Aquaporin expression in breast cancer and their involvement in bleb formation, cell motility and invasion in endocrine resistant variant cells.

Abstract

Estrogen receptor (ER)‑silenced breast cancer cell lines exhibit endocrine resistance and morphological changes from an epithelial to a mesenchymal phenotype. These cells also display increased motility and invasive properties that are further accentuated by exposure to an alkaline pH, exhibiting dynamic plasma membrane blebbing and cytoplasmic streaming. These latter morphological changes are hypothesized to involve substantial water movement across the plasma membrane, contributing to bleb formation; this may involve aquaporin channel proteins(AQPs). AQP1, 3, 4 and5 expression/localization was examined via reverse transcription‑quantitative PCR, western blotting and confocal microscopy in endocrine‑sensitive(YS1.2) and ‑resistant (pII and MDA‑MB‑231) breast cancer cells, as well as normal breast epithelial cells(MCF10A). The effects of osmotic changes on bleb formation were examined via live cell imaging. AQP3 protein expression was knocked down by small interfering RNA(siRNA) transfection, and the effect of its reduced expression on bleb formation, cell motility and invasion were determined via immunofluorescence, scratch and Cultrex assays, respectively. Expression of the four AQPs varied across the different cell lines, and exhibited nuclear, cytoplasmic and membranous localization. Osmotic changes affected the formation of blebs. In pIIcells exposed to alkaline pH, AQP3 was observed to be redistributed from the nucleus into the newly formed blebs. siRNA‑mediated knockdown of AQP3 in pIIcells significantly reduced cellular blebbing induced by alkaline pH, as well as motility and invasion. These data suggested that AQP3, and potentially other aquaporins, may participate in the processes leading to blebbing of endocrine‑resistant cells which is proposed to be a mechanism that drives tumor metastasis.