No abstract

Little is known about leptin's interaction with other circulating proteins which could be important for its biological effects. Sephadex G-100 gel filtration elution profiles of 125 Ileptin-serum complex demonstrated 125 I-leptin eluting in significant proportion associated with macromolecules. The 125 I-leptin binding to circulating macromolecules was specific, reversible, and displaceable with unlabeled leptin (ED 50 : 0.73 Ϯ 0.09 nM, mean Ϯ SEM, n ϭ 3). Several putative leptin binding proteins were detected by leptin-affinity chromatography of which either 80-or 100-kD proteins could be the soluble leptin receptor as ‫ف‬ 10% of the bound 125 I-leptin was immunoprecipitable with leptin receptor antibodies.Significantly higher ( P Ͻ 0.001) proportions of total leptin circulate in the bound form in lean (46.5 Ϯ 6.6%) compared with obese (21.4 Ϯ 3.4%) subjects. In lean subjects with 21% or less body fat, 60-98% of the total leptin was in the bound form. Short-term fasting significantly decreased basal leptin levels in three lean ( P Ͻ 0.0005) and three obese ( P Ͻ 0.005) subjects while refeeding restored it to basal levels. The effects of fasting on free leptin levels were more pronounced in lean subjects (basal vs. 24-h fasting: 19.6 Ϯ 1.9 vs. 1.3 Ϯ 0.4 ng/ml) compared with those in obese subjects (28.3 Ϯ 9.8 vs. 14.7 Ϯ 5.3). No significant ( P Ͼ 0.05) decrease was observed in bound leptin in either group. These studies suggest that in obese individuals the majority of leptin circulates in free form, presumably bioactive protein, and thus obese subjects are resistant to free leptin. In lean subjects with relatively low adipose tissue, the majority of circulating leptin is in the bound form and thus may not be available to brain receptors for its inhibitory effects on food intake both under normal and food deprivation states.

We investigated the response of leptin to short-term fasting and refeeding in humans. A mild decline in subcutaneous adipocyte ob gene mRNA and a marked fall in serum leptin were observed after 36 and 60 h of fasting. The dynamics of the leptin decline and rise were further substantiated in a 6-day study consisting of a 36-h baseline period, followed by 36-h fast, and a subsequent refeeding with normal diet. Leptin began a steady decline from the baseline values after 12 h of fasting, reaching a nadir at 36 h. The subsequent restoration of normal food intake was associated with a prompt leptin rise and a return to baseline values 24 h later. When responses of leptin to fasting and refeeding were compared with that of glucose, insulin, fatty acids, and ketones, a reverse relationship between leptin and beta-OH-butyrate was found. Consequently, we tested whether the reciprocal responses represented a causal relationship between leptin and beta-OH-butyrate. Small amounts of infused glucose equal to the estimated contribution of gluconeogenesis, which was sufficient to prevent rise in ketogenesis, also prevented a fall in leptin. The infusion of beta-OH-butyrate to produce hyperketonemia of the same magnitude as after a 36-h fast had no effect on leptin. The study indicates that one of the adaptive physiological responses to fasting is a fall in serum leptin. Although the mediator that brings about this effect remains unknown, it appears to be neither insulin nor ketones.

One of the key mediators of the hypoxic response in animal cells is the hypoxia-inducible transcription factor-1 (HIF-1) complex, in which the ␣-subunit is highly susceptible to oxygen-dependent degradation. The hypoxic response is manifested in many pathophysiological processes such as tumor growth and metastasis. During hypoxia, cells shift to a primarily glycolytic metabolic mode for their energetic needs. This is also manifested in the HIF-1-dependent up-regulation of many glycolytic genes. Paradoxically, tumor cells growing under conditions of normal oxygen tension also show elevated glycolytic rates that correlate with the increased expression of glycolytic enzymes and glucose transporters (the Warburg effect). A key regulator of glycolytic flux is the relatively recently discovered fructose-2,6-bisphosphate (F-2,6-P2), an allosteric activator of 6-phosphofructo-1-kinase (PFK-1). Steady state levels of F-2,6-P2 are maintained by the bifunctional enzyme PFK-2/F2,6-Bpase, which has both kinase and phosphatase activities. Herein, we show that one isozyme, PFKFB3, is highly induced by hypoxia and the hypoxia mimics cobalt and desferrioxamine. This induction could be replicated by the use of an inhibitor of the prolyl hydroxylase enzymes responsible for the von Hippel Lindau (VHL)-dependent destabilization and tagging of HIF-1␣. The absolute dependence of the PFKFB3 gene on HIF-1 was confirmed by its overexpression in VHL-deficient cells and by the lack of hypoxic induction in mouse embryonic fibroblasts conditionally nullizygous for HIF-1␣.

ABSTRACT. Background. Leptin, the protein product of the ob gene, is produced by the adipocyte and seems to function as a link between adiposity, satiety, and activity. Leptin has also been found to be necessary for pubertal development, conception, and pregnancy in mice, and is increased in prepubertal children, independent of adiposity, suggesting a role in childhood growth and development. This study investigated 100 mother/newborn pairs to determine the role of leptin in neonatal development. Placental tissue was assayed for leptin mRNA to evaluate it as a source of leptin production in utero.Methods. One hundred mother/newborn pairs were enrolled in this study. Radioimmunoassay was performed for leptin on maternal venous and newborn cord blood. Leptin concentrations were measured in 43 children in Tanner stages 1 and 2 as a control group. Placental tissue was obtained from five mothers and assayed for leptin mRNA by reverse transcription/polymerase chain reaction (RT/PCR). Human placental cell lines JAR and JEG-3 were also assayed for leptin mRNA expression.Results. Leptin was present in all newborns studied at a mean concentration of 8.8 ng/mL (؎9.6 standard deviations). Leptin concentrations in cord blood correlated with newborn weight (r ‫؍‬ .51), body mass index (BMI) (r ‫؍‬ .48), and arm fat (r ‫؍‬ .42). There was no correlation between leptin and insulin. When statistically covarying for adiposity for newborns and Tanner stages 1 and 2 children, newborns had greater concentrations of leptin (mean, 10.57 ng/mL) than children (mean, 3.04 ng/mL). Leptin was present in all mothers at a mean value of 28.8 ng/mL (؎22.2 standard deviations). Leptin concentration correlated with prepregnancy BMI (r ‫؍‬ .56), BMI at time of delivery (r ‫؍‬ .74), and arm fat (r ‫؍‬ .73). Maternal leptin correlated with serum insulin (r ‫؍‬ .49). There was no correlation between maternal and newborn leptin concentrations. Thirteen percent of newborns had higher leptin concentrations than their mothers. Placental tissue from five separate placentas expressed leptin mRNA at comparable or greater levels than adipose tissue. Two human trophoblastic placental cell lines, JAR and JEG-3, also expressed leptin mRNA. Conclusions.The correlation between leptin and adiposity found in children and adults was also found in newborns. Serum leptin concentrations in newborns were increased more than three-fold compared with children in Tanner stages 1 and 2 when controlling for adiposity, suggesting that leptin concentrations in the newborn are not explained by adiposity alone. Maternal leptin concentrations correlated with measures of adiposity at delivery but did not correlate with newborn adiposity or leptin. Leptin mRNA was expressed both in placental tissue and in two human placental cell lines. These data suggest that leptin has a role in intrauterine and neonatal development and that the placenta provides a source of leptin for the growing fetus. Pediatrics 1997; 100(1). URL: http://www.pediatrics.org/cgi/content/full/ 100/1/e1; leptin, ...