Tyrosine Aminotransferase In Rats Research Articles

Regulation of glucocorticoid receptors during adaptive liver growth.

Mirex, an organochlorine pesticide, is a potent inducer of liver hyperplasia and hypertrophy in intact (INT) rats. In contrast, mirex elicits predominantly hyperplastic liver growth in adrenalectomized (ADX) rats and hypertrophic liver growth in thyroidectomized (THX) rats. Supplements of glucocorticoids restore liver hypertrophy and inhibit DNA synthesis in ADX mirex-dosed rats. Because responsiveness to glucocorticoids is in part dependent on the number and affinity of glucocorticoid receptors (GR) we have measured hepatic GR levels and dexamethasone inducibility of tyrosine aminotransferase (TAT) in mirex-dosed INT, ADX, and THX rats. Specific [3H]dexamethasone binding sites decreased to 48, 49, and 59% of control values in mirex-dosed INT, ADX, and THX rats, respectively, with no changes in the apparent equilibrium dissociation constant at 48 h postdose. The significant depletion of hepatic cytosolic GR in all experimental groups dosed with mirex failed to impair both uninduced and dexamethasone-induced TAT levels in these groups. It appears that the decrease in steroid binding is neither a result of an interaction between mirex and hepatic GR nor a simple "dilution" of receptors due to the accompanying liver hypertrophy. Taken collectively, these data suggest that despite the ligand-independent downregulation of hepatic GR, responsiveness of hepatocytes from mixed-dosed rats to glucocorticoids is not altered. Therefore, glucocorticoid-mediated liver hypertrophy in mirex-dosed rats probably involves either 1) modulation of steps subsequent to ligand binding and activation translocation of hepatic GR, 2) glucocorticoid effects at extrahepatic sites that release factors that are the actual effectors of parenchymal cell enlargement, or 3) an unconventional receptor-mediated mechanism.

Biological Activity of the Fatty Acid Ester Metabolites of Corticoids*

Fatty acid esters of all families of steroid hormones are known to exist naturally. While their physiological roles are not clear, the C-17-fatty acid esters of estradiol are extremely potent and unusually long-lived estrogens. Thus, it appeared that increased potency would be a logical consequence of the esterification of all of the active steroid hormones. To test this hypothesis we measured the effect of an ester of corticosterone, corticosterone-21-stearate, on the induction of tyrosine aminotransferase in adrenalectomized rats. Surprisingly, while the ester is active compared to the unesterified corticoid corticosterone, there was no difference in either the magnitude or the duration of the induction of this enzyme. To determine whether the C-21-steroidal ester could itself induce this gluconeogenic response, we tested corticosterone-21-oleate and corticosterone-21-stearate as competitors for the binding of [3H] dexamethasone to the glucocorticoid receptor in rat liver cytosol. Both were poor ligands, with binding affinities of about 4% and more than 1%, respectively, compared to corticosterone. From these results, it is doubtful that the esters could act directly in vivo without prior cleavage of the fatty acid. We measured the rate of hydrolysis of corticosterone-21-stearate and estradiol-17-stearate by rat liver esterases. Corticosterone-21-stearate is hydrolyzed at a much greater rate (10- to 25-fold) than estradiol-17-stearate. Consequently, the difference in both potency and duration of response between the ester of the corticoid and that of the estrogen can be explained by the very rapid rate of conversion of the former into the unesterified form. Since the esterification of the corticoids appears not to be related to an increased biological half-life, as it is in the estrogens, the question remains as to the physiological role that they might play.

The effect of age on the induction of tyrosine aminotransferase and tryptophan oxygenase genes by physiological stress

Expression of the genes coding for tyrosine aminotransferase and tryptophan oxygenase in rats is induced in response to cold stress. We have studied the effect of ageing on this induction. The induction of tyrosine aminotransferase activity in young adult rats (10 months) was about twice that observed with old rats (25 months). This difference between the two age groups was also observed when the steady-state level of tyrosine aminotransferase mRNA was measured by hybridization with a specific DNA probe. However, when the transcription rate of the gene was measured by in vitro elongation of nascent RNA in isolated nuclei, no difference was detected. In contrast to the results with tyrosine aminotransferase, induced tryptophan oxygenase enzyme and mRNA levels did not show an age-dependent difference. These results suggest that there is, with ageing, an impairment in post-transcriptional regulation of the synthesis of tyrosine aminotransferase. The regulation of tryptophan oxygenase, on the other hand, is similar in the two age groups.

Age-related changes of liver tyrosine aminotransferase in senescent rats.

Tyrosine aminotransferase was induced in adult and senescent rat liver and its properties studied. We show the appearance of a 'cross-reacting material' for induced tyrosine aminotransferase of old rats compared to basal enzyme; this cross-reacting material can be provoked in adult rats after injection of cycloheximide, and suppressed in adult and old rats after injection of a serine protease inhibitor (tosylphenylalanine chloromethylketone). Other properties of induced tyrosine aminotransferase (thermostability, Km for tyrosine, isoelectrofocusing) are identical except for the proportion of the three forms and their sensitivity to trypsin in the absence of pyridoxal phosphate, which is increased in senescent animals. The suppression of cross-reacting material clearly indicates that it is not due to errors on old rat liver DNA but rather to post-translational modifications. This demonstrates also the role of serine proteases in tyrosine aminotransferase degradation. We suggest that induced enzyme of senescent rats would undergo a conformational change, possibly due to a release of pyridoxal phosphate from the enzymic molecules, which would thus become more susceptible to proteolytic attack than those of adult rats.

Influence of caprolactam on rat-liver tyrosine aminotransferase and tryptophan oxygenase

Caprolactam (2-oxohexamethylenimine), the monomer used in the production of Nylon 6, was tested for its ability to induce tyrosine aminotransferase (TAT) and tryptophan oxygenase (TPO) in rats. When a single oral dose of 1·5 g caprolactam/kg was given, maximum induction occurred in the livers after 6 hr and there were marked increases in enzyme activities after 3 and 24 hr. The induction was dose dependent, with a no-effect level for TAT of 300 mg/kg and for TPO of less than 300 mg/kg. In a paired-feeding study, it was shown that caprolactam inhibited the adaptive changes in liver-protein synthesis that were associated with a decrease in food intake. The induction of TAT by caprolactam did not require endogenous steroid, as increased enzyme activity was observed in adrenalectomized rats. Furthermore, the caprolactam induction did not appear to require RNA synthesis, since the induction was only partially blocked by inhibitors of RNA synthesis, actinomycin D or aflatoxin B 1. It is suggested that caprolactam induces changes in amino acid metabolism that bring about alterations in the activities of these enzymes.

Initial steps in the induction by glucocorticosteroids of rat liver tryptophan oxygenase and tyrosine aminotransferase

Approximately 90 min after the application of dexamethasone, an increase in the activity of the liver enzymes tryptophan oxygenase and tyrosine aminotransferase in adrenalectomized male rats could be measured. As early as 30 sec after the injection of an inducing amount of [ 3H]dexamethasone, significant radioactivity can be found in purified rat liver nuclei. The uptake is linear during the first 15 min, whereas during the same time period, the increase of the radioactivity in the 10,000 g supernatant is biphasic. No time shift could be observed in the uptake of dexamethasone by nuclei as compared to the accumulation in the 10,000 g supernatant. Studies on the time course of sensitivity of the induction process to α-amanitin suggested that the first induced transcripts for tryptophan oxygenase and tyrosine aminotransferase are completed about 25 min after the administration of the hormone. The ensuing processing of the transcripts to the corresponding mRNAs can be blocked by actinomycin D, the actinomycin D-sensitive phase lasting another 15–25 min. A lag phase of approximately 20 min has been calculated from the time of the entry of the first hormone molecules into the nucleus till the initiation of specific tryptophan oxygenase and tyrosine aminotransferase gene transcription.

Effects of morphine on isoenzymes of pyruvate kinase and tyrosine aminotransferase in rat

The acute and chronic effects of morphine on isoenzymes of pyruvate kinase and tyrosine aminotransferase in rat liver have been studied in vivo. Acute administration of morphine inhibits the L-type pyruvate kinase but increases forms II, III and IV of tyrosine aminotransferase in rats which have received a single intraperitoneal injection of morphine 6 hr prior to sacrifice. However both effects are lost after 24 hr. Morphine starts to regain its stimulatory effect on tyrosine aminotransferase at the hr 48 and reaches a maximum at the hr 72. Chronic morphinization leads to similar changes in the total activities of pyruvate kinase and tyrosine aminotransferase but the change in isoenzyme pattern of tyrosine aminotransferase concerns only forms II and III. Results from this study support the hydrocortisone-like property of morphine and indicate that morphine may enhance gluconeogenesis in vivo.

The repression and derepression of hepatic tyrosine aminotransferase by carcinogens

Like hydrocortisone, a single carcinogenic dose of dimethylnitrosamine (50 mg/kg) initiates the induction cycle for hepatic tyrosine aminotransferase in adrenalectomized rats. However, following this initial induction in the presence of dimethylnitrosamine, the enzyme becomes refractory to reinduction by known inducers. The administration of thioacetamide to either adrenalectomized or intact rats leads to an immediate and progressive loss of inducibility by hydrocortisone, dibutyrylcyclic AMP or dimethylnitrosamine. Although the thioacetamide-induced repression was not reversed even up to 10 weeks after the cessation of treatment, it was reversed after the induction of liver regeneration. Both the carcinogen-mediated induction and repression of tyrosine aminotransferase appears to occur by mechanisms which do not involve the corticosteroid-binding proteins which normally mediate the induction by glucocorticoids.

Effect of high- and low-protein diets on the regulation of rat liver tyrosine aminotransferase and tryptophan oxygenase activities by L-tyrosine and L-tryptophan.

Induction of rat liver tyrosine aminotransferase by l-tyrosine and tryptophan oxygenase by l-tryptophan was studied in groups of rats fed on diets containing 18 or 5% protein. The basal activity of hepatic tyrosine aminotransferase of rats receiving 5% protein gradually increased with the age of the animals but that of rats receiving 18% protein did not. l-Tyrosine induced hepatic tyrosine aminotransferase in rats receiving 18% protein when tested at ages from 4 to 20 weeks. When induction by l-tyrosine was carried out in rats receiving the 5% protein diet, significant induction of tyrosine aminotransferase occurred only in 4- or 6-week-old rats. Induction by l-tryptophan of tryptophan oxygenase in liver or the basal activity of this enzyme in liver did not differ between the groups fed on 5 and 18% protein. On changing the diet from 0 to 18% protein, the above-mentioned effects on the induction of hepatic tyrosine aminotransferase were reversed.

Time and level of increased plasma corticosterone necessary for induction of liver tyrosine aminotransferase in rats subjected to trauma.

Time and level of increased plasma corticosterone necessary for induction of liver tyrosine aminotransferase in rats subjected to trauma.

Enhancement by carbon tetrachloride treatment of hepatic tyrosine amino-transferase induction in the presence of net polyribosome breakdown in the rat

Carbon tetrachloride treatment inhibited the glucocorticoid induction of hepatic tryptophan oxygenase (TPO) while simultaneously enhancing the induction of hepatic tyrosine aminotransferase (TAT) in intact or adrenalectomized rats. In addition, treatment with CCl 4 alone increased the activity of hepatic TAT, but the extent of the CCl 4-induced stimulation of TAT was decreased by adrenalectomy or by hypophysectomy. The CCl 4-induced stimulation of TAT was prevented by high doses of actinomycin D but was enhanced further by 5-fluorouracil. CCl 4 also enhanced the labeling of hepatic polyribosomes with [ 14C]orotic acid while simultaneously producing breakdown of heavy polysomes. These findings suggest that CCl 4 treatment may enhance the synthesis of certain hepatic enzymes, possibly via synthesis of new polysomes, while simultaneously inhibiting the induction of other enzymes such as TPO.

The role of cyclic AMP in the induction of tyrosine aminotransferase in neonatal rats: evidence for a release function based on an anomalous effect of puromycin

1. 1. In surgically delivered rat infants the administration of puromycin at 3 or 6 hours after delivery leads to potentiation of the postnatal synthesis of tyrosine aminotransferase but puromycin given at delivery effectively prevents production of enzyme activity. 2. 2. Zone sedimentation analysis shows the enzyme activity ‘ induced ’ by puromycin to be of normal size. 3. 3. Hormonal ‘ cocktails ’ containing insulin, hydrocortisone, and adrenaline produce 3 forms of tyrosine aminotransferase in 2-day postnatal rats, adrenaline produces only I form, and puromycin is an apparent inducer in some litters of animals. All these treatments produce tyrosine aminotransferase of the same sedimentation behaviour. 4. 4. Form C of the enzyme is produced by 5 hours after birth of the rat, and when compared with the multiple forms of the enzyme produced by the triple hormonal cocktail, has the same sedimentation characteristics. 5. 5. In a certain proportion of 2-day postnatal rat litters enzyme activity can be increased by adrenaline or puromycin and both agents either are consistently effective or are both ineffective. 6. 6. In ‘ inducible ’ litters, RNase digestion of either liver extracts or the polysomemicrosome fraction increases the enzyme activity in vitro but has no effect in ‘ pre-induced ’ or ‘ non-inducible ’ preparations. 7. 7. The data are interpreted as indicating the operation of a control mechanism working at an enzyme release step in the tyrosine aminotransferase system and taken together with other work it is suggested that this mechanism operates on form C of tyrosine aminotransferase and is mediated at the intracellular level by cyclic AMP.

Effects of hydrocortisone and growth hormone on tyrosine aminotransferase and tryptophan oxygenase levels in hypophysectomized rats.

The inhibitory effects of highly purified bovine growth hormone (BGH) on the induction of tyrosine aminotransferase (TA) and tryptophan oxygenase (TO) by hydrocortisone in hypophysectomized rats have been studied. A single injection of 25 μg of BGH inhibits the response of TA to HC 44%. The induction of TO by HC is diminished 53% by the administration of only 12.5 μg of BGH. These data indicate that the modifying effects of somatotropin 1) are probably due to BGH alone because of its high chemical and biologic purity, and 2) may be operational at the physiologic level since minute quantities of BGH cause maximal responses. (Endocrinology 86: 1448, 1970)

Analysis of hormonal-dietary interactions and the role of gluconeogenesis in the regulation of tyrosine aminotranferase in rats.

The extent of stimulation of rat liver tyrosine aminotransferase by Cortisol and glucagon is inversely proportional to the basal activity of the enzyme in adrenalectomized and hypophysectomized rats. The basal activity is low in animals fed a high-carbohydrate diet leading to high hormonal stimulation of the enzyme, and high in animals fed a high-protein diet leading to minimal or no hormonal stimulation. The dietary modifications of the hormonal effects have been analyzed into two independent processes. Upon short-term starvation, the enzyme activity declines rapidly in animals previously fed a high-protein diet but rises rapidly in animals previously fed a high-carbohydrate diet. Superimposing a positive (hormonal) effect on these two diametrically opposite processes results in profound modification of the hormonal effect on the enzyme activity. The experiments further suggest that the regulation of tyrosine aminotransferase is coupled with gluconeogenic activity per se rather than with a primary hormonal action at the genetic level.

The hormonal regulation of enzymes in penatal and postnatal rat liver. Effects of adenosine 3',5'-(cyclic)-monophosphate.

1. The administration of glucagon, cAMP [adenosine 3',5'-(cyclic)-monophosphate], BcAMP [6-N-2'-O-dibutyryladenosine 3',5'-(cyclic)-monophosphate] or adrenaline to foetal rats during the last 2 days of gestation evoked the appearance of tyrosine aminotransferase and enhanced the accumulation of glucose 6-phosphatase in the liver. In foetuses 1-2 days younger only BcAMP was effective. After birth liver glucose 6-phosphatase no longer responds to glucagon or BcAMP. Tyrosine aminotransferase is still inducible by these agents in 2-day-old rats, but not in 50-day-old rats. After adrenalectomy of adults glucagon or BcAMP can enhance the induction of the enzyme by hydrocortisone. The results indicate that the ability to synthesize tyrosine aminotransferase and glucose 6-phosphatase when exposed to cAMP develops sooner than the ability to respond to glucagon with an increase in the concentration of cAMP; the responsiveness of enzymes to different hormones changes with age. A scheme illustrating the sequential development of competence in regulating the level of an enzyme is presented. 2. Actinomycin inhibited the effects of glucagon and BcAMP on liver tyrosine aminotransferase and glucose 6-phosphatase in foetal rats. Growth hormone, insulin and hydrocortisone did not enhance the formation of these enzymes. 3. The time-course of accumulation of glucose 6-phosphatase in the kidney is different from that in the liver. Hormones that increase the accumulation in foetal liver do not do so in the kidney of the same foetus or in the livers of postnatal rats.