Down-regulated aquaporin 5 inhibits proliferation and migration of human epithelial ovarian cancer 3AO cells

Abstract

Background: Recent studies suggested that aquaporins 5 (AQP5) was associated with many kinds of cancers and regulated many processes of various kinds of cancer cells. Our previous studies also demonstrated that AQP5 was highly expressed in epithelial ovarian cancer and contributed to the progress of ovarian cancer.

Methods: Lentivirus for knocking-down the expression of AQP5 was prepared and verified by qPCR and Western blotting. Cell counting kit-8 (CCK8) assay and transwell assay were performed to investigate the role of AQP5 on proliferation and migration of 3AO cells. The effects of down-regulating AQP5 on tumorigenesis were tested by tumor xenografts experiments.

Results: An effective lentivirus silencing AQP5 expression was obtained and used in this study. Down-regulating AQP5 inhibited proliferation and migration of cultured human epithelial ovarian cancer 3AO Cell. Furthermore, interfering of AQP5 during tumorigenesis could efficiently decrease the tumor growth in athymic mice.

Conclusions: These findings altogether suggest that AQP5 regulated multi processes in ovarian carcinogenesis and may be an attractive therapeutic target.

Figure 1

Knock-down efficiency of the AQP5 shRNA constructs. (A) Representative images of 3AO cells transfected with indicated plasmids; scale bar, 50 μm. Levels of AQP5 mRNA (B) and protein (C) in 3AO cells transfected with indicated shRNA contrasts. (D) Representative images of 3AO cells infected with indicated lentivirus; scale bar, 50 μm. Expression levels of AQP5 of 3AO cells treated with indicated virus determined by qPCR (E) and western blotting (F).

Figure 2

Knocking-down AQP5 decreases 3AO cells proliferation. 3AO cells infected with AQP5 shRNA lentivirus and various controls. The CCK8 assay demonstrated that cells treated with AQP5 shRNA have lower viability compared with the controls. The experiment was repeated for three times. Statistic significant differences were observed between cells infected with AQP5 shRNA virus and NC/mock cells.

Figure 3

Knocking-down AQP5 inhibits 3AO cells migration. (A) Representative images of 3AO cells with indicated treatments; scale bar, 100 μm (4 ×), 50 μm (10 ×) and 50 μm (20 ×). (B) Migration ability of 3AO with different infections was analyzed by transwell assay. The experiment was performed for three times. Statistic significant differences were noted between cells infected with AQP5 shRNA virus and NC/mock cells (t test, p < 0.01).

Figure 4

Down-regulating AQP5 expression appears to inhibit tumor growth in athymic mice. Tumor growth was observed within 30 days since visible tumor observed. Lentivirus solution or PBS buffer were subcutaneously injected when the average diameter of tumors researched approximately 4 to 6 mm. No tumor growth was observed in mice injected with PBS instead of 3AO cells. Means and standard deviations of tumor volume were calculated from multiple observations in three groups of mice. Statistically significant differences were noted between mice received Lenti-AQP5-shRNA virus injection and mice injected with Lenti-NC virus or PBS buffer (t test, p < 0.01). n = 5 per group.