Endocrinology
Abstract
Genetic defects affecting steroid biosynthesis cause cortisol deficiency and differences of sex development; among them recessive mutations in the steroidogenic enzymes CYP11A1 and CYP11B, whose function is supported by reducing equivalents donated by ferredoxin reductase (FDXR) and ferredoxin. So far, mutations in the mitochondrial flavoprotein FDXR have been associated with a progressive neuropathic mitochondriopathy named FDXR-Related Mitochondriopathy (FRM), but cortisol insufficiency has not been documented. However, FRM patients often experience worsening or demise following stress associated with infections. We investigated two female FRM patients carrying the novel homozygous FDXR mutation p.G437R with ambiguous genitalia at birth and sudden death in the first year of life; they presented with cortisol deficiency and androgen excess compatible with 11-hydroxylase deficiency. In addition, steroidogenic FDXR-variant cell lines reprogrammed from three FRM patients’ fibroblasts displayed deficient mineralocorticoid and glucocorticoid production. Finally, Fdxr-mutant mice allelic to the severe p.R386W human variant, showed reduced progesterone and corticosterone production. Therefore, our comprehensive studies show that human FDXR variants may cause compensated, but possibly life-threatening adrenocortical insufficiency in stress by affecting adrenal glucocorticoid and mineralocorticoid synthesis through direct enzyme inhibition, most likely in combination with disturbed mitochondrial redox balance.
Authors
Emanuele Pignatti, Jesse Slone, María Ángeles Gómez Cano, Teresa Margaret Campbell, Jimmy Vu, Kay-Sara Sauter, Amit V. Pandey, Francisco Martínez-Azorín, Marina Alonso-Riaño, Derek E. Neilson, Nicola Longo, Therina du Toit, Clarissa D. Voegel, Taosheng Huang, Christa E. Flück
Abstract
Glucocorticoid synthesis by adrenal glands (AG) is regulated by the hypothalamic-pituitary-adrenal axis (HPA-axis) to facilitate stress responses when the host is exposed to stimuli. Recent studies have implicated macrophages (MФ) as potential steroidogenic regulators, but the molecular mechanisms by which AG MФ exert such influence remain unclear. In this study, we investigated the role of AG MФ in response to cold challenge or atherosclerotic inflammation as physiologic models of acute or chronic stress. Utilizing single-cell RNA sequencing, we observed dynamic AG MФ polarization toward classical activation and lipid-associated phenotypes following acute or chronic stimulation. Among the transcriptional alterations induced in MФ, Triggering Receptor Expressed on Myeloid (Trem2) was highlighted due to its dramatic upregulation following stress. Conditional deletion of MФ Trem2 revealed a protective role for Trem2 in stress responses. Mechanistically, Trem2 deletion led to increased AG MФ death, abolished the TGFβ-producing capacity of AG MФ, and resulted in enhanced glucocorticoid production. In addition, enhanced glucocorticoid production was replicated by blockade of TGFβ signaling. Together, these observations suggest that AG MФ restrict steroidogenesis through Trem2 and TGFβ, which opens potential avenues for immunotherapeutic interventions targeting the innate immune system to resolve stress-related disorders.
Authors
Yingzheng Xu, Michael T. Patterson, Bastien Dolfi, Alisha Zhu, Adeline Bertola, Patricia R. Schrank, Alexandre Gallerand, Ainsley E. Kennedy, Hannah Hillman, Lynn Dinh, Sia Shekhar, Samuel Tollison, Tyler D. Bold, Stoyan Ivanov, Jesse W. Williams
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
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