Comparison of microRNAs in adipose and muscle tissue from seven indigenous Chinese breeds and Yorkshire pigs

Elucidation of the pig microRNAome is essential for interpreting functional elements of the genome and understanding the genetic architecture of complex traits. Here, we extracted small RNAs from skeletal muscle and adipose tissue, and we compared their expression levels between one Western breed (Yorkshire) and seven indigenous Chinese breeds. We detected the expression of 172 known porcine microRNAs (miRNAs) and 181 novel miRNAs. Differential expression analysis found 92 and 12 differentially expressed miRNAs in adipose and muscle tissue respectively. We found that different Chinese breeds shared common directional miRNA expression changes compared to Yorkshire pigs. Some miRNAs differentially expressed across multiple Chinese breeds, including ssc‐miR‐129‐5p, ssc‐miR‐30 and ssc‐miR‐150, are involved in adipose tissue function. Functional enrichment analysis revealed that the target genes of the differentially expressed miRNAs are associated mainly with signaling pathways rather than metabolic and biosynthetic processes. The miRNA–target gene and miRNA–phenotypic traits networks identified many hub miRNAs that regulate a large number of target genes or phenotypic traits. Specifically, we found that intramuscular fat content is regulated by the greatest number of miRNAs in muscle tissue. This study provides valuable new candidate miRNAs that will aid in the improvement of meat quality and production.     

Metformin inhibits mitochondrial adaptations to aerobic exercise training in older adults

Metformin and exercise independently improve insulin sensitivity and decrease the risk of diabetes. Metformin was also recently proposed as a potential therapy to slow aging. However, recent evidence indicates that adding metformin to exercise antagonizes the exercise‐induced improvement in insulin sensitivity and cardiorespiratory fitness. The purpose of this study was to test the hypothesis that metformin diminishes the improvement in insulin sensitivity and cardiorespiratory fitness after aerobic exercise training (AET) by inhibiting skeletal muscle mitochondrial respiration and protein synthesis in older adults (62 ± 1 years). In a double‐blinded fashion, participants were randomized to placebo (n = 26) or metformin (n = 27) treatment during 12 weeks of AET. Independent of treatment, AET decreased fat mass, HbA1c, fasting plasma insulin, 24‐hr ambulant mean glucose, and glycemic variability. However, metformin attenuated the increase in whole‐body insulin sensitivity and VO₂max after AET. In the metformin group, there was no overall change in whole‐body insulin sensitivity after AET due to positive and negative responders. Metformin also abrogated the exercise‐mediated increase in skeletal muscle mitochondrial respiration. The change in whole‐body insulin sensitivity was correlated to the change in mitochondrial respiration. Mitochondrial protein synthesis rates assessed during AET were not different between treatments. The influence of metformin on AET‐induced improvements in physiological function was highly variable and associated with the effect of metformin on the mitochondria. These data suggest that prior to prescribing metformin to slow aging, additional studies are needed to understand the mechanisms that elicit positive and negative responses to metformin with and without exercise.     

CB1 receptor blockade counters age‐induced insulin resistance and metabolic dysfunction

The endocannabinoid system can modulate energy homeostasis by regulating feeding behaviour as well as peripheral energy storage and utilization. Importantly, many of its metabolic actions are mediated through the cannabinoid type 1 receptor (CB1R), whose hyperactivation is associated with obesity and impaired metabolic function. Herein, we explored the effects of administering rimonabant, a selective CB1R inverse agonist, upon key metabolic parameters in young (4 month old) and aged (17 month old) adult male C57BL/6 mice. Daily treatment with rimonabant for 14 days transiently reduced food intake in young and aged mice; however, the anorectic response was more profound in aged animals, coinciding with a substantive loss in body fat mass. Notably, reduced insulin sensitivity in aged skeletal muscle and liver concurred with increased CB1R mRNA abundance. Strikingly, rimonabant was shown to improve glucose tolerance and enhance skeletal muscle and liver insulin sensitivity in aged, but not young, adult mice. Moreover, rimonabant‐mediated insulin sensitization in aged adipose tissue coincided with amelioration of low‐grade inflammation and repressed lipogenic gene expression. Collectively, our findings indicate a key role for CB1R in aging‐related insulin resistance and metabolic dysfunction and highlight CB1R blockade as a potential strategy for combating metabolic disorders associated with aging.     

Endothelium‐specific CYP2J2 overexpression attenuates age‐related insulin resistance

Ample evidences demonstrate that cytochrome P450 epoxygenase‐derived epoxyeicosatrienoic acids (EETs) exert diverse biological activities, which include potent vasodilatory, anti‐inflammatory, and cardiovascular protective effects. In this study, we investigated the effects of endothelium‐specific CYP2J2 overexpression on age‐related insulin resistance and metabolic dysfunction. Endothelium‐specific targeting of the human CYP epoxygenase, CYP2J2, transgenic mice (Tie2‐CYP2J2‐Tr mice) was utilized. The effects of endothelium‐specific CYP2J2 overexpression on aging‐associated obesity, inflammation, and peripheral insulin resistance were evaluated by assessing metabolic parameters in young (3 months old) and aged (16 months old) adult male Tie2‐CYP2J2‐Tr mice. Decreased insulin sensitivity and attenuated insulin signaling in aged skeletal muscle, adipose tissue, and liver were observed in aged adult male mice, and moreover, these effects were partly inhibited in 16‐month‐old CYP2J2‐Tr mice. In addition, CYP2J2 overexpression‐mediated insulin sensitization in aged mice was associated with the amelioration of inflammatory state. Notably, the aging‐associated increases in fat mass and adipocyte size were only observed in 16‐month‐old wild‐type mice, and CYP2J2 overexpression markedly prevented the increase in fat mass and adipocyte size in aged Tie2‐CYP2J2‐Tr mice, which was associated with increased energy expenditure and decreased lipogenic genes expression. Furthermore, these antiaging phenotypes of Tie2‐CYP2J2‐Tr mice were also associated with increased muscle blood flow, enhanced active‐phase locomotor activity, and improved mitochondrial dysfunction in skeletal muscle. Collectively, our findings indicated that endothelium‐specific CYP2J2 overexpression alleviated age‐related insulin resistance and metabolic dysfunction, which highlighted CYP epoxygenase‐EET system as a potential target for combating aging‐related metabolic disorders.     

Aging and caloric restriction impact adipose tissue,adiponectin, and circulating lipids

Adipose tissue expansion has been associated with system‐wide metabolic dysfunction and increased vulnerability to diabetes, cancer, and cardiovascular disease. A reduction in adiposity is a hallmark of caloric restriction (CR), an intervention that extends longevity and delays the onset of these same age‐related conditions. Despite these parallels, the role of adipose tissue in coordinating the metabolism of aging is poorly defined. Here, we show that adipose tissue metabolism and secretory profiles change with age and are responsive to CR. We conducted a cross‐sectional study of CR in adult, late‐middle‐aged, and advanced‐aged mice. Adiposity and the relationship between adiposity and circulating levels of the adipose‐derived peptide hormone adiponectin were age‐sensitive. CR impacted adiposity but only levels of the high molecular weight isoform of adiponectin responded to CR. Activators of metabolism including PGC‐1a, SIRT1, and NAMPT were differentially expressed with CR in adipose tissues. Although age had a significant impact on NAD metabolism, as detected by biochemical assay and multiphoton imaging, the impact of CR was subtle and related to differences in reliance on oxidative metabolism. The impact of age on circulating lipids was limited to composition of circulating phospholipids. In contrast, the impact of CR was detected in all lipid classes regardless of age, suggesting a profound difference in lipid metabolism. These data demonstrate that aspects of adipose tissue metabolism are life phase specific and that CR is associated with a distinct metabolic state, suggesting that adipose tissue signaling presents a suitable target for interventions to delay aging.     

Metformin blunts muscle hypertrophy in response to progressive resistance exercise training in older adults: A randomized,double‐blind,placebo‐controlled,multicenter trial: The MASTERS trial

Progressive resistance exercise training (PRT) is the most effective known intervention for combating aging skeletal muscle atrophy. However, the hypertrophic response to PRT is variable, and this may be due to muscle inflammation susceptibility. Metformin reduces inflammation, so we hypothesized that metformin would augment the muscle response to PRT in healthy women and men aged 65 and older. In a randomized, double‐blind trial, participants received 1,700 mg/day metformin (N = 46) or placebo (N = 48) throughout the study, and all subjects performed 14 weeks of supervised PRT. Although responses to PRT varied, placebo gained more lean body mass (p = .003) and thigh muscle mass (p < .001) than metformin. CT scan showed that increases in thigh muscle area (p = .005) and density (p = .020) were greater in placebo versus metformin. There was a trend for blunted strength gains in metformin that did not reach statistical significance. Analyses of vastus lateralis muscle biopsies showed that metformin did not affect fiber hypertrophy, or increases in satellite cell or macrophage abundance with PRT. However, placebo had decreased type I fiber percentage while metformin did not (p = .007). Metformin led to an increase in AMPK signaling, and a trend for blunted increases in mTORC1 signaling in response to PRT. These results underscore the benefits of PRT in older adults, but metformin negatively impacts the hypertrophic response to resistance training in healthy older individuals. ClinicalTrials.gov Identifier: NCT02308228.     

Muscle ectopic fat deposition contributes to anabolic resistance in obese sarcopenic old rats through eIF2α activation

Obesity and aging are characterized by decreased insulin sensitivity (IS) and muscle protein synthesis. Intramuscular ceramide accumulation has been implicated in insulin resistance during obesity. We aimed to measure IS, muscle ceramide level, protein synthesis, and activation of intracellular signaling pathways involved in translation initiation in male Wistar young (YR, 6‐month) and old (OR, 25‐month) rats receiving a low‐ (LFD) or a high‐fat diet (HFD) for 10 weeks. A corresponding cellular approach using C2C12 myotubes treated with palmitate to induce intracellular ceramide deposition was taken. A decreased ability of adipose tissue to store lipids together with a reduced adipocyte diameter and a development of fibrosis were observed in OR after the HFD. Consequently, OR fed the HFD were insulin resistant, showed a strong increase in intramuscular ceramide level and a decrease in muscle protein synthesis associated with increased eIF2α phosphorylation. The accumulation of intramuscular lipids placed a lipid burden on mitochondria and created a disconnect between metabolic and regulating pathways in skeletal muscles of OR. In C2C12 cells, palmitate‐induced ceramide accumulation was associated with a decreased protein synthesis together with upregulated eIF2α phosphorylation. In conclusion, a reduced ability to expand adipose tissues was found in OR, reflecting a lower lipid buffering capacity. Muscle mitochondrial activity was affected in OR conferring a reduced ability to oxidize fatty acids entering the muscle cell. Hence, OR were more prone to ectopic muscle lipid accumulation than YR, leading to decreased muscle protein anabolism. This metabolic change is a potential therapeutic target to counter sarcopenic obesity.     

Aging leads to a programmed loss of brown adipocytes in murine subcutaneous white adipose tissue

Insulin sensitivity deteriorates with age, but mechanisms remain unclear. Age‐related changes in the function of subcutaneous white adipose tissue (sWAT) are less characterized than those in visceral WAT. We hypothesized that metabolic alterations in sWAT, which in contrast to epididymal WAT, harbors a subpopulation of energy‐dissipating UCP1+ brown adipocytes, promote age‐dependent progression toward insulin resistance. Indeed, we show that a predominant consequence of aging in murine sWAT is loss of ‘browning’. sWAT from young mice is histologically similar to brown adipose tissue (multilocular, UCP1+), but becomes morphologically white by 12 months of age. Correspondingly, sWAT expression of ucp1 precipitously declines (~300‐fold) between 3 and 12 months. Loss continues into old age (24 months) and is inversely correlated with the development of insulin resistance. Additional age‐dependent changes in sWAT include lower expression of adbr3 and higher expression of maoa, suggesting reduced local adrenergic tone as a potential mechanism. Indeed, treatment with a β3‐adrenergic agonist to compensate for reduced tone rescues the aged sWAT phenotype. Age‐related changes in sWAT are not explained by the differences in body weight; mice subjected to 40% caloric restriction for 12 months are of body weight similar to 3‐month‐old ad lib fed mice, but display sWAT resembling that of age‐matched ad lib fed mice (devoid of brown adipose‐like morphology). Overall, findings identify the loss of ‘browning’ in sWAT as a new aging phenomenon and provide insight into the pathogenesis of age‐associated metabolic disease by revealing novel molecular changes tied to systemic metabolic dysfunction.     

Adipose tissue aging: mechanisms and therapeutic implications

Adipose tissue, which is the crucial energy reservoir and endocrine organ for the maintenance of systemic glucose, lipid, and energy homeostasis, undergoes significant changes during aging. These changes cause physiological declines and age-related disease in the elderly population. Here, we review the age-related changes in adipose tissue at multiple levels and highlight the underlying mechanisms regulating the aging process. We also discuss the pathogenic pathways of age-related fat dysfunctions and their systemic negative consequences, such as dyslipidemia, chronic general inflammation, insulin resistance, and type 2 diabetes (T2D). Age-related changes in adipose tissue involve redistribution of deposits and composition, in parallel with the functional decline of adipocyte progenitors and accumulation of senescent cells. Multiple pathogenic pathways induce defective adipogenesis, inflammation, aberrant adipocytokine production, and insulin resistance, leading to adipose tissue dysfunction. Changes in gene expression and extracellular signaling molecules regulate the aging process of adipose tissue through various pathways. In addition, adipose tissue aging impacts other organs that are infiltrated by lipids, which leads to systemic inflammation, metabolic system disruption, and aging process acceleration. Moreover, studies have indicated that adipose aging is an early onset event in aging and a potential target to extend lifespan. Together, we suggest that adipose tissue plays a key role in the aging process and is a therapeutic target for the treatment of age-related disease, which deserves further study to advance relevant knowledge.Subject terms: Senescence, Endocrine system and metabolic diseases     

Glucose homeostasis in rainbow trout fed a high-carbohydrate diet: metformin and insulin interact in a tissue-dependent manner

Carnivorous fish species such as the rainbow trout (Oncorhynchus mykiss) are considered to be "glucose intolerant" because of the prolonged hyperglycemia experienced after intake of a carbohydrate-enriched meal. In the present study, we use this species to study glucose homeostasis in fish chronically infused with the hypoglycemic agents, insulin, and metformin, and fed with a high proportion of carbohydrates (30%). We analyzed liver, skeletal muscle, and white adipose tissue (WAT), which are insulin- and metformin-specific targets at both the biochemical and molecular levels. Trout infused with the combination of insulin and metformin can effectively utilize dietary glucose at the liver, resulting in lowered glycemia, increased insulin sensitivity, and glucose storage capacity, combined with reduced glucose output. However, in both WAT and skeletal muscle, we observed decreased insulin sensitivity with the combined insulin + metformin treatment, resulting in the absence of changes at the metabolic level in the skeletal muscle and an increased potential for glucose uptake and storage in the WAT. Thus, the poor utilization by rainbow trout of a diet with a high proportion of carbohydrate can at least be partially improved by a combined treatment with insulin and metformin, and the glucose intolerance observed in this species could be, in part, due to some of the downstream components of the insulin and metformin signaling pathways. However, the predominant effects of metformin treatment on the action of insulin in these three tissues thought to be involved in glucose homeostasis remain exclusive in this species.     

Long‐lived hypopituitary Ames dwarf mice are resistant to the detrimental effects of high‐fat diet on metabolic function and energy expenditure

Growth hormone (GH) signaling stimulates the production of IGF‐1; however, increased GH signaling may induce insulin resistance and can reduce life expectancy in both mice and humans. Interestingly, disruption of GH signaling by reducing plasma GH levels significantly improves health span and extends lifespan in mice, as observed in Ames dwarf mice. In addition, these mice have increased adiposity, yet are more insulin sensitive compared to control mice. Metabolic stressors such as high‐fat diet (HFD) promote obesity and may alter longevity through the GH signaling pathway. Therefore, our objective was to investigate the effects of a HFD (metabolic stressor) on genetic mechanisms that regulate metabolism during aging. We show that Ames dwarf mice fed HFD for 12 weeks had an increase in subcutaneous and visceral adiposity as a result of diet‐induced obesity, yet are more insulin sensitive and have higher levels of adiponectin compared to control mice fed HFD. Furthermore, energy expenditure was higher in Ames dwarf mice fed HFD than in control mice fed HFD. Additionally, we show that transplant of epididymal white adipose tissue (eWAT) from Ames dwarf mice fed HFD into control mice fed HFD improves their insulin sensitivity. We conclude that Ames dwarf mice are resistant to the detrimental metabolic effects of HFD and that visceral adipose tissue of Ames dwarf mice improves insulin sensitivity in control mice fed HFD.     

Diabetogenic diet-induced insulin resistance associates with lipid droplet proteins and adipose tissue secretome,but not with sexual dimorphic adipose tissue fat accumulation in Wistar rats

The role of sexual dimorphic adipose tissue fat accumulation in the development of insulin resistance is well known. However, whether vitamin A status and/or its metabolic pathway display any sex- or depot (visceral/subcutaneous)-specific pattern and have a role in sexual dimorphic adipose tissue development and insulin resistance are not completely understood. Therefore, to assess this, 5 weeks old Wistar male and female rats of eight from each sex were provided either control or diabetogenic (high fat, high sucrose) diet for 26 weeks. At the end, consumption of diabetogenic diet increased the visceral fat depots (p < 0.001) in the males and subcutaneous depot (p < 0.05) in the female rats, compared to their sex-matched controls. On the other hand, it caused adipocyte hypertrophy (p < 0.05) of visceral depot (retroperitoneal) in the females and subcutaneous depot of the male rats. Although vitamin A levels displayed sex- and depot-specific increase due to the consumption of diabetogenic diet, the expression of most of its metabolic pathway genes in adipose depots remained unaltered. However, the mRNA levels of some of lipid droplet proteins (perilipins) and adipose tissue secretory proteins (interleukins, lipocalin-2) did display sexual dimorphism. Nonetheless, the long-term feeding of diabetogenic diet impaired the insulin sensitivity, thus affected glucose clearance rate and muscle glucose-uptake in both the sexes of rats. In conclusion, the chronic consumption of diabetogenic diet caused insulin resistance in the male and female rats, but did not corroborate with sexual dimorphic adipose tissue fat accumulation or its vitamin A status.     

Long‐term caloric restriction ameliorates deleterious effects of aging on white and brown adipose tissue plasticity

Age‐related increased adiposity is an important contributory factor in the development of insulin resistance (IR) and is associated with metabolic defects. Caloric restriction (CR) is known to induce weight loss and to decrease adiposity while preventing metabolic risk factors. Here, we show that moderate 20% CR delays early deleterious effects of aging on white and brown adipose tissue (WAT and BAT, respectively) function and improves peripheral IR. To elucidate the role of CR in delaying early signs of aging, young (3 months), middle‐aged (12 months), and old (20 months) mice fed al libitum and middle‐aged and old mice subjected to early‐onset CR were used. We show that impaired plasticity of subcutaneous WAT (scWAT) contributes to IR, which is already evident in middle‐aged mice. Moreover, alteration of thyroid axis status with age is an important factor contributing to BAT dysfunction in middle‐aged animals. Both defects in WAT and BAT/beige cells are ameliorated by CR. Accordingly, CR attenuated the age‐related decline in scWAT function and decreased the extent of fibro‐inflammation. Furthermore, CR promoted scWAT browning. In brief, our study identifies the contribution of scWAT impairment to age‐associated metabolic dysfunction and identifies browning in response to food restriction, as a potential therapeutic strategy to prevent the adverse metabolic effects in middle‐aged animals.     

Modulation of white adipose tissue proteome by aging and calorie restriction

Aging is associated with an accrual of body fat, progressive development of insulin resistance and other obesity comorbidities that contribute to decrease life span. Caloric restriction (CR), which primarily affects energy stores in adipose tissue, is known to extend life span and retard the aging process in animal models. In this study, a proteomic approach combining 2‐DE and MS was used to identify proteins modulated by aging and CR in rat white adipose tissue proteome. Proteomic analysis revealed 133 differentially expressed spots, 57 of which were unambiguously identified by MS. Although CR opposed part of the age‐associated protein expression patterns, many effects of CR were on proteins unaltered by age, suggesting that the effects of CR on adipose tissue are only weakly related to those of aging. Particularly, CR and aging altered glucose, intermediate and lipid metabolism, with CR enhancing the expression of enzymes involved in oxalacetate and NADPH production, lipid biosynthesis and lipolysis. Consistently, insulin‐β and β3‐adrenergic receptors were also increased by CR, which denotes improved sensitivity to lipogenic/lipolytic stimuli. Other beneficial outcomes of CR were an improvement in oxidative stress, preventing the age‐associated decrease in several antioxidant enzymes. Proteins involved in cytoskeleton, iron storage, energy metabolism and several proteins with novel or unknown functions in adipose tissue were also modulated by age and/or CR. Such orchestrated changes in expression of multiple proteins provide insights into the mechanism underlying CR effects, ultimately allowing the discovery of new markers of aging and targets for the development of CR‐mimetics.     

Proteomic and bioinformatic analysis of membrane proteome in type 2 diabetic mouse liver

Type 2 diabetes mellitus (T2DM) is the most prevalent and serious metabolic disease affecting people worldwide. T2DM results from insulin resistance of the liver, muscle, and adipose tissue. In this study, we used proteomic and bioinformatic methodologies to identify novel hepatic membrane proteins that are related to the development of hepatic insulin resistance, steatosis, and T2DM. Using FT‐ICR MS, we identified 95 significantly differentially expressed proteins in the membrane fraction of normal and T2DM db/db mouse liver. These proteins are primarily involved in energy metabolism pathways, molecular transport, and cellular signaling, and many of them have not previously been reported in diabetic studies. Bioinformatic analysis revealed that 16 proteins may be related to the regulation of insulin signaling in the liver. In addition, six proteins are associated with energy stress‐induced, nine proteins with inflammatory stress‐induced, and 14 proteins with endoplasmic reticulum stress‐induced hepatic insulin resistance. Moreover, we identified 19 proteins that may regulate hepatic insulin resistance in a c‐Jun amino‐terminal kinase‐dependent manner. In addition, three proteins, 14–3‐3 protein beta (YWHAB), Slc2a4 (GLUT4), and Dlg4 (PSD‐95), are discovered by comprehensive bioinformatic analysis, which have correlations with several proteins identified by proteomics approach. The newly identified proteins in T2DM should provide additional insight into the development and pathophysiology of hepatic steatosis and insulin resistance, and they may serve as useful diagnostic markers and/or therapeutic targets for these diseases.     

Medium‐Chain Oil Reduces Fat Mass and Down‐regulates Expression of Adipogenic Genes in Rats

Objective: To test the hypothesis that adipose tissue could be one of the primary targets through which medium‐chain fatty acids (MCFAs) exert their metabolic influence. Research Methods and Procedures: Sprague‐Dawley rats were fed a control high‐fat diet compared with an isocaloric diet rich in medium‐chain triglycerides (MCTs). We determined the effects of MCTs on body fat mass, plasma leptin and lipid levels, acyl chain composition of adipose triglycerides and phospholipids, adipose tissue lipoprotein lipase activity, and the expression of key adipogenic genes. Tissue triglyceride content was measured in heart and gastrocnemius muscle, and whole body insulin sensitivity and glucose tolerance were also measured. The effects of MCFAs on lipoprotein lipase activity and adipogenic gene expression were also assessed in vitro using cultured adipose tissue explants or 3T3‐L1 adipocytes. Results: MCT‐fed animals had smaller fat pads, and they contained a considerable amount of MCFAs in both triglycerides and phospholipids. A number of key adipogenic genes were down‐regulated, including peroxisome proliferator activated receptor γ and CCAAT/enhancer binding protein α and their downstream metabolic target genes. We also found reduced adipose tissue lipoprotein lipase activity and improved insulin sensitivity and glucose tolerance in MCT‐fed animals. Analogous effects of MCFAs on adipogenic genes were found in cultured rat adipose tissue explants and 3T3‐L1 adipocytes. Discussion: These results suggest that direct inhibitory effects of MCFAs on adiposity may play an important role in the regulation of body fat development.