Modifications of the GH Axis Reveal Unique Sexually Dimorphic Liver Signatures for Lcn13, Asns, Hamp2, Hao2, and Pgc1a.

Growth hormone (GH) modifies liver gene transcription in a sexually dimorphic manner to meet liver metabolic demands related to sex; thus, GH dysregulation leads to sex-biased hepatic disease. We dissected the steps of the GH regulatory cascade modifying GH-dependent genes involved in metabolism, focusing on the male-predominant genes Lcn13, Asns, and Cyp7b1, and the female-predominant genes Hao2, Pgc1a, Hamp2, Cyp2a4, and Cyp2b9. We explored mRNA expression in 2 settings: (i) intact liver GH receptor (GHR) but altered GH and insulin-like growth factor 1 (IGF1) levels (NeuroDrd2KO, HiGH, aHepIGF1kd, and STAT5bCA mouse lines); and (ii) liver loss of GHR, with or without STAT5b reconstitution (aHepGHRkd, and aHepGHRkd + STAT5bCA). Lcn13 was downregulated in males in most models, while Asns and Cyp7b1 were decreased in males by low GH levels or action, or constant GH levels, but unexpectedly upregulated in both sexes by the loss of liver Igf1 or constitutive Stat5b expression. Hao, Cyp2a4, and Cyp2b9 were generally decreased in female mice with low GH levels or action (NeuroDrd2KO and/or aHepGHRkd mice) and increased in HiGH females, while in contrast, Pgc1a was increased in female NeuroDrd2KO but decreased in STAT5bCA and aHepIGF1kd females. Bioinformatic analysis of RNAseq from aHepGHRkd livers stressed the greater impact of GHR loss on wide gene expression in males and highlighted that GH modifies almost completely different gene signatures in each sex. Concordantly, we show that altering different steps of the GH cascade in the liver modified liver expression of Lcn13, Asns, Cyp7b1, Hao2, Hamp2, Pgc1a, Cyp2a4, and Cyp2b9 in a sex- and gene-specific manner.

The Genomic Landscape of a Restricted ALL Cohort from Patients Residing on the U.S./Mexico Border.

Next-generation sequencing (NGS) has identified unique biomarkers yielding new strategies in precision medicine for the treatment of Acute lymphoblastic leukemia (ALL). Hispanics show marked health disparities in ALL, often absent in clinical trials or cancer research. Thus, it is unknown whether Hispanics would benefit equally from curated data currently guiding precision oncology. Using whole-exome sequencing, nine ALL patients were screened for mutations within genes known to possess diagnostic, prognostic and therapeutic value. Genes mutated in Hispanic ALL patients from the borderland were mined for potentially pathogenic variants within clinically relevant genes. KRAS G12A was detected in this unique cohort and its frequency in Hispanics from the TARGET-ALL Phase II database was three-fold greater than that of non-Hispanics. STAT5B N642H was also detected with low frequency in Hispanic and non-Hispanic individuals within TARGET. Its detection within this small cohort may reflect a common event in this demographic. Such variants occurring in the MAPK and JAK/STAT pathways may be contributing to Hispanic health disparities in ALL. Notable variants in ROS1, WT1, and NOTCH2 were observed in the ALL borderland cohort, with NOTCH2 C19W occurring most frequently. Further investigations on the pathogenicity of these variants are needed to assess their relevance in ALL.

Activation of the MAPK pathway (RASopathies) and partial growth hormone insensitivity.

RASopathies are a heterogeneous group of syndromes caused by germline mutations in genes encoding components of the RAS/MAPK pathway. Postnatal short stature is a cardinal feature of the RASopathies. Although the pathophysiology of these conditions is not fully understood to date, growth hormone insensitivity is one possibility, based on the observation of low IGF-1 values, generally preserved GH secretion and suboptimal growth response to recombinant human GH therapy. In this review, we will discuss the clinical and experimental evidence of GH insensitivity in patients with Noonan syndrome and other RASopathies, as well as their molecular basis.

A novel heterozygous STAT5B variant in a patient with short stature and partial growth hormone insensitivity (GHI).

BACKGROUND: The most frequent monogenic causes of growth hormone insensitivity (GHI) include defects in genes encoding the GH receptor itself (GHR), the signal transducer and activator of transcription (STAT5B), the insulin like-growth factor type I (IGF1) and the acid-labile subunit (IGFALS). GHI is characterized by a continuum of mild to severe post-natal growth failure. OBJECTIVE: To characterize the molecular defect in a patient with short stature and partial GHI. PATIENT AND METHODS: The boy was born at term adequate for gestational age from non-consanguineous normal-stature parents. At 2.2 years, he presented proportionate short stature (height -2.77 SDS), wide forehead and normal mental development. Whole-exome analysis and functional characterization (site-directed mutagenesis, dual luciferase reporter assay, immunofluorescence and western immunoblot) were performed. RESULTS: Biochemical and endocrinological evaluation revealed partial GH insensitivity with normal stimulated GH peak (7.8 ng/mL), undetectable IGF1 and low IGFBP3 levels. Two heterozygous variants in the GH-signaling pathway were found: a novel heterozygous STAT5B variant (c.1896G>T, p.K632N) and a hypomorphic IGFALS variant (c.1642C>T, p.R548W). Functional in vitro characterization demonstrated that p.K632N-STAT5b is an inactivating variant that impairs STAT5b activity through abolished phosphorylation. Remarkably, the patient's immunological evaluation displayed only a mild hypogammaglobulinemia, while a major characteristic of STAT5b deficient patients is severe immunodeficiency. CONCLUSIONS: We reported a novel pathogenic inactivating STAT5b variant, which may be associated with partial GH insensitivity and can present without severe immunological complications in heterozygous state. Our results contribute to expand the spectrum of phenotypes associated to GHI.

Primary Immunodeficiencies Unravel the Role of IL-2/CD25/STAT5b in Human Natural Killer Cell Maturation.

Natural killer (NK) cells play a pivotal role during immunity against viruses and circumstantial evidence also indicates that they can protect the host against developing tumors. Peripheral blood NK cells comprise CD56brightCD16lo/- cells that constitutively express CD25 (IL-2Rα) and CD56dimCD16hi cells that express CD25 upon activation. Using NK cells from two patients, one with a primary immunodeficiency characterized by a homozygous mutation in CD25 (born in year 2007 and studied since she was 3 years old) and one with a homozygous mutation in STAT5b (born in year 1992 and studied since she was 10 years old), we observed that the absence of IL-2 signaling through CD25 promotes the accumulation of CD56brightCD16high NK cells, and that CD56brightCD16lo, CD56brightCD16high, and CD56dimCD16high NK cells of this patient exhibited higher content of perforin and granzyme B, and proliferation capacity, compared to healthy donors. Also, CD56bright and CD56dim NK cells of this patient exhibited a reduced IFN-γ production in response to cytokine stimulation and increased degranulation against K562 cells. Also, the CD25-deficient patient presented a lower frequency of terminally differentiated NK cells in the CD56dimCD16hi NK subpopulation compared to the HD (assessed by CD57 and CD94 expression). Remarkably, CD56dimCD16high NK cells from both patients exhibited notoriously higher expression of CD62L compared to HD, suggesting that in the absence of IL-2 signaling through CD25 and STAT5b, NK cells fail to properly downregulate CD62L during their transition from CD56brightCD16lo/- to CD56dimCD16hi cells. Thus, we provide the first demonstration about the in vivo requirement of the integrity of the IL-2/CD25/STAT5b axis for proper human NK cell maturation.

Growth hormone insensitivity with immune dysfunction caused by a STAT5B mutation in the south of Brazil: evidence for a founder effect

ABSTRACT Homozygous STAT5B mutations causing growth hormone insensitivity with immune dysfunction were described in 10 patients since 2003, including two Brazilian brothers from the south of Brazil. Our objectives were to evaluate the prevalence of their STAT5B mutation in this region and to analyze the presence of a founder effect. We obtained DNA samples from 1,205 local inhabitants, 48 relatives of the homozygous patients and four individuals of another affected family. Genotyping for STAT5B c.424_427del mutation and for two polymorphic markers around it was done through fragment analysis technique. We also determined Y-chromosome and mtDNA haplotypes and genomic ancestry in heterozygous carriers. We identified seven families with STAT5B c.424_427del mutation, with 33 heterozygous individuals. The minor allelic frequency of this mutation was 0.29% in this population (confidence interval 95% 0.08-0.5%), which is significantly higher than the frequency of other pathogenic STAT5B allele variants observed in public databases (p < 0.001). All heterozygous carriers had the same haplotype present in the homozygous patients, found in only 9.4% of non-carriers (p < 0.001), supporting the existence of a founder effect. The Y-chromosome haplotype, mtDNA and genomic ancestry analysis indicated a European origin of this mutation. Our results provide compelling evidence for a founder effect of STAT5B c.424_427del mutation.

Impaired phosphorylation of JAK2-STAT5b signaling in fibroblasts from uremic children.

BACKGROUND: Chronic kidney disease (CKD) in children is characterized by severe growth failure. The growth hormone/insulin-like growth factor-1 (GH/IGF-1) axis in uremic animals shows a post-receptor impaired phosphorylation of Janus kinase 2/signal transducer and activator of transcription (JAK-STAT) proteins. The objective of our study was to characterize the intracellular phosphorylation of JAK-STAT signaling in fibroblasts from children with CKD on chronic peritoneal dialysis (PD). METHODS: Serum GH-binding protein (GHBP), IGF-1 and IGFBP3 were measured in 15 prepubertal CKD stage-5 children on PD. Cytoplasmic JAK2, cytoplasmic/nuclear STAT5b and nuclear IGFBP3, acid-labile subunit (ALS) and IGF-1 mRNA expression were quantified in fibroblasts obtained from skin biopsies before and after stimulation with 200 ng/ml recombinant human growth hormone (rhGH). Phosphorylation activity at both the cytoplasmic and nuclear level was expressed as the ratio phosphorylated (p)/total (t) abundance of the product (p/t) at 30 and 60 min. Fifteen healthy children were recruited as the control group. Values were expressed in arbitrary units (AU) and normalized for comparison. Significance was defined as p < 0.05. RESULTS: Thirty minutes after rhGH stimulus, the cytoplasmic (p/t) JAK2 ratio was significantly lower in patients than in controls [median and interquartile range (IQR): 7.4 (4.56) vs. 20.5 (50.06) AU]. At 60 min after rhGH stimulation, median JAK2 phosphorylation activity was still significantly lower in the patients [7.14 (IQR 3.8) vs. 10.2 (IQR 29.8) AU; p < 0.05]. The increase in the cytoplasmic (p/t) STAT5b/ß-actin ratio was lower at both measurement points in the patients compared to the controls, without reaching statistical significance between groups. Median IGFBP3 mRNA abundance was significantly decreased in fibroblasts from uremic patients 24 h after rhGH stimulation compared to the healthy controls [1.27 (IQR 0.83) vs. 2.37 (IQR 0.80) AU]. Median ALS and IGF-1 mRNA expression changed in response to rhGH stimuli at 24 and 48 h. CONCLUSION: In this study, children with CKD undergoing PD therapy showed an impaired phosphorylation of JAK2/STAT5b signaling in fibroblasts after GH stimulation, as well as impaired IGFBP3 mRNA abundance. Both impairments may be partially responsible for the observed resistance to the growth-promoting actions of GH in chronic kidney failure.

Deficiência da STAT5B: uma nova síndrome de insensibilidade ao hormônio de crescimento associada a acometimento imunológico / STAT5B deficiency: a new growth hormone insensitivity syndrome associated to immunological dysfunction

Uma nova apresentação da insensibilidade ao hormônio de crescimento (IGH), causada por mutações em homozigose no gene STAT5B (transdutor de sinal e ativador de transcrição tipo 5B), foi caracterizada nos últimos anos. Sua particularidade é a associação com quadros de disfunção imunológica grave, sendo o mais característico a pneumonite intersticial linfocítica. A presença concomitante de doenças crônicas imunológicas pode fazer com que a baixa estatura seja erroneamente considerada uma consequência do quadro clínico, levando ao subdiagnóstico dessa forma de IGH. O objetivo desta revisão é divulgar o conhecimento atual sobre essa rara patologia, facilitando o reconhecimento de pacientes com IGH secundária a mutações no gene STAT5B em ambulatórios de endocrinologia e de outras especialidades.

A new presentation of growth hormone insensitivity (GHI) caused by homozygous mutations in STAT5B (signal transducer and activator of transcription 5B) gene has been characterized in the last years. Its particularity is the association with severe immune dysfunction, especially with lymphocytic interstitial pneumonitis. This may mislead physicians into considering short stature as secondary to chronic immunological disease and consequently into underdiagnosing this form of GHI. The objective of this review is to propagate current knowledge about this rare pathology, facilitating the diagnosis of patients with GHI due to STAT5B mutations in endocrinology and other specialties clinics.

El receptor de la hormona de crecimiento humana (hGH) y la proteína de transporte de alta afinidad de la hGH / Human Growth Hormone (GH) Receptor and the High Affinity GH-Binding Protein

La hormona de crecimiento humana (hGH) circula parcialmente unida a su proteína de transporte de alta afinidad (GHBP) la cual resulta del clivaje proteolítico del dominio extracelular del receptor de GH. Recientemente la enzima TACE se identificó como la metaloproteasa responsable del clivaje y liberación de GHBP a circulación. Aunque aún se desconoce la función específica de esta proteína de transporte, distintos trabajos en la literatura demuestran efectos que potencian y efectos inhibitorios sobre la acción de GH. Por otro lado, existen evidencias que demuestran una fuerte relación entre la GHBP y el nivel de receptor de GH en el hígado en situaciones fisiológicas y patológicas. Esto permitió proponer a la determinación de GHBP en suero como un marcador periférico de la abundancia del receptor de GH en los tejidos. La determinación de la concentración de GHBP sería de especial interés para evaluar pacientes con diagnóstico probable de insensibilidad a la acción de GH y orientar el posterior estudio de anormalidades en el gen del receptor de GH. En la presente revisión, también se abordan dificultades metodológicas relacionadas a la medición de GHBP sérica.

Human circulating growth hormone (GH) is partly bound to a high-affinity binding protein (GHBP) which is derived from proteolytical cleavage of the extracellular domain of the GH receptor. Recently, the metalloproteinase TACE has been identified as an important enzyme responsive for inducing GHBP shedding. Although the specific function of GHBP is not fully known, both enhancing and inhibitory roles of this binding protein on GH action have been proposed. Many reports have demonstrated a close relationship between GHBP and the liver GH receptor status in physiological conditions and diseases. Moreover, serum GHBP measurement has been proposed as an useful peripheral index of the GH receptor abundance. Related to the latter, circulating GHBP concentration would be of special interest for the evaluation of GH insensitivity due to GH receptor gene abnormalities. In addition, the present review also focus on methodological problems concerning serum GHBP measurement.