RESUMEN
AIMS: To test the hypothesis that brown adipose tissue (BAT) is a metformin target tissue by investigating in vivo uptake of [11 C]-metformin tracer in mice and studying in vitro effects of metformin on cultured human brown adipocytes. MATERIALS AND METHODS: Tissue-specific uptake of metformin was assessed in mice by PET/CT imaging after injection of [11 C]-metformin. Human brown adipose tissue was obtained from elective neck surgery and metformin transporter expression levels in human and murine BAT were determined by qPCR. Oxygen consumption in metformin-treated human brown adipocyte cell models was assessed by Seahorse XF technology. RESULTS: Injected [11 C]-metformin showed avid uptake in the murine interscapular BAT depot. Metformin exposure in BAT was similar to hepatic exposure. Non-specific inhibition of the organic cation transporter (OCT) protein by cimetidine administration eliminated BAT exposure to metformin, demonstrating OCT-mediated uptake. Gene expression profiles of OCTs in BAT revealed ample OCT3 expression in both human and mouse BAT. Incubation of a human brown adipocyte cell models with metformin reduced cellular oxygen consumption in a dose-dependent manner. CONCLUSION: These results support BAT as a putative metformin target.
Asunto(s)
Tejido Adiposo Pardo/efectos de los fármacos , Hipoglucemiantes/farmacocinética , Metformina/farmacocinética , Consumo de Oxígeno/efectos de los fármacos , Animales , Cimetidina/administración & dosificación , Relación Dosis-Respuesta a Droga , Humanos , Ratones , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Proteínas de Transporte de Catión Orgánico/metabolismo , Tomografía Computarizada por Tomografía de Emisión de Positrones , TranscriptomaRESUMEN
Brown adipose tissue is a promising therapeutic target in metabolic disorders due to its ability to dissipate energy and improve systemic insulin sensitivity and glucose homeostasis. ß-Adrenergic stimulation of brown adipocytes leads to an increase in oxygen consumption and induction of a thermogenic gene program that includes uncoupling protein 1 (Ucp1) and fibroblast growth factor 21 (Fgf21). In kinase inhibitor screens, we have identified glycogen synthase kinase 3 (GSK3) as a negative regulator of basal and ß-adrenergically stimulated Fgf21 expression in cultured brown adipocytes. In addition, inhibition of GSK3 also caused increased Ucp1 expression and oxygen consumption. ß-Adrenergic stimulation triggered an inhibitory phosphorylation of GSK3 in a protein kinase A (PKA)-dependent manner. Mechanistically, inhibition of GSK3 activated the mitogen activated protein kinase (MAPK) kinase 3/6-p38 MAPK-activating transcription factor 2 signaling module. In summary, our data describe GSK3 as a novel negative regulator of ß-adrenergic signaling in brown adipocytes.
Asunto(s)
Adipocitos Marrones/metabolismo , Factores de Crecimiento de Fibroblastos/genética , Glucógeno Sintasa Quinasa 3/genética , Termogénesis/genética , Proteína Desacopladora 1/genética , Animales , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Metabolismo Energético/genética , Regulación de la Expresión Génica/efectos de los fármacos , Glucosa/metabolismo , Resistencia a la Insulina/genética , Sistema de Señalización de MAP Quinasas/genética , Ratones , Consumo de Oxígeno/genética , Cultivo Primario de Células , Inhibidores de Proteínas Quinasas/administración & dosificación , Receptores Adrenérgicos beta/genética , Transducción de Señal/efectos de los fármacosRESUMEN
Brown adipose tissue with its constituent brown adipocytes is a promising therapeutic target in metabolic disorders due to its ability to dissipate energy and improve systemic insulin sensitivity and glucose homeostasis. The molecular control of brown adipocyte differentiation and function has been extensively studied in mice, but relatively little is known about such regulatory mechanisms in humans, which in part is due to lack of human brown adipose tissue derived cell models. Here, we used retrovirus-mediated overexpression to stably integrate human telomerase reverse transcriptase (TERT) into stromal-vascular cell fractions from deep and superficial human neck adipose tissue biopsies from the same donor. The brown and white pre-adipocyte cell models (TERT-hBA and TERT-hWA, respectively) displayed a stable proliferation rate and differentiation until at least passage 20. Mature TERT-hBA adipocytes expressed higher levels of thermogenic marker genes and displayed a higher maximal respiratory capacity than mature TERT-hWA adipocytes. TERT-hBA adipocytes were UCP1-positive and responded to ß-adrenergic stimulation by activating the PKA-MKK3/6-p38 MAPK signaling module and increasing thermogenic gene expression and oxygen consumption. Mature TERT-hWA adipocytes underwent efficient rosiglitazone-induced 'browning', as demonstrated by strongly increased expression of UCP1 and other brown adipocyte-enriched genes. In summary, the TERT-hBA and TERT-hWA cell models represent useful tools to obtain a better understanding of the molecular control of human brown and white adipocyte differentiation and function as well as of browning of human white adipocytes.