Mackenzie C G, Mackenzie J B, Reiss O K, Wisneski J A
J Lipid Res. 1970 Nov;11(6):571-82.
Factors responsible for the high lipogenic activity of rabbit serum were investigated using an assay procedure based on the gravimetric determination of the 24 hr increase in cell lipid. Cellular synthesis of fatty acids was inhibited by the presence of serum in the assay medium. Approximately 90% of the increase in cell lipid produced by serum fractions was due to triglyceride accumulation. Fractionation of rabbit serum by precipitation with ammonium sulfate or by ultracentrifugation in high density medium, both indicated that three-quarters of its lipogenic activity was associated with albumin. The lipoproteins prepared by ultracentrifugation also exhibited about one-half the activity of whole serum. The lipogenic activity of albumin was confirmed by the high potency of the albumin isolated in a nearly pure form from proteins of d>1.21 by precipitation with trichloroacetic acid and extraction with ethanol. As judged from chemical and isotopic analysis, neither the lipid content nor the lipid composition of the albumin was appreciably altered during its isolation. Of the albumin-bound lipids, only the free fatty acids, as determined by DEAE column chromatography, were present in an amount sufficient to account for the observed increase in cell triglycerides. In control experiments with horse serum of low lipogenic activity, the proteins of d>1.21 also possessed low activity in conjunction with a low content of free fatty acid. However, the albumin isolated from the latter preparation exhibited the high lipogenic activity of rabbit serum albumin. Chemical and isotopic analysis of the recovered horse serum albumin revealed that its free fatty acid content was the same as that of rabbit serum albumin. These results indicated that the isolation of horse serum albumin was attended by a substantial increase in its free fatty acid content. When the rabbit serum and horse serum content of media were adjusted to provide equivalent concentrations of albumin-bound fatty acids, the rabbit liver cells grown on the former media accumulated more lipid than cells grown on the latter media. This difference was shown to be due to the higher concentration of albumin per micro mole of fatty acid in horse serum as compared with rabbit serum. Consequently, the albumin to fatty acid ratio also controls the lipogenic activity of a serum. A linear relationship is presented which relates the cell lipid content to the molar ratio of albumin to free fatty acids and to the absolute concentration of free fatty acids in the medium.
采用基于重量法测定细胞脂质24小时增加量的检测方法，对导致兔血清高生脂活性的因素进行了研究。检测培养基中血清的存在会抑制脂肪酸的细胞合成。血清组分产生的细胞脂质增加量中，约90% 是由于甘油三酯积累。通过硫酸铵沉淀或在高密度介质中进行超速离心对兔血清进行分级分离，均表明其四分之三的生脂活性与白蛋白有关。通过超速离心制备的脂蛋白也表现出约为全血清活性一半的活性。通过用三氯乙酸沉淀并乙醇萃取从密度大于1.21的蛋白质中分离出的近乎纯的白蛋白具有高效能，从而证实了白蛋白的生脂活性。从化学和同位素分析判断，白蛋白在分离过程中其脂质含量和脂质组成均未发生明显改变。通过DEAE柱色谱法测定，白蛋白结合的脂质中，只有游离脂肪酸的含量足以解释观察到的细胞甘油三酯增加。在低生脂活性的马血清对照实验中，密度大于1.21的蛋白质活性也较低，且游离脂肪酸含量也低。然而，从后一种制剂中分离出的白蛋白表现出兔血清白蛋白的高生脂活性。对回收的马血清白蛋白进行化学和同位素分析表明，其游离脂肪酸含量与兔血清白蛋白相同。这些结果表明，马血清白蛋白的分离伴随着其游离脂肪酸含量的大幅增加。当将培养基中的兔血清和马血清含量调整为提供等量浓度的白蛋白结合脂肪酸时，在前一种培养基上生长的兔肝细胞比在后一种培养基上生长的细胞积累了更多的脂质。结果表明，这种差异是由于与兔血清相比，马血清中每微摩尔脂肪酸的白蛋白浓度更高。因此，白蛋白与脂肪酸的比例也控制着血清的生脂活性。给出了一种线性关系，该关系将细胞脂质含量与白蛋白与游离脂肪酸的摩尔比以及培养基中游离脂肪酸的绝对浓度相关联。
