Age-related changes in rat hepatic acetyl-coenzyme A carboxylase

Acetyl-CoA carboxylase (ACC) catalyzes the rate-limiting step in the synthesis of long-chain fatty acids. Since aging influences adiposity, we studied the activity of ACC and its mRNA content in livers of 4-, 12-, and 24-month-old male Fischer 344 rats. The mean (+/- SEM) activity of ACC (mU/mg protein) in liver homogenates from 4-month-old rats was 1.01 +/- 0.14. There was an 80% increase in activity (1.83 +/- 0.27) in 12-month-old rats (P < 0.01). However, there was significantly less activity (0.46 +/- 0.06) in livers of 24-month-old rats (P < 0.001). The total activity of ACC (per g liver) followed the same trend. The enzyme from all age groups was purified by avidin-affinity chromatography. The purified preparation migrated as a major protein band (M(r) 262,000) on sodium dodecyl sulfate (SDS)-polyacrylamide gels. The specific activity of the purified preparation was 1.5, 1.8, and 1.8 U/mg for 4-, 12-, and 24-month-old rats, respectively. The alkali-labile phosphate content was 5.66 +/- 0.17, 5.64 +/- 0.21, and 6.21 +/- 0.35 mols P(i)/mole subunit for 4-, 12-, and 24-month-old rats, respectively. These age-related differences were not significant. The hepatic ACC mRNA measured by ribonuclease protection assay when corrected for G3PDH mRNA was significantly reduced in 24-month-old rats (0.24 +/- 0.03) compared with 12-month-old (0.58 +/- 0.04) or 4-month-old rats (0.43 +/- 0.007) P < 0.01. In summary: (i) Aging in rats is associated with significant changes in ACC activity; (ii) the purified ACC preparations from the three age groups had similar specific activity and similar phosphate content; and (iii) the changes in ACC mRNA content of the liver paralleled the changes in total enzyme activity when 12-month-old rats were compared with 24-month-old rats whereas the increase in ACC activity in 12-month-old rats compared with 4-month-old rats could not be ascribed to changes in hepatic mRNA levels. These results indicate that the age-related changes in hepatic ACC occur at a post-translational level during early years of aging and at a pretranslational level at late states of senescence. These changes may contribute to the age-related alterations in body adiposity.     

Response of leptin mRNA to 24-h food deprivation and refeeding is influenced by age in rats

To obtain an insight into the influence of aging on leptin gene expression, the responses of leptin mRNA in retroperitoneal and epididymal adipose tissues and plasma leptin concentrations to 24-h food deprivation and refeeding were examined in 2-, 10- and 24-month-old normal rats. The basal level of leptin gene expression in retroperitoneal adipose tissue was significantly higher in 10- and 24-month-old rats than that in 2-month-old rats, while the level in epididymal adipose tissue was highest in 10-month-old rats for all three age groups. The basal concentrations of plasma leptin was significantly higher in 10- and 24-month-old rats than those in 2-month-old rats. The 24-h food deprivation was followed by a significant reduction in leptin mRNA expression in both retorperitoneal and epididymal adipose tissues for all three age groups. The leptin gene expression was restored to control levels 24 h following refeeding in the 2- and 10-month-old rats, but failed to be restored in the 24-month-old rats. In addition, the time course of recovery for leptin mRNA expression by refeeding to the control levels differed between the retroperitoneal and the epididymal adipose tissue in 2- and 10-month-old rats. The concentrations of plasma leptin 24 h following refeeding were compatible with the leptin mRNA levels in adipose tissues in three age groups. These results suggest that the expression of the leptin gene in response to food-deprivation and refeeding is influenced by an animal's age and that this expression is different for different regions of white adipose tissue.     

Adaptation of ghrelin levels to limit body weight gain in the obese Zucker rat

In this study, we measured the ghrelin, leptin, and insulin variations in lean and obese Zucker fa/fa rats during the acute phase of body weight gain. At 2 months of age, plasma insulin and leptin concentrations in fa/fa rats were, respectively, 470% and 3700% higher than in lean rats (p <0.0001). Plasma ghrelin was significantly lower (-24.6%; p <0.02) than in lean rats. At 6 months of age, ghrelin increased in both genotypes but the difference was no more significant. The inverse correlations existing between ghrelin and either body weight (BW), insulin or leptin at 2 months of age were no more observable in 6-month-old rats. At 6 months of age, the lean rats had the same body weight as the 2-month-old obese rats. In these body weight-matched rats, ghrelin was not correlated with BW but it remained negatively correlated with insulin and leptin. At the same body weight, obese rats had a much lower plasma ghrelin than lean rats (717+/-42 vs. 1754+/-83 pg/ml; p <0.0001). These data indicate that body composition rather than body weight is the primary factor for the down-regulation of the ghrelin system. This down-regulation constitutes a mechanism of defense of the organism against the development of obesity at least during the first part of life.     

Feeding and temperature responses to intravenous leptin infusion are differential predictors of obesity in rats

Obesity is frequently associated with leptin resistance. The present study investigated whether leptin resistance in rats is present before obesity develops, and thus could underlie obesity induced by 16 wk exposure to a liquid, palatable, high-energy diet (HED). Before HED exposure, male Wistar rats (weighing between 330 and 360 g) received intravenous infusions of 20 microg leptin 2 h before dark (approximately 57 microg/kg rat). Relative to saline infusion, this caused a highly variable effect on food intake (ranging between -94 and +129%), with food intake suppression that appeared negatively correlated with HED-induced increases in body weight gain, caloric intake, adiposity, and plasma leptin levels. In contrast, leptin's thermogenic response was positively correlated to body weight gain linked to weights of viscera, but not to adiposity. Before HED exposure, leptin unexpectedly increased food intake in some rats (fi+, n = 8), whereas others displayed the normal reduction in food intake (fi-, n = 7). HED-exposed fi+ rats had higher plasma leptin levels, retroperitoneal fat pad weight, HED intake, and body weight gain than fi- and chow-fed rats. These parameters were also higher in HED-exposed fi-rats relative to chow rats, except for plasma leptin concentrations. It is concluded that leptin's reduced efficacy to suppress food intake could predict obesity on an HED. An unexpected orexigenic effect of leptin might potentially contribute to this as well.     

Hepatic steatosis and plasma dyslipidemia induced by a high-sucrose diet are corrected by an acute leptin infusion.

High sucrose (HS) feeding in rats induces hepatic steatosis and plasma dyslipidemia. In previous reports (Huang W, Dedousis N, Bhatt BA, O'Doherty RM. J Biol Chem 279: 21695-21700, 2004; and Huang W, Dedousis N, Bandi A, Lopaschuk GD, O'Doherty RM. Endocrinology 147: 1480-1487, 2006), our laboratory demonstrated a rapid ( approximately 100 min) leptin-induced decrease in liver and plasma VLDL triglycerides (TG) in lean rats, effects that were abolished in obese rats fed a high-fat diet, a model that also presents with hepatic steatosis and plasma dyslipidemia. To further examine the capacity of acute leptin treatment to improve metabolic abnormalities induced by nutrient excess, hepatic leptin action was studied in rats after 5 wk of HS feeding. HS feeding induced hepatic steatosis (TG+80+/-8%; P=0.001), plasma hyperlipidemia (VLDL-TG+102+/-14%; P=0.001), hyperinsulinemia (plasma insulin +67+/-12%; P=0.04), and insulin resistance as measured by homeostasis model assessment (+125+/-20%; P=0.02), without increases in adiposity or plasma leptin concentration compared with standard chow-fed controls. A 120-min infusion of leptin (plasma leptin 13.6+/-0.7 ng/ml) corrected hepatic steatosis (liver TG-29+/-3%; P=0.003) and plasma hyperlipidemia in HS (VLDL-TG-42+/-4%; P=0.001) and increased plasma ketones (+45+/-3%; P=0.006), without altering plasma glucose, insulin, or homeostasis model assessment compared with saline-infused HS controls. In addition, leptin activated liver phosphatidylinositol 3-kinase (+70+/-18%; P=0.01) and protein kinase B (Akt; +90+/-29%; P=0.02), and inhibited acetyl-CoA carboxylase (40+/-7%; P=0.04) in HS, further demonstrating that hepatic leptin action was intact in these animals. We conclude that 1) leptin action on hepatic lipid metabolism remains intact in HS-fed rats, 2) leptin rapidly reverses hepatic steatosis and plasma dyslipidemia induced by sucrose, and 3) the preservation of hepatic leptin action after a HS diet is associated with the maintenance of low adiposity and plasma leptin concentrations.     

Fetal leptin is a signal of fat mass independent of maternal nutrition in ewes fed at or above maintenance energy requirements

In adults, circulating leptin concentrations are dependent on body fat content and on current nutritional status. However, the relationships among maternal nutrient intake, fetal adiposity, and circulating leptin concentrations before birth are unknown. We investigated the effects of an increase in nutrient intake in the pregnant ewe on fetal adiposity and plasma leptin concentrations during late gestation. Between 115 and 139-141 days gestation (term = 147 +/- 3 days gestation), ewes were fed a diet calculated to provide either maintenance (control, n = 6) or approximately 155% of maintenance requirements (well-fed, n = 8). The fetal fat depots (perirenal and interscapular) were dissected, and the relative proportion of unilocular and multilocular adipocytes in each depot was determined. Maternal plasma glucose and leptin concentrations were significantly increased in well-fed ewes. Fetal plasma glucose concentrations were also higher in the well-fed group (115-139 days gestation: control, 1.65 +/- 0.14 mmol/L; well-fed, 2.00 +/- 0.14 mmol/L; F = 5.76, P < 0.04). There was no effect of increasing maternal feed intake on total fat mass, the relative mass of unilocular fat, or fetal plasma leptin concentrations (115-139 days gestation: control, 5.2 +/- 0.8 ng/ml; well-fed, 4.7 +/- 0.7 ng/ml). However, in both the control and well-fed groups fetal plasma leptin concentrations (y) were positively correlated with the relative mass of unilocular fat (x): y = 1.51x + 1.70; (R = 0.76, P < 0.01). Thus, fetal leptin may play a role as a signal of unilocular fat mass in the fetus when maternal nutrient intake is at or above maintenance requirements.     

Impact of periconceptional nutrition on maternal and fetal leptin and fetal adiposity in singleton and twin pregnancies

It has been proposed that maternal nutrient restriction may alter the functional development of the adipocyte and the synthesis and secretion of the adipocyte-derived hormone, leptin, before birth. We have investigated the effects of restricted periconceptional undernutrition and/or restricted gestational nutrition on fetal plasma leptin concentrations and fetal adiposity in late gestation. There was no effect of either restricted periconceptional or gestational nutrition on maternal or fetal plasma leptin concentrations in singleton or twin pregnancies during late gestation. In ewes carrying twins, but not singletons, maternal plasma leptin concentrations in late gestation were directly related to the change in ewe weight that occurred during the 60 days before mating [maternal leptin = 0.9 (change in ewe weight) + 7.8; r = 0.6, P < 0.05]. In twin, but not singleton, pregnancies, there was also a significant relationship between maternal and fetal leptin concentrations (maternal leptin = 0.5 fetal leptin + 4.2, r = 0.63, P < 0.005). The relative mass of perirenal fat was also significantly increased in twin fetal sheep in the control-restricted group (6.0 +/- 0.5) compared with the other nutritional groups (control-control: 4.1 +/- 0.4; restricted-restricted: 4.4 +/- 0.4; restricted-control: 4.3 +/- 0.3). In conclusion, the impact of maternal undernutrition on maternal plasma leptin concentrations during late gestation is dependent on fetal number. Furthermore, we have found that there is an increased fetal adiposity in the twins of ewes that experienced restricted nutrition throughout gestation, and this may be important in the programming of postnatal adiposity.     

The influence of aging on the stereoselective pharmacokinetics of propranolol in the rat.

The influence of aging on the pharmacokinetics and the tissue distribution of (R)- and of (S)-propranolol was studied in 3-, 12-, and 24-month-old rats. After both iv and oral administration of rac-propranolol, the plasma concentrations were higher for the (R)- than for the (S)-enantiomer. For the tissue concentrations, the reverse was true. The free fraction of (S)-propranolol in plasma was about 4 times larger than that of (R)-propranolol, and this is the main factor responsible for the differences in kinetics between the two enantiomers. There was a suggestion for a difference in tissue binding between the two enantiomers. With aging, the plasma and tissue concentrations of both enantiomers increase, probably due to a decrease in blood clearance. Tissue binding did not change much with aging. Notwithstanding the marked differences between the kinetics of the propranolol enantiomers, the changes which occur with aging affect both enantiomers to the same degree.     

Effects of TNF-alpha neutralization on adipocytokines and skeletal muscle adiposity in the metabolic syndrome

In a prior study, we have shown that tumor necrosis factor (TNF)-alpha neutralization improves inflammatory markers and total adiponectin in patients with the metabolic syndrome, without improving insulin sensitivity. In this study, we sought to extend our understanding of the effects of TNF-alpha neutralization in this human model of obesity by investigating the responses of high-molecular-weight (HMW) adiponectin, resistin, leptin, and muscle adiposity to etanercept in patients with the metabolic syndrome. Fifty-six men and women with the metabolic syndrome enrolled in a double-blind randomized placebo-controlled trial. Circulating concentrations of total and HMW adiponectin, resistin, and leptin were determined at baseline and after 4 wk of treatment with etanercept. Muscle adiposity was measured by computed tomography (CT). Although etanercept increased total adiponectin concentration, the HMW form, which is thought to mediate insulin sensitivity, was unchanged. Thus the ratio of HMW to total adiponectin decreased following etanercept treatment compared with placebo (-0.03 +/- 0.03 vs. 0.06 +/- 0.03, P = 0.02). Resistin tended to decrease in the etanercept-treated group compared with placebo (-0.6 +/- 0.7 vs. 1.2 +/- 0.7 ng/ml, P = 0.06), whereas leptin was not altered. Etanercept decreased muscle attenuation on CT [-0.61 +/- 0.64 Hounsfield units (HU) vs. 1.54 +/- 0.77 HU in placebo, P = 0.04], suggesting an increase in muscle adiposity. Together, these results demonstrate that neutralization of TNF-alpha in obese humans results in differential effects on critical adipokines and body composition indexes. These findings may help to explain the lack of effect on insulin sensitivity and extend our knowledge of the biological effects of TNF-alpha neutralization in obesity.     

Decreased food intake rather than zinc deficiency is associated with changes in plasma leptin, metabolic rate, and activity levels in zinc deficient rats( small star, filled)

This study investigated the hypothesis that the reduced food intake and poor weight gain in zinc deficient rats is due to: increased plasma leptin concentration, increased physical activity and/or increased metabolic rate. Weanling rats were assigned to three groups: controls fed ad libitum (C), zinc deficient (ZD), and pair-fed controls (PF), and tested in a metabolic chamber and activity monitor at baseline and weekly for four weeks. At the end of the study, all groups were compared for differences in plasma leptin concentrations. ZD and PF animals had markedly reduced food intake and weight gain. ZD had reduced stereotypic and locomotor activity compared to PF animals and both groups demonstrated an abolished peri-nocturnal activity spike and were much less active than controls. This was associated with a reduced total metabolic rate by day 30: ZD (0.73 +/- 0.07 kcal/hr, p = 0.0001) and PF (0.83 +/- 0.06 kcal/hr, p = 0.0001) groups vs. controls (1.82 +/- 0.09 kcal/hr). Plasma leptin concentrations in ZD (1.55 +/- 0.06 &mgr;g/L) were lower than controls (2.01 +/- 0.18 &mgr;g/L, p < 0.03), but neither ZD nor controls were statistically different from PF (1.68 +/- 0.05 &mgr;g/L). Both low leptin concentrations and low metabolic rates in the ZD and PF rats were associated with decreased food intake rather than zinc deficiency. The reduced food intake and poor weight gain observed in zinc deficient rats could not be explained by elevated leptin concentrations, hypermetabolism, or increased activity. Low serum leptin concentrations, hypometabolism, and decreased activity are more likely the result of the anorexia of zinc deficiency.     

Chronic leptin administration promotes lipid utilization until fat mass is greatly reduced and preserves lean mass of normal female rats

Leptin is a hormone synthesized and secreted from adipose tissue. To study the physiologic effects of chronic leptin treatment, normal adult female Sprague-Dawley rats were injected subcutaneously for 35 days. Twice daily injections (250 microgram/day, b.i.d.) resulted in a significant (P<0.05) decrease in food intake that was maintained for 10 days before gradually returning to control level by day 21. Leptin decreased body weight by a maximum of 12% of the initial body weight on day 22 and remained reduced for the duration of the treatment. After 35 days of treatment, visible peritoneal adipose tissue was not detected. Body composition analysis showed that chronic injection of leptin resulted in a dramatic decrease in fat content (28+/-2 to 4+/-2 g, P<0.05; mean+/-SEM) while the lean content remained unchanged. Rats pair-fed to the leptin-treated group but treated with vehicle had the same body composition (23+/-3 g fat mass) as that measured for the ad libitum fed controls. Using indirect calorimetry we observed that leptin decreased respiratory quotient and thus increased fat oxidation. Leptin also prevented energy expenditure reduction typically associated with food restriction. Leptin treatment for 35 days decreased plasma triglyceride (0.75+/-0.07 to 0.30+/-0.03 mM, P<0.05), free fatty acid (0.56+/-0.06 to 0.32+/-0.04 mM) and insulin (3.2+/-0.5 to 1. 4+/-0.4 ng/ml, P<0.05) concentrations despite the fact that food intake was normalized by day 35. Withdrawal of leptin triggered hyperphagia indicating that leptin biology remained throughout the duration of the chronic treatment. These data suggest that leptin reduces fat mass by initially decreasing appetite and by maintaining enhanced fat utilization even when food intake has returned to that of vehicle-treated control.     

Effects of fructose and glucose on plasma leptin, insulin, and insulin resistance in lean and VMH-lesioned obese rats

To determine the influence of dietary fructose and glucose on circulating leptin levels in lean and obese rats, plasma leptin concentrations were measured in ventromedial hypothalamic (VMH)-lesioned obese and sham-operated lean rats fed either normal chow or fructose- or glucose-enriched diets (60% by calories) for 2 wk. Insulin resistance was evaluated by the steady-state plasma glucose method and intravenous glucose tolerance test. In lean rats, glucose-enriched diet significantly increased plasma leptin with enlarged parametrial fat pad, whereas neither leptin nor fat-pad weight was altered by fructose. Two weeks after the lesions, the rats fed normal chow had marked greater body weight gain, enlarged fat pads, and higher insulin and leptin compared with sham-operated rats. Despite a marked adiposity and hyperinsulinemia, insulin resistance was not increased in VMH-lesioned rats. Fructose brought about substantial insulin resistance and hyperinsulinemia in both lean and obese rats, whereas glucose led to rather enhanced insulin sensitivity. Leptin, body weight, and fat pad were not significantly altered by either fructose or glucose in the obese rats. These results suggest that dietary glucose stimulates leptin production by increasing adipose tissue or stimulating glucose metabolism in lean rats. Hyperleptinemia in VMH-lesioned rats is associated with both increased adiposity and hyperinsulinemia but not with insulin resistance. Dietary fructose does not alter leptin levels, although this sugar brings about hyperinsulinemia and insulin resistance, suggesting that hyperinsulinemia compensated for insulin resistance does not stimulate leptin production.     

Increased weight gain after ovariectomy is not a consequence of leptin resistance

The positive correlation between leptin and body fat mass has caused some investigators to speculate that leptin resistance contributes to obesity. Loss of ovarian function in human and rat is associated with increased fat mass gain and increased circulating leptin levels. To study whether ovariectomy produces leptin resistance, Sprague-Dawley female rats were ovariectomized or sham operated and injected with leptin for 35 days. Ovariectomy (OVX) produced hyperphagia and increased gain in both lean and fat mass. Daily leptin injections initially decreased food intake significantly, but feeding gradually increased to a stable level by day 16 and remained at that level for the duration of study. Body composition analysis indicated that chronic injection of leptin to OVX rats dramatically decreased (P < 0.05) fat mass [30 +/- 2 (SE) g, vehicle, to 3 +/- 1 g, leptin]. Using indirect calorimetry, we observed that OVX did not change energy expenditure or total level of fuel utilization. Leptin administration increased fat utilization and prevented reduction in calorie expenditure that is typically associated with food restriction. Leptin treatment to OVX rats decreased plasma triglyceride, free fatty acid, and insulin concentrations, whereas glucose concentration was normal. Withdrawal of leptin triggered hyperphagia, indicating that leptin biology remained throughout the duration of the chronic treatment. The same dose of leptin produced qualitatively similar data in sham-operated rats. Thus we concluded that the loss of ovarian function in rats is not associated with a change in leptin sensitivity.     

Effect of aging on 24-hour pattern of stress hormones and leptin in rats

This work analyzes the 24-hour changes of hypothalamic-pituitary-adrenal (HPA) axis activity and leptin release in aged rats. Three- and 22-month-old male Wistar rats were killed at 6 time intervals during a 24-hour cycle (n=8-10 rats/group). Aging augmented plasma ACTH while it decreased plasma and adrenal gland corticosterone levels. Plasma and adrenal corticosterone levels attained high levels during all the scotophase, concomitantly with the maxima in ACTH levels, whereas in aged rats only a brief plasma corticosterone peak at the early scotophase and no time of day variations of adrenal corticosterone were observed. Aging augmented circulating leptin, with a significant interaction "agextime" in the factorial ANOVA, i.e. only in young rats time of day changes were significant, with the lowest values of leptin at the middle of the light period and higher values at night. When plasma leptin was expressed on body weight basis, the age-related differences became not significant but the daily pattern of plasma leptin found in young rats persisted. Plasma and adrenal corticosterone levels correlated significantly with plasma ACTH only in young rats. Likewise, plasma leptin correlated with plasma corticosterone only in young rats. These changes can be attributed to a disrupting effect of aging on the homeostatic mechanisms modulating HPA activity and leptin release.     

Leptin concentrations in the abdominal subcutaneous adipose tissue of patients with anorexia nervosa assessed by in vivo microdialysis

OBJECTIVE: The adipocyte-derived hormone leptin is involved in energy metabolism and body weight regulation. Plasma leptin concentrations are significantly reduced in patients with anorexia nervosa (AN) and with severe malnutrition. Whether reduced plasma leptin is reflected by its decreased production by the adipose tissue is unknown. METHODS: In the present study we measured leptin concentrations locally in the abdominal subcutaneous adipose tissue of 9 female AN patients and 11 healthy controls by in vivo microdialysis. RESULTS: Adipose tissue free leptin levels were not different in patients with AN compared to controls (2.59+/-1.99 vs 2.36+/-0.25 ng/ml, P>0.05). Plasma leptin soluble receptor (sOb-R) levels were significantly higher in patients with AN than in healthy subjects (58.05+/-38.69 vs 12.79+/-5.08 U/ml, P<0.01). The area of adipocyte in AN was considerably smaller than in the controls (183+/-104.01 microm2 compared to 2145.8+/-1003.41). CONCLUSIONS: We conclude that decreased plasma leptin levels in patients with AN are not directly related to dialysate leptin levels in the abdominal subcutaneous adipose tissue.     

Growth hormone-mediated breakdown of body fat: insulin and leptin responses to GH are modulated by diet composition and caloric intake in old rats.

This work was performed to elucidate whether growth hormone (GH)-mediated loss of adipose tissue and responses in plasma insulin and leptin are modulated by diet composition. 12-month-old rats were first fed a high-fat (HF) diet or a low-fat (LF) diet for 14 weeks. After that, GH or saline was administered to rat groups that were maintained on either HF or LF diets or that were switched from the HF to the LF diet. All 6 groups had free access to food. One additional saline group was pair-fed with the GH group that was switched from the HF to the LF diet. The caloric consumption of this latter group was also translated to yet another GH group receiving restricted amounts of the HF diet. GH was given in a total dose of 4 mg/kg/d for three weeks. After sacrifice, blood was collected and tissues were excised. In groups injected with saline, the weight of excised adipose tissue was 60 +/- 4.7, 41 +/- 3.8 and 50 +/- 4.5 g in animals that continued with the HF diet, LF diet, or that were switched from HF to LF, respectively. Corresponding figures after GH treatment were significantly (p < 0.05) decreased to 38 +/- 2.7, 30 +/- 2.3, and 31 +/- 2.7 g, respectively. Pair-feeding had no effect, whereas only 26 +/- 3.0 g of adipose tissue was retrieved in rats fed restricted amounts of HF diet while receiving GH. In this group, plasma insulin and leptin were also significantly (p < 0.05) depressed compared with other GH groups, especially to the group fed the unrestricted HF diet (203 +/- 35 vs. 1345 +/- 160 pmol/l and 9.3 +/- 1.2 vs. 31 +/- 4.4 micro g/l). In conclusion, this study shows that GH mediates breakdown of adipose tissue under a variety of dietary conditions, and that induction of hyperinsulinemia can be prevented if GH treatment is combined with restricted feeding of a diet which is relatively low in carbohydrates and rich in fat. This will also promote a fall of plasma leptin.     

Quantitative determination of cholesterol 5alpha-, 7alpha-, and 7beta-hydroperoxides in rat skin.

An assay method for determination of cholesterol 5alpha-, 7alpha-, and 7beta-hydroperoxides (ChOOHs) in rat skin using high-performance liquid chromatography (HPLC) with a chemiluminescence detector has been developed. In the assay method, free form and free plus ester forms of ChOOHs could be separately determined by HPLC in combination with the treatment of a tissue extract by cholesterol esterase. Lower limits of quantitation for cholesterol 5alpha-, 7alpha-, and 7beta-hydroperoxides were 0.2, 0.1, and 0.5 nmol/g skin, respectively. This assay method showed that (i) good absolute recoveries of ChOOHs from rat skin (80-90% of radiolabeled ChOOHs added to rat skin); (ii) negligible autoxidation of cholesterol caused by the assay procedure (<9.4x10(-5)% of radiolabeled cholesterol added to rat skin); and (iii) good correlation between ChOOHs added to rat skin and ChOOHs determined, indicating this assay method is applicable to quantify ChOOHs in rat skin. By using this assay method, we observed that (i) cholesterol 5alpha-hydroperoxide was detected in skin of rats pretreated with oral doses of pheophorbide a and subsequent visible irradiation; (ii) concentrations of cholesterol 7-hydroperoxides in skin of rats in an ambient light room were not significantly different from those in a dark room for 12 weeks; and (iii) ultraviolet light B irradiation markedly enhanced the concentrations of cholesterol 7-hydroperoxides in the skin of rats.     

Leptin responses to glucose infusions in obesity-prone rats

The secretion of leptin is dually regulated. In fasting animals, plasma leptin concentrations reflect body fat stores, whereas the incremental leptin response to fasting or refeeding most likely reflects insulin-mediated energy flux and metabolism within adipocytes. Impaired secretion of leptin in either pathway could result in obesity. We therefore measured plasma leptin concentrations in fasted animals and plasma leptin concentrations after an intravenous glucose infusion in a rat model of obesity. Young Sprague-Dawley (S-D) and Fischer 344 (F344) rats had similar percent body fat and fasting glucose and fasting leptin concentrations. However, F344 animals had higher insulin concentrations and leptin responses to intravenous glucose than did the S-D animals. The animals were then fed a control or high-fat diet for 6 wk. High-fat fed animals gained more weight and body fat than did the control fed animals. Control and high-fat fed F344 animals gained approximately 40% (P < 0.0001) more weight and >100% (P < 0.01) more body fat than did the S-D animals. Fasting leptin concentrations and leptin concentrations after intravenous glucose infusions and feeding were more than double (P < 0.05) in F344 animals compared with S-D animals. Whether an animal is fed a control or high-fat diet had little effect on the leptin response to intravenous glucose. In conclusion, young, lean F344 animals, before the onset of obesity, demonstrated a greater acute leptin response to intravenous glucose than similarly lean S-D animals. After a 6-wk diet, F344 animals had a greater percent increase in body weight and insulin resistance and exhibited higher fasting leptin concentrations and a greater absolute leptin response to intravenous glucose compared with the S-D animals. The chronic diet (control or high fat) had little impact on the acute leptin response to intravenous glucose. F344 animals exhibit leptin resistance in young, lean animals and after aging and fat accumulation.     

Leptin in type 2 diabetic or myotonic dystrophic women.

BACKGROUND: Insulin resistance is an important determinant of circulating leptin concentrations in humans, but its independent contribution on plasma leptin levels are controversial. In the present study, we characterized plasma leptin levels and their regulation in women with 2 different insulin resistance states: type 2 diabetes and myotonic dystrophy disease, and in controls. MATERIAL AND METHODS: We studied 3 groups of women: 21 type 2 diabetic patients, 20 myotonic dystrophic patients and a control group of 20 normoglycemic subjects, matched in age and body mass index. Body composition, fasting glucose and insulin, IGF-I, IGF-binding protein-3 and leptin were studied. Body composition was measured using a bioelectrical impedance analyser. Insulin sensitivity (in percentage) was modeled according to a computer-based homeostasis model assessment model. Data are expressed in mean +/- SEM. RESULTS: In both groups of patients, glucose concentrations were higher in type 2 diabetic patients than in myotonic dystrophic patients, and insulin concentrations and insulin sensitivity were similar in the 2 groups of patients (82.4 +/- 18.6% in type 2 diabetic patients vs. 69.7 +/- 9.7% in myotonic dystrophic patients, p = 0.2) and lower than in controls. Serum leptin and leptin/fat mass ratio were higher in myotonic dystrophic patients than in type 2 diabetic patients (30 +/- 4.9 ng/ml vs. 17.7 +/- 2.6 ng/ml, p = 0.03 and 2.32 +/- 0.69 ng/ml/kg vs. 1.07 +/- 0.2 ng/ml/kg, p = 0.02, respectively) or those found in controls. In type 2 diabetic patients, leptin concentrations were correlated with body mass index and body fat, and in myotonic dystrophic patients leptin concentrations were correlated with age, body mass index, fasting insulin and lower insulin sensitivity, whereas leptin concentrations were not correlated with body fat. CONCLUSIONS: These findings suggest that leptin concentrations and regulation in myotonic dystrophic patients are different from type 2 diabetes.