Putrescine In Liver Research Articles

Association between remote organ injury and tissue polyamine homeostasis in acute experimental pancreatitis – treatment with a polyamine analogue bismethylspermine

Experimental pancreatitis is associated with activation of polyamine catabolism. The polyamine analog bismethylspermine (Me2Spm) can ameliorate pancreatic injury. We investigated the roles of polyamine catabolism in remote organs during pancreatitis and explored the mechanism of polyamine catabolism by administering Me2Spm. Acute pancreatitis was induced by an infusion of 2 or 6% taurodeoxycholate before Me2Spm administration. Blood, urine and tissues were sampled at 24 and 72h to assess multi-organ injury and polyamine catabolism. The effect of Me2Spm on mortality in experimental pancreatitis was tested separately. Liver putrescine levels were elevated following liver injury. Me2Spm increased the activity of spermidine/spermine N1-acetyltransferase (SSAT) and depleted the spermidine, spermine or putrescine levels. Lung putrescine levels increased, and SSAT and spermine decreased following lung injury. Me2Spm enhanced the activity of SSAT and decreased the spermidine and spermine levels. Renal injury was manifested as an increase in creatinine or a decrease in urine output. Decreases in kidney SSAT, spermidine or spermine and an increase in putrescine were found during pancreatitis. In the 2% taurodeoxycholate model, Me2Spm decreased urine output and raised plasma creatinine levels. Me2Spm increased SSAT and decreased polyamines. Excessive Me2Spm accumulated in the kidney, and greater amounts were found in the 6% taurodeoxycholate model in which this mortality was not reduced by Me2Spm. In the 2% taurodeoxycholate model, Me2Spm dose-dependently induced mortality at 72h.Like pancreatic injury, remote organ injury in pancreatitis is associated with increased putrescine levels. However, Me2Spm could not ameliorate multi-organ injury. Me2Spm administration was associated with significant renal toxicity and induced mortality, suggesting that the current dose is too high and needs to be modified.

Effect of inhibition of ornithine decarboxylase activity in a model of acute hepatocellular necrosis.

The effect of blockade of the enzyme ornithine decarboxylase by difluoromethylornithine (DFMO) on hepatocellular necrosis and survival in rats treated with thioacetamide (TAA) was investigated. In one experiment, the effect of DFMO on survival of rats with TAA-induced acute hepatocellular necrosis was determined. In another experiment, blood and liver specimens were obtained from DFMO or saline-treated rats 24 h after the administration of TAA for determinations of serum alanine aminotransferase (ALT) and liver content of polyamines and microsomal cytochrome P-450 and for assessment of hepatic histology. Liver polyamines were determined by reversed-phase HPLC and microsomal cytochrome P-450 content by dithionite-difference spectroscopy of CO-treated homogenates. The severity of hepatocellular necrosis was scored blindly. TAA-treated rats that received DFMO survived longer than saline-treated controls (P < 0.01). Serum ALT and liver putrescine concentrations were lower and the histological severity of acute hepatocellular necrosis was less in DFMO-treated rats with TAA-induced hepatocellular necrosis than in saline-treated controls (P < 0.05, P < 0.01 and P < 0.05, respectively). Total cytochrome P-450 levels were similar in DFMO and saline-treated rats with TAA-induced hepatocellular necrosis. DFMO increases survival in TAA-induced fulminant hepatic failure by decreasing the severity of acute hepatocellular necrosis. The beneficial effects of DFMO do not appear to be mediated by its effects on polyamine metabolism, but may be attributable to an effect of DFMO on thioacetamide metabolism or on an alternative pathway of ornithine metabolism.

Transgenic mice expressing the human ornithine decarboxylase gene under the control of mouse metallothionein I promoter

We have generated a transgenic mouse line harbouring the human ornithine decarboxylase gene under the control of mouse metallothionein I promoter. Even in the absence of an exposure to heavy metals, ornithine decarboxylase was over-expressed in heart, testis, brain, and especially in liver, of the transgenic animals. An exposure of the transgenic mice to zinc further enhanced the enzyme activity to a level which in liver represented up to 8000-fold increase in comparison with non-transgenic animals. The striking stimulation of liver ornithine decarboxylase activity upon treatment of the transgenic mice with zinc was accompanied by a nearly 150-fold increase in the hepatic putrescine content as compared with similarly treated non-transgenic animals. Even though the liver putrescine concentration reached that of spermidine and spermine in the transgenic animals, the contents of the higher polyamines only transiently increased upon zinc administration and then returned to the basal level. These findings once again indicate that mammalian cells possess extremely powerful regulatory machinery to prevent an over-accumulation of spermidine and spermine in non-dividing cells, and that very high tissue putrescine concentrations can be tolerated, at least for periods of a few days, with seemingly no phenotypic changes.

Daily cycles of putrescine, sperrnidine, and spermine in the liver, pineal gland, Harderian gland, anterior pituitary, and testes of rats kept in LD 12:12

A circadian rhythm in cellular polyamine levels was detected in the liver, pineal gland, anterior pituitary gland, Harderian gland, and testicular seminiferous tubules of male rats fed ad libitum and maintained in a light:dark cycle of LD 12:12 (lights on at 07:00). Liver putrescine content was highest at 24:00, showing a sixfold increase over 12:00 levels. Pineal spermidine and spermine contents reached a maximum at 06:00, late in the dark phase. A similar pattern was also detected in the Harderian gland. In the anterior pituitary, the polyamines putrescine, spermidine and spermine were highest at 18:00, late in the light phase. However, the increase in putrescine was not statistically significant. The three polyamine contents decreased late in the dark phase. In testicular seminiferous tubules putrescine, on the contrary, was highest (about a twofold increase) late in the dark phase.

Induction of spermidine/spermine N1-acetyltransferase in rat tissues by polyamines.

Treatment of rats with spermidine, spermine or sym-norspermidine led to a substantial induction of spermidine/spermine N1-acetyltransferase activity in liver, kidney and lung. The increase in this enzyme, which was determined independently of other acetylases by using a specific antiserum, accounted for all of the increased acetylase activity in extracts from rats treated with these polyamines. Spermine was the most active inducer, and the greatest effect was seen in liver. Liver spermidine/spermine N1-acetyltransferase activity was increased about 300-fold within 6 h of treatment with 0.3 mmol/kg doses of spermine; activity in kidney increased 30-fold and activity in the lung 15-fold under these conditions. The increased spermidine/spermine N1-acetyltransferase activity led to a large increase in the liver putrescine content and a decline in spermidine. These changes are due to the oxidation by polyamine oxidase of the N1-acetylspermidine formed by the acetyltransferase. Our results indicated that spermidine was the preferred substrate in vivo of the acetylase/oxidase pathway for the conversion of the higher polyamines into putrescine. The induction of the spermidine/spermine N1-acetyltransferase by polyamines may provide a mechanism by which excess polyamines can be removed.

Relation between induction of rat hepatic ornithine decarboxylase activity by tumor promoters 12-O-tetradecanoylphorbol-13-acetate and phenobarbital and levels of the polyamines putrescine, spermidine and spermine, in vivo; differential effects of retinyl-acetate.

A single i.p. injection of 12-O-tetradecanoylphorbol-13-acetate (TPA) induced a transient increase in the levels of rat liver putrescine, spermidine and spermine. These polyamine concentrations continuously increased until 6 h, immediately following administration of the tumor promoter. Phenobarbital (PB) induced an increase of the putrescine and spermidine concentrations during the 8 h post administration studied, while spermine reached a plateau after 4 h. When retinyl-acetate (RA) was injected one hour prior to TPA, the increases of the polyamines were considerably inhibited. This treatment with RA partially inhibited further increase of putrescine and spermine by PB. These findings are discussed in respect to the concomitant ornithine decarboxylase (ODC) activities, we have previously reported. I.p. injection of RA alone caused an elevation of ODC activity as well as putrescine and spermidine concentration within 2 h of exposure.

Comparison of 3-isobutyl-1-methylxanthine-induced accumulation of putrescine in rat pancreas and liver

3-Isobutylmethylxanthine (IBMX), a potent phosphodiesterase inhibitor, causes accumulation of putrescine of same magnitude in rat pancreas and liver. IBMX produces increases of acetyl CoA : polymine N′-acetyltransferase (PAT) and of ornithine decarboxylase (ODC) activities in both organs. However ODC activity is 300 times higher in liver than in pancreas. In the latter organ, there is a transient increase of N 1-acetylspermidine, followed by a decrease of spermidine. α-Difluoromethylornithine (DFMO), a potent ODC inhibitor, impairs the accumulation of putrescine in liver but not in pancreas. These results suggest that in pancreas the accumulated putrescine is essentially formed from spermidine, via N 1-acetylation and oxidation, while in liver it is formed from decarboxylation of ornithine. A possible involvement of cAMP in the stimulation of the polyamine interconversion pathway is discussed.

Conversion of exogenous spermidine into putrescine after administration to rats.

Administration of large, but non-toxic doses of spermidine (0.4-1.25 mmol/kg) led to a substantial increase in putrescine in liver, kidney and a number of other tissues including muscle. The increase in putrescine peaked at 6 h after treatment and was completely prevented by administration of cycloheximide 3 h after the spermidine suggesting that the induction of a new protein was required. This protein is likely to be spermidine N1-acetyltransferase which was induced by the treatment with spermidine and increased 3-4-fold in liver and kidney within 6 h. N1-Acetylspermidine was detected in tissues at this time after spermidine treatment and experiments in which labeled spermidine was given indicated that a substantial fraction of the administered spermidine was converted into N1-acetylspermidine and into putrescine. These results suggest that the rise in putrescine after spermidine treatment is brought about by the production of N1-acetylspermidine which is converted into putrescine by the action of polyamine oxidase. The limiting step in this conversion is the activity of the acetylase which is induced in response to the rise in spermidine content. The acetylase/oxidase pathway, therefore, provides a means by which polyamine levels can be regulated and excess polyamine disposed of.

Changes in hepatic polyamine levels during acute and chronic administration of aflatoxin B 1 to rats

A subacute dose of aflatoxin B 1 (3 mg/kg body weight) increases liver putrescine levels within 1 hr after administration, with high levels persisting over 24 hr. Higher doses of the carcinogen elicited larger increases in liver polyamine levels. A marked elevation of putrescine, spermidine and spermine were noted in aflatoxin B 1-induced preneoplastic liver. Pretreatment of rats with phenobarbital prior to aflatoxin B 1 administration resulted in no synergistic or additive effects.

Effects of L-DOPA on putrescine levels in the brain and the liver of the rat

The pharmacological effects of several amino acid precursors of putative neurotransmitters on putrescine levels in the brain and the liver of the rat were studied. L-DOPA increased brain and liver putrescine levels in a dose-dependent manner that reached its maximum effect in 4–6 h. The increase in liver putrescine was associated with a concomitant increase in ornithine decarboxylase activity. L-5-hydroxytryptophan increased liver putrescine but had no effect on brain putrescine levels. D-DOPA and D-5-hydroxytryptophan were both ineffective in altering brain or liver putrescine. The effects of L-DOPA persisted after hypophysectomy and were not associated with changes in tissue levels of S-adenosylmethionine. The repeated administration of L-DOPA for periods of 48 and 96 h resulted in a sustained elevation of putrescine levels in the brain but not in the liver.

Inhibition of polyamine accumulation and deoxyribonucleic acid synthesis in regenerating rat liver

Repeated injections of 1,3-diaminopropane into rats after partial hepatectomy caused a repression-type inhibiton of liver ornithine decarboxylase (EC 4.1.1.17) and totally prevented the marked increases in liver putrescine and spermidine concentrations that normally occur in response to partial hepatectomy. The inhibition of polyamine synthesis by diaminopropane was accompanied by a profound decrease (about 80%) in the synthesis of DNA in the regenerating rat liver without any changes in the synthesis of RNA and total liver protein.

Synthesis and accumulation of polyamines in rat liver regenerating after treatment with carbon tetrachloride

A single intraperitoneal injection of carbon tetrachloride into rats resulted within 12 hours in a marked accumulation of putrescine in liver with a concomitant decrease in the concentration of spermidine. The accumulation of putrescine apparently was partly due to an immense stimulation of ornithine decarboxylase activity occurring at the same time. However, in addition it was found that during the maximal accumulation of putrescine there was a marked incorporation of radioactivity from labelled spermidine to liver putrescine in vivo . The conversion of spermidine to liver putrescine was hardly detectable in control animals. Besides the treatment with carbon tetrachloride, increased conversion of radioactive spermidine to liver putrescine in vivo also occurred after treatment with growth hormone, after partial hepatectomy and after treatment with thioacetamide, i. e. under circumstances characterized by a stimulation of ornithine decarboxylase activity and an increased accumulation of putrescine.

Ornithine decarboxylase activity and the accumulation of putrescine at early stages of liver regeneration

intensive stimulation of ornithine decarboxylase (EC 4.1 .1.17, L-ornithine carboxylase) activity after partial hepatectomy of the rat belongs to the earliest biochemical changes in the regenerating liver remnant [l-4] . Enhanced ornithine decarboxylase activity results in a concomitant accumulation of liver putrescine [ 1,5,6] which in turn is supposed to stimulate the synthesis of spermidine [l] probably by activating the decarboxylation of S-adenosyl-L- methionine (Ado-met), the reaction needed for the synthesis of spermidine [7] . The activity of ornithine decarboxylase is almost maximally stimulated as early as at about 4 hr after partial hepatectomy and remains elevated for several days [2,6]. It has been reported that hypophysectomy, but not adrenalectomy, castration, or thyroidectomy, considerably delays, although does not prevent, the stimulation of ornithine decarboxylase after partial hepatectomy [8] . This latter observation might indicate the involvement of some humoral factor(s) in the regulation of putrescine synthesis in the regenerat- ing rat liver. In the present communication we have studied more closely the synthesis and accumulation of putrescine at early stages of liver regeneration. The results of several series of rats revealed that during the first day of regeneration the stimulation of ornithine decarboxylase appeared to occur in two phases, the first peak of the enzyme activity invariably occurring at 4 hr after the operation independently of the age of the animal. The stimulation of ornithine decarboxyl- ase was closely followed by an accumulation of putrescine. Evidence is also presented indicating that