MicroRNA-575 regulates the biological characteristics of gastric cancer cells by targeting absent in melanoma 2
Objective To investigate the role of absent in melanoma 2(AIM2),an important mol-ecule in the pyroptosis signaling pathway,in gastric cancer(GC),and to further analyze the effect of mi-croRNA(miRNA,miR)/AIM2 axis on the biological characteristics of GC cells by searching for micrornas that regulate AIM2.Methods The limma package of R studio software was used to analyze the differenti-ally expressed genes in GSE113255 and GSE118897 datasets,and the common genes related to pyroptosis were screened by Venn diagram to determine the target genes.Cancer tissues and adjacent tissues of 56 GC patients who were treated in the Gastroenterology Department of Yuncheng Central Hospital Affiliated to Shanxi Medical University from January 2022 to January 2023 were collected to detect the expression of tar-get genes.AIM2 was overexpressed in human GC cell line MKN45,and cell death and proliferation were detected by propidium iodide(PI)staining,cell counting kit-8(CCK-8)assay and Matrigel invasion as-say.Targetscan website was used to predict the mirnas regulating AIM2,and luciferase reporter gene was used to detect the binding between AIM2 promoter region and AIM2.MKN45 cells were transfected with miR-575 mimic(miR-575 mimics)/negative control(NC mimics)or miR-575 inhibitor(miR-575 inhibitor)/negative control(NC inhibitor).PI staining,CCK-8 assay and Matrigel invasion assay were used to detect cell death and proliferation.MKN45 cells were treated with Ad-GFP/Ad-AIM2 and NC mimics/miR-575 mimics.Lactate dehydrogenase release assay,CCK-8 assay and Matrigel invasion assay were used to detect cell death and proliferation.The differences between groups were analyzed by t-test or analysis of variance(ANOVA).Results PI staining,CCK-8 assay and Matrigel invasion assay showed that the number of cell death in Ad-AIM2 group was significantly greater than that in Ad-GFP group[(43.52±3.08)％vs.(17.72±2.21)％,t=6.114,P＜0.01].Cell proliferation and invasion ability were significantly lower than Ad-GFP group(48 h:1.17±0.18 vs.1.45±0.22,t=5.027,P＜0.05;72 h:1.58±0.36 vs.2.31±0.42,t=4.822,P＜0.05).Using Targetscan website prediction and verified by luciferase reporter gene experiment,it was found that miR-575 and AIM2 had complementary nucleotide sequences.After co-transfection of miR-575 mimics and wild type AIM2,the luciferase activity of miR-575 mimics group was significantly lower than that of NC mimics group(0.52±0.06 vs.0.98±0.14,t=5.323,P＜0.05).However,after co-transfection of miR-575 mimics and mutant AIM2,the luciferase activity of cells in miR-575 mimics group was not significantly different from that in NC mimics group(0.93±0.08 vs.1.00±0.12,t=0.972,P＞0.05).Western blotting results showed that The expression levels of apopto-sis-associated speck-like protein(ASC),cleaved cysteinyl aspartate-specific protease(Caspase)-1/Caspase-1 and high mobility group box 1 protein(HMGB1)in Ad-AIM2+miR-575 mimics group were sig-nifiicantly higher than those in Ad-GFP+group miR-575 mimics group(ASC:1.62±0.31 vs.0.73±0.19,t=4.302,P＜0.05;cleaved Caspase-1/Caspase-1:1.44±0.21 vs.0.78±0.16,t=3.198,P＜0.05;HMGB1:1.32±0.14 vs.0.81±0.11,t=5.029,P＜0.05).Conclusion The expression of AIM2 is decreased in GC.miR-575 inhibits pyroptosis and promotes the proliferation and invasion of MKN45 cells by targeting AIM2 expression.
Gastric cancerMicroRNAAbsent in melanoma 2ProliferationPyroptosis
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