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The present study aimed to elucidate how Atlantic salmon adipocytes pre-enriched with palmitic (16:0, PA), oleic (18:1 n -9, OA), or eicosapentaenoic (20:5 n -3, EPA) acid respond to a fasting condition mimicked by nutrient deprivation and glucagon. All experimental groups were supplemented with radiolabeled PA to trace secreted lipids and distribution of radioactivity in different lipid classes. There was a higher content of intracellular lipid droplets in adipocytes pre-enriched with OA than in adipocytes pre-enriched with PA or EPA. In the EPA group, the radiolabeled PA was mainly esterified in phospholipids and triacylglycerols, whereas in the OA and PA groups, the radioactivity was mainly recovered in phospholipids and cholesterol-ester. By subjecting the experimental groups to nutrient-deprived media supplemented with glucagon, lipolysis occurred in all groups, although to a lower extent in the OA group. The lipids were mainly secreted as esterified lipids in triacylglycerols and phospholipids, indicating mobilization in lipoproteins. A significant proportion was secreted as free fatty acids and glycerol. Leptin secretion was reduced in all experimental groups in response to fasting, while the mitochondria area responded to changes in the energy supply and demand by increasing after 3 h of fasting. Overall, different lipid classes in adipocytes influenced their mobilization during fasting.

Background/objective: N 6 -methyladenosine (m 6 A) modification of mRNA plays an important role in regulating adipogenesis. However, its underlying mechanism remains largely unknown., Subjects/methods: Using Jinhua and Landrace pigs as fat and lean models, we presented a comprehensive transcriptome-wide m 6 A profiling in adipose tissues from these two pig breeds. Two differentially methylated genes were selected to explore the mechanisms of m 6 A-mediated regulation of gene function., Results: The ratio of m 6 A/A in the layer of backfat (LB) was significantly higher in Landrace than that in Jinhua. Transcriptome-wide m 6 A profiling revealed that m 6 A modification on mRNA occurs in the conserved sequence motif of RRACH and that the pig transcriptome contains 0.53-0.91 peak per actively expressed transcript. The relative density of m 6 A peaks in the 3'UTR were higher than in 5'UTR. Genes with common m 6 A peaks from both Landrace (L-LB) and Jinhua (J-LB) were enriched in RNA splicing and cellular lipid metabolic process. The unique m 6 A peak genes (UMGs) from L-LB were mainly enriched in the extracellular matrix (ECM) and collagen catabolic process, whereas the UMGs from J-LB are mainly involved in RNA splicing, etc. Lipid metabolism processes were not significantly enriched in the UMGs from L-LB or J-LB. Uncoupling protein-2 (UCP2) and patatin-like phospholipase domain containing 2 (PNPLA2) were two of the UMGs in L-LB. Synonymous mutations (MUT) were conducted to reduce m 6 A level of UCP2 and PNPLA2 mRNAs. Adipogenesis test showed that UCP2-MUT further inhibited adipogenesis, while PNPLA2-MUT promoted lipid accumulation compared with UCP2-WT and PNPLA2-WT, respectively. Further study showed m 6 A negatively mediates UCP2 protein expression and positively mediates PNPLA2 protein expression. m 6 A modification affects the translation of PNPLA2 most likely through YTHDF1, whereas UCP2 is likely neither the target of YTHDF2 nor the target of YTHDF1., Conclusion: Our data demonstrated a conserved and yet dynamically regulated m 6 A methylome in pig transcriptomes and provided an important resource for studying the function of m 6 A epitranscriptomic modification in obesity development.

Betaine, a neutral zwitterionic compound, could regulate intramuscular fat (IMF) deposition and meat quality. However, the efficacy is controversial. Moreover, the regulatory mechanism of betaine on lipid metabolism in skeletal muscle cells remains unclear. Therefore, in this study, we examined the effects and regulatory mechanism of betaine on lipid accumulation in adipogenic-differentiated C2C12 cells. We found that adipogenic-induced C2C12 cells treated with 10 mM betaine for 24 and 48 h had more lipid accumulation than the control group. Real-time PCR and Western blot results revealed that betaine treatment did not alter the expression of lipolysis and lipid oxidation-related genes, but dramatically increased the expression of peroxisome proliferator-activated receptor γ (PPARγ) and its target genes such as fatty acid binding protein 4 (aP2), fatty acid synthase (FAS) and lipoprteinlipase (LPL). Furthermore, betaine combined with PPARγ inhibitor GW9662 treatment showed that betaine elevated C2C12 lipid accumulation through upregulation of PPARγ. Mechanistically, we found that betaine promoted PPARγ expression and lipid accumulation through inhibition of extracellular regulated protein kinases1/2 (ERK1/2) signalling pathway. These results demonstrate that betaine acts through ERK1/2-PPARγ signalling pathway to regulate lipid metabolism in adipogenic-differentiated skeletal muscle cells, which could provide some useful information for controlling muscle lipid accumulation by manipulating ERK1/2 and PPARγ signalling pathway.

Excessive intramyocellular triacylglycerols (IMTGs, muscle lipids) are associated with the abnormal energy metabolism and insulin resistance of skeletal muscle. AMP-activated protein kinase (AMPK), a crucial cellular energy sensor, consists of α, β and γ subunits. Researchers have not clearly determined whether Prkaa1 (also known as AMPKα1) affects IMTG accumulation in skeletal muscle. Here, we show an important role of Prkaa1 in skeletal muscle lipid metabolism. Deletion of muscle Prkaa1 leads to the delayed development of skeletal muscles but does not affect glucose tolerance or insulin sensitivity in animals fed a normal diet. Notably, when animals are fed a high-fat diet, the skeletal muscle of muscle-specific Prkaa1 knockout mice accumulates more lipids than the skeletal muscle of wild-type (WT) mice, with concomitant upregulation of adipogenic gene expressions and downregulation of the expression of genes associated with mitochondrial oxidation. Muscle-specific Prkaa1 ablation also results in hyperlipidemia, which may contribute to the increased IMTG levels. Furthermore, Prkaa1 deletion activates skeletal muscle mTOR signalling, which has a central role in lipid metabolism and mitochondrial oxidation. Collectively, our study provides new insights into the role of Prkaa1 in skeletal muscle. This knowledge may contribute to the treatment of related metabolic diseases.

Dietary fatty acids have different effects on fat deposition in pigs. To clarify the underlying mechanisms of this difference, we compared the metabolism of palmitic (PAM, saturated), oleic (OLA, monounsaturated) and linoleic acid (LNA, polyunsaturated) in porcine adipocytes treated with 100 μM PAM, OLA or LNA. We observed that the adipocytes incubated with LNA accumulated more lipids compared with those treated with PAM and OLA. We then probed the metabolism of these fatty acids in porcine adipocytes by using isotope-labelled fatty acids. The results showed that 42% of the [1- 14 C] LNA, 34% of the [1- 14 C] PAM and 28% of the [1- 14 C] OLA were recovered in the cellular lipids. The gene expression analyses showed that LNA significantly increased the expression of adipogenesis- and oxidation-related genes including PPARγ, C/EBPα, ap2 and NRF1. In addition, the cells incubated with LNA showed a decreased Ser 112 phosphorylation in PPARγ compared to those incubated with PAM and OLA. Furthermore, when PPARγ Ser 112 phosphorylation was inhibited, no significant difference in the triacylglycerol contents in the adipocytes was observed. These results showed the dietary fatty acids had different metabolism pathways in porcine adipocytes, and LNA significantly promoted lipid accumulation, probably by regulating PPARγ phosphorylation in adipocytes.

Skeletal muscle plays important roles in whole-body energy homeostasis. Excessive skeletal muscle lipid accumulation is associated with some metabolic diseases such as obesity and Type 2 Diabetes. The energy sensor AMPK (AMP-activated protein kinase) is a key regulator of skeletal muscle lipid metabolism, but the precise regulatory mechanism remains to be elucidated. Here, we provide a novel mechanism by which AMPK regulates skeletal muscle lipid accumulation through fat mass and obesity-associated protein (FTO)-dependent demethylation of N 6 -methyladenosine (m 6 A). We confirmed an inverse correlation between AMPK and skeletal muscle lipid content. Moreover, inhibition of AMPK enhanced lipid accumulation, while activation of AMPK reduced lipid accumulation in skeletal muscle cells. Notably, we found that mRNA m 6 A methylation levels were inversely correlated with lipid content in skeletal muscle. Furthermore, AMPK positively regulated the m 6 A methylation levels of mRNA, which could negatively regulate lipid accumulation in C2C12. At the molecular level, we demonstrated that AMPK regulated lipid accumulation in skeletal muscle cells by regulating FTO expression and FTO-dependent demethylation of m 6 A. Together, these results provide a novel regulatory mechanism of AMPK on lipid metabolism in skeletal muscle cells and suggest the possibility of controlling skeletal muscle lipid deposition by targeting AMPK or using m 6 A related drugs., Competing Interests: The authors declare no competing financial interests.

n-3 Long-chain PUFA up-regulate intestinal lipid metabolism. However, whether these metabolic effects of PUFA on intestine are mediated by AMP-activated protein kinase (AMPK) remains to be elucidated. To determine the effects of α-linolenic acid (ALA) on intestinal fatty acid (FA) metabolism and whether these effects were affected by AMPK deletion, mice deficient in the catalytic subunit of AMPKα1 or AMPKα2 and wild-type (WT) mice were fed either a high-fat diet (HF) or HF supplemented with ALA (HF-A). The results showed that ALA supplementation decreased serum TAG content in WT mice. ALA also increased mRNA expression of genes (carnitine palmitoyltransferase 1a, acyl-CoA oxidase 1, medium-chain acyl-CoA dehydrogenase, cytochrome P450 4A10 and pyruvate dehydrogenase kinase isoenzyme 4a) involved in intestinal lipid oxidation and mRNA expression of TAG synthesis-related genes (monoacylglycerol O-acyltransferase 2, diacylglycerol O-acyltransferases 1 and 2) in WT mice. Consistent with these, expression levels of phosphorylated AMPKα1 and AMPKα2 were also increased in WT mice after ALA addition. However, in the absence of either AMPKα1 or AMPKα2, ALA supplementation failed to increase intestinal lipid oxidation. In addition, no significant effects of either diet (HF and HF-A) or genotype (WT, AMPKα1(-/-) and AMPKα2(-/-)) on FA uptake in the intestine and faecal TAG output were observed. Our results suggest that AMPK is indispensable for the effects of ALA on intestinal lipid oxidation.

Dysregulation of lipid metabolism has been believed to be central in the development of skeletal muscle insulin resistance. Since first being described in 1989, the role of AMPK in energy metabolism, especially its role in lipid metabolism in skeletal muscle has been well studied. However, some recent literature report that fatty acid oxidation in skeletal muscle is not directly associated with AMPK activation and ACC phosphorylation. To further understand the role of AMPK in lipid metabolism and the development of induced obesity and insulin resistance, muscle specific AMPKα2 knockout mice (mAMPKα2-KO) was employed in this study. The results showed that AMPKα2 ablation in muscle did not exacerbate high fat diet induce obesity in mice. On the contrary, it improved animal glucose tolerance and insulin sensitivity, with reduced triglyceride content in skeletal muscle and fat mass in various adipose tissues, when mice were fed high fat diet for 14 weeks. Gene expression analysis revealed that AMPKα2 knockout up-regulated the expression of genes related to lipid catabolism and down-regulated that of genes related to triglyceride synthesis. More importantly, ablation of AMPKα2 altered the expression of several myokines related to adipogenesis and muscle regeneration. Our data suggest that defect in AMPKα2 signaling does not necessarily lead to the development of muscle insulin resistance and obesity. AMPKα2 may regulate whole body lipid metabolism by regulating myokine secretion., (Copyright © 2015. Published by Elsevier Inc.)

N6‐methyladenosine (m6A), as the most abundant RNA epigenetic modifications, has been shown to play critical roles in various biological functions. Research about enzymes that can catalyze and remove m6A have revealed its comprehensive roles in messenger RNA (mRNA) metabolism and other physiological processes. The "readers" including YTH domain‐containing proteins, hnRNPC, hnRNPG, hnRNPA2B1, IGF2BP1, IGF2BP2, and IGF2BP3, which can affect the fates of mRNA in an m6A‐dependent manner. In this review, we focus on recent advances in the research of the m6A modifications, especially about the latest functions of its writers, erasers, readers in RNA metabolism, cancer, and lipid metabolism. In the end, we provide insights into the underlying molecular mechanisms of m6A modifications. In this review, we focus on recent advances in the research of the m6A modifications, especially about the latest functions of its writers, erasers, readers in RNA metabolism, cancer, and lipid metabolism. In the end, we provide insights into the underlying molecular mechanisms of m6A modifications. [ABSTRACT FROM AUTHOR]