The role of NLRP3 inflammasome in 5-fluorouracil resistance of oral squamous cell carcinoma.
The role of NLRP3 inflammasome in 5-fluorouracil resistance of oral squamous cell carcinoma.
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NLRP3炎症小体在口腔鳞癌5-氟尿嘧啶耐药中的作用
DOI：
10.1186/s13046-017-0553-x
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发表时间：
2017-06-21
期刊：
影响因子：
--
通讯作者：
Chen F
中科院分区：
文献类型：
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作者：
Feng X;Luo Q;Zhang H;Wang H;Chen W;Meng G;Chen F
研究背景5-氟尿嘧啶（5-FU）是一种广泛应用于肿瘤治疗的药物。然而，肿瘤细胞对5-FU的耐药性通常限制了其临床有效性。方法采用真实的时间定量PCR（qRT-PCR）和western blot方法分别检测口腔鳞癌组织中NLRP 3、Caspase 1和IL-1β的mRNA和蛋白表达水平。采用免疫组化法检测石蜡包埋的口腔鳞癌组织中NLRP 3和Ki-67的表达。通过评估NLRP 3和IL-1β在没有或有NLRP 3敲低的OSCC细胞系中的表达，进一步检查5-FU处理与NLRP 3炎性体的表达和活化之间的相关性。MTT法检测细胞活力;流式细胞仪检测5-FU诱导OSCC细胞凋亡及细胞内活性氧（ROS）的变化。通过连续口服4-硝基喹啉1-氧化物在野生型BALB/c、Nlrp 3 −/−和Caspase 1 −/−小鼠中建立致癌物诱导的舌鳞状细胞癌小鼠模型。观察肿瘤发生率并评估肿瘤面积。结果临床分析显示，接受5-FU化疗的OSCC患者组织中NLRP 3炎性体的表达和激活明显增加。多因素考克斯回归分析显示，这种高表达与肿瘤分期和分化程度显著相关，与预后不良相关。此外，在细胞培养系统和异种移植小鼠模型中，5-FU处理增加了OSCC细胞中NLRP 3炎性体的表达和活化。沉默NLRP 3表达可显著抑制OSCC细胞增殖并增强5-FU诱导的OSCC细胞凋亡。进一步研究表明，5-FU诱导的细胞内活性氧促进NLRP 3炎性体的表达和激活，增加白细胞介素（IL）-1β的产生，从而介导化疗耐药性。在致癌物诱导的OSCC模型中，我们发现Nlrp 3 −/−和Caspase 1 −/−小鼠的肿瘤发病率低于野生型小鼠，且发病时间较晚。结论NLRP 3炎性小体在体内外均能促进OSCC对5-FU的耐药，靶向ROS/NLRP 3炎性小体/IL-1β信号通路可能有助于OSCC的5-FU辅助化疗。
Background5-Fluorouracil (5-FU) is a widely used drug for the therapy of cancer. However, the chemoresistance of tumor cells to 5-FU usually limits its clinical effectiveness. In this study, we explored the role of NLRP3 inflammasome in 5-FU resistance of oral squamous cell carcinoma (OSCC).MethodsThe mRNA and protein expression levels of NLRP3, Caspase1 and IL-1β in resected OSCC specimens or cell lines were measured respectively by quantitative real time-PCR (qRT-PCR) and western blot. NLRP3 and Ki-67 expression in paraffin-embedded OSCC tissues was determined by immunohistochemistry. The correlation between 5-FU treatment and the expression and activation of NLRP3 inflammasome was further examined by evaluating NLRP3 and IL-1β expression in OSCC cell lines without or with NLRP3 knocked down. Cell viabilities of OSCC cells were determined by the MTT assay. Apoptosis and intracellular reactive oxygen species (ROS) of OSCC cells induced by 5-FU were measured by the flow cytometer. The carcinogen-induced tongue squamous carcinoma mice model was established by continuous oral administration of 4-nitroquinoline 1-oxide in wild-type BALB/c,Nlrp3−/−andCaspase1−/−mice. Tumor incidence were observed and tumor area were evaluated.ResultsIn the clinical analysis, expression and activation of NLRP3 inflammasome was clearly increased in OSCC tissues of patients who received 5-FU-based chemotherapy. Multivariate Cox regression analysis revealed that this high expression was significantly correlated with tumor stage and differentiation, and was associated with poor prognosis. Moreover, 5-FU treatment increased expression and activation of NLRP3 inflammasome in OSCC cells in a cell culture system and xenograft mouse model. Silencing of NLRP3 expression significantly inhibited OSCC cell proliferation and enhanced 5-FU-induced apoptosis of OSCC cells. Further investigation showed that intracellular ROS induced by 5-FU promoted the expression and activation of NLRP3 inflammasome and increased the production of interleukin (IL)-1β, which then mediated the chemoresistance. With the carcinogen-induced OSCC model, we found less and later tumor incidence inNlrp3−/−andCaspase1−/−mice than wild-type mice. And greater decrease of tumor area was observed in the gene deficient mice treated with 5-FU.ConclusionsOur findings suggest that NLRP3 inflammasome promoted 5-FU resistance of OSCC both in vitro and in vivo, and targeting the ROS/NLRP3 inflammasome/IL-1β signaling pathway may help 5-FU-based adjuvant chemotherapy of OSCC.