Guo Z, Yuan C, Wei-Lavery T, Fang Y, Garvin R A, Nishida H I, Nishida T
The Burnsides Research Laboratory, Department of Food Science & Human Nutrition and Division of Nutritional Sciences, University of Illinois, Urbana, IL 61801, USA.
J Nutr. 1999 Nov;129(11):1984-91. doi: 10.1093/jn/129.11.1984.
Hep G2 cells were used to study the synthesis and secretion of phospholipid transfer protein (PLTP). Upon incubation of the cells at confluence with serum-free Dulbecco's modified Eagle's medium (DMEM), phosphatidylcholine (PC) transfer activity was found to accumulate in the culture media. The PC transfer activity in the media was effectively inhibited by rabbit anti-human PLTP immunoglobulin (Ig)G, thus indicating that the PC transfer activity was due to secreted PLTP. The molecular weight of Hep G2 PLTP was approximately 78 kDa by Western blot analysis, in agreement with the molecular weight obtained for purified human plasma PLTP. The PLTP secreted by Hep G2 also possessed an HDL conversion activity similar to that of human plasma PLTP. The addition of butyrate to the cell culture media resulted in a marked increase in the secretion of PLTP. After 24 h incubation with 4 mmol/L sodium butyrate, a more than twofold increase (P < 0.01) of PC transfer activity in the cell-conditioned media was obtained. The dose-dependent increase in the PC transfer activity in the media upon butyrate treatment was well correlated (r = 0.80, P < 0.01) with that of PLTP mass as determined by immuno-slot blot analysis of cell-conditioned media. The increased secretion of PLTP by Hep G2 treated with sodium butyrate was accompanied by a greater increase in the level of PLTP mRNA in the cells as determined by ribonuclease protection assay. In the presence of 4 mmol/L sodium butyrate, a fourfold increase (P < 0. 01) in mRNA level was obtained at 24 h. No stabilizing effect of butyrate on PLTP mRNA was apparent upon treatment of the cultured cells with the RNA synthesis inhibitor, actinomycin D. Thus, the up-regulatory effect of butyrate on PLTP gene expression seemed to have occurred at the transcriptional level.
采用Hep G2细胞研究磷脂转运蛋白（PLTP）的合成与分泌。将汇合的细胞与无血清的杜尔贝科改良伊格尔培养基（DMEM）一起孵育后，发现培养基中积累了磷脂酰胆碱（PC）转运活性。培养基中的PC转运活性被兔抗人PLTP免疫球蛋白（Ig）G有效抑制，这表明PC转运活性是由分泌的PLTP所致。通过蛋白质印迹分析，Hep G2 PLTP的分子量约为78 kDa，与纯化的人血浆PLTP的分子量一致。Hep G2分泌的PLTP也具有与人血浆PLTP相似的高密度脂蛋白（HDL）转化活性。向细胞培养基中添加丁酸盐导致PLTP分泌显著增加。用4 mmol/L丁酸钠孵育24小时后，细胞条件培养基中的PC转运活性增加了两倍多（P < 0.01）。丁酸盐处理后培养基中PC转运活性的剂量依赖性增加与通过细胞条件培养基的免疫斑点印迹分析测定的PLTP质量增加密切相关（r = 0.80，P < 0.01）。用丁酸钠处理的Hep G2分泌的PLTP增加，同时通过核糖核酸酶保护试验测定的细胞中PLTP mRNA水平也有更大程度的增加。在存在4 mmol/L丁酸钠的情况下，24小时时mRNA水平增加了四倍（P < 0.01）。用RNA合成抑制剂放线菌素D处理培养细胞后，未观察到丁酸盐对PLTP mRNA的稳定作用。因此，丁酸盐对PLTP基因表达的上调作用似乎发生在转录水平。
