Role of Aquaporins in cell proliferation: Functional inhibition of AQP3 with a gold-based compound

Abstract

[Objective] Numerous studies indicated an abnormal AQPs expression in tumor of different origins and a role for these proteins in angiogenesis, cell migration and proliferation had been shown. Recently we verified that the gold (III) complex Auphen significantly inhibits the cell proliferation of AQP3 expressing cells. Then to better understand the role AQPs may play in the cell proliferation process we explore the effects that stable overexpression of these proteins (o-AQPs) produce over the proliferation and cell cycle of wt-PC12 cells as a cellular model.

[Methods] Cell cycle by flow cytometry with propidium iodide and cell proliferation through cell counting and BrdU staining were used. Using Nocodazole we evaluated the cell response to arrest its cell cycle and the resistance to apoptosis by Annexin V staining; and proteomic and transcriptomic techniques were performed to highlight key molecules implicated in cell proliferation which expression may be altered by overexpression of AQPs. Finally, in cells with large expression of AQP3 we explore the effect of Auphen over cyclins expression and cell cycle progression.

[Results] Cells with o-AQPs showed higher cell proliferation rate and larger percentage of cells in phases S and G2/M. After 24h in the presence of Nocodazole, o-AQPs cells exhibited less modification of the cell cycle pattern and lower Annexin V specific staining consistent with a higher resistance to apoptosis. Additionally, in AQP3-expressing cells treated with Auphen, strong arrest of the cell cycle in the S-G2/M phases, in concordance with the analysis of cyclins (A, B1, D1, E) levels was observed. The RT- qPCR analysis comparing o-AQPs cells to wt cells validated interesting changes in the expression of molecules related with cell proliferation, tumor and cell cycle progression, such as, Zeb2, Jun, JunB, NF-kβ, Cxcl9, Cxcl10, TNF, and TNF receptors.

[Conclusions] The significant role of AQPs in the cell proliferation process seems to be connected to increments in the cell cycle turnover. Our results support the view that larger expression of AQPs confers to the cell a more tumor-like phenotype that contributes to explain the presence of these proteins in much different type of tumors. A potential therapeutic effect of Auphen in tumors where cell proliferation can be associated with AQP3 seems promising, but more studies are necessary to clarify this issue.