Lipogenesis at the suckling-weaning transition in liver and brown adipose tissue of the rat

The responses of rat hepatic and brown adipose tissue in vivo lipogenesis to premature (15 days) and normal (21 days) weaning have been correlated to changes in the activities of acetyl-CoA carboxylase and two NADPH-producing enzymes, malic enzyme and glucose-6-phosphate dehydrogenase. Both tissues show an induction of lipogenesis in response to weaning. In the liver, lipogenic flux is closely linked to the activity of acetyl-CoA carboxylase, but not necessarily that of malic enzyme or glucose-6-phosphate dehydrogenase, whereas no such dissociation between enzyme activity and flux rate occurs in brown adipose tissue. Thyroid hormones, implicated in many physiological changes around weaning, do not seem to play a primary role in the adaptation of lipogenesis to the dietary change at this time, although a permissive role in both tissues is possible.     

Effect of clofibrate on malic enzyme and leptin mRNAs level in rat brown and white adipose tissue.

Two previous studies have reported contradictory results regarding the effect of fibrates treatment on obese (ob) gene expression in rodents. The purpose of the present study was to reinvestigate this issue. We examined the effect of clofibrate (fibrate derivative) administration for 14 days to rats on malic enzyme (as an adequate control of fibrates action) and leptin mRNAs level in the white and brown adipose tissues (WAT and BAT, respectively). The malic enzyme activity and malic enzyme mRNA level in white adipose tissue increased significantly after clofibrate feeding. In brown adipose tissue, the drug treatment resulted in depression of malic enzyme activity and malic enzyme mRNA level. Under the same conditions, leptin mRNA level did not change in these tissues. The results presented in this paper provide further evidence that the clofibrate (activator of peroxisome proliferator activated receptor alpha), feeding is without effect on ob gene expression in rat white and brown adipose tissue. Furthermore, the present study demonstrates that clofibrate causes opposite effects on malic enzyme gene expression in WAT (up-regulation) and BAT (down-regulation).     

Organ specific regulation of malic enzyme and hexosemonophosphate shunt dehydrogenases activity by high carbohydrate diet

The effect of starvation-refeeding transitions on the activity of malic enzyme and hexosemonophosphate shunt dehydrogenases in lipogenic and non-lipogenic tissues from rats was investigated. Starvation of the rats caused a decrease of malic enzyme activity in the liver, white and brown adipose tissue. Refeeding of the animals with high carbohydrate diet caused a several fold increase of malic enzyme activity in these tissues. Substitution of high fat for high carbohydrate diet resulted in only a slight increase of malic enzyme activity in the liver, white and brown adipose tissues. In the same rats, no significant effect of starvation-refeeding transition on malic enzyme activity in the kidney cortex, brain, heart, skeletal muscle and spleen was observed. The changes of the activity of hexosemonophosphate shunt dehydrogenases during starvation-refeeding transition essentially paralleled those of malic enzyme in all the tissues examined.     

Evidence that triiodothyronine decreases rat serum leptin concentration by down-regulation of leptin gene expression in white adipose tissue

Conflicting results have been reported regarding the effect of triiodothyronine (T(3)) on serum leptin and adipose tissue leptin gene expression in human and animals. The aim of the present study was to evaluate the effect of administration of increasing doses of T(3) on serum leptin concentration and on leptin mRNA abundance in white adipose tissue of rats. The results presented in this paper indicate that administration of single different doses of T(3) to euthyroid rats resulted dose dependent increases of serum total T(3) concentrations which are associated with a decrease in white adipose tissue leptin mRNA level. The leptin mRNA level in white adipose tissue was negatively correlated with serum total T(3) concentration (r=-0.8, p<0.001). Like white adipose tissue leptin mRNA level, serum leptin concentration decreased after T(3) administration, and was also negatively correlated with the serum T(3) concentration (r=-0.8, p<0.001). In contrast, administration of T(3) to the same rats led to a significant increase in white adipose tissue expression of the malic enzyme gene (malic enzyme activity and malic enzyme mRNA level), a known target gene for T(3). The results indicate that T(3) exerts a selective inhibitory effect on white adipose tissue leptin gene expression in vivo. A conclusion is that T(3) decreases rat serum leptin concentration by down-regulation of leptin gene expression in white adipose tissue.     

Effects of fasting on lipoprotein lipase activity in different depots of white and brown adipose tissues in diet-induced overweight rats

The aim of the present study was to evaluate the effects of 24 hours of starvation on lipoprotein lipase (LPL) activity in various depots of white and brown adipose tissues in control rats and in rats with two different degrees of overweight, both induced by dietary treatment. In control rats, no changes in LPL immunoreactive mass were observed in either white or brown adipose tissues after fasting, whereas the effects of food deprivation on enzyme activity were opposite in white versus brown adipose tissues. The LPL activity response to fasting was impaired by obesity: White adipose depots of cafeteria obese rats showed a lower ability to downregulate LPL during fasting and the increased LPL activity induced by fasting in brown adipose depots was less intense in the obese rats compared with control animals. When the degree of overweight was reduced, the differences between obese and control rats were also attenuated.     

Postnatal appearance of uncoupling protein and formation of thermogenic mitochondria in hamster brown adipose tissue

Brown adipose tissue of developing hamster was characterized by western blotting, enzyme activity measurements and immunoelectron microscopy. During the first postnatal week the tissue contained significant amounts of differentiating mitochondria and comparable quantities of active cytochrome oxidase and ATP synthase. The uncoupling protein appeared on the 7/8th day and its specific content increased 80-times between day 8 and day 17. In parallel, the specific content and activity of cytochrome oxidase increased 3-times but ATP synthase decreased 2-times. The total content of uncoupling protein and of cytochrome oxidase in interscapular brown adipose tissue increased 360- and 11-times, respectively. Analysis of isolated mitochondria showed that the observed differences result mainly from changes of the enzymic equipment of the mitochondrial membrane. During the same interval, propylthiouracil-insensitive "type II' thyroxine 5'-deiodinase activity in brown adipose tissue increased 10-times. It was concluded that the thermogenic function of the hamster brown adipose tissue develops after the first postnatal week due to highly differentiated synthesis of mitochondrial proteins leading to replacement of preexisting, uncoupling protein-lacking nonthermogenic mitochondria by thermogenic ones, similarly as shown in brown adipose tissue of the embryonic mouse and rat (Houst?k, J., et al. (1988) Biochim. Biophys. Acta 935, 19-25).     

Sex-linked changes in immunoreactive glucose-6-phosphate dehydrogenase in rat liver

The level of hepatic immunoreactive glucose-6-phosphate dehydrogenase protein was found to correlate well with the enzyme activity in adult rats fed the stock laboratory diet in a variety of hormonal conditions. The amount of immunoreactive protein and enzyme activity was 2-fold greater in sexually mature female rats compared with aged matched male animals. However, this difference was absent in diabetic animals, and furthermore although triiodothyronine administration to the diabetic male rat could restore the level of enzyme activity to that of the normoglycaemic animal, it was much less effective in the female animal. In contrast, administration of insulin to the normoglycaemic animal increased the level of glucose-6-phosphate dehydrogenase in the female, but was without effect in the male. These results are discussed in relation to the possible role of thyroid status and steroid sex hormones in the regulation of hepatic glucose-6-phosphate dehydrogenase.     

Lipogenesis in the liver and adipose tissue of the cardiomyopathic hamster.

The activities of fatty acid synthetase, citrate cleavage enzyme, glucose-6-phosphate dehydrogenase and malic enzyme were measured in the liver and adipose tissue of cardiomyopathic and normal hamsters at age 33, 68 and 108 days. There was no difference in the activity of hepatic fatty acid synthetase between the diseased animals and the controls at any stage in their development. The activity of glucose-6-phosphate dehydrogenase was not different until age 108 days where it was significantly elevated in the BIO 82.62 strain. Citrate cleavage enzyme in the liver was depressed at all stages in the diseased animals as was malic enzyme. In adipose tissue, all enzyme activities were significantly depressed in the cardiomyopathic animals at the three stages. These data suggest that lipogenesis was depressed in the cardiomyopathic hamster.     

Iodothyronine 5'-deiodinase activity in rat brown adipose tissue during development

Iodothyronine 5'-deiodinase activity in rat brown adipose tissue has a characteristic pattern of developmental changes that is completely different from that of the liver. Fetal brown fat exhibits an extremely high iodothyronine 5'-deiodinase activity that is approx. 10-fold that in adult rats. Even though brown fat iodothyronine 5'-deiodinase activity falls suddenly at birth, there is a new peak in the activity around days 5-7 of life, whereas it remains very low afterwards. Just after birth, brown adipose tissue iodothyronine 5'-deiodinase activity is already capable of stimulation by noradrenaline. The postnatal peak in brown fat iodothyronine 5'-deiodinase correlates with the known increase in the thermogenic activity of the tissue in the neonatal rat, thus reinforcing the suggestion that local 3',3,5-triiodothyronine generation could be an important event related to thermogenesis in brown adipose tissue. However, the high fetal activity was only slightly related to the thermogenic activity of brown fat. Moreover, the increased iodothyronine 5'-deiodinase activity of brown adipose tissue during fetal and neonatal life suggests a substantial contribution by brown fat in the overall extrathyroidal 3',3,5-triiodothyronine production in these physiological periods.     

The actions of dichloroacetic acid on blood glucose, liver glycogen and fatty acid synthesis in obese-hyperglycaemic (ob/ob) and lean mice

Obese-hyperglycaemic mice and lean mice were injected with dichloroacetate to determine the significance of gluconeogenesis in maintaining the hyperglycaemia of obese mice and to investigate the effects of a fall in blood glucose on fatty acid synthesis. One hour after the second of two, hourly, injections of dichloroacetate the blood glucose concentrations in fed and starved lean mice were decreased, whereas in obese mice they were sharply increased. In obese and lean mice, both fed and starved, dichloroacetate decreased plasma lactate but insulin was unchanged. The quantity of liver glycogen was decreased in all dichloroacetate treated mice, with the largest falls in fed and starved obese mice, which had much larger glycogen stores than lean mice. Dichloroacetate treatment decreased the concentration of plasma non-esterified fatty acids in fed and starved obese mice and fed lean mice but not in starved lean mice. Fatty acid synthesis in white (inguinal, subcutaneous) adipose tissue was stimulated by dichloroacetate in fed obese mice and inhibited in fed lean mice. Fatty acid synthesis in brown adipose tissue (scapular) was faster than in white adipose tissue and was less affected by dichloroacetate although the changes were in the same direction as in white adipose tissue. We attribute the increased hyperglycaemia of obese mice treated with dichloroacetate to increased glycogenolysis coupled with a failure to secrete additional insulin in response to the raised blood glucose. This high blood glucose concentration in dichloroacetate treated obese mice may in turn explain the increased fatty acid synthesis in their white adipose tissue.     

Regulation of the activity and synthesis of malic enzyme in 3T3-L1 cells

Malic enzyme activity in differentiated 3T3-L1 cells was about 20-fold greater than activity in undifferentiated cells. A new steady-state level was achieved about 8 days after initiating differentiation of confluent cultures with a 2-day exposure to dexamethasone, isobutylmethylxanthine, and insulin. This increase in enzyme activity resulted from an increase in the mass of malic enzyme as detected by immunotitration of enzyme activity with goat antiserum directed against purified rat liver malic enzyme. Malic enzyme synthesis was undetectable in undifferentiated cells and increased to about 0.2% of soluble protein in differentiated cells, suggesting that the increase in enzyme mass was due primarily to an increase in enzyme synthesis. Thyroid hormone, a potent stimulator of malic enzyme activity in hepatocytes in culture and in liver and adipose tissue in intact animals, decreased or increased malic enzyme activity in differentiating 3T3-L1 cells by about 40% when it was removed or added to the medium, respectively. Insulin, another physiologically important regulator of malic enzyme activity in vivo, had no effect on the initial rate of accumulation of malic enzyme activity in the differentiating cells and caused a 30 to 40% decrease in the final level of enzyme activity in the fully differentiated cells. Cyclic AMP, a potent inhibitor of malic enzyme synthesis in hepatocytes in culture, inhibited this process in 3T3-L1 cells by 30%. Malic enzyme is like several other enzymes in that the large increase in its concentration which accompanies differentiation of 3T3-L1 cells is due to increased synthesis of enzyme protein. However, the hormonal modulation of malic enzyme characteristic of liver and adipose tissue in intact animals does not appear to occur in differentiated 3T3-L1 cells, suggesting that differentiated 3T3-L1 cells may not be an appropriate model system in which to study the hormonal modulation of malic enzyme that occurs in liver and adipose tissue of intact animals.     

Dietary induction of hepatic malic enzyme activity: differentiation of the induction process

The activity of rat liver malic enzyme shows a marked increase when the animals are maintained on a restricted protein diet. Of the NADP-linked dehydrogenases tested (malic enzyme, glucose-6-phosphate dehydrogenase, and isocitrate dehydrogenase), the response is confined only to malic enzyme. Dietary sucrose is not required for the increase in activity, but elevated dietary levels of this disaccharide increase hepatic malic enzyme regardless of dietary protein. Glucose-6-phosphate dehydrogenase activity is increased by dietary sucrose provided adequate dietary protein is supplied. The specificity of the response to lowered dietary protein shown by malic enzyme suggests that the control of the hepatic enzyme is mediated by processes different from those controlling the activity of glucose-6-phosphate dehydrogenase.     

An immunohistochemical and in situ hybridisation study of the postnatal development of uncoupling protein-1 and uncoupling protein-1 mRNA in lamb perirenal adipose tissue

In the lamb, the uncoupling protein-1 (UCP1) content of perirenal adipose tissue at birth is an important factor in heat production by non-shivering thermogenesis and the prevention of hypothermia. This study examines UCP1 gene expression and protein content in perirenal adipose tissue over the first 15 days of life by in situ hybridisation and immunohistochemistry. UCP1 mRNA was detected at birth in 30% of adipocytes, and in approximately 24% of fat cells at 2 days of life. However, by 5 days of age and thereafter UCP1 mRNA was undetectable. Immunoreactive UCP1 was present in all adipocytes at birth and at 2 days of age, and remained detectable in a decreasing proportion of cells until day 10 of life. By 15 days of age no immunoreactive UCP1 was detected and the perirenal adipose tissue had the appearance of white fat. It is concluded that UCP1 gene expression is suppressed in most adipocytes in perirenal adipose tissue of newborn lambs, and gene expression rapidly falls in the remaining adipocytes over the first 5 days of postnatal life. In contrast, immunoreactive UCP1, a characteristic of brown adipose tissue, was present in many adipocytes for up to 10 days of age, suggesting that UCP1 has a long half-life in lambs. All adipocytes in perirenal adipose tissue of newborn lambs appear to be functionally brown, but over the first 2 weeks of postnatal life there is a complete transformation to white adipocytes.     

Hormonal regulation of malic enzyme and glucose-6-phosphate-dehydrogenase expression in fetal brown-adipocyte primary cultures under non-proliferative conditions.

The expression of malic enzyme and glucose-6-phosphate (Glc6P) dehydrogenase was investigated in primary cultures of fetal brown adipocytes after the prolonged presence (6 d or 10 d) of various hormones under non-proliferative conditions. The presence of triiodothyronine for 6 d and 10 d resulted in maturation of the triiodothyronine regulatory mechanism of malic-enzyme expression at the mRNA level. However, triiodothyronine had no effect on Glc6P dehydrogenase expression. Insulin increased malic-enzyme and Glc6P dehydrogenase expression at the mRNA and protein level after 6 d and 10 d of culture. The joint presence of triiodothyronine and insulin produced an additive effect on malic-enzyme expression at the mRNA and protein level after 6 d and 10 d of culture, by two independent mechanisms. Noradrenaline prevented the effect at the protein level after 6 d, but not after 10 d, probably due to loss of the beta-adrenergic response of brown adipocytes after prolonged culture. Triiodothyronine overexpressed the Glc6P dehydrogenase mRNA induced by the presence of insulin at 6 d and 10 d of culture. There was no adrenergic regulation of Glc6P dehydrogenase expression in cultured fetal brown adipocytes, regardless of the time of culture.     

Developmental changes in the activity of lipoprotein lipase (clearing-factor lipase) in rat lung, cardiac muscle, skeletal muscle and brown adipose tissue.

The lipoprotein lipase activity of the lung, skeletal muscle, heart muscle and brown adipose tissue of the rat was studied during the period from late foetal to adult life. The enzyme activity in all four tissues emerged substantially during the first 24th after birth. Subsequently, heart and lung enzyme activity remained relatively constant per unit wet weight of tissue. The enzyme activity present in brown adipose tissue and skeletal muscle was elevated per unit weight of tissue during suckling compared with other periods of life. Delivery of near-term foetuses stimulated the emergence of enzyme activity in all four tissues with the same time course as that evoked by normal delivery. The significance of the presence of the enzyme in the tissues and the activity changes which occurred during development are discussed in relation to possible mechanisms of control.     

Weaning and metabolic regulation in the rat

Premature weaning of rats to high carbohydrate diets causes a variety of short- and long-term changes in lipid metabolism, but the mechanisms involved are unclear. It is likely that interaction of diet with certain emerging hormonal control patterns during weaning might condition metabolic control and (or) subsequent adaptations in the adult organism. This implies that the adaptive responses of infant animals to diet may differ from those of the mature organism. For example, premature weaning leads to early appearance of rat liver malic enzyme (ME), even when fat supplies as much as 65% of the dietary energy; the same diet suppresses ME activity in 45-day-old rats. The levels of plasma glucagon and thyroid hormones are elevated during the weaning period. Several studies have shown that triiodothyronine evokes hepatic ME in suckling rats. Conversely, glucagon infusion into prematurely weaned rats suppresses the early appearance of the enzyme. Premature weaning, regardless of fat intake, leads to a rapid decline in plasma glucagon levels. Since glucagon is known to antagonize the actions of triiodothyronine on liver ME, the interaction of diet with glucagon and thyroid hormones is conceivably part of the mechanism responsible for the early appearance of hepatic malic enzyme, whereby the decline of plasma glucagon permits triiodothyronine to act on liver ME. Insulin probably exerts a permissive action subsequently. The manner in which these events relate to the long-term consequences of premature weaning is unknown.