IMMUNOLIPOSOMES WITH DIFFERENT ACID SENSITIVITIES AS PROBES FOR THE CELLULAR ENDOCYTIC PATHWAY
IMMUNOLIPOSOMES WITH DIFFERENT ACID SENSITIVITIES AS PROBES FOR THE CELLULAR ENDOCYTIC PATHWAY
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DOI：
10.1016/0005-2736(89)90453-7
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发表时间：
1989-12-11
期刊：
影响因子：
--
通讯作者：
HUANG, L
中科院分区：
文献类型：
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作者：
COLLINS, D;MAXFIELD, F;HUANG, L
将二油酰磷脂酰乙醇胺（DOPE）与油酸（OA）、棕榈酰同型半胱氨酸（PHC）或二棕榈酰琥珀酰甘油（DPSG）结合，制备了具有不同酸敏感性的ph敏感脂质体。DOPE/OA脂质体对酸最敏感，而DOPE/DPSG脂质体对酸最不敏感。与小鼠L929细胞孵育的DOPE/OA脂质体通过改变脂质组成降低了这些脂质体的ph敏感性。利用白喉毒素片段A作为细胞质传递的标记物，我们发现ph敏感性免疫脂质体的传递动力学与脂质体修饰的酸敏感性密切相关。免疫脂质体内化后5-15分钟的1/2达到pH 6-6.2。相比之下，内体酸化到pH 5.0需要更长的时间（t1/2 .apprxeq）。25分钟)。我们还使用了全细胞零点技术（Yamishiro和Maxfield (1987) J. cell Biol. 105, 2713-2721）来直接测定内噬免疫脂质体所遇到的平均pH值。我们发现，零点法确定的酸化过程比DTA数据估计的酸化速度要慢。这很可能是由于测量的DTA递送是由那些首先到达具有足够酸度的内体的脂质体完成的。我们的数据表明，DOPE/PHC免疫脂质体在早期内体传递，而DOPE/DPSG免疫脂质体在晚期内体传递。具有改变的组成和酸敏感性的DOPE/OA免疫脂质体通过其他两种免疫脂质体之间的动力学中间体传递。因此，ph敏感脂质体是研究核内体酸化动力学的有用探针。
By combining dioleoylphosphatidylethanolamine (DOPE) with oleic acid (OA), palmitoylhomocysteine (PHC) or dipalmitoylsuccinylglycerol (DPSG) we have prepared pH-sensitive liposomes with different acid sensitivities. DOPE/OA liposomes are the most acid sensitive, while DOPE/DPSG liposomes are the least acid sensitive. Incubation of DOPE/OA liposomes with mouse L929 cells reduces the pH-sensitivity of these liposomes by altering the lipid composition. Using diphtheria toxin fragment A as a marker for cytoplasmic delivery, we find that the delivery kinetics of pH-sensitive immunoliposomes closely correlates with the modified acid sensitivities of the liposomes. Immunoliposomes encounter pH 6-6.2 with a t1/2 of 5-15 min after internalization. By contrast, acidification of the endosomes to pH 5.0 takes longer (t1/2 .apprxeq. 25 min). We also used a whole cell null point technique (Yamishiro and Maxfield (1987) J. Cell Biol. 105, 2713-2721) to directly determine the average pH encountered by the endocytosed immunoliposomes. We find that acidification determined by the null point method proceeds less rapidly than that estimated from DTA delivery data. This is likely to the fact that the measured DTA delivery is done by those liposomes which first arrive at the endosomes with sufficient acidity. Our data suggests that DOPE/PHC immunoliposomes deliver at the early endosome while DOPE/DPSG immunoliposomes deliver at the late endosomes. The DOPE/OA immunoliposomes, with the altered composition and acid sensitivity, deliver with a kinetics intermediate between the other two immunoliposomes. Thus, pH-sensitive liposomes represents useful probes for studying the kinetics of endosome acidification.