RESUMEN
Essential fructosuria (EF) is a benign, asymptomatic, autosomal recessive condition caused by loss-of-function variants in the ketohexokinase gene and characterized by intermittent appearance of fructose in the urine. Despite a basic understanding of the genetic and molecular basis of EF, relatively little is known about the long-term clinical consequences of ketohexokinase gene variants. We examined the frequency of ketohexokinase variants in the UK Biobank sample and compared the cardiometabolic profiles of groups of individuals with and without these variants alone or in combination. Study cohorts consisted of groups of participants defined based on the presence of one or more of the five ketohexokinase gene variants tested for in the Affymetrix assays used by the UK Biobank. The rs2304681:G>A (p.Val49Ile) variant was present on more than one-third (36.8%) of chromosomes; other variant alleles were rare (<1%). No participants with the compound heterozygous genotype present in subjects exhibiting the EF phenotype in the literature (Gly40Arg/Ala43Thr) were identified. The rs2304681:G>A (p.Val49Ile), rs41288797 (p.Val188Met), and rs114353144 (p.Val264Ile) variants were more common in white versus non-white participants. Otherwise, few statistically or clinically significant differences were observed after adjustment for multiple comparisons. These findings reinforce the current understanding of EF as a rare, benign, autosomal recessive condition.
Asunto(s)
Alelos , Fructoquinasas/genética , Variación Genética , Anciano , Bancos de Muestras Biológicas , Femenino , Fructoquinasas/deficiencia , Errores Innatos del Metabolismo de la Fructosa , Genotipo , Heterocigoto , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Reino UnidoRESUMEN
Angiotensin converting enzyme inhibitors, among them captopril, improve survival following myocardial infarction (MI). The mechanisms of captopril action remain inadequately understood due to its diverse effects on multiple signalling pathways at different time periods following MI. Here we aimed to establish the role of captopril in late-stage post-MI remodelling. Left anterior descending artery (LAD) ligation or sham surgery was carried out in male C57BL/6J mice. Seven days post-surgery LAD ligated mice were allocated to daily vehicle or captopril treatment continued over four weeks. To provide comprehensive characterization of the changes in mouse heart following MI a 3D light sheet imaging method was established together with automated image analysis workflow. The combination of echocardiography and light sheet imaging enabled to assess cardiac function and the underlying morphological changes. We show that delayed captopril treatment does not affect infarct size but prevents left ventricle dilation and hypertrophy, resulting in improved ejection fraction. Quantification of lectin perfused blood vessels showed improved vascular density in the infarct border zone in captopril treated mice in comparison to vehicle dosed control mice. These results validate the applicability of combined echocardiographic and light sheet assessment of drug mode of action in preclinical cardiovascular research.
Asunto(s)
Inhibidores de la Enzima Convertidora de Angiotensina/farmacología , Captopril/farmacología , Infarto del Miocardio/tratamiento farmacológico , Función Ventricular Izquierda/efectos de los fármacos , Animales , Modelos Animales de Enfermedad , Ecocardiografía , Ventrículos Cardíacos/diagnóstico por imagen , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/patología , Humanos , Masculino , Ratones , Microscopía , Infarto del Miocardio/diagnóstico por imagen , Infarto del Miocardio/genética , Infarto del Miocardio/patologíaRESUMEN
Whether vitamin D is chemopreventive and/or has potential therapeutically in prostate cancer is unresolved. One confounding factor is that many prostate cancers express a TMPRSS2:ERG fusion gene whose expression is increased both by androgens and by vitamin D receptor (VDR) activation. Two challenges that limit VDR agonist use clinically are hypercalcemia and the cooperation of VDR with ERG to hyper-induce the 1α,25-dihydroxyvitamin D3 metabolizing enzyme, CYP24A1, thus reducing VDR activity. Using the VCaP TMPRSS2:ERG positive cell line as a model, we found that a nonsecosteroidal CYP24A1 resistant VDR agonist, VDRM2, substantially reduces growth of xenograft tumors without inducing hypercalcemia. Utilizing next generation RNA sequencing, we found a very high overlap of 1,25D(OH)2D3 and VDRM2 regulated genes and by drawing upon previously published datasets to create an ERG signature, we found activation of VDR does not induce ERG activity above the already high basal levels present in VCaP cells. Moreover, we found VDR activation opposes 8 of the 10 most significant ERG regulated Hallmark gene set collection pathways from Gene Set Enrichment Analysis (GSEA). Thus, a CYP24A1 resistant VDR agonist may be beneficial for treatment of TMPRSS2:ERG positive prostate cancer; one negative consequence of TMPRSS2:ERG expression is inactivation of VDR signaling.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Receptores de Calcitriol/agonistas , Serina Endopeptidasas/genética , Regulador Transcripcional ERG/genética , Regulador Transcripcional ERG/metabolismo , Animales , Línea Celular Tumoral , Proliferación Celular , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Xenoinjertos , Humanos , Masculino , Ratones , Modelos Biológicos , Proteína Oncogénica p55(v-myc)/metabolismo , Receptores de Calcitriol/metabolismo , Transducción de Señal/efectos de los fármacos , Transcriptoma , Carga Tumoral , Vitamina D/análogos & derivados , Vitamina D/química , Vitamina D/farmacología , Vitamina D3 24-Hidroxilasa/genética , Vitamina D3 24-Hidroxilasa/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The high regenerative capacity of adult skeletal muscle relies on a self-renewing depot of adult stem cells, termed muscle satellite cells (MSCs). Androgens, known mediators of overall body composition and specifically skeletal muscle mass, have been shown to regulate MSCs. The possible overlapping function of androgen regulation of muscle growth and MSC activation has not been carefully investigated with regards to muscle regeneration.Therefore, the aim of this study was to examine coinciding androgen-mediated genetic changes in an in vitro MSC model and clinically relevant in vivo models. A gene signature was established via microarray analysis for androgen-mediated MSC engagement and highlighted several markers including follistatin (FST), IGF-1, C-X-C chemokine receptor 4 (CXCR4), hepatocyte growth factor (HGF) and glucocorticoid receptor (GR). In an in vivo muscle atrophy model, androgen re-supplementation significantly increased muscle size and expression of IGF-1, FST, and HGF, while significantly decreasing expression of GR. Biphasic gene expression profiles over the 7-day re-supplementation period identified temporal androgen regulation of molecular targets involved in satellite cell engagement into myogenesis. In a muscle injury model, removal of androgens resulted in delayed muscle recovery and regeneration. Modifications in the androgen signaling gene signature, along with reduced Pax7 and MyoD expression, suggested that limited MSC activation and increased inflammation contributed to the delayed regeneration. However, enhanced MSC activation in the androgen-deplete mouse injury model was driven by an androgen receptor (AR) agonist. These results provide novel in vitro and in vivo evidence describing molecular targets of androgen signaling, while also increasing support for translational use of AR agonists in skeletal muscle recovery and regeneration.
Asunto(s)
Andrógenos/metabolismo , Músculo Esquelético/citología , Músculo Esquelético/fisiología , Regeneración , Transducción de Señal , Animales , Biomarcadores/metabolismo , Línea Celular , Folistatina/genética , Regulación de la Expresión Génica/efectos de los fármacos , Ligandos , Masculino , Ratones , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/lesiones , Ratas , Receptores Androgénicos/metabolismo , Receptores CXCR4/genética , Receptores de Glucocorticoides/genética , Regeneración/efectos de los fármacos , Células Satélite del Músculo Esquelético/citología , Células Satélite del Músculo Esquelético/efectos de los fármacos , Células Satélite del Músculo Esquelético/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Skeletal muscle includes many individual fibers with diverse phenotypes. A barrier to understanding muscle glucose uptake at the cellular level has been the absence of a method to measure glucose uptake by single fibers from mammalian skeletal muscle. This study's primary objective was to develop a procedure to measure glucose uptake by single fibers from rat skeletal muscle. Rat epitrochlearis muscles were incubated ex vivo with [(3)H]-2-deoxy-d-glucose, with or without insulin or AICAR, before isolation of ~10-30 single fibers from each muscle. Fiber type (myosin heavy chain [MHC] isoform) and glucose uptake were determined for each single fiber. Insulin-stimulated glucose uptake (which was cytochalasin B inhibitable) varied according to MHC isoform expression, with ~2-fold greater values for IIA versus IIB or IIX fibers and ~1.3-fold greater for hybrid (IIB/X) versus IIB fibers. In contrast, AICAR-stimulated glucose uptake was ~1.5-fold greater for IIB versus IIA fibers. A secondary objective was to assess insulin resistance of single fibers from obese versus lean Zucker rats. Genotype differences were observed for insulin-stimulated glucose uptake and inhibitor κB (IκB)-ß abundance in single fibers (obese less than lean), with decrements for glucose uptake (44-58%) and IκB-ß (25-32%) in each fiber type. This novel method creates a unique opportunity for future research focused on understanding muscle glucose uptake at the cellular level.
Asunto(s)
Regulación de la Expresión Génica/fisiología , Glucosa/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Cadenas Pesadas de Miosina/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/farmacología , Animales , Desoxiglucosa/metabolismo , Genotipo , Glucosa/genética , Quinasa I-kappa B/genética , Quinasa I-kappa B/metabolismo , Insulina/farmacología , Resistencia a la Insulina , Masculino , Cadenas Pesadas de Miosina/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Isoformas de Proteínas , Ratas , Ratas Wistar , Ratas Zucker , Ribonucleótidos/farmacologíaRESUMEN
The primary purpose of this study was to assess the feasibility of applying a novel approach to measure myosin heavy chain (MHC) isoform expression, glucose uptake, fiber volume, and protein abundance in single muscle fibers of adult (9 months) and old (25 months) rats. Epitrochlearis muscle fibers were successfully isolated and analyzed for MHC isoform expression, glucose uptake, fiber volume, and protein (COXIV, APPL1, IκB-ß) abundance. Insulin-stimulated glucose uptake by single fibers did not differ between age groups, but there was a significant difference between fiber types (IIA > IIX > IIB/X ≈ IIB). There were also significant main effects of fiber type on APPL1 (IIX > IIB) and COXIV (IIA > IIX > IIB/X ≈ IIB) abundance, and IIB fibers were significantly larger than IIA fibers. This study established the feasibility of a new approach for assessing age-related differences in muscle at the single-fiber level and demonstrated the magnitude and rank order for fiber-type differences in insulin-stimulated glucose uptake of 9-month-old and 25-month-old rats.
Asunto(s)
Glucosa/metabolismo , Resistencia a la Insulina/fisiología , Fibras Musculares de Contracción Rápida/metabolismo , Fibras Musculares de Contracción Lenta/metabolismo , Cadenas Pesadas de Miosina/metabolismo , Envejecimiento/metabolismo , Animales , Estudios de Factibilidad , Immunoblotting , Técnicas In Vitro , Masculino , Isoformas de Proteínas , Ratas , Ratas Endogámicas , Sarcopenia/metabolismoRESUMEN
Skeletal muscle is a heterogeneous tissue. To further elucidate this heterogeneity, we probed relationships between myosin heavy chain (MHC) isoform composition and abundance of GLUT4 and four other proteins that are established or putative GLUT4 regulators [Akt substrate of 160 kDa (AS160), Tre-2/Bub2/Cdc 16-domain member 1 (TBC1D1), Tethering protein containing an UBX-domain for GLUT4 (TUG), and RuvB-like protein two (RUVBL2)] in 12 skeletal muscles or muscle regions from Wistar rats [adductor longus, extensor digitorum longus, epitrochlearis, gastrocnemius (mixed, red, and white), plantaris, soleus, tibialis anterior (red and white), tensor fasciae latae, and white vastus lateralis]. Key results were 1) significant differences found among the muscles (range of muscle expression values) for GLUT4 (2.5-fold), TUG (1.7-fold), RUVBL2 (2.0-fold), and TBC1D1 (2.7-fold), but not AS160; 2) significant positive correlations for pairs of proteins: GLUT4 vs. TUG (R = 0.699), GLUT4 vs. RUVBL2 (R = 0.613), TUG vs. RUVBL2 (R = 0.564), AS160 vs. TBC1D1 (R = 0.293), and AS160 vs. TUG (R = 0.246); 3) significant positive correlations for %MHC-I: GLUT4 (R = 0.460), TUG (R = 0.538), and RUVBL2 (R = 0.511); 4) significant positive correlations for %MHC-IIa: GLUT4 (R = 0.293) and RUVBL2 (R = 0.204); 5) significant negative correlations for %MHC-IIb vs. GLUT4 (R = -0.642), TUG (R = -0.626), and RUVBL2 (R = -0.692); and 6) neither AS160 nor TBC1D1 significantly correlated with MHC isoforms. In 12 rat muscles, GLUT4 abundance tracked with TUG and RUVBL2 and correlated with MHC isoform expression, but was unrelated to AS160 or TBC1D1. Our working hypothesis is that some of the mechanisms that regulate GLUT4 abundance in rat skeletal muscle also influence TUG and RUVBL2 abundance.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Proteínas Activadoras de GTPasa/metabolismo , Músculo Esquelético/metabolismo , Cadenas Pesadas de Miosina/metabolismo , Isoformas de Proteínas/metabolismo , Proteínas/metabolismo , Factores de Transcripción/metabolismo , Animales , Proteínas Portadoras/metabolismo , ADN Helicasas/metabolismo , Masculino , Ratas , Ratas WistarRESUMEN
The slow-twitch soleus, but not fast-twitch muscle, of old vs. adult rats has previously been demonstrated to become insulin resistant for in vivo glucose uptake. We probed cellular mechanisms for the age effect by assessing whether insulin resistance for glucose uptake was an intrinsic characteristic of the muscle ex vivo and by analyzing key insulin signaling steps. We hypothesized that isolated soleus and epitrochlearis (fast-twitch) muscles from old (25 mo) vs. adult (9 mo) male Fisher-344 x Brown Norway rats would have insulin resistance for Akt2 Thr308 phosphorylation (pAkt2Thr308), AS160 phosphorylation Thr642 (pAS160Thr642), and atypical PKC (aPKCzeta/lambda) activity corresponding in magnitude to the extent of insulin resistance for [3H]-2-deoxyglucose (2-DG) uptake. Epitrochlearis insulin-stimulated 2-DG uptake above basal values was unaltered by age, and epitrochlearis pAkt2Thr308, pAS160Thr642, and aPKCzeta/lambda activity were not significantly different in adult vs. old rats. Conversely, insulin-stimulated 2-DG uptake by the soleus of old vs. adult rats was reduced with 1.2 nM (42%) and 30 nM (28%) insulin concomitant with an age-related decline in pAkt2Thr308 of the insulin-stimulated soleus. There were no age effects on pAS160Thr642 or aPKCzeta/lambda activity or abundance of Akt2, AS160, GLUT4 or Appl1 protein in either muscle. The results suggest the possibility that an age-related decline in pAkt2Thr308, acting by a mechanism other than reduced pAS160Thr642, may play a role in the insulin resistance in the soleus of old rats. Skeletal muscle insulin resistance in old age is distinctive compared with other insulin-resistant rodent models that are not selective for greater insulin resistance in the soleus vs. the epitrochlearis.