软坚散结方下调miR-182表达调控化疗启动EMT在抗肺癌复发转移作用机制研究

The nature of easy recurrence and metastasis after surgery remains one of the most challenging for lung cancer treatment. Because of definciency of healthy Qi, then blood stasis, and phlegm accumulation.The therapy of resolving hard lump could help to prevent the occurrence and propagation of this “phlegm stagnation”.From our previous study, we found the formula of RuanJianSanJie (RJSJ)effectively inhibited recurrence and metastasis after lung cancer surgery on the nude mice and down-regulated the expression level of miR-182.Meanwhile,Some studies have showed that miR-182 plays an double effect role in the progress of lung cancer, and still need further study. In this present study, firstly, we try to clarify the potential explanation of the mechanism behind the role of miR-182and RJSJ in the actions of adhesion, migration, invasion and metastasis of lung cancer. For this purpose, we need to observe the differences of expression of miR-182 between lung cancer cell lines with the potential of proliferation, migration and metastasis in vitro studies and perform transwell migration assay and invasion assay as well through technique of cell biology and molecular biology. We also sclient the miR-182 to evaluate the regulation mechanism of EMT process induced by chemotherapy . On the other hand, we need to carry out a series of in vivo studies which serve to prove the effect of formula RJSJ on prevention the recurrence and metastasis of lung cancer after surgery. For instance, we can dynamically observe the change of metastatic lesion on the NCI-H460 xenograted nude mice by IVIS Lumina and other parameters like the number of nudes in lung. The mRNA and protein level of EMT related genes such as Twist, Snial,E-cadherin, N-cadherin, etc. will be detected by Molecular biology. We hypothesize the reason behind the inhibition effect of formala RJSJ on recurrence and metastasis of post-operative lung cancer is that, it may down regulate miR-182 though chemotherapy-induced EMT for RJSJ formula in lung cancer.Furthermore, our study aim to lay the foundation for the clinical treatment and the development of new targeted drug.

肺癌术后易复发转移是临床治疗的难点。正气不足，痰瘀毒聚是肺癌发生的关键，软坚散结法从源头截断“痰浊”滋生，阻止肿瘤复发转移。前期研究发现，软坚散结方能下调miR-182表达并抑制肺癌术后的复发转移。而miR-182可能在肺癌的发展中发挥双重作用，仍需深入研究。本研究一方面利用细胞生物学和分子生物学技术，借助Transwell和划痕实验等观察mi-R182表达以及软坚散结方干预后对肺癌细胞粘附、迁移和侵袭的影响；沉默miR-182表达评价其在化疗诱导的EMT进程中的调控作用。另一方面，建立裸鼠肺癌术后复发转移模型，利用小动物活体成像系统，动态监测肺结节的发生，分子生物学检测EMT相关Twist、Snail、E-cadherin、N-cadherin等的表达。阐明软坚散结方可能通过下调mi-R182表达调控化疗触发的EMT抑制肺癌复发转移的机制，为指导肺癌临床治疗以及开发新型靶向药物奠定基础。

肺癌术后易复发转移是临床治疗的难点。正气不足，痰瘀毒聚是肺癌发生的关键，软坚散结法从源头截断“痰浊”滋生，阻止肿瘤复发转移。前期研究发现，软坚散结方能下调miR-182表达并抑制肺癌术后的复发转移。而miR-182可能在肺癌的发展中发挥双重作用，仍需深入研究。本研究一方面利用细胞生物学和分子生物学技术，借助Transwell和划痕实验等观察mi-R182表达以及软坚散结方干预后对肺癌细胞粘附、迁移和侵袭的影响；过表达或者沉默miR-182表达评价其在EMT进程中的调控作用。另一方面，建立裸鼠肺癌术后复发转移模型，利用小动物活体成像系统，监测肺转移瘤的发生，分子生物学检测EMT相关Twist、Snail、E-cadherin、N-cadherin等的表达。阐明软坚散结方通过下调mi-R182表达靶向EPAS1调控EMT进程抑制肺癌复发转移的机制，为指导肺癌临床治疗以及开发新型靶向药物奠定基础。