BK polyomavirus infection promotes growth and aggressiveness in bladder cancer

Abstract

Background: Recent studies have confirmed the integration of the BK polyomavirus (BKPyV) gene into the cellular genome of urothelial carcinomas in transplant recipients, further confirming the correlation between BKPyV and urothelial carcinomas after transplantation. However, the role BKPyV infections play in the biological function of bladder cancer remains unclear.

Methods: We developed a BKPyV-infected bladder cancer cell model and a mice tumor model to discuss the role of BKPyV infections.

Results: Our research proves that BKPyV infections promote the proliferation, invasion and migration of bladder cancer cells, while the activation of β-catenin signaling pathway is one of its mediation mechanisms.

Conclusions: We first described BKPyV infection promotes the proliferation, invasion and migration of bladder cancer. We verified the role of β-catenin signaling pathway and Epithelial-Mesenchymal Transition effect in BKPyV-infected bladder cancer. These results provide meaningful information towards the diagnosis and treatment of clinical bladder cancer.

Keywords: BK polyomavirus; Bladder cancer; Cell aggressiveness; Cell growth.

Fig. 1

BKPyV infection in bladder cancer cells is a non-lytic infection (MOI 2; 48 h post infection). a Immunofluorescence staining was performed (red: LTag, blue, DNA, and images were taken at 400× magnification). b, c BKPyV infection significantly increased growth in T24 and HTB-9 cells, as measured by CCK-8. The effect was significantly reversed by KYA1797K. All graphs represent the mean ± SD obtained from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001; + BKV versus control; + BKV + KYA1797K versus + BKV; Student’s t-test or one-way ANOVA

Fig. 2

BKPyV infection promotes proliferation, invasion and migration of bladder cancer cells. a, b BKPyV infection significantly increased growth in T24 and HTB-9 cells, as measured by colony formation. c–h BKPyV infection significantly promoted migration of T24 and HTB-9 cells, as measured by the Transwell migration assay (c, d) and wound healing assay (e–h) after infected with BKPyV 48 h. i, j Transwell invasion assay showed that BKPyV infection significantly promoted invasiveness of T24 and HTB-9 cells. These effect was significantly reversed by KYA1797K. All images were taken at 100× magnification. All graphs represent the mean ± SD obtained from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001; + BKV versus control; + BKV + KYA1797K versus + BKV; Student’s t-test or one-way ANOVA

Fig. 3

BKPyV infection enhances bladder tumor growth and metastasis. a T24, HTB-9 and their respective BKPyV-infected cells were injected subcutaneously into nude mice with subsequent intraperitoneal injection of KYA1797K (20 mg/kg) (n = 5). Tumor volumes (b, c) of mice were measured every 5 days. BKPyV infection significantly promoted tumor growth in vivo. The effect was significantly reversed by KYA1797K. Tumors were subjected to H&E staining (d) (200×) and TUNEL immunohistochemistry (e, f) (200×). TUNEL staining showed that BKPyV infection significantly induced tumor apoptosis. The effect was significantly reversed by KYA1797K. g Representative images of liver, metastatic nodules (indicated by arrows) are shown. h Livers were subjected to H&E staining (original magnification,40×; smaller image at lower left, 200×). Liver metastasis was found only in the BKPyV-infected group. The effect was significantly reversed by KYA1797K. All graphs represent the mean ± SD obtained from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001; + BKPyV versus control; + BKPyV +KYA1797K versus + BKPyV; Student’s t-test or one-way ANOVA

Fig. 4

BKPyV infection enhances β-catenin signaling pathway activation and EMT effect in bladder cancer cells. a-f Expression levels of β-catenin, cMYC, and EMT-related proteins investigated by western blotting. β-Actin was used as a loading control. The effect was significantly reversed by β-catenin inhibitor (KYA1797K). All graphs represent the mean ± SD obtained from three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001; + BKPyV versus control; + BKPyV +KYA1797K versus + BKPyV; Student’s t-test or one-way ANOVA