Various liver diseases, including hepatocellular carcinoma (HCC), have been linked to mitochondrial dysfunction, reduction of reactive oxygen species (ROS) and elevation of nitric oxide (NO). In this study, we subjected the human liver mitochondrial proteome to an extensive quantitative proteomic profiling analysis and molecular characterization to identify potential signatures indicative of cancer cell growth and progression. A sequential proteomic analysis identified 2452 mitochondrial proteins, of which 1464 and 2010 were classified as non-tumor and tumor (HCC) mitochondrial proteins, respectively, with 1022 overlaps. Further metabolic mapping of the HCC mitochondrial proteins narrowed our biological characterization to four proteins, namely ALDH4A1, LRPPRC, ATP5C1 and ALDH6A1. The latter protein, a mitochondrial methylmalonate semialdehyde dehydrogenase (ALDH6A1), was most strongly suppressed in HCC tumor regions (~10-fold decrease) in contrast to LRPPRC (~6-fold increase), and was predicted to present in plasma. Accordingly, we selected ALDH6A1 for a functional analysis and engineered Hep3B cells to overexpress this protein, called ALDH6A1-O/E cells. Since ALDH6A1 is predicted to be involved in mitochondrial respiration, we assessed changes in the levels of NO and ROS in the overexpressed cell lines. Surprisingly, in ALDH6A1-O/E cells, NO was decreased nearly 50% but ROS was increased at the similar level, while the former was restored by treatment with S-nitroso-N-acetyl-penicillamine. The lactate levels were also decreased relative to control cells. Propidium iodine staining suggested that the decrease in NO and increase in ROS in ALDH6A1-O/E cells could be caused by depolarization of the mitochondrial membrane potential (ΔΨ). Taken together, our results suggest that hepatic neoplastic transformation appear to suppress the expression of ALDH6A1, which is accompanied by a respective increase and decrease in NO and ROS in cancer cells. Given the close link between ALDH6A1 suppression and abnormal cancer cell growth, this protein may serve as a potential molecular signature or biomarker of hepatocarcinogenesis and treatment responses. Mass spectrometry data are available via ProteomeXchange with identifier PXD014252.

中文翻译：

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通过线粒体蛋白质组的定量分析鉴定 ALDH6A1 作为肝细胞癌的潜在分子特征。

包括肝细胞癌 (HCC) 在内的各种肝脏疾病与线粒体功能障碍、活性氧 (ROS) 减少和一氧化氮 (NO) 升高有关。在这项研究中，我们对人类肝脏线粒体蛋白质组进行了广泛的定量蛋白质组学分析和分子表征，以确定指示癌细胞生长和进展的潜在特征。连续蛋白质组学分析鉴定出 2452 种线粒体蛋白，其中 1464 和 2010 种分别被归类为非肿瘤和肿瘤 (HCC) 线粒体蛋白，有 1022 种重叠。HCC 线粒体蛋白的进一步代谢图谱将我们的生物学特征缩小到四种蛋白质，即 ALDH4A1、LRPPRC、ATP5C1 和 ALDH6A1。后一种蛋白质，一种线粒体甲基丙二酸半醛脱氢酶 (ALDH6A1)，与 LRPPRC（​​增加约 6 倍）相比，在 HCC 肿瘤区域中受到最强烈的抑制（减少约 10 倍），并且预计存在于血浆中。因此，我们选择 ALDH6A1 进行功能分析，并改造 Hep3B 细胞以过度表达这种蛋白质，称为 ALDH6A1-O/E 细胞。由于预计 ALDH6A1 参与线粒体呼吸，我们评估了过表达细胞系中 NO 和 ROS 水平的变化。令人惊讶的是，在 ALDH6A1-O/E 细胞中，NO 减少了近 50%，但 ROS 以相似的水平增加，而前者通过 S-亚硝基-N-乙酰青霉胺处理恢复。相对于对照细胞，乳酸水平也降低了。碘化丙啶染色表明 ALDH6A1-O/E 细胞中 NO 的减少和 ROS 的增加可能是由线粒体膜电位 (ΔΨ) 的去极化引起的。总之，我们的结果表明肝脏肿瘤转化似乎抑制了 ALDH6A1 的表达，这伴随着癌细胞中 NO 和 ROS 的相应增加和减少。鉴于 ALDH6A1 抑制与异常癌细胞生长之间的密切联系，这种蛋白质可能作为肝癌发生和治疗反应的潜在分子特征或生物标志物。质谱数据可通过 ProteomeXchange 获得，标识符为 PXD014252。我们的研究结果表明，肝脏肿瘤转化似乎抑制了 ALDH6A1 的表达，这伴随着癌细胞中 NO 和 ROS 的相应增加和减少。鉴于 ALDH6A1 抑制与异常癌细胞生长之间的密切联系，这种蛋白质可能作为肝癌发生和治疗反应的潜在分子特征或生物标志物。质谱数据可通过 ProteomeXchange 获得，标识符为 PXD014252。我们的研究结果表明，肝脏肿瘤转化似乎抑制了 ALDH6A1 的表达，这伴随着癌细胞中 NO 和 ROS 的相应增加和减少。鉴于 ALDH6A1 抑制与异常癌细胞生长之间的密切联系，这种蛋白质可能作为肝癌发生和治疗反应的潜在分子特征或生物标志物。质谱数据可通过 ProteomeXchange 获得，标识符为 PXD014252。