scholarly journals Growth Hormone Receptor (Ghr) 6ω Pseudoexon Activation: A Novel Cause Of Severe Growth Hormone Insensitivity (Ghi)

2021 ◽  
Author(s):  
Emily Cottrell ◽  
Avinaash Maharaj ◽  
Jack Williams ◽  
Sumana Chatterjee ◽  
Grazia Cirillo ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Growth Hormone Receptor ◽  
Growth Failure ◽  
Index Patient ◽  
Postnatal Growth ◽  
The Novel ◽  
Growth Hormone Insensitivity ◽  
In Vitro Splicing ◽  
The Impact

Abstract Context Severe forms of Growth Hormone Insensitivity (GHI) are characterized by extreme short stature, dysmorphism and metabolic anomalies. Objective Identification of the genetic cause of growth failure in 3 ‘classical’ GHI subjects. Design A novel intronic GHR variant was identified, and in vitro splicing assays confirmed aberrant splicing. A 6Ω pseudoexon GHR vector and patient fibroblast analysis assessed the consequences of the novel pseudoexon inclusion and the impact on GHR function. Results We identified a novel homozygous intronic GHR variant (g.5:42700940T>G, c.618 + 836T> G), 44bp downstream of the previously recognized intronic 6Ψ GHR pseudoexon mutation in the index patient. Two siblings also harbored the novel intronic 6Ω pseudoexon GHR variant in compound heterozygosity with the known GHR c.181C>T (R43X) mutation. In vitro splicing analysis confirmed inclusion of a 151bp mutant 6Ω pseudoexon not identified in wild-type constructs. Inclusion of the 6Ω pseudoexon causes a frameshift resulting in a non-functional truncated GHR lacking the transmembrane and intracellular domains. The truncated 6Ω pseudoexon protein demonstrated extracellular accumulation and diminished activation of STAT5B signaling following growth hormone stimulation. Conclusion Novel GHR 6Ω pseudoexon inclusion results in loss of GHR function consistent with a severe GHI phenotype. This represents a novel mechanism of Laron syndrome and is the first deep intronic variant identified causing severe postnatal growth failure. The 2 kindreds originate from the same town in Campania, Southern Italy, implying common ancestry. Our findings highlight the importance of studying variation in deep intronic regions as a cause of monogenic disorders.

scholarly journals Phenotypic spectrum and responses to recombinant human IGF1 (rhIGF1) therapy in patients with homozygous intronic pseudoexon growth hormone receptor mutation

2018 ◽  
Vol 178 (5) ◽  
pp. 481-489 ◽  
Author(s):  
Sumana Chatterjee ◽  
Lucy Shapiro ◽  
Stephen J Rose ◽  
Talat Mushtaq ◽  
Peter E Clayton ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Growth Hormone Receptor ◽  
Elevated Serum ◽  
Growth Failure ◽  
Height Velocity ◽  
Phenotypic Data ◽  
Growth Hormone Insensitivity ◽  
Pakistani Origin ◽  
Pre Treatment ◽  
Serum Gh

Background Patients with homozygous intronic pseudoexon GH receptor (GHR) mutations (6Ψ) have growth hormone insensitivity (GHI) (growth failure, IGF1 deficiency and normal/elevated serum GH). We report 9 patients in addition to previously described 11 GHR 6Ψ patients and their responses to rhIGF1 therapy. Methods 20 patients (12 males, 11 families, mean age 4.0 ± 2.2 years) were diagnosed genetically in our centre. Phenotypic data and responses to rhIGF1 treatment were provided by referring clinicians. Continuous parametric variables were compared using Student t-test or ANOVA. Results 10/20 (50%) had typical facial features of GHI, 19/20 (95%) from consanguineous families and 18/20 (90%) of Pakistani origin. At diagnosis, mean height SDS: −4.1 ± 0.95, IGF1 SDS: −2.8 ± 1.4; IGFBP3 SDS: −3.0 ± 2.1 and mean basal and peak GH levels: 11.9 µg/L and 32.9 µg/L, respectively. 1/12 who had IGF1 generation test, responded (IGF1: 132–255 ng/mL). 15/20 (75%; 11M) received rhIGF1 (mean dose: 114 µg/kg twice daily, mean duration: 5.3 ± 2.5 years). Mean baseline height velocity of 4.7 ± 1.1 cm/year increased to 7.4 ± 1.8 cm/year (P = 0.001) during year 1 of therapy. Year 3 mean height SDS (−3.2 ± 1.0) was higher than pre-treatment height SDS (−4.3 ± 0.8) (P = 0.03). Mean cumulative increase in height SDS after year 5 was 1.4 ± 0.9. Difference between target height (TH) SDS and adult or latest height SDS was less than that of TH SDS and pre-treatment height SDS (2.1 ± 1.2 vs 3.0 ± 0.8; P = 0.02). Conclusion In addition to phenotypic heterogeneity in the cohort, there was mismatch between clinical and biochemical features in individual patients with 6Ψ GHR mutations. rhIGF1 treatment improved height outcomes.

scholarly journals Consequences of maternal undernutrition for fetal and postnatal hepatic insulin-like growth factor-I, growth hormone receptor and growth hormone binding protein gene regulation in the rat

1998 ◽  
Vol 20 (3) ◽  
pp. 313-326 ◽  
Author(s):  
SM Woodall ◽  
NS Bassett ◽  
PD Gluckman ◽  
BH Breier
Keyword(s):  
Growth Hormone ◽  
Growth Hormone Receptor ◽  
Growth Failure ◽  
Postnatal Growth ◽  
Exon 2 ◽  
Maternal Undernutrition ◽  
Igf I ◽  
Exon 1 ◽  
Ad Libitum ◽  
Postnatal Growth Failure

The mechanisms that contribute to postnatal growth failure following intrauterine growth retardation (IUGR) are poorly understood. We demonstrated previously that nutritional deprivation in the pregnant rat leads to IUGR in offspring, postnatal growth failure and to changes in endocrine parameters of the somatotrophic axis. The present study examines the effects of maternal undernutrition (30% of the ad libitum available diet; IUGR group) throughout pregnancy on hepatic insulin-like growth factor-I (IGF-I), growth hormone receptor (GHR) and GH-binding protein (GHBP) gene expression using solution hybridisation/RNase protection assays (RPAs). Animals were killed at fetal (E22, term=23 days) and postnatal (birth, days 5, 9, 15, 21) ages, livers were collected and RNA extracted for RPAs. Results demonstrate the presence of all IGF-I mRNAs resulting from transcription start sites (ss) in exon 1 (ss1/2, ss3, ss2 spliced), exon 2, the two IGF-I E-domain variants (Ea and Eb) as well as GHR and GHBP mRNAs in hepatic tissue at E22 in both the ad libitum fed and IUGR offspring. In the postnatal liver, IGF-I ss1/2, ss3, ss2 spliced, Ea and Eb IGF-I variants as well as GHR and GHBP mRNA transcripts increased in abundance from birth to day 21. IGF-I exon 2 transcripts were relatively constant from E22 until postnatal day 15, then increased at postnatal day 21 in both the ad libitum fed and IUGR offspring. The expressions of all hepatic IGF-I leader exon ss and Ea domain variants were significantly reduced in IUGR offspring (P<0.05) from E22 to postnatal day 9. In contrast, relative abundance of hepatic IGF-I Eb variants, GHR and GHBP mRNAs were unaltered in IUGR offspring compared with the ad libitum fed animals. Whether these postnatal effects of undernutrition are a direct consequence of IUGR or whether they are related, in part, to differences in postnatal food intake remains to be investigated. In summary, we have demonstrated that hepatic IGF-I ss within exon 1 and exon 2 are coordinately regulated. Use of exon 1 ss increased during normal development and decreased with IUGR without changes in GHR or GHBP gene expression. Eb transcripts, thought to represent GH-dependent endocrine regulation of IGF-I, were unchanged in IUGR. These results suggest a possible postreceptor defect in GH action as a consequence of IUGR.

Three novel Growth Hormone Receptor (GHR) splicing mutations causing a spectrum of Growth Hormone Insensitivity

2019 ◽  
Author(s):  
Emily Cottrell ◽  
Avinaash Maharaj ◽  
Tasneem Ladha ◽  
Sumana Chatterjee ◽  
Anna Grandone ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Hormone Receptor ◽  
Growth Hormone Receptor ◽  
Splicing Mutations ◽  
Growth Hormone Insensitivity

Periodic interactions of GH-releasing factor and somatostatin can augment GH release in vitro

1987 ◽  
Vol 253 (5) ◽  
pp. E508-E514
Author(s):  
J. Weiss ◽  
M. J. Cronin ◽  
M. O. Thorner
Keyword(s):  
Growth Hormone ◽  
Human Growth Hormone ◽  
Human Growth ◽  
Male Rat ◽  
Base Line ◽  
Additive Effects ◽  
Growth Hormone Releasing Factor ◽  
The Impact

Growth hormone (GH) is secreted as pulses in vivo. To understand the signals governing this periodicity, we have established a perifusion-based model of pulsatile GH release. Male rat anterior pituitaries were dispersed and perifused with pulses of human growth hormone-releasing factor-(1--40) (GHRF), with or without a continuous or discontinuous somatostatin tonus. An experiment was composed of a 1-h base-line collection followed by four 3-h cycles; each contained single or paired 10-min infusion(s) of 3 nM GHRF. In testing the impact of somatostatin, the protocol was identical except that 0.3 nM somatostatin was added 30 min into the base-line period and then was either continued throughout the study or withdrawn during the periods of GHRF infusion. GH base lines with somatostatin were lower than vehicle base lines (P less than 0.05). GHRF pulses generated consistent peaks of GH release between 200 and 300 ng. min-1. (10(7) cells)-1, and these peaks were not altered by continuous somatostatin. In contrast, withdrawal of somatostatin during GHRF administration elicited markedly higher GH peaks (P less than 0.05) and more total GH release (P less than 0.05). This response could not be accounted for by the additive effects of GHRF and somatostatin withdrawal.

361 GROWTH HORMONE RECEPTOR MUTANT PIGS PRODUCED BY USING THE CLUSTERED REGULARLY INTERSPACED SHORT PALINDROMIC REPEATS (CRISPR) AND CRISPR-ASSOCIATED SYSTEMS IN IN VITRO-PRODUCED ZYGOTES

2015 ◽  
Vol 27 (1) ◽  
pp. 269 ◽  
Author(s):  
M. Kurome ◽  
M. Dahlhoff ◽  
S. Bultmann ◽  
S. Krebs ◽  
H. Blum ◽  
...  
Keyword(s):  
Growth Hormone ◽  
Hormone Receptor ◽  
Growth Hormone Receptor ◽  
Large Animal Model ◽  
Large Animal ◽  
Single Mutation ◽  
Wild Type ◽  
Ghr Gene ◽  
Normal Fertilization

Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) technology is considered as an efficient strategy for generating gene edited large animals, such as pigs. Compared to somatic cell nuclear transfer, this new technology offers a relatively simple way to generate mutant pigs by direct injection of RNA into the cytoplasm of zygotes. Moreover, the use of in vitro produced zygotes would provide a highly effective and practical method for the production of porcine disease models for biomedical research. Here we examined the production efficiency of growth hormone receptor (GHR) mutant pigs by the combination of the CRISPR/Cas system and in vitro produced zygotes. In vitro maturation (IVM) of oocytes was performed as described previously (Kurome et al., Meth. Mol. Biol., in press). In all experiments, the same batch of frozen sperm was used. After IVM, around 20 oocytes with expanded cumulus cells were incubated with 5 × 104 spermatozoa in a 100-μL drop of porcine fertilization medium for 7 h. In vitro-produced embryos were assessed by the ratio of normal fertilization (eggs with 2 pronuclei) and blastocyst formation at Day 7. The Cas9 mRNA and a single guide RNA, recognising a short sequence of 20 base pairs in exon 3 of the GHR gene, were injected directly into the cytoplasm of the embryos 8.5 to 9.5 h after IVF. Injected embryos were transferred laparoscopically to recipient pigs, and 86.4% (57/66) of sperm-penetrated oocytes (66/96) exhibited normal fertilization. Incidence of polyspermy was relatively low (9/66, 13.6%). Developmental ability of in vitro-produced embryos to the blastocyst stage was 17.4% (24/138). In total, 426 RNA-injected embryos were transferred into 2 recipients, one of which became pregnant and gave birth to 8 piglets. All piglets were clinically healthy and developed normally. In 3 out of 8 piglets (37.5%), mutations were introduced. Next-generation sequencing revealed that all of them were mosaics: one with a single mutation (22% wild-type/78% mutant) and 2 piglets with 2 different mutations (80% wild-type/2% mutant_1/18% mutant_2 and 94% wild-type/4% mutant_1/2% mutant_2). Four out of 5 mutations caused a frameshift in the GHR gene. Our study reports for the first time generation of GHR mutant pigs by the use of the CRISPR/Cas system in in vitro-produced zygotes. Because all GHR mutant offspring were mosaic, Cas9 activation probably occurred after the 1-cell stage under our experimental conditions. The founder animal with the highest proportion of mutant GHR alleles will be used for breeding to establish a large animal model for Laron syndrome.This work is supported by the German Research Council (TR-CRC 127).

scholarly journals Comparison of Oxidative Status of Human Milk, Human Milk Fortifiers and Preterm Infant Formulas

Foods ◽  
2019 ◽  
Vol 8 (10) ◽  
pp. 458 ◽  
Author(s):  
Pozzo ◽  
Cirrincione ◽  
Russo ◽  
Karamać ◽  
Amarowicz ◽  
...  
Keyword(s):  
Human Milk ◽  
Antioxidant Capacity ◽  
Preterm Infant ◽  
Bovine Milk ◽  
Growth Failure ◽  
Postnatal Growth ◽  
Infant Formulas ◽  
Low Birth Weight Infants ◽  
Multivariate Statistical

Preterm and low birth weight infants require specific nutrition to overcome the accumulated growth deficit, and to prevent morbidities related to postnatal growth failure. In order to guarantee an adequate nutrient-intake, mother’s own milk, when available, or donor human milk, are usually fortified with additional nutrients, in particular proteins. Fortification with processed ingredients may result in additional intake in oxidative compounds, deriving from extensive heat treatments, that are applied during processing. The aim of the present work was to compare the in vitro antioxidant activity and oxidative compound content conveyed by different preterm infant foods and fortifiers, namely raw and pasteurized human milk, two different preterm infant formulas, three bovine milk-based fortifiers and two experimental donkey milk-based fortifiers. Univariate and multivariate statistical analyses revealed significant differences between the different products. The use of human milk minimizes the intake of dietary oxidative compound in comparison to infant formulas, irrespective of pasteurization or fortification, especially as far as malondialdehyde content is concerned. The addition of fortifiers to human milk increases its antioxidant capacity, and the choice of the protein source (hydrolysed vs. whole proteins) differently impacted the resulting total antioxidant capacity of the diet.

Post-receptor IGF1 insensitivity restricted to the MAPK pathway in a Silver–Russell syndrome patient with hypomethylation at the imprinting control region on chromosome 11

2012 ◽  
Vol 166 (3) ◽  
pp. 543-550 ◽  
Author(s):  
Luciana R Montenegro ◽  
Andrea C Leal ◽  
Debora C Coutinho ◽  
Helena P L Valassi ◽  
Mirian Y Nishi ◽  
...  
Keyword(s):  
Mapk Pathway ◽  
Growth Failure ◽  
Postnatal Growth ◽  
Chromosome 11 ◽  
Gene Expressions ◽  
Gh Treatment ◽  
Poor Growth ◽  
Mapk Activation ◽  
Silver Russell Syndrome

BackgroundHypomethylation of the paternal imprinting center region 1 (ICR1) is the most frequent molecular cause of Silver–Russell syndrome (SRS). Clinical evidence suggests that patients with this epimutation have mild IGF1 insensitivity.ObjectiveTo assess in vitro IGF1 action in fibroblast culture from a patient with SRS and IGF1 insensitivity.MethodsFibroblast cultures from one patient with SRS due to ICR1 demethylation and controls were established. The SRS patient has severe growth failure, elevated IGF1 level, and poor growth rate during human recombinant GH treatment. IGF1 action was assessed by cell proliferation, AKT, and p42/44-MAPK phosphorylation. Gene expression was determined by real-time PCR.ResultsDespite normal IGF1R sequence and expression, fibroblast proliferation induced by IGF1 was 50% lower in SRS fibroblasts in comparison with controls. IGF1 and insulin promoted a p42/44-MAPK activation in SRS fibroblasts 40 and 36%, respectively, lower than that in control fibroblasts. On the other hand, p42/44-MAPK activation induced by EGF stimulation was only slightly reduced (75% in SRS fibroblasts in comparison with control), suggesting a general impairment in MAPK pathway with a greater impairment of the stimulation induced by insulin and IGF1 than by EGF. A PCR array analysis disclosed a defect in MAPK pathway characterized by an increase in DUSP4 and MEF2C gene expressions in patient fibroblasts.ConclusionA post-receptor IGF1 insensitivity was characterized in one patient with SRS and ICR1 hypomethylation. Although based on one unique severely affected patient, these results raise an intriguing mechanism to explain the postnatal growth impairment observed in SRS patients that needs confirmation in larger cohorts.

scholarly journals Postnatal Growth Failure, Short Life Span, and Early Onset of Cellular Senescence and Subsequent Immortalization in Mice Lacking the Xeroderma Pigmentosum Group G Gene

1999 ◽  
Vol 19 (3) ◽  
pp. 2366-2372 ◽  
Author(s):  
Yoshi-Nobu Harada ◽  
Naoko Shiomi ◽  
Manabu Koike ◽  
Masahito Ikawa ◽  
Masaru Okabe ◽  
...  
Keyword(s):  
Early Onset ◽  
Xeroderma Pigmentosum ◽  
Excision Repair ◽  
Growth Failure ◽  
Postnatal Growth ◽  
Additional Function ◽  
Xeroderma Pigmentosum Group G ◽  
Postnatal Growth Failure ◽  
Deficient Mice

ABSTRACT The xeroderma pigmentosum group G (XP-G) gene (XPG) encodes a structure-specific DNA endonuclease that functions in nucleotide excision repair (NER). XP-G patients show various symptoms, ranging from mild cutaneous abnormalities to severe dermatological impairments. In some cases, patients exhibit growth failure and life-shortening and neurological dysfunctions, which are characteristics of Cockayne syndrome (CS). The known XPG protein function as the 3′ nuclease in NER, however, cannot explain the development of CS in certain XP-G patients. To gain an insight into the functions of the XPG protein, we have generated and examined mice lacking xpg (the mouse counterpart of the humanXPG gene) alleles. The xpg-deficient mice exhibited postnatal growth failure and underwent premature death. SinceXPA-deficient mice, which are totally defective in NER, do not show such symptoms, our data indicate that XPG performs an additional function(s) besides its role in NER. Our in vitro studies showed that primary embryonic fibroblasts isolated from thexpg-deficient mice underwent premature senescence and exhibited the early onset of immortalization and accumulation of p53.

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