Advances in genetic etiology of isolated growth hormone deficiency / 国际儿科学杂志

Isolated growth hormone deficiency(IGHD)is a growth disorder characterized by short stature.The etiology and pathogenesis of IGHD are still not fully understood.IGHD can be caused by congenital(heredity and/or malformations)or acquired(tumors, physical trauma, inflammation, brain infections, or radiation therapy)factors.The most common genes in its genetic etiology are the growth hormone 1(GH1)and growth hormone-releasing hormone receptor(GHRHR). In rare cases, IGHD may be caused by mutations in transcription factors such as HESX1, SOX3, OTX2, POU1F1, etc.The disease phenotype of IGHD patients is highly variable.Correct diagnosis and early treatment are crucial for the long-term prognosis of IGHD patients.This review mainly discusses advance of IGHD gene mutation and disease phenotype.

The state of Sergipe contribution to GH research: from Souza Leite to Itabaianinha syndrome

ABSTRACT In the late 19th century, José Dantas de Souza Leite, a physician born in Sergipe, published the first detailed clinical description of acromegaly under the guidance of the French neurologist Pierre Marie. In 2014, the Brazilian Society of Endocrinology and Metabolism created the "José Dantas de Souza Leite Award", which is granted every two years to a Brazilian researcher who has contributed to the development of endocrinology. In 2022, the award was given to another physician from Sergipe, Manuel Hermínio de Aguiar Oliveira, from the Federal University of Sergipe for the description of "Itabaianinha syndrome" in a cohort of individuals with isolated GH deficiency due to a homozygous inactivating mutation in the GH-releasing hormone receptor gene. This research, which was carried out over almost 30 years, was performed in partnership with Roberto Salvatori from Johns Hopkins University and in collaboration with other researchers around the world. This review article tells the story of Souza Leite, some milestones in the history of GH, and summarizes the description of Itabaianinha syndrome.

Genotyping and Nucleotide Sequences of Growth Hormone Releasing Hormone and Its Receptor Genes in Egyptian Buffalo.

Aim: The hypothalamic hormone, growth hormone-releasing hormone, is the principal stimulator of pituitary growth hormone (GH) synthesis and secretion. GHRH and its receptor (GHRHR) provide important functions in the regulation of the GH axis and in the development and proliferation of pituitary somatotropic axis. This study aimed to identify the genotypes and nucleotide sequences of two multifunctional genes; growth hormone-releasing hormone (GHRH) and its receptor (GHRHR) in Egyptian buffalo. Methodology: Genomic DNA was extracted from blood samples of 100 healthy buffaloes maintained at the Mahlet Mussa and El-Gmeasa herds from 2010 to 2012. PCR was performed using primers flanking a 296-bp fragment from GHRH gene and a 425-bp fragment from GHRHR gene of Egyptian buffalo. The PCR-amplified fragments were digested with HaeIII (GHRH) and Eco57I (GHRHR), electrophoresed and analyzed on agarose gels stained with ethidium bromide. The two amplified fragments were also sequenced and aligned with published sequences. Results: Depending on the presence of the restriction site at 241

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.

Neuroendocrine Regulation of Growth Hormone Secretion / 대한소아내분비학회지

The regulation of growth hormone (GH) secretion is, to a larger extent, controlled by three hypothalamic hormones: GH-releasing hormone (GHRH), somatostatin, and ghrelin. Each binds to G protein-linked membrane receptors through which signaling occurs. We used a series of genetic and transgenic animal models with perturbations of individual compounds of the GH regulatory system to study somatotrope signaling. Impaired GH signaling is present in the lit mouse, which has a GHRH receptor (GHRH-R) mutation, and the dw rat, which has a post-receptor signaling defect. Both models also have impaired response to GH secretagogues (GHS), implying an interaction between the two signaling systems. The spontaneous dwarf rat (SDR), in which a mutation of the GH gene results in total absence of the hormone, shows characteristic changes in the hypothalamic regulatory hormones due to an absence of GH feedback and alterations in the expression of each of their pituitary receptors. Treatment of SDRs with GHRH and a GHS has allowed demonstration of a stimulatory effect GHRH on GHRH-R and GHS-R, and somatostatin receptor type 2 (sst2) expression and an inhibitory effect on sst5 expression. GH also modifies the expression of these receptors, though its effects are seen at later time periods and appear to be indirect. In the absence of GH negative feedback, both hypothalamic and pituitary expression is altered to favor stimulation of GH synthesis and release. However, in the presence of GH negative feedback, both hypothalamic and pituitary expression is altered to favor suppression of GH synthesis and release. Loss of liver insulin-like growth factor I (IGF-I) feedback on the hypothalamic-pituitary system increases GH secretion, which, in turn, stimulates liver growth. Depletion of liver-derived IGF-I increases the expression and sensitivity of pituitary GHRH-R and GHS-R. The major site of action of liver-derived IGF-I in the regulation of GH secretion is at the pituitary level. Neuropeptide Y (NPY) is not required for basal regulation of the GH axis. NPY is required for fasting-induced suppression of GHRH and SRIH expression. NPY is also required for fasting-induced augmentation of pituitary GHS-R mRNA. Overall, the results indicate a complex regulation of GH secretion in which somatotrope receptor, as well as ligand expression, exerts an important physiological role.

Ectopic Co-expression of Growth Hormone Releasing Hormone and Pituitary Adenylate Activating Peptide in Skeletal Muscle Enhance Animal Growth / 生物化学与生物物理进展

Growth hormone (GHRH) and pitutary adenylate cyclase activating peptide (PACAP) are the members of the PACAP/Glucagon superfamily,who are related in both sequence and function.Their stimulation of GH secretion and animal growth is concerned.A series of expression plasmid,pIRES1-GHRH-PACAP (P-G-P),plRESI-GHRH (P-G) and plRESI-PACAP(P-P),were constructed,extracted and purified,then transfected into CHO cell line with Lipofectamine.The expression was examined by RT-PCR,dot-ELISA and Western blotting.The biological activity of expression products was detected in rats.At 8 h after injection of transfection supematant,serum IGF-I concentrations in P-G-P group were significantly higher than that in other groups(P ＜ 0.05).PLGA encapsulating plasmid microspheres were prepared and injected intramuscularly into rabbit legs.Growth behavior and IGF-1 level were measured at day 0,15,30 and 45 after injection.Greater body weights gain and higher serum 1GF- [ levels were observed in three plasmid microsphere injection groups,compared with control group.At day 30,the body weight gain in P-G-P group was greater than saline group (81%,P＜ 0.01),P-G mierosphere group (15%,P＜ 0.05) and P-P microsphere group (7%,P＞ 0.05),serum IGF-I concentration in P-G-P microsphere group showed a 16.68% increase to P-G microsphere (P ＞ 0.05),a 17.14% increase to P-P microsphere(P ＞ 0.05) and a 50.46% increase to control (P ＜ 0.05).These results suggest that co-expression of GHRH and PACAP in one expression plasmid might exert an additive stimulation of GH secretion and growth when delivered into rabbit skeletal muscle with PLGA mierosphere.The results may provide a new approach to regulate animal growth.

Role of Glucocorticoids in Fasting-induced Changes in Hypothalamic and Pituitary Components of the Growth Hormone (GH)-axis

To directly test if elevated glucocorticoids are required for fasting-induced regulation of growth hormone (GH)-releasing hormone (GHRH), GHRH receptors (GHRH-R) and ghrelin receptors (GHS-R) expression, male rats were bilaterally adrenalectomized or sham operated. After 7 days, animals were fed ad libitum or fasted for 48 h. Bilateral adrenalectomy increased hypothalamic GHRH to 146% and decreased neuropeptide Y (NPY) mRNA to 54% of SHAM controls. Pituitary GHRH-R and GHS-R mRNA levels were decreased by adrenalectomy to 30% and 80% of sham-operated controls. In sham- operated rats, fasting suppressed hypothalamic GHRH (49%) and stimulated NPY (166%) mRNA levels, while fasting increased pituitary GHRH-R (391%) and GHS-R (218%) mRNA levels. However, in adrenalectomized rats, fasting failed to alter pituitary GHRH-R mRNA levels, while the fasting-induced suppression of GHRH and elevation of NPY and GHS-R mRNA levels remained intact. In fasted adrenalectomized rats, corticosterone replacement increased GHRH-R mRNA levels and intensified the fasting-induced decrease in GHRH, but did not alter NPY or GHS-R response. These data suggest that elevated glucocorticoids mediate the effects of fasting on hypothalamic GHRH and pituitary GHRH-R expression, while glucocorticoids are likely not the major determinant in fasting-induced increases in hypothalamic NPY and pituitary GHS-R expression.

Effect of one month ketoconazole treatment on GH, cortisol and ACTH release after ghrelin, GHRP-6 and GHRH administration in patients with cushing’s disease

GH responses to ghrelin, GHRP-6, and GHRH in Cushing’s disease (CD) are markedly blunted. There is no data about the effect of reduction of cortisol levels with steroidogenesis inhibitors, like ketoconazole, on GH secretion in CD. ACTH levels during ketoconazole treatment are controversial. The aims of this study were to compare the GH response to ghrelin, GHRP-6, and GHRH, and the ACTH and cortisol responses to ghrelin and GHRP-6 before and after one month of ketoconazole treatment in 6 untreated patients with CD. Before treatment peak GH (mg/L; mean ± SEM) after ghrelin, GHRP-6, and GHRH administration was 10.0 ± 4.5; 3.8 ± 1.6, and 0.6 ± 0.2, respectively. After one month of ketoconazole there was a significant decrease in urinary cortisol values (mean reduction: 75 percent), but GH responses did not change (7.0 ± 2.0; 3.1 ± 0.8; 0.9 ± 0.2, respectively). After treatment, there was a significant reduction in cortisol (mg/dL) responses to ghrelin (before: 30.6 ± 5.2; after: 24.2 ± 5.1). No significant changes in ACTH (pg/mL) responses before (ghrelin: 210.9 ± 69.9; GHRP-6: 199.8 ± 88.8) and after treatment (ghrelin: 159.7 ± 40.3; GHRP-6: 227 ± 127.2) were observed. In conclusion, after short-term ketoconazole treatment there are no changes in GH or ACTH responses, despite a major decrease of cortisol levels. A longer period of treatment might be necessary for the recovery of pituitary function.

Na doença de Cushing (DC), as respostas do GH à ghrelina, ao GHRP-6 e ao GHRH estão diminuídas. Não existem dados sobre o efeito da redução dos níveis de cortisol, após cetoconazol, na secreção de GH na DC. Nessa situação, os níveis de ACTH são variáveis. Os objetivos do estudo são comparar as respostas do GH à administração de ghrelina, GHRP-6 e GHRH, e de ACTH e cortisol à ghrelina e ao GHRP-6 antes e após um mês de tratamento com cetoconazol em 6 pacientes com DC não tratados. Antes do tratamento, o pico de GH (mg/L; média ± EPM) após a administração de ghrelina, GHRP-6 e GHRH foi de 10,0 ± 4,5; 3,8 ± 1,6 e 0,6 ± 0,2, respectivamente. Após um mês de cetoconazol, ocorreu diminuição significante do cortisol urinário (redução média: 75 por cento), mas as respostas de GH permaneceram inalteradas (7,0 ± 2,0; 3,1 ± 0,8; 0,9 ± 0,2, respectivamente). Após o tratamento, houve redução da resposta de cortisol (mg/dL) à ghrelina (antes: 30,6 ± 5,2; após: 24,2 ± 5,1), mas não ocorreram mudanças nas respostas de ACTH (pg/mL) (ghrelina antes: 210,9 ± 69,9; após: 159,7 ± 40,3; GHRP-6 antes: 199,8 ± 88,8; após: 227 ± 127,2). Assim, o tratamento a curto prazo com cetoconazol não modificou as respostas de GH ou ACTH, apesar da redução do cortisol. Para a recuperação da função hipofisária deve ser necessário um período de tratamento maior.

Changes in Somatostatin Receptor mRNA Levels by G Protein Mutation in GH3 Cells Which Show Responsiveness to Growth Hormone-Releasing Hormone / 대한내분비학회지

BACKGROUNDS: GH3 cells lack growth hormone(GH)-releasing hormone(GHRH) receptors. In this study, GH3 cells permanently transfected with human GHRH receptor cDNA(GH3-GHRHR cells), were established in order to examine the effects of GHRH and G protein mutation(gsp oncogene) on the levels of somatostatin receptor mRNA. METHODS: GH3 cells were permanently transfected with a plasmid expressing human GHRH receptor cDNA. The GHRH receptor mRNA was detected by RT-PCR. The responsiveness to GHRH was evaluated using a GHRH binding assay, Western blot analysis, Northern blot analysis, and measurements of the intracellular cAMP levels and GH release. Cells were transiently transfected with the gsp oncogene, and then treated with GHRH or octreotide for 4h. The sst1 and sst2 mRNA levels were measured using real-time RT-PCR analyses. RESULTS: GHRH receptor mRNA was detected in the GH3 cells permanently transfected with human GHRH receptor cDNA. The GHRH binding assay showed that GHRH was bound to the GH3-GHRHR cells. The GHRH treatment increased the intracellular cAMP levels, GH release, GH mRNA levels, and MAPK activity, as well as the levels of sst1 and sst2 mRNA. Transient expression of the gsp oncogene for 48h increased the cAMP, GH release, and levels of sst1 and sst2 mRNA. In the gsp-transfected GH3-GHRHR cells, GHRH stimulation resulted in decreases in the magnitude of the increase in the levels of sst1 and sst2 mRNA compared to those transfected with a control vector. Octreotide treatment did not alter the levels of sst1 and sst2 mRNA in either the control or gsp-transfected cells. CONCLUSION: These results suggest that GH3 cells permanently transfected with the GHRH receptor are useful in the in vitro studies on the actions of GHRH. The gsp oncogene was shown to increases the levels of sst1 and sst2 mRNA in GH3 cells, but these findings are unlikely to be the major mechanism by which gsp-positive pituitary tumors show a greater response to somatostatin. The discrepancy between the in vivo and these in vitro results should be examined further.

Modulation of Pituitary Somatostatin Receptor Subtype (sst1-5) mRNA Levels by Growth Hormone (GH) -Releasing Hormone in Purified Somatotropes

We have previously reported that expression of the somatostatin receptor subtypes, sst1-5, is differentially regulated by growth hormone (GH) -releasing hormone (GHRH) and forskolin (FSK), in vitro. GHRH binds to membrane receptors selectively located on pituitary somatotropes, activates adenylyl cyclase (AC) and increases sst1 and sst2 and decreases sst5 mRNA levels, without significantly altering the expression of sst3 and sst4. In contrast FSK directly activates AC in all pituitary cell types and increases sst1 and sst2 mRNA levels and decreases sst3, sst4 and sst5 expression. Two explanations could account for these differential effects: 1) GHRH inhibits sst3 and sst4 expression in somatotropes, but this inhibitory effect is masked by expression of these receptors in unresponsive pituitary cell types, and 2) FSK inhibits sst3 and sst4 expression levels in pituitary cell types other than somatotropes. To differentiate between these two possibilities, somatotropes were sequentially labeled with monkey anti-rat GH antiserum, biotinylated goat anti-human IgG, and streptavidin-PE and subsequently purified by fluorescent-activated cell sorting (FACS). The resultant cell population consisted of 95% somatotropes, as determined by GH immunohistochemistry using a primary GH antiserum different from that used for FACS sorting. Purified somatotropes were cultured for 3 days and treated for 4 h with vehicle, GHRH (10 nM) or FSK (10micrometer). Total RNA was isolated by column extraction and specific receptor mRNA levels were determined by semi-quantitative multiplex RT-PCR. Under basal conditions, the relative expression levels of the various somatostatin receptor subtypes were sst2> sst5> sst3=sst1> sst4. GHRH treatment increased sst1 and sst2 mRNA levels and decreased sst3, sst4 and sst5 mRNA levels in purified somatotropes, comparable to the effects of FSK on purified somatotropes and mixed pituitary cell cultures. Taken together, these results demonstrate that GHRH acutely modulates the expression of all somatostatin receptor subtypes within GH-producing cells and its actions are likely mediated by activation of AC.

GROWTH HORMONE-RELEASING HORMONE 1-44 STIMULATES GROWTH VELOCITY IN HYPOTHALAMIC GROWTH HORMONE-DEMCIENT PATIENTS / 中华内分泌代谢杂志

10?g/L and 5 patients with serum GH peak between 5 - 10?g/L in 5-day GHRH stimulation test were about the same. There was negative correlation between chronological age and growth velocity after treatment with GHRH 1 -44 for 6 months (r = 0.7078, P