Zhang Ying, Qian He-Sheng, Hu Gengwei, Wang Lu, Zhu Yiping
Department of Oncology, Fuyang Cancer Hospital, Fuyang, China.
Department of Cardiovascular Medicine, The First Affiliated Hospital with Nanjing Medical University, Nanjing, China.
J Gastrointest Oncol. 2024 Jun 30;15(3):862-872. doi: 10.21037/jgo-24-283. Epub 2024 Jun 27.
Defects in DNA damage repair can cause genetic mutations, which in turn can cause different types of cancers. Chromatin remodeling complexes, which help repair damaged DNA, can cause the chromatin structure to change as a result of DNA damage. ARID1A may play a role in the process of DNA damage repair, and arid1a may be related to the occurrence and development of gastric cancer (GC). This study aimed to investigate the mechanism of ARID1A regulating the DNA damage repair of gastric adenocarcinoma cell lines AGS and SGC-7901 and its effect on migration, proliferation and apoptosis.
The expression of ARID1A plasmid was detected by Western blot and real-time polymerase chain reaction (PCR). The effect of etoposide (ETO) on the survival rate of AGS and SGC-7901 gastric adenocarcinoma cell lines was detected by MTT assay. The DNA double-strand break model was established by ETO and then passed through the comet assay and immunofluorescence co-localization to observe DNA damage; western blot method was used to detect the effect of ARID1A on the expression of related proteins in DNA damage repair pathway in gastric adenocarcinoma cells; scratch test and colony formation experiments were used to observe ARID1A migration and proliferation of gastric adenocarcinoma cells. The flow cytometry was used to detect the effect of ARID1A on apoptosis of gastric adenocarcinoma cells.
The expression of mRNA and protein was increased after transfection of ARID1A plasmid. ETO was confirmed by MTT assay to inhibit cell survival in a dose-dependent manner. After the DNA double-strand break model was established by ETO, the expression levels of phospho-ataxia telangiectasia mutated (p-ATM) protein increased in the overexpressed ARID1A group. Meanwhile, the overexpressed ARID1A group had a shortened tail moment, and γ-H2AX and ARID1A co-localized in the DNA damage site of the nucleus. The over-expressed ARID1A group had weaker wound healing ability, reduced number of clone formation, and increased apoptosis rate.
ARID1A may repair DNA double-strand breaks caused by ETO by p-ATM pathway; ARID1A can inhibit the migration and proliferation of gastric adenocarcinoma cells and promote apoptosis. Our findings indicate that could serve as a therapeutic target and biomarker for GC patients.
DNA损伤修复缺陷可导致基因突变，进而引发不同类型的癌症。染色质重塑复合物有助于修复受损的DNA，其可因DNA损伤导致染色质结构发生改变。ARID1A可能在DNA损伤修复过程中发挥作用，且arid1a可能与胃癌（GC）的发生发展有关。本研究旨在探讨ARID1A调控胃腺癌细胞系AGS和SGC-7901的DNA损伤修复的机制及其对迁移、增殖和凋亡的影响。
采用蛋白质免疫印迹法（Western blot）和实时聚合酶链反应（PCR）检测ARID1A质粒的表达。采用MTT法检测依托泊苷（ETO）对AGS和SGC-7901胃腺癌细胞系存活率的影响。用ETO建立DNA双链断裂模型，然后通过彗星试验和免疫荧光共定位观察DNA损伤；采用蛋白质免疫印迹法检测ARID1A对胃腺癌细胞DNA损伤修复途径中相关蛋白表达的影响；采用划痕试验和集落形成实验观察ARID1A对胃腺癌细胞迁移和增殖的影响。采用流式细胞术检测ARID1A对胃腺癌细胞凋亡的影响。
转染ARID1A质粒后，mRNA和蛋白表达增加。MTT法证实ETO以剂量依赖方式抑制细胞存活。用ETO建立DNA双链断裂模型后，过表达ARID1A组中磷酸化共济失调毛细血管扩张突变蛋白（p-ATM）的表达水平升高。同时，过表达ARID1A组的尾矩缩短，γ-H2AX与ARID1A在细胞核的DNA损伤位点共定位。过表达ARID1A组的伤口愈合能力较弱，克隆形成数量减少，凋亡率增加。
ARID1A可能通过p-ATM途径修复ETO引起的DNA双链断裂；ARID1A可抑制胃腺癌细胞的迁移和增殖并促进凋亡。我们的研究结果表明， 可作为GC患者的治疗靶点和生物标志物。
