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. 2022 May 16:77:100044.
doi: 10.1016/j.clinsp.2022.100044. eCollection 2022.

IPO7 promotes pancreatic cancer progression via regulating ERBB pathway

Ming Li  1 , Dongqiang Xu  2 , Yijun Zhan  3 , Shiyun Tan  1
Affiliations

IPO7 promotes pancreatic cancer progression via regulating ERBB pathway

Ming Li et al. Clinics (Sao Paulo). .

Abstract

Background: Importin 7 (IPO7) belongs to the Importin β family and is implicated in the progression of diverse human malignancies. This work is performed to probe the role of IPO7 in pancreatic cancer development and its potential downstream mechanisms.

Methods: IPO7 expression in PC and paracancerous tissues were measured using Immunohistochemistry (IHC) staining and qRT-PCR. Western blotting was utilized to detect the expression level of IPO7 in PC cells and immortalize the pancreatic ductal epithelial cell line. After constructing the IPO7 overexpression and knockdown models, the effect of IPO7 on the proliferation of PC cells was analyzed by the CCK-8 and EdU assay. The migration and invasion of PC cells were examined by wound healing assay and Transwell experiment. The apoptosis rate of PC cells was analyzed by flow cytometry and TUNEL assay. The Gene Set Enrichment Analysis (GSEA) was used to determine the enrichment pathways of IPO7. The effect of IPO7 on the ERBB2 expression was determined using Western blotting. A xenograft mouse model was applied to investigate the carcinogenic effect of IPO7 in vivo.

Results: IPO7 expression was remarkably elevated in the cancer tissues of PC patients. IPO7 overexpression remarkably enhanced PC cell proliferation, migration and invasion and suppressed apoptosis, while knockdown of IPO7 exerted the opposite effect. Mechanistically, IPO7 facilitated the malignant phenotype of PC cells by up-regulating ERBB2 expression. In addition, knockdown of IPO7 inhibited tumor growth and lung metastasis in vivo.

Conclusion: IPO7 can act as an oncogenic factor and accelerate PC progression by modulating the ERBB pathway.

Keywords: ERBB pathway; IPO7; Pancreatic cancer.

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Conflict of interest statement

Conflicts of interest The authors declare no conflicts of interest.

Figures

Fig 1
Fig. 1
IPO7 expression in PC tissues and cell lines. (A‒B) IPO7 expression was up-regulated in PC tissues and the expression of IPO7 was correlated with the prognosis of PC patients by searching the Gene Expression Analysis Interactive Analysis (GEIPA) database. (C‒D). Immunohistochemistry showed that IPO7 was highly expressed in PC tissues. (E) qRT-PCR showed that IPO7 mRNA expression was up-regulated in PC tissues. F-G. qRT-PCR and Western blotting showed that IPO7 expression was up-regulated in 4 PC cell lines (PANC-1, HPAC, BxPC-3 and Capan-2) compared with the normal pancreatic ductal epithelial cell line (hTERT-HPNE). All of the experiments were performed in triplicate (**p < 0.01; ***p < 0.001).
Fig 2
Fig. 2
Effect of IPO7 on the proliferation and apoptosis of PC cells. (A‒B) qRT-PCR and Western blotting were adopted to verify the transfection efficiency of pcDNA-IPO7 and IPO7-siRNAs. (C‒E) CCK-8 assay and EdU experiment showed that PC cell proliferation was enhanced in pcDNA-IPO7 group and decreased in the IPO7-siRNA#1 and IPO7-siRNA#2 group. (F‒G) Flow cytometry and TUNEL assay showed that IPO7 overexpression inhibited PC cell apoptosis, and IPO7-siRNA#1 or IPO7-siRNA#2 could promote cell apoptosis. All of the experiments were performed in triplicate (*p < 0.05; **p < 0.01; ***p < 0.001).
Fig 3
Fig. 3
Effect of IPO7 on the migration and invasion of PC cells. (A) Wound healing experiment showed that PC cell motility was promoted in the pcDNA-IPO7 group and decreased in the IPO7-siRNA#1 and IPO7-siRNA#2 group. (B‒C) The Transwell experiment showed that the migration and invasion of PC cells was increased in the pcDNA-IPO7 group and decreased in the IPO7-siRNA#1 and IPO7-siRNA#2 group. All of the experiments were performed in triplicate (**p < 0.01).
Fig 4
Fig. 4
The regulatory effect of IPO7 on the ERBB pathway. (A) GSEA showed that ERBB signal pathway was related with IPO7 in PC. (B‒C) qRT-PCR and Western blotting showed that ERBB2 expression was up-regulated in pcDNA-IPO7 group and down-regulated in IPO7-siRNA#1 and IPO7-siRNA#2 group. All of the experiments were performed in triplicate (*p < 0.05).
Fig 5
Fig. 5
The effect of IPO7 on the development of PC in vivo. PANC-1 cells with IPO7 knockdown or the control PANC-1 cells were inoculated into the mice via subcutaneous injection or tail vein. (A) Tumor volumes were decreased in the IPO7-siRNA group. (B) Tumor weight were decreased in the IPO7-siRNA group. (C) qRT-PCR showed that IPO7 mRNA and ERBB mRNA expression levels were decreased in the tumor tissue of the mice in IPO7 knockdown group. (D) The lung metastasis of the mice was ameliorated in IPO7 knockdown group, and HE staining was utilized to evaluated the histopathological changes of the lung of the mice (*p < 0.05).
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