NLRP3炎性体在细菌脂多糖与煤焦沥青诱导炎症相关肺癌发生中的促进作用及其机制

Lung cancer is the malignant tumor with the highest morbidity and mortality in the world. Studies demonstrated that chronic inflammation could promote initiation and development of lung cancer, and NLRP3 inflammasome is not only involved in the formation of inflammation, but also tumors. However, there is not any report on the role of NLRP3 inflammasome in occurrence of inflammation-related lung cancer, as well as the mechanism is poorly understood. In this study, firstly, human bronchial epithelial cells (BEAS-2B) are utilized as the in vitro model induced by lipopolysaccharide (LPS) and coal tar pitch extract (CTPE), RNA interference is used to explore the role of NLRP3 inflammasome in LPS/CTPE-induced tumorigenic transformation of BEAS-2B cells. Secondly, wide type (WT) mice and NLRP3-/- mice are utilized to build animal model in vivo also induced by LPS and CTPE, the rate of lung cancer occurrence at many stages, especially at early stage, in NLRP3-/- mice is compared with that in WT mice to verify the role of NLRP3 inflammasome. Inflammsome activation-related factors are determined to investigate the mechanism in vitro and in vivo. It’s meaningful to explore the role and the mechanism of NLRP3 inflammasome in initiation of inflammation-related lung cancer, which will contribute to the warning of lung cancer at early stage and the prevention of initiation of lung cancer.

肺癌是世界上发病率和死亡率最高的恶性肿瘤，目前研究发现慢性炎症能够促进肺癌的发生发展；NLRP3炎性体不仅参与炎症的形成，而且与肿瘤密切相关，但NLRP3炎性体在炎症相关肺癌发生中的作用及相关机制未见报道。本研究预采用细菌脂多糖和煤焦沥青加热烟气提取物为染毒剂，作用于人支气管上皮细胞系（BEAS-2B）构建炎症相关肺癌的体外模型，通过RNA干扰技术检测NLRP3炎性体在BEAS-2B细胞恶性转化中的作用，并分析细胞因子和NLRP3炎性体激活相关因子的表达；同时构建炎症相关肺癌的动物模型，通过比较野生型小鼠和NLRP3基因敲除小鼠（NLRP3-/-小鼠）在炎症相关肺癌发生的各个阶段，特别是早期阶段的肺癌发生情况，从体内、体外研究综合分析NLRP3炎性体在细菌脂多糖和煤焦沥青诱导炎症相关肺癌发生中的作用和可能机制，为肺癌的早期预警和干预提供新的思路。

肺癌是世界上发病率和死亡率最高的恶性肿瘤，环境污染和吸烟诱发的慢性肺部炎症能够促进肺癌的发生发展，NLRP3炎性体与炎症和肿瘤均关系密切，但其在炎症相关肺癌发生中的作用未见报道。本研究采用细菌脂多糖（Lipopolysaccharide , LPS）和煤焦沥青烟气提取物（Coal tar pitch extract, CTPE）联合染毒人支气管上皮细胞（BEAS-2B），单独CTPE、LPS、二甲基亚砜（DMSO）和生理盐水（Saline）作为对照，待细胞生长至30代，检测各组细胞平板克隆形成情况，结果发现LPS+CTPE组与单独CTPE组细胞平板克隆形成率均明显高于Saline组，且LPS+CTPE组细胞平板克隆形成率高于单独CTPE组(P<0.05)；Western blot检测 LPS+CTPE染毒BEAS-2B细胞Cleaved caspase-1 p10蛋白表达水平和Confocol检测NLRP3炎性复合体形成的阳性细胞率均高于单独CTPE染毒的细胞(P<0.05)。同时采用LPS和CTPE成分中代表性物质--苯并(a)芘[B(a)P]通过快速吸入式气管滴注给药系统联合染毒野生型小鼠（WT）和NLRP3基因敲除小鼠（NLRP3-/-），染毒后30周，发现LPS+ B(a)P染毒WT小鼠、NLRP3-/-小鼠肺部肉眼可见瘤体数、瘤体长径和病理切片癌巢数均比单独B(a)P染毒同种系的小鼠瘤体数多、瘤体长径长、癌巢数多(P<0.05)，而无论是LPS+ B(a)P组还是B(a)P组，NLRP3-/-小鼠与WT小鼠相比，其肺部肉眼可见瘤体数减少、瘤体长径短、癌巢数减少(P<0.05)。与单独B(a)P组相比，LPS+ B(a)P染毒WT小鼠肺组织IL-1β mRNA的表达水平明显减少(P<0.05)，而IL-18 mRNA的表达水平明显增高(P<0.05)。本研究成功构建了炎症相关肺癌的细胞模型和动物模型，验证了慢性肺部炎症能够促进肺癌的发生，并为研究肺癌、特别是炎症相关肺癌提供了良好的实验平台；体外实验和体内实验结果均显示NLPR3及其NLRP3炎性体活化参与介导LPS/ CTPE或LPS/B(a)P诱导的炎症相关肺癌发生，为探索肺癌的发生机制提供新的思路，为肺癌的早期干预提供新的线索。