Sanaei Masumeh, Kavoosi Fraidoon, Hosseini Farzane
Research Center for Non-Communicable Diseases, Jahrom University of Medical Sciences, Jahrom, Iran.
Student of Research Committee, Jahrom University of Medical Sciences, Jahrom, Iran.
Iran J Pharm Res. 2020 Fall;19(4):193-202. doi: 10.22037/ijpr.2020.112223.13614.
Tumorigenesis must be understood as a summary of altered genetic and genomic changes resulting in the inactivation of tumor suppressor genes (TSGs). One of the characterizations of epigenetic alterations is DNA methylation. Epigenetic alteration of the p16INK4a, p14ARF, p15INK4b and DNA methyltransferase 1 gene (DNMT1) expression occurs in hepatocellular carcinoma (HCC) and pancreatic cancer frequently. DNA methyltransferase inhibitors (DNMTIs), such as zebularine, play a significant effect on the demethylation and reactivation of TSGs. This study aimed to investigate the effect of zebularine on p16INK4a, p14ARF, p15INK4b, and DNA methyltransferase 1 gene expression, cell growth inhibition, and apoptosis induction in HCC PLC/PRF5 and pancreatic cancer PA-TU-8902 cell lines. Both cell lines were cultured and treated with zebularine at different times. The MTT assay, real-time quantitative reverse-transcription polymerase chain reaction (qRT-PCR), and flow cytometry were used to determine cell viability, gene expression, and apoptotic cells, respectively. The result indicated that zebularine inhibited cell growth of both cell lines significantly as time- and dose-dependent manner ( 0.007). The agent induced significant down-regulation of DNMT1 and up-regulation of p16INK4a, p14ARF, p15INK4b ( 0.028). Besides, it had a significant apoptosis effect on both cell lines ( 0.001). This compound had a strong significant effect on PLC/PRF5 in comparison to PA-TU-8902 cells. Concluding, zebularine inhibited PLC/PRF5 and PA-TU-8902 cell growth and induced apoptosis in these cell lines. The most likely mechanism underlying the zebularine played its role involves down-regulation of DNMT1 and up-regulation of p16INK4a, p14ARF, and p15INK4b genes.
肿瘤发生必须被理解为导致肿瘤抑制基因（TSGs）失活的遗传和基因组变化改变的总结。表观遗传改变的特征之一是DNA甲基化。p16INK4a、p14ARF、p15INK4b和DNA甲基转移酶1基因（DNMT1）表达的表观遗传改变在肝细胞癌（HCC）和胰腺癌中频繁发生。DNA甲基转移酶抑制剂（DNMTIs），如泽布勒林，对TSGs的去甲基化和重新激活具有显著作用。本研究旨在探讨泽布勒林对HCC PLC/PRF5和胰腺癌PA-TU-8902细胞系中p16INK4a、p14ARF、p15INK4b和DNA甲基转移酶1基因表达、细胞生长抑制和凋亡诱导的影响。两种细胞系均在不同时间用泽布勒林培养和处理。MTT法、实时定量逆转录聚合酶链反应（qRT-PCR）和流式细胞术分别用于测定细胞活力、基因表达和凋亡细胞。结果表明，泽布勒林以时间和剂量依赖性方式显著抑制两种细胞系的细胞生长（0.007）。该药物诱导DNMT1显著下调，p16INK4a、p14ARF、p15INK4b显著上调（0.028）。此外，它对两种细胞系均有显著的凋亡作用（0.001）。与PA-TU-8902细胞相比，该化合物对PLC/PRF5有强烈的显著作用。总之，泽布勒林抑制PLC/PRF5和PA-TU-8902细胞生长并诱导这些细胞系凋亡。泽布勒林发挥作用的最可能机制涉及DNMT1的下调和p16INK4a、p14ARF和p15INK4b基因的上调。
