Saito Tsuyoshi, Oda Yoshinao, Kawaguchi Ken-ichi, Sugimachi Keishi, Yamamoto Hidetaka, Tateishi Naomi, Tanaka Kazuhiro, Matsuda Shuichi, Iwamoto Yukihide, Ladanyi Marc, Tsuneyoshi Masazumi
Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Oncogene. 2004 Nov 11;23(53):8629-38. doi: 10.1038/sj.onc.1207960.
We have recently reported frequent E-cadherin gene mutations in synovial sarcoma (SS), suggesting mutational inactivation of E-cadherin as a potential mechanism of spindle cell morphology in SS, a spindle cell sarcoma that shows areas of glandular epithelial differentiaton in some cases (biphasic SS) and only pure spindle cell morphology in most cases (monophasic SS). However, the mechanism of downregulation of E-cadherin in SS remains unknown. To further address this issue, we analysed the mechanisms of E-cadherin silencing in 40 SS. Genetic and epigenetic changes in the E-cadherin gene, and the expression level of its transcriptional repressor Snail were examined by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP), methylation-specific PCR, and real-time quantitative PCR, respectively. Expression of E-cadherin was examined by RT-PCR and immunohistochemistry. We also examined ELF3, a transcription factor associated with epithelial differentiation in SS in a previous cDNA microarray, by RT-PCR. E-cadherin and ELF3 transcripts were detected, respectively, in 27/40 (67.5%) and in 25/40 (62.5%) of SS, and these epithelial-related genes were almost always coexpressed. Hypermethylation of the promoter of the E-cadherin gene was detected in five cases (12.5%) in SS; however, E-cadherin was silenced at mRNA level in only one of the five cases. E-cadherin missense mutations were observed in five cases (12.5%) of SS. In SS, all five cases with E-cadherin missense mutations had the SYT-SSX1 fusion and were monophasic tumors, suggesting a relationship between the SYT-SSX fusion type and E-cadherin missense mutation (P=0.07). E-cadherin mRNA expression in SS was associated with reduced Snail expression level (P=0.03). E-cadherin membranous expression was observed in 14/40 (35.0%) of SS, and was also correlated with SYT-SSX1 fusion type and biphasic histology. ELF3 was confirmed to be more highly expressed in biphasic than monophasic SS by real-time quantitative PCR. These results suggest that in SS the loss of E-cadherin expression occurs either by Snail trans-repression or by inactivating mutations. Thus, E-cadherin downregulation is associated with the loss or absence of glandular epithelial differentiation in certain SS.
我们最近报道了滑膜肉瘤（SS）中频繁出现E-钙黏蛋白基因突变，提示E-钙黏蛋白的突变失活是SS中梭形细胞形态形成的潜在机制，SS是一种梭形细胞肉瘤，在某些病例中表现为腺上皮分化区域（双相性SS），而在大多数病例中仅表现为单纯的梭形细胞形态（单相性SS）。然而，SS中E-钙黏蛋白下调的机制仍不清楚。为了进一步解决这个问题，我们分析了40例SS中E-钙黏蛋白沉默的机制。分别通过聚合酶链反应-单链构象多态性（PCR-SSCP）、甲基化特异性PCR和实时定量PCR检测E-钙黏蛋白基因的遗传和表观遗传变化及其转录抑制因子Snail的表达水平。通过RT-PCR和免疫组织化学检测E-钙黏蛋白的表达。我们还通过RT-PCR检测了ELF3，它是先前cDNA微阵列中与SS上皮分化相关的转录因子。在40例SS中，分别有27例（67.5%）和25例（62.5%）检测到E-钙黏蛋白和ELF3转录本，并且这些上皮相关基因几乎总是共表达的。在5例SS（12.5%）中检测到E-钙黏蛋白基因启动子的高甲基化；然而，在这5例中只有1例在mRNA水平上E-钙黏蛋白沉默。在5例SS（12.5%）中观察到E-钙黏蛋白错义突变。在SS中，所有5例有E-钙黏蛋白错义突变的病例都有SYT-SSX1融合，并且是单相性肿瘤，提示SYT-SSX融合类型与E-钙黏蛋白错义突变之间存在关联（P=0.07）。SS中E-钙黏蛋白mRNA表达与Snail表达水平降低相关（P=0.03）。在40例SS中有14例（35.0%）观察到E-钙黏蛋白膜表达，并且也与SYT-SSX1融合类型和双相组织学相关。通过实时定量PCR证实ELF3在双相性SS中比单相性SS表达更高。这些结果表明，在SS中，E-钙黏蛋白表达的缺失是通过Snail反式抑制或失活突变发生的。因此，E-钙黏蛋白下调与某些SS中腺上皮分化的丧失或缺乏相关。
