Jia-Jun Tian, Su-Mei Lu, Liang Yu, Ju-Ke Ma, Ya-Kui Mu, Hai-Bo Wang, Wei Xu
Department of Otolaryngology-Head and Neck Surgery, Provincial Hospital affiliated to Shandong University, Jinan 250021, PR, China.
Institute of Eye & Otolaryngology, Shandong Clinic Research Institute, Jinan 250021, PR, China.
Head Neck Oncol. 2012 Mar 27;4:7. doi: 10.1186/1758-3284-4-7.
The objective of this study was to evaluate the efficacy of Nimesulide, a selective cyclooxygenase-2 (COX-2) inhibitor, on the growth of hypopharyngeal carcinoma cells (FaDu) in vitro, and investigate its potential mechanism.
After FaDu cells were treated with graded concentrations of Nimesulide for divergent time, sensitivity of cells to drug treatment was analyzed by MTT assay. Morphological changes of FaDu cells in the presence of Nimesulide were observed by acridine orange cytochemistry staining. Proliferating cells were detected using the 5-Bromo-2'-deoxy-uridine (BrdU) incorporation assay. Following cells were subjected to Nimesulide (500 μmol/l) for 6 h, 12 h and 24 h, the percentage of apoptosis was examined by flow cytometry. We detected COX-2 and Survivin expression change by RT-PCR and Western blot, and analyzed the correlation of them with the growth of FaDu cells. Additionally, we also analyzed Caspase-3, Bcl-2 and Bax expressions as markers to investigate the related pathway of Nimesulide-indued apoptosis.
Compared with the control group, the viabilities rates were decreased by Nimesulide in time- and dose-dependent manners, typical morphological changes of apoptotic cells were observed in the Nimesulide-treatment groups, Nimesulide could suppress the proliferation of FaDu cells significantly. The percentage of apoptosis in FaDu cells were markedly increased after Nimesulide-treatment for 6 h, 12 h and 24 h. Nimesulide down-regulated the Survivin and COX-2 expressions at mRNA and protein levels in FaDu cells. Additional analyses indicated that Bcl-2 expression was significantly decreased and the expressions of Caspase-3 as well as Bax were increased at both mRNA and protein levels.
Based on the induction of apoptosis and suppression of proliferation, Nimesulide could inhibit the growth of FaDu cells. Furthermore, the suppression of Survivin expression may play an important role in Nimesulide-induced growth inhibition. Nimesulide could act as an effective therapeutic agent for hypopharyngeal carcinoma therapy.
本研究的目的是评估选择性环氧化酶-2（COX-2）抑制剂尼美舒利对下咽癌细胞（FaDu）体外生长的疗效，并探讨其潜在机制。
用不同浓度的尼美舒利处理FaDu细胞不同时间后，通过MTT法分析细胞对药物处理的敏感性。用吖啶橙细胞化学染色观察尼美舒利存在时FaDu细胞的形态变化。使用5-溴-2'-脱氧尿苷（BrdU）掺入法检测增殖细胞。细胞分别用500μmol/l尼美舒利处理6小时、12小时和24小时后，通过流式细胞术检测凋亡百分比。通过RT-PCR和蛋白质印迹法检测COX-2和Survivin表达变化，并分析它们与FaDu细胞生长的相关性。此外，我们还分析了Caspase-3、Bcl-2和Bax的表达作为标志物，以研究尼美舒利诱导凋亡的相关途径。
与对照组相比，尼美舒利以时间和剂量依赖性方式降低细胞活力，在尼美舒利处理组中观察到凋亡细胞的典型形态变化，尼美舒利可显著抑制FaDu细胞的增殖。尼美舒利处理6小时、12小时和24小时后，FaDu细胞的凋亡百分比明显增加。尼美舒利在mRNA和蛋白质水平下调FaDu细胞中Survivin和COX-2的表达。进一步分析表明，Bcl-2表达显著降低，Caspase-3和Bax在mRNA和蛋白质水平的表达均增加。
基于诱导凋亡和抑制增殖，尼美舒利可抑制FaDu细胞的生长。此外，Survivin表达的抑制可能在尼美舒利诱导的生长抑制中起重要作用。尼美舒利可作为下咽癌治疗的有效治疗剂。
