Reduced Perinatal Leptin Availability May Contribute to Adverse Metabolic Programming in a Rat Model of Uteroplacental Insufficiency.

Leptin availability in perinatal life critically affects metabolic programming. We tested the hypothesis that uteroplacental insufficiency and intrauterine stress affect perinatal leptin availability in rat offspring. Pregnant rats underwent bilateral uterine vessel ligation (LIG; n = 14), sham operation (SOP; n = 12), or no operation (controls, n = 14). Fetal livers (n = 180), placentas (n = 180), and maternal blood were obtained 4 hours (gestational day [E] 19), 24 hours (E20), and 72 hours (E22) after surgery. In the offspring, we took blood samples on E22 (n = 44), postnatal day (P) 1 (n = 29), P2 (n = 16), P7 (n = 30), and P12 (n = 30). Circulating leptin (ELISA) was significantly reduced in LIG (E22, P1, P2) and SOP offspring (E22). Postnatal leptin surge was delayed in LIG but was accelerated in SOP offspring. Placental leptin gene expression (quantitative RT-PCR) was reduced in LIG (E19, E20, E22) and SOP (E20, E22). Hepatic leptin receptor (Lepr-a, mediating leptin degradation) gene expression was increased in LIG fetuses (E20, E22) only. Surprisingly, hypoxia-inducible factors (Hif; Western blot) were unaltered in placentas and were reduced in the livers of LIG (Hif1a, E20; Hif2a, E19, E22) and SOP (Hif2a, E19) fetuses. Gene expression of prolyl hydroxylase 3, a factor expressed under hypoxic conditions contributing to Hif degradation, was increased in livers of LIG (E19, E20, E22) and SOP (E19) fetuses and in placentas of LIG and SOP (E19). In summary, reduced placental leptin production, increased fetal leptin degradation, and persistent perinatal hypoleptinemia are present in intrauterine growth restriction offspring, especially after uteroplacental insufficiency, and may contribute to perinatal programming of leptin resistance and adiposity in later life.

Duplicated leptin receptors in two species of eel bring new insights into the evolution of the leptin system in vertebrates.

Since its discovery in mammals as a key-hormone in reproduction and metabolism, leptin has been identified in an increasing number of tetrapods and teleosts. Tetrapods possess only one leptin gene, while most teleosts possess two leptin genes, as a result of the teleost third whole genome duplication event (3R). Leptin acts through a specific receptor (LEPR). In the European and Japanese eels, we identified two leptin genes, and for the first time in vertebrates, two LEPR genes. Synteny analyses indicated that eel LEPRa and LEPRb result from teleost 3R. LEPRb seems to have been lost in the teleost lineage shortly after the elopomorph divergence. Quantitative PCRs revealed a wide distribution of leptins and LEPRs in the European eel, including tissues involved in metabolism and reproduction. Noticeably, leptin1 was expressed in fat tissue, while leptin2 in the liver, reflecting subfunctionalization. Four-month fasting had no impact on the expression of leptins and LEPRs in control European eels. This might be related to the remarkable adaptation of silver eel metabolism to long-term fasting throughout the reproductive oceanic migration. In contrast, sexual maturation induced differential increases in the expression of leptins and LEPRs in the BPG-liver axis. Leptin2 was strikingly upregulated in the liver, the central organ of the reproductive metabolic challenge in teleosts. LEPRs were differentially regulated during sexual maturation, which may have contributed to the conservation of the duplicated LEPRs in this species. This suggests an ancient and positive role of the leptin system in the vertebrate reproductive function. This study brings new insights on the evolutionary history of the leptin system in vertebrates. Among extant vertebrates, the eel represents a unique case of duplicated leptins and leptin receptors as a result of 3R.

Association of leptin/receptor and TNF-α gene variants with adolescent obesity in Malaysia.

BACKGROUND: Leptin (LEP) G-2548A (rs7799039), leptin receptor (LEPR) Q223R (rs1137101) and tumor necrosis factor (TNF)-α G-308A (rs1800629) gene variants have been reported to be associated with obesity, although results for subjects from different countries have been controversial. The aim of this study was to determine the prevalence of overweight and obesity in Malaysian adolescents and the association of these polymorphisms with overweight and obese or over-fat adolescents. METHODS: A total of 613 adolescents (241 Malay, 219 Chinese, 153 Indian) were enrolled. Anthropometric measurements of body mass index (BMI) and body fat percentage were used to classify subjects as controls (non-overweight/obese or normal fat) or as cases (overweight/obese or over-fat). Genomic DNA was extracted from oral buccal mucosa cells for genotyping using polymerase chain reaction-restriction fragment length polymorphism and data obtained were statistically analyzed. RESULTS: A total of 23.3% of subjects were overweight/obese whereas 11.4% were over-fat; there were significantly more overweight/obese and over-fat Indian and Malay adolescents compared to Chinese (P < 0.001). A allele was the minor one for LEPR Q223R and TNF-α G-308A in all ethnic groups, whereas G allele was minor for LEP G-2548A in Chinese and Malay adolescents, except for Indian adolescents. Indian male adolescents with AA genotype for LEP G-2548A were associated with overweight/obesity (P = 0.025; odds ratio, 3.64; 95% confidence interval: 1.15-11.54). Despite the lack of association observed for LEPR Q223R and TNF-α G-308A, Indian and Chinese subjects with AA risk genotype for LEPR Q223R/LEP G-2548A and TNF-α G-308A/LEP G-2548A, respectively, had increased mean BMI (P = 0.049, P = 0.016). CONCLUSIONS: Genotype distribution and association of these polymorphisms with overweight/obesity vary between ethnic groups and genders. Nevertheless, the LEP G-2548A risk allele may be associated with overweight/obese Indian male adolescents in Malaysia.

Role of leptin receptors in granulosa cells during ovulation.

Leptin is an important hormone influencing reproductive function. However, the mechanisms underpinning the role of leptin in the regulation of reproduction remain to be completely deciphered. In this study, our objective is to understand the mechanisms regulating the expression of leptin receptor (Lepr) and its role in ovarian granulosa cells during ovulation. First, granulosa cells were collected from superovulated mice to profile mRNA expression of Lepr isoforms (LeprA and LeprB) throughout follicular development. Expression of LeprA and LeprB was dramatically induced in the granulosa cells of ovulating follicles at 4 h after human chorionic gonadotropin (hCG) treatment. Relative abundance of both mRNA and protein of CCAAT/enhancer-binding protein ß (Cebpß) increased in granulosa cells from 1 to 7 h post-hCG. Furthermore, chromatin immunoprecipitation assay confirmed the recruitment of Cebpß to Lepr promoter. Thus, hCG-induced transcription of Lepr appears to be regulated by Cebpß, which led us to hypothesise that Lepr may play a role during ovulation. To test this hypothesis, we used a recently developed pegylated superactive mouse leptin antagonist (PEG-SMLA) to inhibit Lepr signalling during ovulation. I.p. administration of PEG-SMLA (10 µg/g) to superovulated mice reduced ovulation rate by 65% compared with control treatment. Although the maturation stage of the ovulated oocytes remained unaltered, ovulation genes Ptgs2 and Has2 were downregulated in PEG-SMLA-treated mice compared with control mice. These results demonstrate that Lepr is dramatically induced in the granulosa cells of ovulating follicles and this induction of Lepr expression requires the transcription factor Cebpß. Lepr plays a critical role in the process of ovulation by regulating, at least in part, the expression of the important genes involved in the preovulatory maturation of follicles.

Transcriptional Characterization of Porcine Leptin and Leptin Receptor Genes.

The leptin (LEP) and its receptor (LEPR) regulate food intake and energy balance through hypothalamic signaling. However, the LEP-LEPR axis seems to be more complex and its expression regulation has not been well described. In pigs, LEP and LEPR genes have been widely studied due to their relevance. Previous studies reported significant effects of SNPs located in both genes on growth and fatness traits. The aim of this study was to determine the expression profiles of LEP and LEPR across hypothalamic, adipose, hepatic and muscle tissues in Iberian x Landrace backcrossed pigs and to analyze the effects of gene variants on transcript abundance. To our knowledge, non porcine LEPR isoforms have been described rather than LEPRb. A short porcine LEPR isoform (LEPRa), that encodes a protein lacking the intracellular residues responsible of signal transduction, has been identified for the first time. The LEPRb isoform was only quantifiable in hypothalamus while LEPRa appeared widely expressed across tissues, but at higher levels in liver, suggesting that both isoforms would develop different roles. The unique LEP transcript showed expression in backfat and muscle. The effects of gene variants on transcript expression revealed interesting results. The LEPRc.1987C>T polymorphism showed opposite effects on LEPRb and LEPRa hypothalamic expression. In addition, one out of the 16 polymorphisms identified in the LEPR promoter region revealed high differential expression in hepatic LEPRa. These results suggest a LEPR isoform-specific regulation at tissue level. Conversely, non-differential expression of LEP conditional on the analyzed polymorphisms could be detected, indicating that its regulation is likely affected by other mechanisms rather than gene sequence variants. The present study has allowed a transcriptional characterization of LEP and LEPR isoforms on a range of tissues. Their expression patterns seem to indicate that both molecules develop peripheral roles apart from their known hypothalamic signal transduction function.

Postnatal growth after intrauterine growth restriction alters central leptin signal and energy homeostasis.

Intrauterine growth restriction (IUGR) is closely linked with metabolic diseases, appetite disorders and obesity at adulthood. Leptin, a major adipokine secreted by adipose tissue, circulates in direct proportion to body fat stores, enters the brain and regulates food intake and energy expenditure. Deficient leptin neuronal signalling favours weight gain by affecting central homeostatic circuitry. The aim of this study was to determine if leptin resistance was programmed by perinatal nutritional environment and to decipher potential cellular mechanisms underneath.We clearly demonstrated that 5 months old IUGR rats develop a decrease of leptin sentivity, characterized by no significant reduction of food intake following an intraperitoneal injection of leptin. Apart from the resistance to leptin injection, results obtained from IUGR rats submitted to rapid catch-up growth differed from those of IUGR rats with no catch-up since we observed, for the first group only, fat accumulation, increased appetite for food rich in fat and increased leptin synthesis. Centrally, the leptin resistant state of both groups was associated with a complex and not always similar changes in leptin receptor signalling steps. Leptin resistance in IUGR rats submitted to rapid catch-up was associated with alteration in AKT and mTOR pathways. Alternatively, in IUGR rats with no catch-up, leptin resistance was associated with low hypothalamic expression of LepRa and LepRb. This study reveals leptin resistance as an early marker of metabolic disorders that appears before any evidence of body weight increase in IUGR rats but whose mechanisms could depend of nutritional environment of the perinatal period.

Adipocytokine expression in placentas from pre-eclamptic and chronic hypertensive patients.

OBJECTIVE: Growing evidence suggests that concentration of the adipocytokines leptin and adiponectin may be affected by risk of hypertension during pregnancy. Leptin and leptin receptor gene expression has been studied in placentas obtained from pre-eclamptic patients, but not in those with chronic high blood pressure (CHBP). Adiponectin receptors remain unstudied in placentas obtained from hypertensive patients. METHODS: Therefore, we investigated relative mRNA expression of selected adipocytokine genes (leptin, leptin receptors (LEPRA, LEPRB, LEPRC, LEPRD) and adiponectin receptors (ADIPOR1, ADIPOR2)) in placental tissues from women with pre-eclampsia (n = 6) or CHBP (n = 8). Placentas from 28 normotensive patients were analyzed as controls. mRNA extracted from biopsies taken from the maternal and fetal sides of the placenta was investigated using real-time polymerase chain reaction. RESULTS: Compared with controls, significant increases in leptin mRNA expression were seen in placentas from pre-eclamptic patients on the maternal (p = 0.01) and fetal (p = 0.02) sides, and in placentas from CHBP mothers on the fetal side (p = 0.001). Maternal-side tissue from CHBP patients was not significantly different from that of controls (p = 0.08), but this might be due to the small sample size. No significant differences were seen in mRNA expression for most of the adipocytokine receptors tested for hypertensive cases compared with controls. However, there was a decrease in LEPRC (pre-eclamptic, maternal side, p = 0.03) and LEPRD (pre-eclamptic, maternal side, p = 0.01; CHBP, fetal side, p = 0.009) in case-control analysis. CONCLUSIONS: This pilot study shows that increases seen in leptin expression in placentas from hypertensive mothers might be a consequence of defects in placentation associated with this disease, and motivates further region-specific adipocytokine gene expression analysis across this organ.