Nagler Rafael, Cohen Shiri, Gavish Moshe
Department of Neuroscience, Rappaport Family Institute for Research in the Medical Sciences, Technion-Israel Institute of Technology, 31096, Haifa, Israel.
J Cell Biochem. 2015 Dec;116(12):2786-92. doi: 10.1002/jcb.25221.
Lung cancer is prevalent in cigarette smokers. The mitochondrial membrane translocator protein (TSPO), is thought to protect cells from free radical damage. We examined the effect of cigarette smoke (CS) (containing free radicals) alone and in the presence of saliva (containing redox active free iron), on survival of H1299 lung cancer cells and on their mitochondrial characteristics, and whether TSPO binding was influenced by CS and by saliva. We exposed H1299 cells to CS in the presence/absence of saliva and also characterized TSPO binding in the cells using [3H]PK 11195 as a radioligand. CS induced a significant drop in mitochondrial potential (ΔΨm), while addition of saliva did not lead to further loss of ΔΨm (42.5% vs. 39.85%). Scatchard analysis of the saturation curve of [3H]PK 11195 binding (0.2-6 nM final concentration) yielded a straight-line plot (R = 0.9). Average Bmax value was 3274 ± 787 fmol/mg of protein, and average Kd value was 9.2 ± 1.3 nM. Benzodiazepine diazepam partially prevented decrease in cell survival following exposure to CS and redox active iron containing media (saliva) while benzodiazepine clonazepam did not, indicating that this effect is TSPO-specific. Exposure of cells to CS resulted in alternation of biomolecules expressed by CLs peroxidation, reduction of TSPO binding, and depletion of the mitochondrial potential. This irreversible damage was enhanced in the presence of saliva. All these modulations may result in cellular death increase following CS exposure, enhanced in the presence of saliva.
肺癌在吸烟者中很常见。线粒体膜转运蛋白（TSPO）被认为可以保护细胞免受自由基损伤。我们研究了单独的香烟烟雾（CS）（含有自由基）以及在唾液（含有具有氧化还原活性的游离铁）存在的情况下，对H1299肺癌细胞存活率及其线粒体特征的影响，以及TSPO结合是否受到CS和唾液的影响。我们将H1299细胞在有/无唾液的情况下暴露于CS中，并使用[3H]PK 11195作为放射性配体来表征细胞中的TSPO结合。CS导致线粒体膜电位（ΔΨm）显著下降，而添加唾液并未导致ΔΨm进一步丧失（分别为42.5%和39.85%）。对[3H]PK 11195结合饱和曲线（终浓度0.2 - 6 nM）进行Scatchard分析得到一条直线图（R = 0.9）。平均最大结合量（Bmax）值为3274 ± 787 fmol/mg蛋白质，平均解离常数（Kd）值为9.2 ± 1.3 nM。苯二氮䓬地西泮部分预防了暴露于CS和含氧化还原活性铁的培养基（唾液）后细胞存活率的下降，而苯二氮䓬氯硝西泮则没有，这表明这种作用是TSPO特异性的。细胞暴露于CS导致由CLs过氧化表达的生物分子发生改变、TSPO结合减少以及线粒体膜电位耗竭。在唾液存在的情况下，这种不可逆损伤会增强。所有这些调节可能导致CS暴露后细胞死亡增加，在唾液存在时这种增加更为明显。
