应用抑制性消减杂交技术鉴定肿瘤转移抑制相关基因

Identification of genes associated with human osteosarcoma metastasis suppression using suppression subtractive hybridization

目的 克隆与肿瘤转移抑制相关的基因 ,探讨成骨肉瘤转移的分子机理。方法 采用抑制性消减杂交技术构建了人成骨肉瘤低转移细胞株 SOSP- 96 0 7的消减文库 ,对部分克隆进行了测序和同源性分析 ,用 Northern杂交及反转录 PCR技术证实了克隆的表达情况。结果 在低转移细胞株 SOSP- 96 0 7中高表达的克隆中有 2个克隆与人端粒结合因子 2 (telomeric repeat binding factor2 ,TERF2 )同源。Northern杂交及反转录 PCR证实这个基因只在低转移细胞中表达 ,而在高转移人成骨肉瘤细胞株 SOSP-M和裸鼠肺转移结节中均未表达。结论 TERF2可能在维持肿瘤细胞自身相对稳定、抑制骨肉瘤演进中起重要作用。

Objective: To identify genes associated with metastasis suppression and to investigate the molecular mechanism of osteosarcoma metastasis. Methods: The subtracted cDNA library of low metastatic human osteosarcoma cell line SOSP-9607 was constructed using suppression subtractive hybridization. Partial clones were sequenced. The acquired sequence data were aligned against the GenBank nucteotide database using Blastn to search for sequence matches. The interested clone was used to perform Northern blot and reverse transcriptase-PCR (RT-PCR) analysis on mRNA isolated from low metastatic cell line SOSP-9607 and OS-9901, high metastatic cell line SOSP-M and three pulmonic metastatic nodules of nude mice. Results: A cDNA clone from low metastatic cell line SOSP-9607 subtracted cDNA library was identified as telomeric repeat binding factor 2 (TERF2) by sequence analysis and Blastn search. Northern blot and RT-PCR analysis demonstrated that TERF2 expressed highly in low metastatic cell line SOSP-9607 and OS-9901, but not in high metastatic cell line SOSP-M and three pulmonic metastatic nodules. Conclusion: TERF2 may be important for suppressing metastasis of osteosarcoma.