Punnonen E L, Reunanen H
Department of Cell Biology, University of Jyväskylä, Finland.
Exp Mol Pathol. 1990 Feb;52(1):87-97. doi: 10.1016/0014-4800(90)90061-h.
The microtubule inhibitor vinblastine causes accumulation of autophagic vacuoles in many cell types. In hepatocytes, many of the accumulated vacuoles are nascent, which has been interpreted to suggest that vinblastine acts by inhibiting the fusion of hydrolase-containing lysosomes with early autophagic vacuoles. However, our previous results suggested that, in Ehrlich ascites cells, vinblastine causes accumulation mainly of older autophagic vacuoles (AVs). This study was undertaken to further characterize the mode of action of vinblastine in these cells. The vinblastine-accumulated AVs were quantified by electron-microscopic morphometry. In addition, the effects of inhibitors of autophagic segregation (leucine, histidine, and 3-methyladenine) on the vinblastine-induced accumulation of autophagic vacuoles were studied. Protein degradation was measured using [14C]valine. Vinblastine caused accumulation of advanced autophagic vacuoles but did not increase the rate of protein degradation. The volume density of early vacuoles remained at the control level. The amino acids retarded but did not prevent the accumulation of autophagic vacuoles, whereas 3-methyladenine almost completely prevented the accumulation. The results suggest that in Ehrlich ascites cells vinblastine acts by inhibiting the maturation of advanced autophagic vacuoles into residual bodies and by stimulating the formation of new autophagic vacuoles. However, 3-methyladenine almost completely prevents the formation of new autophagic vacuoles in the presence of vinblastine. In conclusion, in Ehrlich ascites cells, vinblastine does not prevent the entry of hydrolases into autophagic vacuoles. This calls into question the importance of microtubules in the transport of lysosomal enzymes into autophagic vacuoles.
微管抑制剂长春碱可使多种细胞类型中自噬泡积累。在肝细胞中，许多积累的空泡是新生的，这被解释为表明长春碱通过抑制含水解酶的溶酶体与早期自噬泡的融合发挥作用。然而，我们之前的结果表明，在艾氏腹水癌细胞中，长春碱主要导致较老的自噬泡（AVs）积累。本研究旨在进一步阐明长春碱在这些细胞中的作用方式。通过电子显微镜形态计量学对长春碱积累的自噬泡进行定量。此外，还研究了自噬分离抑制剂（亮氨酸、组氨酸和3 - 甲基腺嘌呤）对长春碱诱导的自噬泡积累的影响。使用[14C]缬氨酸测量蛋白质降解。长春碱导致晚期自噬泡积累，但未提高蛋白质降解速率。早期空泡的体积密度维持在对照水平。氨基酸可延缓但不能阻止自噬泡积累，而3 - 甲基腺嘌呤几乎完全阻止了积累。结果表明，在艾氏腹水癌细胞中，长春碱通过抑制晚期自噬泡成熟为残余小体以及刺激新的自噬泡形成发挥作用。然而，在长春碱存在的情况下，3 - 甲基腺嘌呤几乎完全阻止了新自噬泡的形成。总之，在艾氏腹水癌细胞中，长春碱并不阻止水解酶进入自噬泡。这对微管在溶酶体酶向自噬泡运输中的重要性提出了质疑。
