Autosomal Dominant Growth Hormone Deficiency (Type II).

Isolated growth hormone deficiency (IGHD) is the commonest pituitary hormone deficiency resulting from congenital or acquired causes, although for most patients its etiology remains unknown. Among the known factors, heterozygous mutations in the growth hormone gene (GH1) lead to the autosomal dominant form of GHD, also known as type II GHD. In many cohorts this is the commonest form of congenital isolated GHD and is mainly caused by mutations that affect the correct splicing of GH-1. These mutations cause skipping of the third exon and lead to the production of a 17.5-kDa GH isoform that exerts a dominant negative effect on the secretion of the wild type GH. The identification of these mutations has clinical implications for the management of patients, as there is a well-documented correlation between the severity of the phenotype and the increased expression of the 17.5-kDa isoform. Patients with type II GHD have a variable height deficit and severity of GHD and may develop additional pituitary hormone defiencies over time, including ACTH, TSH and gonadotropin deficiencies. Therefore, their lifelong follow-up is recommended. Detailed studies on the effect of heterozygous GH1 mutations on the trafficking, secretion and action of growth hormone can elucidate their mechanism on a cellular level and may influence future treatment options for GHD type II.

SOX2 haploinsufficiency is associated with slow progressing hypothalamo-pituitary tumours.

SOX2 is an early developmental transcription factor and marker of stem cells that has recently been implicated in the development of the pituitary gland. Heterozygous SOX2 mutations have been described in patients with hypopituitarism and severe ocular abnormalities. In the majority of published cases, the pituitary gland is either small or normal in size. Here, we report two unrelated patients with SOX2 haploinsufficiency (a heterozygous gene deletion and a novel c.143TC>AA/p.F48X mutation) who developed nonprogressive pituitary tumors of early onset, suggesting a congenital etiology. The truncating mutation resulted in significant loss of function and impaired nuclear localization of the mutant protein, in addition to a failure to repress ß-catenin transcriptional activity in vitro. This is the first indication that SOX2 haploinsufficiency is implicated in the generation of pituitary tumors with distinct clinical characteristics, possibly mediated via its effects on the Wnt signaling pathway.

Development of the Pituitary Gland.

The development of the anterior pituitary gland occurs in distinct sequential developmental steps, leading to the formation of a complex organ containing five different cell types secreting six different hormones. During this process, the temporal and spatial expression of a cascade of signaling molecules and transcription factors plays a crucial role in organ commitment, cell proliferation, patterning, and terminal differentiation. The morphogenesis of the gland and the emergence of distinct cell types from a common primordium are governed by complex regulatory networks involving transcription factors and signaling molecules that may be either intrinsic to the developing pituitary or extrinsic, originating from the ventral diencephalon, the oral ectoderm, and the surrounding mesenchyme. Endocrine cells of the pituitary gland are organized into structural and functional networks that contribute to the coordinated response of endocrine cells to stimuli; these cellular networks are formed during embryonic development and are maintained or may be modified in adulthood, contributing to the plasticity of the gland. Abnormalities in any of the steps of pituitary development may lead to congenital hypopituitarism that includes a spectrum of disorders from isolated to combined hormone deficiencies including syndromic disorders such as septo-optic dysplasia. Over the past decade, the acceleration of next-generation sequencing has allowed for rapid analysis of the patient genome to identify novel mutations and novel candidate genes associated with hypothalmo-pituitary development. Subsequent functional analysis using patient fibroblast cells, and the generation of stem cells derived from patient cells, is fast replacing the need for animal models while providing a more physiologically relevant characterization of novel mutations. Furthermore, CRISPR-Cas9 as the method for gene editing is replacing previous laborious and time-consuming gene editing methods that were commonly used, thus yielding knockout cell lines in a fraction of the time. © 2020 American Physiological Society. Compr Physiol 10:389-413, 2020.

Recent advances in central congenital hypothyroidism.

Central congenital hypothyroidism (CCH) may occur in isolation, or more frequently in combination with additional pituitary hormone deficits with or without associated extrapituitary abnormalities. Although uncommon, it may be more prevalent than previously thought, affecting up to 1:16 000 neonates in the Netherlands. Since TSH is not elevated, CCH will evade diagnosis in primary, TSH-based, CH screening programs and delayed detection may result in neurodevelopmental delay due to untreated neonatal hypothyroidism. Alternatively, coexisting growth hormones or ACTH deficiency may pose additional risks, such as life threatening hypoglycaemia. Genetic ascertainment is possible in a minority of cases and reveals mutations in genes controlling the TSH biosynthetic pathway (TSHB, TRHR, IGSF1) in isolated TSH deficiency, or early (HESX1, LHX3, LHX4, SOX3, OTX2) or late (PROP1, POU1F1) pituitary transcription factors in combined hormone deficits. Since TSH cannot be used as an indicator of euthyroidism, adequacy of treatment can be difficult to monitor due to a paucity of alternative biomarkers. This review will summarize the normal physiology of pituitary development and the hypothalamic-pituitary-thyroid axis, then describe known genetic causes of isolated central hypothyroidism and combined pituitary hormone deficits associated with TSH deficiency. Difficulties in diagnosis and management of these conditions will then be discussed.

Novel application of luciferase assay for the in vitro functional assessment of KAL1 variants in three females with septo-optic dysplasia (SOD).

KAL1 is implicated in 5% of Kallmann syndrome cases, a disorder which genotypically overlaps with septo-optic dysplasia (SOD). To date, a reporter-based assay to assess the functional consequences of KAL1 mutations is lacking. We aimed to develop a luciferase assay for novel application to functional assessment of rare KAL1 mutations detected in a screen of 422 patients with SOD. Quantitative analysis was performed using L6-myoblasts stably expressing FGFR1, transfected with a luciferase-reporter vector containing elements of the FGF-responsive osteocalcin promoter. The two variants assayed [p.K185N, p.P291T], were detected in three females with SOD (presenting with optic nerve hypoplasia, midline and pituitary defects). Our novel assay revealed significant decreases in transcriptional activity [p.K185N: 21% (p < 0.01); p.P291T: 40% (p < 0.001)]. Our luciferase-reporter assay, developed for assessment of KAL1 mutations, determined that two variants in females with hypopituitarism/SOD are loss-of-function; demonstrating that this assay is suitable for quantitative assessment of mutations in this gene.

Isolated growth hormone deficiency (GHD) in childhood and adolescence: recent advances.

The diagnosis of GH deficiency (GHD) in childhood is a multistep process involving clinical history, examination with detailed auxology, biochemical testing, and pituitary imaging, with an increasing contribution from genetics in patients with congenital GHD. Our increasing understanding of the factors involved in the development of somatotropes and the dynamic function of the somatotrope network may explain, at least in part, the development and progression of childhood GHD in different age groups. With respect to the genetic etiology of isolated GHD (IGHD), mutations in known genes such as those encoding GH (GH1), GHRH receptor (GHRHR), or transcription factors involved in pituitary development, are identified in a relatively small percentage of patients suggesting the involvement of other, yet unidentified, factors. Genome-wide association studies point toward an increasing number of genes involved in the control of growth, but their role in the etiology of IGHD remains unknown. Despite the many years of research in the area of GHD, there are still controversies on the etiology, diagnosis, and management of IGHD in children. Recent data suggest that childhood IGHD may have a wider impact on the health and neurodevelopment of children, but it is yet unknown to what extent treatment with recombinant human GH can reverse this effect. Finally, the safety of recombinant human GH is currently the subject of much debate and research, and it is clear that long-term controlled studies are needed to clarify the consequences of childhood IGHD and the long-term safety of its treatment.

Variations in PROKR2, but not PROK2, are associated with hypopituitarism and septo-optic dysplasia.

CONTEXT: Loss-of-function mutations in PROK2 and PROKR2 have been implicated in Kallmann syndrome (KS), characterized by hypogonadotropic hypogonadism and anosmia. Recent data suggest overlapping phenotypes/genotypes between KS and congenital hypopituitarism (CH), including septo-optic dysplasia (SOD). OBJECTIVE: We screened a cohort of patients with complex forms of CH (n = 422) for mutations in PROK2 and PROKR2. RESULTS: We detected 5 PROKR2 variants in 11 patients with SOD/CH: novel p.G371R and previously reported p.A51T, p.R85L, p.L173R, and p.R268C-the latter 3 being known functionally deleterious variants. Surprisingly, 1 patient with SOD was heterozygous for the p.L173R variant, whereas his phenotypically unaffected mother was homozygous for the variant. We sought to clarify the role of PROKR2 in hypothalamopituitary development through analysis of Prokr2(-/-) mice. Interestingly, these revealed predominantly normal hypothalamopituitary development and terminal cell differentiation, with the exception of reduced LH; this was inconsistent with patient phenotypes and more analogous to the healthy mother, although she did not have KS, unlike the Prokr2(-/-) mice. CONCLUSIONS: The role of PROKR2 in the etiology of CH, SOD, and KS is uncertain, as demonstrated by no clear phenotype-genotype correlation; loss-of-function variants in heterozygosity or homozygosity can be associated with these disorders. However, we report a phenotypically normal parent, homozygous for p.L173R. Our data suggest that the variants identified herein are unlikely to be implicated in isolation in these disorders; other genetic or environmental modifiers may also impact on the etiology. Given the phenotypic variability, genetic counseling may presently be inappropriate.

Phenotype-genotype correlations in congenital isolated growth hormone deficiency (IGHD).

Isolated growth hormone deficiency (IGHD) may be congenital, often due to genetic mutations, or acquired as a result of other factors such as cranial irradiation. The commonest genes implicated in its genetic etiology are those encoding growth hormone (GH1) and the receptor for GH-releasing hormone (GHRHR). Rarely, IGHD may be caused by mutations in transcription factors (HESX1, SOX3, OTX2) or be the first presentation before the development of other pituitary hormone deficiencies. IGHD has been classified in four genetic forms (type IA, IB, II and III). Despite the increasing number of genes implicated in the etiology of IGHD, mutations in known genes account only for a small percentage of cases; therefore, other as yet unidentified factors may be implicated in its etiology. Although there is no strict genotype/phenotype correlation in patients with IGHD, there are some emerging patterns that may guide us towards a genetic diagnosis of the condition. There is increasing understanding that the phenotype of patients with IGHD is highly variable and sometimes even evolving, dictating the need for long term follow-up in these cases.

Septo-optic dysplasia and other midline defects: the role of transcription factors: HESX1 and beyond.

Septo-optic dysplasia (SOD) is a highly heterogeneous condition comprising variable phenotypes including midline and forebrain abnormalities, optic nerve and pituitary hypoplasia. Most instances of SOD are sporadic and several aetiologies including drug and alcohol abuse have been suggested to account for the pathogenesis of the condition. However, a number of familial cases have been described with an increasing number of mutations in developmental transcription factors including HESX1, SOX2, SOX3 and OTX2 being implicated in its aetiology. These factors are essential for normal forebrain/pituitary development, and disruptions to these genes could account for the features observed in SOD and other midline disorders. The variable phenotypes observed within the condition are most likely due to the varying contributions of genetic and environmental factors. This review will discuss the current knowledge about SOD. Further study of these and other novel factors may shed light on the complex aetiology of this condition.

Increased transactivation associated with SOX3 polyalanine tract deletion in a patient with hypopituitarism.

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

Absence of GH-releasing hormone (GHRH) mutations in selected patients with isolated GH deficiency.

CONTEXT: Although numerous reports of mutations in GH1 and GHRHR (GHRH receptor) causing isolated GH deficiency (IGHD) have been published, mutations in GHRH itself have not been hitherto reported but are obvious candidates for GH deficiency. OBJECTIVE: The aim of this study was to identify mutations in GHRH in a large cohort of patients with IGHD. PATIENTS AND METHODS: DNA was isolated from 151 patients diagnosed with IGHD at national and international centers. Seventy-two patients fulfilled all the following criteria: severe short stature (height sd score ≤ -2.5), low peak GH after stimulation (peak ≤ 5 ng/ml), eutopic posterior pituitary lobe, and absence of mutations in GH1 and GHRHR and therefore were strong candidates for GHRH mutations. The coding sequence and splice sites of GHRH were amplified by PCR with intronic primers and sequenced. RESULTS: In five of 151 patients (four of 42 from Brazil), the GHRH c.223 C>T, p.L75F change was identified in heterozygosity. This variant has been previously reported as a polymorphism and is more frequent in African than European and Asian populations. Six allelic variants (five novel) that do not predict change of amino acids or splice sites were identified in five patients: c.147 C>T, p.S49S, IVS1 -70 G>A, IVS1 -74 T>C, IVS3 -47 del1, and IVS3 +7 G>A /IVS3+41 G>A. No functional mutations were found in this cohort. CONCLUSIONS: GHRH mutations were not identified in a selected cohort of patients with IGHD, suggesting that, if they exist, they may be an extremely rare cause of IGHD. Other, as-yet-unidentified genetic factors may be implicated in the genetic etiology of IGHD in our cohort.

Novel FGF8 mutations associated with recessive holoprosencephaly, craniofacial defects, and hypothalamo-pituitary dysfunction.

CONTEXT: Fibroblast growth factor (FGF) 8 is important for GnRH neuronal development with human mutations resulting in Kallmann syndrome. Murine data suggest a role for Fgf8 in hypothalamo-pituitary development; however, its role in the etiology of wider hypothalamo-pituitary dysfunction in humans is unknown. OBJECTIVE: The objective of this study was to screen for FGF8 mutations in patients with septo-optic dysplasia (n = 374) or holoprosencephaly (HPE)/midline clefts (n = 47). METHODS: FGF8 was analyzed by PCR and direct sequencing. Ethnically matched controls were then screened for mutated alleles (n = 480-686). Localization of Fgf8/FGF8 expression was analyzed by in situ hybridization in developing murine and human embryos. Finally, Fgf8 hypomorphic mice (Fgf8(loxPNeo/-)) were analyzed for the presence of forebrain and hypothalamo-pituitary defects. RESULTS: A homozygous p.R189H mutation was identified in a female patient of consanguineous parentage with semilobar HPE, diabetes insipidus, and TSH and ACTH insufficiency. Second, a heterozygous p.Q216E mutation was identified in a female patient with an absent corpus callosum, hypoplastic optic nerves, and Moebius syndrome. FGF8 was expressed in the ventral diencephalon and anterior commissural plate but not in Rathke's pouch, strongly suggesting early onset hypothalamic and corpus callosal defects in these patients. This was consolidated by significantly reduced vasopressin and oxytocin staining neurons in the hypothalamus of Fgf8 hypomorphic mice compared with controls along with variable hypothalamo-pituitary defects and HPE. CONCLUSION: We implicate FGF8 in the etiology of recessive HPE and potentially septo-optic dysplasia/Moebius syndrome for the first time to our knowledge. Furthermore, FGF8 is important for the development of the ventral diencephalon, hypothalamus, and pituitary.

Genetic causes and treatment of isolated growth hormone deficiency-an update.

Isolated growth hormone deficiency is the most common pituitary hormone deficiency and can result from congenital or acquired causes, although the majority of cases are idiopathic with no identifiable etiology. Known genes involved in the genetic etiology of isolated growth hormone deficiency include those that encode growth hormone (GH1), growth-hormone-releasing hormone receptor (GHRHR) and transcription factor SOX3. However, mutations are identified in a relatively small percentage of patients, which suggests that other, yet unidentified, genetic factors are involved. Among the known factors, heterozygous mutations in GH1 remain the most frequent cause of isolated growth hormone deficiency. The identification of mutations has clinical implications for the management of patients with this condition, as individuals with heterozygous GH1 mutations vary in phenotype and can, in some cases, develop additional pituitary hormone deficiencies. Lifelong follow-up of these patients is, therefore, recommended. Further studies in the genetic etiology of isolated growth hormone deficiency will help to elucidate mechanisms implicated in the control of growth and may influence future treatment options. Advances in pharmacogenomics will also optimize the treatment of isolated growth hormone deficiency and other conditions associated with short stature, for which recombinant human growth hormone is a licensed therapy.

Two cases of myomectomy complicated by intravascular hemolysis and renal failure: disseminated intravascular coagulation or hemolytic uremic syndrome?

OBJECTIVE: To present two cases of myomectomy complicated by intravascular hemolysis leading to acute renal failure and discuss the differential diagnosis and possible mechanism. DESIGN: Case report. SETTING: Minimally Invasive Therapy Unit, University Department of Obstetrics and Gynecology. PATIENT(S): Two premenopausal patients with uterine fibroids. INTERVENTION(S): Both patients underwent otherwise uncomplicated myomectomies, one by laparotomy and one by laparoscopy, with tourniquets around the uterine and ovarian vessels being used to control intraoperative bleeding. MAIN OUTCOME MEASURE(S): Renal function in the postoperative period. RESULT(S): Both patients developed a very rare complication after surgery of severe thrombocytopenia with microangiopathic hemolytic anemia leading to acute renal failure. One patient made a full recovery within weeks but the other still has reduced renal function almost 2 years after the surgery. The differential diagnosis consisted of disseminated intravascular coagulation or hemolytic uremic syndrome. CONCLUSION(S): The etiology of thrombotic microangiopathy in these patients was unclear, but disruption and manipulation of fibroids during surgery may have led to the dissemination of pro-coagulant tissue factor containing particles leading to disseminated intravascular coagulation or hemolytic uremic syndrome, perhaps aggravated by utero-ovarian ischemia caused by the tourniquets.

Genetic forms of hypopituitarism and their manifestation in the neonatal period.

The anterior pituitary gland is a central regulator of growth, reproduction and homeostasis. The development of the pituitary gland depends on the sequential temporal and spatial expression of transcription factors and signalling molecules. Naturally occurring and transgenic murine models have demonstrated a role for many of these molecules in the aetiology of congenital hypopituitarism. These include the transcription factors HESX1, PROP1, POU1F1, LHX3, LHX4, PITX1, PITX2, OTX2, SOX2 and SOX3. Mutations in any of the genes involved in pituitary development may result in congenital hypopituitarism, which manifests as the deficiency in one or more pituitary hormones. The phenotype can be highly variable and may consist of isolated hypopituitarism, or more complex disorders such as septo-optic dysplasia (SOD) and holoprosencephaly. Neonates with congenital hypopituitarism may present with non-specific symptoms, with or without associated developmental defects such as ocular, midline and genital abnormalities. Alternatively, they may be initially asymptomatic but at risk of developing pituitary hormone deficiencies over time. The overall incidence of mutations in known transcription factors in patients with hypopituitarism is low, indicating that many genes remain to be identified. Their characterization will further elucidate the pathogenesis of this complex condition and will shed light on normal pituitary development.

The role of SOX proteins in normal pituitary development.

Pituitary development is a complex process that depends on the co-ordinated spatial and temporal expression of transcription factors and signalling molecules that culminates in the formation of a complex organ that secretes six hormones from five different cell types. Given the fact that all distinct hormone producing cells arise from a common ectodermal primordium, the patterning, architecture and plasticity of the gland is impressive. Among the transcription factors involved in the early steps of pituitary organogenesis are SOX2 and SOX3, members of the SOX family that are emerging as key players in many developmental processes. Studies in vitro and in vivo in transgenic animal models have helped to elucidate their expression patterns and roles in the developing hypothalamo-pituitary region. It has been demonstrated that they may be involved in pituitary development either directly, through shaping of Rathke's pouch, or indirectly affecting signalling from the diencephalon. Their role has been further underlined by the pleiotropic effects of their mutations in humans that range from isolated hormone deficiencies to panhypopituitarism and developmental abnormalities affecting many organ systems. However, the exact mechanism of action of SOX proteins, their downstream targets and their interplay within the extensive network that regulates pituitary development is still the subject of a growing number of studies. The elucidation of their role is crucial for the understanding of a number of processes that range from developmental mechanisms to disease phenotypes and tumorigenesis.

Acanthosis nigricans and insulin sensitivity in patients with achondroplasia and hypochodroplasia due to FGFR3 mutations.

BACKGROUND AND AIMS: Acanthosis nigricans (AN) has been reported in association with severe skeletal dysplasias due to activating mutations in FGFR3, including thanatophoric dysplasia, severe achondroplasia (ACH) with developmental delay and AN (SADDAN syndrome), and Crouzon syndrome with AN. There are isolated reports of patients with ACH and AN. In this series, we report clinical and biochemical data on five male patients, four with ACH and one with hypochondroplasia (HCH), who developed AN without SADDAN. METHODS AND RESULTS: We compared the results of a 1.75 g/kg oral glucose tolerance test performed in patients with ACH/HCH and AN with age-, sex-, and puberty-matched short children. Three of the patients were treated with recombinant human GH (dose range, 45-50 microg/kg/d), one patient had discontinued treatment 6 months before presentation, and one had never been treated. All patients had a fasting plasma glucose of less than 6 mmol/liter, and no patient had a plasma glucose greater than 7.8 mmol/liter at 2 h after ingestion of a glucose load. Although body mass index was higher in patients with skeletal dysplasia (28.9 +/- 7.3 vs. 20 +/- 0.6 kg/m(2); P = 0.01), mean fasting plasma insulin concentration was greater in controls (14.4 +/- 4.8 vs. 6.0 +/- 4.5 mU/liter; P = 0.03), as was homeostasis assessment index for insulin resistance (2.5 +/- 0.9 vs. 1.17 +/- 0.8; P = 0.05). CONCLUSION: Our findings suggest that the development of AN in patients with ACH/HCH is not due to insulin insensitivity either on its own or secondary to treatment with recombinant human GH. Whether the AN is due to altered melanocyte function in these individuals remains to be established.

Expanding the spectrum of mutations in GH1 and GHRHR: genetic screening in a large cohort of patients with congenital isolated growth hormone deficiency.

CONTEXT: It is estimated that 3-30% of cases with isolated GH deficiency (IGHD) have a genetic etiology, with a number of mutations being reported in GH1 and GHRHR. The aim of our study was to genetically characterize a cohort of patients with congenital IGHD and analyze their characteristics. PATIENTS AND METHODS: A total of 224 patients (190 pedigrees) with IGHD and a eutopic posterior pituitary were screened for mutations in GH1 and GHRHR. To explore the possibility of an association of GH1 abnormalities with multiple pituitary hormone deficiencies, we have screened 62 patients with either multiple pituitary hormone deficiencies (42 pedigrees), or IGHD with an ectopic posterior pituitary (21 pedigrees). RESULTS: Mutations in GH1 and GHRHR were identified in 41 patients from 21 pedigrees (11.1%), with a higher prevalence in familial cases (38.6%). These included previously described and novel mutations in GH1 (C182X, G120V, R178H, IVS3+4nt, a>t) and GHRHR (W273S, R94L, R162W). Autosomal dominant, type II IGHD was the commonest form (52.4%), followed by type IB (42.8%) and type IA (4.8%). Patients with type II IGHD had highly variable phenotypes. There was no difference in the endocrinology or magnetic resonance imaging appearance between patients with and without mutations, although those with mutations presented with more significant growth failure (height, -4.7 +/- 1.6 SDS vs. -3.4 +/- 1.7 SDS) (P = 0.001). There was no apparent difference between patients with mutations in GH1 and GHRHR. CONCLUSIONS: IGHD patients with severe growth failure and a positive family history should be screened for genetic mutations; the evolving endocrinopathy observed in some of these patients suggests the need for long-term follow-up.