Fulbright R M, Axelrod D, Dunham W R, Marcelo C L
Biophysics Research Division, University of Michigan, Ann Arbor 48109-1055, USA.
Exp Cell Res. 1997 May 25;233(1):128-34. doi: 10.1006/excr.1997.3553.
The fluorescent probe diI was used to study the lateral mobility of lipids in in vitro strains of living adult human keratinocytes grown in four different media. One medium was essential fatty acid deficient (EFAD) and low in calcium ion, a medium known to yield cells that proliferate rapidly and contain lipid with extremely low levels of essential fatty acids. Two other media were supplemented with essential fatty acids (FAS), media that are known to result in cells that grow more slowly and have normalized fatty acid proportions. A fourth medium consisted of 1 microM all-trans-retinoic acid added to the fatty acid-supplemented medium (FAS-RA), a medium known to produce cells that are highly proliferative, with a growth rate greater than that of the FAS strains and similar to that of the EFAD strains. The keratinocytes grown in these four media were studied using the fluorescence recovery after photobleaching (FRAP) technique to determine the lateral diffusion rate of diI in the plasma membranes. Our results showed a positive correlation between growth rate and diffusion coefficient (D): the diffusion coefficient of diI was higher in the EFAD or FAS-RA cells than in the FAS cells. The measurement of D among the FAS cells fell into two groups. One group was similar to the single group seen in the EFAD cells, but the other group was composed of much lower D values. The other FRAP parameters (mobile fraction and bleach depth) were larger in the "slow" group than in the "fast" group. This trend of negative correlation between these parameters and D was also found within the fast group. These results are interpreted in terms of possible changes in membrane structure or morphology that might be indirectly associated with the fatty acid alterations, including the possible presence of areas in senescing keratinocytes where plasma membranes collapse to form an interacting system of lipid bilayers.
荧光探针二碘甲磺酸钠（diI）用于研究在四种不同培养基中培养的成人活体角质形成细胞体外菌株中脂质的侧向流动性。一种培养基缺乏必需脂肪酸（EFAD）且钙离子含量低，这种培养基已知会产生快速增殖且含有极低水平必需脂肪酸的脂质的细胞。另外两种培养基添加了必需脂肪酸（FAS），已知这种培养基会导致细胞生长更缓慢且脂肪酸比例正常化。第四种培养基是在添加脂肪酸的培养基（FAS-RA）中添加1微摩尔全反式维甲酸，这种培养基已知会产生高度增殖的细胞，其生长速率大于FAS菌株且与EFAD菌株相似。使用光漂白后荧光恢复（FRAP）技术研究在这四种培养基中生长的角质形成细胞，以确定diI在质膜中的侧向扩散速率。我们的结果表明生长速率与扩散系数（D）之间呈正相关：diI在EFAD或FAS-RA细胞中的扩散系数高于FAS细胞。FAS细胞中D的测量结果分为两组。一组与EFAD细胞中看到的单一组相似，但另一组由低得多的D值组成。“慢”组中的其他FRAP参数（可移动部分和漂白深度）比“快”组中的大。在快速组中也发现了这些参数与D之间的这种负相关趋势。这些结果可根据膜结构或形态的可能变化来解释，这些变化可能与脂肪酸改变间接相关，包括在衰老角质形成细胞中可能存在质膜塌陷形成脂质双层相互作用系统的区域。
