Michihara Akihiro, Toda Ken, Kubo Takuo, Fujiwara Yoshiteru, Akasaki Kenji, Tsuji Hiroshi
Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Hiroshima, Japan.
Biol Pharm Bull. 2005 Jun;28(6):947-51. doi: 10.1248/bpb.28.947.
Chloroquine has been used as an anti-malarial drug and is known as a lysosomotropic amine as well. The effects of chloroquine on lysosomal integrity in cultured rat hepatocytes were studied by measuring lysosomal enzyme beta-glucuronidase (beta-G) or lysosomal membrane glycoprotein (lamp-1) in Percoll density gradient fractions, in the cytosolic fraction obtained from cells permeabilized by digitonin or in the cytosolic fraction obtained by conventional cell fractionation. The distribution of beta-G on a Percoll density gradient in chloroquine-treated cells was approximately similar to that of a cytosolic protein, mevalonate pyrophosphate decarboxylase, in nontreated cells. Lamp-1 was decreased in the lysosomal fractions on a Percoll density gradient in chloroquine-treated cells, and was increased in the plasma membrane fraction, as compared with the levels in nontreated cells. Furthermore, after cells were cultured in the presence and absence of chloroquine, the proportions of beta-G activity in the cytosolic fraction obtained from the digitonin-permeabilized cells were 19% and 4%, while those in the cytosolic fraction obtained by conventional cell fractionation were 54% and 26%, respectively. From these findings, we infer that chloroquine caused the disruption of lysosomes in the living cells, and that lysosomes treated with chloroquine were easily disrupted by homogenization or centrifugation during cell fractionation.
氯喹一直被用作抗疟药物，同时也被认为是一种溶酶体亲和胺。通过测量Percoll密度梯度分级分离物中的溶酶体酶β-葡萄糖醛酸酶（β-G）或溶酶体膜糖蛋白（lamp-1）、从经洋地黄皂苷通透处理的细胞中获得的胞质部分或通过常规细胞分级分离获得的胞质部分，研究了氯喹对培养的大鼠肝细胞溶酶体完整性的影响。氯喹处理细胞中β-G在Percoll密度梯度上的分布与未处理细胞中胞质蛋白甲羟戊酸焦磷酸脱羧酶的分布大致相似。与未处理细胞相比，氯喹处理细胞中Percoll密度梯度上溶酶体部分的lamp-1减少，而质膜部分的lamp-1增加。此外，在有和没有氯喹存在的情况下培养细胞后，从经洋地黄皂苷通透处理的细胞中获得的胞质部分中β-G活性的比例分别为19%和4%，而通过常规细胞分级分离获得的胞质部分中β-G活性的比例分别为54%和26%。从这些发现中，我们推断氯喹导致活细胞中的溶酶体破坏，并且用氯喹处理的溶酶体在细胞分级分离过程中容易被匀浆或离心破坏。
