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High neonatal growth hormone (GH) secretion has been described in several species. However, the neuroendocrine mechanisms behind this surge remain unknown. Thus, the pattern of postnatal GH secretion was investigated in mice and rats. Blood GH levels were very high on postnatal day (P)1 and progressively decreased until near zero by P17 in C57BL/6 mice without sex differences. This pattern was similar to that observed in rats, except that female rats showed higher GH levels on P1 than males. In comparison, follicle-stimulating hormone exhibited higher secretion in females during the first 3 weeks of life. Hypothalamic Sst mRNA and somatostatin neuroendocrine terminals in the median eminence were higher in P20/P21 mice than in newborns. Knockout mice for GH-releasing hormone (GHRH) receptor showed no GH surge, whereas knockdown mice for the Sst gene displayed increased neonatal GH peak. Leptin deficiency caused only minor effects on early-life GH secretion. GH receptor ablation in neurons or the entire body did not affect neonatal GH secretion, but the subsequent reduction in blood GH levels was attenuated or prevented by these genetic manipulations, respectively. This phenotype was also observed in knockout mice for the insulin-like growth factor-1 (IGF-1) receptor in GHRH neurons. Moreover, glucose-induced hyperglycemia overstimulated GH secretion in neonatal mice. In conclusion, GH surge in the first days of life is not regulated by negative feedback loops. However, neonatal GH secretion requires GHRH receptor, and is modulated by somatostatin and blood glucose levels, suggesting that this surge is controlled by hypothalamic-pituitary communication., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com. See the journal About page for additional terms.)

Small extracellular vesicles (sEV) have emerged as a novel mode of intercellular material transport and information transmission. It has been suggested hormones may regulate the production and function of sEV. However, the specific impact of growth hormone-releasing hormone (GHRH) on pituitary sEV production and the role of sEV in the regulation of the GHRH-GH-IGF axis has not been previously reported. The results of the present study demonstrated that GHRH increased the production of pituitary sEV by promoting the expression of Rab27a. More importantly, GHRH induced alterations in protein and miRNA levels within GH3-sEV components. Notably, GH3-sEV with GHRH treatment exhibited the enhanced ability to impede BRL 3A cell proliferation and the expression of IGF-1. Conclusively, for the first time, we corroborate the influence of GHRH on pituitary sEV, thereby presenting novel evidence for how sEV participates in the balance of the GHRH-GH-IGF axis. Importantly, this study provides new insight into a novel balance mechanism mediated by sEV within the endocrine system., (© 2024. The Author(s).)

The development of resistance remains the primary challenge in treating castration-resistant prostate cancer (CRPC). GHRH receptors (GHRH-R), which are coupled to G-proteins (GPCRs), can mediate EGFR transactivation, offering an alternative pathway for tumour survival. This study aimed to evaluate the effects of the GHRH-R antagonist MIA-690, in combination with the EGFR inhibitor Gefitinib, on cell viability, adhesion, gelatinolytic activity, and the cell cycle in advanced prostate cancer PC-3 cells. The findings demonstrate a synergistic effect between MIA-690 and Gefitinib, leading to the inhibition of cell viability, adhesion, and metalloprotease activity. Cell cycle analysis suggests that both compounds induce cell cycle arrest, both individually and in combination. Furthermore, similar effects of the GHRH-R antagonist MIA-690 combined with Gefitinib were observed in PC-3 tumours developed by subcutaneous injection in athymic nude mice 36 days post-inoculation. These results indicate that combined therapy with a GHRH-R antagonist and an EGFR inhibitor exerts a stronger antitumor effect compared to monotherapy by preventing transactivation between EGFR and GHRH-R in CRPC.

Acromegaly due to ectopic secretion of growth hormone-releasing hormone (GHRH) is a rare disorder. The signs and symptoms of ectopic acromegaly are indistinguishable from acromegaly due to a somatotroph adenoma. A 35-year-old female presented with secondary amenorrhea for 10 years, intermittent headache, and reduced vision in both eyes for 4 years, which worsened over 4 months before presentation. Additionally, she was diagnosed with uncontrolled diabetes mellitus. On examination, she had coarse facial features, a fleshy nose, and acral enlargement. She had diminished visual acuity (left>right) and bitemporal hemianopia on perimetry. Biochemical investigations revealed elevated IGF-1 [588 ng/ml, reference range (RR) 100-242], markedly elevated basal growth hormone (>80 ng/ml; RR, 0.12-9.88), and hyperprolactinemia in the tumoral range (832 ng/ml; RR, 5-25). MRI sella demonstrated a 22×30×34mm sellar-suprasellar mass with T2 hypointensity. Chest imaging revealed a 75×87×106mm left lung mass, which was found to be a well-differentiated neuroendocrine tumor (NET) on biopsy. Plasma GHRH levels were elevated [38,088 ng/l; RR, <250-300], and a diagnosis of ectopic acromegaly secondary to lung neuroendocrine tumor was considered. During workup, the patient developed in-hospital pituitary apoplexy, which improved with medical management. After a left pneumonectomy, her clinical features of acromegaly improved, her diabetes underwent remission, and there was a marked reduction in plasma GHRH and pituitary size. Histopathology was suggestive of a neuroendocrine tumor, with immunohistochemistry positive for GHRH and negative for prolactin. Her final diagnosis was ectopic acromegaly due to GHRH secreting a lung NET with pituitary somatotroph and lactotroph pituitary hyperplasia and apoplexy in the hyperplastic pituitary., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Gupta, Kasaliwal, Das, Sharma, Kaur, Vasiljevic, Raverot, Korbonits and Dutta.)

Purpose: It is unclear whether the age-related decline in the somatotropic axis stems from a reduced growth hormone (GH) production in the pituitary gland, or from a peripheral origin akin to an acquired GH resistance. With the help of a GHRH/arginine test, high-aged multimorbid hospitalized patients with IGF-I deficiency are to be tested to determine whether there is primarily a pituitary GH deficiency in the sense of a somatopause., Methods: Seventeen multimorbid patients (eleven men and six women) with a mean age of 82 years, with IGF-I concentrations below two standard deviations of 30-year-old men and women were identified. Patients suffered from a variety of common age-related stable diseases including coronary artery disease, chronic liver or kidney disease, chronic heart failure as well as acute conditions e.g., urosepsis or endocarditis. To assess the somatotropic axis they underwent a GHRH/arginine test. Results were evaluated using descriptive statistics., Results: In average, the peak concentration of GH after stimulation was 14.8 µg/L with a range from 2.76 to 47.4 µg/L. Taking into account both, gender and BMI (with a mean of 26.5 kg/m²) for each participant, the pituitary gland was adequately stimulated in 16 out of the 17 patients. No patient reported common side effects related to the GHRH/arginine test., Conclusion: The somatotroph pituitary gland retains its secretory capacity in the advanced aged. Therefore, age does not seem to be the driving pacemaker for the functional decline of the somatotropic axis within the aged population., (© 2024. The Author(s).)

Growth hormone-releasing hormone-deficient (GHRH-KO) mice have previously been characterized by lower body weight, disproportionately high body fat accumulation, preferential metabolism of lipids compared to carbohydrates, improved insulin sensitivity, and an extended lifespan. That these mice are long-lived and insulin-sensitive conflicts with the notion that adipose tissue accumulation drives the health detriments associated with obesity (i.e., diabetes), and indicates that GH signaling may be necessary for the development of adverse effects linked to obesity. This prompts investigation into the ultimate effect of diet-induced obesity on the lifespan of these long-lived mice. To this end, we initiated high-fat feeding in mid and late-life in GHRH-KO and wild-type (WT) mice. We carried out extensive lifespan analysis coupled with glucose/insulin tolerance testing and indirect calorimetry to gauge the metabolic effect of high-fat dietary stress through adulthood on these mice. We show that under high-fat diet (HFD) conditions, GHRH-KO mice display extended lifespans relative to WT controls. We also show that GHRH-KO mice are more insulin-sensitive and display less dramatic changes in their metabolism relative to WT mice, with GHRH-KO mice fed HFD displaying respiratory exchange ratios and glucose oxidation rates comparable to control-diet fed GHRH-KO mice, while WT mice fed HFD showed significant reductions in these parameters. Our results indicate that GH deficiency protects against the adverse effects of diet-induced obesity in later life., (© 2024 The Author(s). Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Growth hormone-releasing hormone antagonists (GHRHAnt) have been associated with antitumor and antioxidative activities. The present study investigates for the first time the effects of those compounds towards pro-inflammatory cytokine expression in a murine model of cecal ligation and puncture (CLP) - induced sepsis. The results indicate that GHRHAnt JV-1-36 significantly suppressed IL-1α, IL-6, and pSTAT3 activation in septic lungs. Moreover, GHRHAnt treatment reduced bronchoalveolar lavage fluid (BALF) protein concentration, suggesting a protective effect of that compound in sepsis-induced lung edema. Based on those findings, it is suggested that GHRHAnt may represent an exciting new therapeutic possibility in sepsis-induced endotoxemia and lung injury., Competing Interests: Declaration of competing interest The authors declare no conflict of interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Growth hormone-releasing hormone (GHRH) has been widely shown to stimulate growth hormone (GH) production via binding to GHRH receptor GHRHR in various species of vertebrates, but information regarding the functional roles of GHRH and GHRHR in the protochordate amphioxus remains rather scarce. We showed here that two mature peptides, BjGHRH-1 and BjGHRH-2, encoded by BjGHRH precursor, and a single BjGHRHR protein were identified in the amphioxus Branchiostoma. japonicum. Like the distribution profiles of vertebrate GHRHs and GHRHRs, both the genes Bjghrh and Bjghrhr were widely expressed in the different tissues of amphioxus, including in the cerebral vesicle, Hatschek's pit, neural tube, gill, hepatic caecum, notochord, testis and ovary. Moreover, both BjGHRH-1 and BjGHRH-2 interacted with BjGHRHR, and triggered the cAMP/PKA signal pathway in a dose-dependent manner. Importantly, BjGHRH-1 and BjGHRH-2 were both able to activate the expression of GH-like gene in the cells of Hatschek's pit. These indicate that a functional vertebrate-like GHRH-GHRHR axis had already emerged in amphioxus, which is a seminal innovation making physiological divergence including reproduction, growth, metabolism, stress and osmoregulation possible during the early evolution of vertebrates., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Hematological and oncological diseases are still among the leading causes of childhood mortality. Expression of growth hormone-releasing hormone (GHRH) and its receptors (GHRH-R) has been previously demonstrated in various human tumors, but very limited findings are available about the presence and potential function of GHRH-Rs in oncological and hematological disorders of children. In this study, we aimed to investigate the expression of mRNA for GHRH and splice variant 1 (SV) of GHRH-R in 15 pediatric hematological/oncological specimens by RT-PCR. The presence and binding characteristics of GHRH-R protein were also studied by Western blot and ligand competition assays. Of the fifteen specimens studied, eleven pediatric samples (73%) showed the expression of mRNA for GHRH. These eleven samples also expressed mRNA for GHRH receptor SV1. GHRH-R protein was found to be expressed in two benign tumor samples and five malignant tumors examined by Western blot. The presence of specific, high affinity binding sites on GHRH-R was demonstrated in all of the seven human pediatric solid tumor samples investigated. Our results show that the expression of GHRH and SV1 of GHRH-R in hemato-oncological diseases in children can pave the way for further investigation of GHRH-Rs as potential molecular targets for diagnosis and therapy.

Dysregulation of growth hormone (GH) signaling consistently leads to increased lifespan in laboratory rodents, yet the precise mechanisms driving this extension remain unclear. Understanding the molecular underpinnings of the beneficial effects associated with GH deficiency could unveil novel therapeutic targets for promoting healthy aging and longevity. In our pursuit of identifying metabolites implicated in aging, we conducted an unbiased lipidomic analysis of serum samples from growth hormone-releasing hormone knockout (GHRH-KO) female mice and their littermate controls. Employing a targeted lipidomic approach, we specifically investigated ceramide levels in GHRH-KO mice, a well-established model of enhanced longevity. While younger GHRH-KO mice did not exhibit notable differences in serum lipids, older counterparts demonstrated significant reductions in over one-third of the evaluated lipids. In employing the same analysis in liver tissue, GHRH-KO mice showed pronounced downregulation of numerous ceramides and hexosylceramides, which have been shown to elicit many of the tissue defects that accompany aging (e.g., insulin resistance, oxidative stress, and cell death). Additionally, gene expression analysis in the liver tissue of adult GHRH-KO mice identified substantial decreases in several ceramide synthesis genes, indicating that these alterations are, at least in part, attributed to GHRH-KO-induced transcriptional changes. These findings provide the first evidence of disrupted ceramide metabolism in a long-lived mammal. This study sheds light on the intricate connections between GH deficiency, ceramide levels, and the molecular mechanisms influencing lifespan extension., (© 2024 The Author(s). Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.

Growth hormone-releasing hormone (Ghrh) neurons in the dorsomedial ventromedial hypothalamic nucleus (VMNdm) express the metabolic transcription factor steroidogenic factor-1 and hypoglycemia-sensitive neurochemicals of diverse chemical structures, transmission modes, and temporal signaling profiles. Ghrh imposes neuromodulatory control of coexpressed transmitters. Multiple metabolic sensory mechanisms are employed in the brain, including screening of the critical nutrient glucose or the energy currency ATP. Here, combinatory laser-catapult-microdissection/single-cell multiplex qPCR tools were used to investigate whether these neurons possess molecular machinery for monitoring cellular metabolic status and if these biomarkers exhibit sex-specific sensitivity to insulin-induced hypoglycemia. Data show that hypoglycemia up- (male) or downregulated (female) Ghrh neuron glucokinase (Gck) mRNA; Ghrh gene silencing decreased baseline and hypoglycemic patterns of Gck gene expression in each sex. Ghrh neuron glucokinase regulatory protein (Gckr) transcript levels were respectively diminished or augmented in hypoglycemic male vs female rats; this mRNA profile was decreased by Ghrh siRNA in both sexes. Gene transcripts encoding catalytic alpha subunits of the energy monitor 5-AMP-activated protein kinase (AMPK), i.e., Prkaa1 and 2, were increased by hypoglycemia in males, yet only the former mRNA was hypoglycemia-sensitive in females. Ghrh siRNA downregulated baseline and hypoglycemia-associated Prkaa subunit mRNAs in males but elicited divergent changes in Prkaa2 transcripts in eu- vs hypoglycemic females. Results provide unique evidence that VMNdm Ghrh neurons express the characterized metabolic sensor biomarkers glucokinase and AMPK and that the corresponding gene profiles exhibit distinctive sex-dimorphic transcriptional responses to hypoglycemia. Data further document Ghrh neuromodulation of baseline and hypoglycemic transcription patterns of these metabolic gene profiles.

The negative coordination of growth hormone secretagogue receptor (GHS-R) and growth hormone-releasing hormone receptor (GHRH-R) involves in the repair processes of cellular injury. The allosteric U- or H-like modified GHRH dimer Grinodin and 2Y were comparatively evaluated in normal Kunming mice and hamster infertility models induced by CPA treatment. 1-3-9 µg of Grinodin or 2Y per hamster stem-cell-exhaustion model was subcutaneously administered once a week, respectively inducing 75-69-46 or 45-13-50 % of birth rates. In comparison, the similar mole of human menopausal gonadotropin (hMG) or human growth hormone (hGH) was administered once a day but caused just 25 or 20 % of birth rates. Grinodin induced more big ovarian follicles and corpora lutea than 2Y, hMG, hGH. The hMG-treated group was observed many distorted interstitial cells and more connective tissues and the hGH-treated group had few ovarian follicles. 2Y had a plasma lifetime of 21 days and higher GH release in mice, inducing lower birth rate and stronger individual specificity in reproduction as well as only promoting the proliferation of mesenchymal-stem-cells (MSCs) in the models. In comparison, Grinodin had a plasma lifetime of 30 days and much lower GH release in mice. It significantly promoted the proliferation and activation of ovarian MSCs together with the development of follicles in the models by increasing Ki67 and GHS-R expressions, and decreasing GHRH-R expression in a dose-dependent manner. However, the high GH and excessive estrogen levels in the models showed a dose-dependent reduction in fertility. Therefore, unlike 2Y, the low dose of Grinodin specifically shows low GHS-R and high GHRH-R expressions thus evades GH and estrogen release and improves functions of organs, resulting in an increase of fertility., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Inflammation, demyelination, and axonal damage to the central nervous system (CNS) are the hallmarks of multiple sclerosis (MS) and its representative animal model, experimental autoimmune encephalomyelitis (EAE). There is scientific evidence for the involvement of growth hormone (GH) in autoimmune regulation. Previous data on the relationship between the GH/insulin like growth factor-1 (IGF-1) axis and MS/EAE are inconclusive; therefore, the aim of our study was to investigate the changes in the GH axis during acute monophasic EAE. The results show that the gene expression of Ghrh and Sst in the hypothalamus does not change, except for Npy and Agrp , while at the pituitary level the Gh , Ghrhr and Ghr genes are upregulated. Interestingly, the cell volume of somatotropic cells in the pituitary gland remains unchanged at the peak of the disease. We found elevated serum GH levels in association with low IGF-1 concentration and downregulated Ghr and Igf1r expression in the liver, indicating a condition resembling GH resistance. This is likely due to inadequate nutrient intake at the peak of the disease when inflammation in the CNS is greatest. Considering that GH secretion is finely regulated by numerous central and peripheral signals, the involvement of the GH/IGF-1 axis in MS/EAE should be thoroughly investigated for possible future therapeutic strategies, especially with a view to improving EAE disease.

Ectopic acromegaly is a rare condition caused by extrapituitary central or peripheral neuroendocrine tumours (NET) that hypersecrete GH or, more commonly, GHRH. It affects less than 1% of acromegaly patients and a misdiagnosis of classic acromegaly can lead to an inappropriate pituitary surgery. Four types of ectopic acromegaly have been described: 1) Central ectopic GH-secretion: Careful cross-sectional imaging is required to exclude ectopic pituitary adenomas. 2) Peripheral GH secretion: Extremely rare. 3) Central ectopic GHRH secretion: Sellar gangliocytomas immunohistochemically positive for GHRH are found after pituitary surgery. 4) Peripheral GHRH secretion: The most common type of ectopic acromegaly is due to peripheral GHRH-secreting NETs. Tumours are large and usually located in the lungs or pancreas. Pituitary hyperplasia resulting from chronic GHRH stimulation is difficult to detect or can be misinterpreted as pituitary adenoma in the MRI. Measurement of serum GHRH levels is a specific and useful diagnostic tool. Surgery of GHRH-secreting NETs is often curative., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The prospect that the ventromedial hypothalamic nucleus (VMN) transcription factor steroidogenic factor-1/NR5A1 (SF-1) may exert sex-dimorphic control of glucose counterregulation is unresolved. Recent studies in male rats show that SF-1 regulates transcription of co-expressed hypoglycemia-sensitive neurochemicals in dorsomedial VMN growth hormone-releasing hormone (Ghrh) neurons. Gene knockdown and laser-catapult-microdissection/single-cell multiplex qPCR techniques were used here in a female rat model to determine if SF-1 control of Ghrh neuron transmitter marker, energy sensor, and estrogen receptor (ER) variant mRNAs varies according to sex. Data show that in females, hypoglycemia elicits a gain of SF-1 inhibitory control of VMNdm Ghrh neuron Ghrh and Ghrh-receptor gene profiles and loss of augmentation of glutaminase transcription; SF-1 gene silencing diminished eu- and hypoglycemic patterns of neuronal nitric oxide gene transcription. SF-1 imposes divergent control of baseline and hypoglycemic glutamate decarboxylase 65 (GAD)-1 (stimulatory) versus GAD2 (inhibitory) mRNAs in that sex. SF-1 stimulates baseline VMNdm Ghrh neuron PRKAA1/AMPKα1 and PRKAA2/AMPKα2 gene expression, yet causes opposite changes in these gene profiles during hypoglycemia. SF-1 exerts glucose-dependent control of ER-alpha and G-protein-coupled ER-1 transcription, but blunts ER-beta gene profiles during eu- and hypoglycemia. In females, SF-1 knockdown did not affect hypercorticosteronemia or hyperglucagonemia, but blunted hypoglycemic suppression of growth hormone secretion. Results show that SF-1 expression is critical for female rat VMNdm Ghrh neuron counterregulatory neurochemical, AMPK catalytic subunit, and ER gene transcription responses to hypoglycemia. Sex differences in direction of SF-1 control of distinctive gene profiles may result in observed disparities in SF-1 regulation of counterregulatory hormone secretion between sexes.

Ventromedial hypothalamic nucleus (VMN) growth hormone-releasing hormone (Ghrh) neurotransmission shapes counterregulatory hormone secretion. Dorsomedial VMN Ghrh neurons express the metabolic-sensitive transcription factor steroidogenic factor-1/NR5A1 (SF-1). In vivo SF-1 gene knockdown tools were used here to address the premise that in male rats, SF-1 may regulate basal and/or hypoglycemic patterns of Ghrh, co-transmitter biosynthetic enzyme, and estrogen receptor (ER) gene expression in these neurons. Single-cell multiplex qPCR analyses showed that SF-1 regulates basal profiles of mRNAs that encode Ghrh and protein markers for neurochemicals that suppress (γ-aminobutyric acid) or enhance (nitric oxide; glutamate) counterregulation. SF-1 siRNA pretreatment respectively exacerbated or blunted hypoglycemia-associated inhibition of glutamate decarboxylase 67 (GAD 67 /GAD1) and - 65 (GAD 65 /GAD2) transcripts. Hypoglycemia augmented or reduced nitric oxide synthase and glutaminase mRNAs, responses that were attenuated by SF-1 gene silencing. Ghrh and Ghrh receptor transcripts were correspondingly refractory to or increased by hypoglycemia, yet SF-1 knockdown decreased both gene profiles. Hypoglycemic inhibition of ER-alpha and G protein-coupled-ER gene expression was amplified by SF-1 siRNA pretreatment, whereas as ER-beta mRNA was amplified. SF-1 knockdown decreased (corticosterone) or elevated [glucagon, growth hormone (GH)] basal counterregulatory hormone profiles, but amplified hypoglycemic hypercorticosteronemia and -glucagonemia or prevented elevated GH release. Outcomes document SF-1 control of VMN Ghrh neuron counterregulatory neurotransmitter and ER gene transcription. SF-1 likely regulates Ghrh nerve cell receptivity to estradiol and release of distinctive neurochemicals during glucose homeostasis and systemic imbalance. VMN Ghrh neurons emerge as a likely substrate for SF-1 control of glucose counterregulation in the male rat.

Growth hormone (GH)-releasing hormone (GHRH) has been suggested to play a crucial role in brain function. We aimed to further investigate the effects of a novel GHRH antagonist of the Miami (MIA) series, MIA-602, on emotional disorders and explore the relationships between the endocrine system and mood disorders. In this context, the effects induced by MIA-602 were also analyzed in comparison to vehicle-treated mice with GH deficiency due to generalized ablation of the GHRH gene (GHRH knock out (GHRHKO)). We show that the chronic subcutaneous administration of MIA-602 to wild type (+/+) mice, as well as generalized ablation of the GHRH gene, is associated with anxiolytic and antidepressant behavior. Moreover, immunohistochemical and Western blot analyses suggested an evident activation of Nrf2, HO1, and NQO1 in the prefrontal cortex of both +/+ mice treated with MIA-602 (+/+ MIA-602) and homozygous GHRHKO (-/- control) animals. Finally, we also found significantly decreased COX-2 , iNOS , NFkB , and TNF-α gene expressions, as well as increased P-AKT and AKT levels in +/+ MIA-602 and -/- control animals compared to +/+ mice treated with vehicle (+/+ control). We hypothesize that the generalized ablation of the GHRH gene leads to a dysregulation of neural pathways, which is mimicked by GHRH antagonist treatment.

Neonatal malnutrition is one of the most common causes of neurological disorders. However, the mechanism of action of the factors associated with neonatal nutrition in the brain remains unclear. In this study, we focused on fibroblast growth factor (FGF) 21 to elucidate the effects of malnutrition on the neonatal brain. FGF21 is an endocrine factor produced by the liver during lactation which is the main source of nutrition during the neonatal period. In this study, malnourishment during nursing mice induced decreased levels of Fgf21 mRNA in the liver and decreased levels of FGF21 in the serum. RNA-seq analysis of neonatal mouse brain tissue revealed that FGF21 controlled the expression of Kalrn-201 in the neonatal mouse brain. Kalrn-201 is a transcript of Kalirin, a Ras homologous guanine nucleotide exchange factor at the synapse. In mouse neurons, FGF21 induced the expression of Kalirin-7 (a Kalirin isoform) by down-regulating Kalrn-201. FGF21-induced Kalirin-7 stimulated neurite outgrowth in Neuro-2a cells. FGF21 also induced Growth hormone-releasing hormone (GHRH) expression in Neuro-2a cells. Kalirin-7 and GHRH expression induced by FGF21 was altered by inhibiting the activity of SH2-containing tyrosine phosphatase (SHP2) which is located downstream of the FGF receptor (FGFR). Additionally, malnourished nursing induced intron retention of the SHP2 gene (Ptpn11), resulting in the alteration of Kalirin-7 and GHRH expression by FGF21 signaling. Ptpn11 intron retention is suggested to be involved in regulating SHP2 activity. Taken together, these results suggest that FGF21 plays a critical role in the induction of neuronal neurite outgrowth and GHRH secretion in the neonatal brain, and this mechanism is regulated by SHP2. Thus, Ptpn11 intron retention induced by malnourished nursing may be involved in SHP2 activity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

The ventromedial hypothalamic nucleus (VMN) controls glucose counter-regulation, including pituitary growth hormone (GH) secretion. VMN neurons that express the transcription factor steroidogenic factor-1/NR5A1 (SF-1) participate in glucose homeostasis. Research utilized in vivo gene knockdown tools to determine if VMN growth hormone-releasing hormone (Ghrh) regulates hypoglycemic patterns of glucagon, corticosterone, and GH outflow according to sex. Intra-VMN Ghrh siRNA administration blunted hypoglycemic hypercorticosteronemia in each sex, but abolished elevated GH release in males only. Single-cell multiplex qPCR showed that dorsomedial VMN (VMNdm) Ghrh neurons express mRNAs encoding Ghrh, SF-1, and protein markers for glucose-inhibitory (γ-aminobutyric acid) or -stimulatory (nitric oxide; glutamate) neurotransmitters. Hypoglycemia decreased glutamate decarboxylase 67 (GAD 67 ) transcripts in male, not female VMNdm Ghrh/SF-1 neurons, a response that was refractory to Ghrh siRNA. Ghrh gene knockdown prevented, in each sex, hypoglycemic down-regulation of Ghrh/SF-1 nerve cell GAD 65 transcription. Ghrh siRNA amplified hypoglycemia-associated up-regulation of Ghrh/SF-1 neuron nitric oxide synthase mRNA in male and female, without affecting glutaminase gene expression. Ghrh gene knockdown altered Ghrh/SF-1 neuron estrogen receptor-alpha (ERα) and ER-beta transcripts in hypoglycemic male, not female rats, but up-regulated GPR81 lactate receptor mRNA in both sexes. Outcomes infer that VMNdm Ghrh/SF-1 neurons may be an effector of SF-1 control of counter-regulation, and document Ghrh modulation of hypoglycemic patterns of glucose-regulatory neurotransmitter along with estradiol and lactate receptor gene transcription in these cells. Co-transmission of glucose-inhibitory and -stimulatory neurochemicals of diverse chemical structure, spatial, and temporal profiles may enable VMNdm Ghrh neurons to provide complex dynamic, sex-specific input to the brain glucose-regulatory network., (Copyright © 2023 IBRO. Published by Elsevier Ltd. All rights reserved.)

Dysregulation of Th17 cell differentiation and pathogenicity contributes to multiple autoimmune and inflammatory diseases. Previously growth hormone releasing hormone receptor (GHRH-R) deficient mice have been reported to be less susceptible to the induction of experimental autoimmune encephalomyelitis. Here, we show GHRH-R is an important regulator of Th17 cell differentiation in Th17 cell-mediated ocular and neural inflammation. We find that GHRH-R is not expressed in naïve CD4 + T cells, while its expression is induced throughout Th17 cell differentiation in vitro. Mechanistically, GHRH-R activates the JAK-STAT3 pathway, increases the phosphorylation of STAT3, enhances both non-pathogenic and pathogenic Th17 cell differentiation and promotes the gene expression signatures of pathogenic Th17 cells. Enhancing this signaling by GHRH agonist promotes, while inhibiting this signaling by GHRH antagonist or GHRH-R deficiency reduces, Th17 cell differentiation in vitro and Th17 cell-mediated ocular and neural inflammation in vivo. Thus, GHRH-R signaling functions as a critical factor that regulates Th17 cell differentiation and Th17 cell-mediated autoimmune ocular and neural inflammation., (© 2023. The Author(s).)

Growth hormone-releasing hormone (GHRH) regulates the synthesis of growth hormone from the anterior pituitary gland, and it is involved in inflammatory responses. On the other hand, GHRH antagonists (GHRHAnt) exhibit the opposite effects, resulting in endothelial barrier enhancement. Exposure to hydrochloric acid (HCL) is associated with acute and chronic lung injury. In this study, we investigate the effects of GHRHAnt in HCL-induced endothelial barrier dysfunction, utilizing commercially available bovine pulmonary artery endothelial cells (BPAEC). Cell viability was measured by utilizing 3-(4,5-dimethylthiazol2-yl)- 2,5-diphenyltetrazolium bromide (MTT) assay. Moreover, fluorescein isothiocyanate (FITC)-dextran was used to assess barrier function. Our observations suggest that GHRHAnt exert protective effects against HCL-induced endothelial breakdown, since those peptides counteract HCL-triggered paracellular hyperpermeability. Based on those findings, we propose that GHRHAnt represent a new therapeutic approach towards HCL-induced endothelial injury., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Purpose: Growth hormone-releasing hormone (GHRH) is a hypothalamic hormone, which regulates growth hormone release from the anterior pituitary gland. GHRH antagonists (GHRHAnt) are anticancer agents, which also exert robust anti-inflammatory activities in malignancies. GHRHAnt exhibit anti-oxidative and anti-inflammatory effects in vascular endothelial cells, indicating their potential use against disorders related to barrier dysfunction (e.g. sepsis). Herein, we aim to investigate the effects of GHRHAnt against lung endothelial hyperpermeability., Methods: The in vitro effects of GHRHAnt in H 2 O 2 -induced endothelial barrier dysfunction were investigated in bovine pulmonary artery endothelial cells (BPAEC). Electric cell-substrate impedance sensing (ECIS) was utilized to measure transendothelial resistance, an indicator of barrier function., Results: Our results demonstrate that GHRHAnt protect against H 2 O 2 -induced endothelial barrier disruption via P53 and cofilin modulation. Both proteins are crucial modulators of vascular integrity. Moreover, GHRHAnt prevent H 2 O 2 - induced decrease in transendothelial resistance., Conclusions: GHRHAnt represent a promising therapeutic intervention towards diseases related to lung endothelial hyperpermeability, such as acute respiratory distress syndrome - related or not to COVID-19 - and sepsis. Targeted medicine for those potentially lethal disorders does not exist., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Aims: To test the hypothesis that the activation of the growth hormone-releasing hormone (GHRH) receptor signalling pathway within the myocardium both prevents and reverses diastolic dysfunction and pathophysiologic features consistent with heart failure with preserved ejection fraction (HFpEF). Impaired myocardial relaxation, fibrosis, and ventricular stiffness, among other multi-organ morbidities, characterize the phenotype underlying the HFpEF syndrome. Despite the rapidly increasing prevalence of HFpEF, few effective therapies have emerged. Synthetic agonists of the GHRH receptors reduce myocardial fibrosis, cardiomyocyte hypertrophy, and improve performance in animal models of ischaemic cardiomyopathy, independently of the growth hormone axis., Methods and Results: CD1 mice received 4- or 8-week continuous infusion of angiotensin-II (Ang-II) to generate a phenotype with several features consistent with HFpEF. Mice were administered either vehicle or a potent synthetic agonist of GHRH, MR-356 for 4-weeks beginning concurrently or 4-weeks following the initiation of Ang-II infusion. Ang-II-treated animals exhibited diastolic dysfunction, ventricular hypertrophy, interstitial fibrosis, and normal ejection fraction. Cardiomyocytes isolated from these animals exhibited incomplete relaxation, depressed contractile responses, altered myofibrillar protein phosphorylation, and disturbed calcium handling mechanisms (ex vivo). MR-356 both prevented and reversed the development of the pathological phenotype in vivo and ex vivo. Activation of the GHRH receptors increased cAMP and cGMP in cardiomyocytes isolated from control animals but only cAMP in cardiac fibroblasts, suggesting that GHRH-A exert differential effects on cardiomyocytes and fibroblasts., Conclusion: These findings indicate that the GHRH receptor signalling pathway(s) represents a new molecular target to counteract dysfunctional cardiomyocyte relaxation by targeting myofilament phosphorylation and fibrosis. Accordingly, activation of GHRH receptors with potent, synthetic GHRH agonists may provide a novel therapeutic approach to management of the myocardial alterations associated with the HFpEF syndrome., Competing Interests: Conflict of interest: A.V.S. and J.M.H. are listed as co-inventors on patents on GHRH analogues which were assigned to the University of Miami and Veterans Affairs Department. J.M.H. previously owned equity in Biscayne Pharmaceuticals, license of intellectual property used in this study. Biscayne Pharmaceuticals did not provide funding for this study. J.M.H. reported having a patent for cardiac cell-based therapy. He holds equity in Vestion Inc. and maintains a professional relationship with Vestion Inc. as a consultant and member of the Board of Directors and Scientific Advisory Board. J.M.H. is the Chief Scientific Officer, a compensated consultant and advisory board member for Longeveron, and holds equity in Longeveron. He is also the co-inventor of intellectual property licensed to Longeveron. Longeveron LLC and Vestion Inc. did not participate in funding this work. J.M.H.’s relationships are disclosed to the University of Miami, and a management plan is in place. All other authors have declared that no conflict of interest exists., (© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Purpose: Growth hormone-releasing hormone (GHRH) is a 44-amino acid peptide that regulates growth hormone (GH) secretion. We hypothesized that GHRH receptor (GHRH-R) in alveolar type 2 (AT2) cells could modulate pro-inflammatory and possibly subsequent pro-fibrotic effects of lipopolysaccharide (LPS) or cytokines, such that AT2 cells could participate in lung inflammation and fibrosis. Methods: We used human alveolar type 2 (iAT2) epithelial cells derived from induced pluripotent stem cells (iPSC) to investigate how GHRH-R modulates gene and protein expression. We tested iAT2 cells' gene expression in response to LPS or cytokines, seeking whether these mechanisms caused endogenous production of pro-inflammatory molecules or mesenchymal markers. Quantitative real-time PCR (RT-PCR) and Western blotting were used to investigate differential expression of epithelial and mesenchymal markers. Result: Incubation of iAT2 cells with LPS increased expression of IL1-β and TNF-α in addition to mesenchymal genes, including ACTA2, FN1 and COL1A1. Alveolar epithelial cell gene expression due to LPS was significantly inhibited by GHRH-R peptide antagonist MIA-602. Incubation of iAT2 cells with cytokines like those in fibrotic lungs similarly increased expression of genes for IL1-β, TNF-α, TGFβ-1, Wnt5a, smooth muscle actin, fibronectin and collagen. Expression of mesenchymal proteins, such as N-cadherin and vimentin, were also elevated after prolonged exposure to cytokines, confirming epithelial production of pro-inflammatory molecules as an important mechanism that might lead to subsequent fibrosis. Conclusion: iAT2 cells clearly expressed the GHRH-R. Exposure to LPS or cytokines increased iAT2 cell production of pro-inflammatory factors. GHRH-R antagonist MIA-602 inhibited pro-inflammatory gene expression, implicating iAT2 cell GHRH-R signaling in lung inflammation and potentially in fibrosis.

The discovery of hypothalamic hormones propelled exciting advances in pharmacotherapy and improved life quality worldwide. Growth hormone-releasing hormone (GHRH) is a crucial element in homeostasis maintenance, and regulates the release of growth hormone from the anterior pituitary gland. Accumulating evidence suggests that this neuropeptide can also promote malignancies, as well as inflammation. Our review is focused on the role of that 44 - amino acid peptide (GHRH) and its antagonists in inflammation and vascular function, summarizing recent findings in the corresponding field. Preclinical studies demonstrate the protective role of GHRH antagonists against endothelial barrier dysfunction, suggesting that the development of those peptides may lead to new therapies against pathologies related to vascular remodeling (eg, sepsis, acute respiratory distress syndrome). Targeted therapies for those diseases do not exist., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Growth hormone (GH) secretion is controlled by short and long negative feedback loops. In this regard, both GH (short-loop feedback) and insulin-like growth factor 1 (IGF-1; long-loop feedback) can target somatotropic cells of the pituitary gland and neuroendocrine hypothalamic neurons to regulate the GH/IGF-1 axis. GH-releasing hormone (GHRH)-expressing neurons play a fundamental role in stimulating pituitary GH secretion. However, it is currently unknown whether IGF-1 action on GHRH-expressing cells is required for the control of the GH/IGF-1/growth axis. In the present study, we investigated the phenotype of male and female mice carrying ablation of IGF-1 receptor (IGF1R) exclusively in GHRH cells. After weaning, both male and female GHRHΔIGF1R mice exhibited increases in body weight, lean body mass, linear growth, and length of long bones (tibia, femur, humerus, and radius). In contrast, the percentage of body fat was similar between control and GHRHΔIGF1R mice. The higher body growth of GHRHΔIGF1R mice can be explained by increases in mean GH levels, GH pulse amplitude, and pulse frequency, calculated from 36 blood samples collected from each animal at 10-minute intervals. GHRHΔIGF1R mice also showed increased hypothalamic Ghrh mRNA levels, pituitary Gh mRNA expression, hepatic Igf1 expression, and serum IGF-1 levels compared with control animals. Furthermore, GHRHΔIGF1R mice displayed significant alterations in the sexually dimorphic hepatic gene expression profile, with a prevailing feminization in most genes analyzed. In conclusion, our findings indicate that GHRH neurons represent a key and necessary site for the long-loop negative feedback that controls the GH/IGF-1 axis and body growth., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Antagonists of growth hormone-releasing hormone (GHRH) inhibit the growth of various tumors, including endometrial carcinomas (EC). However, tumoral receptors that mediate the antiproliferative effects of GHRH antagonists in human ECs have not been fully characterized. In this study, we investigated the expression of mRNA for GHRH and splice variants (SVs) of GHRH receptors (GHRH-R) in 39 human ECs and in 7 normal endometrial tissue samples using RT-PCR. Primers designed for the PCR amplification of mRNA for the full length GHRH-R and SVs were utilized. The PCR products were sequenced, and their specificity was confirmed. Nine ECs cancers (23%) expressed mRNA for SV1, three (7.7%) showed SV2 and eight (20.5%) revealed mRNA for SV4. The presence of SVs for GHRH-Rs could not be detected in any of the normal endometrial tissue specimens. The presence of specific, high affinity GHRH-Rs was also demonstrated in EC specimens using radioligand binding studies. Twenty-four of the investigated thirty-nine tumor samples (61.5%) and three of the seven corresponding normal endometrial tissues (42.9%) expressed mRNA for GHRH ligand. Our findings suggest the possible existence of an autocrine loop in EC based on GHRH and its tumoral SV receptors. The antiproliferative effects of GHRH antagonists on EC are likely to be exerted in part by the local SVs and GHRH system.

Background: Growth Hormone Releasing Hormone (GHRH), 44 amino acids containing hypothalamic hormone, retains the biological activity by its first 29 amino acids. GHRH (NH2 1-29) peptide antagonists inhibit the growth of prostate, breast, ovarian, renal, gastric, pancreatic cancer in vitro and in vivo. Aptamers, single-strand RNA, or DNA oligonucleotides are capable of binding to target molecules with high affinity. Our aim in this study is to synthesize and select X-aptamers against both GHRH NH2 (1-29) and GHRH NH2 (1-44) and demonstrate synthesized aptamers' target binding activity as well as serum stability., Methods and Results: Aptamers against GHRH NH2 (1-44) and NH2 (1-29) peptides were synthesized, and binding affinity (Kd) of 24 putative X-aptamers was determined by the dot-blot method, co-immunofluorescence staining and, SPR analysis. The serum stability of TKY.T1.08, TKY1.T1.13, TKY.T2.08, TKY.T2.09 X-aptamers was 90-120 h, respectively. The dose-dependent binding of TKY1.T1.13, TKY.T2.08, TKY.T2.09 X-aptamers on GHRHR in MIA PaCa-2 was approved by co-IF assay results. Moreover, SPR analysis indicated the Kd (4.75, 1.21, and 4.0 nM) levels of TKY2.T1.13, TKY.T2.08, TKY.T2.09 putative X-aptamers, respectively., Conclusion: Our results illustrate the synthesis of 24 putative X-aptamers against both GHRH NH2 (1-44) and NH2 (1-29) peptides and TKY1.T1.13, TKY.T2.08, TKY.T2.09 X-aptamers have high serum stability, high target binding potential with low Kd levels., Competing Interests: The authors have declared that no competing interests exist.

Background/aims: To evaluate the effect and possible mechanism of growth hormone releasing hormone (GHRH) on chondrocytes of osteoarthritis (OA)., Methods: Articular chondrocytes were cultured and the expression of GHRH receptor in chondrocytes was detected. Then recombinant adenovirus GHRH (Ad-GHRH) was transfected to one group of chondrocytes. The expression of collagen type II, matrix metalloproteinase 13 (MMP-13) and signal transducer and activator of transcription 3 (STAT3) in each experimental group was determined by Western blot., Results: The GHRH receptor was expressed in chondrocytes, and this provided a basis for further study of the role of GHRH in chondrocytes. Cell proliferation of the Ad-GHRH group was significantly higher than that of the OA group by CCK-8 assay. Compared with the OA-group, the protein expression of MMP‑13 was decreased in the Ad-GHRH group. Compared with the OA-group, the protein expression of collagen type II, phosphorylated STAT3 (P-STAT3) were increased in the Ad-GHRH group., Conclusion: Our results show that the GHRH receptor is expressed in chondrocytes. GHRH can promote the proliferation of chondrocytes and the synthesis of type II collagen, and increase the extracellular matrix, which is achieved by phosphorylated STAT3 protein.

Accumulating evidence suggests that the influence on developmental traits might have long-term effects on aging and health later in life. Metformin is a widely used drug for treating type 2 diabetes and is also used for delaying sexual maturation in girls with precocious puberty. The current report focuses on investigating the effects of metformin on development and metabolic traits. Heterogeneous mice (UM-HET3) were treated with i.p. metformin between the ages of 15 and 56 days. Our results show that body weight and food consumption were increased in both sexes, and sexual maturation was delayed in females. Tail length and circulating insulin-like growth factor 1 (IGF1) levels were significantly increased in both sexes. No significant difference was found in insulin tolerance test, but glucose tolerance was significantly reduced in the males. Circulating adiponectin and insulin levels were altered by metformin treatment in a sex-specific manner. Analysis of quantitative insulin sensitivity check index (QUICKI) suggests that metformin treatment increased insulin sensitivity in female pups, but had opposite effect in male pups. This study revealed that early life metformin treatment alters development and metabolism of mice in both sex-specific and non-specific manners. These effects of metformin may have long-term impacts on aging-related traits., (Copyright © 2021 Elsevier B.V. All rights reserved.)

DON is commonly found in foods and feeds; it presents health risks, especially an increase of growth inhibition in humans, particularly infants and young children. However, there are relatively few research studies devoted to the mechanism of DON-mediated growth retardation. Interestingly, our results showed that DON does not cause any significant production of ROS but results in a persistent and significant release of NO with iNOS increasing activity, mitochondrial ultrastructural changes and decreasing ΔΨm. Moreover, the significant decreases in GH production and secretion induced by DON were dose-dependent, accompanied by an increase of caspase 3, 8 and 9, IL-11, IL-lβ and GHRH. NO scavenging agent (haemoglobin) and free radical scavenging agent (N-acetylcysteine) partially reversed mitochondrial damage, and Z-VAD-FMK increased the levels of GH and decreased the levels of caspase 3, 8 and 9, while haemoglobin decreased the levels of caspase 3, 8 and 9, indicating that NO is the primary target of DON-mediated inhibition. Present research study firstly demonstrated that NO is a key mediator of DON-induced growth inhibition and plays critical roles in the interference of GH transcription and synthesis. The current research is conducive to future research on the molecular mechanisms of DON-induced growth inhibition in humans, especially children., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Age-related diseases such as cardiovascular diseases portend disability, increase health expenditures, and cause late-life mortality. Synthetic agonists of growth hormone-releasing hormone (GHRH) exhibit several favorable effects on heart function and remodeling. Here we assessed whether GHRH agonist MR409 can modulate heart function and systemic parameters in old mice. Starting at the age of 15 months, mice were injected subcutaneously with MR409 (10 µg/day, n = 8) or vehicle (n = 7) daily for 6 months. Mice treated with MR409 showed improvements in exercise activity, cardiac function, survival rate, immune function, and hair growth in comparison with the controls. More stem cell colonies were grown out of the bone marrow recovered from the MR409-treated mice. Mitochondrial functions of cardiomyocytes (CMs) from the MR409-treated mice were also significantly improved with more mitochondrial fusion. Fewer β-gal positive cells were observed in endothelial cells after 10 passages with MR409. In Doxorubicin-treated H9C2 cardiomyocytes, cell senescence marker p21 and reactive oxygen species were significantly reduced after cultured with MR409. MR409 also improved cellular ATP production and oxygen consumption rate in Doxorubicin-treated H9C2 cells. Mitochondrial protein OPA1 long isoform was significantly increased after treatment with MR409. The effects of MR409 were mediated by GHRH receptor and protein kinase A (PKA). In short, GHRH agonist MR409 reversed the aging-associated changes with respect of heart function, mobility, hair growth, cellular energy production, and senescence biomarkers. The improvement of heart function may be related to a better mitochondrial functions through GHRH receptor/cAMP/PKA/OPA1 signaling pathway and relieved cardiac inflammation., (© 2021 Wiley Periodicals LLC.)

Ischemic stroke can induce neurogenesis. However, most stroke-generated newborn neurons cannot survive. It has been shown that MR-409, a potent synthetic agonistic analog of growth hormone-releasing hormone (GHRH), can protect against some life-threatening pathological conditions by promoting cell proliferation and survival. The present study shows that long-term treatment with MR-409 (5 or 10 μg/mouse/d) by subcutaneous (s.c.) injection significantly reduces the mortality, ischemic insult, and hippocampal atrophy, and improves neurological functional recovery in mice operated on for transient middle cerebral artery occlusion (tMCAO). Besides, MR-409 can stimulate endogenous neurogenesis and improve the tMCAO-induced loss of neuroplasticity. MR-409 also enhances the proliferation and inhibits apoptosis of neural stem cells treated with oxygen and glucose deprivation-reperfusion. The neuroprotective effects of MR-409 are closely related to the activation of AKT/CREB and BDNF/TrkB pathways. In conclusion, the present study demonstrates that GHRH agonist MR-409 has remarkable neuroprotective effects through enhancing endogenous neurogenesis in cerebral ischemic mice.

Growth hormone-releasing hormone (GHRH) is a hypothalamic hormone, which regulates the secretion of growth hormone (GH) from the anterior pituitary gland. The effects of GHRH extend beyond the GH-insulin-like growth factor I axis, and that neuropeptide has been involved in the potentiation of several malignancies and other inflammatory disorders. The development of GHRH antagonists (GHRHAnt) delivers an exciting possibility to counteract the pathogenesis of the GHRH-related effects in human pathophysiology, especially when considered that GHRHAnt support endothelial barrier integrity. Those GHRHAnt-mediated effects are exerted at least in part due to the suppression of major inflammatory pathways, and the modulation of major cytoskeletal components. In the present study, we measured the production of reactive oxygen species (ROS) in bovine pulmonary artery endothelial cells, human cerebral microvascular endothelial cells, and human lung microvascular endothelial cells exposed to GHRH or a commercially available GHRHAnt. Our findings reveal the antioxidative effects of GHRHAnt in all three cell lines, which express GHRH receptors. The redox status of NIH/3T3 cells, which do not produce GHRH receptors, was not significantly affected by GHRH or GHRHAnt. Hence, the application of GHRHAnt in pathologies related to increased ROS production should be further investigated., (© 2021 Wiley Periodicals LLC.)

The hormone secretion of GHRH-GH-IGF-1 axis in animals was decreased as aging. These hormones play an important role in maintaining bone mass and bone structure, and also affect the normal structure and function of the skin. We used plasmid-based technology to deliver growth hormone releasing hormone (GHRH) to elderly mice. In the current study, 80 and 120 μg/kg pVAX-GHRH plasmid expression plasmid were injected into old mice, the serum GHRH and insulin-like growth factor-1(IGF-1) content were increased within three weeks (P < 0.05). In the groups of 80 and 120 μg/kg plasmid, the content of procollagen type I N-terminal pro-peptide (PINP) in the serum was increased(P < 0.05), and the content of C-terminal telopeptides of type I collagen (CTX-1) in the serum was reduced significantly (P < 0.05). Furthermore, the expression of osteoprotegerin (OPG) and osteocalcin (OCN) in the femur also was increased(P < 0.05). The bone mineral density(BMD)、trabecular bone volume (BV/TV) and trabecular number(Tb.N) of mouse femur were increased significantly (P < 0.05) and trabecular separation(Tb.Sp) was decreased(P < 0.05). There were more trabecular bones in the bone marrow cavity and the trabecular bones are thicker in the groups of 80 and 120 μg/kg plasmid relative to control. The superoxide dismutase (SOD) content in the skin was increased(P < 0.05), and the malondialdehyde (MDA) content was reduced significantly (P < 0.05). Meanwhile, the skin moisture content also increased significantly(P < 0.05). Moreover, the expression of matrix metalloproteinase 3(MMP3) and matrix metalloproteinase 9(MMP9) was decreased in the skin(P < 0.05). The thickness of the dermis and epidermis of the skin had increased significantly(P < 0.05). Skin structure is more dense and complete in the two groups. These results indicate that 80 and 120 μg/kg plasmid-mediated GHRH supplementation can improve osteoporosis and skin aging in aged mice., (Copyright © 2021. Published by Elsevier Ltd.)

The endoplasmic reticulum (ER) stress is one of the mechanisms related to decreased insulin secretion and beta cell death, contributing to the progress of type 2 diabetes mellitus (T2D). Thus, investigating agents that can influence this process would help prevent the development of T2D. Recently, the growth-hormone-releasing hormone (GHRH) action has been demonstrated in INS-1E cells, in which it increases cell proliferation and insulin secretion. As the effects of GHRH and its agonists have not been fully elucidated in the beta cell, we proposed to investigate them by evaluating the role of the GHRH agonist, MR-409, in cells under ER stress. Our results show that the agonist was unable to ameliorate or prevent ER stress. However, cells exposed to the agonist showed less oxidative stress and greater survival even under ER stress. The mechanisms by which GHRH agonist, MR-409, leads to these outcomes require further investigation., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Insulin receptor substrate-1 (Irs1) is one of the major substrates for insulin receptor and insulin-like growth factor-1 (IGF-1) receptor tyrosine kinases. Systemic Irs1-deficient mice show growth retardation, with resistance to insulin and IGF-1, although the underlying mechanisms remain poorly understood. For this study, we generated mice with brain-specific deletion of Irs1 (NIrs1KO mice). The NIrs1KO mice exhibited lower body weights, shorter bodies and bone lengths, and decreased bone density. Moreover, the NIrs1KO mice exhibited increased insulin sensitivity and glucose utilization in the skeletal muscle. Although the ability of the pituitary to secrete growth hormone (GH) remained intact, the amount of hypothalamic growth hormone-releasing hormone (GHRH) was significantly decreased and, accordingly, the pituitary GH mRNA expression levels were impaired in these mice. Plasma GH and IGF-1 levels were also lower in the NIrs1KO mice. The expression levels of GHRH protein in the median eminence, where Irs1 antibody staining is observed, were markedly decreased in the NIrs1KO mice. In vitro, neurite elongation after IGF-1 stimulation was significantly impaired by Irs1 downregulation in the cultured N-38 hypothalamic neurons. In conclusion, brain Irs1 plays important roles in the regulation of neurite outgrowth of GHRH neurons, somatic growth, and glucose homeostasis., (© 2021 by the American Diabetes Association.)

Purpose: Early institution of GH therapy in children with Prader-Willi syndrome (PWS) yields beneficial effects on their phenotype and is associated with a persistent improvement of body composition, both in the transition age and in adulthood. Reports from GH stimulation testing in PWS adults, however, suggest that GH deficiency (GHD) is not a universal feature of the syndrome, and the current Consensus Guidelines suggest to perform a reassessment of persistent GHD so as to continue GH therapy after reaching adult height. Few data about GH responsiveness to stimulation testing throughout the transitional period in PWS are available to date. Thus, we investigated the prevalence of GHD in a large cohort of patients with PWS during the transition phase., Patients and Methods: One hundred forty-one PWS patients, 72 females and 69 males, aged 15.4-24.9 years, were evaluated by dynamic testing with growth hormone-releasing hormone (GHRH) plus arginine (GHRH + ARG). To define GHD, both BMI-dependent and BMI-independent diagnostic cut-off limits were considered., Results: According to BMI-dependent criteria, 10.7% of normal weight (NW), 18.5% of overweight and 22.1% of obese PWS maintained a status of GHD. Similar results were obtained by adopting a cut-off limit specific for the adult age (26.2%), as well as criteria for the transition phase in NW subjects (25%)., Conclusion: Our study shows that about 20% of patients with PWS fulfilled the criteria for GHD during the transitional age, suggesting the need of an integrated analysis of GH/IGF-I axis, in the context of the general clinical picture and other endocrine abnormalities, in all subjects after attainment of final stature.

In healthy conditions, prepubertal growth follows an individual specific growth channel. Growth hormone (GH) is undoubtedly the major regulator of growth. However, the homeostatic regulation to maintain the individual specific growth channel during growth is unclear. We recently hypothesized a body weight sensing homeostatic regulation of body weight during adulthood, the gravitostat. We now investigated if sensing of body weight also contributes to the strict homeostatic regulation to maintain the individual specific growth channel during prepubertal growth. To evaluate the effect of increased artificial loading on prepubertal growth, we implanted heavy (20% of body weight) or light (2% of the body weight) capsules into the abdomen of 26-day-old male rats. The body growth, as determined by change in biological body weight and growth of the long bones and the axial skeleton, was reduced in rats bearing a heavy load compared with light load. Removal of the increased load resulted in a catch-up growth and a normalization of body weight. Loading decreased hypothalamic growth hormone releasing hormone mRNA, liver insulin-like growth factor (IGF)-1 mRNA, and serum IGF-1, suggesting that the reduced body growth was caused by a negative feedback regulation on the somatotropic axis and this notion was supported by the fact that increased loading did not reduce body growth in GH-treated rats. Based on these data, we propose the gravitostat hypothesis for the regulation of prepubertal growth. This states that there is a homeostatic regulation to maintain the individual specific growth channel via body weight sensing, regulating the somatotropic axis and explaining catch-up growth., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society.)

Mice with disruptions of growth hormone-releasing hormone (GHRH) or growth hormone receptor (GHR) exhibit similar phenotypes of prolonged lifespan and delayed age-related diseases. However, these two models respond differently to calorie restriction indicating that they might carry different and/or independent mechanisms for improved longevity and healthspan. In order to elucidate these mechanisms, we generated GHRH and GHR double-knockout mice (D-KO). In the present study, we focused specifically on the characteristics of female D-KO mice. The D-KO mice have reduced body weight and enhanced insulin sensitivity compared to wild-type (WT) controls. Growth retardation in D-KO mice is accompanied by decreased GH expression in pituitary, decreased circulating IGF-1, increased high-molecular-weight (HMW) adiponectin, and leptin hormones compared to WT controls. Generalized linear model-based regression analysis, which controls for body weight differences between D-KO and WT groups, shows that D-KO mice have decreased lean mass, bone mineral density, and bone mineral content, but increased adiposity. Indirect calorimetry markers including oxygen consumption, carbon dioxide production, and energy expenditure were significantly lower in D-KO mice relative to the controls. In comparison with WT mice, the D-KO mice displayed reduced respiratory exchange ratio (RER) values only during the light cycle, suggesting a circadian-related metabolic shift toward fat utilization. Interestingly, to date survival data suggest extended lifespan in D-KO female mice., (© 2021 The Authors. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

The classic concept of how pituitary GH is regulated by somatostatin and GHRH has changed in recent years, following the discovery of peripheral hormones involved in the regulation of energy homeostasis and mineral homeostasis. These hormones are ghrelin, nesfatins, and klotho. Ghrelin is an orexigenic hormone, released primarily by the gastric mucosa, although it is widely expressed in many different tissues, including the central nervous system and the pituitary. To be active, ghrelin must bind to an n-octanoyl group (n = 8, generally) on serine 3, forming acyl ghrelin which can then bind and activate a G-protein-coupled receptor leading to phospholipase C activation that induces the formation of inositol 1,4,5-triphosphate and diacylglycerol that produce an increase in cytosolic calcium that allows the release of GH. In addition to its direct action on somatotrophs, ghrelin co-localizes with GHRH in several neurons, facilitating its release by inhibiting somatostatin, and acts synergistically with GHRH stimulating the synthesis and secretion of pituitary GH. Gastric ghrelin production declines with age, as does GH. Klotho is an anti-aging agent, produced mainly in the kidneys, whose soluble circulating form directly induces GH secretion through the activation of ERK1/2 and inhibits the inhibitory effect that IGF-I exerts on GH. Children and adults with untreated GH-deficiency show reduced plasma levels of klotho, but treatment with GH restores them to normal values. Deletions or mutations of the Klotho gene affect GH production. Nesfatins 1 and 2 are satiety hormones, they inhibit food intake. They have been found in GH3 cell cultures where they significantly reduce the expression of gh mRNA and that of pituitary-specific positive transcription factor 1, consequently acting as inhibitors of GH production. This is a consequence of the down-regulation of the cAMP/PKA/CREB signaling pathway. Interestingly, nesfatins eliminate the strong positive effect that ghrelin has on GH synthesis and secretion. Throughout this review, we will attempt to broadly analyze the role of these hormones in the complex world of GH regulation, a world in which these hormones already play a very important role., Competing Interests: The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Devesa.)

Therapies for heart failure with preserved ejection fraction (HFpEF) are lacking. Growth hormone-releasing hormone agonists (GHRH-As) have salutary effects in ischemic and nonischemic heart failure animal models. Accordingly, we hypothesized that GHRH-A treatment ameliorates chronic kidney disease (CKD)-induced HFpEF in a large-animal model. Female Yorkshire pigs ( n = 16) underwent 5/6 nephrectomy via renal artery embolization and 12 wk later were randomized to receive daily subcutaneous injections of GHRH-A (MR-409; n = 8; 30 µg/kg) or placebo ( n = 8) for 4 to 6 wk. Renal and cardiac structure and function were serially assessed postembolization. Animals with 5/6 nephrectomy exhibited CKD (elevated blood urea nitrogen [BUN] and creatinine) and faithfully recapitulated the hemodynamic features of HFpEF. HFpEF was demonstrated at 12 wk by maintenance of ejection fraction associated with increased left ventricular mass, relative wall thickness, end-diastolic pressure (EDP), end-diastolic pressure/end-diastolic volume (EDP/EDV) ratio, and tau, the time constant of isovolumic diastolic relaxation. After 4 to 6 wk of treatment, the GHRH-A group exhibited normalization of EDP ( P = 0.03), reduced EDP/EDV ratio ( P = 0.018), and a reduction in myocardial pro-brain natriuretic peptide protein abundance. GHRH-A increased cardiomyocyte [Ca 2+ ] transient amplitude ( P = 0.009). Improvement of the diastolic function was also evidenced by increased abundance of titin isoforms and their ratio ( P = 0.0022). GHRH-A exerted a beneficial effect on diastolic function in a CKD large-animal model as demonstrated by improving hemodynamic, structural, and molecular characteristics of HFpEF. These findings have important therapeutic implications for the HFpEF syndrome., Competing Interests: Competing interest statement: A.V.S and J.M.H. are listed as co-inventors on patents on GHRH analogs, which were assigned to the University of Miami and Veterans Affairs Department. J.M.H. previously owned equity in Biscayne Pharmaceuticals, licensee of intellectual property used in this study. Biscayne Pharmaceuticals did not provide funding for this study. J.M.H. is the chief scientific officer, a compensated consultant, and advisory board member for Longeveron and holds equity in Longeveron. J.M.H. is also the co-inventor of intellectual property licensed to Longeveron. Longeveron did not play a role in the design, conduct, or funding of the study. J.M.H.’s relationships are reported to the University of Miami, and an appropriate management plan is in place. I.H.S. contributed to this manuscript in her personal capacity. The opinions expressed in this article are the author's own and do not reflect the view of the National Institutes of Health, the Department of Health and Human Services, or the United States government.

Obese individuals often show low growth hormone (GH) secretion, which leads to reduced lipid mobilization and further fat accumulation. Pharmacological approaches to increase GH levels in obese individuals by GH injection or GH-releasing hormone receptor agonist showed promising effects on fat reduction. However, side effects on glucose metabolism and the heavy costs on making large peptides hindered their clinical application. Here, we tested whether stimulation of endogenous GH secretion by a synthetic GH secretagogue receptor (GHSR) agonist, hexarelin, improved the metabolism in a hyperphagic obese mouse model. Male melanocortin 4 receptor knockout mice (MC4RKO) were pair-fed and received continuous hexarelin (10.56 μg/day) or vehicle infusion by an osmotic pump for 3-4 weeks. Hexarelin treatment significantly increased the pulsatile GH secretion without detectable alteration on basal GH secretion in MC4RKO mice. The treated mice showed increased lipolysis and lipid oxidation in the adipose tissue, and reduced de novo lipogenesis in the liver, leading to reduced visceral fat mass, reduced triglyceride content in liver, and unchanged circulating free fatty acid levels. Importantly, hexarelin treatment improved the whole-body insulin sensitivity but did not alter glucose tolerance, insulin levels, or insulin-like growth factor 1 (IGF-1) levels. The metabolic effects of hexarelin were likely through the direct action of GH, as indicated by the increased expression level of genes involved in GH signaling pathways in visceral adipose tissues and liver. In conclusion, hexarelin treatment stimulated the pulsatile GH secretion and reduced the fat accumulation in visceral depots and liver in obese MC4RKO mice with improved insulin sensitivity without altered levels of insulin or IGF-1. It provides evidence for managing obesity by enhancing pulsatile GH secretion through activation of GHSR in the pituitary gland., (© 2020 Federation of American Societies for Experimental Biology.)

To clarify the role of growth hormone (GH) in the immune system of fish, we examine the comparative effect of GH and Growth Hormone Release Factor (GRF) on leukocytes culture of the head kidney of Atlantic salmon and the SHK-1 cell line. There are studies that associate the growth hormone (GH) / insulin-like growth factor (IGF) axis with the immune regulation of fish. However, there is no evidence that GH and GRF stimulate Atlantic salmon leukocyte cell lines, where there areńt reports on expression changes in different immune response markers. Thus, we investigated the effect of GH and GRF in Atlantic salmon leukocytes extracted from head kidney and the SHK-1 cell line on the different immune response markers such as: NLRC5, NLRC3, IL-1β, TNF-α, and IL-8 through qPCR. Our data suggest that GH increases the expression of NLRC5, NLRC3, and IL-1β mainly at 16 h post-stimulation in Atlantic salmon leukocytes. This indicates differential regulation between the two models used, helping us to better understand the independent action of GH on the immune system and the GH / IGF axis for future research., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Anterior pituitary somatotropes are important metabolic sensors responding to leptin by secreting growth hormone (GH). However, reduced leptin signals caused by fasting have not always correlated with reduced serum GH. Reports show that fasting may stimulate or reduce GH secretion, depending on the species. Mechanisms underlying these distinct somatotrope responses to fasting remain unknown. To define the somatotrope response to decreased leptin signaling we examined markers of somatotrope function over different time periods of fasting. Male mice were fasted for 24 and 48 h, with female mice fasted for 24 h compared to fed controls ad libitum. Body weight and serum glucose were reduced in both males and females, but, unexpectedly, serum leptin was reduced only in males. Furthermore, in males, serum GH levels showed a biphasic response with significant reductions at 24 h followed by a significant rise at 48 h, which coincided with the rise in serum ghrelin levels. In contrast, females showed an increase in serum GH at 24 h. We then explored mechanisms underlying the differential somatotrope responses seen in males and observed that pituitary levels of Gh mRNA increased, with no distinction between acute and prolonged fasting. By contrast, the Ghrhr mRNA (encoding GH releasing hormone receptor) and the Ghsr mRNA (encoding the ghrelin receptor) were both greatly increased at prolonged fasting times coincident with increased serum GH. These findings show sex differences in the somatotrope and adipocyte responses to fasting and support an adaptive role for somatotropes in males in response to multiple metabolic signals.

Hypoglycemia is a frequent complication of diabetes, limiting therapy and increasing morbidity and mortality. With recurrent hypoglycemia, the counterregulatory response (CRR) to decreased blood glucose is blunted, resulting in hypoglycemia-associated autonomic failure (HAAF). The mechanisms leading to these blunted effects are only poorly understood. Here, we report, with ISH, IHC, and the tissue-clearing capability of iDISCO+, that growth hormone releasing hormone (GHRH) neurons represent a unique population of arcuate nucleus neurons activated by glucose deprivation in vivo. Repeated glucose deprivation reduces GHRH neuron activation and remodels excitatory and inhibitory inputs to GHRH neurons. We show that low glucose sensing is coupled to GHRH neuron depolarization, decreased ATP production, and mitochondrial fusion. Repeated hypoglycemia attenuates these responses during low glucose. By maintaining mitochondrial length with the small molecule mitochondrial division inhibitor-1, we preserved hypoglycemia sensitivity in vitro and in vivo. Our findings present possible mechanisms for the blunting of the CRR, significantly broaden our understanding of the structure of GHRH neurons, and reveal that mitochondrial dynamics play an important role in HAAF. We conclude that interventions targeting mitochondrial fission in GHRH neurons may offer a new pathway to prevent HAAF in patients with diabetes.

In normal individuals, pituitary somatotrophs optimise body composition by responding to metabolic signals from leptin. To identify mechanisms behind the regulation of somatotrophs by leptin, we used Cre-LoxP technology to delete leptin receptors (LEPR) selectively in somatotrophs and developed populations purified by fluorescence-activated cell sorting (FACS) that contained 99% somatotrophs. FACS-purified, Lepr-null somatotrophs showed reduced levels of growth hormone (GH), growth hormone-releasing hormone receptor (GHRHR), and Pou1f1 proteins and Gh (females) and Ghrhr (both sexes) mRNAs. Pure somatotrophs also expressed thyroid-stimulating hormone (TSH) and prolactin (PRL), both of which were reduced in pure somatotrophs lacking LEPR. This introduced five gene products that were targets of leptin. In the present study, we tested the hypothesis that leptin is both a transcriptional and a post-transcriptional regulator of these gene products. Our tests showed that Pou1f1 and/or the Janus kinase/signal transducer and activator of transcription 3 transcriptional regulatory pathways are implicated in the leptin regulation of Gh or Ghrhr mRNAs. We then focused on potential actions by candidate microRNAs (miRNAs) with consensus binding sites on the 3' UTR of Gh or Ghrhr mRNAs. Somatotroph Lepr-null deletion mutants expressed elevated levels of miRNAs including miR1197-3p (in females), miR103-3p and miR590-3p (both sexes), which bind Gh mRNA, or miRNA-325-3p (elevated in both sexes), which binds Ghrhr mRNA. This elevation indicates repression of translation in the absence of LEPR. In addition, after detecting binding sites for Musashi on Tshb and Prl 3' UTR, we determined that Musashi1 repressed translation of both mRNAs in in vitro fluc assays and that Prl mRNA was enriched in Musashi immunoprecipitation assays. Finally, we tested ghrelin actions to determine whether its nitric oxide-mediated signalling pathways would restore somatotroph functions in deletion mutants. Ghrelin did not restore either GHRH binding or GH secretion in vitro. These studies show an unexpectedly broad role for leptin with respect to maintaining somatotroph functions, including the regulation of PRL and TSH in subsets of somatotrophs that may be progenitor cells., (© 2020 British Society for Neuroendocrinology.)

Börjeson-Forssman-Lehmann syndrome (BFLS) is an intellectual disability and endocrine disorder caused by plant homeodomain finger 6 ( PHF6 ) mutations. Individuals with BFLS present with short stature. We report a mouse model of BFLS, in which deletion of Phf6 causes a proportional reduction in body size compared with control mice. Growth hormone (GH) levels were reduced in the absence of PHF6. Phf6 - /Y animals displayed a reduction in the expression of the genes encoding GH-releasing hormone (GHRH) in the brain, GH in the pituitary gland and insulin-like growth factor 1 (IGF1) in the liver. Phf6 deletion specifically in the nervous system caused a proportional growth defect, indicating a neuroendocrine contribution to the phenotype. Loss of suppressor of cytokine signaling 2 (SOCS2), a negative regulator of growth hormone signaling partially rescued body size, supporting a reversible deficiency in GH signaling. These results demonstrate that PHF6 regulates the GHRH/GH/IGF1 axis., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2020. Published by The Company of Biologists Ltd.)

Growth hormone-releasing hormone (GHRH) is secreted primarily from the hypothalamus, but other tissues, including the lungs, produce it locally. GHRH stimulates the release and secretion of growth hormone (GH) by the pituitary and regulates the production of GH and hepatic insulin-like growth factor-1 (IGF-1). Pituitary-type GHRH-receptors (GHRH-R) are expressed in human lungs, indicating that GHRH or GH could participate in lung development, growth, and repair. GHRH-R antagonists (i.e., synthetic peptides), which we have tested in various models, exert growth-inhibitory effects in lung cancer cells in vitro and in vivo in addition to having anti-inflammatory, anti-oxidative, and pro-apoptotic effects. One antagonist of the GHRH-R used in recent studies reviewed here, MIA-602, lessens both inflammation and fibrosis in a mouse model of bleomycin lung injury. GHRH and its peptide agonists regulate the proliferation of fibroblasts through the modulation of extracellular signal-regulated kinase (ERK) and Akt pathways. In addition to downregulating GH and IGF-1, GHRH-R antagonist MIA-602 inhibits signaling pathways relevant to inflammation, including p21-activated kinase 1-signal transducer and activator of transcription 3/nuclear factor-kappa B (PAK1-STAT3/NF-κB and ERK). MIA-602 induces fibroblast apoptosis in a dose-dependent manner, which is an effect that is likely important in antifibrotic actions. Taken together, the novel data reviewed here show that GHRH is an important peptide that participates in lung homeostasis, inflammation, wound healing, and cancer; and GHRH-R antagonists may have therapeutic potential in lung diseases.