Role of adipocyte-derived lipoprotein lipase in adipocyte hypertrophy
Role of adipocyte-derived lipoprotein lipase in adipocyte hypertrophy
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DOI：
10.1186/1743-7075-4-22
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发表时间：
2007-10-30
影响因子：
4.5
通讯作者：
Orlando, Robert A.
中科院分区：
文献类型：
--
作者：
Gonzales, Amanda M.;Orlando, Robert A.
背景：脂肪细胞摄取的大部分可用脂肪酸来自于脂蛋白脂酶(LPL)介导的循环脂蛋白颗粒的水解。体内研究旨在确定脂肪细胞来源的LPL在脂肪组织的脂肪储存功能中的确切作用，由于激活内源性脂肪酸合成的代偿机制，一直无法提供确凿的证据。为了解决这一认识上的差距，我们利用成熟的脂肪细胞分化培养模型，测量了降低脂肪细胞LPL表达对细胞内脂质积累的影响。方法：将针对小鼠LPL的siRNA导入3T3-LI脂肪细胞。用实时定量聚合酶链式反应检测LPL的表达，用比色法测定底物(对硝基苯丁酸酯)水解后细胞表面相关LPL酶的活性。定量逆转录聚合酶链式反应检测载脂蛋白CII和CIII的表达。结果：在3T3-LI前体脂肪细胞分化过程中，LPL基因表达增加6倍，细胞表面相关LPL酶活性增加2倍。与LPL表达的增加相平行，我们发现细胞内脂质的增加类似于10倍，这表明脂肪细胞来源的LPL表达与脂肪储存之间存在直接关联。接下来，我们通过siRNA转染降低了脂肪细胞中LPL的表达，以直接量化脂肪细胞来源的LPL对脂肪储存的贡献，这种处理将LPL mRNA的表达和细胞表面相关的LPL酶活性降低到接近未处理对照组的50%，而细胞内脂质水平降低了80%。外源添加纯化的LPL(以恢复细胞外脂肪分解活性)或棕榈酸酯(作为游离脂肪酸的来源)到siRNA处理的细胞中，使细胞内的脂质水平恢复到未处理对照的测量水平。我们还发现，脂肪细胞表达载脂蛋白CII和CIII，此外，在LPL表达降低的细胞中，apoCII/apoCIII的比率基本保持不变。结论：我们提供了脂肪细胞来源的LPL是有效摄取和储存脂肪酸所必需的证据，并且脂肪细胞表达自己的apoCII和apoCIII来源来调节细胞外LPL的活性。这些发现表明，脂肪细胞能够产生必要的酶成分和辅助因子，以有效地储存脂肪，而不依赖于血管来源。
Background: A major portion of available fatty acids for adipocyte uptake is derived from lipoprotein lipase (LPL)-mediated hydrolysis of circulating lipoprotein particles. In vivo studies aimed at identifying the precise role of adipocyte-derived LPL in fat storage function of adipose tissue have been unable to provide conclusive evidence due to compensatory mechanisms that activate endogenous fatty acid synthesis. To address this gap in knowledge, we have measured the effect of reducing adipocyte LPL expression on intracellular lipid accumulation using a well-established cultured model of adipocyte differentiation.Methods: siRNA specific for mouse LPL was transfected into 3T3-LI adipocytes. Expression of LPL was measured by quantitative real-time PCR and cell surface-associated LPL enzymatic activity was measured by colorimetric detection following substrate (p-nitrophenyl butyrate) hydrolysis. Apolipoprotein CII and CIII expression ratios were also measured by qRT- PCR. Intracellular lipid accumulation was quantified by Nile Red staining.Results: During differentiation of 3T3-LI pre-adipocytes, LPL mRNA expression increases 6-fold resulting in a 2-fold increase in cell surface-associated LPL enzymatic activity. Parallel to this increase in LPL expression, we found that intracellular lipids increased similar to 10-fold demonstrating a direct correlation between adipocyte-derived LPL expression and lipid storage. We next reduced LPL expression in adipocytes using siRNA transfections to directly quantify the contributions of adipocyte-derived LPL to lipid storage, This treatment reduced LPL mRNA expression and cell surface-associated LPL enzymatic activity to similar to 50% of non-treated controls while intracellular lipid levels were reduced by 80%. Exogenous addition of purified LPL (to restore extracellular lipolytic activity) or palmitate (as a source of free fatty acids) to siRNA-treated cells restored intracellular lipid levels to those measured for non-treated controls. We also found that adipocytes express apolipoprotein CII and CIII and, in addition, the apoCII/apoCIII ratio remains largely unchanged in cells with reduced LPL expression.Conclusion: We provide evidence that adipocyte-derived LPL is required for efficient fatty acid uptake and storage, and that adipocytes express their own source of apoCII and apoCIII for regulating extracellular LPL activity. These findings demonstrate that adipocytes are capable of producing the necessary enzymatic components and co-factors for efficient lipid storage independent of vascular sources.