver although the dose used is known to inhibit protein synthesis almost completely in vivo. It is concluded that enzymes concerned in the biosynthesis of NAD and NADP do not “turn-over” so rapidly that suppression of their biosynthesis for a 1 hr period significantly affects the overall content of nicotinamide adenine dinucleotides in the liver.

Effect of azaserine on the incorporation of 14C-labeled purines and pyrimidines into the acid-soluble and nucleic acid fractions of Trypanosoma equiperdum

Azaserine decreased the incorporation of 14C-labeled adenine, guanine, hypoxanthine, uracil, and orotic acid into the nucleic acid fraction and increased the incorporation of these compounds into the acid-soluble fraction of Trypanosoma equiperdum incubated in vitro. The amino acids l-glutamine, l-tryptophan, l-leucine, and l-tyrosine partially prevented the inhibition produced by azaserine on the incorporation of ade- nine- 14C into the nucleic acid fraction. Azaserine did not interfere with the incorporation of adenine- 14C into the acid-soluble adenine- and guanine-containing compounds but did inhibit the incorporation of uracil- 14C into acid-soluble cytosine-containing nucleotides and the UMP and CMP components of RNA. These results suggest that in T. equiperdum azaserine inhibits the conversion of UMP to CMP, and as a result of this inhibition there is a secondary inhibition of the biosynthesis of RNA. The extremely limited ability of T. equiperdum to anabolize tritiated cytosine or cytidine suggests that this protozoan species depends primarily upon the de novo pathway for the biosynthesis of CMP and indicates a possible site for exploitation with the appropriate antimetabolites.

Effects of purine antagonists on the metabolism of other purine antagonists

The effects of azaserine, thioguanine and 6-mercaptopurine on the metabolism of 4-aminopyrazolopyrimidine- 14C and of 4-aminopyrazolopyrimidine on the metabolism of thioguanine- 14C and of 6-mercaptopurine- 14C have been studied in Ehrlich ascites carcinoma and ascites sarcoma-180. Azaserine, thioguanine and 6-mercaptopurine not only enhanced the inhibition of tumor growth by 4-aminopyrazolopyrimidine, but also increased the amount of incorporation of the analog into tumor polynucleotides. Enhancement of the size of the pool of 4-aminopyrazolopyrimidine mononucloetides was evident during dual treatment with azaserine and thioguanine. The amount of incorporation of radioactivity from labeled 6-mercaptopurine into polynucleotides was decreased by 4-aminopyrazolopyrimidine, while little change in the size of the labeled mononucleotide pool was evident. The therapeutic effectiveness of thioguanine was increased by the co-administration of 4-aminopyrazolopyrimidine and the amount of incorporation of thioguanine into nucleic acids was somewhat increased by 4-aminopyrazolopyrimidine. These results are discussed with respect to present hypotheses concerning the anabolic conversion of purine analogs into their active derivatives.

Effect of Azaserine on the Biosynthesis of Diphosphopyridine Nucleotide in Mouse

Effect of Azaserine on the Biosynthesis of Diphosphopyridine Nucleotide in Mouse

THE LYMPHOMAS: COMBINED CLINIC AT THE COLLEGE OF PHYSICIANS AND SURGEONS, COLUMBIA UNIVERSITY, NEW YORK, N. Y.

Excerpt Dr. Alfred Gellhorn: Today we discuss a group of diseases, the lymphomas, various aspects of which are being productively investigated in many parts of the world. During our presentation we...

Effect of azaserine (o-diazoacetyl-L-serine) on the pyridine nucleotide levels of mouse liver.

THE amount of diphosphopyridine nucleotide in mouse liver has been shown to increase upon the intraperitoneal injection of a number of pyridine derivatives1,2; nicotinamide is the most effective, raising the level 8–10 times. Although nicotinic acid is as effective as many of the analogues, it appears to inhibit the synthesis when administered with nicotinamide, as compared with nicotinamide alone. Inhibition of synthesis from nicotinamide injection has also been reported with the administration of the purine antagonists 6-mercaptopurine and thioguanine1,3. These inhibitions appear to affect only the level of diphosphopyridine nucleotide attained from nicotinamide injection and the rate of its subsequent return to the normal level, but do not appear to affect the basal level normally found in the liver.

The Effect of Azaserine against Plasmodium lophurae in Chicks and Attempts to Antagonize This Effect with Metabolites

N2CH-C-O-CH2-CH-COOH I I I 0 NH2 O-diazoacetyl-1-serine It is an amino acid analog, has antitumor activity (Sugiura and Stock, 1955), acts as an antimetabolite with certain microorganisms in vitro (Kaplan and Stock, 1954), and has suppressive activity in avian malaria (Coffey et al, 1954). These facts encouraged further study of the antimalarial activity of azaserine and suggested that the testing of various metabolites for ability to antagonize its activity in vivo might yield useful information on the physiology of the plasmodia and biological activity of the drug. The results of such work are reported here.

Amoebicidal action of azaserine.

IN 1954, a group of workers reported on a new tumour-inhibitory substance1. This substance, called azaserine, also possessed antibacterial activity against numerous microbial species2. Chemically, this substance is O-diazoacetyl-L-serine3. It has been reported that azaserine is a profound inhibitor of formate incorporation into the nucleic acids4. Since the role of nucleic acid synthesis is particularly important in Endamoeba histolytica 5,6 the effect of azaserine on the amœbae was studied.

Studies on the mode of action of azaserine

1. 1. Azaserine-induced inhibition of E. coli was significantly prevented by ( a) 4-amino-5-imidazolecarboxamide (AIC), adenine, guanine, hypoxanthine, or xanthine; ( b) methionine; and ( c) glutamine; and, as has already been observed by others, by phenylalanine, tyrosine, or tryptophan. 2. 2. In studies carried out on the effect of azaserine on purine synthesis in the tumors, intestines, livers, and spleens of mice bearing Sarcoma 180, it was found that azaserine inhibited utilization of formate-C 14 and glycine-1-C 14 for purine synthesis, failed to affect utilization of C 14-labeled adenine, AIC, and hypoxanthine, and failed to affect utilization of formate for thymine biosynthesis. Azaserine also inhibited utilization of formate-C 14 and glycine-1-C 14 for purine synthesis by heterologous implants of a human sarcoma growing in the rat. 3. 3. The results indicate that azaserine inhibits purine synthesis in the intact animal at a stage prior to the formation of 5-amino-4-imidazole-carboxamide ribotide.