脂肪细胞的分化形成与天然产物调控

研究脂肪生成的机理及其调控过程对于防治动物肥胖引起的相关疾病、改善肉品风味和质量以及畜牧生产效率具有重要意义。脂肪细胞起源于多能性骨髓间充质干细胞（MSCs）,接受细胞外刺激因子后迅速引起早期脂肪调节因子、C/EBPβ和C/EBPδ的表达,并传递信息至PPARγ和C/EBPα等转录因子,促进细胞的分化和脂滴形成。在此过程中,众多转录因子和细胞周期蛋白参与前体脂肪细胞到脂肪细胞的分化与成脂过程。植物来源的多种天然产物,如多酚类、生物碱、萜类、醌类化合物在脂肪细胞分化和抑制脂肪生成过程中发挥重要的调控作用,对这些天然化合物的深入和成药研究有望成为具有抑制细胞内脂滴聚集的潜在药物。     

INS、GLN和LP对体外培养牛脂肪细胞内HSL、LP的mRNA丰度的影响

为了阐明胰岛素（INS）、胰高血糖素（GLN）和瘦蛋白（LP）对牛脂肪细胞内激素敏感酯酶（HSL）、LP的mRNA表达的调控作用，本试验采用体外原代培养牛脂肪细胞的培养液中添加不同浓度的胰岛素（0、5、10、20、50、100mmol／L）、胰高血糖素（O、25、100、200、500、1000pg／mL）和瘦蛋白（O、2．5、5、10、50、100ng／mL），培养12h后，应用建立的实时荧光定量PCR（FQ-PCR）方法检测牛脂肪细胞内HSL、LP的mRNA丰度。结果表明：胰岛素对牛脂肪细胞内HSL的mRNA表达呈现剂量依赖性抑制作用，对LP的mRNA表达呈现剂量依赖性促进作用。胰高血糖素对牛脂肪细胞内HSL的mRNA表达呈现剂量依赖性促进作用，对牛脂肪细胞内LP的mRNA表达无作用。瘦蛋白对牛脂肪细胞内HSL的mRNA表达先呈现剂量依赖性促进作用，后呈现抑制作用。但对牛脂肪细胞内LP的mRNA表达呈现剂量依赖性抑制作用。     

不同生物素水平对3T3-L1脂肪细胞脂肪合成相关基因转录表达的影响

【目的】诱导3T3-L1细胞分化为3T3-L1脂肪细胞,研究不同生物素水平对3T3-L1脂肪细胞脂肪合成相关基因转录表达的影响。【方法】利用三联诱导法将3T3-L1细胞经诱导分化为3T3-L1脂肪细胞。当3T3-L1脂肪细胞密集后分别采用0(对照组)、0.2、0.5、1 μmol/L生物素处理,分别在12 h、24 h与48 h时检测细胞上清液中PK mRNA、GLUT-4 mRNA、FAS mRNA及ACC1 mRNA相对表达量。【结果】在试验12 h时:各试验组GLUT-4、PK、ACC1 mRNA相对表达量均极显著(P＜0.01)高于对照组,1 μmol/L组与0.5 μmol/L组FAS mRNA相对表达量极显著(P＜0.01)高于对照组与0.2 μmol/L组;在试验24 h时:1 μmol/L组GLUT-4 mRNA与PK mRNA相对表达量显著(P＜0.05)高于对照组,1 μmol/L组ACC1 mRNA相对表达量极显著(P＜0.01)高于对照组,0.2 μmol/L组与0.5 μmol/L组ACC1 mRNA相对表达量显著(P＜0.05)高于对照组;在试验48 h时:1 μmol/L组GLUT-4 mRNA相对表达量显著(P＜0.05)高于对照组,1 μmol/L 组PK mRNA相对表达量极显著(P＜0.01)高于对照组,0.5 μmol/L组FAS mRNA相对表达量极显著(P＜0.05)高于1 μmol/L组,1 μmol/L组FAS mRNA相对表达量显著(P＜0.05)高于对照组,1 μmol/L组ACC1 mRNA相对表达量极显著(P＜0.01)高于其它组。【结论】生物素的添加可以提升脂肪细胞GLUT-4、PK、FAS与ACC1 mRNA相对表达量,且当以1 μmol/L浓度作用时对GLUT-4、PK与ACC1提升效果最佳,以0.5 μmol/L浓度作用时对FAS提升效果最佳。     

Overexpression of Krueppel like factor 3 promotes subcutaneous adipocytes differentiation in goat Capra hircus

Previous research reported that KLF3 plays different roles in the regulation of adipose deposition across species. However, the exact function of KLF3 in goat subcutaneous adipocyte remains unknown. Here, the goat KLF3 gene was firstly cloned and showed that the mRNA sequence of the goat KLF3 gene was 1,264 bp (GenBank accession number: KU041753.1) and its coding sequence was 1,037 bp, encoding 345 amino acids with three classic zinc finger domains of KLFs family at its C-terminus. The alignment of the amino acid sequence of KLF3 among various species demonstrated that goat had the highest homology to that of sheep, presenting 99.4% similarity, while the homology similarity to that of mice presented only 93.62% in contrast. Furthermore, KLF3 had highest mRNA level in fat tissue and lowest level in the heart in comparison. Additionally, the mRNA level of KLF3 gradually tended to increase during adipogenesis. Interestingly, overexpression of KLF3 increased lipid accumulation. In line with this, the gain-of-function of KLF3 dramatically elevated the mRNA levels of TG synthetic genes and adipogenic maker genes (p < .01) . Moreover, overexpression of KLF3 upregulated all the potential target genes, except for C/EBPα. These results suggested that KLF3 is a positive regulator for subcutaneous adipocyte differentiation in goats.     

N-carbamoylglutamate improves lipid metabolism,inflammation, and apoptosis responses in visceral adipocytes of Japanese seabass (Lateolabrax japonicus), in vivo and in vitro

This study applied in vivo and in vitro methods to investigate the effect of dietary N-carbamoylglutamate (NCG) on lipid metabolism, inflammation and apoptosis related-gene expression in visceral adipose tissue and isolated adipocytes of Japanese seabass (Lateolabrax japonicus). A basal diet and a test diet supplemented with 720 mg/kg NCG were fed to the fish for 10 weeks. During the growth trial, no mortality and no significant differences in growth performance were observed in fish between the 2 groups (P > 0.05). Plasma Arg content and mRNA level of argininosuccinate synthetase (ASS) in adipose tissue were significantly increased, which indicated that NCG inclusion promoted endogenous Arg synthesis. Thereafter, the potential effects of NCG treatment on lipid metabolism-related genes expression were studied through in vivo and in vitro methods. In the present study, we successfully established a primary adipocytes culture system and isolated pre-adipocytes in vitro of Japanese seabass for the first time. Both the results in vivo and in vitro showed that NCG treatment decreased the mRNA levels of genes related to adipogenesis (fatty acid synthase, FASN), cholesterol synthesis (3-hydroxy-3-methylglutaryl-CoA reductase, HMGCR) and fat deposition (lipoprotein lipase [LPL] and leptin), which revealed the underlying mechanism of NCG on reducing fat deposition. The results of this study demonstrated that NCG inclusion reduced the expression of inflammatory and apoptosis cytokines markedly in vivo and in vitro. In conclusion, NCG did exert beneficial effects on ameliorating adipogenesis, inflammation and apoptosis via promoting Arg endogenous synthesis in Japanese seabass.     

lncRNA-6617调控猪肌内前体脂肪细胞分化的筛选与功能研究

旨在探究lncRNA-6617的生物学特性及其对猪肌内前体脂肪细胞分化的影响,以及初步探究其上游调节机制,为后续深入研究猪肌内脂肪沉积调控机制提供参考依据。本研究以饲养在相同条件下的1、90、180日龄健康马身公猪(n=3)和大白公猪(n=3)为研究对象,利用RNA-seq技术筛选脂肪型猪(马身猪)和瘦肉型猪(大白猪)肌肉特异表达的lncRNAs,并通过生物信息学分析、RT-PCR及Sanger测序鉴定lncRNA-6617生物学特性及其猪组织中的表达特征。在猪肌内前体脂肪细胞中干扰lncRNA-6617,检测lncRNA-6617对肌内脂肪细胞分化能力的影响。随后,检测m⁶A修饰相关酶在不同品种猪中的表达差异,研究lncRNA-6617的上游调控机制。结果显示,lncRNA-6617位于猪18号染色体,RAMP3基因反义链的第3内含子区域,无蛋白编码能力,主要分布于细胞核; lncRNA-6617在猪所检测各组织中均有表达,背部皮下脂肪中表达量最高,且在马身猪肌肉组织中的表达极显著高于大白猪(P < 0.01);在猪肌内前体脂肪细胞分化过程中,lncRNA-6617表达量呈上升趋势,且在分化第5天表达量达到最高; 干扰lncRNA-6617诱导分化7 d后,分化的成熟脂肪细胞明显减少,成脂关键基因CEBPα、PPARγ和aP2的表达均极显著下调(P < 0.01)。m⁶A修饰的去甲基化酶FTO在马身猪背最长肌和股二头肌中的表达极显著高于大白猪(P < 0.01),在腰大肌中的表达显著高于大白猪(P < 0.05),上游调节因子ZNF217在马身猪肌肉组织中的表达极显著高于大白猪(P < 0.01),与lncRNA-6617表达模式一致。干扰FTO后,lncRNA-6617的表达极显著下调(P < 0.01)。本研究筛选了在马身猪肌肉组织中显著高表达的lncRNA-6617,并深入研究了lncRNA-6617促进猪肌内前体脂肪细胞分化的功能及其上游调节机制,丰富了脂肪细胞生成的表观调控网络。     

六堡茶对胰岛素抵抗3T3-L1脂肪细胞糖脂代谢的影响

采用软脂酸诱导3T3-L1脂肪细胞胰岛素抵抗后,给予不同剂量的六堡茶水提物治疗,探讨六堡茶对胰岛素抵抗3T3-L1脂肪细胞的糖脂代谢的影响。结果表明,1 mmol·L^-1的软脂酸对3T3-L1脂肪细胞作用24 h后,葡萄糖摄取量减少,建立胰岛素抵抗模型。六堡茶对模型细胞干预24 h后,与模型组比较,细胞培养基的残留液中葡萄糖和游离脂肪酸的含量降低;对糖脂代谢相关关键酶（己糖激酶、6-磷酸果糖激酶-1、丙酮酸激酶、甘油-3-磷酸酰基转移酶）和葡萄糖转运子基因表达都有上调作用,并下调乙酰辅酶A羧化酶、蛋白酪氨酸磷酸酯酶1B基因表达。低剂量的六堡茶下调肉碱脂酰转移酶-I基因的表达,但是高剂量能显著上调肉碱脂酰转移酶-I基因表达。实验证明了六堡茶能够提高胰岛素抵抗3T3-L1脂肪细胞对葡萄糖的摄取能力,同时促进细胞内糖脂代谢,六堡茶有改善3T3-L1脂肪细胞胰岛素抵抗的作用。     

荞麦多酚干预对高脂膳食诱导小鼠的降脂作用及其调控机制

【背景】目前已有相关动物实验证实荞麦对于高脂膳食引起的肥胖和脂代谢紊乱具有良好的干预作用。同时研究表明,棕色脂肪适应性产热可以有效改善人体能量新陈代谢。因此,增加棕色脂肪活性、促进白色脂肪棕色化可作为一种预防肥胖和改善能量代谢疾病的有效途径。【目的】采用原粮及从荞麦中鉴定出的4种主要酚类物质按在原粮中的含量比例配制成多酚干预物进行干预,研究荞麦多酚通过调节白色脂肪棕色化对高脂膳食小鼠的干预及改善作用,系统研究荞麦中发挥降脂功能的主要酚类物质及其对脂代谢的调控机制。【方法】通过UHPLC-Q-Orbitrap质谱结合数据库对荞麦多酚提取物中的酚类物质进行鉴定,并通过相对定量将其中含量最多的主要多酚按其在荞麦原粮中的含量比例配制成多酚复合物。通过添加荞麦饲料以及荞麦多酚复合物干预C57BL/6J小鼠饮食,研究荞麦酚类物质改善脂质代谢的作用。以10%、20%、40%比例的荞麦替换基础饲料和2.5 mg·mL^-1荞麦多酚复配物对小鼠进行饮食干预,探究荞麦对小鼠体重、脏器指数和血脂水平的调节作用。并通过Western-blot和q-PCR探究荞麦饲料及多酚干预对小鼠皮下脂肪...     

不同生物素水平对3T3-L1脂肪细胞脂肪分解及相关基因表达的影响

为研究不同生物素水平对3T3-L1脂肪细胞脂肪分解及相关基因表达的影响,本试验利用三联诱导法将3T3-L1细胞经诱导分化为3T3-L1脂肪细胞。当3T3-L1脂肪细胞密集后分别采用0（对照组）、0.2、0.5、1.0 μmol/L生物素处理,分别在12、24与48 h时检测细胞上清液中甘油释放量、脂肪甘油三酯水解酶（ATGL）、激素敏感性脂肪酶（HSL）、脂滴包被蛋白A （perilipin A）及脂肪分化相关蛋白（ADRP）相对表达量。结果显示,在12 h时,1.0 μmol/L组甘油释放量、ATGL基因mRNA相对表达量均极显著低于对照组（P<0.01）,0.5、1.0 μmol/L组HSL基因mRNA相对表达量显著低于对照组（P<0.05）,1.0 μmol/L组perilipin A、ADRP基因mRNA相对表达量均极显著高于对照组与0.2 μmol/L组（P<0.01）;在24 h时,各试验组甘油释放量、1.0 μmol/L组ATGL基因mRNA相对表达量均极显著低于对照组（P<0.01）,0.5、1.0 μmol/L组perilipin A基因mRNA相对表达量极显著高于对照组与0.2 μmol/L组（P<0.01）;1.0 μmol/L组ADRP基因mRNA相对表达量极显著高于其他组（P<0.01）;在48 h时,1.0 μmol/L组甘油释放量、ATGL基因mRNA相对表达量均极显著或显著低于对照组（P<0.01;P<0.05）,1.0 μmol/L组perilipin A基因mRNA相对表达量显著高于对照组与0.2 μmol/L组（P<0.05）,0.5、1.0 μmol/L组ADRP基因mRNA相对表达量极显著高于对照组（P<0.01）。综上所述,生物素可通过促进perilipin A与ADRP表达,同时抑制ATGL与HSL表达,达到抑制脂肪分解的效果,且浓度为1.0 μmol/L时效果最佳。     

Lipid Inhibitory Effect of (−)-loliolide Isolated from Sargassum horneri in 3T3-L1 Adipocytes: Inhibitory Mechanism of Adipose-Specific Proteins

Sargassum horneri (S. horneri) is a well-known brown seaweed widely distributed worldwide. Several biological activities of S. horneri have been reported. However, its effects on lipid metabolism and the underlying mechanisms remain elusive. In the present study, we examined the inhibitory effect of the active compound “(−)-loliolide ((6S,7aR)-6-hydroxy-4,4,7a-trimethyl-5,6,7,7a-tetrahydro-1-benzofuran-2(4H)-one (HTT))” from S. horneri extract on lipid accumulation in differentiated adipocytes. MTT assays demonstrated that (−)-loliolide is not toxic to 3T3-L1 adipocytes in a range of concentrations. (−)-loliolide significantly reduced intracellular lipid accumulation in the differentiated phase of 3T3-L1 adipocytes as shown by Oil Red O staining. Western blot analysis revealed that (−)-loliolide increased the expression of lipolytic protein phospho-hormone-sensitive lipase (p-HSL) and thermogenic protein peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1). Additionally, (−)-loliolide decreased expression of adipogenic and lipogenic proteins, including sterol regulatory element-binding protein-1 (SREBP-1), peroxisome proliferator-activated receptor-γ (PPAR-γ), CCAAT/enhancer-binding protein-α (C/EBP-α), and fatty acid-binding protein 4 (FABP4) in 3T3-L1 adipocytes. These results indicate that (−)-loliolide from S. horneri could suppress lipid accumulation via regulation of antiadipogenic and prolipolytic mechanisms in 3T3-L1 cells. Considering the multifunctional effect of (−)-loliolide, it can be useful as a lipid-lowering agent in the management of patients who suffer from obesity.