AIM2 Promotes Gastric Cancer Cell Proliferation via the MAPK Signaling Pathway

Abstract

Background: Gastric cancer (GC) is a highly prevalent tumor type. The dysregulated expression of melanoma deficiency factor 2 (AIM2) has been observed in a range of tumor types. Herein, we explore the role of AIM2 in the regulation of GC progression.

Methods: Gastric cancer cells BGC-823 and MGC-803 in logarithmic growth phase were divided into blank group (control), Control group (NC) and SH-AIM2 group, respectively. Control group and SH-AIM2 group were transfected with AIM2 NC and SH-AIM2, respectively. Nude mice were divided into blank group (control) and SH-AIM2 group, and the treatment methods were the same as above. Differential AIM2 expression in GC tissues was assessed via bioinformatics analyses, after which western blotting was used for analyzing the AIM2 levels in tumor and paracancerous tissues from five stomach cancer patients. In addition, qPCR and protein imprinting were used to assess AIM2 expression levels in GC cells, and AIM2 knockdown was conducted in MGC-803 and BGC-823cells, after which colony formation and EdU incorporation assay were utilized to assess cell proliferation. The oncogenic role of AIM2 was then assessed in mice and validated through immunohistochemical analyses.

Results: GC tissues and cell lines exhibited marked AIM2 overexpression. AIM2 knockdown significantly impaired GC cell proliferation and migration, as confirmed through in vitro assays. In vivo experiments showed that both the increment ability and invasion and migration ability of AIM2 knockdown group were significantly lower than that of control and NC the change of AIM2 protein level would affect the change of MAPK pathway related protein level.

Conclusions: AIM2 knockdown markedly suppresses the proliferation, migration, as well as invasion of GC cells via the inhibition of MAPK signaling, thereby slowing tumor progression. Overall, these results suggest that further analyses of AIM2 may offer clinically valuable insights that can aid in the treatment of human GC.

Figure 1

AIM2 is upregulated in gastric cancer. (^∗P < 0.05, Data obtained from three independent are represented as means and standard errors.) (a) AIM2 expression in GC tumor (n = 375) and paracancerous normal control tissues (n = 32) was assessed. (b) AIM2 expression was quantified in pairs of GC tumors and control tissues. (c) Assessment of AIM2 ROC curve sensitivity and specificity. (d) Western blotting was used to assess AIM2 protein levels in matched tumor (T) and normal (N) tissue samples. AIM2 is upregulated in GC cell lines.

Figure 2

Assessment and manipulation of AIM2 expression in GC cell lines (^∗P < 0.05, Data obtained from three independent are represented as means and standard errors.) (a, b) AIM2 levels were assessed in the indicated GC cells via Western blotting and qPCR. (c, d) An shRNA construct was used to knock down AIM2 expression in GC823 and MGC 803 cells, as confirmed via qPCR.

Figure 3

Inhibition of GC cell malignancy by the AIM2 knockdown. (^∗P < 0.05, Data obtained from three independent are represented as means and standard errors.) (a) BGC-823 and MGC-803 GC cells were separated into control, NC, and sh-AIM2 treatment groups and used in EdU incorporation assays to evaluate cell proliferation. (b) The average colony counts were decreased by the AIM2 knockdown in colony formation assay. (c) A wound healing assessment was employed for appraising the influences of AIM2 knockdown on GC cell migration. (d) Transwell assessments were utilized for evaluating the impact of AIM2 knockdown on GC cell invasion.

Figure 4

AIM2 knockdown suppresses excessive MAPK signaling activity. (^∗P < 0.05, ^∗∗P < 0.01, Data obtained from three independent are represented as means and standard errors.) (a-d) ERK, JNK, and P38 phosphorylation levels were assessed via Western blotting in BGC823 and MGC803 cells, with GAPDH as a normalization control. AIM2 facilitates GC tumor progression in vivo.

Figure 5

AIM2 knockdown suppresses in vivo tumor proliferation. (^∗P < 0.05, Data obtained from three independent are represented as means and standard errors.) (a-c) Nude mice were implanted with control or AIM2-knockdown MGC803 cells. Tumor volume and weight were then measured. (d) Phosphorylated p-ERK, p-JNK and p-p38levels in control and AIM2-knockdown tumors were assessed via immunohistochemistry.