Protection of acute myeloblastic leukemia cells against apoptotic cell death by high glutathione and gamma-glutamylcysteine synthetase levels during etoposide-induced oxidative stress.
Protection of acute myeloblastic leukemia cells against apoptotic cell death by high glutathione and gamma-glutamylcysteine synthetase levels during etoposide-induced oxidative stress.
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在依托泊苷诱导的氧化应激过程中，高谷胱甘肽和γ-谷氨酰半胱氨酸合成酶水平可保护急性髓细胞白血病细胞免受细胞凋亡。
DOI：
10.1023/a:1008382912096
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发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Koistinen,P
中科院分区：
文献类型：
--
作者：
Siitonen,T;Alaruikka,P;Mäntymaa,P;Savolainen,ER;Kavanagh,TJ;Krejsa,CM;Franklin,CC;Kinnula,V;Koistinen,P
背景足叶乙甙通过诱导细胞凋亡介导其细胞毒作用。因此，调节细胞凋亡的机制也应该影响耐药性。氧化剂和抗氧化剂已被证明参与细胞凋亡的调节。我们有兴趣在研究是否急性髓细胞白血病（AML）细胞依托泊苷的反应是介导的氧化应激和谷胱甘肽levels.Patients和methodsTwo亚克隆的OCI/AML-2细胞系，这是依托泊苷敏感（ES），和依托泊苷耐药（ER），建立了作者在大学的欧卢，并用作模型。细胞凋亡的测定包括磷脂酰丝氨酸的外化（如膜联蛋白V结合所证明的），和胱天蛋白酶活化（如聚（ADP-核糖）聚合酶的裂解所指示的（蛋白质印迹法）。通过流式细胞术分析过氧化物形成。谷氨酰胺和γ-谷氨酰半胱氨酸合成酶（γ-GCS）的水平分别通过免疫印迹法和Westernblotting. ResultsEtoposide诱导的细胞凋亡是明显的ES亚克隆治疗后12小时，但在ER亚克隆后24小时明显。ER亚克隆的谷胱甘肽和γ-GCS水平高于ES亚克隆。足叶乙甙增加过氧化物的形成在两个亚克隆12小时后曝光。在依托泊苷暴露期间，ES亚克隆中观察到谷胱甘肽显着消耗，而ER亚克隆中的谷胱甘肽水平保持不变。在暴露于依托泊苷24小时期间，在两个亚克隆中均未观察到γ-GCS诱导。此外，γ-GCS的催化亚基在凋亡过程中被切割，同时细胞内谷胱甘肽耗尽。当谷胱甘肽被耗尽的治疗与buthiflavine sulfoximine，γ-GCS的直接抑制剂，依托泊苷的敏感性增加，特别是在ER亚clonel.ConclusionsThe结果强调的意义，谷胱甘肽的生物合成的AML细胞依托泊苷的反应。依托泊苷暴露期间介导谷胱甘肽耗竭的分子机制可能包括γ-GCS催化亚基的裂解。
BackgroundEtoposide mediates its cytotoxicity by inducing apoptosis. Thus, mechanisms which regulate apoptosis should also affect drug resistance. Oxidants and antioxidants have been shown to participate in the regulation of apoptosis. We were interested in studying whether responsiveness of acute myeloblastic leukemia (AML) cells to etoposide is mediated by oxidative stress and glutathione levels.Patients and methodsTwo subclones of the OCI/AML-2 cell line which are etoposide-sensitive (ES), and etoposideresistant (ER), were established by the authors at the University of Oulu, and used as models. Assays for apoptosis included externalization of phosphatidylserine (as evidenced by annexin V binding), and caspase activation as indicated by cleavage of poly(ADP-ribose)polymerase (Western blotting). Peroxide formation was analyzed by flow cytometry. Glutathione and gamma-glutamylcysteine synthetase (γ-GCS) levels were determined spectrophotometrically and by Western blotting, respectively.ResultsEtoposide-induced apoptosis was evident 12 hours after treatment in the ES subclone, but was apparent in the ER subclone only after 24 hours. The basal glutathione and γ-GCS levels were higher in the ER than the ES subclone. Etoposide increased peroxide formation in both subclones after 12-hour exposure. Significant depletion of glutathione was observed in the ES subclone during etoposide exposure, while glutathione levels were maintained in the ER subclone. In neither of the subclones was induction of γ-GCS observed during 24-hour exposure to etoposide. Furthermore, the catalytic subunit of γ-GCS was cleaved during apoptosis, concurrent with depletion of intracellular glutathione. When glutathione was depleted by treatment with buthionine sulfoximine, a direct inhibitor of γ-GCS, the sensitivity to etoposide was increased, particularly in the ER subclone.ConclusionsThe results underline the significance of glutathione biosynthesis in the responsiveness of AML cells to etoposide. The molecular mechanisms mediating glutathione depletion during etoposide exposure might include the cleavage of the catalytic subunit of γ-GCS.
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影响因子：
8.8
作者：
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DOI：
--
发表时间：
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期刊：
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