Effects of hormones on polyamine biosynthesis in cultured sheep adipose tissue slices.
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Adenosylmethionine Decarboxylase↗
Biomedical subjects
A M Snoswell
Publications and source records attributed to A M Snoswell.
Novel effects of growth hormone on polyamine biosynthesis in sheep adipose tissue.
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Adipose Tissue↗
An animal model of systemic carnitine deficiency produced by haemodialysis of sheep.
1. Sheep, which had previously been surgically prepared with cannulae in various vessels to monitor substrate and metabolite exchanges across all the major organs, were connected to a haemodialysis machine and their blood was dialysed at an average rate of 6.23 ml/min/kg body weight. 2. Dialysis for 4 hr reduced the blood free carnitine concentrations to approx. 50% of the initial values and the concentrations returned to the initial values after 18 hr recovery. 3. Carnitine balance studies showed that approx. twice the amount of carnitine lost from the blood during dialysis passed into the dialysate indicating that carnitine was also lost from the extracellular fluid. 4. The average blood concentration of short-chain acylcarnitines did not vary significantly during dialysis or during the recovery phase. However, an output of short-chain acylcarnitines by the liver at 3 and 18 hr recovery and an uptake by the hind-body at 18 hr recovery was observed. 5. These results suggest that haemodialysis of sheep provides a useful model of systemic carnitine deficiency and suggest that treatment with acetylcarnitine or propionylcarnitine could be an efficient means of supplying carnitine in carnitine replacement therapy.
Animals↗
Partial purification and properties of the isozymes of S-adenosylmethionine synthetase from sheep liver.
Three forms of AdoMet synthetase were separated from sheep liver. The apparent molecular weights of the native isozymes were 122,000, 62,400 and 70,800 for the alpha-, beta 1- and beta 2-form, respectively and beta 1 was the predominant form. The alpha-form exhibited negative cooperativity with [S] 0.5 values of 31 microM for methionine and 62 microM for ATP; while the two beta-forms exhibited positive cooperativity with [S]0.5 values for methionine of 82 microM and 70 microM and those for ATP of 572 microM and 505 microM for the beta 1- and beta 2-form, respectively. Dimethylsulfoxide markedly stimulated the activities of the two beta-forms at low methionine concentrations. However, at high methionine levels, it inhibited the activity of the beta 2-form but not that of the beta 1-form. The effect of dimethylsulfoxide on the alpha-form was not significant. AdoMet was inhibitory at high concentrations. However, it had a slight stimulatory effect on the two beta-forms at low concentrations when methionine level was also low. These results suggest that AdoMet synthetase is a regulatory enzyme and the reaction rate in vivo can be directly influenced by substrate and product concentrations.
Animals↗
Determination of creatinine in whole blood, plasma, and urine by high-performance liquid chromatography.
A sensitive method for the specific determination of creatinine in whole blood, plasma, and urine with high precision and accuracy is described. Samples were deproteinized by addition of acetonitrile and analyzed by high-performance liquid chromatography using a cation-exchange column with a mobile phase of 9% acetonitrile in 40 mM ammonium phosphate (pH 4.0). The recoveries of creatinine added to blood and plasma were almost complete, ranging from 99 to 101%. The coefficients of variation were very small, 1.6% for blood and plasma and 1.5% for urine. Samples can be assayed in 11-min intervals subsequent to the initial injection. As little as 2 microliter of blood or plasma or 0.02 microliter of urine is sufficient for chromatographic analysis. The present method was successfully used for the accurate measurement of small arterial-venous differences of creatinine concentrations in blood across body organs and showed that in the sheep creatinine is produced in the muscles and is excreted by the kidneys. The method is also suitable for routine analysis of creatinine in clinical laboratories.
Animals↗
Quantitative study on creatine metabolism in sheep tissues.
Creatine metabolism was quantitated in sheep tissues by measurement of the amount of creatine and creatinine flowing into and out of various tissues using multi-catheterized sheep. The results showed that sheep derived no creatine from the digestive tract. The total renal clearance of creatine plus creatinine was about 18 mmoles per day. Daily creatine synthesis in the sheep was some 15 mmoles, of which 80% occurred in the liver. There was a daily uptake of about 5.4 mmoles of creatine and an output of 7.8 mmoles of creatinine by the hindlimbs of the sheep. This difference suggests that the skeletal muscle may be able to synthesize a significant amount of creatine. However, creatine metabolism in the heart, lung and brain was much less active.
Animals↗
Choline biosynthesis in sheep. Evidence for extrahepatic synthesis.
1. Choline production by various tissues of the sheep was measured by determining venous and arterial free choline concentrations in blood samples taken from various vessels in conscious multicannulated sheep. 2. Significant production of free choline occurred in the upper and lower body regions, and specifically in the heart, brain and hind-limb muscles of sheep, but there was no significant uptake or output of phosphatidylcholine across these tissues, as determined by arterio-venous differences. 3. In contrast, in the rat there were no significant arterio-venous differences in the concentrations of free choline or phosphatidylcholine across the hind-body. 4. Synthesis of phosphatidylcholine from endogenous phosphatidylethanolamine and S-adenosyl-L-[methyl-14C]methionine was measured in experiments in vitro using microsomal preparations from a variety of sheep and rat tissues. 5. The biosynthetic activity was highest in liver from sheep and rats, although the activity in sheep microsomal preparations was about one-quarter of that in rat microsomal preparations. 6. Microsomal preparations from sheep lung, kidney, gut epithelium, brain, heart and skeletal muscles also showed considerable biosynthetic activity, but in the rat the activity was virtually confined to the liver. 7. Overall, the results show a significant production of choline in extrahepatic tissues of the sheep, with skeletal muscle contributing some 60% of this extrahepatic activity. Thus the extrahepatic production of choline in the sheep, together with the extensive reutilization of bile choline, can explain the maintenance of the large endogenous body pool of choline in this species.
Animals↗
The enterohepatic recycling of bile choline in sheep.
1. Measurement of unesterified choline in blood samples taken from five conscious multi-cannulated sheep indicated a significant production of unesterified choline by the alimentary tract, as judged by the portal venous minus arterial difference and significant uptake by the liver, as judged from the portal venous minus hepatic venous and arterial minus hepatic venous differences. 2. A mean liver blood flow rate of 1.68 +/- 0.22 1/min for the five sheep was determined by bromosulphophthalein clearance and, combined with the differences in unesterified choline across organs, gave a production rate of free choline of 9.1 mmol/day by the alimentary tract and an uptake by the liver of 13.2 mmol/day. 3. Infusion of [methyl-3H]choline chloride into the portal vein of a sheep over 1 hr and subsequent isolation of the bile for several days showed over 70% cumulative recovery of the radioactivity in the choline moiety of bile phosphatidylcholine over a 120 hr period. 4. Subsequent infusion 17 days later of bile lipid [3H]choline via a duodenal fistula also gave approx. 70% cumulative recovery of radioactivity in the choline moiety of newly secreted bile phosphatidylcholine in 120 hr. 5. These results show a very extensive enterohepatic recirculation of bile choline in the sheep, which is in contrast to the situation in monogastric animals.
Animals↗
Methyl group metabolism in sheep.
1. Sheep have a very low intake of methyl nutrients in the post-ruminant state, due to the almost complete degradation of dietary choline by rumen microorganisms, the lack of dietary creatine and the relatively low content of methionine in microbial proteins. 2. Methylneogenesis provides a major source of labile methyl groups in post-ruminant sheep and impairment of the methylneogenesis leads to a marked reduction of the labile methyl pool. 3. S-Adenosylmethionine (AdoMet) metabolism via transmethylation is most active in sheep liver and pancreas and is regulated by the availability of methionine and intracellular ratios of AdoMet to S-adenosylhomocysteine (AdoHcy). 4. Adaptive mechanisms which arise as a consequence of the poor methyl nutrition in post-ruminant sheep are a marked reduction of labile methyl catabolism and an increase in the capacity of methylneogenesis.
Adaptation, Physiological↗
Influence of dietary cholesterol on desaturase enzymes of rat liver microsomes.
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Animals↗
Quantitative evaluation and regulation of S-adenosylmethionine-dependent transmethylation in sheep tissues.
The overall rates of S-adenosylmethionine (AdoMet)-dependent transmethylation were estimated in various tissues from the initial rate of S-adenosylhomocysteine (AdoHcy) plus AdoMet accumulation after blocking hydrolysis of AdoHcy. The rates were found to differ widely among the tissues of sheep and the highest rate was in the pancreas, being 600 times higher than that in the muscle. Sheep liver possessed approximately 75% of total-body capacity for transmethylation although the transmethylation rate was approximately half that in rat liver. The minimum estimate of daily requirement of AdoMet for transmethylation for adult sheep was approximately 18 mmol, far in excess of methionine intake. Methionine loading elevated AdoMet levels only in the tissues with a high or moderate rate of transmethylation. The kinetic properties of major methyltransferases in sheep liver along with tissue distribution of AdoMet and AdoHcy suggest that transmethylation rate is subject to physiological regulation by tissue levels of AdoMet and AdoHcy.
Animals↗
Developmental changes in the activities of enzymes related to methyl group metabolism in sheep tissues.
The activities of choline oxidase and betaine-homocysteine methyltransferase increased markedly in pre-ruminant lamb liver after birth and subsequently decreased when the lambs reached the ruminant state, while the developmental changes in hepatic 5-methyl-H4folate-homocysteine methyltransferase were negatively correlated with those of betaine-homocysteine methyltransferase. Hepatic phospholipid methyltransferase was elevated almost four-fold by the 10th postnatal day, but declined thereafter. Hepatic glycine methyltransferase in one-day-old lambs increased 55-fold, compared with that of fetuses, and thereafter decreased dramatically with age. Guanidoacetate methyltransferase, glycine methyltransferase and betaine-homocysteine methyltransferase in sheep pancreas increased markedly with age and were many times higher than the hepatic enzymes in adult sheep. Choline oxidase, betaine-homocysteine methyltransferase, cystathionine beta-synthase and glycine methyltransferase in adult sheep liver were much lower than those in rat. These results illustrate the conservative features of methyl group metabolism in postruminant sheep.
Aging↗
Perturbation of methionine metabolism in sheep with nitrous-oxide-induced inactivation of cobalamin.
Exposure of sheep to 36% nitrous oxide for 8 days (2-hr per day) led to 90%, 82% and 74% inhibition of 5-methyltetrahydrofolate-homocysteine methyltransferase in the liver, heart and brain, respectively, while there was no significant decrease in the activity of methylmalonyl-CoA mutase. There was also no change of betaine-homocysteine methyltransferase activity. The level of plasma methionine in nitrous-oxide-exposed sheep fell to 30% of its initial value. S-Adenosylmethionine level was reduced to 50% of the control value in the liver, and was also significantly decreased in the heart, but not in the brain. Excretion of formiminoglutamic acid and homocystine was also observed in the urine of sheep exposed to nitrous oxide. These results demonstrate that inhibition of 5-methyltetrahydrofolate-homocysteine methyltransferase causes a pronounced perturbation of methionine metabolism in sheep, suggesting that dietary methionine plus methionine synthesized from the methyl groups of betaine are not sufficient to meet the methyl needs for biological methylation reactions in this species and, in turn, emphasizing the role of 5-methyltetrahydrofolate-homocysteine methyltransferase in methionine synthesis in the sheep.
5-Methyltetrahydrofolate-Homocysteine S-Methyltran↗
Comparative studies on the methionine synthesis in sheep and rat tissues.
The important features of the enzymes involved in methionine synthesis in sheep were found to be the low activity of betaine-homocysteine methyltransferase and the high activity of 5-methyltetrahydrofolate-homocysteine methyltransferase. The rate of the methionine synthesis in sheep liver was significantly lower than that in rats due to the low activity of hepatic betaine-homocysteine methyltransferase. The hepatic methionine recycling was stimulated by the addition of betaine in both species. These results indicate that in sheep 5-methyltetrahydrofolate-homocysteine methyltransferase plays a significant role in hepatic methionine synthesis along with betaine-homocysteine methyltransferase. In contrast, in the rat hepatic system methionine synthesis is virtually dependent on betaine-homocysteine methyltransferase.
5-Methyltetrahydrofolate-Homocysteine S-Methyltran↗
A comparison of the influence of diets high in saturated versus unsaturated fatty acids on lipid composition and glucose-6-phosphatase activity of rat liver microsomes.
Glucose-6-phosphatase (G6Pase) activity in microsomes (M-SF) prepared from livers of rats fed a saturated fat-enriched diet has been observed to be 20-30% higher than in microsomes (M-PUF) prepared from rats fed a polyunsaturated fat-enriched diet. This difference was persisted throughout the 49 days experimental period. Neutral lipids of M-PUF contained a significantly higher concentration of 18:2 and 20:4, and a lower concentration of 16:0, 18:0 and 18:1 whereas in phospholipids 18:2 and 20:4 accumulated at the expense of 16:0 as compared to M-SF. Total phospholipid content from M-PUF was 30% lower than that from M-SF but the proportional phospholipid composition remained the same. An increase in cholesterol content was observed in M-PUF and this increase was localized in the ester fraction of cholesterol. These observations clearly indicate that manipulations of dietary lipids can influence both the fatty acid profiles and enzyme function of hepatic microsomal membranes.
Animals↗
Disturbance of methyl group metabolism in alloxan-diabetic sheep.
Alloxan-induced diabetes results in changes in the activities of a number of enzymes related to methyl group metabolism in sheep. Decreases in the activities of phospholipid methyltransferase and betaine-homocysteine methyltransferase in diabetic sheep liver indicate a reduced rate of choline synthesis and oxidation. A 65-fold increase in the activity of glycine methyltransferase and a 4-fold rise in the activity of gamma-cystathionase in diabetic sheep liver with elevated urinary excretion of cyst(e)ine suggest that catabolism of the methyl group of methionine and homocysteine was enhanced in the diabetic state.
5-Methyltetrahydrofolate-Homocysteine S-Methyltran↗
Regulation of methyl group metabolism in lactating ewes.
The effect of lactation on a number of enzymes involved in transmethylation reactions and the secretion of major methyl compounds into milk have been examined in sheep. The activities of hepatic phospholipid methyltransferase and 5-methyltetrahydrofolate-homocysteine methyltransferase were significantly higher in lactating ewes, compared with those in non-lactating ewes, while the activity of both hepatic and pancreatic glycine methyltransferase was significantly lower in the lactating state. No differences were observed in the activities of hepatic guanidoacetate methyltransferase, betaine-homocysteine methyltransferase and cystathionine beta-synthase on lactation. These results suggest that the extra demand for methyl groups for the secretion of methyl compounds in the milk is facilitated by enhancing the rate of de novo methyl group synthesis and lowering the rate of physiologically nonessential methylation.
Animals↗
Dependence on blood acetate concentration of the metabolic effects of ethanol in perfused rat liver.
In livers from fed rats perfused with recirculating blood, infusion of ethanol produced an inhibition of ketogenesis followed by substantially increased production. Perfusate lactate concentrations fell markedly following the increase in ketone body formation. In these experiments perfusate acetate rose continuously, reaching a concentration of 10 mM at 70 min, while in a control liver concentrations remained very low. During non-recirculating perfusion with 3 mM acetate there was output of lactate, whereas at 10 mM acetate ketogenesis was greatly stimulated and there was net lactate uptake. These data support the concept that there is a concentration of acetate in the region of 5 mM, below which it competes with lactate for lipogenesis. Above this level acetate may penetrate the mitochondrion and stimulate ketogenesis and gluconeogenesis. Effects of ethanol in vitro may depend on the concentrations of acetate attained in the experimental system.
Acetates↗