Feng Yu, Chen Ke, Pan Liangbin, Jiang Wei, Pang Pei, Mao Guocai, Zhang Biao, Chen Shaomu
Department of Thoracic Surgery, the First Affiliated Hospital of Soochow University, Suzhou, China.
Department of Pathology, the First Affiliated Hospital of Soochow University, Suzhou, China.
J Thorac Dis. 2021 Jan;13(1):299-311. doi: 10.21037/jtd-20-3461.
This study aimed to investigate the relationship between RNA polymerase II subunit 5 (RPB5)-mediating protein (RMP) and clinicopathological characteristics of non-small cell lung cancer (NSCLC) patients by measuring the expression level of RMP in human NSCLC tissues and cell lines. At the same time, we studied the impact of RMP on the biological function of cancer, providing strong support for gene targeted therapy of NSCLC.
Real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot were used to determine the expression levels of messenger (m)RNA and protein in NSCLC cell lines and tissues. Cell counting kit 8 (CCK8) assay and flow cytometry were selected to detect cell proliferation, cycle and apoptosis. The wound healing assay was chosen to detect the migration and invasion ability of cells. The xenograft model was performed to study the function of RMP in vivo. Immunohistochemical (IHC) staining showed the levels of RMP, Bcl-2, Bax and caspase-3.
First, mRNA and protein levels of RMP were relatively overexpressed in NSCLC cells. Compared with the corresponding normal tissues, the mRNA and protein levels of RMP were significantly higher in human NSCLC tissues. Concurrently, we found that the expression of RMP was related to the status of lymph nodes (LNs) in cancer tissues and T stage. Then, RMP overexpression promoted the proliferation of A549. At the same time, RMP provided A549 cells the ability to resist chemotherapy and radiotherapy; when A549 cells were treated with gefitinib and radiation, RMP reduced apoptosis. We also found that RMP can protect A549 from G2 block caused by radiation. Over-irradiated RMP-overexpressed A549 cells had lower Bcl2-associated X protein (Bax) levels and higher B-cell lymphoma 2 (Bcl-2) levels. The migration and invasion ability of A549 cells was increased by RMP. Finally, RMP can promote tumor growth by increasing Bcl-2 levels and decreasing Bax and caspase-3 levels in the xenograft model.
There is potential for RMP to develop into a diagnostic and therapeutic target for NSCLC.
本研究旨在通过检测人非小细胞肺癌（NSCLC）组织及细胞系中RNA聚合酶II亚基5（RPB5）介导蛋白（RMP）的表达水平，探讨其与NSCLC患者临床病理特征的关系。同时，研究RMP对肿瘤生物学功能的影响，为NSCLC的基因靶向治疗提供有力支持。
采用实时定量逆转录聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法检测NSCLC细胞系及组织中信使（m）RNA和蛋白质的表达水平。选用细胞计数试剂盒8（CCK8）法和流式细胞术检测细胞增殖、周期及凋亡情况。采用划痕实验检测细胞的迁移和侵袭能力。建立异种移植模型研究RMP在体内的功能。免疫组织化学（IHC）染色检测RMP、Bcl-2、Bax和半胱天冬酶-3的水平。
首先，RMP的mRNA和蛋白质水平在NSCLC细胞中相对过表达。与相应正常组织相比，人NSCLC组织中RMP的mRNA和蛋白质水平显著升高。同时，我们发现RMP的表达与癌组织中的淋巴结（LNs）状态和T分期有关。其次，RMP过表达促进A549细胞增殖。同时，RMP赋予A549细胞抗化疗和放疗的能力；当A549细胞用吉非替尼和辐射处理时，RMP减少细胞凋亡。我们还发现RMP可保护A549细胞免受辐射引起的G2期阻滞。过度照射的RMP过表达A549细胞中Bcl2相关X蛋白（Bax）水平较低，B细胞淋巴瘤2（Bcl-2）水平较高。RMP增强了A549细胞的迁移和侵袭能力。最后，在异种移植模型中，RMP可通过提高Bcl-2水平、降低Bax和半胱天冬酶-3水平促进肿瘤生长。
RMP有潜力发展成为NSCLC的诊断和治疗靶点。
