Mechanism of anti-lipolytic action of acipimox in isolated rat adipocytes

Summary Acipimox is commonly used to treat hyper-triglyceridaemia in non-insulin-dependent diabetic patients, but its precise mechanism of action has yet to be elucidated. We examined the in vitro effects of acipimox on the lipolytic regulatory cascade in epididymal adipocytes isolated from Wistar rats. Acipimox inhibited the lipolytic rate stimulated by adenosine deaminase (1 U/ml) in a concentration-dependent manner, reaching a near-basal value at 10 mol/l acipimox. Lipolysis activated by sub-maximal levels of isoproterenol in combination with adenosine deaminase (20 mU/ml) was significantly (p<0.05) decreased by 100 mol/l acipimox, whereas, in the absence of adenosine deaminase, 100 mol/l acipimox showed no significant (p>0.05) inhibition. These findings suggested that the anti-lipolytic mechanism regulated by adenosine may also be regulated by acipimox. Acipimox diminished the intracellular cyclic AMP level produced by 25 nmol/l isoproterenol in the presence of adenosine deaminase (20 mU/ml) in a concentration-dependent manner. At the same level of stimulation, acipimox inhibited the cyclic AMP-dependent protein kinase activity ratio and lipolytic rate over the same concentration range, with significant (p<0.05) reductions occurring at and above, 0.5 mol/l and 10 mol/l acipimox, respectively. Western blotting showed that upon lipolytic stimulation (1 U/ml adenosine deaminase; 100 nmol/l isoproterenol) a threefold increase in the lipolytic rate was accompanied by a significant (p<0.05) rise in hormone-sensitive lipase associated with the lipid fraction. Acipimox (1 mmol/l) and insulin (1 nmol/l) re-distributed hormone-sensitive lipase back to the cytosol, with a corresponding significant (p<0.05) loss from the fat cake fraction of adipocyte homogenates. In conclusion, the anti-lipolytic action of acipimox is mediated through suppression of intracellular cyclic AMP levels, with the subsequent decrease in cyclic AMP-dependent protein kinase activity, leading to the reduced association of hormone-sensitive lipase with triacylglycerol substrate in the lipid droplet of adipocytes.Abbreviations NIDDM Non-insulin-dependent diabetes mel-litus - HSL hormone-sensitive lipase - cAMP adenosine 35-cyclic monophosphate - A-kinase cyclic AMP-dependent protein kinase - DTT dithiothreitol - PIA N⁶-[R-(–)-1-methyl-2-phenethyl]-adenosine - Hepes 4-(2-hydroxyethyl)-1-piperazi-neethanesulphonic acid - BSA bovine serum albumin - PCV packed cell volume - Ro 20-1724 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone - MOPS 3-(N-morpholino) propanesulphonic acid - ADA adenosine deaminase - NEFA non-esterified fatty acids - KRH Krebs-Ringer-Hepes - TCA trichloroacetic acid - PBS phosphate buffered saline - IC₅₀ half-maximal inhibition     

Ex vivo study of incorporation into adipocytes and lipolysis-inhibition effect of polycyclic aromatic hydrocarbons

We have previously shown that benzo[a]pyrene (B[a]P) administrated at extremely low dose can cause weight gain in mice and that the increase in adipose tissue mass is due to inhibition of β-adrenergic stimulation of lipolysis. Moreover we have suggested that in addition to its endocrine properties, adipose tissue act as a reservoir for many chemical carcinogens including Polycyclic Aromatic Hydrocarbons (PAHs).     

促酰化蛋白对3T3-L1脂肪细胞炎性反应的影响

目的 观察3T3-L1成熟脂肪细胞胰岛素抵抗状态下促酰化蛋白(acylation stimulating protein,ASP)对炎性因子(IL-6、MCP-1、MIP-1α和TNF-α)分泌水平以及炎性信号因子(JNK1、IKKβ)蛋白表达的影响。 方法 3T3-L1成熟脂肪细胞分别给予不同浓度(0、0.125、0.5、1.0mmol/L)油酸(C18∶ 1)或棕榈酸(C16∶ 0)温育过夜,诱导胰岛素抵抗,在此基础上给予ASP刺激,用ELISA测定IL-6、MCP-1、 MIP-1α和TNF-α 4种细胞炎性因子的分泌水平,采用Western blot法检测JNK1和KKβ蛋白表达。 结果 油酸诱导的胰岛素抵抗下,ASP刺激的脂肪细胞IL-6和MCP-1的分泌水平轻度下调,但差异无统计学意义;而ASP刺激的脂肪细胞MIP-1α和TNF-α的分泌水平显著下降,MIP-1α和TNF-α的分泌分别下调57%(P<0.05)和48%(P<0.05)(1.0mmol/L油酸组)。棕榈酸诱导的胰岛素抵抗下,ASP刺激的脂肪细胞IL-6分泌水平呈下降趋势,最大下调50% IL-6(P<0.05);22% MCP-1(P>0.05),35% MIP-1α(P>0.05)和38%TNF-α(P>0.05)的分泌。高浓度(1.0mmol/L) 油酸和棕榈酸诱导的脂肪细胞胰岛素抵抗状态下,ASP刺激的炎性信号蛋白JNK1蛋白表达显著下调,分别为34%(P<0.05)和54%(P<0.01)。但IKKβ蛋白表达差异无统计学意义。 结论 在脂肪酸诱导的脂肪细胞胰岛素抵抗状态下,ASP在一定程度上调节炎性因子分泌,参与调节JNK、IKK/NF-κB信号通路中重要信号分子的功能,ASP参与了脂肪细胞脂毒性-炎性反应的调控。     

一种新的脂肪细胞源性血管舒张因子

血压调节的研究多年来经过许多变革,最初的工作多集中在各种具有血管活性的神经、内分泌因子及其受体、受体亚型上。上世纪80年代起,随着人们发现内皮源性的一氧化氮可扩张血管,便开始将研究兴趣转向血管内皮的调节作用。此间,研究者并未重视血管外周组织可能具有分泌功能,研究     

人牙周膜细胞体外定向诱导分化为脂肪细胞的研究

目的:体外定向诱导人牙周膜细胞分化为脂肪细胞,探讨人牙周膜细胞的多向分化潜能。方法:体外分离、培养人牙周膜细胞,用地塞米松、吲哚美辛、3-异丁基-1-甲基黄嘌呤(IBMX)和胰岛素诱导其向脂肪细胞定向分化。倒置相差显微镜、透射电子显微镜观察形态结构的变化,采用油红O染色法进行鉴定。结果:经成脂诱导分化培养后6 d,人牙周膜细胞内有脂滴出现。随着诱导培养时间的延长,脂滴逐渐增多并融合成脂泡,细胞由梭形变为圆形和椭圆形,体积增大。透射电子显微镜下见脂泡。油红O染色显示细胞内有中性脂肪形成。结论:人牙周膜细胞在体外可以分化为脂肪细胞,具有多向分化的潜能。     

Exercise-induced galanin release facilitated GLUT4 translocation in adipocytes of type 2 diabetic rats

Although galanin has been shown to increase insulin sensitivity in skeletal muscle of rats, there is no literature available about the effect of galanin on Glucose Transporter 4 (GLUT4) translocation from intracellular membrane pools to plasma membranes in adipocytes of type 2 diabetic rats. In the present study M35, a galanin antagonist was used to elucidate whether exercise-induced galanin release increased GLUT4 translocation in adipocytes of streptozotocin-induced diabetic rats. The present findings showed that plasma galanin levels after swimming training in all four trained groups were higher compared with each sedentary control. M35 treatment had an inhibitory effect on glucose infusion rates in the euglycemic-hyperinsulinemic clamp test and GLUT4 mRNA expression levels in adipocytes. Moreover, M35 treatment reduced GLUT4 concentration in both plasma membranes and total cell membranes. The ratios of GLUT4 contents in plasma membranes to total cell membranes in four drug groups were lower compared with each control. These data demonstrate a beneficial role of endogenous galanin to transfer GLUT4 from internal stores to plasma membranes in adipocytes of type 2 diabetic rats. Galanin plays a significant role in regulation of glucose metabolic homeostasis and is an important hormone relative to diabetes.     

Effect of dehydroepiandrosterone on insulin sensitivity in Otsuka Long-Evans Tokushima-fatty rats

In order to clarify the effect of dehydroepiandrosterone (DHEA) on improvement of insulin resistance, we examined the effects of overexpression of wild-type protein kinase C-ζ (wt-PKCζ)/3-phosphoinositide-dependent protein kinase-1 (wt-PDK1) and kinase-inactive PKCζ/PDK1 (ΔPKCζ/ΔPDK1) on DHEA-induced [³H]2-deoxyglucose (DOG) uptake using the electroporation method in rat adipocytes. Overexpression of wt-PKCζ and wt-PDK1 significantly increased in DHEA-induced [³H]2-DOG uptake. Wortmannin completely suppressed DHEA-induced [³H]2-DOG uptake in wt-PKCζ- and wt-PDK1-transfected adipocytes. Overexpression of neither ΔPKCζ nor ΔPDK1 increased DHEA-induced [³H]2-DOG uptake. Otsuka Long-Evans fatty rats (OLETF), animal models of type 2 diabetes, and Long-Evans Tokushima rats (LETO) as control, were treated with 0.4% DHEA for 2 weeks. Insulin-induced [³H]2-DOG uptakes, activations of PI 3-kinase and PKCζ of adipocytes were significantly increased in DHEA-treated OLETF rats. Moreover, plasma glucose levels in OLETF rats after treatment with DHEA for 2 weeks were significantly lower than treatment without DHEA, but not in LETO rats. These results indicate that DHEA treatment may improve glucose tolerance through a PI 3-kinase-PKCζ pathway and downregulates adiposity in OLETF rats.     

脂肪分泌物对脂肪间充质干细胞的诱导研究

目的研究脂肪分泌物对脂肪间充质干细胞(ADSCs)的诱导作用,探讨脂肪再生机制。方法利用组织块贴壁法和细胞流式技术分离、鉴定ADSCs,收集含有脂肪分泌物的培养基(CM)作为条件培养基并用来诱导ADSCs(CM诱导组),化学培养基诱导组作为阳性对照组,用普通培养基组作为空白对照组,通过细胞形态学观察脂滴的形成和Real-time PCR技术在基因水平上对其可靠性进行体外验证。用可溶性明胶海绵作为ADSCs的支架来研究脂肪分泌物在体内对ADSCs的诱导作用,并作HE切片观察。结果流式细胞术证实所培养的细胞为ADSCs。在体外实验中,CM诱导4d时,CM诱导组中的ADSCs胞浆内出现了明显的脂滴,Real-time PCR显示,与空白对照组比较,相关的成脂基因mRNA表达量也明显增加(P<0.05);体内实验HE染色结果可以观察到CM诱导组中有明显的血管化生成。结论脂肪组织分泌物中可能含有能诱导ADSCs分化形成成熟脂肪细胞以及血管化的目标因子。