Cotadutide (GLP-1/Glucagon dual receptor agonist) modulates hypothalamic orexigenic and anorexigenic neuropeptides in obese mice.

The hypothalamic neuropeptides linked to appetite and satiety were investigated in obese mice treated with cotadutide (a dual receptor agonist of glucagon-like peptide 1 (GLP-1R)/Glucagon (GCGR)). Twelve-week-old male C57BL/6 mice were fed a control diet (C group, n = 20) or a high-fat diet (HF group, n = 20) for ten weeks. Each group was further divided, adding cotadutide treatment and forming groups C, CC, HF, and HFC for four additional weeks. The hypothalamic arcuate neurons were labeled by immunofluorescence, and protein expressions (Western blotting) for neuropeptide Y (NPY), proopiomelanocortin (POMC), agouti-related protein (AgRP), and cocaine- and amphetamine-regulated transcript (CART). Cotadutide enhanced POMC and CART neuropeptides and depressed NPY and AGRP neuropeptides. In addition, gene expressions (RT-qPCR) determined that Lepr (leptin receptor) and Calcr (calcitonin receptor) were diminished in HF compared to C but enhanced in CC compared to C and HFC compared to HF. Besides, Socs3 (suppressor of cytokine signaling 3) was decreased in HFC compared to HF, while Sst (somatostatin) was higher in HFC compared to HF; Tac1 (tachykinin 1) and Mc4r (melanocortin-4-receptor) were lower in HF compared to C but increased in HFC compared to HF. Also, Glp1r and Gcgr were higher in HFC compared to HF. In conclusion, the findings are compelling, demonstrating the effects of cotadutide on hypothalamic neuropeptides and hormone receptors of obese mice. Cotadutide modulates energy balance through the gut-brain axis and its associated signaling pathways. The study provides insights into the mechanisms underlying cotadutide's anti-obesity effects and its possible implications for obesity treatment.

AMPK/mTOR pathway significance in healthy liver and non-alcoholic fatty liver disease and its progression.

Obesity is related to several organs, but the liver is particularly affected. Adenosine monophosphate-activated protein kinase (AMPK) is a cellular energy sensor and regulator of liver lipid dysfunction and glucose metabolism. The mechanistic target of rapamycin (mTOR) is a protein kinase regulating cell growth, survival, metabolism, and immunity. Together, these pathways are involved in obesity, insulin resistance, non-alcoholic fatty liver disease (NAFLD) and its progression, and autophagy. During energy demand, liver kinase B (LKB) phosphorylation helps activate the AMPK/mTOR pathways. Likewise, the protein forkhead box O family (FOXO) negatively regulates adipogenesis by binding to the promoter sites of peroxisome proliferator-activated receptor-gamma coactivator 1-alpha, initiating adipogenesis. In addition, acetyl-CoA carboxylase, which regulates de novo lipogenesis, is linked to LKB and FOXO in developing NAFLD. The kinase complex, consisting of Unc-51-like autophagy-activating kinase 1 or 2 (ULK1, ULK2) by stimulating autophagy, and eliminating fat droplets in NAFLD, is regulated by mTORC1 and negatively regulated by AMPK that suppresses liver lipogenesis and increases fatty acid oxidation. Also, ULK1 is essential for initiating phagophore formation, establishing macrophagy, and generating autophagosomes. The selective breakdown of lipid droplets through macroautophagy, or macrolipophagy, occurs on a cellular energy level using free fatty acids. In addition, mTORC1 promotes lipogenesis by activating sterol regulatory element-binding protein. Finding new components and novel regulatory modes in signaling is significant for a better understanding of the AMPK/mTOR pathways, potentially facilitating the development of future diagnostic and therapeutic strategies for NAFLD and its progression to non-alcoholic steatohepatitis, cirrhosis, and hepatocellular carcinoma.

Pancreatic islet remodeling in cotadutide-treated obese mice.

Obesity and type 2 diabetes mellitus (T2DM) cause morphofunctional alterations in pancreatic islet alpha and beta cells. Therefore, we hypothesize that the new GLP-1/Glucagon receptor dual agonist cotadutide may benefit islet cell arrangement and function. Twelve-week-old C57BL/6 male mice were fed a control diet (C, 10 % kJ fat) or a high-fat diet (HF, 50 % kJ fat) for ten weeks. Then, the animals were divided into four groups for an additional 30 days and daily treated with subcutaneous cotadutide (30 nmol/kg) or vehicle: C, CC (control+cotadutide), HF, and HFC (high-fat+cotadutide). Cotadutide led to weight loss and reduced insulin resistance in the HFC group, increasing insulin receptor substrate 1 and solute carrier family 2 gene expressions in isolated islets. Also, cotadutide enhanced transcriptional factors related to islet cell transdifferentiation, decreasing aristaless-related homeobox and increasing the paired box 4 and 6, pancreatic and duodenal homeobox 1, v-maf musculoaponeurotic fibrosarcoma oncogene family protein A, neurogenin 3, and neurogenic differentiation 1. In addition, cotadutide improved the proliferating cell nuclear antigen, NK6 homeobox 1, B cell leukemia/lymphoma 2, but lessening caspase 3. Furthermore, cotadutide mitigated the endoplasmic reticulum (ER) stress-responsive genes, reducing transcription factor 4, DNA-damage-inducible transcript 3, and growth arrest and DNA-damage-inducible 45. In conclusion, our data demonstrated significant beneficial actions of cotadutide in DIO mice, such as weight loss, glycemic control, and insulin resistance improvement. In addition, cotadutide counteracted the pathological adaptive cellular arrangement of the pancreatic islet in obese mice, improving the markers of the transdifferentiating pathway, proliferation, apoptosis, and ER stress.

Cotadutide effect in liver and adipose tissue in obese mice.

Obesity, adipose tissue inflammation, and nonalcoholic fatty liver disease (NAFLD) are associated with insulin resistance and type 2 diabetes (T2D). Cotadutide is a dual agonist GLP-1/glucagon, currently in a preclinical study phase 2 that presents an anti-obesity effect. Diet-induced obese (DIO) C57BL/6 mice were treated for 4 weeks with cotadutide (30 nm/kg once a day at 14:00 h). The study focused on epididymal white adipose tissue (eWAT), liver (NAFLD), inflammation, lipid metabolism, AMP-activated protein kinase (AMPK)/mechanistic target of rapamycin (mTOR) pathways, and the endoplasmic reticulum (ER) stress. As a result, cotadutide controlled weight gain, glucose intolerance, and insulin resistance and showed beneficial effects on plasma markers in DIO mice (triacylglycerol, total cholesterol, alanine aminotransferase, and aspartate aminotransferase, leptin, adiponectin, monocyte chemoattractant protein-1, resistin, interleukin-6, tumor necrosis factor-alpha). Also, cotadutide lessened liver fat accumulation, eWAT proinflammatory markers, and ER stress. In addition, cotadutide improved lipid metabolism genes in eWAT, fatty acid synthase, peroxisome proliferator-activated receptor gamma and mitigates adipocyte hypertrophy and apoptosis. Furthermore, the effects of cotadutide were related to liver AMPK/mTOR pathway and ER stress. In conclusion, cotadutide induces weight loss and treats glucose intolerance and insulin resistance in DIO mice. In addition, cotadutide shows beneficial effects on liver lipid metabolism, mitigating steatosis, inflammation, and ER stress. Besides, in adipocytes, cotadutide decreases hypertrophy and reduces apoptosis. These actions rescuing the AMPK and mTOR pathway, improving lipid metabolism, and lessening NAFLD, inflammation, and ER stress in both eWAT and liver of DIO mice indicate cotadutide as a potentially new pharmacological treatment for T2D and associated obesity.

Treatment with semaglutide, a GLP-1 receptor agonist, improves extracellular matrix remodeling in the pancreatic islet of diet-induced obese mice.

AIMS: The extracellular matrix (ECM) is fundamental for the normal endocrine functions of pancreatic islet cells and plays key roles in the pathophysiology of type 2 diabetes. Here we investigated the turnover of islet ECM components, including islet amyloid polypeptide (IAPP), in an obese mouse model treated with semaglutide, a glucagon-like peptide type 1 receptor agonist. MAIN METHODS: Male one-month-old C57BL/6 mice were fed a control diet (C) or a high-fat diet (HF) for 16 weeks, then treated with semaglutide (subcutaneous 40 µg/kg every three days) for an additional four weeks (HFS). The islets were immunostained and gene expressions were assessed. KEY FINDINGS: Comparisons refer to HFS vs HF. Thus, IAPP immunolabeling and beta-cell-enriched beta-amyloid precursor protein cleaving enzyme (Bace2, -40 %) and heparanase immunolabeling and gene (Hpse, -40 %) were mitigated by semaglutide. In contrast, perlecan (Hspg2, +900 %) and vascular endothelial growth factor A (Vegfa, +420 %) were enhanced by semaglutide. Also, semaglutide lessened syndecan 4 (Sdc4, -65 %) and hyaluronan synthases (Has1, -45 %; Has2, -65 %) as well as chondroitin sulfate immunolabeling, and collagen type 1 (Col1a1, -60 %) and type 6 (Col6a3, -15 %), lysyl oxidase (Lox, -30 %) and metalloproteinases (Mmp2, -45 %; Mmp9, -60 %). SIGNIFICANCE: Semaglutide improved the turnover of islet heparan sulfate proteoglycans, hyaluronan, chondroitin sulfate proteoglycans, and collagens in the islet ECM. Such changes should contribute to restoring a healthy islet functional milieu and should reduce the formation of cell-damaging amyloid deposits. Our findings also provide additional evidence for the involvement of islet proteoglycans in the pathophysiology of type 2 diabetes.

Brown adipose tissue as an endocrine organ: updates on the emerging role of batokines.

Brown adipose tissue (BAT) remains active in adults, oxidizing fatty acids or glucose and releasing energy in the form of heat. Brown adipocytes and enhanced thermogenesis are targets for treating obesity and its comorbidities. BAT shows high synthesis activity and secretes several signaling molecules. The brown adipokines, or batokines, take action in an autocrine, paracrine, and endocrine manner. Batokines have a role in the homeostasis of the cardiovascular system, central nervous system, white adipose tissue, liver, and skeletal muscle and exert beneficial effects on BAT. The systemic function of batokines gives BAT an endocrine organ profile. Besides, the batokines Fibroblast Growth Factor-21, Vascular Endothelial Growth Factor A, Bone Morphogenetic Protein 8, Neuregulin 4, Myostatin, and Interleukin-6 emerge as targets to treat obesity and its comorbidities, deserving attention. This review outlines the role of six emerging batokines on BAT and their cross-talk with other organs, focusing on their physiological significance and diet-induced changes.

The mTORC1/AMPK pathway plays a role in the beneficial effects of semaglutide (GLP-1 receptor agonist) on the liver of obese mice.

PURPOSE: The liver regulates lipid metabolism. Decreasing mTOR (mechanistic target of rapamycin complex 1) and enhancing AMPK (AMP-activated protein kinase) help degrade hepatic diet-induced accumulated lipids. Therefore, the glucagon-like peptide type 1 receptor agonist (GLP-1) is indicated to treat obesity-related liver metabolic alterations. Then, we investigated the effects of semaglutide (recent GLP-1) by analyzing the liver mTORC1/AMPK pathway genes in obese mice. BASIC PROCEDURES: C57BL/6 male mice were separated into two groups and submitted for 16 weeks of obesity induction. Then they were treated for an additional four weeks with semaglutide (subcutaneous, 40 µg/kg once every three days). The groups formed were: C, control group; CS, control group plus semaglutide; HF, high-fat group; HFS, high-fat group plus semaglutide. Next, the livers were dissected, and rapidly fragments of all lobes were kept and frozen at -80° C for analysis (RT-qPCR). MAIN FINDINGS: Liver markers for the mTOR pathway associated with anabolism and lipogenesis de novo were increased in the HF group compared to the C group but comparatively attenuated by semaglutide. Also, liver markers for the AMPK pathway, which regulates chemical pathways involving the cell's primary energy source, were impaired in the HF group than in the C group but partly restored by semaglutide. CONCLUSION: the mTOR pathway was attenuated, and the insulin signaling and the AMPK pathway were enhanced by semaglutide, ameliorating the liver gene expressions related to the metabolism of obese mice. These findings are promising in delaying the progression of nonalcoholic fatty liver disease.

Intermittent fasting, high-intensity interval training, or a combination of both have beneficial effects in obese mice with nonalcoholic fatty liver disease.

Intermittent fasting (IF) and high-intensity interval training (HIIT) are procedures that might mitigate the effects of nonalcoholic fatty liver disease. Two groups of 3-month-old C57BL/6 male mice were fed for 16 weeks with a control (C) or high-fat (HF) diet. In the last 4 weeks of the study, IF, HIIT, and IF/HIIT were implemented. Obese HF animals showed liver fat accumulation with macro-, and micro-vesicular steatosis and inflammatory infiltrate. IF and HIIT successfully reduced liver steatosis in the HF-derived groups. IF, HIIT, and IF/HIIT were beneficial in improving glucose metabolism in both C-derived and HF-derived groups. High levels observed in plasmatic and liver levels of total cholesterol and triacylglycerol in the HF group compared to the C group were mitigated by IF, HIIT, and IF/HIIT. IF decreased adiponectin and increased leptin and insulin in the HF group. HIIT improved adiponectin and leptin. IF chances liver gene expressions: increased interleukin-6 (IL-6) in the C IF group, reduced IL-6, and PAI-1 in the HF group. IF/HIIT reduced IL-6, MCP-1, and PAI-1. IF and HIIT enhanced hepatic beta-oxidation. However, lipogenesis was reduced by IF and HIIT in the HF-derived groups. In conclusion, IF and HIIT benefit weight loss, hormones, glucose tolerance/insulin resistance, liver steatosis/inflammation, fatty acid oxidation, and lipogenesis. Furthermore, the IF groups showed beneficial effects more often and intensely than HIIT ones. The IF/HIIT combination was slightly more efficient than IF, indicating that IF is the primary intervening factor benefiting the obese mouse liver.

Obese mice weight loss role on nonalcoholic fatty liver disease and endoplasmic reticulum stress treated by a GLP-1 receptor agonist.

BACKGROUND/OBJECTIVES: The weight loss following Semaglutide treatment, a GLP-1 receptor agonist, might be responsible for some effects observed on the nonalcoholic fatty liver disease of obese mice. SUBJECTS/METHODS: Two groups of C57BL/6 male mice (n = 30/group) were fed the diets Control (C) or high-fat (HF) for 16 weeks. Then, separated into six new groups for an additional four weeks (n = 10/group) and treated with Semaglutide (S, 40 µg/kg) or paired feeding (PF) with S groups (C; C-S; C-PF; HF; HF-S; HF-PF). RESULTS: Semaglutide reduced energy consumption leading to weight loss. Simultaneously it improved glucose intolerance, glycated hemoglobin, insulin resistance/sensitivity, plasma lipids, and gastric inhibitory polypeptide. Semaglutide and paired feeding mitigated liver steatosis and adipose differentiation-related protein (Plin2) expression. Semaglutide also improved hormones and adipokines, reduced lipogenesis and inflammation, and increased beta-oxidation. Semaglutide lessened liver glucose uptake and endoplasmic reticulum (ER) stress. Among the 14 genes analyzed, 13 were modified by Semaglutide (93 %, six genes were changed exclusively by Semaglutide, and seven other genes were affected by the combination of Semaglutide and paired feeding). In seven genes, the paired diet showed no effect (50% of the genes tested). No marker was affected exclusively by paired feeding. CONCLUSIONS: Semaglutide and the consequent weight loss reduced obese mice liver inflammation, insulin resistance, and ER stress. However, weight loss alone did show few or no action on some significant study findings, like liver steatosis, leptin, insulin, resistin, and amylin. Furthermore, hepatic inflammation mediated by MCP-1 and partially by TNF-alpha and IL6 were also not reduced by weight loss. Furthermore, weight loss alone did not lessen hepatic lipogenesis as determined by the findings of SREBP-1c, CHREBP, PPAR-alpha, and SIRT1. Semaglutide was implicated in improving glucose uptake and lessening ER stress by reducing GADD45, independent of weight loss.

Pancreatic islet cells disarray, apoptosis, and proliferation in obese mice. The role of Semaglutide treatment.

There are significant injuries of pancreatic islets due to obesity and insulin resistance. Therefore, GLP-1 receptor agonists like Semaglutide might benefit the islet structural remodeling and its endocrine function in diet-induced obese mice. One-month-old male C57BL/6 mice were allotted into two dietary groups (n = 60/group) and fed for 16 weeks a control diet (C) or a high‒fat diet (HF). Then, for an additional four weeks, the main groups were resampled to include treatment (Semaglutide, S, 40 µg/kg), or paired feed with the treated group (PF), totaling six groups (n = 20/group): C, CS, CPF, HF, HFS, HFPF. Biochemistry, stereology, immunohistochemistry/immunofluorescence, confocal microscopy, and RT-qPCR were used in the study. The mouse model reproduced metabolism and bodily changes due to diet-induced obesity. Pancreatic islet hypertrophy was observed with alpha- and beta-cell remodeling, cell disarray, and apoptosis. Semaglutide increased islet cell proliferation and recovered islet size and alpha- and beta-cell masses. The changes include recovery of glucose and hormone levels, reduction of pro-inflammatory markers, improvement of pancreatic duodenal homeobox 1 (PDX-1), glucose transporter 2 (GLUT-2), v-maf musculoaponeurotic fibrosarcoma oncogene homolog A (MAF-A), and peroxisome proliferator-activated receptors (PPAR) -gamma. In conclusion, damage to the pancreatic islet caused by insulin resistance and the attempt to adapt the islet of obese mice involved different pathways, especially the pro-inflammatory pathway, PDX1, and PPAR-alpha and gamma. Semaglutide showed beneficial effects on these pathways, reducing the lesion on the islet. However, the weight loss influence of Semaglutide was of little relevance in the pancreatic islet.

Mice as an animal model for the study of adipose tissue and obesity / Ratones como modelo animal para el estudio del tejido adiposo y la obesidad

SUMMARY: The study of adipose tissue has gained increasing importance in the biomedical area due to its implications for health and obesity. Obesity is a situation of great concern mainly in the Western world due to its high prevalence and morbidity. Experimental studies on obesity often need a model where it is possible to carry out experiments, drug testing, and other therapeutic procedures, which are typically not possible in humans. Although several animals are used for obesity studies, rodents are by far the most used animals, and among rodents, mice are particularly indicated for this investigation. This mini review will introduce the challenging classification of obesity in rodents, paralleling human obesity, defining and classifying what an obese mouse is. The text will differentiate between white adipose tissue (WAT, aimed at endocrine secretion and lipogenesis) and brown adipose tissue (BAT, aimed at thermogenesis) and describe the browning process of white adipocytes in an adaptation to increase thermogenesis. The text will also describe the various types of body fat in mice with their differences and indications for investigation and teach how to recognize and dissect these fats. At the end of this introductory reading, the young researcher is expected to have acquired sufficient knowledge to start an experimental investigative project on obesity.

RESUMEN: El estudio del tejido adiposo ha ganado una importancia creciente en el área biomédica por sus implicaciones para la salud y la obesidad. La obesidad es una situación de gran preocupación, principalmente, en el mundo occidental debido a su alta prevalencia y morbilidad. Los estudios experimentales sobre la obesidad a menudo necesitan un modelo en el que sea posible realizar experimentos, pruebas de drogas y otros procedimientos terapéuticos, que normalmente no son posibles en humanos. Aunque se utilizan varios animales para estudios de obesidad, los roedores son, con mucho, los animales más utilizados y, entre los roedores, los ratones están especialmente indicados para esta investigación. Esta mini revisión presenta la desafiante clasificación de la obesidad en roedores, en paralelo con la obesidad humana, definiendo y clasificando qué es un ratón obeso. El texto diferencia entre tejido adiposo blanco (WAT, destinado a la secreción endocrina y lipogénesis) y tejido adiposo marrón (BAT, destinado a la termogénesis) y describe el proceso de pardeamiento de los adipocitos blancos en una adaptación para aumentar la termogénesis. El texto también describe los diversos tipos de grasa corporal en ratones con sus diferencias e indicaciones para la investigación y enseña cómo reconocer y diseccionar estas grasas. Al final de esta lectura introductoria, se espera que el joven investigador haya adquirido los conocimientos suficientes para iniciar un proyecto de investigación experimental sobre la obesidad.

Liver structural injury in leptin-deficient (ob/ob) mice: lipogenesis, fibrogenesis, inflammation, and apoptosis / Lesión estructural del hígado en ratones con deficiencia de leptina (ob/ob): lipogénesis, fibrogénesis, inflamación y apoptosis

SUMMARY: Nonalcoholic fatty liver disease (NAFLD) might progress the steatosis to nonalcoholic steatohepatitis (NASH), reaching a cirrhosis state and possibly hepatocellular carcinoma. The liver of three-month-old C57BL/6J mice (wild-type, WT group, n=10) and leptin- deficient obese mice (ob/ob group, n=10) were studied, focusing on the mechanisms associated with the activation of the hepatic stellate cells (HSCs) and pro-fibrogenesis. The obese ob/ob animals' liver showed steatosis, increased lipogenesis gene expressions, inflammation, increased pro-inflammatory gene expressions, inflammatory infiltrate, and potential apoptosis linked to a high Caspase 3 expression. In ob/ob mice, liver sections were labeled in the fibrotic zones by anti-alpha-smooth muscle actin (alpha-SMA) and anti-Reelin, but not in the WT mice. Moreover, the alpha-SMA gene expression was higher in the ob/ob group's liver than the WT group. The pro-fibrogenic gene expressions were parallel to anti- alpha-SMA and anti-Reelin immunofluorescence, suggesting HSCs activation. In the ob/ob animals, there were increased gene expressions involved with lipogenesis (Peroxisome proliferator-activated receptor-gamma, Cell death-inducing DFFA-like effector-c, Sterol regulatory element-binding protein-1c, and Fatty acid synthase), pro-fibrogenesis (Transforming growth factor beta1, Smad proteins- 3, Yes-associated protein-1, Protein platelet-derived growth factor receptor beta), pro-inflammation (Tumor necrosis factor-alpha, and Interleukin-6), and apoptosis (Caspase 3). In conclusion, the results in obese ob/ob animals provide a clue to the events in humans. In a translational view, controlling these targets can help mitigate the hepatic effects of human obesity and NAFLD progression to NASH.

RESUMEN: La enfermedad del hígado graso no alcohólico (HGNA) puede progresar de la esteatosis a esteatohepatitis no alcohólica (ENA), alcanzando un estado de cirrosis y posiblemente carcinoma hepatocelular. Se estudió el hígado de ratones C57BL / 6J de tres meses de edad (tipo salvaje, grupo WT, n = 10) y ratones obesos con deficiencia de leptina (grupo ob/ob, n = 10), centrándose en los mecanismos asociados con la activación de las células estrelladas hepáticas (HSC) y profibrogénesis. El hígado de los animales obesos ob/ob mostró esteatosis, aumento de la expresión génica de la lipogénesis, inflamación, aumento de la expresión génica proinflamatoria, infiltrado inflamatorio y posible apoptosis ligada a una alta expresión de Caspasa 3. En ratones ob/ob, las sec- ciones de hígado se marcaron en las zonas fibróticas con anti-alfa- actina de músculo liso (alfa-SMA) y anti-Reelin, pero no en los ratones WT. Además, la expresión del gen alfa-SMA fue mayor en el hígado del grupo ob/ob que en el grupo WT. Las expresiones génicas profibrogénicas fueron paralelas a la inmunofluorescencia anti-alfa-SMA y anti-Reelin, lo que sugiere la activación de las HSC. En los animales ob/ob, hubo un aumento de las expresiones génicas involucradas con la lipogénesis (receptor activado por proliferador de peroxisoma gamma, efector c similar a DFFA inductor de muerte celular, proteína de unión al elemento regulador de esterol-1c y sintasa de ácidos grasos), pro-fibrogénesis (factor de crecimiento transformante beta 1, proteínas Smad-3, proteína-1 asociada a Yes, receptor beta del factor de crecimiento derivado de plaquetas de proteínas), proinflamación (factor de necrosis tumoral alfa e interleucina-6) y apoptosis (caspasa 3). ). En conclusión, los resultados en animales obesos ob/ob proporcionan una pista de los eventos en humanos. Desde un punto de vista traslacional, el control de estos objetivos puede ayudar a mitigar los efectos hepáticos de la obesidad humana y la progresión de HGNA a ENA.

Nutritional research and fetal programming: parental nutrition influences the structure and function of the organs / Investigación nutricional y programación fetal: la nutrición parental influye en la estructura y función de los órganos

SUMMARY: Rodents are animals extensively used in biomedical and nutrition research, a necessary step before the research in humans. The composition and type of administration of the experimental diets are relevant and should be thought, considering each type of animal used in the research. It is particularly important to consider, among others, the metabolic differences between species and food needs in macro- and micronutrients to avoid possible bias. The American Institute of Nutrition (AIN) made recommendations for rodents, adapted to the period of growth (AIN-93G), which are pivotal in fetal programming studies. The experiments can be compared among different studies and better translated into humans, considering these limitations in the nutrition of parents and offspring. The review addresses different compositions of experimental food for rodents during development with the ability to induce fetal programming in the offspring and chronic diseases in adulthood due to the nutrition of the mother and father. The 'developmental origins of health and disease' (DOHaD) concept due to maternal nutrition is commented considering the protein restriction, vitamin D restriction, obesity, and intake of fructose or fish-oil. The 'paternal origins of health and disease transmission' (POHaD), because of the nutritional state of the father, were also analyzed in the review, primarily considering the obesity of the father. The review proposes some diet compositions to experimental research considering varied nutritional situations, hoping to assist young researchers or researches not familiar with experimental diet manipulations in the elaboration of the projects.

RESUMEN: Los roedores son animales utilizados frecuentemente en la investigación biomédica y nutricional, un paso necesario antes de la investigación en humanos. La composición y el tipo de administración de las dietas experimentales son relevantes y se debe considerar cada tipo de animal utilizado en los estudios. Es particularmente importante considerar las diferencias metabólicas entre las especies y las necesidades alimentarias de macro y micronutrientes para evitar posibles sesgos. El Instituto Americano de Nutrición (AIN) estableció recomendaciones para los roedores, adaptadas al período de crecimiento (AIN-93G), que son fundamentales en los estudios de programación fetal. Los experimentos se pueden comparar entre diferentes estudios y aplicar en humanos, considerando estas limitaciones en la nutrición de padres e hijos. La revisión aborda diferentes composiciones de alimentos para estudios experimentales en roedores durante su desarrollo, con la capacidad de inducir programación fetal en la descendencia y enfermedades crónicas en la adultez, considerando la nutrición de los padres. El concepto de 'orígenes del desarrollo de la salud y la enfermedad' (DOHaD) debido a la nutrición materna se comenta considerando la restricción de proteínas, la restricción de vitamina D, la obesidad y la ingesta de fructosa o aceite de pescado. Los 'orígenes paternos de la salud y transmisión de enfermedades' (POHaD), debido al estado nutricional del padre, también fueron analizados considerando principalmente la obesidad del padre. La revisión propone algunas composiciones dietéticas a la investigación experimental considerando situaciones nutricionales variadas, con la esperanza de ayudar a jóvenes investigadores o investigadores no familiarizados con las manipulaciones experimentales de la dieta en la elaboración de los proyectos.

Intermittent fasting benefits on alpha- and beta-cell arrangement in diet-induced obese mice pancreatic islet.

AIMS: There is a pancreatic islet adaptation in obese subjects, resulting in insulin resistance and diabetes type 2. We studied the effect of intermittent fasting (IntF) on the islet structure of diet-induced obese (DIO) mice. METHODS: Three-month-old male mice fed a control diet (C, 10% Kcal fat) or a high-fat diet (HF, 50% Kcal fat) for two months (n = 20 each group). Then, half of each group did IntF (alternating 24 h fed/24 h fast), continuing in their diets four more weeks: C, C-IntF, HF, HF-IntF. Islets were prepared to microscopy or isolated for molecular analysis. RESULTS: HF group (vs. C group) showed hyperglycemia, hyperinsulinemia, hyperleptinemia, hypoadiponectinemia, glucose intolerance, insulin resistance, and islet hypertrophy with a consequent higher both the alpha-cell and beta-cell masses. In the HF group (vs. C), there was low PDX1 (pancreatic and duodenal homeobox 1), and IntF did not alter PDX1. There was a low p-AKT/AKT ratio (protein kinase B), and IntF enhanced it. Also, tumor suppressor p53 was increased, and IntF decreased it. IL (interleukin) -6 was higher in the HF group (vs. C), and HF-IntF (vs. C-IntF). Any significant change in NFkB was seen among groups. CONCLUSIONS: IntF improves pancreatic islet structure in DIO mice, even with continued HF diet intake, primarily considering on the alpha- and beta-cell masses regulation, then improving insulin signaling and decreasing cell apoptosis. Future research should explore whether the shortening of the IntF extend could maintain the benefits observed in the long term.

Pancreatic islet stereology: estimation of beta cells mass / Estereología de los islotes pancreáticos: estimación de la masa de células beta

Obesity and its comorbidities are becoming epidemic in the Western world. Beta cell mass estimation is an important indicator to track the progression of insulin resistance/type 2 diabetes, particularly in experimental studies, where it can be performed with stereological tools in an unbiased way. In this work, we present a simple protocol that can contribute to doing the practice of estimating the mass of beta cells more frequent and reproducible. As with any quantitative study, the necessary precautions regarding sampling and randomness must be respected.

La obesidad y sus comorbilidades se están convirtiendo en una epidemia en el mundo occidental. La estimación de la masa de células beta es un indicador importante para rastrear la progresión de la resistencia a la insulina/diabetes tipo 2, particularmente en estudios experimentales, donde se puede realizar con herramientas estereológicas de manera imparcial. En este trabajo presentamos un protocolo simple que puede contribuir a que la práctica de estimar la masa de células beta sea más frecuente y reproducible. Como en cualquier estudio cuantitativo, deben respetarse las precauciones necesarias con respecto al muestreo y la aleatoriedad.

Metformin enhances mitochondrial biogenesis and thermogenesis in brown adipocytes of mice.

AIMS: We studied the effect of metformin on the brown adipose tissue (BAT) in a fructose-rich-fed model, focusing on BAT proliferation, differentiation, and thermogenic markers. MAIN METHODS: C57Bl/6 mice received isoenergetic diets for ten weeks: control (C) or high-fructose (F). For additional eight weeks, animals received metformin hydrochloride (M, 250 mg/kg/day) or saline. After sacrifice, BAT and white fat pads were prepared for light microscopy and molecular analyses. KEY FINDINGS: Body mass gain, white fat pads, and adiposity index were not different among the groups. There was a reduction in energy intake in the F group and energy expenditure in the F and FM groups. Metformin led to a more massive BAT in both groups CM and FM, associated with a higher adipocyte proliferation (ß1-adrenergic receptor, proliferating cell nuclear antigen, and vascular endothelial growth factor), and differentiation (PR domain containing 16, bone morphogenetic protein 7), in part by activating 5' adenosine monophosphate-activated protein kinase. Metformin also enhanced thermogenic markers in the BAT (uncoupling protein type 1, peroxisome proliferator-activated receptor gamma coactivator-1 alpha) through adrenergic stimuli and fibroblast growth factor 21. Metformin might improve mitochondrial biogenesis in the BAT (nuclear respiratory factor 1, mitochondrial transcription factor A), lipolysis (perilipin, adipose triglyceride lipase, hormone-sensitive lipase), and fatty acid uptake (lipoprotein lipase, cluster of differentiation 36, adipocyte protein 2). SIGNIFICANCE: Metformin effects are not linked to body mass changes, but affect BAT thermogenesis, mitochondrial biogenesis, and fatty acid uptake. Therefore, BAT may be a metformin adjuvant target for the treatment of metabolic disorders.

Weight loss enhances hepatic antioxidant status in a NAFLD model induced by high-fat diet.

Nonalcoholic fatty liver disease (NAFLD) is a benign condition that can progress to more severe liver damage in a process mediated, in part, by disturbances in redox balance. Additionally, some argue that it is set to become the main cause of end-stage liver disease in the near future. Here, we investigated whether diet-induced weight loss is able to reverse hepatic lipid accumulation and reduce oxidative stress in liver from C57BL/6 mice fed a high-fat (HF) diet. Male C57BL/6 mice were divided into 4 groups: standard chow (SC; 10% energy from fat, 16 weeks); HF (50% energy from fat, 16 weeks); SC-HF (SC for 8 weeks followed by HF for 8 weeks); and HF-SC (HF for 8 weeks followed by SC for 8 weeks). The HF diet during 8 (SC-HF) and 16 weeks (HF) downregulated messenger RNA levels and protein expression of Nrf2 and endogenous antioxidant enzymes (superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase) in the liver; caused liver steatosis; affected liver function markers; increased intra-abdominal and subcutaneous adipose tissue; and induced glucose intolerance and hypercholesterolemia compared with controls (SC). Diet-induced weight loss significantly reduced the intrahepatic lipid accumulation, improved glucose tolerance, and restored both gene and protein expression of the antioxidant enzymes. Our findings suggest that a dietary intervention aimed to induce weight loss may exert protective effects in NAFLD as it can reduce hepatic oxidative stress and intrahepatic lipid accumulation, which can hinder the progression of this condition to more severe states.

Eicosapentaenoic acid (EPA) vs. Docosahexaenoic acid (DHA): Effects in epididymal white adipose tissue of mice fed a high-fructose diet.

BACKGROUND: Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been demonstrated to be beneficial for many diseases, including those associated with the metabolic syndrome (e.g. insulin resistance and hypertension). Nevertheless, not only their actions are not entirely understood, but also their only effects were not yet elucidated. Therefore, we aimed to compare the effects of EPA and DHA, alone or in combination, on the epididymal white adipose tissue (WAT) metabolism in mice fed a high-fructose diet. METHODS: 3-mo-old C57Bl/6 mice were fed a control diet (C) or a high-fructose diet (HFru). After three weeks on the diets, the HFru group was subdivided into four new groups for another five weeks: HFru, HFru+EPA, HFru+DHA, and HFru-EPA+DHA (n=10/group). Besides evaluating biometric and metabolic parameters of the animals, we measured the adipocyte area and performed molecular analyses (inflammation and lipolysis) in the epididymal WAT. RESULTS: The HFru group showed adipocyte hypertrophy, inflammation, and uncontrolled lipolysis. The treated animals showed a reversion of adipocyte hypertrophy, inhibition of inflammation with activation of anti-inflammatory mediators, and regularization of lipolysis. Overall, the beneficial effects were more marked with DHA than EPA. CONCLUSION: Although the whole-body metabolic effects were similar between EPA and DHA, DHA appeared to be the central actor in WAT metabolism, modulating pro and anti-inflammatory pathways and alleviating adipocytes abnormalities. Therefore, when considering fructose-induced adverse effects in WAT, the most prominent actions were observed with DHA.

Lean vs. obese mice: the ventral prostate revisited / Ratones magros vs. ratones obesos: revisión de la próstata ventral

Obese mice (C57BL/6J-ob/ob) do not express leptin and develops hyperphagia, decreased energy expenditure, obesity, hyperglycemia, hyperinsulinemia, hypothermia, and infertility. Obesity causes reproductive dysfunction with negative impacts on prostatic structure and fertility. We aimed to compare the structure and molecular aspects of the ventral prostate between of lean and obese (ob/ob) mice. Three months old male lean and obese mice had their prostates dissected and prepared for light microscopy and immunofluorescence. In comparison to the lean mouse, the obese mouse showed a substantial structural modification in the ventral prostate starting with an atrophy of the prostate ventral lobe. Histologically, the acini showed a reduction in size, and in the lumen, we found a mixed secretion PAS positive and negative. Epithelial changes consisted of a hypertrophied acinar epithelium with intraepithelial neoplasia focuses. Also, we observed a marked expression of PCNA and Caspase3 in the epithelium indicating even cellular proliferation as cell death. The stroma showed a high activity of the extracellular matrix remodeling with marked deposition of collagen fibers and smooth muscle cells. Around the ventral region, we observed an increase in the presence of adipose tissue. The expressions of interleukin 6 and tumor necrosis factor alpha were present in the ventral prostate of the obese mice indicating inflammation. In conclusion, obesity negatively modulates prostate in ob/ob mice, directly affecting cellular and structural mechanisms necessary for the maintenance of prostate and reproductive structure.

Los ratones obesos (C57BL / 6J-ob / ob) no expresan leptina y desarrollan hiperfagia, disminución del gasto energético, obesidad, hiperglucemia, hiperinsulinemia, hipotermia e infertilidad. La obesidad causa disfunción reproductiva con impacto negativo sobre la estructura prostática y la fertilidad. El objetivo de nuestro trabajo consistió en comparar la estructura y los aspectos moleculares de la próstata ventral en ratones magros y obesos (ob/ob). Se disecaron las próstatas de ratones machos obesos, de tres meses de edad, y fueron preparadas para visualizarlas por microscopía óptica e inmunofluorescencia. En comparación con el ratón magro, el ratón obeso mostró una sustancial modificación estructural en la próstata ventral comenzando con una atrofia del lóbulo ventral de la próstata. Histológicamente, los acinos mostraron una reducción de tamaño, y en el lumen, encontramos una secreción mixta PAS positiva y negativa. Los cambios epiteliales consistieron en un epitelio acinar hipertrofiado con focos de neoplasia intraepitelial. Además, se observó una marcada expresión de PCNA y Caspase3 en el epitelio, que indica tanto la proliferación celular como la muerte celular. El estroma mostró una alta remodelación de la matriz extracelular con marcada deposición de fibras de colágeno y células de músculo liso. Alrededor de la región ventral, se observó un aumento en la presencia de tejido adiposo. Las expresiones de interleuquina 6 y factor de necrosis tumoral alfa estaban presentes en la próstata ventral de los ratones obesos indicando inflamación. En conclusión, la obesidad modula negativamente la próstata en los ratones ob/ob, afectando directamente los mecanismos celulares y estructurales necesarios para el mantenimiento de la estructura de la próstata y la reproducción.