Ghrelin生物学功能的研究进展

Ghrelin是一种在大鼠和人胃内新发现的生长激素促分泌素受体(GHS-R)的内源性配基,有28个氨基酸,起促生长激素释放作用。当Ghrelin与位于垂体和下丘脑的GHS-R结合后,产生一系列生物学效应。本文从生长激素释放、机体生长发育、食欲和采食量、能量代谢、胃酸分泌等方面综述了Ghrelin的生物学功能,并初步探讨其在畜牧业上的应用前景。     

ghrelin对生殖系统的调节作用

ghrelin是新近发现的一个含有28个氨基酸残基的多肽,是生长激素促分泌素受体(GHS-R)的天然配体,除具有调节GH分泌和能量平衡的功能之外,尚有其他许多功能。近年来体外或体内试验研究表明,ghrelin对生殖激素如LH、PRL具有一定的调节作用;另外,ghrelin及其受体系统广泛存在于生殖系统中。提示这一新发现的激素可能对生殖系统具有重要的调节作用。文章就ghrelin对生殖系统调节作用的研究进展加以综述。     

Purification and characterization of feline ghrelin and its possible role

Ghrelin, a novel 28-amino acid peptide with an n-octanoyl modification at Ser3, has been isolated from rat and human stomach as an endogenous ligand for the growth hormone secretagogue receptor. Here, we purified feline ghrelin and examined its possible physiological role in cats. The major active form of feline ghrelin is a 28-amino acid peptide octanoylated (C8:0) at Ser3; except for one amino acid residue replacement, this structure is identical to those of rat and human ghrelins. However, much structural divergence in peptide length and fatty acid modification was observed in feline ghrelin: peptides consisting of 27 or 26 amino acids lacking Gln14 and/or Arg28 were found, and the third serine residue was modified by octanoic acid (C8:0), decanoic acid (10:0), or unsaturated fatty acids (C8:1, C10:1 and C10:2). In agreement with the structural divergence, two kinds of cDNA with different lengths were isolated. Administration of synthetic rat ghrelin increased plasma growth hormone levels in cats, with a potency similar to that in rat or human. Plasma levels of ghrelin in cats increased approximately 2.5-fold after fasting. The present study indicates the existence of structural divergence in feline ghrelin and suggests that, as in other animals, ghrelin may play important roles in GH release and feeding in cats.     

Ghrelin stimulates myogenic differentiation in a mouse muscle satellite cell line and in primary cultures of bovine myoblasts

Ghrelin is an acylated hormone that influences food intake, energy metabolism and reproduction, among others. Ghrelin may also stimulate proliferating myoblast cell differentiation and multinucleated myotube fusion. The aim of this work was to assess the effect of human ghrelin (hGHRL) and human ghrelin fragment 1-18 (hGHRL1-18) on myoblast differentiation by means of mRNA expression and protein level. Two types of cells were tested, the cell line i28 obtained from mouse skeletal muscle and primary cultures of bovine myoblasts. Both ghrelin and its N-terminal fragment hGHRL1-18 were used at concentrations of 0, 0.01, 0.1, 1, 10 and 100 nm. Treatments were applied to pre-confluent cultures and were maintained for 4 days. We determined that between 0.1 and 100 nm, hGHRL and hGRHL1-18 had similar effects on myogenic differentiation of i28 cells (p < 0.01). On the other hand, only the higher concentrations (10 and 100 nm) of hGHRL stimulated bovine myoblast differentiation. These results could be attributed to the presence, in both i28 cells and in bovine myoblasts, of the mRNA for GHS-R1a and CD36 receptors. The use of ghrelin in livestock production is still questionable because of the limited effects shown in this study, and additional research is needed in this field.     

A role of ghrelin in cancerogenesis

Ghrelin is a 28 amino-acid multi-functional peptide hormone, which was identified as a natural ligand of the growth hormone secretagogue receptor (GHS-R). Pituitary growth hormone-releasing activity in both animals and humans has been well documented. It has various biological functions, including regulation of appetite and body weight, control of energy homeostasis, modulation of cardiovascular and gastrointestinal system and anti-inflammatory effect. However, both ghrelin and its receptor (GHS-R) are widely distributed in various tumors, which strongly implies their role in neoplastic cell growth trough autocrine/paracrine mechanism. Multiple studies have demonstrated the role of ghrelin in cancer cells proliferation, differentiation, invasiveness and apoptosis inhibition. The ghrelin axis is more complex than it was originally thought and consist of several compounds that might interact with each other and affect ghrelin activities. Here, we provide an overview of the ghrelin and its receptor role in tumor progression.     

Inhibition of growth hormone secretagogue receptor antagonist, [D-Lys-3]-GHRP-6, in adipogenesis of ovine and rat adipocytes

Ghrelin and growth hormone secretagogues receptor (GHS‐R or ghrelin receptor) have been reported as being one of the factors of adipogenesis in adipocytes. To investigate the involvement of ghrelin and GHS‐R in adipocytes, the effect of the GHS‐R antagonist, [D‐Lys‐3]‐GHRP‐6 (His‐D‐Trp‐D‐Lys‐Trp‐D‐Phe‐Lys‐NH₂), on the process of adipogenesis in ovine and rat adipocytes was evaluated. [D‐Lys‐3]‐GHRP‐6 (10^?7 mol/L) significantly inhibited adipogenic differentiation of ovine and rat preadipocytes prepared from adipose tissues. The level of peroxisome proliferator activated receptor (PPAR)‐γ2 mRNA, an adipogenic marker, was decreased during the differentiation of adipocytes treated with [D‐Lys‐3]‐GHRP‐6 for 10 days. Ghrelin stimulated adipogenesis, also causing an increment of glycerol‐3‐phosphate dehydrogenase and upregulation of PPAR‐γ2. Furthermore, the antilipolytic effect of ghrelin was attenuated by treatment with [D‐Lys‐3]‐GHRP‐6 in both types of isolated adipocytes. Overall, the results of the present study highlight that GHS‐R in adipogenesis can be blocked by treatment with [D‐Lys‐3]‐GHRP‐6.     

Circulating ghrelin and leptin concentrations and growth hormone secretagogue receptor abundance in liver, muscle, and adipose tissue of beef cattle exhibiting differences in composition of gain

Data from species other than cattle indicate that ghrelin and GH secretagogue receptor (GHS-R) could play a key role in fat deposition, energy homeostasis, or glucose metabolism by directly affecting liver and adipose tissue metabolism. Beef steers (n = 72) were used to test the hypothesis that plasma ghrelin and leptin concentrations and abundance of the GHS-R in liver, muscle, and adipose tissues differ in steers exhibiting differences in composition of gain. At trial initiation (d 0), 8 steers were slaughtered for initial carcass composition. The remaining 64 steers were stratified by BW, allotted to pen, and treatment was assigned randomly to pen. Steers were not implanted with anabolic steroids. Treatments were 1) a low-energy (LE) diet fed during the growing period (0 to 111 d) followed by a high-energy (HE) diet during the finishing period (112 to 209 d; LE-HE) or 2) the HE diet for the duration of the trial (1 to 209 d; HE-HE). Eight steers per treatment were slaughtered on d 88, 111, 160, and 209. Carcass ninth, tenth, and eleventh rib sections were dissected for chemical composition and regression equations were developed to predict compositional gain. Liver, muscle, and subcutaneous adipose tissues were frozen in liquid nitrogen for subsequent Western blotting for GHS-R. Replicate blood samples collected before each slaughter were assayed for ghrelin and leptin concentrations. When compared at a common compositional fat end-point, the rate of carcass fat accretion (g·kg of shrunk BW(-1)) was greater (P < 0.001) in HE-HE steers whereas the rate of carcass protein accretion (g·kg of shrunk BW(-1)) was less (P < 0.001) compared with LE-HE steers. When compared at a common compositional fat end-point, plasma leptin, ghrelin, and insulin concentrations were greater (P < 0.05) for HE-HE compared with LE-HE steers. Abundance of the GHS-R, to which ghrelin binds, increased over time in liver and adipose tissue but did not differ as a result of treatment. Plasma ghrelin concentrations were increased for cattle continuously fed the HE diet as they became increasingly fatter; however, abundance of the GHS-R in liver, muscle, and subcutaneous adipose tissue was not different between treatment groups. The role of ghrelin in cattle metabolism warrants further investigation as it could have a significant effect on composition of BW gain, feed efficiency, and metabolic disorders such as ketosis and fatty liver.     

Effect of intrahypothalamic injections of norepinephrine and serotonin on plasma arginine vasotocin and mesotocin in the White Leghorn cockerel

1. Saline (10 microliters), norepinephrine (NE) and Serotonin (5-HT), 500 nmol each, were injected into the anterior third ventricle (A3V; n = 7) or the posterior third ventricle (P3V; n = 11) of ananesthetised, unrestrained White Leghorn cockerels. Plasma arginine vasotocin (AVT) and mesotocin (MT) were measured 20, 60 and 120 min after injection. 2. Injection of NE into both the A3V and P3V had no significant effect on either plasma AVT or plasma MT at any of the sampling times. 3. Administration of 5-HT into the A3V significantly increased plasma MT about two-fold 20 min following injection. At 120 min time, plasma MT returned to normal. 4. In P3V birds, 5-HT had no effect on plasma MT in the first 20 min, but a significant increase in plasma MT occurred 60 to 120 min after injection. The magnitude of the response was lower than in the A3V cockerels. 5. Plasma AVT was not affected by 5-HT administration into the A3V at any of the sampling times, but 5-HT administration into the P3V caused significant rises in plasma AVT at 120 min. 6. Serotonergic, but not noradrenergic, induction of neurohypophysial peptide secretion was demonstrated.     

Role for des-acyl ghrelin in the responsiveness of plasma hormones and metabolites to ghrelin in Holstein steers

Gastric-derived peptide hormone ghrelin is known for its potent growth hormone (GH) stimulatory effects. The acyl-modification on N-terminal Ser(3) residue is reported to be important to stimulate the ghrelin receptor, GH secretagogue-receptor type1a (GHS-R1a). However, major portion of circulating ghrelin lacks in acylation, and some biological properties of des-acyl ghrelin have been reported in monogastric animals. In the present study, the responsiveness of plasma hormones and metabolites to ghrelin in steers was characterized, and role for des-acyl ghrelin in these changes was investigated. The repeated intravenous administrations of bovine ghrelin (1.0 microg/kg BW) every 2h for 8h to Holstein steers significantly increased the plasma acylated ghrelin, total ghrelin, GH, insulin and NEFA levels. The GH responses in peak values and area under the curves (AUCs) were attenuated by repeated injections of ghrelin, however, the responses of plasma total ghrelin were similar. Plasma insulin AUC decreased after fourth injection of ghrelin while plasma NEFA AUCs gradually increased by repeated injections of ghrelin. Pretreatment of des-acyl ghrelin (10.0 microg/kg BW) 5 min prior to the single injection of ghrelin (1.0 microg/kg BW) did not affect the ghrelin-induced hormonal changes. Moreover, the responses of plasma GH to bovine and porcine ghrelin, which differ in C-terminal amino acid residues, were similar in calves. These data show that (1) GH release was attenuated by repeated administration of ghrelin, (2) ghrelin regulates glucose and fatty acid metabolism probably via different pathway, and (3) des-acyl ghrelin is unlikely the antagonist for ghrelin to induce endocrine effects in Holstein steers.     

Comparison of effects of leptin and ghrelin on porcine ovarian granulosa cells

The aim of our studies was to compare the roles of leptin and ghrelin in the direct control of proliferation, apoptosis, and secretory activity by porcine ovarian cells. In our in vitro experiments, we analyzed the effects of leptin and ghrelin treatments (at 0, 1, 10, or 100 ng/mL medium) on the accumulation of proliferation-related peptides (PCNA, cyclin B1, MAP kinase [MAPK]) and apoptosis-associated peptides (Bax, caspase 3, p53), and on progesterone secretion by cultured porcine granulosa cells, using immunocytochemistry, SDS PAGE-Western immunoblotting, and radioimmunoassay (RIA). Leptin stimulated proliferation (PCNA, cyclin B1, MAPK), apoptosis (Bax, p53), and progesterone secretion. Ghrelin promoted proliferation (PCNA, cyclin B1, MAPK) and progesterone secretion but suppressed apoptosis (Bax, caspase 3, p53). These observations suggest that both leptin and ghrelin directly control proliferation, apoptosis, and secretory activity by porcine ovarian cells. At the level of the ovary, in contrast to the hypothalamo-hypophysial system, leptin and ghrelin may have similar action in promoting granulosa cell proliferation and progesterone secretion, but they may be antagonistic to one another (leptin, stimulator; ghrelin, inhibitor) in controlling apoptosis.     

Oxyntomodulin increases the concentrations of insulin and glucose in plasma but does not affect ghrelin secretion in Holstein cattle under normal physiological conditions

Ghrelin, the natural ligand of the growth hormone secretagogue receptor (GHS-R1a), has been shown to stimulate growth hormone (GH) secretion. Regulation of ghrelin secretion in ruminants is not well studied. We investigated the effects of oxyntomodulin (OXM) and secretin on the secretions of ghrelin, insulin, glucagon, glucose, and nonesterified fatty acids (NEFA) in pre-ruminants (5 wk old) and ruminants (10 wk old) under normal physiological (feeding) conditions. Eight male Holstein calves (pre-ruminants: 52 ± 1 kg body weight [BW]; and ruminants: 85 ± 1 kg BW) were injected intravenously with 30 μg of OXM/kg BW, 50 μg of secretin/kg BW, and vehicle (0.1% bovine serum albumin [BSA] in saline as a control) in random order. Blood samples were collected, and plasma hormones and metabolites were analyzed using a double-antibody radioimmunoassay system and commercially available kits, respectively. We found that OXM increased the concentrations of insulin and glucose but did not affect the concentrations of ghrelin in both pre-ruminants and ruminants and that there was no effect of secretin on the concentrations of ghrelin, insulin, and glucose in these calves. We also investigated the dose-response effects of OXM on the secretion of insulin and glucose in 8 Holstein steers (401 ± 1 d old, 398 ± 10 kg BW). We found that OXM increased the concentrations of insulin and glucose even at physiological plasma concentrations, with a minimum effective dose of 0.4 μg/kg for the promotion of glucose secretion and 2 μg/kg for the stimulation of insulin secretion. These findings suggest that OXM takes part in glucose metabolism in ruminants.     

Performance and serum parameters of calves (Bos taurus) subject to milk restriction associated with supplementation with 2-hydroxy-4-(methylthio)butanoic acid

Our aim with this study was to evaluate the consumption, performance, quantitative characteristics of carcasses, biochemical profile, plasma levels of ghrelin and leptin, expression of the receptor for ghrelin (GHS-R1a) in the hypothalamus and duodenum, and the number of goblet cells in the duodenum of calves subjected to milk volume restriction and supplemented with 2-hydroxy-4-(methylthio)butanoic acid (HMTBa). We used 21 Holstein mixed-breed calves, aged between 3 and 15 d with an average weight of 36.8 kg, and housed in pens with troughs for hay, concentrate, and water. The study included two consecutive experimental periods (first period [P1] and second period [P2]) of 21 d each, with 7 d of adaptation to the diet and facilities. The calves were distributed in a completely randomized design in three treatments with seven repetitions. 1) Control: 6 liters of milk/d during P1 and 6 liters of milk/day during P2; 2) RES (milk restriction): 3 liters of milk/day during P1 and 6 liters of milk/day during P2; and 3) RES + HMTBa: 3 liters of milk/day during P1 and 6 liters of milk/day during P2 + 3.3 g of HMTBa/day in both periods. HMTBa was supplied in milk, and the amount of concentrated ration and hay provided and leftovers were recorded daily to estimate dry matter (DM) and crude protein consumption. Mean daily weight gain (DWG), final weight (FW), and feed conversion (FC) were obtained at the beginning and at the end of each 21-d period. Plasma concentrations of ghrelin and leptin, triglycerides, total protein, urea, lactate, creatinine, alkaline phosphatase, and cholesterol were measured for P1 and P2 at the end of each 21-d period. At the end of P2, animals were slaughtered; sections of the duodenum were collected to evaluate the expression of GHS-R1a and quantity of goblet cells; hypothalamus was used to evaluate the expression of GHS-R1a; rumen was used to evaluate the thickness of epithelium and keratin and the density, height, and width of ruminal papillae. In P1, total DM consumption, FW, DWG, glucose, and triglycerides were lower in the RES and RES + HMTBa groups (P < 0.001). In P2, there was an improvement in the FC of the RES + HMTBa group (compared with Control and RES groups) and a lower urea concentration in the RES group (compared with Control and RES + HMTBa groups) (P < 0.001). No differences were observed among groups regarding hormonal concentrations, histological parameters, and GHS-R1a expression in the duodenum and hypothalamus. Therefore, milk restriction combined with HMTBa supplementation promoted greater compensatory gain by a mechanism independent of changes in GHS-R1a expression and hormone levels of ghrelin and leptin.     

Combined administration of ghrelin and GHRH synergistically stimulates GH release in Holstein preweaning calves

Ghrelin is a gut peptide which participates in growth regulation through its somatotropic, lipogenic and orexigenic effects. Synergism of ghrelin and growth hormone-releasing hormone (GHRH) on growth hormone (GH) secretion has been reported in humans and rats, but not in domestic animals in vivo. In this study, effects of a combination of ghrelin and GHRH on plasma GH and other metabolic parameters, and changes in plasma active and total ghrelin levels were studied in Holstein bull calves before and after weaning. Six calves were intravenously injected with vehicle (0.1% BSA-saline), ghrelin (1 microg/kg BW), GHRH (0.25 microg/kg BW) or a combination of ghrelin plus GHRH at the age of 5 weeks and 10 weeks (weaning at 6 weeks of age). Ghrelin stimulated GH release with similar potency as GHRH and their combined administration synergistically stimulated GH release in preweaning calves. After weaning, GH responses to ghrelin and GHRH became greater compared with the values of preweaning calves, but a synergistic effect of ghrelin and GHRH was not observed. The GH areas under the concentration curves for 2h post-injection were greater in weaned than in preweaning calves (P<0.05) if ghrelin or GHRH were injected alone, but were similar if ghrelin and GHRH were injected together. Basal plasma active and total ghrelin levels did not change around weaning, but transiently increased after ghrelin injection. Basal plasma insulin, glucose and non-esterified fatty acid levels were reduced after weaning, but no changes by treatments were observed. In conclusion, ghrelin and GHRH synergistically stimulated GH release in preweaning calves, but this effect was lost after weaning.     

禽类Ghrelin研究进展

Ghrelin是从小鼠的胃中提纯并鉴定了的一种促生长激素分泌受体的特定内源性配体,是一种新的脑肠肽,由胃黏膜分泌,能影响哺乳动物的摄食、饮水和消化道的活动以及动物体内其他激素的分泌、从而影响其生长发育,关于Ghrelin在禽类的报道较少.为了更全面的了解禽类Ghrelin的研究作进展,为开展禽类Ghrelin的研究提供...     

Tryptophan enhances ghrelin expression and secretion associated with increased food intake and weight gain in weanling pigs

These studies were conducted to determine whether ghrelin, a 28-amino acid peptide produced mainly by the stomach, was involved in tryptophan-mediated appetite stimulation in swine. In experiment 1, 36 crossbred (Long WhitexLarge White) barrows were used in a 2x3 factorial design to determine the effects of food intake (ad libitum versus limit fed) and tryptophan level (0.12%, 0.19% and 0.26%) on growth performance as well as ghrelin expression, plasma insulin, ghrelin and leptin levels. Ad libitum fed pigs gained more weight, but had poorer feed conversion than limit fed pigs. Weight gain, food intake and feed conversion all improved with increased ingestion of dietary tryptophan. Ad libitum feeding increased plasma insulin. Plasma insulin was unaffected by the level of dietary tryptophan. However, plasma leptin was significantly lower in pigs fed 0.19% tryptophan compared to those fed 0.12% tryptophan. Plasma ghrelin levels and ghrelin mRNA level in gastric fundus and duodenun was significantly higher in pigs fed 0.19% and 0.26% tryptophan diet compared with pigs fed 0.12%. In the second experiment, 18 crossbred barrows were divided into three treatments involving oral infusion of saline, tryptophan (40mg/kg BW) or 5-hydroxytryptophan (40mg/kg BW). Plasma ghrelin levels at 20, 40 and 60min after infusion of tryptophan were higher than after saline and 5-hydroxytryptophan infusion, 5-hydroxytryptophan infusion induced lower food intake than saline infusion, and tryptophan infusion increased food intake 2, 8 and 24h after infusion. In conclusion, oral tryptophan ingestion increased ghrelin expression in gastric fundus and plasma ghrelin level.     

大鼠GTH细胞PKC对FSH-β mRNA表达的影响

为研究大鼠促卵泡素(FSH)分泌的受体后信号转导机制,将GTH细胞用PMA或H7处理后,用GnRH脉冲刺激,再用实时荧光定量PCR方法测定细胞FSH-βmRNA表达的Ct值,并与空白对照组比较。结果表明,PMA能显著降低GTH细胞FSH-βmRNA表达的Ct值,H7则曾能显著升高,说明GTH细胞PKC活性显著影响FSH-βmRNA的表达,FSH-βmRNA的表达随着PKC活性的升高而显著升高,随着PKC活性的降低而显著降低。因此,PKC-Ca2+是GnRH脉冲刺激所引起的FSH-βmRNA表达的受体后的信号转导途径。     

Plasma concentrations of arginine vasotocin and mesotocin following pentobarbital anaesthesia and carotid cannulation in domestic fowl

1. Plasma concentrations of arginine vasotocin (AVT) and mesotocin (MT), arterial pH and pCO2, body temperature, respiratory rate, heart rate, mean arterial blood pressure, packed cell volume and plasma osmolality were monitored in birds for 28 h following anaesthesia and carotid cannulation. 2. Plasma MT concentration decreased after 6 h of surgical recovery but there were no differences between 6 and 28 h of recovery. 3. In contrast, plasma AVT concentration was raised only after 27 h, but no differences were observed between 3 and 28 h of recovery. 4. Anaesthesia and/or carotid cannulation apparently suppressed AVT and enhanced MT release, but no significant change in either hormone was observed after 6 h of surgical recovery under the conditions specified in this study.