Inactivation of primary antioxidant enzymes in mouse keratinocytes by photodynamically generated singlet oxygen
Inactivation of primary antioxidant enzymes in mouse keratinocytes by photodynamically generated singlet oxygen
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DOI：
10.1089/ars.2006.8.1307
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发表时间：
2006-07-01
影响因子：
6.6
通讯作者：
Domann, Frederick E.
中科院分区：
文献类型：
--
作者：
Luo, Jun;Li, Ling;Domann, Frederick E.
细胞抗氧化酶可保护机体免受内源性或外源性促氧化剂的伤害。单线态氧(O-1(2))是一种氧的活性形式，可以在正常和病理生理条件下在体内产生，也可以通过光敏化学物质产生，如在光动力治疗中。我们假设光动力产生的O-1(2)会降低细胞抗氧化剂的酶活性。为了验证这一假设，我们用光敏剂Photofrin加可见光处理培养的小鼠表皮角质形成细胞产生O-1(2)，然后用凝胶内和分光光度测定酶活性测定CuZnSOD、MnSOD和过氧化氢酶的活性。我们的结果表明，角质形成细胞经Photofrin加可见光处理后，细胞内CuZnSOD、MnSOD和过氧化氢酶的活性显著降低。与之相反，单独用Photofrin或仅用可见光处理的对照细胞的CuZnSOD、MnSOD和过氧化氢酶的活性没有受到影响。尽管酶活性水平下降，但所有三种主要抗氧化酶的蛋白质水平在光动力处理后保持不变，正如Western blotting所确定的那样。O-1(2)猝灭剂L-组氨酸对光动力产生的细胞内CuZnSOD、MnSOD和过氧化氢酶的失活具有保护作用。这些实验的结论是，在有核的哺乳动物细胞中，主要的细胞抗氧化酶CuZnSOD、MnSOD和过氧化氢酶可以被光动力产生的O-1(2)灭活。这些发现可能有助于未来抗肿瘤辅助疗法的发展，这种疗法利用光动力产生O-1(2)来灭活抗氧化防御，目的是使肿瘤细胞对促氧化剂生成药物敏感。
Cellular antioxidant enzymes protect against damage caused by exposure to endogenous or exogenous prooxidants. Singlet oxygen (O-1(2)) is a reactive form of oxygen that can be produced in vivo either in normal and pathophysiologic conditions or by photosensitizing chemicals, as during photodynamic treatment. We hypothesized that photodynamically generated O-1(2) would decrease the enzymatic activities of cellular antioxidants. To test this hypothesis, we treated cultured mouse epidermal keratinocytes with the photosensitizer Photofrin plus visible light to produce O-1(2), and then measured CuZnSOD, MnSOD, and catalase activities with both in-gel and spectrophotometric enzyme activity assays. Our results demonstrated that the enzymatic activities of cellular CuZnSOD, MnSOD, and catalase were significantly decreased after keratinocytes were treated with Photofrin plus visible light. By contrast, the enzymatic activities of cellular CuZnSOD, MnSOD, and catalase were unaffected in control cells treated with Photofrin only or visible light only. Despite the decreased levels of enzymatic activities, the protein levels of all three primary antioxidant enzymes remained constant after photodynamic treatment, as determined by Western blotting. L-Histidine, a O-1(2) quencher, protected against the inactivation of cellular CuZnSOD, MnSOD, and catalase enzymes induced by photodynamically generated 10,. The conclusion from these experiments is that the primary cellular antioxidant enzymes CuZnSOD, MnSOD, and catalase can be inactivated by photodynamically generated O-1(2) in nucleated mammalian cells. These findings may be useful in the future development of antineoplastic adjuvant therapies that use photodynamic generation of O-1(2) to inactivate antioxidant defenses with a goal of sensitizing tumor cells to prooxidant-generating drugs.