Ján Knopp

Regulation of cytosolic malate dehydrogenase by juvenile hormone in Drosophila melanogaster

The Drosophila malate dehydrogenase, or malic enzyme (ME) encoded by the Men gene, is a non-mitochondrial enzyme recovered in the cytosolic fraction. By using mutation in the Men gene and deficiencies uncovering this locus, we could show that the ME activity recovered in cytosolic fractions originates exclusively from the Men gene located at map position 87D-1 on the right arm of the 3rd chromosome. We found that juvenile hormone (JH) can induce ME activity by two mechanisms. One mechanism corresponds to a direct effect of JH on the enzyme, whose activity was enhanced by a two-fold factor in the absence of transcription and translation. This enhancement can be noticed 1 h after JH treatment and lasts for approx 3–4 h. The other mechanism involves the transcription of the MEN gene. In the absence of inhibitors the induction of ME activity by JH is increased by a three to fourfold factor and extends over a period of 10–16 h. Since induction of ME activity by JH and JH analogs displayed a dose-response curve, specific for each tested component, we concluded that the hormonal action could be mediated through a receptor. The use of two temperature sensitive mutations deficient in the production of ecdysteroid, ecd1 and su(f)ts67g, revealed that ME response to JH requires the presence of a minimal level of the steroid hormone ecdysone, showing a complex hormonal regulatory circuit in the execution of the JH response.

Insulin and Catecholamines Act at Different Stages of Rat Liver Regenerationa

Insulin and catecholamines are known to exert effects on hepatocyte growth and metabolism. The binding of insulin, the plasma levels of insulin (INS), and the plasma catecholamine levels of epinephrine (EPI) and norepinephrine (NE) were measured during liver regeneration after partial hepatectomy (PH). A significant decrease (p < 0.05) of INS receptor binding capacity was found at 1, 2, and 3 days after operation. A single insulin injection (2.5 IU/kg body weight) at 24 h after sham operation or partial hepatectomy did not affect these changes of INS binding to hepatocytes. The plasma insulin and glucose levels were similar in both hepatectomized and sham-operated rats. Within 20 min after liver resection or sham operation, plasma NE and EPI concentrations increased rapidly. Then, a significant decrease was observed in plasma catecholamine levels at 1 h after laparotomy and PH. In both groups, laparotomized and partially hepatectomized plasma levels of NE at 4 h reached control values and remained unchanged at the 4- and 24-h periods. After PH, the levels of EPI remained elevated at 4 h in comparison with laparotomy. Adrenal tyrosine hydroxylase mRNA levels were significantly elevated at 4 h in both PH and sham-operated groups. These results suggest that signals that are initiated by catecholamines and transduced through second messengers presumably participate in the trigger mechanism of liver regeneration, while insulin (considered as a secondary mitogen) enhances a stimulus for liver regeneration.

Iodine Concentration and Content in the Organs of Rat during Postnatal Development

It was found that the thyroid attains its maximum relative weight during the first three weeks of postnatal life. The concentration of thyroidal iodine declines during the 3rd week; the highest value

Effect of Varying Iodine Intake on its Concentration in Rat Tissues during Early Development

From the day of parturition, three groups of lactating rats and their young were kept on a laboratory diet containing 60 ng iodine/g of food and drank water with an iodine concentration of 5, 100 and 500 ng/ml, respectively. The first two groups of sucklings showed a relatively decreasing and the third a relatively increasing iodine intake (μg I/day/100 g body weight). Iodine intake in food was found to affect iodine concentration but had no effect on its decline in the tissues during postnatal life. Consequently, it is presumed that this decline is genetically controlled.

Stress Is Associated with Inhibition of Type I Iodothyronine 5′‐Deiodinase Activity in Rat Liver

Abstract: Type I iodothyronine deiodinase (5′‐DI) generates the thyromimetically active hormone 3,5,3′‐triiodothyronine (T3) by reductive monodeiodination of the phenolic ring of L‐thyroxine (T4). The present study was undertaken to compare the effects of cold stress (4°C) alone or in combination with immobilization stress (IMO), insulin treatment (5 IU/kg, i.p.) or 2‐deoxy‐D‐glucose (2DG)‐induced intracellular glucopenia on the activity of 5′‐DI in rat liver. Cold stress either for 24 h or 28 days when compared to that in the nonstressed group of rats significantly reduced (P < 0.001) the activity of 5′‐DI in liver. In comparison with cold stressed rats for 28 days, an additional decrease in 5′‐DI activity was observed when those rats underwent 1 × IMO in cold for 2 h (P < 0.001) or insulin treatment (P < 0.05). A significant decrease in 5′‐DI activity was found also in rats maintained at room temperature that underwent a single 1 × IMO for 2 h (P < 0.001) or insulin treatment (P < 0.01) when compared to nonstressed animals. In comparison with nonstressed rats, no significant change of the 5′‐DI activity was observed after 2DG application (500 mg/kg, i.p.) at room temperature. In conclusion, cold stress and/or 1 × IMO, insulin treatment, or 1 × IMO at room temperature markedly affects reductive monodeiodination of T4, and thus reduces the concentration of biologically active T3 in liver.

Nuclear 3,5,3′-triiodothyronine receptors and malic enzyme activity in liver of rats fed fish oil or cocoa butter

The role of different fat supplements (fish oil and cocoa butter), applied intragastrically for 3 weeks, on specific binding of T3 by rat liver nuclear receptors was examined. Fish oil (rich in n-3 polyunsaturated fatty acids) supplementation resulted in a significant (P < 0.05) enhancement of the maximal binding capacity (MBC) for T3 with a diminution of the apparent association constant (Ka). No changes in nuclear receptor MBC or Ka were observed after cocoa butter administration. Moreover, the rate of fatty acid synthesis in the liver was evaluated by measuring malic enzyme (EC. 1.1.1.40.) activity in rat liver cytosol, which was found to be decreased in both the cocoa butter- (P < 0.002) and the fish oil- (P < 0.001) treated rats. Interestingly, in fish oil-supplemented animals, not even a higher MBC value for the T3 receptors in liver nuclei and specific binding of T3 to its receptor were able to restore the activity of malic enzyme in liver cytosol. The findings of the same level of thyroid hormones—thyroxine (T4) and triiodothyronine (T3)—in blood of all groups showed that there was apparently no effect of fat supplementation on the thyroid function. However, it may be hypothesized that the decrease of malic enzyme activity in both fat-treated groups may have resulted from a decreased T4 to T3 conversion in liver cells. The data suggest the following: (a) increased dietary fat intake is accompanied by lower lipogenesis in the liver in relation to the degree of fat unsaturation; (b) although polyunsaturated dietary fat increases T3 receptor binding to liver nuclei; (c) thus, an uncoupling between T3 binding and action on malic enzyme activity during raised fish oil intake cannot be ruled out.

Ontogenetic Development of Deiodinase and Aminotransaminase in Rat Thyroid Gland

Thyroid aminotransaminase and deiodinase in the infant rat were examined during the postnatal period of life. Both enzymes were found on the first day after delivery with various activities, low the deiodinase and high the aminotransaminase. The activity of enzymes increased during the third postnatal week. These changes of enzymes probably represent changes in the functional state of the hormonogenesis in the thyroid gland which maturates in this period of life.

Effect of 3,5,3′-triiodothyronine on cytosolic cAMP-dependent, cAMP-independent protein kinase and nuclear DNA-associated kinase activity in proliferating rat liver under different nutritional regimens

Cytosolic cAMP-dependent, cAMP-independent protein kinase (PK) and DNA-associated nuclear kinase activity were compared in the proliferating liver of rats: (1) fed with commercially available basal laboratory diet; (2) fasted (2 days) and then fed with basal laboratory diet (2 days); (3) fasted (2 days), supplied with 30% glucose (1 day), and then realimented with basal laboratory diet; (4) fasted (2 days), supplied with 30% glucose (1 day), and then given a single dose of 3,5,3′-tri-iodothyronine (T 3 ) with realimentation (1 day with basal laboratory diet). The incorporation of 14 C-thymidine into rat liver DNA was increased in animals supplemented with glucose ( P 3 ) injection. Fasting significantly decreased both cytosolic cAMP-dependent and cAMP-independent PK activities in the rat liver, but not DNA-associated kinase activity, when compared with the ad libitum fed controls. Both of the cytosolic kinase activities were restored by glucose, while nuclear DNA-associated kinase activity was not altered. T 3 did not change the cytosolic kinase activities but significantly increased the DNA-associated kinase activity. The data obtained by Linewiever-Burk analysis of the cAMP-dependent protein kinase activity suggest that the lower activity in liver of fasted rats restored by glucose was not due to the induction of the enzyme. The specific binding of T 3 to rat liver nuclear receptors was characterized; the equilibrium association constant (Ka) and the maximal binding capacity (MBC) were evaluated. Fasting for 48 hr decreased MBC ( P 3 may play a role as a modulator of liver cell proliferation, potentially via nuclear DNA-associated kinase activity. cAMP-dependent and cAMP-independent cytosolic kinase activities may be necessary for permitting DNA synthesis stimulated by nutritional manipulations, but these cytosolic kinase activities are clearly not sufficient for stimulating elevated DNA synthesis: increased DNA-associated kinase activity appears to be critical. Our results also support the idea that the mechanism of hepatocyte proliferation in rats caused by a nutritional regimen may differ from that found in the liver regenerating after partial hepatectomy.

Nuclear Receptors for Thyroxine in Rat Liver during Postnatal Development in Relation to DNA-Dependent RNA Polymerase Activity

Specific binding of 125I-thyroxine on liver nuclear extract of rat fetuses aged 15 and 18 days, of young rats aged 1, 2, 7, 12-14 and 18 days and of adult rats was quantified. The highest value representing the capacity of specific 125I-thyroxine binding was noted in rats aged 7 days. A marked diminution of specific 125I-thyroxine binding was demonstrated in rats aged 12-14 days. Practically the same values of Ka in liver nuclear extract were found in fetuses aged 18 days (Ka = 4.44 X 10(9) 1 mol-1) and in a group of young animals aged 7 days (Ka = 4.17 X 10(9) 1 mol-1). In nuclear extract of the animals aged 12-14 days and in adult rats diminished values (Ka = 2.72 X 10(9) 1 mol-1 and Ka = 1.18 X 10(9) 1 mol-1, respectively) were noted in comparison with the above-mentioned groups of rats. A positive correlation between the binding of 125I-thyroxine and Mn2+-activated DNA-dependent RNA polymerase activity in liver nuclei of rats aged 4 and 14 days and adult rats was found (r = 0.700). These results support the suggestion that thyroxine in the perinatal period of life plays an important role in the induction of mRNA.