PHORBOL ESTERS AND HORSERADISH-PEROXIDASE STIMULATE PINOCYTOSIS AND REDIRECT THE FLOW OF PINOCYTOSED FLUID IN MACROPHAGES
PHORBOL ESTERS AND HORSERADISH-PEROXIDASE STIMULATE PINOCYTOSIS AND REDIRECT THE FLOW OF PINOCYTOSED FLUID IN MACROPHAGES
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DOI：
10.1083/jcb.100.3.851
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发表时间：
1985-01-01
影响因子：
7.8
通讯作者：
SILVERSTEIN, SC
中科院分区：
文献类型：
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作者：
SWANSON, JA;YIRINEC, BD;SILVERSTEIN, SC
Lucifer Yellow CH （LY）是一种很好的液相胞饮检测探针。它在巨噬细胞空泡系统内积累，不降解，并且在6.0 mg/ml浓度下无毒。其摄取被抑制在0度。C.巯基乙酸酯诱导的小鼠腹腔巨噬细胞对LY呈曲线摄取动力学。在培养基中加入LY后，有一段非常快速的细胞积累（在1 mg/ml LY下，每小时1400 ng LY/mg蛋白）。这种积累速率与胞饮作用引起的液体流入速率最为接近。在60分钟内，LY的积累速率减慢到250 ng/mg / h的稳态速率，然后持续到18 h。脉冲追踪实验显示，稳态条件下积累速率的降低是由于LY的外排。进入细胞的LY只有20%被保留；其余的被释放回介质中。射流有快、慢两部分；每一种反应在动力学上都可以表征为一级反应。其动力学与Besterman等人所描述的相似，Besterman等人将液相胞饮作用解释为至少涉及2个室，一个小的，快速周转的室，另一个明显较大的，填充和清空缓慢的室。为了寻找控制细胞内液体运输的过程，研究了用辣根过氧化物酶（HRP）或肿瘤启动子肉豆酸酯磷酸酯（PMA）处理巨噬细胞后的胞饮作用。HRP常被用作液相胞饮的标记物，观察到它能刺激巨噬细胞中LY的积累速度。PMA使LY积累速率立即增加4- 7倍。HRP和PMA都通过刺激内流和减少快速循环的内化流体的百分比来增加LY积累。更大比例的内吞液体进入缓慢排空的腔室（推测为溶酶体）。由于细胞的大量外排，标记物积累的测量不能准确地估计液体胞饮症的发生率。通过细胞质的水和溶质的胞浆流动在脂质体形成以外的点受到调节。
Lucifer Yellow CH (LY) is an excellent probe for fluid-phase pinocytosis. It accumulates within the macrophage vacuolar system, is not degraded, and is not toxic at concentrations of 6.0 mg/ml. Its uptake is inhibited at 0.degree. C. Thioglycollate-elicited mouse peritoneal macrophages were found to exhibit curvilinear uptake kinetics of LY. Upon addition of LY to the medium, there was a brief period of very rapid cellular accumulation of the dye (1400 ng of LY/mg protein per h at 1 mg/ml LY). This rate of accumulation most closely approximates the rate of fluid influx by pinocytosis. Within 60 min, the rate of LY accumulation slowed to a steady-state rate of 250 ng/mg protein per h which then continued for up to 18 h. Pulse-chase experiments revealed that the reduced rate of accumulation under steady-state conditions was due to efflux of LY. Only 20% of LY taken into the cells was retained; the remainder was released back into the medium. Efflux has 2 components, rapid and slow; each can be characterized kinetically as a 1st-order reaction. The kinetics are similar to those described by Besterman et al. who interpret fluid-phase pinocytosis as involving at least 2 compartments, one small, rapidly turnover over compartment and another apparently larger one which fills and empties slowly. To search for processes that control intracellular fluid traffic, pinocytosis was studied after treatment of macrophages with horseradish peroxidase (HRP) or with the tumor promoter phorbol myristate acetate (PMA). HRP, often used as a marker for fluid-phase pinocytosis, was observed to stimulate the rate of LY accumulation in macrophages. PMA caused an immediate 4- to 7-fold increase in the rate of LY accumulation. Both HRP and PMA increased LY accumulation by stimulating influx and reducing the percentage of internalized fluid that is rapidly recycled. A greater proportion of endocytosed fluid passes into the slowly emptying compartment (presumed lysosomes). Because of the considerable efflux by cells, measurement of marker accumulation inaccurately estimates the rate of fluid pinocytosis. Pinocytic flow of water and solutes through cytoplasm is subject to regulation at points beyond the formation of pinosomes.