癌基因 GAEC1 调控 CAPN10 的表达，其表达水平可预测食管鳞癌患者的生存情况。
Oncogene GAEC1 regulates CAPN10 expression which predicts survival in esophageal squamous cell carcinoma.
机构信息
State Key Laboratory of Chirosciences, Lo Ka Chung Centre for Natural Anti-cancer Drug Development, Department of Applied Biology and Chemical Technology, the Hong Kong Polytechnic University, Hong Kong, China.
出版信息
World J Gastroenterol. 2013 May 14;19(18):2772-80. doi: 10.3748/wjg.v19.i18.2772.
AIM
To identify the downstream regulated genes of GAEC1 oncogene in esophageal squamous cell carcinoma and their clinicopathological significance.
METHODS
The anti-proliferative effect of knocking down the expression of GAEC1 oncogene was studied by using the RNA interference (RNAi) approach through transfecting the GAEC1-overexpressed esophageal carcinoma cell line KYSE150 with the pSilencer vector cloned with a GAEC1-targeted sequence, followed by MTS cell proliferation assay and cell cycle analysis using flow cytometry. RNA was then extracted from the parental, pSilencer-GAEC1-targeted sequence transfected and pSilencer negative control vector transfected KYSE150 cells for further analysis of different patterns in gene expression. Genes differentially expressed with suppressed GAEC1 expression were then determined using Human Genome U133 Plus 2.0 cDNA microarray analysis by comparing with the parental cells and normalized with the pSilencer negative control vector transfected cells. The most prominently regulated genes were then studied by immunohistochemical staining using tissue microarrays to determine their clinicopathological correlations in esophageal squamous cell carcinoma by statistical analyses.
RESULTS
The RNAi approach of knocking down gene expression showed the effective suppression of GAEC1 expression in esophageal squamous cell carcinoma cell line KYSE150 that resulted in the inhibition of cell proliferation and increase of apoptotic population. cDNA microarray analysis for identifying differentially expressed genes detected the greatest levels of downregulation of calpain 10 (CAPN10) and upregulation of trinucleotide repeat containing 6C (TNRC6C) transcripts when GAEC1 expression was suppressed. At the tissue level, the high level expression of calpain 10 protein was significantly associated with longer patient survival (month) of esophageal squamous cell carcinoma compared to the patients with low level of calpain 10 expression (37.73 ± 16.33 vs 12.62 ± 12.44, P = 0.032). No significant correction was observed among the TNRC6C protein expression level and the clinocopathologcial features of esophageal squamous cell carcinoma.
CONCLUSION
GAEC1 regulates the expression of CAPN10 and TNRC6C downstream. Calpain 10 expression is a potential prognostic marker in patients with esophageal squamous cell carcinoma.
目的
鉴定 GAEC1 癌基因在食管鳞状细胞癌中的下游调控基因及其临床病理意义。
方法
通过用靶向 GAEC1 的 pSilencer 载体转染过表达 GAEC1 的食管癌细胞系 KYSE150，采用 RNA 干扰（RNAi）方法研究敲低 GAEC1 表达的抗增殖作用，随后用 MTS 细胞增殖试验和流式细胞术进行细胞周期分析。然后从亲本、靶向 GAEC1 的 pSilencer 载体转染和 pSilencer 阴性对照载体转染的 KYSE150 细胞中提取 RNA，进一步分析不同基因表达模式。然后使用 Human Genome U133 Plus 2.0 cDNA 微阵列分析比较亲本细胞与 pSilencer 阴性对照载体转染细胞，确定 GAEC1 表达受抑制时差异表达的基因。然后通过组织微阵列的免疫组织化学染色研究最显著调节的基因，通过统计分析确定其在食管鳞状细胞癌中的临床病理相关性。
结果
RNAi 方法敲低基因表达显示，在食管鳞状细胞癌细胞系 KYSE150 中有效抑制 GAEC1 表达，导致细胞增殖抑制和凋亡群体增加。用于鉴定差异表达基因的 cDNA 微阵列分析检测到，当 GAEC1 表达受抑制时，钙蛋白酶 10 (CAPN10) 的下调水平最高，三核苷酸重复包含 6C (TNRC6C) 转录本的上调水平最高。在组织水平上，与 CAPN10 低表达的患者相比，CAPN10 蛋白高水平表达的食管鳞状细胞癌患者的生存时间（月）更长（37.73 ± 16.33 对 12.62 ± 12.44，P = 0.032）。TNRC6C 蛋白表达水平与食管鳞状细胞癌的临床病理特征之间没有观察到显著相关性。
结论
GAEC1 调节 CAPN10 和 TNRC6C 的表达。钙蛋白酶 10 表达可能是食管鳞状细胞癌患者的潜在预后标志物。
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