Abstract. Changes in pancreatic exocrine enzyme activities were studied in different forms of experimental diabetes in rats. The effects of adrenalectomy on these changes were measured. The early actions of insulin on pancreatic enzyme activities and on incorporation of (4,5‐3H)‐leucine into amylase were also determined. The main results are: Alloxan‐diabetes leads to a decrease in amylase activity, a decreased rate of amylase synthesis and an increase in the activities of trypsinogen and chymotrypsinogen as described by Palla et al. [9]. Only the decrease in amylase activity correlates with the increase in blood glucose concentration. Adrenalectomy does not reverse the changes in the activities of exocrine pancreatic enzymes induced by diabetes. Diazoxide‐diabetes is accompanied by an increase in all pancreatic enzyme activities, most probably due to the inhibition of enzyme secretion. After treatment with streptozotozin a significant decrease in pancreatic amylase activity appears after 36 h. Amylase activity continues to fall during the following days, in contrast the increase in the activities of proteolytic enzymes is not significant until the 4th day. Insulin treatment of severely diabetic, non‐ketotic rats leads, as early as 90 min. after injection, to a significant increase in pancreatic amylase activity with no change in the other exocrine enzyme activities. A decrease in all enzyme activities, seen 6 h after insulin, is due to enhanced enzyme secretion. As soon as 2 h after injection insulin leads to a significant increase in the incorporation of (4,5‐3H)‐leucine into pancreatic amylase but not into total pancreatic protein. This increase is completely abolished by actinomycin D. Short‐term effects of adrenalectomy (20 h) and effects of substitution with 6‐α‐fluoro‐16α‐methyl‐prednisolon on the incorporation of (4,5‐3H)‐leucine into pancreatic amylase and total pancreatic protein result mainly from changes in the size of the cold leucine pool. According to these results insulin regulates pancreatic amylase synthesis mainly at the level of transcription. Insulin plays a permissive role in pancreatic amylase synthesis and is not involved in short‐term regulation of amylase synthesis in the non‐diabetic state. Clinical findings in juvenile diabetics indicate a similar type of regulation for amylase synthesis.
At external concentrations of 50,UM, L-methionine was rapidly taken up by hepatocytes, whereas almost no S-adenosylmethionine (SAM) was removed from the incubation medium. SAM did not enter the intracellular water space but equilibrated with a very small pool, which was most likely to be situated on the external side of the plasma membrane. Methyl groups from external L-methionine, but not from external SAM, were incorporated into total and nuclear RNA. A significant incorporation of methyl groups into phospholipids occurred not only with methionine but also with SAM. After subfractionation of hepatocytes it became evident that methyl groups from SAM were mainly incorporated into plasma-membrane phospholipids, and that phospholipid methylation in other cellular compartments resulted from contamination with plasma membrane. The pattern of methylation of the various phospholipid species with SAM as precursor was different from that obtained with L-methionine. In contrast with external L-methionine, external SAM did not enter the intracellular SAM pool. According to these results a transport system for SAM does not exist in rat hepatocytes, although methyl groups from external SAM can phospholipids from the outside.Studies concerning transmethylation processes in intact cells or tissues can be done by using [Me-3H]or [Me-14C1-methionine. After entering the cell the labelled methionine is converted into the corresponding labelled SAM, which then mixes with the internal pool of unlabelled SAM. From this pool methyl groups are transferred by specific methyltransferases on to their physiological acceptors.Since the early experiments of Stekol et al. (1958), who observed an increased formation of methylated products by rat liver slices in the presence of exogenous SAM, various groups have looked for a specific uptake system for SAM. This would offer the opportunity to label the internal SAM pool directly and allow measurements of transfer of methyl groups on to proteins without interference Abbreviations used: Hepes, 4-(2-hydroxyethyl)l-piperazine-ethanesulphonic acid; PtdEtn, phosphatidylmonoethanolamine; Ptd[EtnI2, phosphatidyldiethanolamine; PtdCho, phosphatidylcholine; SAM, S-adenosyl-L-methionine; SDS, sodium dodecyl sulphate.be incorporated into plasma-membrane with the incorporation of labelled methionine via protein synthesis.An uptake system for SAM has indeed been described for yeast cells by Svihla & Schlenk (1960) and by Spence (1971) and was characterized by Murphy & Spence (1972). The transport was temperature-and energy-dependent, had an apparent Km for SAM of 3.3,pM and was inhibited by S-adenosylhomocysteine and S-adenosylethionine. Similar findings were obtained by Nakamura & Schlenk (1974). Stramentinoli et al. (1978a,b) found a transport system for SAM in rabbit erythrocytes. Zappia et al. (1978) described the transport of exogenous SAM into isolated perfused rat livers, and Stramentinoli et al. (1978) claimed even that exogenous SAM was able to protect rat livers against galactosamineinduced liver...
Whole liver and isolated liver mitochondria are able to release free acetate, especially under conditions of increased fatty acid oxidation. In the present paper it is shown that rat liver contains acetyl-CoA deacylase (EC 3.1.2.1) activity (0.72,umol/min per g wet wt.of liver at 30°C and 0.5 mM-acetyl-CoA). At 0.5 mM-acetyl-CoA 73 % of total enzyme activity was found in the mitochondria, 8 % in the lysosomal fraction and 19% in the postmicrosomal supernatant. Mitochondrial subfractionation shows that mitochondrial acetyl-CoA deacylase activity is restricted to the matrix space. Mitochondrial acetylCoA deacylase showed almost no activity with either butyryl-or hexanoyl-CoA. AcetylCoA hydrolase activity from purified rat liver lysosomes exhibited a very low affinity for acetyl-CoA (apparent Km >15 mm compared with an apparent Km value of 0.5 mm for the mitochondrial enzyme) and reacted at about the same rate with acetyl-, n-butyryland hexanoyl-CoA. We could not confirm the findings of Costa & Snoswell [(1975) Biochem. J. 152, 167-172] according to which mitochondrial acetyl-CoA deacylase was considered to be an artifact resulting from the combined actions of acetyl-CoA-Lcarnitine acetyltransferase (EC 2.3.1.7) and acetylcarnitine hydrolase. The results are in line with the concept that free acetate released by the liver under physiological conditions stems from the intramitochondrial deacylation of acetyl-CoA.
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