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Resumen del trabajo presentado al 13th International Symposium on Insulin Receptor and Insulin Action, celebrado en Niza (Francia) del 20 al 22 de abril de 2017., [Question]: Brown adipose tissue (BAT), a specialized fat that dissipates energy to produce heat, plays an important role in the regulation of energy balance. We designed a pharmacological intervention protocol with a GLP-1R agonist and GLP-1R/ GCGR dual agonist to investigate their effects in preventing diet-induced obesity in m ice by enhancing BAT activation, browning and insulin signaling in differentiated brown adipocytes (BA). [Methods]: 8 week-old male C57Bl/6 mice were fed chow or high fat diet (HFD) for 10 weeks. HFD-fed mice were subsequently injected (s.c. every 2 days) vehicle or a GLP-1R agonist (10 nmol/kg) or G49, a GLP-1R/GCGR dual agonist (4.4 mg/kg). Parameters assessing obesity, glucose homeostasis, b-cell function, BAT activation and browning were analyzed following treatment for 3-6 weeks. lnsulin signaling was assayed in differentiated BA. [Results]: Following 3 weeks of treatment, body weight loss was greater in m ice injected G49 compared to those injected GLP-1R agonist. PET analysis revealed cold-induced activation of BAT by both drug treatments, but the effect was higher in G49-treated group. These differential effects were maintained following 6 weeks of treatment, at which time adiposity and macrophage inftltration of eWAT were reduced and UCP1 immunostaining in BAT and iWAT was increased. Also, the effects of G49 in inducing mRNA levels of BAT activation markers and browning in iWAT and eWAT were higher in m ice injected G49. A stronger decrease in plasma glucose and insulin was detected in G49-treated animals versus those receiving GLP-1 R agonist in parallel with improvement of islet morphology. The higher efftcacy of G49 was tested in differentiated BA by its effect in counteracting the impairment in insulin signaling induced by pro-inflammatory cytokines. [Conclusions]: Our results strongly suggest a novel role of an oxyntomodulin-like dual acting GLP-1R/GCGR agonist in reducing obesity and improve insulin sensitivity by increasing energy expenditure due to its effects in BAT and beta-cell function.

Trabajo presentado al CIBERDEM 8th Annual Meeting, celebrado en Barcelona (España) del 17 al 19 de mayo de 2017.

Resumen del trabajo presentado presentado al CIBERDEM Annual Meeting, celebrado en Cerdanyola del Vallès, Barcelona (España) del 11 al 13 de mayo de 2016.-- et al., [Background and aims]: Since non-alcoholic steatohepatitis (NASH) is associated with impaired liver regeneration, we aimed to investigate the effects of a dual acting glucagon-like peptide-1(GLP1)/glucagon (GCG) receptor agonist, termed G49, on NASH and hepatic regeneration. [Methods]: C57/Bl6 male mice fed chow or methionine and choline-deficient (MCD) diet for one week were divided into 4 groups: C (chow diet plus vehicle), MCD (MCD diet plus vehicle), C+G49 (chow diet plus G49) and M+G49 (MCD diet plus G49). Following 2 weeks of treatment, partial hepatectomy (PH) was performed and mice were maintained on the same treatment schedule for an additional 2 weeks. Treatment effects on liver function, hepatic regeneration and comprehensive genomic and metabolic profiling was conducted throughout the study. [Results]: NASH significantly improved in the M+G49 group manifested by reduced inflammation, steatosis, oxidative stress, apoptosis and increased mitochondria biogenesis. G49 treatment was also associated with a replenishment of hepatic glucose stores due to enhanced gluconeogenesis and reduced glucose utilization via pentose phosphate cycle (PPC) and oxidative metabolism. Following PH, treatment with G49 increased survival rates, restored the cytokine-mediated priming phase and enhanced proliferative capacity and hepatic regeneration ratio in mice on MCD diet. All NASH markers remained decreased in M+G49 mice after PH and glucose utilization was shifted to PPC and oxidative metabolism. Initiating G49 treatment immediately after PH was also effective in alleviating the pathological changes induced by the MCD diet. [Conclusion]: Dual acting GLP-1R/GCGR agonists such as G49 represent a novel therapeutic approach for patients with NASH and particularly in those requiring PH.

RNA interference (RNAi) is an exciting new tool to effect acute in vivo knockdown of genes for pharmacological target validation. Testing the application of this technology to metabolic disease targets, three RNAi delivery methods were compared in two frequently utilized preclinical models of obesity and diabetes, the diet-induced obese (DIO) and B6.V-Lep/J (ob/ob) mouse. Intraperitoneal (i.p.) and high pressure hydrodynamic intravenous (i.v.) administration of naked siRNA, and low pressure i.v. administration of shRNA-expressing adenovirus were assessed for both safety and gene knockdown efficacy using constructs targeting cJun N-terminal kinase 1 (JNK1). Hydrodynamic delivery of siRNA lowered liver JNK1 protein levels 40% in DIO mice, but was accompanied by iatrogenic liver damage. The ob/ob model proved even more intolerant of this technique, with hydrodynamic delivery resulting in severe liver damage and death of most animals. While well-tolerated, i.p. injections of siRNA in DIO mice did not result in any knockdown or phenotypic changes in the mice. On the other hand, i.v. injected adenovirus expressing shRNA potently reduced expression of JNK1 in vivo by 95% without liver toxicity. In conclusion, i.p. and hydrodynamic injections of siRNA were ineffective and/or inappropriate for in vivo gene targeting in DIO and ob/ob mice, while adenovirus-mediated delivery of shRNA provided a relatively benign and effective method for exploring liver target silencing.

Resumen del póster presentado a la 74th Scientific Sesions of American Diabetes Association, celebrada en San Francisco-California (US) del 13 al 17 de junio de 2104., A number of epidemiologic and clinical studies have shown an close between non-alcoholic fatty Iiver disease (NAFLD), insulin resistance and impaired Iiver function Since partial hepatectomy (PH) is a common surgical intervention to treat Iiver diseases, we designed a phamacological intervention with a PEGylated dual acting GLPlR and GCGR co-agonist to investigate the effect of this drug on hepatic regeneration in NAFLD. Eight week-old male mice on the C57BL/6 background were divided in 4 groups. Groups 1 and 2 were maintained on a chow diet (CHD) and groups 3 and 4 with choline-methionine-deficient (MCD) diet, an established model of steatohepatitis (NASH). After 1 week, groups 2 and 4 were injected with the co-agonist at 4 mg/kg every 2 days for 2 weeks Then, PH was performed and mice were maintained with similar intervention until sacrifice. Survival rate following PH increased with co-agonist treatment in CHD and MCD groups compared with vehicle-treated mice. In mice sacrificed at 48 h post-PH, higher DNA synthesis was found in the CHD and MCD plus co-agonist groups compared to mice receiving vehicle. Two weeks after PH, hepatic regeneration index significantly improved in mice with CHD or MCD diet plus co-agonist compared to the same groups treated with vehicle. Coagonist treatment in mice receiving MCD diet decreased transaminases and improved Iiver histology. Serum tryglicerides (TG) decreased in the MCD compared to CHD mice, reflecting lower capacity of the Iiverto export VLDL during NASH. However, the MCD plus co-agonist group restored TG to near levels of CHD mice. Parameters that assessed hepatic inflammation, oxidative stress, fibrosis and apoptosis were increased by the MCD diet and ameliorated by co-agonist treatment These data suggest a novel role of an Dxyntomodulin-like dual acting GLPlR and GCGR co-agonist in alleviating NASH and improving Iiver regeneration following PH and may offer potential therapeutic application in patients with NASH., Supported by Medlmmune.

Resumen del póster presentado a la Conferencia del Centro Nacional de Investigaciones Cardiovasculares (CNIC) titulada: "Energy homeostasis and metabolic disease", celebrada en Madrid del 7 al 8 de noviembre de 2014., [Objectives]: Partial hepatectomy (PH) is a common surgical intervention to treat liver diseases, particularly hepatocellular carcinoma. Since, a number of epidemiologic and clinical studies have shown a close association between non-alcoholic fatty liver disease (NAFLD) and response to PH, we designed a pharmacological intervention protocol with a PEGylated derivated of oxyntomodulyn aimed to investigate the effect of this drug on hepatic regeneration in a NAFLD model. [Methods]: 8 week-old male mice on the C57BU6 background were divided in 4 groups. Groups 1 and 2 were maintained on a chow diet (CHD) and groups 3 and 4 were fed with choline-methionine-deficient (MCD) diet. After 1 week, groups 2 and 4 were injected (S.C.) with oxyntomodulyn at 4 mg/kg every 2 days. Then, PH was performed and mice were maintained with the same diet and pharmacological intervention until they were sacrificed at several time-periods. Parameters that assess liver function and hepatic regeneration were analyzed. [Results]:S urvival rate following PW was increased with oxyntomodulyn in both dietary groups. Higher rate of DNA synthesis was found in the CHD and MCD oxyntomodulyn treated animals after 48 h post-PH. Hepatic regeneration index, analyzed 2 weeks after PH, was significantly improved in mice receiving CHD or MCD diet plus oxyntomodulyn as compared with not treated animals. In addition, oxyntomodulyn treatment was associated with decreased transaminase levels and improved liver histology in mice to receiving MCD diet. Moreover, several parameters that assessed hepatic inflammation, oxidative stress, fibrosis and apoptosis were increased by the MCD diet and ameliorated by oxyntomodulyn treatment. [Conclusions]: Taken together, these results strongly suggest a novel role of an oxyntomodulin-like dual acting GLPl R and GCGR co-agonist in alleviating NASH, improving liver regeneration following PH and may offer a potential therapeutic application in patients with NASH.