Increased Expression of Protease-Activated Receptor 4 and Trefoil Factor 2 in Human Colorectal Cancer
Increased Expression of Protease-Activated Receptor 4 and Trefoil Factor 2 in Human Colorectal Cancer
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DOI：
10.1371/journal.pone.0122678
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发表时间：
2015-04-13
期刊：
影响因子：
3.7
通讯作者：
Zhang, Yun
中科院分区：
文献类型：
--
作者：
Yu, Guoyu;Jiang, Ping;Zhang, Yun
蛋白酶激活受体4（PAR 4）是G蛋白偶联受体家族的一员，在胃癌和食管鳞癌中表达降低，而在前列腺癌中表达增加。三叶因子2（TFF 2）是一种在胃粘膜中组成性表达的小分子肽，在胃粘膜修复中起保护作用。TFF 2表达的改变也与胃肠道癌的发生有关。TFF 2已被证实通过PAR 4促进细胞迁移，但PAR 4和TFF 2在结直肠癌进展中的作用仍不清楚。本研究采用实时荧光定量PCR（n = 38）、免疫印迹（n = 38）和组织芯片（n = 66）检测结直肠癌组织中PAR 4和TFF 2的表达水平。结直肠癌组织中PAR 4和TFF 2的mRNA和蛋白表达水平均显著高于癌旁组织，尤其是在淋巴结阳性和低分化癌组织中。结肠直肠癌细胞LoVo显示出对TFF 2的反应增加，如通过PAR 4表达后的细胞侵袭所评估的。然而，PAR 4表达干预后，PAR 4阳性大肠癌细胞HT-29对TFF 2的细胞侵袭反应降低。基因组亚硫酸氢盐测序显示结直肠癌组织中PAR 4启动子低甲基化，而正常粘膜中PAR 4启动子高甲基化，提示启动子低甲基化与PAR 4表达增高相关。以上结果表明，PAR 4和TFF 2在结直肠癌组织中表达上调，PAR 4的过表达可能与其启动子低甲基化有关。而TFF 2促进过表达PAR 4的LoVo细胞的侵袭活性，并且当在HT-29细胞中敲低PAR 4时，这种作用显著降低。我们的研究结果将有助于进一步探讨蛋白酶激活受体（PARs）和三叶因子（TFFs）在结直肠癌发生发展中的作用及其分子机制。
Protease-activated receptor 4 (PAR4), a member of G-protein coupled receptors family, was recently reported to exhibit decreased expression in gastric cancer and esophageal squamous cancer, yet increased expression during the progression of prostate cancer. Trefoil factor 2 (TFF2), a small peptide constitutively expressed in the gastric mucosa, plays a protective role in restitution of gastric mucosa. Altered TFF2 expression was also related to the development of gastrointestinal cancer. TFF2 has been verified to promote cell migration via PAR4, but the roles of PAR4 and TFF2 in the progress of colorectal cancer are still unknown. In this study, the expression level of PAR4 and TFF2 in colorectal cancer tissues was measured using real-time PCR (n = 38), western blotting (n = 38) and tissue microarrays (n = 66). The mRNA and protein expression levels of PAR4 and TFF2 were remarkably increased in colorectal cancer compared with matched noncancerous tissues, especially in positive lymph node and poorly differentiated cancers. The colorectal carcinoma cell LoVo showed an increased response to TFF2 as assessed by cell invasion upon PAR4 expression. However, after intervention of PAR4 expression, PAR4 positive colorectal carcinoma cell HT-29 was less responsive to TFF2 in cell invasion. Genomic bisulfite sequencing showed the hypomethylation of PAR4 promoter in colorectal cancer tissues and the hypermethylation in the normal mucosa that suggested the low methylation of promoter was correlated to the increased PAR4 expression. Taken together, the results demonstrated that the up-regulated expression of PAR4 and TFF2 frequently occurs in colorectal cancer tissues, and that overexpression of PAR4 may be resulted from promoter hypomethylation. While TFF2 promotes invasion activity of LoVo cells overexpressing PAR4, and this effect was significantly decreased when PAR4 was knockdowned in HT-29 cells. Our findings will be helpful in further investigations into the functions and molecular mechanisms of Proteinase-activated receptors (PARs) and Trefoil factor factors (TFFs) during the progression of colorectal cancer.