Growth Hormone Receptor Exon 3 Isoforms and Their Implication in Growth Disorders and Treatment

Human recombinant growth hormone (hGH) has been used to treat short stature in several different conditions, but considerable inter-individual variation in short- and long-term growth response exists. Pharmacogenomics can provide important insights into hGH therapy. The GH receptor (GHR) is the first key molecule mediating GH action. In the past 3 years, a common GHR polymorphism reflecting the presence (GHRf1) or absence (GHRd3) of exon 3 has been under intensive investigation regarding its influence on the response to hGH therapy. Studies that evaluated response to GH treatment determined by these two GHR isoforms in children with GH deficiency, girls with Turner syndrome, children born small for gestational age and patients with acromegaly showed that patients carrying the GHRd3 allele demonstrated a greater GH sensitivity than patients homozygous for the GHRf1 allele. Other studies presented contradictory data, however, which may be caused by confounding factors such as small sample sizes and differences in experimental design. This GHR exon 3 genotype is the first identified genetic factor found to modulate the individual response to GH therapy. This article reviews the historical aspects and pharmacogenetic studies published to date in relation to this GHR polymorphism. The analyses of present and future validation studies may define the use of this and other polymorphisms in clinical practice, moving from pharmacogenetics to routine application and allowing individualization of hGH doses to optimize final outcome. Copyright (C) 2009 S. Karger AG, Basel

Increased Transactivation Associated with SOX3 Polyalanine Tract Deletion in a Patient with Hypopituitarism

Backgound and Aims: Correct gene dosage of SOX3 is critical for the development of the hypothalamo-pituitary axis. Both overdosage of SOX3, as a result of gene duplication, and loss of function resulting from expansion of the first polyalanine (PA) tract are associated with variable degrees of hypopituitarism, with or without mental retardation. The aim of this study was to further investigate the contribution of SOX3 in the etiology of hypopituitarism and the mechanisms involved in the phenotypic variability. Methods: We screened 154 patients with congenital hypopituitarism and an undescended posterior pituitary for mutations in SOX3 and variability in the length of the first PA tract. In addition, 300 patients with variable septooptic dysplasia were screened for variability of the PA tract. Results: We report a novel 18-base pair deletion (p.A243_A248del6, del6PA) in a female patient with hypopituitarism resulting in a 2-fold increase in transcriptional activation in vitro, compared with wild-type SOX3. We also identified a previously reported seven-alanine expansion (p.A240_A241ins7, +7PA) in two male siblings with isolated GH deficiency and a distinct phenotype, in addition to the nonsynonymous variant p.R5Q in an unrelated individual; this appears to have no functional effect on the protein. In contrast to +7PA, del6PA maintained its ability to repress beta-catenin mediated transcription in vitro. Conclusion: This is the first study to report that PA tract deletions associated with hypopituitarism have functional consequences in vitro, possibly due to increased activation of SOX3 target genes. In addition, we have expanded the phenotypic spectrum associated with PA tract expansion (+7PA) mutations to include panhypopituitarism or isolated GH deficiency, with or without mental retardation. (J Clin Endocrinol Metab 96: E685-E690, 2011)

Novel Heterozygous Nonsense GLI2 Mutations in Patients with Hypopituitarism and Ectopic Posterior Pituitary Lobe without Holoprosencephaly

Context: GLI2 is a transcription factor downstream in Sonic Hedgehog signaling, acting early in ventral forebrain and pituitary development. GLI2 mutations were reported in patients with holoprosencephaly (HPE) and pituitary abnormalities. Objective: The aim was to report three novel frameshift/nonsense GLI2 mutations and the phenotypic variability in the three families. Setting: The study was conducted at a university hospital. Patients and Methods: The GLI2 coding region of patients with isolated GH deficiency (IGHD) or combined pituitary hormone deficiency was amplified by PCR using intronic primers and sequenced. Results: Three novel heterozygous GLI2 mutations were identified: c. 2362_2368del p. L788fsX794 (family 1), c. 2081_2084del p. L694fsX722 (family 2), and c. 1138 G > T p. E380X (family 3). All predict a truncated protein with loss of the C-terminal activator domain. The index case of family 1 had polydactyly, hypoglycemia, and seizures, and GH, TSH, prolactin, ACTH, LH, and FSH deficiencies. Her mother and seven relatives harboring the same mutation had polydactyly, including two uncles with IGHD and one cousin with GH, TSH, LH, and FSH deficiencies. In family 2, a boy had cryptorchidism, cleft lip and palate, and GH deficiency. In family 3, a girl had hypoglycemia, seizures, excessive thirst and polyuria, and GH, ACTH, TSH, and antidiuretic hormone deficiencies. Magnetic resonance imaging of four patients with GLI2 mutations and hypopituitarism showed a hypoplastic anterior pituitary and an ectopic posterior pituitary lobe without HPE. Conclusion: We describe three novel heterozygous frameshift or nonsense GLI2 mutations, predicting truncated proteins lacking the activator domain, associated with IGHD or combined pituitary hormone deficiency and ectopic posterior pituitary lobe without HPE. These phenotypes support partial penetrance, variable polydactyly, midline facial defects, and pituitary hormone deficiencies, including diabetes insipidus, conferred by heterozygous frameshift or nonsense GLI2 mutations. (J Clin Endocrinol Metab 95: E384-E391, 2010)

Controversies regarding the effects of growth hormone on the heart

Growth hormone (GH) profoundly affects the developing and adult myocardium. Adult patients with GH deficiency (GHD) and GH excess (acromegaly) provide important models in which to understand the effects of GH in adult cardiac physiology. An increasing body of clinical and experimental evidence illustrates the specific physiological abnormalities that are likely associated with the excess cardiovascular mortality observed in both acromegaly and GHD. Because human GH replacement is now available to treat adults with GHD, new questions emerge about the long-term cardiovascular effects of replacement therapy. In multiple trials, GH therapy for congestive heart failure has been proved ineffective in the absence of preexisting GHD. Case reports suggest that, in the setting of GHD, GH therapy can exert a potent beneficial effect on congestive heart failure. Long-term studies addressing cardiovascular morbidity and mortality are needed to assess the role of GH therapy for GHD.

Clinical and molecuar characterization of Brazilian patients with growth hormone gene deletions

Genomic DNA from 23 patients with isolated growth hormone (GH) deficiency (12 males and 11 females: heights -4.9 ± 1.4 SDS) was screened for GH gene deletions by restriction endonuclease analysis of polymerase chain reaction amplification products. Three unrelated patients had typical features of severe GH deficiency and deletions (6.7 kb in two and 7.6 kb in one) of the GH gene. The two patients with 6.7-kb deletions developed growth-attenuating anti-GH antibodies whereas the patient with the 7.6-kb deletion continued to grow with GH replacement therapy. Our finding that 3/23 (~13%) Brazilian subjects had GH gene deletions agrees with previous studies of severe isolated GH deficiency subjects in other populations. Two of three subjects (67%) with deletions developed blocking antibodies despite administration of exogenous GH at low doses. Interestingly, only 1/10 of cases with affected relatives or parental consanguinity had GH-1 gene deletions

Deficiência de hormônio do crescimento após radioterapia por meduloblastoma na infância: relato de caso

A radiação do crânio para tratamento das neoplasias do sistema nervoso central na infância pode evoluir com sequelas neuroendócrinas, sendo a deficiência de hormônio do crescimento (GH) com retardo do crescimento linear, uma das mais frequentes. Relatamos o caso de menino de 10 anos com cefaléia occipital associada a vertigem, náuseas e vômitos. A tomografia do crânio demonstrou processo expansivo no hemisfério cerebelar esquerdo, que foi retirado cirurgicamente. O exame histopatológico revelou meduloblastoma e o paciente foi submetido a radioterapia crânio-espinhal. Evoluiu sem recidiva da neoplasia e sem déficits neurológicos durante 4 anos. Apresentou retardo do crescimento estatural, sendo confirmada a hipótese de deficiência de GH. Atualmente, encontra-se em uso de GH 0,1 U/kg/dia, tendo apresentado incremento de 4cm na estatura em 6 meses. O presente caso destaca a importância do acompanhamento criterioso de pacientes submetidos à radiação do crânio para tratamento oncológico na infância, visto que podem evoluir com deficiências neuroendócrinas e serem beneficiados com reposição hormonal.

Growth hormone response to growth hormone-releasing peptide-2 in growth hormone-deficient Little mice

OBJECTIVE: To investigate a possible direct, growth hormone-releasing, hormone-independent action of a growth hormone secretagogue, GHRP-2, in pituitary somatotroph cells in the presence of inactive growth hormone-releasing hormone receptors. MATERIALS AND METHODS: The responses of serum growth hormone to acutely injected growth hormone-releasing P-2 in lit/litmice, which represent a model of GH deficiency arising frommutated growth hormone-releasing hormone-receptors, were compared to those observed in the heterozygous (lit/+) littermates and wild-type (+/+) C57BL/6J mice. RESULTS: After the administration of 10 mcg of growth hormone-releasing P-2 to lit/lit mice, a growth hormone release of 9.3 +/- 1.5 ng/ml was observed compared with 1.04 +/- 1.15 ng/ml in controls (p<0.001). In comparison, an intermediate growth hormone release of 34.5 +/- 9.7 ng/ml and a higher growth hormone release of 163 +/- 46 ng/ml were induced in the lit/+ mice and wild-type mice, respectively. Thus, GHRP-2 stimulated growth hormone in the lit/lit mice, and the release of growth hormone in vivo may be only partially dependent on growth hormone-releasing hormone. Additionally, the plasma leptin and ghrelin levels were evaluated in the lit/lit mice under basal and stimulated conditions. CONCLUSIONS: Here, we have demonstrated that lit/lit mice, which harbor a germline mutation in the Growth hormone-releasing hormone gene, maintain a limited but statistically significant growth hormone elevation after exogenous stimulation with GHRP-2. The present data probably reflect a direct, growth hormone-independent effect on Growth hormone S (ghrelin) stimulation in the remaining pituitary somatotrophs of little mice that is mediated by growth hormone S-R 1a.

The consequences of growth hormone-releasing hormone receptor haploinsufficiency for bone quality and insulin resistance

Objective Growth hormone (GH)/insulin-like growth factor (IGF) axis and insulin are key determinants of bone remodelling. Homozygous mutations in the GH-releasing hormone receptor (GHRHR) gene (GHRHR) are a frequent cause of genetic isolated GH deficiency (IGHD). Heterozygosity for GHRHR mutation causes changes in body composition and possibly an increase in insulin sensitivity, but its effects on bone quality are still unknown. The objective of this study was to assess the bone quality and metabolism and its correlation with insulin sensitivity in subjects heterozygous for a null mutation in the GHRHR. Patients and methods A cross-sectional study was performed on 76 normal subjects (68.4% females) (N/N) and 64 individuals (64.1% females) heterozygous for a mutation in the GHRHR (MUT/N). Anthropometric features, quantitative ultrasound (QUS) of the heel, bone markers [osteocalcin (OC) and CrossLaps], IGF-I, glucose and insulin were measured, and homeostasis model assessment of insulin resistance (HOMAIR) was calculated. Results There were no differences in age or height between the two groups, but weight (P = 0.007) and BMI (P = 0.001) were lower in MUT/N. There were no differences in serum levels of IGF-I, glucose, T-score or absolute values of stiffness and OC, but insulin (P = 0.01), HOMAIR (P = 0.01) and CrossLaps (P = 0.01) were lower in MUT/N. There was no correlation between OC and glucose, OC and HOMAIR in the 140 individuals as a whole or in the separate MUT/N or N/N groups. Conclusions This study suggests that one allele mutation in the GHRHR gene has a greater impact on energy metabolism than on bone quality.

Growth hormone response to growth hormone-releasing peptide-2 in growth hormone-deficient Little mice

OBJECTIVE: To investigate a possible direct, growth hormone-releasing, hormone-independent action of a growth hormone secretagogue, GHRP-2, in pituitary somatotroph cells in the presence of inactive growth hormonereleasing hormone receptors. MATERIALS AND METHODS: The responses of serum growth hormone to acutely injected growth hormone-releasing P-2 in lit/litmice, which represent a model of GH deficiency arising frommutated growth hormone-releasing hormonereceptors, were compared to those observed in the heterozygous (lit/+) littermates and wild-type (+/+) C57BL/6J mice. RESULTS: After the administration of 10 mcg of growth hormone-releasing P-2 to lit/lit mice, a growth hormone release of 9.3±1.5 ng/ml was observed compared with 1.04±1.15 ng/ml in controls (p<0.001). In comparison, an intermediate growth hormone release of 34.5±9.7 ng/ml and a higher growth hormone release of 163±46 ng/ml were induced in the lit/+ mice and wild-type mice, respectively. Thus, GHRP-2 stimulated growth hormone in the lit/lit mice, and the release of growth hormone in vivo may be only partially dependent on growth hormone-releasing hormone. Additionally, the plasma leptin and ghrelin levels were evaluated in the lit/lit mice under basal and stimulated conditions. CONCLUSIONS: Here, we have demonstrated that lit/lit mice, which harbor a germline mutation in the Growth hormone-releasing hormone gene, maintain a limited but statistically significant growth hormone elevation after exogenous stimulation with GHRP-2. The present data probably reflect a direct, growth hormone-independent effect on Growth hormone S (ghrelin) stimulation in the remaining pituitary somatotrophs of little mice that is mediated by growth hormone S-R 1a.

Il craniofaringioma nel bambino e nell'adolescente. Luci ed ombre durante il follow-up

Introduction. Craniopharyngioma (CF) is a malformation of the hypothalamicpituitary region and it is the most common nonglial cerebral tumor in children with an high overall survival rate. In some case severe endocrinologic and metabolic sequelae may occur during follow up. 50% of patients (pts), in particular those with radical removal of suprasellar lesions, develop intractable hyperphagia and morbid obesity, with dyslypidemia and high cardiovascular risk. We studied the auxological and metabolic features of a series of 29 patients (18 males) treated at a mean age of 7,6 years, followed up in our Centre from 1973 to 2008 with a mean follow up of 8,3 years. Patients features at the onset. 62% of pts showed as first symptoms of disease visual impairment and neurological disturbancies (headache); 34% growth arrest; 24% signs of raised intracranial pressure and 7% diabetes insipidus. Diagnosis. Diagnosis of CF was reached finally by TC or MRI scans which showed endo-suprasellar lesion in 23 cases and endosellar tumour in 6 cases. Treatment and outcome. 25/29 pts underwent surgical removal of CF (19 by transcranial approach and 6 by endoscopic surgery); 4 pts underwent stereotactic surgery as first line therapy. 3 pts underwent local irradiation with yttrium-90, 5 pts post surgery radiotherapy. 45% of pts needed more than one treatment procedure. Results. After CF treatment all patients suffered from 3 or more pituitary hormone deficiencies and diabetes insipidus. They underwent promptly substitutive therapy with corticosteroids, l-thyroxine and desmopressin. In 28/29 pts we found growth hormone (GH) deficiency. 20/28 pts started GH substitutive therapy and 15 pts reached final height(FH) near target height(TH). 8 pts were not GH treated for good growth velocity, even without GH, or for tumour residual. They reached in 2 cases FH over TH showing the already known phenomenon of growth without GH. 38% of patients showed BMI SDS >2 SDS at last assessment, in particular pts not GH treated (BMI 2,5 SDS) are more obese than GH treated (BMI 1,2 SDS). Lipid panel of 16 examined pts showed significative differencies among GH treated (9 pts) and not treated (7 pts) with better profile in GH treated ones for Total Cholesterol/C-HDL and C-LDL/C-HDL. We examined intima media thickness of common carotid arteries in 11 pts. 3/4 not GH treated pts showed ultrasonographic abnormalities: calcifications in 2 and plaque in 1 case. Of them 1 pt was only 12,6 years old and already showed hypothalamic obesity with hyperphagia, high HOMA index and dyslipidemia. In the GH treated group (7) we found calcifications in 1 case and a plaque in another one. GH therapy was started in the young pt with carotid calcifications, with good improvement within 6 months of treatment. 5/29 pts showed hypothalamic obesity, related to hypothalamic damage (type of surgical treatment, endo-suprasellar primitive lesion, recurrences). 48% of patients recurred during follow up ( mean time from treatment: 3 years) and underwent, in some cases up to 4 transcranial surgical treatments. GH seems not to increase recurrence rate since 40% of GH treated recurred vs 66,6% of not GH treated pts. Discussion. Our data show the extereme difficulties that occur during follow up of craniopharyngioma treated patients. GH therapy should be offered to all patients even with good growth velocity after CF treatment, to avoid dislypidemia and reduce cardiovascular risk. The optimal therapy is not completely understood and whether gross tumor removal or partial surgery is the best option remains to be decided only on one patient tumour features and hypothalamic involvement. In conclusion the gold standard treatment of CF remains complete tumour removal, when feasible, or partial resection to preserve hypothalamic function in endosuprasellar large neoplasms.

Growth hormone replacement therapy regulates microRNA-29a and targets involved in insulin resistance

Replacement of growth hormone (GH) in patients suffering from GH deficiency (GHD) offers clinical benefits on body composition, exercise capacity, and skeletal integrity. However, GH replacement therapy (GHRT) is also associated with insulin resistance, but the mechanisms are incompletely understood. We demonstrate that in GH-deficient mice (growth hormone-releasing hormone receptor (Ghrhr)(lit/lit)), insulin resistance after GHRT involves the upregulation of the extracellular matrix (ECM) and the downregulation of microRNA miR-29a in skeletal muscle. Based on RNA deep sequencing of skeletal muscle from GH-treated Ghrhr(lit/lit) mice, we identified several upregulated genes as predicted miR-29a targets that are negative regulators of insulin signaling or profibrotic/proinflammatory components of the ECM. Using gain- and loss-of-function studies, five of these genes were confirmed as endogenous targets of miR-29a in human myotubes (PTEN, COL3A1, FSTL1, SERPINH1, SPARC). In addition, in human myotubes, IGF1, but not GH, downregulated miR-29a expression and upregulated COL3A1. These results were confirmed in a group of GH-deficient patients after 4 months of GHRT. Serum IGF1 increased, skeletal muscle miR-29a decreased, and miR-29a targets were upregulated in patients with a reduced insulin response (homeostatic model assessment of insulin resistance (HOMA-IR)) after GHRT. We conclude that miR-29a could contribute to the metabolic response of muscle tissue to GHRT by regulating ECM components and PTEN. miR-29a and its targets might be valuable biomarkers for muscle metabolism following GH replacement. KEY MESSAGES GHRT most significantly affects the ECM cluster in skeletal muscle from mice. GHRT downregulates miR-29a and upregulates miR-29a targets in skeletal muscle from mice. PTEN, COL3A1, FSTL1, SERPINH1, and SPARC are endogenous miR-29a targets in human myotubes. IGF1 decreases miR-29a levels in human myotubes. miR-29a and its targets are regulated during GHRT in skeletal muscle from humans.

The role of growth hormone replacement in a growth hormone deficient patient with underlying cardiomyopathy and severe congestive heart failure

It has been reported that-growth hormone (GH) deficiency induced cardiomyopathy responds to growth hormone replacement therapy. We describe the case of a middle-aged male with cardiomyopathic heart failure and growth hormone deficiency of the adult secondary to surgical panhypopituitarism. We demonstrate clinical and hemodynamic improvement of cardiac function with growth hormone replacement therapy despite underlying structural heart disease. Copyright (C) 2005 by the International Society for Heart and Lung Transplantation.

Effectiveness of the Combined Recombinant Human Growth Hormone and Gonadotropin-Releasing Hormone Analog Therapy in Pubertal Patients with Short Stature due to SHOX Deficiency

Context: Isolated heterozygous SHOX defects are the most frequent monogenic cause of short stature, and combined therapy with recombinant human GH (rhGH) and GnRH analog (GnRHa) in pubertal patients has been suggested, but there are no data on final height. Objective: The aim of the study was to analyze adult height after rhGH and GnRHa therapy in patients with SHOX haploinsufficiency. Patients: Ten peripubertal patients with isolated SHOX defects participated in the study. Intervention: Five patients were followed without treatment, and five were treated with rhGH (50 mu g/kg/d) and depot leuprolide acetate (3.75 mg/month). Main Outcome Measures: Adult height SD score (SDS) was measured. Results: All patients followed without treatment had marked downward growth shift during puberty (height SDS, -1.2 +/- 0.7 at 11.4 +/- 1.4 yr; adult height SDS, -2.5 +/- 0.5). Conversely, four of five patients treated with rhGH for 2 to 4.9 yr associated to GnRHa for 1.4 to 5.8 yr improved their height SDS from -2.3 +/- 1.3 at 11.8 +/- 2.1 yr to a final height SDS of -1.7 +/- 1.6. The difference between the mean height SDS at the first evaluation and final height SDS was statistically significant in nontreated vs. treated patients (mean height SDS change, -1.2 +/- 0.4 vs. 0.6 +/- 0.4, respectively; P < 0.001). Conclusion: A gain in adult height of patients with isolated SHOX defects treated with combined rhGH and GnRHa therapy was demonstrated for the first time, supporting this treatment for children with SHOX defects who have just started puberty to avoid the loss of growth potential observed in these patients during puberty. (J Clin Endocrinol Metab 95: 328-332, 2010)

Obesity due to Melanocortin 4 Receptor (MC4R) Deficiency Is Associated with Increased Linear Growth and Final Height, Fasting Hyperinsulinemia, and Incompletely Suppressed Growth Hormone Secretion

Context: Melanocortin receptor 4 (MC4R) deficiency is characterized by increased linear growth greater than expected for the degree of obesity. Objective: The objective of the investigation was to study the somatotroph axis in obese MC4R-deficient patients and equally obese controls. Patients and Methods: We obtained anthropometric measurements and insulin concentrations in 153 MC4R-deficient subjects and 1392 controls matched for age and severity of obesity. We measured fasting IGF-I, IGF-II, IGF binding protein (IGFBP)-1, IGFBP-3, and acid-labile subunit levels in a subset of 33 MC4R-deficient patients and 36 control subjects. We examined pulsatile GH secretion in six adult MC4R-deficient subjects and six obese controls. Results: Height so score was significantly greater in MC4R-deficient children under 5 yr of age compared with controls (mean +/- SEM: 2.3 +/- 0.06 vs. 1.8 +/- 0.04, P < 0.001), an effect that persisted throughout childhood. Final height (cm) was greater in MC4R-deficient men (mean +/- SEM 173 +/- 2.5 vs. 168 +/- 2.1, P < 0.001) and women (mean 165 +/- 2.1 vs. 158 +/- 1.9, P < 0.001). Fasting IGF-I, IGF-II, acid-labile subunit, and IGFBP-3 concentrations were similar in the two groups. GH levels were markedly suppressed in obese controls, but pulsatile GH secretion was retained in MC4R deficiency. The mean maximal GH secretion rate per burst (P < 0.05) and mass per burst (P < 0.05) were increased in MC4R deficiency, consistent with increased pulsatile and total GH secretion. Fasting insulin levels were markedly elevated in MC4R-deficient children. Conclusions: In MC4R deficiency, increased linear growth in childhood leads to increased adult final height, greater than predicted by obesity alone. GH pulsatility is maintained in MC4R deficiency, a finding consistent with animal studies, suggesting a role for MC4R in controlling hypothalamic somatostatinergic tone. Fasting insulin levels are significantly higher in children carrying MC4R mutations. Both of these factors may contribute to the accelerated growth phenotype characteristic of MC4R deficiency. (J Clin Endocrinol Metab 96: E181-E188, 2011)

3-M syndrome associated with growth hormone deficiency: 18 year follow-up of a patient.

3-M syndrome is a rare autosomal recessive disorder that causes short stature, unusual facial features and skeletal abnormalities. Mutations in the CUL7, OBSL1 and CCDC8 genes could be responsible for 3-M syndrome.Here we describe the growth and evolution of dismorphic features of an Italian boy with 3-M syndrome and growth hormone deficiency (GHD) from birth until adulthood. He was born full term with a very low birth weight (2400 g=-3.36 standard deviation score, SDS) and length (40.0 cm =-6.53 SDS). At birth he presented with a broad, fleshy nose with anteverted nostrils, thick and patulous lips, a square chin, curvilinear shaped eyebrows without synophrys, short thorax and long slender bones. Then, during childhood tall vertebral bodies, hip dislocation, transverse chest groove, winged scapulae and hyperextensible joints became more evident and the diagnosis of 3-M syndrome was made; this was also confirmed by the finding of a homozygous deletion in exon 18 of the CUL7 gene, which has not been previously described.The patient also exhibited severe GHD (GH <5 ng/ml) and from the age of 18 months was treated with rhGH. Notwithstanding the early start of therapy and good compliance, his growth rate was always very low, except for the first two years of treatment and he achieved a final height of 132 cm (-6.42 SDS).