Hepatocellular carcinoma (HCC) is the most common liver cancer and is highly malignant. Male prevalence and frequent activation of the Ras signaling pathway are distinct characteristics of HCC. However, the underlying mechanisms remain to be elucidated. By exploring Hras12V transgenic mice showing male-biased hepatocarcinogenesis, we performed a high-throughput comparative proteomic analysis based on tandem-mass-tag (TMT) labeling combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) on the tissue samples obtained from HCC (T) and their paired adjacent precancerous (P) of Hras12V transgenic male and female mice (Ras-Tg) and normal liver (W) of wild-type male and female mice (Non-Tg). The further validation and investigation were performed using quantitative real-time PCR and western blot. Totally, 5193 proteins were quantified, originating from 5733 identified proteins. Finally, 1344 differentially expressed proteins (DEPs) (quantified in all examined samples; |ratios| ≥ 1.5, p < 0.05) were selected for further analysis. Comparison within W, P, and T of males and females indicated that the number of DEPs in males was much higher than that in females. Bioinformatics analyses showed the common and unique cluster-enriched items between sexes, indicating the common and gender-disparate pathways towards HCC. Expression change pattern analysis revealed HCC positive/negative-correlated and ras oncogene positive/negative-correlated DEPs and pathways. In addition, it showed that the ras oncogene gradually and significantly reduced the responses to sex hormones from hepatocytes to hepatoma cells and therefore shrunk the gender disparity between males and females, which may contribute to the cause of the loss of HCC clinical responses to the therapeutic approaches targeting sex hormone pathways. Additionally, gender disparity in the expression levels of key enzymes involved in retinol metabolism and terpenoid backbone/steroid biosynthesis pathways may contribute to male prevalence in hepatocarcinogenesis. Further, the biomarkers, SAA2, Orm2, and Serpina1e, may be sex differences. In conclusion, common and unique DEPs and pathways toward HCC initiated by ras oncogene from sexually dimorphic hepatocytes provide valuable and novel insights into clinical investigation and practice.

中文翻译：

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蛋白质组学分析揭示了性别依赖性肝癌发生中的常见、独特和系统特征。

肝细胞癌（HCC）是最常见的肝癌，恶性程度高。男性患病率和 Ras 信号通路的频繁激活是 HCC 的明显特征。然而，潜在的机制仍有待阐明。通过探索 Hras12V 转基因小鼠显示男性偏向肝癌的发生，我们对组织样本进行了基于串联质量标签 (TMT) 标记结合液相色谱-串联质谱 (LC-MS/MS) 的高通量比较蛋白质组学分析从 Hras12V 转基因雄性和雌性小鼠 (Ras-Tg) 的 HCC (T) 及其配对的相邻癌前病变 (P) 和野生型雄性和雌性小鼠 (Non-Tg) 的正常肝脏 (W) 中获得。使用定量实时 PCR 和蛋白质印迹进行进一步的验证和调查。共定量5193种蛋白质，源自 5733 个已鉴定的蛋白质。最后，选择了 1344 种差异表达蛋白 (DEP)（在所有检查的样本中量化；|比率| ≥ 1.5，p < 0.05）进行进一步分析。男性和女性在 W、P 和 T 内的比较表明，男性的 DEP 数量远高于女性。生物信息学分析显示了两性之间常见和独特的聚类丰富项目，表明了 HCC 的常见和性别不同的途径。表达变化模式分析显示 HCC 阳性/阴性相关和 ras 致癌基因阳性/阴性相关 DEP 和途径。此外，还表明ras癌基因逐渐显着降低了肝细胞对肝癌细胞对性激素的反应，从而缩小了男女之间的性别差异，这可能导致 HCC 对针对性激素途径的治疗方法失去临床反应的原因。此外，参与视黄醇代谢和萜类骨架/类固醇生物合成途径的关键酶表达水平的性别差异可能有助于男性在肝癌发生中的患病率。此外，生物标志物 SAA2、Orm2 和 Serpina1e 可能是性别差异。总之，由来自性二态性肝细胞的 ras 癌基因引发的常见和独特的 DEP 和通往 HCC 的途径为临床研究和实践提供了有价值和新颖的见解。参与视黄醇代谢和萜类骨架/类固醇生物合成途径的关键酶表达水平的性别差异可能有助于男性在肝癌发生中的流行。此外，生物标志物 SAA2、Orm2 和 Serpina1e 可能是性别差异。总之，由来自性二态性肝细胞的 ras 癌基因引发的常见和独特的 DEP 和通往 HCC 的途径为临床研究和实践提供了有价值和新颖的见解。参与视黄醇代谢和萜类骨架/类固醇生物合成途径的关键酶表达水平的性别差异可能有助于男性在肝癌发生中的流行。此外，生物标志物 SAA2、Orm2 和 Serpina1e 可能是性别差异。总之，由来自性二态性肝细胞的 ras 癌基因引发的常见和独特的 DEP 和通往 HCC 的途径为临床研究和实践提供了有价值和新颖的见解。