Risé P, Regazzoni C, Caruso D, Colombo C, Galli G, Galli C
Istituto di Scienze Farmacologiche, Università di Milano.
Ital J Biochem. 1995 Jul-Aug;44(4):169-82.
We have supplemented THP-1 cells, a human monocytic leukemia cell line, with arachidonic acid (AA), containing [3H8] AA, 1-25 microM, for up to 24 hours, and explored the time and concentration dependent patterns of incorporation in cell lipid classes and subclasses. Twenty-five microM AA consisted of deuterated AA ([2H8] AA), containing also [3H8] AA. Phospholipids (PL) were separated by HPLC with UV and radiodetection, and the fatty acids (FA) methyl esters were analyzed by GC. [2H8] AA pentafluorobenzyl-esters from individual lipid classes were obtained and analyzed by GC-MS. Incorporation of AA in cell lipids increased linearly with increasing concentrations, whereas 22:4 and 22:5 accumulated only at 25 microM AA. Up to 10 microM AA, more than 95% of the FA was incorporated in PL, whereas at 25 microM AA a significant proportion of the exogenous FA was incorporated in triglycerides (TG) and in diacyl phosphatidylcholine (PC). The time-course of AA incorporation showed that the peak was at 3 hours, with minimal incorporation in TG, in the presence of 5 microM, whereas the peak occurred at 6 hours, with about 50 percent incorporation in TG, with 25 microM. The data indicate that the range of AA concentrations and the time course of the incorporation of this FA in cell structural lipids are critical.
我们用含有[3H8]花生四烯酸（AA）的1 - 25微摩尔花生四烯酸对人单核细胞白血病细胞系THP - 1细胞进行了长达24小时的补充，并探索了其在细胞脂质类别和亚类中掺入的时间和浓度依赖性模式。25微摩尔的AA由氘代AA（[2H8] AA）组成，其中也含有[3H8] AA。通过高效液相色谱结合紫外和放射性检测分离磷脂（PL），并通过气相色谱分析脂肪酸（FA）甲酯。从各个脂质类别中获得[2H8] AA五氟苄酯并通过气相色谱 - 质谱分析。AA在细胞脂质中的掺入量随浓度增加呈线性增加，而22:4和22:5仅在25微摩尔AA时积累。在高达10微摩尔AA时，超过95%的FA掺入PL中，而在25微摩尔AA时，相当一部分外源FA掺入甘油三酯（TG）和二酰基磷脂酰胆碱（PC）中。AA掺入的时间进程表明，在5微摩尔存在下，峰值出现在3小时，TG中的掺入量最少；而在25微摩尔时，峰值出现在6小时，TG中的掺入量约为50%。数据表明，AA浓度范围以及该FA掺入细胞结构脂质的时间进程至关重要。
