Mills J W, Lubin M
Am J Physiol. 1986 Feb;250(2 Pt 1):C319-24. doi: 10.1152/ajpcell.1986.250.2.C319.
We examined the effect of N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (DBcAMP) on the volume and cytoskeleton of confluent cultures of Madin-Darby canine kidney (MDCK) cells. A 90-min exposure to 1 mM DBcAMP resulted in a 20% reduction in volume as measured by [14C]-urea water space. The volume in cells exposed to isobutylmethylxanthine (IBMX, 0.1 mM) was reduced by 24%. In control cultures F-actin, revealed by staining with nitrobenzoxadiazole-phallacidin, was found at the base of the cell as fibers, at the junctional region as a circumferential band, and on the apical cell surface as a mottled fluorescence. A dense pattern of microtubules, revealed by indirect immunofluorescence, was seen throughout the cell. Exposure to DBcAMP for 90 min resulted in a change of F-actin fibers into dense bundles near the periphery of the cell. This effect was even more striking when cells were exposed to IBMX. Cytochalasin B disrupted F-actin and resulted in a volume reduction similar to that in DBcAMP. Neither DBcAMP nor IBMX affected the distribution of microtubules. Moreover, colchicine, which completely disrupted the microtubules, did not change MDCK cell volume. The results suggest that DBcAMP and F-actin play a role in volume control in MDCK cells.
我们研究了N6,2'-O-二丁酰腺苷3',5'-环一磷酸（DBcAMP）对汇合的麦迪逊-达比犬肾（MDCK）细胞培养物的体积和细胞骨架的影响。通过[14C]-尿素水空间测量，暴露于1 mM DBcAMP 90分钟导致体积减少20%。暴露于异丁基甲基黄嘌呤（IBMX，0.1 mM）的细胞体积减少了24%。在对照培养物中，用硝基苯并恶二唑-鬼笔环肽染色显示的F-肌动蛋白在细胞底部呈纤维状，在连接区域呈环形带，在细胞顶端表面呈斑驳荧光。通过间接免疫荧光显示，整个细胞中可见密集的微管模式。暴露于DBcAMP 90分钟导致F-肌动蛋白纤维在细胞周边附近变成密集束。当细胞暴露于IBMX时，这种效应更加明显。细胞松弛素B破坏F-肌动蛋白并导致体积减少，类似于DBcAMP处理的情况。DBcAMP和IBMX均未影响微管的分布。此外，完全破坏微管的秋水仙碱并未改变MDCK细胞体积。结果表明，DBcAMP和F-肌动蛋白在MDCK细胞的体积控制中起作用。
