JMJD2B/KDM4B inactivation in adipose tissues accelerates obesity and systemic metabolic abnormalities

Obesity is a serious global health issue; however, the roles of genetics and epigenetics in the onset and progression of obesity are still not completely understood. The aim of this study was to determine the role of Kdm4b, which belongs to a subfamily of histone demethylases, in adipogenesis and fat metabolism in vivo. We established conditional Kdm4b knockout mice. Inactivation of Kdm4b in adipocytes (K4bKO) induced profound obesity in mice on a high fat diet (HFD). The HFD‐fed K4bKO mice exhibited an increased volume of fat mass and higher expression levels of adipogenesis‐related genes. In contrast, the genes involved in energy expenditure and mitochondrial functions were down‐regulated. Supporting these findings, the energy expenditure of Kdm4b‐deficient cells was markedly decreased. In addition, progression of glucose intolerance and hepatic steatosis with hepatocellular damages was observed. These data indicate that Kdm4b is a critical regulator of systemic metabolism via enhancing energy expenditure in adipocytes.     

Involvement of receptor-interacting protein 140 in estrogen-mediated osteoclasts differentiation,apoptosis, and bone resorption

Estrogen withdrawal following menopause results in an increase of osteoclasts formation and bone resorption, which is one of the most important mechanisms of postmenopausal osteoporosis. Recently, growing evidence has suggested that receptor-interacting protein 140 was implicated in estrogen-regulated metabolic disease, including fat metabolism and lipid metabolism. However, little is known regarding the role of receptor-interacting protein 140 in the regulation of bone metabolic by estrogen. In the present study, Western blotting disclosed that estrogen brings a significant increasing expression of receptor-interacting protein 140 in osteoclasts, but not in osteoblasts and bone marrow mesenchymal stem cells. Furthermore, analysis of TRAP staining and bone resorption assay showed that depletion of receptor-interacting protein 140 could significantly alleviate the inhibitory effects of estrogen on osteoclasts formation and bone resorption activity. Moreover, estrogen could induce osteoclasts apoptosis by increasing receptor-interacting protein 140 expression through the Fas/FasL pathway. Taken together, receptor-interacting protein 140 might be a critical player in estrogen-mediated osteoclastogenesis and bone resorption.     

Identification of a novel role for Drosophila MESR4 in lipid metabolism

Drosophila provides a powerful genetic model to analyze lipid metabolism. Drosophila has an adipose‐like organ called the fat body, which plays a crucial role in energy homeostasis. Here, we conducted a fat body‐specific misexpression screen to identify genes involved in lipid metabolism. We found that over‐expression of a nuclear protein with nine C₂H₂ type zinc‐finger motifs and a PHD‐finger, Misexpression suppressor of ras 4 (MESR4), reduces lipid accumulation in the fat body, whereas MESR4 knockdown increases it. We further show that MESR4 up‐regulates the expression of major lipases, which may account for the reduction in lipid storage in the fat body and the release of free fatty acids (FFAs) in the body. These results suggest that MESR4 acts as an important upstream regulator of energy homeostasis.     

resistin基因过表达影响3T3-L1脂肪细胞的脂质代谢

目的观察resistin基因过表达对3T3-L1脂肪细胞的脂质代谢、糖代谢的影响。方法构建大鼠resistin真核表达载体并转染3T3-L1前体脂肪细胞,获得稳定表达resistin基因的细胞株;采用油红O染色,观察脂肪细胞分化及脂质积聚情况;采用逆转录PCR技术,检测脂肪细胞分化标志基因及葡萄糖转运体4（glucose transporter4,GLUT4）基因表达变化;采用全自动生化仪比色法,检测脂肪细胞内甘油三酯（triglyceride,TG）、游离脂肪酸（free fatty acids,FFAs）的含量变化。结果（1）resistin基因过表达脂肪细胞中,脂滴出现时间提前,且细胞内布满了小而多的圆形脂滴;（2）resistin基因过表达脂肪细胞中,分化中、晚期标志基因C/EBPα、FAS的mRNA表达水平明显上调,分化早期标志基因Pref-1的表达则明显下调;（3）re-sistin基因过表达脂肪细胞中,胞质内TG、FFAs含量均显著增加;（4）resistin基因过表达脂肪细胞中,分化第2、4、8d的GLUT4基因mRNA表达水平间无显著变化,与正常脂肪细胞中的表达水平差异也无统计学意义。结论resistin基因过表达能够显著干扰3T3-L1脂肪细胞的脂质代谢,有助于肥胖和胰岛素抵抗的发生,而并不影响GLUT4基因的表达。     

Large-scale RNAi screens add both clarity and complexity to Drosophila NF-κB signaling

NF-κB signaling is an immune response mechanism remarkably conserved through phylogeny. The genetically tractable model animal Drosophila melanogaster is an important model organism for studying NF-κB signaling in the immune response. Fruit flies have two NF-κB signaling pathways: the Toll and the Imd pathway. Traditional genetic screens have revealed many important aspects about the regulation of Drosophila NF-κB signaling and have helped us to also understand the immune response in humans. For example, the discovery that Toll like receptors are the main immune signaling molecules in mammals was based on work in flies. During the past decade high throughput RNA interference (RNAi)-based screening in cultured Drosophila cells has become a common method for identifying novel genes required for numerous cellular processes including NF-κB signaling. These screens have identified many novel positive and negative regulators of Drosophila NF-κB signaling thus enhancing our understanding of these signaling cascades.     

Life span and stress resistance of Caenorhabditis elegans are differentially affected by glutathione transferases metabolizing 4-hydroxynon-2-enal

The lipid peroxidation product 4-hydroxynon-2-enal (4-HNE) forms as a consequence of oxidative stress, and acts as a signaling molecule or, at superphysiological levels, as a toxicant. The steady-state concentration of the compound reflects the balance between its generation and its metabolism, primarily through glutathione conjugation. Using an RNAi-based screen, we identified in Caenorhabditis elegans five glutathione transferases (GSTs) capable of catalyzing 4-HNE conjugation. RNAi knock-down of these GSTs (products of the gst-5, gst-6, gst-8, gst-10, and gst-24 genes) sensitized the nematode to electrophilic stress elicited by exposure to 4-HNE. However, interference with the expression of only two of these genes (gst-5 and gst-10) significantly shortened the life span of the organism. RNAi knock-down of the other GSTs resulted in at least as much 4-HNE adducts, suggesting tissue specificity of effects on longevity. Our results are consistent with the oxidative stress theory of organismal aging, broadened by considering electrophilic stress as a contributing factor. According to this extended hypothesis, peroxidation of lipids leads to the formation of 4-HNE in a chain reaction which amplifies the original damage. 4-HNE then acts as an "aging effector" via the formation of 4-HNE-protein adducts, and a resulting change in protein function.     

受体相互作用蛋白激酶RIP1在细胞信号转导途径中作用的研究进展

 受体相互作用蛋白1是一类丝氨酸/苏氨酸蛋白激酶,是一种重要的细胞信号转导分子,从其被发现开始就成为了细胞信号转导的研究热点,但多集中在其在细胞凋亡中的作用,近年来的研究表明,受体相互作用蛋白1不仅参与了细胞的凋亡,还参与了细胞存活,细胞程序性坏死等多种信号的转导,是一类重要的调节细胞生存或死亡的信号分子,本文对受体相互作用蛋白1功能的研究进展做简要综述。     

Development of nutritional obesity in transgenic mice with an adrenergic receptivity in adipose tissue comparable with that of humans

Obesity is characterized by an excessive development of fat mass which is a consequence of increased fat cell size and/or fat cell number. Several hormones and neurotransmitters are regulators of adipose tissue development and metabolism. Among them, catecholamines play a major role by acting through alpha 2- and beta-adrenergic receptors. Stimulation of alpha 2-adrenergic receptors induce inhibition of lipolysis in mature adipocytes as well as preadipocyte proliferation. The antilipolytic effect mediated by alpha 2-adrenergic receptors is in part responsible for the weak lipid mobilization of some fat deposits in humans (subcutaneous fat in particular). Changes in beta- and alpha 2-adrenergic receptors ratio and function have been proposed to explain the lipolytic disturbances described in some obese subjects. Human and rodent adipocytes differ considerably with respect to the balance between beta- and alpha 2-adrenergic receptors. Human adipocytes express mainly alpha 2- but very few beta 3-adrenergic receptors while the reverse is true for rodent adipocytes. Since no suitable animal model was available to study the contribution of alpha 2/beta-adrenergic balance in adipocytes in vivo, we combined gene targeting and transgenic approaches to create a mice with increased alpha 2/beta-adrenergic ratio in adipose tissue. Specifically, we have generated transgenic mice strains on a beta 3-adrenergic receptor knock-out background which express human alpha 2-adrenergic receptors. No particular phenotype was observed in mice maintained in normal diet whereas when fed a high fat diet, transgenic mice increased significantly body weight and fat mass. These results underline the physiologic relevance of the interaction of the presence of alpha 2-adrenergic receptors with a high fat diet in the control of adipose tissue development.     

绝经后骨质疏松症与脂代谢的相关性

动脉粥样硬化常与骨质疏松症共存，进一步分析发现脂代谢指标与骨代谢指标有相关性，而且多数用于治疗骨质疏松症的药物同时对血脂有调节作用，所以推测脂代谢与骨代谢有相关性。其机制可能有：脂肪酸可以促进成骨细胞向脂肪细胞的转化及促进前破骨细胞向破骨细胞分化，调节骨髓间质细胞分化的转录因子及信号转导途径，脂代谢与骨代谢的遗传基因有相关性，高脂血症后骨髓内脂肪细胞增多增大压迫微血管使骨头供血明显减少等。     

A genetic screen identifies the Triple T complex required for DNA damage signaling and ATM and ATR stability

In response to DNA damage, cells activate a complex signal transduction network called the DNA damage response (DDR). To enhance our current understanding of the DDR network, we performed a genome-wide RNAi screen to identify genes required for resistance to ionizing radiation (IR). Along with a number of known DDR genes, we discovered a large set of novel genes whose depletion leads to cellular sensitivity to IR. Here we describe TTI1 (Tel two-interacting protein 1) and TTI2 as highly conserved regulators of the DDR in mammals. TTI1 and TTI2 protect cells from spontaneous DNA damage, and are required for the establishment of the intra-S and G2/M checkpoints. TTI1 and TTI2 exist in multiple complexes, including a 2-MDa complex with TEL2 (telomere maintenance 2), called the Triple T complex, and phosphoinositide-3-kinase-related protein kinases (PIKKs) such as ataxia telangiectasia-mutated (ATM). The components of the TTT complex are mutually dependent on each other, and act as critical regulators of PIKK abundance and checkpoint signaling.     

Generation of RNAi libraries for high-throughput screens

The completion of the genome sequencing for several organisms has created a great demand for genomic tools that can systematically analyze the growing wealth of data. In contrast to the classical reverse genetics approach of creating specific knockout cell lines or animals that is time-consuming and expensive, RNA-mediated interference (RNAi) has emerged as a fast, simple, and cost-effective technique for gene knockdown in large scale. Since its discovery as a gene silencing response to double-stranded RNA (dsRNA) with homology to endogenous genes in Caenorhabditis elegans (C elegans), RNAi technology has been adapted to various high-throughput screens (HTS) for genome-wide loss-of-function (LOF) analysis. Biochemical insights into the endogenous mechanism of RNAi have led to advances in RNAi methodology including RNAi molecule synthesis, delivery, and sequence design. In this article, we will briefly review these various RNAi library designs and discuss the benefits and drawbacks of each library strategy.     

Rev-erbα and Rev-erbβ coordinately protect the circadian clock and normal metabolic function

The nuclear receptor Rev-erbα regulates circadian rhythm and metabolism, but its effects are modest and it has been considered to be a secondary regulator of the cell-autonomous clock. Here we report that depletion of Rev-erbα together with closely related Rev-erbβ has dramatic effects on the cell-autonomous clock as well as hepatic lipid metabolism. Mouse embryonic fibroblasts were rendered arrhythmic by depletion of both Rev-erbs. In mouse livers, Rev-erbβ mRNA and protein levels oscillate with a diurnal pattern similar to that of Rev-erbα, and both Rev-erbs are recruited to a remarkably similar set of binding sites across the genome, enriched near metabolic genes. Depletion of both Rev-erbs in liver synergistically derepresses several metabolic genes as well as genes that control the positive limb of the molecular clock. Moreover, deficiency of both Rev-erbs causes marked hepatic steatosis, in contrast to relatively subtle changes upon loss of either subtype alone. These findings establish the two Rev-erbs as major regulators of both clock function and metabolism, displaying a level of subtype collaboration that is unusual among nuclear receptors but common among core clock proteins, protecting the organism from major perturbations in circadian and metabolic physiology.     

Genetic polymorphisms in fatty acid metabolism genes and colorectal cancer

Colorectal cancer (CRC) is a leading cause of cancer death worldwide. Epidemiological risk factors for CRC included dietary fat intake; consequently, the role of genes in the fatty acid biosynthesis and metabolism pathways is of particular interest. Moreover, hyperlipidaemia has been associated with different type of cancer and serum lipid levels could be affected by genetic factors, including polymorphisms in the lipid metabolism pathway. The aim of this study is to assess the association between single-nucleotide polymorphisms (SNPs) in fatty acid metabolism genes, serum lipid levels, body mass index (BMI) and dietary fat intake and CRC risk; 30 SNPs from 8 candidate genes included in fatty acid biosynthesis and metabolism pathways were genotyped in 1780 CRC cases and 1864 matched controls from the Molecular Epidemiology of Colorectal Cancer study. Information on clinicopathological characteristics, lifestyle and dietary habits were also obtained. Logistic regression and association analysis were conducted. Several LIPC (lipase, hepatic) polymorphisms were found to be associated with CRC risk, although no particular haplotype was related to CRC. The SNP rs12299484 showed an association with CRC risk after Bonferroni correction. We replicate the association between the T allele of the LIPC SNP rs1800588 and higher serum high-density lipoprotein levels. Weak associations between selected polymorphism in the LIPC and PPARG genes and BMI were observed. A path analysis based on structural equation modelling showed a direct effect of LIPC gene polymorphisms on colorectal carcinogenesis as well as an indirect effect mediated through serum lipid levels. Genetic polymorphisms in the hepatic lipase gene have a potential role in colorectal carcinogenesis, perhaps though the regulation of serum lipid levels.     

脂类代谢和运输涉及的基因与大鼠肝再生的相关性分析

目的在基因转录水平了解脂类代谢和运输相关基因在大鼠肝再生(LR)中的表达变化和模式。方法用搜集网站资料和查阅相关论文等方法获得参与脂类代谢和运输基因,用大鼠基因组230 2.0芯片检测它们在大鼠再生肝中的表达情况,用比较手术和假手术中基因表达的差异性确定肝再生相关基因。结果初步证实上述基因中193个基因与肝再生相关。肝再生早期[部分肝切除(PH)后0.5~4h]、前期(PH后6~12h)、中期(PH后12~66h)、后期(PH后72~168h)等4个阶段起始表达的基因数为113、20、66和1;基因的总表达次数为250、205、796和293。共上调852次,下调630次,分为27种表达方式。肝再生早期和前期胆汁酸代谢相关基因转录减弱;早期和后期糖皮质激素分解相关基因转录增强;前期和中期磷脂合成相关基因转录增强,磷脂分解相关基因转录减弱;中期脂肪酸、白三烯和鞘糖脂合成相关基因转录增强,甘油三酯和磷脂酰肌醇代谢相关基因转录增强,鞘糖脂分解相关基因转录减弱;中期和后期前列腺素合成和脂肪酸分解相关基因转录增强;几乎在整个肝再生中性激素、糖皮质激素和孕酮合成相关基因转录增强,鞘磷脂代谢相关基因转录增强,脂类运输相关基因转录增强,胆固醇代谢相关基因转录减弱。结论肝再生中脂类代谢和运输变化较大,与肝再生密切相关。     

积雪草酸改善小鼠脂肪细胞胰岛素抵抗的机制研究

 目的： 观察番石榴叶三萜化合物积雪草酸对小鼠3T3-L1前脂肪细胞增殖、分化以及胰岛素抵抗脂肪细胞糖脂代谢的影响并探讨其作用机制。方法：MTT法检测药物对3T3-L1前脂肪细胞增殖的影响;油红O染色法观察药物对其分化的影响。地塞米松诱导建立脂肪细胞胰岛素抵抗模型,药物干预后采用葡萄糖氧化酶法检测培养液中葡萄糖消耗量;比色法检测游离脂肪酸浓度;ELISA法检测脂联素水平;Western blotting法检测过氧化物酶体增殖物激活受体γ（PPARγ）和蛋白酪氨酸磷酸酶1B(PTP1B)蛋白表达的变化。结果：与溶媒对照组相比,积雪草酸在10~100 μmol/L时能显著促进3T3-L1前脂肪细胞增殖,但明显抑制其分化（P<0.05或P<0.01）;在30和100 μmol/L时,无论是基础状态还是胰岛素刺激状态,均能显著增加胰岛素抵抗脂肪细胞葡萄糖的消耗,减少游离脂肪酸的产生（P<0.05）;其对胰岛素抵抗脂肪细胞的脂联素分泌和PPARγ蛋白表达无明显影响（P>0.05）,但能显著下调PTP1B蛋白的表达（P<0.05或P<0.01）。结论：积雪草酸能显著改善脂肪细胞胰岛素抵抗,增加胰岛素抵抗脂肪细胞葡萄糖的消耗和减少游离脂肪酸的产生,其机制可能是其下调胰岛素信号转导的负性调节因子PTP1B的表达,增强胰岛素信号转导,从而改善胰岛素抵抗。