Endocrinology
Abstract
Caloric restriction improves metabolic health, but is often complicated by bone loss. We studied bone parameters in humans during a 10-day fast and identified candidate metabolic regulators of bone turnover. P1NP, a bone formation marker, decreased within 3 days of fasting. Whereas dual-energy X-ray absorptiometry measures of bone mineral density were unchanged after 10 days of fasting, high-resolution peripheral quantitative CT demonstrated remodeling of bone microarchitecture. Pathway analysis of longitudinal metabolomics data identified one-carbon metabolism as fasting-dependent. In cultured osteoblasts, we tested the functional significance of one-carbon metabolites modulated by fasting, finding that methionine — which surged after 3 days of fasting — impacted markers of osteoblast cell state in a concentration dependent manner, in some instances exhibiting a U-shaped response with both low and high concentrations driving putative anti-bone responses. Administration of methionine to mice for 5 days recapitulated some fasting effects on bone, including a reduction in serum P1NP. In conclusion, a 10-day fast in humans led to remodeling of bone microarchitecture, potentially mediated by a surge in circulating methionine. These data support an emerging model that points to a window of optimal methionine exposure for bone health.
Authors
Tânia Amorim, Naveen G.V. Kumar, Natalie L. David, William Dion, Trishya Pagadala, Nandini K. Doshi, Bokai Zhu, Andrey Parkhitko, Matthew L. Steinhauser, Pouneh K. Fazeli
Abstract
MAPK activating death domain (MADD) is a multifunctional protein regulating small GTPases RAB3 and RAB27, MAPK signaling, and cell survival. Polymorphisms in the MADD locus are associated with glycemic traits, but patients with biallelic variants in MADD manifest a complex syndrome affecting nervous, endocrine, exocrine, and hematological systems. We identified a homozygous splice site variant in MADD in 2 siblings with developmental delay, diabetes, congenital hypogonadotropic hypogonadism, and growth hormone deficiency. This variant led to skipping of exon 30 and in-frame deletion of 36 amino acids. To elucidate how this mutation causes pleiotropic endocrine phenotypes, we generated relevant cellular models with deletion of MADD exon 30 (dex30). We observed reduced numbers of β cells, decreased insulin content, and increased proinsulin-to-insulin ratio in dex30 human embryonic stem cell–derived pancreatic islets. Concordantly, dex30 led to decreased insulin expression in human β cell line EndoC-βH1. Furthermore, dex30 resulted in decreased luteinizing hormone expression in mouse pituitary gonadotrope cell line LβT2 but did not affect ontogeny of stem cell–derived GnRH neurons. Protein-protein interactions of wild-type and dex30 MADD revealed changes affecting multiple signaling pathways, while the GDP/GTP exchange activity of dex30 MADD remained intact. Our results suggest MADD-specific processes regulate hormone expression in pancreatic β cells and pituitary gonadotropes.
Authors
Kristiina Pulli, Jonna Saarimäki-Vire, Pekka Ahonen, Xiaonan Liu, Hazem Ibrahim, Vikash Chandra, Alice Santambrogio, Yafei Wang, Kirsi Vaaralahti, Anna-Pauliina Iivonen, Johanna Känsäkoski, Johanna Tommiska, Yasmine Kemkem, Markku Varjosalo, Sanna Vuoristo, Cynthia L. Andoniadou, Timo Otonkoski, Taneli Raivio
Abstract
Background. Upper body obesity (UBO) results in insulin resistance with regards to free fatty acid (FFA) release; how this differs by fat depot and sex between UBO and lean adults is unknown. We tested the hypothesis that insulin suppression of FFA release from the splanchnic bed, leg fat and upper body non-splanchnic (UBNS) adipose tissue would be impaired in UBO. Methods. Fourteen UBO (7 men, 7 women) and 14 healthy, normal weight (7 men, 7 women) volunteers participated in studies that included femoral artery, femoral vein and hepatic vein catheterization. We then measured leg and splanchnic plasma flow as well as FFA kinetics (using isotopic tracers) under overnight fasting, low- and high-dose insulin infusion using the insulin clamp technique. Results. We found the expected insulin resistance in UBO; the most quantitatively important difference between UBO and lean adults was greater FFA release from UBNS adipose tissue when plasma insulin concentrations are in the post-prandial, physiological range. There were obesity, but not sex differences in the regulation of splanchnic FFA release and sex differences in the regulation of leg FFA release. Conclusion. Reversing the defects in insulin-regulated UBNS adipose tissue FFA release would have the greatest impact on systemic FFA abnormalities in UBO. Trial Registration: (not applicable) Funding: These studies were supported by grants DK45343 and DK40484 from the U.S. Public Health Service, and the Novo Nordic Foundation (grant numbers NNF18OC0031804 and NNF16OC0021406) and the Independent Research Fund Denmark (grant number 8020-00420B).
Authors
Søren Nielsen, Michael D. Jensen
Abstract
Fibroblast Growth Factor 23 (FGF23) production has recently been shown to increase downstream of G⍺q/11-PKC signaling in osteocytes. Inactivating mutations in the gene encoding G⍺11 (GNA11) cause familial hypocalciuric hypercalcemia (FHH) due to impaired calcium-sensing receptor signaling. We explored the impact of G⍺11 deficiency on FGF23 production in mice with heterozygous (Gna11+/–) or homozygous (Gna11–/–) ablation of Gna11. Both Gna11+/– and Gna11–/– mice demonstrated hypercalcemia and mildly raised parathyroid hormone levels, consistent with FHH. Strikingly, these mice also displayed increased serum levels of total and intact FGF23 and hypophosphatemia. Gna11–/– mice showed augmented Fgf23 mRNA levels in the liver and heart, but not in bone or bone marrow, and evidence of systemic inflammation with elevated serum IL1β levels. Furin gene expression was significantly increased in the Gna11–/– liver, suggesting enhanced FGF23 cleavage despite the observed rise in intact FGF23 levels. Gna11–/– mice had normal renal function and reduced serum levels of glycerol-3-phosphate, excluding kidney injury as the primary cause of elevated intact FGF23 levels. Thus, G⍺11 ablation caused systemic inflammation and excess serum FGF23 in mice, suggesting that FHH patients, at least those with GNA11 mutations, may be at risk for these complications.
Authors
Birol Ay, Sajin Marcus Cyr, Kaitlin Klovdahl, Wen Zhou, Christina M. Tognoni, Yorihiro Iwasaki, Eugene P. Rhee, Alpaslan Dedeoglu, Petra Simic, Murat Bastepe
Abstract
Silver-Russell syndrome (SRS) is a heterogeneous disorder characterized by intrauterine and postnatal growth retardation. HMGA2 variants are a rare cause of SRS and its functional role in human linear growth is unclear. Patients with suspected SRS negative for 11p15LOM/mUPD7 underwent whole-exome and/or targeted-genome sequencing. Mutant HMGA2 protein expression and nuclear localization were assessed. Two Hmga2-knockin mouse models were generated. Five clinical SRS patients harbored HMGA2 variants with differing functional impacts: 2 stop-gain nonsense variants (c.49G>T, c.52C>T), c.166A>G missense variant, and 2 frameshift variants (c.144delC, c.145delA) leading to an identical, extended-length protein. Phenotypic features were highly variable. Nuclear localization was reduced/absent for all variants except c.166A>G. Homozygous knockin mice recapitulating the c.166A>G variant (Hmga2K56E) exhibited a growth-restricted phenotype. An Hmga2Ter76-knockin mouse model lacked detectable full-length Hmga2 protein, similarly to patient 3 and 5 variants. These mice were infertile, with a pygmy phenotype. We report a heterogeneous group of individuals with SRS harboring variants in HMGA2 and describe the first Hmga2 missense knockin mouse model (Hmga2K56E) to our knowledge causing a growth-restricted phenotype. In patients with clinical features of SRS but negative genetic screening, HMGA2 should be included in next-generation sequencing testing approaches.
Authors
Avinaash V. Maharaj, Emily Cottrell, Thatchawan Thanasupawat, Sjoerd D. Joustra, Barbara Triggs-Raine, Masanobu Fujimoto, Sarina G. Kant, Danielle van der Kaay, Agnes Clement-de Boers, Alice S. Brooks, Gabriel Amador Aguirre, Irene Martín del Estal, María Inmaculada Castilla de Cortázar Larrea, Ahmed Massoud, Hermine A. van Duyvenvoorde, Christiaan De Bruin, Vivian Hwa, Thomas Klonisch, Sabine Hombach-Klonisch, Helen L. Storr
Abstract
Patients with mutations in the thyroid hormone (TH) cell transporter MCT8 gene develop severe neuro-psychomotor retardation known as the Allan-Herndon-Dudley syndrome (AHDS). It is assumed that this is caused by a reduction in TH signaling in the developing brain, and treatment remains understandably challenging. Given species differences in brain TH transporters and the limitations of studies in mice, we generated brain organoids (BOs) using human iPSCs from MCT8-deficient patients. We found that MCT8-deficient BOs exhibit (i) impaired T3 transport in developing neural cells, as assessed through deiodinase-3-mediated T3 catabolism, (ii) reduced expression of genes involved in neurogenesis and neuronal maturation, and (iii) reduced T3-inducibility of TH-regulated genes. In contrast, the TH-analogs 3,5-diiodothyropropionic acid and 3,3’,5-triiodothyroacetic acid triggered normal responses (induction/repression of T3-responsive genes) in MCT8-deficient BOs, constituting a proof-of-concept that lack of T3 transport underlies the pathophysiology of AHDS, demonstrating the clinical potential for TH analogues to be used in treating AHDS patients. MCT8-deficient BOs represent a species-specific relevant preclinical model that can be utilized to screen drugs with potential benefits as personalized therapeutics for AHDS patients.
Authors
Federico Salas-Lucia, Sergio Escamilla, Antonio C. Bianco, Alexandra Dumitrescu, Samuel Refetoff
Abstract
In humans, Type 2 Diabetes Mellitus (T2DM) shows a higher prevalence in men compared to women, phenotype that has been attributed to a lower peripheral insulin sensitivity in men. Whether sex-specific differences in pancreatic β-cell function also contribute is largely unknown. Here we characterized the electrophysiological properties of β-cells in intact mouse male and female islets. Elevation of glucose concentration above 5 mM triggers an electrical activity with a similar glucose dependence in β-cells of both sexes. However, female β-cells have a more depolarized membrane potential and increased firing frequency compared to males. The higher membrane depolarization in female β-cells is caused by ~50% smaller Kv2.1 K+ currents compared to males but otherwise unchanged KATP, Ca2+-activated BK and SK, and background TASK1/TALK1 K+ current densities. In female β-cells the higher depolarization causes a membrane potential-dependent inactivation of the voltage-gated Ca2+ channels (CaV) resulting in reduced Ca2+ entry. Nevertheless, this reduced Ca2+ influx is offset by the higher action potential firing frequency. Since exocytosis of insulin granules does not show a sex-specific difference we conclude that the higher electrical activity promotes insulin release in females improving glucose tolerance.
Authors
Noelia Jacobo-Piqueras, Tamara Theiner, Stefanie M. Geisler, Petronel Tuluc
Abstract
Glucose homeostasis is achieved via complex interactions between the endocrine pancreas and other peripheral tissues and glucoregulatory neurocircuits in the brain that remain incompletely defined. Within the brain, neurons in the hypothalamus appear to play a particularly important role. Consistent with this notion, we report evidence that (pro)renin receptor (PRR) signaling within a subset of tyrosine hydroxylase (TH) neurons located in the hypothalamic paraventricular nucleus (PVNTH neurons) is a physiological determinant of the defended blood glucose level. Specifically, we demonstrate that PRR deletion from PVNTH neurons restores normal glucose homeostasis in mice with diet-induced obesity (DIO). Conversely, chemogenetic inhibition of PVNTH neurons mimics the deleterious effect of DIO on glucose. Combined with our finding that PRR activation inhibits PVNTH neurons, these findings suggest that in mice, (a) PVNTH neurons play a physiological role in glucose homeostasis, (b) PRR activation impairs glucose homeostasis by inhibiting these neurons, and (c) this mechanism plays a causal role in obesity-associated metabolic impairment.
Authors
Shiyue Pan, Lucas A.C. Souza, Caleb J. Worker, Miriam E. Reyes Mendez, Ariana Julia B. Gayban, Silvana G. Cooper, Alfredo Sanchez Solano, Richard N. Bergman, Darko Stefanovski, Gregory J. Morton, Michael W. Schwartz, Yumei Feng Earley
Abstract
Pseudohypoparathyroidism type 1B (PHP1B) is caused by aberrant genomic imprinting at the GNAS gene. Defining the underlying genetic cause in new patients is challenging because various genetic alterations (e.g., deletions, insertions) within the GNAS genomic region, including the neighboring STX16 gene, can cause PHP1B, and the genotype-epigenotype correlation has not been clearly established. Here, by analyzing PHP1B patients with a wide variety of genotypes and epigenotypes, we identified a GNAS differentially methylated region (DMR) of distinct diagnostic value. This region, GNAS AS2, was hypomethylated in patients with genetic alterations located centromeric but not telomeric of this DMR. The AS2 methylation status was captured by a single probe of the methylation-sensitive multiplex ligation-dependent probe amplification (MS-MLPA) assay utilized to diagnose PHP1B. In human embryonic stem cells, where NESP55 transcription regulates GNAS methylation status on the maternal allele, AS2 methylation depended on two imprinting control regions (STX16-ICR and NESP-ICR) essential for NESP55 transcription. These results suggest that the AS2 methylation status in PHP1B patients reflects the position at which the genetic alteration affects NESP55 transcription during an early embryonic period. Therefore, AS2 methylation levels enable mechanistic PHP1B categorization based on genotype-epigenotype correlation and thus help identify the underlying molecular defect in patients.
Authors
Yorihiro Iwasaki, Monica Reyes, Harald Jüppner, Murat Bastepe
Abstract
Suppression of glucagon hypersecretion can normalize hyperglycemia during type 1 diabetes (T1D). Activating erythropoietin-producing human hepatocellular receptor type-A4 (EphA4) on α cells reduced glucagon hypersecretion from dispersed α cells and T1D islets from both human donor and mouse models. We synthesized a high-affinity small molecule agonist for the EphA4 receptor, WCDD301, which showed robust plasma and liver microsome metabolic stability in both mouse and human preparations. In islets and dispersed islet cells from nondiabetic and T1D human donors, WCDD301 reduced glucagon secretion comparable to the natural EphA4 ligand, Ephrin-A5. In diabetic NOD and streptozotocin-treated mice, once-daily oral administration of WCDD301 formulated with a time-release excipient reduced plasma glucagon and normalized blood glucose for more than 3 months. These results suggest that targeting the α cell EphA4 receptor by sustained release of WCDD301 is a promising pharmacologic pathway for normalizing hyperglycemia in patients with T1D.
Authors
Farzad Asadi, Subhadra C. Gunawardana, Roland E. Dolle, David W. Piston
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