Superoxide-Generating Nox5α Is Functionally Required for the Human T-Cell Leukemia Virus Type 1-Induced Cell Transformation Phenotype.
Superoxide-Generating Nox5α Is Functionally Required for the Human T-Cell Leukemia Virus Type 1-Induced Cell Transformation Phenotype.
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人类 T 细胞白血病病毒 1 型诱导的细胞转化表型在功能上需要产生超氧化物的 Nox5α。
DOI：
10.1128/jvi.00983-15
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发表时间：
2015
期刊：
影响因子：
--
通讯作者：
Kamata T.
中科院分区：
文献类型：
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作者：
Shigemura T;Shiohara M;Kato M;Furuta S;Kaneda K;Morishita K;Hasegawa H;Fujii M;Gorlach A;Koike K;Kamata T.
材料和方法细胞系和试剂。 HTLV-1 感染的 T 细胞系（MT1、MT2、MT4 和 HUT102）(8, 21)、HTLV-1 未感染的 T 细胞系（HUT78、H9、Jurkat、Molt-4 和 Molt-17）(21)、HTLV-II 感染的细胞系 (Mot) 和 Bcr-Abl 阳性骨髓性白血病细胞品系(K562)维持在补充有10%胎牛血清(FBS)的RPMI 1640中。二苯基碘鎓 (DPI)、n-乙酰半胱氨酸 (NAC)、吡咯烷二硫代氨基甲酸酯 (PDTC)、AG490、STI-571（伊马替尼）和 STAT5 抑制剂购自 Calbiochem。兔抗 Nox5 抗体如前所述制备 (22) 或购自 Novus Biologicals。针对磷酸-AKT (Thr308)、AKT、磷酸细胞外信号调节激酶 (ERK)(Thr202/Tyr204)、ERK、STAT5 和磷酸-STAT5 (Tyr694) 的抗体购自 Cell Signaling。 pGD210-Bcr-Abl 由 T. Tauchi 和 pcDNA3 提供。 1-myc-STAT5B-CA，作者：K. Ikuta。人类标本。研究方案经信州大学人类伦理审查委员会批准，所有样本均在获得患者知情同意后采集。如前所述，通过 Ficoll-Hypaque 梯度离心 (Amersham Bioscience) 纯化来自健康志愿者和 ATL 患者的外周血单核细胞 (PBMC) (8)。
MATERIALS AND METHODSCell lines and reagents. HTLV-1-infected T-cell lines (MT1, MT2, MT4, and HUT102)(8, 21), HTLV-1-uninfected T-cell lines (HUT78, H9, Jurkat, Molt-4, and Molt-17)(21), a HTLV-II-infected cell line (Mot) and a Bcr-Abl-positive myeloid leukemia cell line (K562) were maintained in RPMI 1640 supplemented with 10% fetal bovine serum (FBS). Diphenyleniodonium (DPI), n-acetyl cysteine (NAC), pyrrolidine dithiocarbamate (PDTC), AG490, STI-571 (imatinib), and STAT5 inhibitor were purchased from Calbiochem. Rabbit anti-Nox5 antibodies were produced as described previously (22) or purchased from Novus Biologicals. Antibodies against phospho-AKT (Thr308), AKT, phospho-extracellular signal-regulated kinase (ERK)(Thr202/Tyr204), ERK, STAT5, and phospho-STAT5 (Tyr694) were purchased from Cell Signaling. pGD210-Bcr-Abl was provided by T. Tauchi and pcDNA3. 1-myc-STAT5B-CA by K. Ikuta.Human specimens. The study protocol was approved by the Human Ethics Review Committee of Shinshu University, and all samples were collected after obtaining informed consent from patients. Peripheral blood mononuclear cells (PBMC) from healthy volunteers and ATL patients were purified by Ficoll-Hypaque gradient centrifugation (Amersham Bioscience) as described previously (8).