Yang Run-Wei, Zeng Ying-Yue, Wei Wen-Ting, Cui Yan-Mei, Sun Hui-Ying, Cai Yue-Long, Nian Xin-Xin, Hu Yun-Teng, Quan Yu-Ping, Jiang Sheng-Lu, Wang Meng, Zhao Ya-Li, Qiu Jun-Feng, Li Ming-Xuan, Zhang Jia-Huan, He Mei-Rong, Liang Li, Ding Yan-Qing, Liao Wen-Ting
Department of Pathology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, Guangdong, China.
Department of Pathology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong, China.
J Exp Clin Cancer Res. 2016 Sep 27;35(1):152. doi: 10.1186/s13046-016-0426-8.
Transducin-like enhancer of Split3 (TLE3) serves as a transcriptional corepressor during cell differentiation and shows multiple roles in different kinds of cancers. Recently, TLE3 together with many other genes involved in Wnt/β-catenin pathway were detected hyper-methylated in colorectal cancer (CRC). However, the potential role and the underlying mechanism of TLE3 in CRC progression remain scarce.
Gene expression profiles were analyzed in The Cancer Genome Atlas (TCGA) microarray dataset of 41 normal colorectal intestine tissues and 465 CRC tissues. Western blot and Real-time Quantitative PCR (RT-qPCR) were respectively performed to detect protein and mRNA expression in 8 pairs of CRC tissue and matched adjacent normal mucosa. Immunohistochemistry (IHC) was conducted to evaluate TLE3 protein expression in 105 paraffin-embedded, archived human CRC tissues from patients, whose survival data were analyzed with Kaplan-Meier method. In vitro experiments including MTT assay, colony formation assay, and soft agar formation assay were used to investigate the effects of TLE3 on CRC cell growth and proliferation. Additionally, subcutaneous tumorigenesis assay was performed in nude mice to confirm the effects of TLE3 in vivo. Furthermore, gene set enrichment analysis (GSEA) was run to explore potential mechanism of TLE3 in CRC, and then we measured the distribution of CRC cell cycle phases and apoptosis by flow cytometry, as well as the impacts of TLE3 on MAPK and AKT signaling pathways by Western blot and RT-qPCR.
TLE3 was significantly down-regulated in 465 CRC tissues compared with 41 normal tissues. Both protein and mRNA expressions of TLE3 were down-regulated in CRC compared with matched adjacent normal mucosa. Lower expression of TLE3 was significantly associated with poorer survival of patients with CRC. Besides, knock down of TLE3 promoted CRC cell growth and proliferation, while overexpression of TLE3 showed suppressive effects. Furthermore, overexpression of TLE3 caused G1-S phase transition arrest, inhibition of MAPK and AKT pathways, and up-regulation of p21Cip1/WAF1 and p27Kip1.
This study indicated that TLE3 repressed CRC proliferation partly through inhibition of MAPK and AKT signaling pathways, suggesting the possibility of TLE3 as a biomarker for CRC prognosis.
分裂样增强子3（TLE3）在细胞分化过程中作为转录共抑制因子，在多种癌症中发挥多种作用。最近，在结直肠癌（CRC）中检测到TLE3与许多其他参与Wnt/β-连环蛋白通路的基因发生了高甲基化。然而，TLE3在CRC进展中的潜在作用和潜在机制仍然缺乏。
在癌症基因组图谱（TCGA）的41例正常结直肠组织和465例CRC组织的微阵列数据集中分析基因表达谱。分别进行蛋白质免疫印迹和实时定量PCR（RT-qPCR）检测8对CRC组织和匹配的相邻正常黏膜中的蛋白质和mRNA表达。进行免疫组织化学（IHC）评估105例石蜡包埋的存档人类CRC组织中TLE3蛋白的表达，并用Kaplan-Meier方法分析患者的生存数据。体外实验包括MTT法、集落形成实验和软琼脂形成实验，用于研究TLE3对CRC细胞生长和增殖的影响。此外，在裸鼠中进行皮下成瘤实验以证实TLE3在体内的作用。此外，进行基因集富集分析（GSEA）以探索TLE3在CRC中的潜在机制，然后通过流式细胞术测量CRC细胞周期阶段和凋亡的分布，以及通过蛋白质免疫印迹和RT-qPCR检测TLE3对MAPK和AKT信号通路的影响。
与41例正常组织相比，465例CRC组织中TLE3显著下调。与匹配的相邻正常黏膜相比，CRC中TLE3的蛋白质和mRNA表达均下调。TLE3表达较低与CRC患者较差的生存率显著相关。此外，敲低TLE3促进CRC细胞生长和增殖，而TLE3过表达则显示出抑制作用。此外，TLE3过表达导致G1-S期转变停滞，抑制MAPK和AKT通路，并上调p21Cip1/WAF1和p27Kip1。
本研究表明，TLE3部分通过抑制MAPK和AKT信号通路抑制CRC增殖，提示TLE3作为CRC预后生物标志物的可能性。
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