The glucocorticoid receptor (GR) is a major drug target in inflammatory disease. However, chronic glucocorticoid (GC) treatment leads to disordered energy metabolism, including increased weight gain, adiposity, and hepatosteatosis — all programs modulated by the circadian clock. We demonstrated that while antiinflammatory GC actions were maintained irrespective of dosing time, the liver was significantly more GC sensitive during the day. Temporal segregation of GC action was underpinned by a physical interaction of GR with the circadian transcription factor REVERBa and co-binding with liver-specific hepatocyte nuclear transcription factors (HNFs) on chromatin. REVERBa promoted efficient GR recruitment to chromatin during the day, acting in part by maintaining histone acetylation, with REVERBa-dependent GC responses providing segregation of carbohydrate and lipid metabolism. Importantly, deletion of Reverba inverted circadian liver GC sensitivity and protected mice from hepatosteatosis induced by chronic GC administration. Our results reveal a mechanism by which the circadian clock acts through REVERBa in liver on elements bound by HNF4A/HNF6 to direct GR action on energy metabolism.
Giorgio Caratti, Mudassar Iqbal, Louise Hunter, Donghwan Kim, Ping Wang, Ryan M. Vonslow, Nicola Begley, Abigail J. Tetley, Joanna L. Woodburn, Marie Pariollaud, Robert Maidstone, Ian J. Donaldson, Zhenguang Zhang, Louise M. Ince, Gareth Kitchen, Matthew Baxter, Toryn M. Poolman, Dion A. Daniels, David R. Stirling, Chad Brocker, Frank Gonzalez, Andrew S.I. Loudon, David A. Bechtold, Magnus Rattray, Laura C. Matthews, David W. Ray
Renin cells are crucial for survival: they control fluid-electrolyte and blood pressure homeostasis, vascular development, regeneration, and oxygen delivery to tissues. During embryonic development, renin cells are progenitors for multiple cell types which retain the memory of the renin phenotype. When there is a threat to survival, those descendants are transformed and reenact the renin phenotype to restore homeostasis. We tested the hypothesis that the molecular memory of the renin phenotype resides in unique regions and states of these cells’ chromatin. Using renin cells at various stages of stimulation, we identified regions in the genome where the chromatin is open for transcription, mapped histone modifications characteristic of active enhancers such as H3K27ac and deposition of transcriptional activators such a Med1 whose deletion results in ablation of Renin expression and low blood pressure. Using the rank ordering of super-enhancers, epigenetic re-writing, and enhancer deletion analysis, we found that renin cells harbor a unique set of super-enhancers that determine their identity. The most prominent Renin super-enhancer may act as a chromatin sensor of signals that convey the physiologic status of the organism and is responsible for the transformation of renin cell descendants to the renin phenotype, a fundamental process to ensure homeostasis.
Maria Florencia Martinez, Silvia Medrano, Evan A. Brown, Turan Tufan, Stephen Shang, Nadia Bertoncello, Omar Guessoum, Mazhar Adli, Brian C. Belyea, Maria Luisa S. Sequeira Lopez, R. Ariel Gomez
Movement of circulating fatty acids (FAs) to parenchymal cells requires their transfer across the endothelial cell (EC) barrier. The multi-ligand receptor cluster of differentiation 36 (CD36) facilitates tissue FA uptake and is expressed in ECs and parenchymal cells such as myocytes and adipocytes. Whether tissue uptake of FAs is dependent on EC or parenchymal cell CD36, or both, is unknown. Using a cell-specific deletion approach, we show that EC, but not parenchymal cell CD36 deletion increased fasting plasma FAs and postprandial triglycerides. EC-Cd36 knockout mice had reduced uptake of radiolabeled long chain FAs into heart, skeletal muscle, and brown adipose tissue; these uptake studies were replicated using [11C]palmitate PET scans. High fat diet-fed EC-CD36 deficient mice had improved glucose tolerance and insulin sensitivity. Both EC and cardiomyocyte (CM) deletion of CD36 reduced heart lipid droplet accumulation after fasting, but CM deletion did not affect heart glucose or FA uptake. Heart expression of several genes modulating glucose metabolism and insulin action increased with EC-CD36 deletion, but decreased with CM deletion. In conclusion, EC CD36 acts as a gatekeeper for parenchymal cell FA uptake, with important downstream effects on glucose utilization and insulin action.
Ni-Huiping Son, Debapriya Basu, Dmitri Samovski, Terri A. Pietka, Vivek S. Peche, Florian Willecke, Xiang Fang, Shui-Qing Yu, Diego Scerbo, Hye Rim Chang, Fei Sun, Svetlana Bagdasarov, Konstantinos Drosatos, Steve T. Yeh, Adam E. Mullick, Kooresh I. Shoghi, Namrata Gumaste, KyeongJin Kim, Lesley-Ann M. Huggins, Tenzin Lhakhang, Nada A. Abumrad, Ira J. Goldberg
The biological activity of 24R,25-dihydroxyvitamin D3 [24R,25(OH)2D3] remains controversial, but it has been suggested that it contributes to fracture healing. Cyp24a1–/– mice, synthesizing no 24R,25(OH)2D3, show suboptimal endochondral ossification during fracture repair, with smaller callus and reduced stiffness. These defects were corrected by 24R,25(OH)2D3 treatment, but not by 1,25-dihydroxyvitamin D3. Microarrays with Cyp24a1–/– callus mRNA identified FAM57B2 as a mediator of the 24R,25(OH)2D3 effect. FAM57B2 produced lactosylceramide (LacCer) upon specific binding of 24R,25(OH)2D3. Fam57b inactivation in chondrocytes (Col2-Cre Fam57bfl/fl) phenocopied the callus formation defect of Cyp24a1–/– mice. LacCer or 24R,25(OH)2D3 injections restored callus volume, stiffness, and mineralized cartilage area in Cyp24a1-null mice, but only LacCer rescued Col2-Cre Fam57bfl/fl mice. Gene expression in callus tissue suggested that the 24R,25(OH)2D3/FAM57B2 cascade affects cartilage maturation. We describe a previously unrecognized pathway influencing endochondral ossification during bone repair through LacCer production upon binding of 24R,25(OH)2D3 to FAM57B2. Our results identify potential new approaches to ameliorate fracture healing.
Corine Martineau, Roy Pascal Naja, Abdallah Husseini, Bachar Hamade, Martin Kaufmann, Omar Akhouayri, Alice Arabian, Glenville Jones, René St-Arnaud
BACKGROUND. A common germline variant in HSD3B1(1245A>C) encodes for a hyperactive 3β-hydroxysteroid dehydrogenase 1 (3βHSD1) missense that increases metabolic flux from extragonadal precursor steroids to DHT synthesis in prostate cancer. Enabling of extragonadal DHT synthesis by HSD3B1(1245C) predicts for more rapid clinical resistance to castration and sensitivity to extragonadal androgen synthesis inhibition. HSD3B1(1245C) thus appears to define a subgroup of patients who benefit from blocking extragonadal androgens. However, abiraterone, which is administered to block extragonadal androgens, is a steroidal drug that is metabolized by 3βHSD1 to multiple steroidal metabolites, including 3-keto-5α-abiraterone, which stimulates the androgen receptor. Our objective was to determine if HSD3B1(1245C) inheritance is associated with increased 3-keto-5α-abiraterone synthesis in patients. METHODS. First, we characterized the pharmacokinetics of 7 steroidal abiraterone metabolites in 15 healthy volunteers. Second, we determined the association between serum 3-keto-5α-abiraterone levels and HSD3B1 genotype in 30 patients treated with abiraterone acetate (AA) after correcting for the determined pharmacokinetics. RESULTS. Patients who inherit 0, 1, and 2 copies of HSD3B1(1245C) have a stepwise increase in normalized 3-keto-5α-abiraterone (0.04 ng/ml, 2.60 ng/ml, and 2.70 ng/ml, respectively; P = 0.002). CONCLUSION. Increased generation of 3-keto-5α-abiraterone in patients with HSD3B1(1245C) might partially negate abiraterone benefits in these patients who are otherwise more likely to benefit from CYP17A1 inhibition. FUNDING. Prostate Cancer Foundation Challenge Award, National Cancer Institute.
Mohammad Alyamani, Hamid Emamekhoo, Sunho Park, Jennifer Taylor, Nima Almassi, Sunil Upadhyay, Allison Tyler, Michael P. Berk, Bo Hu, Tae Hyun Hwang, William Douglas Figg, Cody J. Peer, Caly Chien, Vadim S. Koshkin, Prateek Mendiratta, Petros Grivas, Brian Rini, Jorge Garcia, Richard J. Auchus, Nima Sharifi
Tumor Necrosis Factor (TNF) is an important mediator in numerous inflammatory diseases, e.g., in inflammatory bowel diseases (IBD). In IBD, acute increases in TNF production can lead to disease flares. Glucocorticoids (GCs), which are steroids that bind and activate the glucocorticoid receptor (GR), are able to protect animals and humans against acute TNF-induced inflammatory symptoms. Mice with a poor transcriptional response of GR-dimer-dependent target genes were studied in a model of TNF-induced lethal inflammation. In contrast to the GRwt/wt mice, these GRdim/dim mice displayed a significant increase in TNF sensitivity and a lack of protection by the GC dexamethasone (DEX). Unchallenged GRdim/dim mice had a strong interferon-stimulated gene (ISG) signature, along with STAT1 upregulation and phosphorylation. This ISG signature was gut specific and, based on our studies with antibiotics, depended on the gut microbiota. GR dimers directly bound to short DNA sequences in the STAT1 promoter known as inverted repeat negative GRE (IR-nGRE) elements. Poor control of STAT1 in GRdim/dim mice led to failure to repress ISG genes resulting in excessive necroptosis induction by TNF. Our findings support a critical interplay between gut microbiota, interferons, necroptosis and GR in both the basal response to acute inflammatory challenges and in the pharmacological intervention by GCs.
Marlies Ballegeer, Kelly Van Looveren, Steven Timmermans, Melanie Eggermont, Sofie Vandevyver, Fabien Thery, Karen Dendoncker, Jolien Souffriau, Jolien Vandewalle, Lise Van Wyngene, Riet De Rycke, Nozomi Takahashi, Peter Vandenabeele, Jan Tuckermann, Holger M. Reichardt, Francis Impens, Rudi Beyaert, Karolien De Bosscher, Roosmarijn E. Vandenbroucke, Claude Libert
BACKGROUND. Recombinant leptin (metreleptin) ameliorates hyperphagia and metabolic abnormalities in leptin-deficient humans with lipodystrophy. We aimed to determine whether metreleptin improves glucose and lipid metabolism in humans when food intake is held constant. METHODS. Patients with lipodystrophy were hospitalized for 19 days with food intake held constant by controlled diet in an inpatient metabolic ward. In a non-randomized cross-over design, previously metreleptin-treated patients (n = 8) were continued on-metreleptin for five days, and off-metreleptin for the next 14 days (withdrawal cohort). This order was reversed in metreleptin-naïve patients (n = 14), who were restudied after six months of metreleptin treatment on an ad libitum diet (initiation cohort). Outcomes included insulin sensitivity by hyperinsulinemic-euglycemic clamp, fasting glucose and triglycerides, lipolysis measured using isotopic tracers, and liver fat by magnetic resonance spectroscopy. RESULTS. With food intake constant, peripheral insulin sensitivity decreased by 41% after stopping metreleptin for 14 days (withdrawal cohort) and increased by 32% after starting metreleptin for 14 days (initiation cohort). In the initiation cohort only, metreleptin decreased fasting glucose by 11%, triglycerides by 41%, and increased hepatic insulin sensitivity. Liver fat decreased from 21.8% to 18.7%. In the initiation cohort, lipolysis did not change independent of food intake, but decreased after six months on metreleptin on an ad libitum diet by 30% (palmitate turnover) to 35% (glycerol turnover). CONCLUSION. Using lipodystrophy as a human model of leptin deficiency and replacement, we showed that metreleptin improves insulin sensitivity, and decreases hepatic and circulating triglycerides, independent of its effects on food intake. TRIAL REGISTRATION. ClinicalTrials.gov, NCT01778556. FUNDING. This research was supported by the intramural research program of the National Institute of Diabetes and Digestive and Kidney Diseases.
Rebecca J. Brown, Areli Valencia, Megan Startzell, Elaine Cochran, Peter J. Walter, H. Martin Garraffo, Hongyi Cai, Ahmed M. Gharib, Ronald Ouwerkerk, Amber B. Courville, Shanna Bernstein, Robert J. Brychta, Kong Y. Chen, Mary Walter, Sungyoung Auh, Phillip Gorden
Spermatogenesis is regulated by the 2 pituitary gonadotropins, luteinizing hormone (LH) and follicle-stimulating hormone (FSH). This process is considered impossible without the absolute requirement of LH-stimulated testicular testosterone (T) production. The role of FSH remains unclear because men and mice with inactivating FSH receptor (FSHR) mutations are fertile. We revisited the role of FSH in spermatogenesis using transgenic mice expressing a constitutively strongly active FSHR mutant in a LH receptor–null (LHR-null) background. The mutant FSHR reversed the azoospermia and partially restored fertility of Lhr–/– mice. The finding was initially ascribed to the residual Leydig cell T production. However, when T action was completely blocked with the potent antiandrogen flutamide, spermatogenesis persisted. Hence, completely T-independent spermatogenesis is possible through strong FSHR activation, and the dogma of T being a sine qua non for spermatogenesis may need modification. The mechanism for the finding appeared to be that FSHR activation maintained the expression of Sertoli cell genes considered androgen dependent. The translational message of our findings is the possibility of developing a new strategy of high-dose FSH treatment for spermatogenic failure. Our findings also provide an explanation of molecular pathogenesis for Pasqualini syndrome (fertile eunuchs; LH/T deficiency with persistent spermatogenesis) and explain how the hormonal regulation of spermatogenesis has shifted from FSH to T dominance during evolution.
Olayiwola O. Oduwole, Hellevi Peltoketo, Ariel Poliandri, Laura Vengadabady, Marcin Chrusciel, Milena Doroszko, Luna Samanta, Laura Owen, Brian Keevil, Nafis A. Rahman, Ilpo T. Huhtaniemi
We have previously reported that the fractalkine (FKN)/CX3CR1 system represents a novel regulatory mechanism for insulin secretion and β cell function. Here, we demonstrate that chronic administration of a long-acting form of FKN, FKN-Fc, can exert durable effects to improve glucose tolerance with increased glucose-stimulated insulin secretion and decreased β cell apoptosis in obese rodent models. Unexpectedly, chronic FKN-Fc administration also led to decreased α cell glucagon secretion. In islet cells, FKN inhibited ATP-sensitive potassium channel conductance by an ERK-dependent mechanism, which triggered β cell action potential (AP) firing and decreased α cell AP amplitude. This results in increased glucose-stimulated insulin secretion and decreased glucagon secretion. Beyond its islet effects, FKN-Fc also exerted peripheral effects to enhance hepatic insulin sensitivity due to inhibition of glucagon action. In hepatocytes, FKN treatment reduced glucagon-stimulated cAMP production and CREB phosphorylation in a pertussis toxin–sensitive manner. Together, these results raise the possibility of use of FKN-based therapy to improve type 2 diabetes by increasing both insulin secretion and insulin sensitivity.
Matthew Riopel, Jong Bae Seo, Gautam K. Bandyopadhyay, Pingping Li, Joshua Wollam, Heekyung Chung, Seung-Ryoung Jung, Anne Murphy, Maria Wilson, Ron de Jong, Sanjay Patel, Deepika Balakrishna, James Bilakovics, Andrea Fanjul, Artur Plonowski, Duk-Su Koh, Christopher J. Larson, Jerrold M. Olefsky, Yun Sok Lee
The compensatory proliferation of insulin-producing β cells is critical to maintaining glucose homeostasis at the early stage of type 2 diabetes. Failure of β cells to proliferate results in hyperglycemia and insulin dependence in patients. To understand the effect of the interplay between β cell compensation and lipid metabolism upon obesity and peripheral insulin resistance, we eliminated LDL receptor–related protein 1 (LRP1), a pleiotropic mediator of cholesterol, insulin, energy metabolism, and other cellular processes, in β cells. Upon high-fat diet exposure, LRP1 ablation significantly impaired insulin secretion and proliferation of β cells. The diminished insulin signaling was partly contributed to by the hypersensitivity to glucose-induced, Ca2+-dependent activation of Erk and the mTORC1 effector p85 S6K1. Surprisingly, in LRP1-deficient islets, lipotoxic sphingolipids were mitigated by improved lipid metabolism, mediated at least in part by the master transcriptional regulator PPARγ2. Acute overexpression of PPARγ2 in β cells impaired insulin signaling and insulin secretion. Elimination of Apbb2, a functional regulator of LRP1 cytoplasmic domain, also impaired β cell function in a similar fashion. In summary, our results uncover the double-edged effects of intracellular lipid metabolism on β cell function and viability in obesity and type 2 diabetes and highlight LRP1 as an essential regulator of these processes.
Risheng Ye, Ruth Gordillo, Mengle Shao, Toshiharu Onodera, Zhe Chen, Shiuhwei Chen, Xiaoli Lin, Jeffrey A. SoRelle, Xiaohong Li, Miao Tang, Mark P. Keller, Regina Kuliawat, Alan D. Attie, Rana K. Gupta, William L. Holland, Bruce Beutler, Joachim Herz, Philipp E. Scherer