White J R, Ishizaka T, Ishizaka K, Sha'afi R
Proc Natl Acad Sci U S A. 1984 Jul;81(13):3978-82. doi: 10.1073/pnas.81.13.3978.
Rat mast cells, passively sensitized with monoclonal mouse IgE antibody, were stimulated with multi-valent antigen, and an increase in cytoplasmic Ca2+ was determined by using the fluorescent probe quin-2. The increase in quin-2 fluorescence reached maximum within 20 sec after the antigen challenge and then gradually declined. A substantial increase in quin-2 fluorescence was observed in the presence of EGTA, indicating that bridging of cell-bound IgE antibody molecules by antigen induced not only Ca2+ influx but also mobilization of intracellular Ca2+. Phosphatidylserine added to the medium enhanced both the antigen-induced histamine release and the increase in quin-2 fluorescence and slowed the rate at which the quin-2 signal returned to basal levels. Both the antigen-induced increase in quin-2 fluorescence and histamine release were inhibited by pretreatment of mast cells with inhibitors of methyltransferases, theophylline, or cromoglycate. It was also found that methyltransferase inhibitors and theophylline inhibited not only stimulus-dependent calcium influx but also release of bound calcium from intracellular stores. Other secretagogues, compound 48/80 (1 microgram/ml) and Ca ionophore A23187 (0.1 microM), induced a rapid increase in cytoplasmic Ca2+ in rat mast cells and subsequent histamine release. In contrast, the cocarcinogenic compound phorbol 12-myristate 13-acetate caused histamine release without increasing the quin-2 fluorescence.
用单克隆小鼠IgE抗体被动致敏的大鼠肥大细胞，用多价抗原刺激，并用荧光探针喹啉-2测定细胞质Ca2+的增加。抗原攻击后20秒内，喹啉-2荧光增加达到最大值，然后逐渐下降。在存在乙二醇双乙醚二胺四乙酸（EGTA）的情况下，观察到喹啉-2荧光有显著增加，这表明抗原使细胞结合的IgE抗体分子桥联不仅诱导了Ca2+内流，还动员了细胞内的Ca2+。添加到培养基中的磷脂酰丝氨酸增强了抗原诱导的组胺释放和喹啉-2荧光的增加，并减缓了喹啉-2信号恢复到基础水平的速率。肥大细胞用甲基转移酶抑制剂、茶碱或色甘酸预处理可抑制抗原诱导的喹啉-2荧光增加和组胺释放。还发现甲基转移酶抑制剂和茶碱不仅抑制刺激依赖性钙内流，还抑制细胞内储存中结合钙的释放。其他促分泌剂，化合物48/80（1微克/毫升）和钙离子载体A23187（0.1微摩尔），在大鼠肥大细胞中诱导细胞质Ca2+迅速增加并随后释放组胺。相比之下，促癌化合物佛波醇12-肉豆蔻酸酯13-乙酸酯引起组胺释放而不增加喹啉-2荧光。
