Wang Rui, Li Bin, Wang Xi, Lin Fang, Gao Ping, Cheng Shi-Yin, Zhang Hui-Zhong
Clinical Laboratory Department, Tangdu Hospital, Fourth Military Medical University, Xinsi Road, 710038 Xi'an, Shaanxi Province, PR China.
Auris Nasus Larynx. 2009 Jun;36(3):332-9. doi: 10.1016/j.anl.2008.08.006. Epub 2008 Nov 13.
X-linked inhibitor of apoptosis protein (XIAP) is a novel member of the inhibitors of apoptosis (IAPs) family. The overexpression of XIAP is asscociated with radioresistance of human malignancies. The purpose of the present study was to investigate the effect of shRNA-targeted XIAP on the proliferation, apoptosis and radiosensitivity of human laryngeal carcinoma cells (Hep-2).
A siRNA expression vector (pSilencer4.1-XIAPshRNA) was constructed and stably transfected into human laryngeal carcinoma cells (Hep-2). The downregulation of XIAP expression was evaluated by RT-PCR and Western blot analyses. Then, we investigated the effect of XIAP-shRNA on the proliferation, cell cycle changes and apoptosis in vitro of Hep-2 cells. Finally, the radiosensitivity of Hep-2 cells was investigated by clonogenic cell survival assay.
We established stably transfected cell line (Hep-2/XIAPshRNA) in which the expression of XIAP gene was downregulated. The cell viability of Hep-2/XIAP-RNA cells was obviously decreased compared with that of untransfected Hep-2 cells. Morever, XIAP-shRNA induced cell arrest in the G(0)/G(1) phase of cell cycle by flow cytometry analysis. Results of TUNEL assay indicated that Hep-2 cells stably transfected pSilencer4.1-XIAP-shRNA showed obvious apoptosis characters. Furthermore, the downregulation of XIAP expression could lead to significant radiosensitivity enhancement in laryngeal carcinoma cells.
RNAi-mediated downregulation of XIAP expression can inhibit proliferation, induce apoptosis and diminish the radioresistance of laryngeal carcinoma cells, so combined therapy with XIAP inhibition and radiation may be a potential strategy for the treatment of laryngeal carcinoma.
X连锁凋亡抑制蛋白（XIAP）是凋亡抑制蛋白（IAPs）家族的一个新成员。XIAP的过表达与人恶性肿瘤的放射抗性相关。本研究的目的是探讨靶向XIAP的短发夹RNA（shRNA）对人喉癌细胞（Hep-2）增殖、凋亡及放射敏感性的影响。
构建siRNA表达载体（pSilencer4.1-XIAPshRNA）并稳定转染人喉癌细胞（Hep-2）。通过逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹分析评估XIAP表达的下调情况。然后，我们研究了XIAP-shRNA对Hep-2细胞体外增殖、细胞周期变化和凋亡的影响。最后，通过克隆形成细胞存活试验研究Hep-2细胞的放射敏感性。
我们建立了XIAP基因表达下调的稳定转染细胞系（Hep-2/XIAPshRNA）。与未转染的Hep-2细胞相比，Hep-2/XIAP-RNA细胞的细胞活力明显降低。此外，通过流式细胞术分析，XIAP-shRNA诱导细胞周期停滞在G(0)/G(1)期。末端脱氧核苷酸转移酶介导的缺口末端标记（TUNEL）分析结果表明，稳定转染pSilencer4.1-XIAP-shRNA的Hep-2细胞表现出明显的凋亡特征。此外，XIAP表达的下调可导致喉癌细胞放射敏感性显著增强。
RNA干扰介导的XIAP表达下调可抑制喉癌细胞增殖、诱导凋亡并降低其放射抗性，因此XIAP抑制与放疗联合治疗可能是治疗喉癌的一种潜在策略。
