Karaczyn Aldona, Ivanov Sergey, Reynolds Mindy, Zhitkovich Anatoly, Kasprzak Kazimierz S, Salnikow Konstantin
Laboratory of Comparative Carcinogenesis, National Cancer Institute at Frederick, Frederick, Maryland 21702, USA.
J Cell Biochem. 2006 Apr 1;97(5):1025-35. doi: 10.1002/jcb.20705.
Exposure of human lung cells to carcinogenic nickel compounds in the presence of oxygen up-regulated carbonic anhydrase IX (CA IX) and NDRG1/Cap43, both known as intrinsic hypoxia markers and cancer-associated genes. This suggests that factors other than a shortage of oxygen may be involved in this induction. Both proteins can also be induced in the presence of oxygen by culturing these cells to a high density without medium change. The intracellular ascorbate measurements revealed its rapid depletion in both metal- and density-exposed cells. Nickel exposure caused strong activation of HIF-1alpha and HIF-2alpha proteins, underscoring activation of HIF-1-dependent transcription. In contrast, cell density-dependent transcription was characterized by minor induction of HIF-1alpha or HIF-2alpha. Moreover, the up-regulation of NDRG1/Cap43 in HIF-1alpha deficient fibroblasts suggested the involvement of different transcription factor(s). The repletion of intracellular ascorbate reversed the induction of CA IX and NDRG1/Cap43 caused by cell density or nickel exposure. Thus, the loss of intracellular ascorbate triggered the induction of both tumor markers by two different conditions in the presence of oxygen. Ascorbate is delivered to lung cells via the SVCT2 ascorbate transporter, which was found to be sensitive to nickel or cell density. Collectively these findings establish the importance of intracellular ascorbate levels for the regulation of expression of CA IX and NDRG1/Cap43. We suggest, that, in addition to low oxygenation, insufficient supply of ascorbate or its excessive oxidation in tumors, can contribute to the induction of hypoxia-associated proteins via both HIF-dependent and independent mechanisms.
在有氧条件下，将人肺细胞暴露于致癌镍化合物中，会使碳酸酐酶IX（CA IX）和NDRG1/Cap43上调，这两种蛋白均为已知的内源性缺氧标志物和癌症相关基因。这表明除缺氧外的其他因素可能参与了这种诱导过程。通过将这些细胞高密度培养且不更换培养基，在有氧条件下也可诱导这两种蛋白。细胞内抗坏血酸的测量结果显示，在金属暴露和密度暴露的细胞中其含量均迅速减少。镍暴露导致HIF-1α和HIF-2α蛋白强烈激活，突出了HIF-1依赖性转录的激活。相比之下，细胞密度依赖性转录的特征是HIF-1α或HIF-2α的诱导作用较小。此外，在HIF-1α缺陷的成纤维细胞中NDRG1/Cap43的上调表明存在不同的转录因子参与其中。细胞内抗坏血酸的补充逆转了由细胞密度或镍暴露引起的CA IX和NDRG1/Cap43的诱导。因此，细胞内抗坏血酸的缺失在有氧条件下通过两种不同情况触发了这两种肿瘤标志物的诱导。抗坏血酸通过SVCT2抗坏血酸转运体输送到肺细胞，该转运体对镍或细胞密度敏感。这些发现共同确立了细胞内抗坏血酸水平对CA IX和NDRG1/Cap43表达调控的重要性。我们认为，除了低氧状态外，肿瘤中抗坏血酸供应不足或其过度氧化，可通过HIF依赖性和非依赖性机制促进缺氧相关蛋白的诱导。
