胆固醇激活p38 MAPK信号通路调控大肠癌干性表型及改造肝转移灶微环境的机制研究

The high mortality of colorectal cancer (CRC) is closely related to metastasis, especially liver metastasis. Our previous data shows there is a positive correlation between cholesterol level and the incidence of intestinal polyp, CRC as well as liver metastasis. And also, cholesterol will promote the stem-like phenotypes including cell migration, tumor-sphere formation and tumorigenesis of colorectal cancer cells. Moreover, hyperlipidemia may lead to chronic inflammation of liver as well. p38 MAPK signal pathway has been proved to play an important role in inflammation, tumor, and hyperlipidemia-related disease. Thus we hypothesize that the promotion of colorectal cancer stem-like phenotype and inflammation caused by cholesterol may lead to the remodeling of pre-metastatic niche in liver, resulting in the positive correlation between hyperlipidemia and liver metastasis. And this is also a common clinical phenomenon. Based on our previous study, this research intends to further clarify the impact of cholesterol on the stem-like phenotype of colorectal cancer cells. We used A/J mouse to investigate the role of hyperlipidemia in colorectalcarcinogenesis. The impact of hyperlipidemia on chronic hepatitis was determined by NAFLD mouse model and clinical pathology. Site-directed mutagenesis followed by clinical sample study was also performed to identify the specific target of hyperlipidemia on p38 MAPK pathway. Our study provides experimental basis and intervention molecules for further research to prevent colorectalcarcinogenesis and liver metastasis caused by hyperlipidemia. And it is of great significance in scientific research and potential value on clinical practice.

大肠癌的高死亡率与其转移、尤其是肝转移高度相关。我们前期数据证实胆固醇水平与肠息肉、大肠癌、大肠癌肝转移的发生率正相关；并且胆固醇可提高大肠癌细胞迁移、克隆球形成、动物致瘤等干性表型；同时高脂可导致肝脏的慢性非特异性炎症。p38 MAPK信号通路在炎症、肿瘤、高脂相关疾病中起着重要作用。我们推测：胆固醇可促进大肠癌干性表型，同时促进炎症而改造肝脏的转移前微环境，从而造成高胆固醇与大肠癌肝转移正相关这一临床现象。本课题拟在先前研究基础上，明确胆固醇促进大肠癌细胞的干性表型，以A/J鼠自发成瘤模型验证高胆固醇促进大肠癌发生；基于脂肪肝动物模型及临床病理组织明确高胆固醇对肝脏的慢性炎症；定点突变等实验明确胆固醇调控p38 MAPK信号通路的具体靶点，并以临床标本加以验证。上述科学问题的解决，为阻断高脂引起的大肠癌及其肝转移提供实验依据及干预分子，具有重要的科学意义和潜在的临床应用价值。

结直肠癌是全球第三高发的恶性肿瘤，我国的结直肠癌发病率也逐年增高。代谢模式重构及菌群失衡等因素均能促进结直肠癌的发生及进展。我们从GEO数据库中获得GSE146771数据集，通过联合差异分析、GSEA分析、GSVA分析、伪时间序列分析、细胞间通讯网络分析及SCENIC分析共同探究结直肠癌内免疫细胞和恶性细胞内脂代谢重塑情况，发现结直肠癌内CD8+ T细胞在耗竭过程、巨噬细胞在向肿瘤相关巨噬细胞分化过程以及恶性细胞中均表现出以脂代谢重塑为主的代谢模式的重构，且通过对恶性细胞的进一步分析发现，ERR基因主导了恶性细胞的脂代谢重塑过程。近百例结直肠癌标本的组织芯片也显示ERRα在肿瘤细胞中的表达显著升高。且体内外实验也证实ERRα可促进结直肠癌细胞的迁移、克隆球及致瘤等能力。通过CHIP-seq分析ERRα调控脂代谢的下游靶基因，进行了进一步的生物信息学分析。此外，我们构建了高脂饮食动物模型，16S测序分析发现高脂饮食后肠道出现以革兰阴性菌显著增多为特征的菌群失衡变化。并且菌群失衡会引起肠粘膜炎症及肠屏障损害，增加结直肠癌发生的风险。同时，也会诱发肝脏炎症，造成肝脏微环境的改变，便于结直肠癌发生肝转移。与此同时，通过对结直肠癌肝转移及非肝转移患者粪便数据的16S分析，发现发生肝转移的患者肠道中脱硫弧菌丰度明显升高。为此，我们构建脱硫弧菌灌胃小鼠模型，实验结果显示脱硫弧菌肠道中优势定植能诱发肠粘膜炎症及肠粘膜屏障受损，且灌胃脱硫弧菌后肝脏出现明显损伤炎症反应且肝脏微环境进一步得到了塑造，便于结直肠癌细胞生长定植。我们的研究进一步明确了脂代谢重塑、其相关调控分子ERRα及菌群紊乱与结直肠癌的联系，为后续降低结直肠癌的发生、提高结直肠癌的治疗效果提供了更多的选择依据，具有较大的临床转化价值。