小蘖碱抑制游离脂肪酸诱导小鼠肝实质细胞脂肪变性

目的:探讨小蘖碱(Berberine)对游离脂肪酸(free fatty acids,FFAs)诱导的小鼠肝实质细胞脂肪变性的影响。方法:胶原酶灌注分离BALB/c小鼠原代肝实质细胞并体外培养。分对照组,高脂组,高脂加小蘖碱处理组。体外测定细胞内甘油三酯的含量。利用油红染色观察细胞的脂肪样变性。通过Western印迹法检测肝实质细胞内MAPK相关信号通路磷酸化的变化。实时定量PCR检测肝实质细胞中与脂肪化密切相关的mi R-122的表达和相关靶基因的表达改变。结果:与高脂组比较,小蘖碱处理组肝实质细胞内甘油三酯含量降低,脂肪颗粒减少,脂肪变性明显改善,并具有明显的剂量效应,小蘖碱能够抑制FFAs诱导的JNK通路磷酸化。Q-PCR结果表明小蘖碱能够促进肝实质细胞内mi R-122的表达,并降低脂肪化相关基因Dgat2的表达。结论:小蘖碱能够显著改善高脂诱发的肝脂肪变性,抑制JNK通路磷酸化,其机制可能同mi R-122通路相关。     

Lipocalin 13 protein protects against hepatic steatosis by both inhibiting lipogenesis and stimulating fatty acid β-oxidation

Obesity is associated with hepatic steatosis, partially due to increased lipogenesis and decreased fatty acid β-oxidation in the liver; however, the underlying mechanism of abnormal lipid metabolism is not fully understood. We reported previously that obesity is associated with LCN13 (lipocalin 13) deficiency. LCN13 is a lipocalin family member involved in glucose metabolism, and LCN13 deficiency appears to contribute to hyperglycemia in obese mice. Here, we show that LCN13 is also an important regulator of lipogenesis and β-oxidation in the liver. In primary hepatocytes, recombinant LCN13 directly suppressed lipogenesis and increased fatty acid β-oxidation, whereas neutralization of endogenous LCN13 had an opposite effect. Transgenic overexpression of LCN13 protected against hepatic steatosis in mice with either dietary or genetic (ob/ob) obesity. LCN13 transgenic overexpression also improved hyperglycemia, glucose intolerance, and insulin resistance in ob/ob mice. Short-term LCN13 overexpression via an adenovirus-mediated gene transfer similarly attenuated hepatic steatosis in db/db mice. LCN13 inhibited the expression of important lipogenic genes and stimulated the genes that promote β-oxidation. These results suggest that LCN13 decreases liver lipid levels by both inhibiting hepatic lipogenesis and stimulating β-oxidation. LCN13 deficiency is likely to contribute to fatty liver disease in obese mice.     

Fat/Vessel-derived Secretory Protein (Favine)/CCDC3 Is Involved in Lipid Accumulation

We previously identified a novel gene encoding Favine/CCDC3 (NCBI protein entry {"type":"entrez-protein","attrs":{"text":"NP_083080","term_id":"58037351","term_text":"NP_083080"}}NP_083080), a possible secretory factor, the mRNA of which is highly expressed in adipose tissue and the aorta. The Favine mRNA levels are increased in the course of differentiation of rat primary adipocytes and are more elevated in the adipose tissue of genetically obese and diet-induced obese mice than in lean mice. However, its biological function has not yet been elucidated until now. Here, we tested the hypothesis that Favine is involved in lipid metabolism in adipocytes. We found that overexpression of Favine promoted 3T3-L1 adipocyte differentiation. To further investigate the function of Favine in vivo, we generated Favine knock-out (KO) mice. Favine KO mice exhibited a lean phenotype as they aged. The weights of white adipose tissue and liver were less, and adipocyte size was smaller in Favine KO mice compared with wild-type littermates (WT). Expression levels of lipogenic genes, such as fatty-acid synthase (FAS), acetyl-CoA carboxylase α (ACC1), and diacylglycerol O-acyltransferase-2 (Dgat2), were decreased in adipose tissue of Favine KO mice. In 1-year-old mice, Favine deficiency decreased the number of inflammatory cells in white adipose tissue and diminished hepatic steatosis. In vitro, deficiency of Favine attenuated differentiation of primary adipocytes. Taken together, these data demonstrate that Favine has adipogenic and lipogenic effects on adipocytes.     

Human 1-acylglycerol-3-phosphate O-acyltransferase isoforms 1 and 2: biochemical characterization and inability to rescue hepatic steatosis in Agpat2(-/-) gene lipodystrophic mice

Loss-of-function mutations in 1-acylglycerol-3-phosphate O-acyltransferase (AGPAT) 2 in humans and mice result in loss of both the white and brown adipose tissues from birth. AGPAT2 generates precursors for the synthesis of glycerophospholipids and triacylglycerols. Loss of adipose tissue, or lipodystrophy, results in hyperinsulinemia, diabetes mellitus, and severe hepatic steatosis. Here, we analyzed biochemical properties of human AGPAT2 and its close homolog, AGPAT1, and we studied their role in liver by transducing their expression via recombinant adenoviruses in Agpat2(-/-) mice. The in vitro substrate specificities of AGPAT1 and AGPAT2 are quite similar for lysophosphatidic acid and acyl-CoA. Protein homology modeling of both the AGPATs with glycerol-3-phosphate acyltransferase 1 (GPAT1) revealed that they have similar tertiary protein structure, which is consistent with their similar substrate specificities. When co-expressed, both isoforms co-localize to the endoplasmic reticulum. Despite such similarities, restoring AGPAT activity in liver by overexpression of either AGPAT1 or AGPAT2 in Agpat2(-/-) mice failed to ameliorate the hepatic steatosis. From these studies, we suggest that the role of AGPAT1 or AGPAT2 in liver lipogenesis is minimal and that accumulation of liver fat is primarily a consequence of insulin resistance and loss of adipose tissue in Agpat2(-/-) mice.     

Dietary modulation of clostridial cluster XIVa gut bacteria (Roseburia spp.) by chitin-glucan fiber improves host metabolic alterations induced by high-fat diet in mice

Recent studies have provided new evidence that alterations in the composition of the gut microbiota — known as dysbiosis — participate in the development of obesity. The aim of the present study was to investigate the ability of chitin-glucan (CG) from a fungal source to modulate both the gut microbiota and glucose and lipid metabolism in high-fat (HF) diet-induced obese mice. Supplementation of the HF diet with fungal CG (10% w/w) induced caecal enlargement with prominent changes in gut microbiota: it restored the number of bacteria from clostridial cluster XIVa including Roseburia spp., which were decreased due to HF feeding. Furthermore, CG treatment significantly decreased HF-induced body weight gain, fat mass development, fasting hyperglycemia, glucose intolerance, hepatic triglyceride accumulation and hypercholesterolemia, independently of the caloric intake. All those parameters were negatively correlated with specific bacteria of clostridial cluster XIVa, i.e., Roseburia spp. (Pearson's correlations analysis). In contrast to prebiotics that more specifically target the bifidobacteria species, CG effects on obesity appear to be independent of the incretin glucagon-like peptide 1 (GLP-1) production, since portal GLP-1 and proglucagon (its precursor) expression were not modified by the dietary intervention. In conclusion, our findings support the view that chronic consumption of CG has potential beneficial effects with respect to the development of obesity and associated metabolic diabetes and hepatic steatosis, through a mechanism related to the restoration of the composition and/or the activity of gut bacteria, namely, bacteria from clostridial cluster XIVa.     

Cannabidiol protects liver from binge alcohol-induced steatosis by mechanisms including inhibition of oxidative stress and increase in autophagy

Acute alcohol drinking induces steatosis, and effective prevention of steatosis can protect liver from progressive damage caused by alcohol. Increased oxidative stress has been reported as one mechanism underlying alcohol-induced steatosis. We evaluated whether cannabidiol, which has been reported to function as an antioxidant, can protect the liver from alcohol-generated oxidative stress-induced steatosis. Cannabidiol can prevent acute alcohol-induced liver steatosis in mice, possibly by preventing the increase in oxidative stress and the activation of the JNK MAPK pathway. Cannabidiol per se can increase autophagy both in CYP2E1-expressing HepG2 cells and in mouse liver. Importantly, cannabidiol can prevent the decrease in autophagy induced by alcohol. In conclusion, these results show that cannabidiol protects mouse liver from acute alcohol-induced steatosis through multiple mechanisms including attenuation of alcohol-mediated oxidative stress, prevention of JNK MAPK activation, and increasing autophagy.     

Cloning and functional characterization of a novel mitochondrial N-ethylmaleimide-sensitive glycerol-3-phosphate acyltransferase (GPAT2)

Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial and rate-limiting step in glycerolipid synthesis. Several mammalian GPAT activities have been recognized, including N-ethylmaleimide (NEM)-sensitive isoforms in microsomes and mitochondria and an NEM-resistant form in mitochondrial outer membrane (GPAT1). We have now cloned a second mitochondrial isoform, GPAT2 from mouse testis. The open-reading frame encodes a protein of 798 amino acids with a calculated mass of 88.8kDa and 27% amino acid identity to GPAT1. Testis mRNA expression was 50-fold higher than in liver or brown adipose tissue, but the specific activity of NEM-sensitive GPAT in testis mitochondria was similar to that in liver. When Cos-7 cells were transiently transfected with GPAT2, NEM-sensitive GPAT activity increased 30%. Confocal microscopy confirmed a mitochondrial location. Incubation of GPAT2-transfected Cos-7 cells with trace (3 microM; 0.25 microCi) [1-(14)C]oleate for 6h increased incorporation of [(14)C]oleate into TAG 84%. In contrast, incorporation into phospholipid species was lower than in control cells. Although a polyclonal antibody raised against full-length GPAT1 detected an approximately 89-kDa band in liver and testis from GPAT1 null mice and both 89- and 80-kDa bands in BAT from the knockout animals, the GPAT2 protein expressed in Cos-7 cells was only 80 kDa. In vitro translation showed a single product of 89 kDa. Unlike GPAT1, GPAT2 mRNA abundance in liver was not altered by fasting or refeeding. GPAT2 is likely to have a specialized function in testis.