Cancer Research Center, Qilu Hospital of Shandong University, No. 107 Cultural West Road, Jinan, 250012, Shandong, China.
No.2 Comprehensive Department, Qilu Hospital of Shandong University, Jinan, 250012, Shandong, China.
Pathol Oncol Res. 2019 Oct;25(4):1487-1495. doi: 10.1007/s12253-018-0504-7. Epub 2018 Nov 1.
This study aimed to investigate the expression of secreted phosphoprotein 1 (SPP1) on lung cancer cells and explore its underlying mechanism on autophagy and apoptosis which effect the development of lung cancer cells. GSE19804 related to lung cancer cells was screened from Gene Expression Omnibus (GEO) database, and we screened the 47 pairs of differential expressed mRNAs in lung cancer cells and adjacent tissues using microarray analysis. The expression of the core gene SPP1 was detected by qRT-PCR and western-blot. The transfection efficiency of lung cancer cells was detected by qRT-PCR and the expression of transfected group was tested by western-blot. Cell proliferation after transfection was tested by MTT assay and plate cloning experiment. The apoptosis rate of each transfection group was detected by flow cytometry. We use western-blot to test protein expression of autophagy-related proteins Beclin-1, LC3-I, LC3-II and p62 of each transfected group. Through analysis of GSE19804,the heat map showed SPP1 was the highest expressed in tumor tissues. qRT-PCR and western-blot detected SPP1 expression in lung cancer tissues was higher than that in normal adjacent tissues and was significantly increased in lung cancer cell lines. After transfection with pcDNA3.1-SPP1 (p-SPP1 group), siRNA1-SPP1 (siRNA1 group) and siRNA2-SPP1 (siRNA2 group), showed different expression of SPP1. Up-regulation of SPP1 enhanced cell viability and promoted tumor cell proliferation, while knockdown of SPP1 inhibited tumor cell proliferation. From the results of apoptosis rate, SPP1 inhibited the tumor cell apoptosis. However, in normal lung cell, SPP1 had no effect on cell proliferation and apoptosis. And to test autophagy-related proteins, we found that overexpression of SPP1 inhibited autophagy. High expression of SPP1 inhibited autophagy and apoptosis to promote the development of small cell lung cancer cells.
本研究旨在探讨分泌磷蛋白 1（SPP1）在肺癌细胞中的表达，并探讨其对自噬和凋亡的潜在机制，这些机制影响肺癌细胞的发展。从基因表达综合数据库（GEO）筛选与肺癌细胞相关的 GSE19804，通过微阵列分析筛选肺癌细胞及相邻组织中差异表达的 47 对 mRNAs。qRT-PCR 和 Western blot 检测核心基因 SPP1 的表达。qRT-PCR 检测肺癌细胞的转染效率，Western blot 检测转染组的表达。MTT 检测和平板克隆实验检测转染后细胞增殖情况。流式细胞术检测各组细胞凋亡率。Western blot 检测各组细胞自噬相关蛋白 Beclin-1、LC3-I、LC3-II 和 p62 的蛋白表达。通过对 GSE19804 的分析，热图显示 SPP1 在肿瘤组织中表达最高。qRT-PCR 和 Western blot 检测 SPP1 在肺癌组织中的表达高于正常相邻组织，在肺癌细胞系中明显升高。转染 pcDNA3.1-SPP1(p-SPP1 组)、siRNA1-SPP1(siRNA1 组)和 siRNA2-SPP1(siRNA2 组)后，SPP1 表达水平不同。上调 SPP1 增强细胞活力，促进肿瘤细胞增殖，而下调 SPP1 抑制肿瘤细胞增殖。从凋亡率结果来看，SPP1 抑制肿瘤细胞凋亡。然而，在正常肺细胞中，SPP1 对细胞增殖和凋亡没有影响。检测自噬相关蛋白发现，SPP1 过表达抑制自噬。SPP1 的高表达抑制自噬和凋亡，促进小细胞肺癌细胞的发展。
