Prinetti Alessandro, Basso Luisa, Appierto Valentina, Villani Maria Grazia, Valsecchi Manuela, Loberto Nicoletta, Prioni Simona, Chigorno Vanna, Cavadini Elena, Formelli Franca, Sonnino Sandro
Center of Excellence on Neurodegenerative Diseases, Study Center for the Biochemistry and Biotechnology of Glycolipids, Department of Medical Chemistry, Biochemistry and Biotechnology, University of Milan, 20090 Segrate, Italy.
J Biol Chem. 2003 Feb 21;278(8):5574-83. doi: 10.1074/jbc.M207269200. Epub 2002 Dec 16.
In the present work, we studied the effects of fenretinide (N-(4-hydroxyphenyl)retinamide (HPR)), a hydroxyphenyl derivative of all-trans-retinoic acid, on sphingolipid metabolism and expression in human ovarian carcinoma A2780 cells. A2780 cells, which are sensitive to a pharmacologically achievable HPR concentration, become 10-fold more resistant after exposure to increasing HPR concentrations. Our results showed that HPR was able to induce a dose- and time-dependent increase in cellular ceramide levels in sensitive but not in resistant cells. This form of resistance in A2780 cells was not accompanied by the overexpression of multidrug resistance-specific proteins MDR1 P-glycoprotein and multidrug resistance-associated protein, whose mRNA levels did not differ in sensitive and resistant A2780 cells. HPR-resistant cells were characterized by an overall altered sphingolipid metabolism. The overall content in glycosphingolipids was similar in both cell types, but the expression of specific glycosphingolipids was different. Specifically, our findings indicated that glucosylceramide levels were similar in sensitive and resistant cells, but resistant cells were characterized by a 6-fold lower expression of lactosylceramide levels and by a 6-fold higher expression of ganglioside levels than sensitive cells. The main gangliosides from resistant A2780 cells were identified as GM3 and GM2. The possible metabolic mechanisms leading to this difference were investigated. Interestingly, the mRNA levels of glucosylceramide and lactosylceramide synthases were similar in sensitive and resistant cells, whereas GM3 synthase mRNA level and GM3 synthase activity were remarkably higher in resistant cells.
在本研究中，我们研究了阿维A酸（N-(4-羟苯基)视黄酰胺（HPR）），一种全反式维甲酸的羟苯基衍生物，对人卵巢癌A2780细胞鞘脂代谢和表达的影响。A2780细胞对药理学上可达到的HPR浓度敏感，但在暴露于不断增加的HPR浓度后，其耐药性提高了10倍。我们的结果表明，HPR能够在敏感细胞而非耐药细胞中诱导细胞神经酰胺水平呈剂量和时间依赖性增加。A2780细胞的这种耐药形式并非伴随着多药耐药特异性蛋白MDR1 P-糖蛋白和多药耐药相关蛋白的过表达，其mRNA水平在敏感和耐药的A2780细胞中并无差异。HPR耐药细胞的特征是鞘脂代谢整体改变。两种细胞类型中糖鞘脂的总体含量相似，但特定糖鞘脂的表达不同。具体而言，我们的研究结果表明，葡萄糖神经酰胺水平在敏感和耐药细胞中相似，但耐药细胞的特征是乳糖神经酰胺水平的表达比敏感细胞低6倍，神经节苷脂水平的表达比敏感细胞高6倍。耐药A2780细胞的主要神经节苷脂被鉴定为GM3和GM2。我们研究了导致这种差异的可能代谢机制。有趣的是，葡萄糖神经酰胺和乳糖神经酰胺合酶的mRNA水平在敏感和耐药细胞中相似，而GM3合酶mRNA水平和GM3合酶活性在耐药细胞中显著更高。
