Evidence that cyclophilin-A protects cells against oxidative stress
Evidence that cyclophilin-A protects cells against oxidative stress
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DOI：
10.1042/0264-6021:3410127
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发表时间：
1999-07-01
影响因子：
4.1
通讯作者：
Crompton, M
中科院分区：
文献类型：
--
作者：
Doyle, V;Virji, S;Crompton, M
亲环素-A是一个与环孢素A结合的肽基-脯氨酸顺式-非-异构酶家族的胞浆异构体。本研究探讨了亲环素-A在化学缺血和叔丁基氢过氧化氢诱导的坏死性细胞死亡中的作用。18聚体反义硫代寡核苷酸被用来靶向新生大鼠心肌细胞亲环素-a mRNA的翻译区。该寡核苷酸作用24小时后，以免疫学和酶学标准判断，细胞内亲环素-A的含量至少减少了93%。在酶测定中，用荧光法测定了小体积(10亩L)细胞提取液中的肽基-脯氨酸顺式-反式异构酶活性。用抗亲环素-A多克隆抗体等电聚焦和SDS/PAGE制备免疫印迹。亲环素A抑制对氰化物加3-脱氧葡萄糖诱导的细胞死亡无影响。然而，亲环素-A抑制的细胞对叔丁基氢过氧化氢明显更敏感。环孢菌素A在两种类型的细胞中对过氧化氢都有一定的抵抗作用，但在亲环素A抑制的细胞中保护作用更强，其中环孢菌素A使细胞存活时间增加2倍。结论是，两种亲环素亚型以完全不同的方式参与了过氧化氢诱导的细胞死亡。亲环素-A具有保护作用。另一种异构体，可能是线粒体亲环素-D，有一种有害的ColE，因此被环孢菌素A阻断会产生保护作用。
Cyclophilin-A is the cytosolic isoform of a family of peptidyl-proline cis-rr uns-isomerases that bind cyclosporin A. This study investigates the role of cyclophilin-A in necrotic cell death, induced by chemical ischaemia and by t-butylhydroperoxide. An 18-mer antisense phosphorothioate oligodeoxynucleotide was used to target a translated region of cyclophilin-a mRNA in rat neonatal cardiomyocytes. After a 24 h exposure to the oligonucleotide, the amount of cyclophilin-A in the cells was decreased by at least 93% as judged bq immunological and enzymic criteria. For the enzyme assays, peptidyl proline cis-trans-isomerase activity was measured fluorimetrically in small (10 mu l) volumes of cell extract. Immunoblots were developed with a polyclonal anti-cyclophilin-A antibody after sample isoelectric focusing and SDS/PAGE. Cyclophilin-A suppression had no effect on cyanide-plus-3-deoxyglucose-induced cell death. However, cyclophilin-A-suppressed cells were markedly more sensitive to t-butylhydroperoxide. Cyclosporin A conferred some resistance to the peroxide in both types of cell, bur protection was greater in cyclophilin-A-suppressed cells, where cyclosporin A increased the survival time 2-fold. It is concluded that two cyclophilin isoforms are involved, in quite different ways, in peroxide-induced cell death. Cyclophilin-A has a protective role. Another isoform, possibly mitochondrial cyclophilin-D, has a deleterious cole, such that blockade by cyclosporin A leads to protection.