曲古菌素A抑制HL-60细胞端粒酶活性及hTERT的表达并诱导凋亡

曲古菌素A抑制HL-60细胞端粒酶活性及hTERT的表达并诱导凋亡

周咏明, 郭伟, 周浩, 李慧玉, 刘黎琼, 姚军霞, 郑金娥, 郭天南, 黄士昂^*

1.华中科技大学同济医学院附属协和医院干细胞研究和应用中心, 湖北武汉 430022;
2.华中科技大学同济医学院附属同济医院内分泌科, 湖北武汉 430030;
3.华中科技大学同济医学院附属协和医院血液科, 湖北武汉 430022

收稿日期:2005-10-10 修回日期:2006-05-10 出版日期:2006-06-15 发布日期:2006-06-15
通讯作者: 黄士昂*

Trichostatin A Induces Apoptosis by Inhibiting Telomerase Activity and Expression of Telomerase Reverse Transcriptase in HL-60 Cells

ZHOU Yong-ming, GUO Wei, ZHOU Hao, LI Hui-yu, LIU Li-qiong, YAO Jun-xia, ZHENG Jin-e, GUO Tian-nan, HUANG Shi-ang^*

1.Center for Stem Cell Research and Application, Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China;
2.Department of Endocrinology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China;
3.Department of Hematology, Uinion Hospital, Tongji medical College, Huazhong University of Science and Technology, Wuhan 430022, China

Received:2005-10-10 Revised:2006-05-10 Online:2006-06-15 Published:2006-06-15
Contact: HUANG Shi-ang*

摘要/Abstract

摘要：

目的研究组蛋白去乙酰化酶抑制剂曲古菌素A(trichostatin A, TSA)抑制HL-60细胞端粒酶活性及亚单位hTERT的表达并诱导凋亡的机制。方法采用MTT法和倒置相差显微镜观察不同浓度曲古菌素A对HL-60细胞的抑制作用, 流式细胞仪检测600 nmol·L^-1TSA作用后的细胞凋亡情况, TRAP-ELISA法检测端粒酶活性变化, RT-PCR分析端粒酶三个亚单位的mRNA表达情况。结果 TSA对HL-60细胞的抑制作用具有时间和剂量依赖性, AnnexinV/PI双染色法检测凋亡显示, 600 nmol·L^-1TSA作用48 h后, 细胞凋亡率为42.6%。600 nmol·L^-1TSA作用12、24和48 h后, 端粒酶活性分别下降到1.95±0.25、1.73±0.12和1.52±0.09。RT-PCR显示, 端粒酶逆转录酶hTERT表达下降, 而端粒酶RNA模板(hTR)和端粒酶相关蛋白(hTP1)表达无明显改变。结论 TSA抑制HL-60细胞中端粒酶活性, 并诱导凋亡, 其机制可能与曲古菌素A下调hTERT转录水平有关。

关键词: 曲古菌素A, 曲古菌素A, 曲古菌素A, 细胞凋亡, 细胞凋亡, 细胞凋亡, 端粒酶, 端粒酶, 端粒酶, 端粒酶逆转录酶, 端粒酶逆转录酶, 端粒酶逆转录酶

Abstract: Aim To investigate the effects of trichostatin A (TSA) on telomerase activity and the expression of human telomerase reverse transcriptase (hTERT) during apoptosis in vitro and the mechanisms in HL-60 cells. Methods The proliferative activity of HL-60 cells was assessed by MTT assay. Cell apoptosis was analyzed by flow cytometry. Telomerase activity was examined by TRAP-ELISA. The expression of telomerase subunits was analyzed by RT-PCR. Results A time- and dose-dependent inhibition was detected in HL-60 cells treated with TSA. After treatment with 600 nmol·L^-1 TSA for 48 h, the apoptosis rate in HL-60 cells was 42.6% and telomerase activity decreased 1.95±0.25, 1.73±0.12 and 1.52±0.09 for 12, 24 and 48 h, respectively. The expression of hTERTmRNA decreased. No significant changes were observed in the expression of hTRmRNA and hTP1mRNA. Conclusion TSA inhibits telomerase activity and induces apoptosis in HL-60 cells. The underlying mechanism may be related to the down-regulation of hTERT transcription.

Key words: trichostatin A, trichostatin A, apoptosis, apoptosis, telomerase, telomerase, human telomerase reverse transcriptase, human telomerase reverse transcriptase

中图分类号:

R963

R965

Supporting: Foundation item: National Science Foundation for Distinguished Youth Scholar (30225038) and the Major State Basic Research Development Program of China (00CB510103).
^*Corresponding author. Tel.: 86-27-61252507

周咏明, 郭伟, 周浩, 李慧玉, 刘黎琼, 姚军霞, 郑金娥, 郭天南, 黄士昂^*. 曲古菌素A抑制HL-60细胞端粒酶活性及hTERT的表达并诱导凋亡[J]. .

ZHOU Yong-ming, GUO Wei, ZHOU Hao, LI Hui-yu, LIU Li-qiong, YAO Jun-xia, ZHENG Jin-e, GUO Tian-nan, HUANG Shi-ang^*. Trichostatin A Induces Apoptosis by Inhibiting Telomerase Activity and Expression of Telomerase Reverse Transcriptase in HL-60 Cells[J]. .

参考文献

[1] Robbins AR, Jablonski SA, Yen TJ , et al. Inhibitors of histone deacetylases alter kinetochore assembly by disrupting pericentromeric heterochromatin [J]. Cell Cycle, 2005, 4 (5) : 717-726.
[2] Hu J , Colburn NH. Histone deacetylase inhibition downregulates cyclin D1 transcription by inhibiting nuclear factor-kappaB/p65 DNA binding [J]. Mol Cancer Res, 2005, 3 (2) : 100-109.
[3] Garcia-Morales P, Gomez-Martinez A, Carrato A, et al. Histone deacetylase inhibitors induced caspase-independent apoptosis in human pancreatic adenocarcinoma cell lines [J]. Mol Cancer Ther, 2005, 4 (8) : 1222-1230.
[4] Romanski A, Bacic B, Bug G, et al. Use of a novel histone deacetylase inhibitor to induce apoptosis in cell lines of acute lymphoblastic leukemia [J]. Haematologica, 2004, 89 (4) : 419-426.
[5] Marks P, Rifkind RA, Richon VM, et al. Histone deacetylases and cancer: causes and therap ies [J]. Nat Rev Cancer, 2001, 1 (3) : 194-202.
[6] Kim DH, Kim M, Kwon HJ. Histone deacetylase in carcinogenesis and its inhibitors as anti-cancer agents [J]. J Biochem Mol Biol, 2003, 36 (1) : 110-119.
[7] Takakura M, Kyo S, Kanaya T, et al. Cloning of human telomerase catalytic subunit (hTERT) gene promoter and identification of proximal core promoter sequences essential for transcrip tional activation in immortalized and cancer cells [J]. Cancer Res, 1999, 59 (3) : 551-557.
[8] Binz N, Shalaby T, Rivera P, et al. Telomerase inhibition, telomere shortening, cell growth supp ression and induction of apoptosis by telomestatin in childhood neuroblastoma cells [J]. Eur J Cancer, 2005, 41 (18) : 2873-2881.
[9] Zhu B, Zhang LH, Zhao YM, et al. 8-Chloroadenosine induced HL-60 cell growth inhibition, differentiation, and G(0)/G(1) arrest involve attenuated cyclin D1 and telomerase and up-regulated p21 (WAF1/CIP1 ) [J]. J Cell Biochem , 2006, 97 (1) : 166-177.
[10] Crowe DL, Nguyen DC, Ohannessian A. Mechanism of telomerase repression during terminal differentiation of normal epithelial cells and squamous carcinoma lines [J]. Int J Oncol, 2005, 27 (3) : 847-854.
[11] Tomlinson RL, Ziegler TD, Supakorndej T, et al. Cell cycle-regulated trafficking of human telomerase to telomeres [J]. Mol Biol Cell, 2006, 17 (2) : 955-965.
[12] Jady BE, Richard P, Bertrand E, et al. Cell cycle-dependent recruitment of telomerase RNA and Cajal bodies to human telomeres [J]. Mol Biol Cell, 2006, 17 (2) : 944-954.
[13] Richon VM, Sandhoff TW, Rifkind RA, et al. Histone deacetylase inhibitor selectively induces P^21WAF1 expression and gene-associated histone acetylation [J]. Proc Natl Acad Sci USA, 2000, 97 (18) : 10014-10019.
[14] Kim MS, Kwon HJ, Lee YM, et al. Histone deacetylases induce angiogenesis by negative regulation of tumor suppressor genes [J]. Nat Med, 2001, 7 (4) : 437-443.
[15] Endoh T, Tsuji N, Asanuma K, et al. Survivin enhances telomerase activity via up-regulation of specificity protein 1- and c-Myc-mediated human telomerase reverse transcriptase gene transcription [J]. Exp Cell Res, 2005, 305 (2) : 300-311.
[16] Artandi SE, Attardi LD. Pathways connecting telomeres and P⁵³ in senescence, apop tosis, and cancer [J]. Biochem Biophys Res Commun, 2005, 331 (3) : 881-890.
[17] Tsurutani J , Soda H, Oka M, et al. Antip roliferative effects of the histone deacetylase inhibitor FR901228 on small-cell lung cancer lines and drug-resistant sublines [J]. Int J Cancer, 2003, 104 (2) : 238-242.
[18] Bartoli A, Fettucciari K, Fetriconi I, et al. Effect of trichostatin A and 5’-azacytidine on transgene reactivation in U937 transduced cells [J]. Pharmacol Res, 2003, 48 (1) : 111-118.
[19] Kosugi H, Towatari M, Hatano S, et al. Histone deacetylase inhibitors are the potent inducer/enhancer of differentiation in acute myeloid leukemia: a new approach to anti-leukemia therapy[J]. Leukemia, 1999, 13 (9) : 1316-1324.
[20] Matsutani N, Yokozaki H, Tahara E, et al. Expression of MRE11 complex (MRE11, RAD50, NBS1) and hRap1 and its relation with telomere regulation, telomerase activity in human gastric carcinomas [J]. Pathobiology, 2001, 69 (4) : 219-224.
[21] Nagao K, Katsumata K, Aizawa Y, et al. Differential alternative splicing expressions of telomerase reverse transcrip tase in gastrointestinal cell lines [J]. Oncol Rep, 2004, 11 (1) : 127-131.
[22] Guo QL, L in SS, You QD, et al. Inhibition of human telomerase reverse transcrip tase gene exp ression by gambogic acid in human hepatoma SMMC-7721 cells [J]. Life Sci, 2006, 78 (11) : 1238-1245.
[23] Wang Z, Kyo S, Maida Y, et al. Tamoxifen regulates human telomerase reverse transcrip tase (hTERT) gene expression differently in breast and endometrial cancer cells [J]. Oncogene, 2002, 21 (22) : 3517-3524.