Effects of central infusion and immunoneutralization of growth hormone on the timing of puberty and plasma leptin levels in the female rat

Growth hormone (GH) levels increase during puberty though its role in puberty onset is still unclear. An interaction is suggested between GH and leptin, as triggering factor of puberty. To evaluate the role of GH on the timing of puberty and its relation with leptin, we centrally administered recombinant human GH (rhGH; 1 microg/day) to normally fed or food-restricted (FR) prepubertal female rats, and monitored time of vaginal opening (VO). Median time of VO was equally postponed in FR animals and in normally fed rhGH-infused rats: median time of VO was respectively 35 and 34 vs. 27 d. Central infusion of rhGH in FR rats partially restored the delay in VO. Plasma leptin levels were increased in rhGH-infused animals, normally fed or FR. Centrally infused anti-rat GH (0.6 microg/day) did not affect plasma leptin levels, but advanced median time of VO (25 vs. 28 d) in pair-fed female rats but not in ad lib-fed animals. The effects of the centrally infused compounds appear to depend on the dietary regime imposed on the prepubertal animals. Furthermore, plasma leptin levels show no direct or predictive relation to the time of VO. The data indicate an involvement of GH in puberty onset, but do not explain the mechanism employed.     

Growth hormone-releasing factor (GRF) induced growth hormone advances puberty in female buffaloes

Exogenous bovine growth hormone-releasing factor (bGRF) at the dose rate of 10 microg/100 kg body weight was administered intravenously (i.v.) to six Murrah buffalo heifers as treatment group, while another six buffalo heifers served as control group which received the vehicle (0.9% NaCl solution) at an interval of 15 days for a period of 9 months to study the effect of bGRF on puberty onset associated with temporal hormonal changes in peri-pubertal buffalo heifers. Blood samples were collected at 3-day interval from all the animals during the experimental period and plasma harvested was assayed for growth hormonal (GH), luteinizing hormone (LH) and progesterone. The day that plasma progesterone was greater than 1.0 ng/ml for three consecutive sampling days was defined as the day of puberty. Exogenous bGRF administration increased (P = 0.02) plasma GH concentration in treatment group over control group during the treatment of bGRF as well as during the peri-pubertal period. Plasma progesterone concentrations increased transiently earlier (P = 0.05) by 58.5 days in bGRF-treated buffaloes than that in the control group. However, plasma LH concentrations were unaffected by the treatment of bGRF (P = 0.48). Both plasma GH and LH in the buffalo heifers increased (P < 0.01) over time preceding puberty and the higher hormonal concentrations were maintained during the onset of puberty, and thereafter, the concentrations of both the hormones declined (P < 0.05) after puberty. GH and LH were positively correlated both before puberty (r = +0.59 and +0.63; P < 0.05 for control and treatment group, respectively) and after puberty (r = +0.42 and +0.46; P < 0.05 for control and treatment group, respectively) indicating the interaction and/or close relationship of GH and LH in the mechanism of puberty in buffalo species.     

Leptin gene expression, circulating leptin, and luteinizing hormone pulsatility are acutely responsive to short-term fasting in prepubertal heifers: relationships to circulating insulin and insulin-like growth factor I(1)

In the present study, we tested the hypothesis that short-term fasting would reduce leptin gene expression, circulating leptin, and LH pulsatility in prepubertal heifers in association with a decrease in circulating concentrations of insulin and insulin-like growth factor I (IGF-I). Twelve prepubertal crossbred heifers (mean +/- SD = 315 +/- 5 kg body weight) were assigned randomly to one of two treatments in two replicates: 1) control; normal feed consumption (n = 6) and 2) fasted; 48 h of total feed restriction (n = 6). Blood samples were collected at 15-min intervals for 8 h on Days 0 and 2 of the experiment and twice on Day 1. Subcutaneous fat samples were collected before treatment onset (Day -1) and at the end of the intensive blood sampling on Day 2. Acute feed restriction markedly reduced leptin mRNA in adipose tissue (P < 0.01) and circulating concentrations of leptin (P < 0.05), IGF-I (P < 0.01), and insulin (P = 0.05) as compared with controls on Day 2. Moreover, the treatment x day interaction (P < 0.076) and within-day contrasts (expressed as a percentage of Day 0 values) revealed that the mean frequency of LH pulses in the fasted group was lower (P < 0.06) than in controls on Day 2. Neither mean concentrations of growth hormone (GH) nor GH secretory dynamics were affected by acute feed restriction. Fasting-mediated decreases in leptin gene expression and circulating leptin, in association with reductions in secretion of IGF-I, insulin, and LH, provide a basis for investigating leptin as a hormone signaling energy status to the central reproductive axis in cattle.     

The consequences of altered somatotropic system on reproduction

Although the primary control of gonadotropin secretion is by the hypothalamic GnRH and the gonadal function is controlled by the pituitary gonadotropins and prolactin, the emerging evidence suggests a vital role of the somatotropic axis, growth hormone (GH), and insulin-like growth factor-I (IGF-I) in the control of the pituitary and gonadal functions. It has been shown that GH deficiency, GH resistance, and experimental alterations in IGF-I secretion modify folliculogenesis, ovarian maturation, ovulation, and pregnancy, and in the male, GH/IGF-I plays an important role in spermatogenesis and the Leydig cell function. The primary focus of this review is to examine the role of GH/ IGF-I on the onset of puberty, fertility, pituitary, and gonadal endocrine functions. A number of studies have revealed that fertility is affected in GH-deficient dwarf and in IGF-I gene-ablated mice, possibly due to subnormal function of either the pituitary gland or the gonads. In the female GH receptor gene knockout (GHR-KO) mice, there was impairment in follicular development, ovulation rate, sexual maturation, production of and responsiveness to pheromonal signals, and the corpus luteum function. In IGF-I-deficient male GHR-KO mice, puberty is delayed, spermatogenesis is affected, and neuroendocrine-gonadal function is attenuated. Similarly, in some of the human Laron syndrome patients, puberty is delayed due to GH resistance. These data suggest that, in addition to GnRH and gonadotropins, GH/IGF-I influences the pituitary and gonadal functions in animals and humans.     

Normal growth spurt and final height despite low levels of all forms of circulating insulin-like growth factor-I in a patient with acid-labile subunit deficiency

BACKGROUND: In a recently described patient with acid-labile subunit (ALS) deficiency, the inability to form ternary complexes resulted in a marked reduction in circulating total insulin-like growth factor (IGF)-I, whereas skeletal growth was only marginally affected. To further study the role of circulating versus locally produced IGF-I in skeletal growth in this patient, we now describe in detail growth changes and their relationship with several components of the circulating IGF system. DESIGN AND METHODS: We followed growth and development up to the final height in a patient with complete ALS deficiency and determined both spontaneous and growth hormone (GH)-stimulated changes in the IGF system, including measurements of total, free and bioactive IGF-I, total IGF-II and insulin-like growth factor binding protein (IGFBP)-1, IGFBP-2 and IGFBP-3. RESULTS: The patient had a delayed growth and pubertal onset. Six months of GH treatment had no effect on growth. At the age of 19.3 years, he spontaneously completed puberty and had a normal growth spurt for a late adolescent (peak height velocity of 8.4 cm/year). A normal final height was attained at 21.3 years (167.5 cm; -0.78 SDS). During as well as after puberty, basal levels of total, free and bioactive IGF-I were low, as were total IGF-II, IGFBP-1, IGFBP-2 and IGFBP-3. GH treatment for 6 months normalized free IGF-I and increased bioactive IGF-I, but had no effect on growth velocity. CONCLUSIONS: This case story shows that in the presence of complete ALS deficiency, a height within normal limits can be obtained despite low levels of all forms of circulating IGF-I. Furthermore, the patient presented a delayed but normal growth spurt without any marked increment of circulating IGF-I.     

Up-regulation of lymphocytic growth hormone secretion during the luteal phase of cycle and early pregnancy

Growth hormone (GH) has been shown to be produced and secreted by peripheral immune cells. Therefore, we studied the release of GH by lymphocytes, during various stages of pregnancy and estrous cycle in the cow. The effect of leptin on the lymphocytic GH release was also investigated. Estradiol-17β and progesterone concentrations in plasma were measured in all animals to confirm their reproductive status. Growth hormone levels measured in cell cultures during early pregnancy (days 60-80) and during the luteal phase were greater (p ≤ 0.01) than levels during follicular phase or mid (days 100-160) and late (days 240-245) pregnancy. Leptin treatment stimulated (p ≤ 0.05) lymphocytic GH release during mid-pregnancy when the basal GH levels were low. Changes in lymphocytic GH release and elevation of lymphocytic GH secretion by leptin during pregnancy and the absence of such effects in estrous cycle may indicate that leptin modulation of lymphocytic GH plays a role in the regulation of immune response during pregnancy.     

Gonadotropins increase concentrations of immunoreactive insulin-like growth factor-I in porcine follicular fluid in vivo

To evaluate the regulation of ovarian insulin-like growth factor-I (IGF-I) during follicular growth in vivo, we measured the concentration of this peptide in follicular fluid (FFL) of immature gilts during the induction of follicular development by pregnant mare's serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG). FFL concentrations of immunoreactive (i) IGF-I were compared with those of intrafollicular steroids and with concentrations of iIGF-I, estradiol (E2), and porcine growth hormone (GH) in serum. PMSG, administered at Time 0, induced a significant (p less than 0.01), time-dependent increase in intrafollicular iIGF-I that peaked 72 h after administration of the hormone, before the administration of hCG. During the first 72 h, the changes in ovarian iIGF-I paralleled those for progesterone and E2. After the administration of hCG at 72 h, FFL levels of E2 fell, those of iIGF-I remained constant, and progesterone rose. Serum E2 concentrations paralleled those in FFL. Since serum GH and IGF-I levels rise during spontaneous puberty in some species, these levels were also monitored. However, a significant treatment effect on serum GH and iIGF-I was not demonstrated. In summary, ovarian concentrations of iIGF-I are increased by gonadotropic hormones in vivo. The absence of concomitant changes in circulating levels of iIGF-I and GH suggests that the gonadotropin effects are exerted directly on the ovary. These results, together with more abundant data regarding secretion and action of IGF-I in cultured granulosa cells, suggest that IGF-I may function in an autocrine or paracrine fashion to amplify the actions of gonadotropins at an ovarian level.     

The role of insulin-like growth factor-I in neuroendocrine function and the consequent effects on sexual maturation: inferences from animal models

It is known that growth hormone (GH) plays an important role in growth and development.Additionally, emerging evidence suggest that it also influences hypothalamic-pituitary-gonadal function. We have found that GH from different species has different effects in mice. In rodents, human GH (hGH) binds to both GH and prolactin (PRL) receptors; it has both somatotrophic and lactotrophic effects. Since PRL has a profound effect on neuroendocrine function, the results obtained from hGH treatment or from transgenic animals expressing the hGH gene reflect PRL-like effects of this hormone. However, bovine GH (bGH) is purely somatogenic and therefore the effects of bGH represent the function of the natural GH produced in rodents. Furthermore, our studies in mice and rats have shown that not all effects of GH are stimulatory and the duration of exposure of the hypothalamo-hypophyseal-gonadal system to GH might influence the secretions of gonadotropins and gonadal steroids. In humans, excess productions of GH in acromegaly and GH resistance in Laron syndrome adversely affect reproduction. Similarly, it has been demonstrated that in transgenic mice expressing various GH genes, in insulin-like growth factor-I (IGF-I) gene-knockout mice, in GH receptor gene-disrupted (GHR-KO) mice, and in Ames dwarf mice the onset of puberty and/or fertility is altered. Therefore, excess or subnormal secretion of GH can affect reproduction. We have shown that the hypothalamic-pituitary functions are affected in transgenic mice expressing the GH genes, Ames dwarf mice and in GH receptor gene knockout mice. The majority of the GH effects are mediated via IGF-I and the aforementioned effects may be due to the GH-induced IGF-I secretion or due to the absence of this peptide production. It is important to realize that the syntheses and actions of IGF binding proteins are controlled by IGF-I. Furthermore, some IGF binding proteins can inhibit IGF-I action. Therefore, the concentrations of IGF binding proteins and the ratio of these binding proteins and IGF-I within the body might play a pivotal role in modulating IGF-I effects on the neuroendocrine-gonadal system.     

Metabolic consequences of growth hormone treatment in paediatric practice

No metabolic side-effects of clinical significance have been reported during a 5-year study of growth hormone (GH) therapy in children with GH deficiency, Turner syndrome, idiopathic short stature or chronic renal insufficiency. In particular, insulin levels increase but remain within the normal range, as do glucose and haemoglobin A(1c). A recent study showed that the effects of growth on insulin sensitivity in prepubertal children with idiopathic short stature represent the changes in carbohydrate tolerance observed during normal adolescence. Thus, GH treatment may lead to prolongation of the physiological state of insulin resistance observed in normal puberty. Insulin levels during the fasting state and 2 h after a standard glucose load showed no further rise after the first 3 years of continuous GH therapy. The hyperinsulinaemia observed during GH therapy may, therefore, amplify the anabolic effects of insulin on protein metabolism during puberty.     

Time course of changes in plasma concentrations of the growth related hormones during protein restriction in the domestic fowl (Gallus domesticus)

Experiments were conducted to evaluate the possible role of circulating growth hormones triiodothyronine (T3), thyroxine (T4), and insulin-like growth factor I (somatomedin-C; IGF-I) in the elevation of plasma growth hormone (GH) which occurs in protein-restricted chickens. Plasma hormone changes were determined over a 2-week period of protein depletion by feeding a 5% protein diet as well as a similar period of protein repletion with a 20% protein diet. The rise in plasma GH was observed in two separate studies. Plasma concentrations of T4, T3, and IGF-I were all depressed in protein-restricted chicks prior to or concurrent with the GH elevation. In the protein repletion time course study, T4 and T3 concentrations were normalized prior to or concurrent with plasma GH normalization. However, IGF-I concentrations in repleted chicks did not return to control levels until after normal levels of GH were observed. These data suggest that thyroid hormones may play a greater role in the regulation of GH secretion during periods of malnourishment than IGF-I; the latter being currently thought to be a peripherally circulating inhibitor of GH release in animals.     

Prepubertal OVX increases IGF-I expression and bone accretion in C57BL/6J mice

It is generally well accepted that the pubertal surge in estrogen is responsible for the rapid bone accretion that occurs during puberty and that this effect is mediated by an estrogen-induced increase in growth hormone (GH)/insulin-like growth factor (IGF) action. To test the cause and effect relationship between estrogen and GH/IGF, we evaluated the consequence of ovariectomy (OVX) in prepubertal mice (C57BL/6J mice at 3 wk of age) on skeletal changes and the GH/IGF axis during puberty. Contrary to our expectations, OVX increased body weight (12-18%), bone mineral content (11%), bone length (4%), bone size (3%), and serum, liver, and bone IGF-I (30-50%) and decreased total body fat (18%) at 3 wk postsurgery. To determine whether estrogen is the key ovarian factor responsible for these changes, we performed a second experiment in which OVX mice were treated with placebo or estrogen implants. In addition to observing similar results compared with our first experiment, estrogen treatment partially rescued the increased body weight and bone size and completely rescued body fat and IGF-I levels. The increased bone accretion in OVX mice was due to increased bone formation rate (as determined by bone histomorphometry) and increased serum procollagen peptide. In conclusion, contrary to the known estrogen effect as an initiator of GH/IGF surge and thereby pubertal growth spurt, our findings demonstrate that loss of estrogen and/or other hormones during the prepubertal growth period effect leads to an increase in IGF-I production and bone accretion in mice.     

Serum leptin, insulin-like growth factor-I components and sex-hormone binding globulin. Relationship with sex, age and body composition in healthy population

Leptin plays an important role in the regulation of food intake and thermogenesis, regulates long term energy balance and reproductive function and its concentrations are closely linked to body mass index. Leptin secretion is influenced by many factors and the age-related changes in different hormones might modify circulating leptin concentrations. Sex dimorphism in leptin concentrations has been clearly shown in previous studies and its concentrations were lower in men than in women in all decades of life. Insulin growth factor-I (IGF-I) is a peptide growth factor that is present in all types of physiologic fluids and is also produced by connective tissue cell types and its autocrine/paracrine secretion is nearly always present within tissues. There is a physiological decline of the growth hormone (GH)/IGF-I axis with ageing and in addition, insulin, thyroid hormones and the supply of dietary energy may directly regulate the circulating levels of the IGFs and growth hormone binding protein (GHBP). Furthermore, there is no doubt that GH participates in the regulation of body composition, and with advanced age there is a decrease in muscle and an increase in adiposity associated with a decline in GH and total IGF-I. The biological activities of the IGF ligands are modulated by the family of high affinity GHBP. Sex hormone binding globulin (SHBG) concentrations are thought to be regulated primarily through opposing actions of sex steroids on hepatic SHBG production, with oestrogen stimulating and androgen inhibiting SHBG production, and thyroid hormones are also a potent stimulator of SHBG production concentrations. Some studies support an independent IGFBP3 contribution to SHBG variability and these findings are compatible with the hypothesis that some of the anabolic effects ascribed to the GH/IGF axis may be caused by SHBG-mediated changes in testosterone activity or SHBG/total testosterone index.     

Alterations in the growth hormone-insulin-like growth factor axis in insulin dependent diabetes mellitus.

The growth hormone (GH)-insulin-like growth factor (IGF) axis and insulin are major anabolic effectors in promoting weight gain and linear growth. These two anabolic systems are interlinked at many levels, thus abnormalities in one of these systems effect the other causing disordered metabolic homeostasis. Insufficient portal insulinization in insulin dependent diabetes mellitus (IDDM) results in hepatic GH resistance and increased production of IGF-binding proteins-1 (IGFBP-1) and IGFBP-2. GH resistance is reflected by decreased hepatic IGF-I production. In addition, changes in other GH-dependent proteins are also observed in IDDM. Increased proteolysis of IGFBP-3 results in reduction of intact IGFBP-3. Serum ALS levels are also slightly diminished in untreated diabetic patients. Hepatic resistance to GH is, at least in part, caused by diminished GH receptors as reflected by diminished circulating GHBP levels. In addition, there is also evidence from experimental and human studies suggesting post-receptor defect(s) in GH action. As a result of these changes, circulating total and free IGF-I levels are decreased during insulinopenia. Lack of negative feed-back effect of IGF-I on GH secretion causes GH hypersecretion which increases hyperglycemia by decreasing sensitivity to insulin. GH hypersecretion in poorly controlled diabetic patients may play a role in the pathogenesis of diabetic vascular complications. Most of these abnormalities in the GH-IGF axis in diabetes are reversed by effective insulinization of the patient. Addition of IGF-I treatment to insulin in adolescents with IDDM allows correction of GH hypersecretion, improves insulin sensitivity and glycemic control, and decreases insulin requirements. The effect of IGF-I treatment on diabetic complications has yet to be seen.     

Nutrition-induced differences in body composition, compensatory growth and endocrine status in growing pigs

In this experiment, we assessed the effect of amino acid (AA) intake restriction in entire male Yorkshire pigs between 15 and 38 kg BW (restriction phase) on BW gain, body composition and plasma levels of blood urea nitrogen (BUN), cortisol, insulin-like growth factor I (IGF-I), growth hormone (GH) and leptin during the subsequent re-alimentation phase. During the restriction phase, 36 pigs were allotted to one of two dietary treatments: adequate AA intake (control) or AA-limiting diets (AA-30%). Thereafter, pigs were fed common non-limiting diets up to 110 kg BW. Throughout the experiment, pigs were scale-fed at 90% of the estimated voluntary daily digestible energy intake. At the end of the restriction phase, pigs on AA-30% had lesser BW gain (650 v. 784 g/day; P < 0.001), loin area (LA; 12.2 v. 14.2 cm2; P < 0.001), BUN (4.6 v. 6.3 mg/dl; P < 0.02), lesser plasma levels of IGF-I (440 v. 640 ng/m; P < 0.001) and cortisol (8.2 v. 19.2 μg/dl; P < 0.001), greater backfat thickness (BF; 7.56 v. 6.56 mm; P < 0.02), and greater plasma levels of leptin (2.7 v. 1.8 ng/ml; P = 0.027) and GH (3.3 v. 2.0 ng/ml; P = 0.05) than pigs on control. During the re-alimentation phase, previously restricted pigs showed full compensatory growth (CG) in terms of BW gain (1170 v. 1077 g/day; P < 0.002), whole-body protein deposition (Pd) (179 v. 163 g/day; P < 0.001) as well as physical and chemical body composition (whole-body lipid to body protein mass ratio, LB/PB; 1.14 v. 1.15; P > 0.10). Besides GH at 45 kg BW (4.2 v. 2.4 ng/ml; P = 0.066), there were no effects of previous AA intake restriction on leptin, IGF-I and BUN during the re-alimentation phase (P > 0.10). Plasma cortisol and IGF-I levels may act as an indicator of AA-induced restriction in Pd in growing pigs. Plasma BUN level does not appear as a sensitive indicator for compensatory Pd. Plasma leptin and GH levels allow for the involvement of the brain in controlling chemical body composition. Full CG was observed during the energy-dependent phase of Pd in growing pigs and might be driven by a target LB/PB, possibly mediated via plasma leptin, IGF-I and GH levels.     

Inhibitory action of leptin on early follicular growth differs in immature and adult female mice.

In order to investigate the action of leptin on early follicular growth, preantral follicles, 95-115 microm in diameter were mechanically isolated from the ovaries of BDF1 hybrid immature (11-day-old) and adult (8-wk-old) mice, and cultured for 4 days in vitro. Follicular growth was assessed by daily changes in follicular diameter and by the amount of estradiol and immunoreactive (IR)-inhibin released into the culture medium at Day 4. Preantral follicles from immature mice showed a significant development in follicular growth as a result of stimulation by GH (1 mIU/ml), insulin-like growth factor (IGF)-I (100 ng/ml) + FSH (100 mIU/ml), and GH (1 mIU/ml) + FSH (100 mIU/ml). Although leptin at concentrations of 1-1000 ng/ml did not have any significant effect on follicular growth stimulated by IGF-I or GH, it significantly inhibited follicular growth in a dose-related manner when follicles were stimulated by IGF-I + FSH and GH + FSH, respectively, suggesting that leptin attenuated the additive effect of FSH. On the other hand, preantral follicles from adult mice were cultured in the presence of FSH, and FSH-dependent follicular growth was inhibited by leptin in a dose-related manner. Because FSH stimulates cAMP production, we investigated the involvement of cAMP in the inhibitory mechanisms of leptin. Preantral follicles from immature and adult mice were cultured in the presence of either 8-Br-cAMP or forskolin. Both 8-Br-cAMP and forskolin significantly increased follicular diameter and hormone secretion in both immature and adult mice. However, 8-Br-cAMP and forskolin-stimulated follicle growth and hormone secretion were significantly inhibited in immature mice by coadministration of leptin, whereas growth of preantral follicles from adult mice was not inhibited by addition of leptin to cultures. These results indicate that leptin causes an inhibitory effect on the early follicular development of both immature and adult mice, but the inhibitory mechanisms of leptin are different.     

Efficacy and safety of long-term continuous growth hormone treatment of children born small for gestational age

Twelve years of growth hormone (GH) therapy of short children born small for gestational age (SGA) have demonstrated that GH is an effective and well-tolerated therapy. Most children will reach a normal adult height (AH). AH of 55 SGA adolescents was comparable for those treated with a GH dose of 1 or 2 mg/m2 (approximately 0.033 or 0.066 mg/kg) per day, mean (SD) AH SDS being -1.2 (0.7) and -0.8 (0.7), respectively. GH therapy had no influence on the age at onset, the progression of puberty, duration of puberty and pubertal height gain. GH therapy induced higher fasting and glucose-stimulated insulin levels after 1 and 6 years, but 6 months after GH stop, all levels returned to normal. At baseline mean systolic blood pressure was significantly increased, but both systolic and diastolic blood pressure decreased significantly during 6 years of GH and remained so after GH stop. GH therapy demonstrated a beneficial effect on serum lipid profiles, body composition, bone mineral density and head growth. Treatment with 2 mg GH/m2 per day induced mean serum IGF-I levels of +2 SDS, whereas IGF-I levels remained within the normal range with 1 mg GH/m2 per day. In conclusion, long-term GH therapy of short SGA children with 1 mg/m2 per day appears to be effective and safe. Since the future consequences of high serum IGF-I levels during long-term GH therapy with 2 mg/m2 per day are as yet unknown, it seems safer to treat short prepubertal SGA children with a GH dose of 1 mg/m2 per day when children are to be treated continuously for many years.     

Hormonal Regulation of Leptin mRNA Expression and Preadipocyte Recruitment and Differentiation in Porcine Primary Cultures of S-V Cells

The hormonal regulation of leptin mRNA expression and the association between leptin expression and adipocyte differentiation were examined in primary cultures of porcine S-V cells with Northern blot and immunocytochemical analysis. Seeding for 3 days with fetal bovine serum (FBS) with varying levels of dexamethasone (Dex) increased levels of leptin mRNA in a dosedependent manner in parallel with increases in the proportion of preadipocytes (AD-3 positive cells; AD-3, a preadipocyte marker). Six-day treatment with 10 or 850 nM insulin after FBS+Dex treatment resulted in a similar increase in leptin mRNA expression and morphological differentiation. However, significantly lower levels of leptin mRNA and smaller fat cells were observed in cultures treated with 1 nM insulin or 10 nM insulin-like growth factor-I (IGF-I). Dex-induced increases in leptin mRNA levels and AD-3 cell numbers were blocked completely by the addition of transforming growth factor-β (TGF-β) to FBS+Dex-treated cultures. However TGF-β significantly increased fat cell size and leptin mRNA expression when added to ITS (insulin, 850 nM; transferrin, 5 μg/ml; and selenium, 5 ug/mL) treated cultures during the lipid-filling stage. When added with FBS+DEX for the first 3 days, growth hormone (GH) did not influence the Dex-induced increase in AD-3 cells and leptin mRNA expression, but GH reduced leptin mRNA levels when added with insulin for 6 days after FBS+Dex. These results demonstrated that regulation of leptin mRNA expression by Dex, insulin, IGF-I, TGF-β, and GH may be associated with changes in preadipocyte number and fat cell size.     

Inhibition of growth hormone-stimulated lipolysis by somatostatin, insulin, and insulin-like growth factors (somatomedins) in vitro

The effects of somatostatin, insulin, insulin-like growth factor I (IGF-I), and insulin-like growth factor II (IGF-II)/MSA on growth hormone (GH) (1 microgram/ml)-induced lipolysis were examined employing chicken adipose tissue in vitro. Basal and GH-stimulated glycerol release were inhibited by somatostatin (1 ng/ml) and by IGF-II/MSA (10 and 100 ng/ml). Insulin and IGF-I (10 and 100 ng/ml) completely inhibited the lipolytic response to GH without affecting basal glycerol release. Insulin and IGF-I were equipotent in inhibiting GH-induced lipolysis while IGF-II is only 16% as potent as insulin.     

Effects of leptin replacement on hypothalamic-pituitary growth hormone axis function and circulating ghrelin levels in ob/ob mice

Leptin-deficient obese mice (ob/ob) have decreased circulating growth hormone (GH) and pituitary GH and ghrelin receptor (GHS-R) mRNA levels, whereas hypothalamic GH-releasing hormone (GHRH) and somatostatin (SST) expression do not differ from lean controls. Given the fact that GH is suppressed in diet-induced obesity (a state of hyperleptinemia), it remains to be determined whether the absence of leptin contributes to changes in the GH axis of ob/ob mice. Therefore, to study the impact of leptin replacement on the hypothalamic-pituitary GH axis of ob/ob mice, leptin was infused for 7 days (sc), resulting in circulating leptin levels that were similar to wild-type controls (approximately 1 ng/ml). Leptin treatment reduced food intake, body weight, and circulating insulin while elevating circulating n-octanoyl ghrelin concentrations. Leptin treatment did not alter hypothalamic GHRH, SST, or GHS-R mRNA levels compared with vehicle-treated controls. However, leptin significantly increased pituitary GH and GHRH-R expression and tended to enhance circulating GH levels, but this latter effect did not reach statistical significance. In vitro, leptin (1 ng/ml, 24 h) did not affect pituitary GH, GHRH-R, or GHS-R mRNA but did enhance GH release. The in vivo effects of leptin on circulating hormone and pituitary mRNA levels were not replicated by pair feeding ob/ob mice to match the food intake of leptin-treated mice. However, leptin did prevent the fall in hypothalamic GHRH mRNA and circulating IGF-I levels observed in pair-fed mice. These results demonstrate that leptin replacement has positive effects on multiple levels of GH axis function in ob/ob mice.     

Plasma ghrelin levels in patients with end-stage renal disease

Ghrelin is an acylated peptide stimulating secretion of the growth hormone (GH). It was originally isolated from the rat stomach as an endogenous ligand for the growth hormone secretagogue receptor. Although being predominantly produced by endocrine cells of the gastric fundus, its secretion has been found in various tissues including the kidney. To study the influence of renal failure on plasma ghrelin levels we examined 16 patients with end-stage renal disease (ESRD) receiving hemodialysis (8 men and 8 women) and 19 controls (10 men and 9 women). Both groups were comparable in age and BMI. In all subjects we assessed plasma levels of ghrelin, leptin, soluble leptin receptor, insulin, IGF-I, IGFBP-1, IGFBP-3 and IGFBP-6. Ghrelin levels were significantly higher in the group of dialyzed patients (4.49+/-0.74 vs. 1.79+/-0.15 ng/ml; p<0.001). These patients had significantly higher levels of GH, IGFBP-1, IGFBP-6, leptin and percentage of body fat (p<0.05). In the group of patients with ESRD plasma ghrelin levels positively correlated with IGFBP-1 (p<0.01). In the control group, ghrelin positively correlated with GH concentrations (p<0.01) and negatively correlated with the levels of insulin and creatinine (p<0.05). In conclusion, patients with ESRD have higher ghrelin concentrations, which might be caused by a decreased excretion/metabolism of ghrelin in the kidney during renal failure.