摘要
目的 :探讨小鼠端粒酶RNA(mTR)基因的反义寡核苷酸 (ASODN)对体外培养的大鼠A型精原细胞端粒酶活性及其mTR亚基表达的影响。 方法 :以脂质体LipofectAMINE 2 0 0 0 (LF 2 0 0 0 )介导将硫代磷酸修饰的端粒酶mTR的ASODN、正义寡核苷酸 (SODN)、随机寡核苷酸 (RODN)及单纯脂质体组分别转染体外分离纯化的SD大鼠A型精原细胞 ,采用生物发光技术和TRAP SYBR Green染色法检测精原细胞中端粒酶活性的改变 ,原位杂交检测mTR基因mRNA的表达。 结果 :端粒酶mTR的ASODN明显抑制精原细胞端粒酶活性 (P <0 .0 1) ;mTR ASODN作用于精原细胞 2 4h后 ,mTR基因mRNA的表达水平显著下调。单纯脂质体组及SODN、RODN对照组均无此抑制作用 (P >0 .0 5 )。 结论 :硫代修饰的端粒酶mTR ASODN可使精原细胞端粒酶活性明显受到抑制 ,其机制可能是在mTR基因转录水平影响精原细胞端粒酶活性。
Objectives: To study the inhibitory effects of mouse telomerase RNA (mTR) antisense oligodeoxynucleotide(ASODN) on telomerase activity in rat spermatogonia. Methods: 9-mer phosphorothioate mTR-ASODN was encapsulated by LipofectAMINE 2000(LF 2000) and transfected to type A spermatogonia in Sprague Dawley (SD) rat. Telomerase activity was detected by aid of TRAP-SYBR-Green staining and Bioluminescence techniquein type A spermatogonia treated or untreated with ASODN. Results: mTR-ASODN conjugated with LF 2000 could significantly inhibit telomerase activity of spermatogonia(P< 0.01). mTR mRNA level also decreased while the spermatogonia were treated with ASODN for 24 h. No change of telomerase activity and apoptosis were observed when SODN, RODN or single LF 2000 was used. Conclusions: Antisense oligodeoxynucleotide of mTR conjugated with LF 2000 could significantly inhibit telomerase activity of spermatogonia. mTR-ASODN might inhibit telomerase activity of spermatogonia at transcription level.
出处
《中华男科学杂志》
CAS
CSCD
2003年第6期421-424,428,共5页
National Journal of Andrology
基金
卫生部科研基金资助项目 (98 1 136 )
