Ma Xing, Wang Jiening, Zhuang Juhua, Ma Xiaokun, Zheng Ni, Song Yanan, Xia Wei
Department of Nuclear Medicine, The Seventh People's Hospital of Shanghai University of Traditional Chinese Medicine, Shanghai 200137, P.R. China.
Central Laboratory, The Seventh People's Hospital of Shanghai University of Traditional Chinese Medicine, Shanghai 200137, P.R. China.
Oncol Lett. 2020 Jul;20(1):257-265. doi: 10.3892/ol.2020.11569. Epub 2020 Apr 23.
The aim of the present study was to investigate the role of prolyl 4-hydroxylase beta polypeptide (P4HB) in the chemoresistance of liver cancer. Drug-resistant liver cancer cell lines, such as HepG2/adriamycin (ADR) cells, were treated and screened using adriamycin. Gene interference was used to silence the expression of in liver cancer cells. Cell viability, invasiveness and migration were assessed using CCK8, Transwell and wound healing assays, respectively. In addition, changes to key genes and proteins in the epithelial-mesenchymal transition (EMT) and β-catenin/Snail pathway were analyzed using reverse transcription-quantitative PCR and western blotting. Drug-resistant HepG2/ADR cells were successfully cultivated; the IC to ADR for HepG2/ADR and HepG2 cell lines was 4.85 and 0.61 µM, respectively. HepG2/ADR cells exhibited higher invasion and migration abilities compared with HepG2 cells (P<0.05). E-cadherin mRNA and protein expression levels in HepG2/ADR cells were decreased significantly, whereas P4HB, N-cadherin and vimentin mRNA and protein levels were significantly increased compared with HepG2 cells (all P<0.05). Knockdown of P4HB significantly decreased cell viability and the invasion and migration ability of HepG2/ADR cells. In addition, P4HB knockdown enhanced E-cadherin mRNA and protein expression levels, whereas N-cadherin, vimentin, total β-catenin, nuclear β-catenin and Snail mRNA and protein levels were significantly decreased (all P<0.05). Overall, the present study demonstrated that EMT and β-catenin/Snail pathway influence P4HB modulation in liver cancer chemoresistance.
本研究的目的是探讨脯氨酰4-羟化酶β多肽（P4HB）在肝癌化疗耐药中的作用。使用阿霉素处理并筛选耐药肝癌细胞系，如HepG2/阿霉素（ADR）细胞。采用基因干扰技术沉默肝癌细胞中P4HB的表达。分别使用CCK8、Transwell和伤口愈合实验评估细胞活力、侵袭能力和迁移能力。此外，采用逆转录定量PCR和蛋白质印迹法分析上皮-间质转化（EMT）和β-连环蛋白/Snail通路中关键基因和蛋白质的变化。成功培养出耐药HepG2/ADR细胞；HepG2/ADR和HepG2细胞系对阿霉素的半数抑制浓度（IC）分别为4.85和0.61μM。与HepG2细胞相比，HepG2/ADR细胞表现出更高的侵袭和迁移能力（P<0.05）。与HepG2细胞相比，HepG2/ADR细胞中E-钙黏蛋白的mRNA和蛋白质表达水平显著降低，而P4HB、N-钙黏蛋白和波形蛋白的mRNA和蛋白质水平显著升高（均P<0.05）。敲低P4HB可显著降低HepG2/ADR细胞的活力、侵袭和迁移能力。此外，敲低P4HB可增强E-钙黏蛋白的mRNA和蛋白质表达水平，而N-钙黏蛋白、波形蛋白、总β-连环蛋白、核β-连环蛋白和Snail的mRNA和蛋白质水平显著降低（均P<0.05）。总体而言，本研究表明EMT和β-连环蛋白/Snail通路影响肝癌化疗耐药中P4HB的调控。
