Manuel Pombo

Influence of sex, age and adrenergic pathways on the growth hormone response to GHRP‐6

OBJECTIVE His‐dTrp‐Ala‐Trp‐dPhe‐Lys‐NH2 (GHRP‐6) is a synthetic compound that releases GH in a dose‐related and specific manner in several species including man. To further characterize the effects of GHRP‐6 on GH secretion in normal human subjects, we assessed plasma GH levels following GHRP‐6 administration in normal male adult subjects, normal female adult subjects at different stages of their menstrual cycle and in normal prepubertal male and female children. We also studied the influence of adrenergic pathways on GHRP‐6 induced GH secretion in normal adult male subjects.

Hormonal Control of Growth Hormone Secretion

Growth hormone secretion by the somatotroph cells depends upon the interaction between hypothalamic regulatory peptides, target gland hormones and a variety of growth factors acting in a paracrine or autocrine fashion. This review will be focused on recent data regarding the mechanism by which growth hormone-releasing hormone (GHRH) influences somatotroph cell function and the physiological role played by Ghrelin and leptin in the regulation of growth hormone (GH) secretion. It is well established that binding of GHRH to its receptor leads to activation of protein kinase A (PKA). More recently, it was found that GHRH can also activate mitogen-activated protein (MAP) kinase both in pituitary cells and in a cell line overexpressing the GHRH receptor. Whether somatotroph adenomas, either with or without a GS-alpha mutation, have alterations in some of the components of the activation of the MAP kinase pathway remains to be known. The recent isolation of Ghrelin, the endogenous ligand of the growth hormone secretagogue receptor, can be considered a landmark in the GH field, which opens up the possibility of gaining greater insight into our understanding of the mechanisms involved in the regulation of GH secretion and somatic growth. Indeed, preliminary evidences indicate that this peptide exerts a marked stimulatory effect on plasma GH levels in both rats and humans. Finally, it is well known that GH secretion is markedly influenced by nutritional status. Leptin has emerged as an important adipose tissue-generated signal that is involved in the regulation of GH secretion, thus providing an integrated regulatory system of growth and metabolism. Although the effects of leptin on GH secretion in humans remain to be clarified, indirect evidences indicate that it may play an inhibitory role.

Functional Consequences of Seven Novel Mutations in the CYP11B1 Gene: Four Mutations Associated with Nonclassic and Three Mutations Causing Classic 11β-Hydroxylase Deficiency

CONTEXT Steroid 11beta-hydroxylase (CYP11B1) deficiency (11OHD) is the second most common form of congenital adrenal hyperplasia (CAH). Cases of nonclassic 11OHD are rare compared with the incidence of nonclassic 21-hydroxylase deficiency. OBJECTIVE The aim of the study was to analyze the functional consequences of seven novel CYP11B1 mutations (p.M88I, p.W116G, p.P159L, p.A165D, p.K254_A259del, p.R366C, p.T401A) found in three patients with classic 11OHD, two patients with nonclassic 11OHD, and three heterozygous carriers for CYP11B1 mutations. METHODS We conducted functional studies employing a COS7 cell in vitro expression system comparing wild-type (WT) and mutant CYP11B1 activity. Mutants were examined in a computational three-dimensional model of the CYP11B1 protein. RESULTS All mutations (p.W116G, p.A165D, p.K254_A259del) found in patients with classic 11OHD have absent or very little 11beta-hydroxylase activity relative to WT. The mutations detected in patients with nonclassic 11OHD showed partial functional impairment, with one patient being homozygous (p.P159L; 25% of WT) and the other patient compound heterozygous for a novel mild p.M88I (40% of WT) and the known severe p.R383Q mutation. The two mutations detected in heterozygous carriers (p.R366C, p.T401A) also reduced CYP11B1 activity by 23 to 37%, respectively. CONCLUSION Functional analysis results allow for the classification of novel CYP11B1 mutations as causative for classic and nonclassic 11OHD, respectively. Four partially inactivating mutations are predicted to result in nonclassic 11OHD. These findings double the number of mild CYP11B1 mutations previously described as associated with mild 11OHD. Our data are important to predict phenotypic expression and provide important information for clinical and genetic counseling in 11OHD.

Regulation of ghrelin secretion and action

The pulsatile release of growth hormone (GH) from anterior pituitary gland is regulated by the interplay of at least two hypothalamic hormones, GH-releasing hormone (GHRH) and somatostatin, via their engagement with specific cell surface receptors on the anterior pituitary somatotroph. Furthermore, release of GH in vivo may also be controlled by a third type of receptor, the growth hormone secretagogue receptor, a G-protein-coupled receptor, called GHS receptor type 1a (GHS-R1a), which was identified in the pituitary and the hypothalamus in humans using a nonpeptidyl growth hormone secretagogue (MK-0677). Ghrelin, the endogenous ligand for the GHS-R1a, is a 28-amino-acid peptide isolated from human stomach that is modified by a straight chain octanoyl group covalently linked to Ser3, which is essential for its endocrine activity. This hormone, predominantly expressed and secreted by the stomach, has a dual action on GH secretion and food intake, showing interdependency between these actions. The finding that fasting and food intake, respectively, increase and decrease the secretion of ghrelin suggests that this hormone may be the bridge connecting somatic growth and body composition with energy metabolism, and appears to play a role in the alteration of energy homeostasis and body weight in pathophysiological states such as hypothyroidism and hyperthyroidism. Despite this, little is known about the intracellular signaling through which ghrelin exerts its regulatory actions. Activation of intracellular calcium mobilization is one of the earliest known cellular signals elicited by ghrelin. In HEK-293 cells expressing the GHS-R1a, ghrelin induces a biphasic cytosolic calcium elevation characterized by a spike phase of the response, which reflects Ins(1,4,5)P3-dependent calcium mobilization of intracellular stores, and a sustained phase of the response, which is due to calcium influx across the plasma membrane triggered by aperture of capacitative calcium channels (store-operated calcium channels). Upon repeated administration, ghrelin showed a marked suppression of ghrelin-mediated elevations of intracellular calcium. This homologous desensitization represents an important physiological mechanism that modulates receptor responsiveness and acts as an information filter for intracellular signaling system. The discovery of ghrelin adds a new component to the complex machinery responsible for regulation of GH secretion in connection with the regulation of appetite and energy homeostasis.

High variability in CYP21A2 mutated alleles in Spanish 21‐hydroxylase deficiency patients, six novel mutations and a founder effect

Objective To detect common as well as rare and novel CYP21A mutations in 21‐hydroxylase deficiency patients. To estimate the distribution of mutations and compare them with other European studies. To construct haplotypes linked to a recurrent novel mutation.

Growth Hormone Secretagogues: The Clinical Future

Growth hormone (GH) releasing hexapeptide (GHRP)-6 and other peptidergic and non-peptidergic compounds collectively designated GH secretagogues (GHS) are potent releasers of GH in man. Their clinical future may be envisioned in three areas: therapy of GH-deficient (GHD) states, diagnosis of GHD, and non-endocrinological actions. As therapeutic agents and compared with GH itself, GHS have the disadvantage of lower potency but have a more physiological and safer profile of GH secretion. GHS administration could be indicated for states in which medium GH doses have been shown to be effective. As a diagnostic tool, the combined administration of GH releasing hormone plus GHRP-6, both at saturating doses, is currently the most powerful releaser of GH, devoid of side effects and convenient for the patient; it may also be an alternative to the insulin tolerance test for the diagnosis of GHD in adult patients. Their potential action at cardiovascular level is highly promising. Although the clinical future of GH releasing substances is appealing, probably the most relevant contribution has yet to be discovered. Once the endogenous ligand of the GHS receptor is identified, we will have an insight into the real hypothalamic control of GH secretion in man. With this knowledge it is likely that some diagnostic and therapeutic actions that are commonly undertaken will significantly change.

High serum leptin levels in children with type 1 diabetes mellitus : Contribution of age, BMI, pubertal development and metabolic status

Children with diabetes mellitus are prone to develop obesity and to experience a delay in onset of the pubertal process. In order to understand the role of leptin in these abnormalities, serum leptin levels were analysed in children with type 1 diabetes mellitus.

GLUCOCORTICOID DEFICIENCY WITH ACHALASIA OF THE CARDIA AND LACK OF LACRIMATION

Four recent reports describe a multisystem disorder in which ACTH insensitivity is associated with achalasia and alacrima. We report studies on a male patient with this rare triad. The patient had alacrima from birth; isolated glucocorticoid deficiency had been diagnosed at 3·5 years of age and achalasia at age 6. The possibility that this syndrome could be due to a parasympathetic degeneration has already been proposed; the cause of the glucocorticoid deficiency, however, remains unclear. Parasympathetic function in other areas was investigated to determine whether there might be a more generalized abnormality. Specific cardiac tests of parasympathetic function showed that parasympathetic input to the heart was affected in the patient, while the same tests in an Addisonian child were normal. We show, then, a hitherto undetected parasympathetic abnormality in a patient with this syndrome, suggesting a generalized disturbance of this system. On this basis we may hypothesize that the glucocorticoid failure may be a consequence of the loss of parasympathetic input to the adrenal gland, although this remains to be demonstrated experimentally.

Plasma growth hormone response to growth hormone‐releasing hexapeptide (GH‐RP‐6) in children with short stature

Eighteen children with short stature were evaluated for growth hormone (GH) reserve after pharmacological tests and a single iv injection of GH‐RP‐6. These children were divided into two groups: 10 were diagnosed as having idiopathic GH deficiency by classical stimulation tests (group A) and the remaining 8 (group B) were considered growth‐retarded children with normal GH secretion, following conventional stimulation, but reduced endogenous GH secretion. The results were compared with a group of 12 normal children. As a group, patients in group A showed a lower GH response to GH‐RP‐6, while patients in group B had a similar response as normal controls. However, on an individual basis, a considerable degree of overlapping in responses among the three groups was evident. These data indicate that, on an individual basis, GH‐RP‐6 testing is not of diagnostic value in children suspected of having idiopathic GH deficiency.GH deficiency, GH‐RP‐6, short stature

Hypochondroplasia and acanthosis nigricans: a new syndrome due to the p.Lys650Thr mutation in the fibroblast growth factor receptor 3 gene?

BACKGROUND Hypochondroplasia (HCH) is a skeletal dysplasia inherited in an autosomal dominant manner due, in most cases, to mutations in the fibroblast growth factor receptor 3 (FGFR3). Acanthosis nigricans (AN) is a velvety and papillomatous pigmented hyperkeratosis of the skin, which has been recognized in some genetic disorders more severe than HCH involving the FGFR3 gene. OBJECTIVE AND DESIGN After initial study of the proband, who had been consulted for short stature and who also presented AN, the study was extended to the patients mother and to 12 additional family members. METHODS Clinical, biochemical and radiological studies were performed on the family. In addition, exons 11 and 13 of FGFR3 were analyzed. RESULTS The proband and ten relatives presented HCH plus AN and the analysis of FGFR3 showed the p.Lys650Thr mutation. The members with normal phenotypes were non-carriers of the mutation. CONCLUSION This is the first report of a large pedigree with the clinical phenotype of HCH plus AN due to a FGFR3 mutation, p.Lys650Thr. This finding demonstrates the coexistence of both conditions due to the same mutation and it might represent a true complex, which should be further established by searching for AN in mild HCH patients or for HCH in patients with AN.