Department of Human Anatomy, Taishan Medical University, 2 Ying Sheng Dong Lu, Taian 271000, China.
Department of Pathology, Affiliated Hospital of Taishan Medical University, Taian 271000, China.
J Immunol Res. 2018 Sep 30;2018:4735752. doi: 10.1155/2018/4735752. eCollection 2018.
A previous study showed that a downexpression of protease-activated receptor 4 (PAR4) is associated with the development of esophageal squamous cell carcinoma (ESCC). In this study, we explored the relationship between PAR4 activation and the expression of p16, and elucidated the underlying mechanisms in PAR4 inducing the tumor suppressor role in ESCC. ESCC cell lines (EC109 and TE-1) were treated with PAR4-activating peptide (PAR4-AP). Immunohistochemistry for DNA methyltransferase 1 (DNMT1) and histone deacetylase 2 (HDAC2) was performed in 26 cases of ESCC tissues. We found that DNMT1 and HDAC2 immunoreactivities in ESCC were significantly higher than those in adjacent noncancerous tissues. PAR4 activation could suppress DNMT1 and HDAC2, as well as increase p16 expressions, whereas silencing PAR4 dramatically increased HDAC2 and DNMT1, as well as reduced p16 expressions. Importantly, the chromatin immunoprecipitation-PCR (ChIP-PCR) data indicated that treatment of ESCC cells with PAR4-AP remarkably suppressed DNMT1 and HDAC2 enrichments on the p16 promoter. Furthermore, we demonstrated that activation of PAR4 resulted in an increase of p38/ERK phosphorylation and activators for p38/ERK enhanced the effect of PAR4 activation on HDAC2, DNMT1, and p16 expressions, whereas p38/ERK inhibitors reversed these effects. Moreover, we found that activation of PAR4 in ESCC cells significantly inhibited cell proliferation and induced apoptosis. These findings suggest that PAR4 plays a potential tumor suppressor role in ESCC cells and represents a potential therapeutic target of this disease.
先前的研究表明，蛋白酶激活受体 4（PAR4）的下调表达与食管鳞状细胞癌（ESCC）的发展有关。在这项研究中，我们探讨了 PAR4 激活与 p16 表达之间的关系，并阐明了 PAR4 诱导 ESCC 中肿瘤抑制作用的潜在机制。用 PAR4 激活肽（PAR4-AP）处理 ESCC 细胞系（EC109 和 TE-1）。对 26 例 ESCC 组织中的 DNA 甲基转移酶 1（DNMT1）和组蛋白去乙酰化酶 2（HDAC2）进行免疫组织化学染色。我们发现 ESCC 中的 DNMT1 和 HDAC2 免疫反应性明显高于相邻的非癌组织。PAR4 激活可抑制 DNMT1 和 HDAC2，并增加 p16 的表达，而 PAR4 沉默则明显增加 HDAC2 和 DNMT1，并降低 p16 的表达。重要的是，染色质免疫沉淀-PCR（ChIP-PCR）数据表明，用 PAR4-AP 处理 ESCC 细胞可显著抑制 p16 启动子上的 DNMT1 和 HDAC2 富集。此外，我们证明 PAR4 的激活导致 p38/ERK 磷酸化增加，p38/ERK 的激活剂增强了 PAR4 激活对 HDAC2、DNMT1 和 p16 表达的影响，而 p38/ERK 抑制剂则逆转了这些影响。此外，我们发现 ESCC 细胞中 PAR4 的激活可显著抑制细胞增殖并诱导细胞凋亡。这些发现表明 PAR4 在 ESCC 细胞中发挥潜在的肿瘤抑制作用，是该疾病的潜在治疗靶点。
