Objective: To explore the expression of FAT1 in esophageal squamous cell carcinoma (ESCC) tissues, and its effect on cell proliferation. Methods: The expression levels of FAT1 protein in human ESCC tissues and matched adjacent normal tissues were determined by immunohistochemistry (IHC). Lentivirus based knockdown of FAT1 was carried out in YSE2 and Colo680N cell lines and 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H tetrazolium bromide (MTT) assays was performed to examine the effect of FAT1 on the proliferation of these ESCC cells. Colony formation assay was used to detect the colony formation ability. Flow cytometry was performed to analyze the cell cycle and apoptosis. The expression levels of cell cycle markers in FAT1 knock out ESCC cell lines were detected by real-time quantitative reverse transcription polymerase chain reaction(qRT-PCR) and Western blot. Results: The relative expression of FAT1 in ESCC tissues was 66.97±21.53, significantly lower than 78.13±16.76 of adjacent normal tissues(P<0.05). Knockdown of FAT1 promoted cell proliferation and colony formation. In YSE2 cell, the division time in negative control (NC) group was (1 570±51) min, significantly longer than (1 356±31) min in shFAT1 group. In Colo680N cell, division time in NC group was (1 532±53) min, significantly longer than (1 290±30) min in shFAT1 group (P<0.05). Knockdown of FAT1 promoted G1-to S-phase transition and resulted in the upregulation of CDK4/CDK6/CCND1. Conclusion: FAT1 inhibits the proliferation and G1-to S-phase transition of ESCC cells through regulating the protein expression of CDK4/CDK6/CCND1 complex.
目的： 探讨食管鳞癌患者中FAT1的表达情况及其对细胞生长的调控机制。 方法： 收集食管鳞癌组织标本制作组织芯片，采用免疫组化检测FAT1在食管鳞癌及癌旁正常组织中的表达。在高表达FAT1食管鳞癌细胞系YSE2和Colo680N中，应用慢病毒感染法干扰FAT1后，采用四甲基偶氮唑蓝法和平板克隆形成实验观察FAT1对细胞生长的影响。流式细胞术检测敲低FAT1的表达对细胞周期和凋亡率的影响。Western blot检测敲低FAT1的表达对细胞周期标志物表达的影响。 结果： FAT1在食管鳞癌及癌旁正常组织中的相对表达水平分别为66.97±21.53和78.13±16.76，差异有统计学意义(P<0.05)。敲低FAT1表达的细胞生长速度加快，克隆形成能力增强。在YSE2细胞中，对照组细胞分裂1次的时间为(1 570±51)min，敲低FAT1表达后细胞分裂时间为(1 356±31)min；在Colo680N细胞中，对照组细胞分裂1次时间为(1 532±53)min，敲低FAT1表达后细胞分裂时间为(1 290±30)min，差异均有统计学意义(P<0.05)。敲低FAT1表达促进细胞周期由G(1)期向S期转化，上调CDK4/CDK6/CCND1复合物的表达。 结论： FAT1通过调节食管鳞癌细胞CDK4 /CDK6 /CCND1复合物的表达，抑制细胞周期由G(1)期向S期转化，抑制细胞增殖。.
Keywords: Carcinoma, squamous cell; Cell cycle; Cell proliferation; Esophageal neoplasms; FAT1.