Altered purinergic receptor calcium signalling associated with hypoxia in MDA-MB-468 breast cancer cells [Poster]

Introduction: Hypoxia is a common feature of the microenvironment of some breast cancers. Hypoxia can induce epithelial-mesenchymal transition (EMT) a process that can convert breast cancer cells into a more invasive phenotype. We have previously shown an association between epidermal growth factor (EGF)-mediated EMT and alterations in purinergic receptor-mediated calcium signalling and levels of puringeric receptor mRNA (e.g. up-regulation of P2X5).

Aims: 1. To compare changes in ATP-induced calcium transients in MDA-MB-468 breast cancer cells in normoxia and hypoxia. 2. To assess mRNA levels of a panel of purinergic receptors in MDA-MB-468 cells in normoxia and hypoxia.

Method: MDA-MB-468 cells were placed in a hypoxic incubator (0.1% O₂) for 24 hours at 37°C. Cytosolic free Ca²⁺ ([Ca²⁺]_CYT) levels were assessed using a Fluorescent Imaging Plate Reader (FLIPR). Changes in mRNA levels were evaluated by quantitative real-time RT-PCR.

Results: Hypoxia-induced EMT was confirmed by changes in EMT markers including vimentin, N-cadherin, snail, and CD24. The nature of the cytosolic calcium response to ATP was altered with hypoxia (e.g. EC₅₀ 0.5 μM innormoxia vs 1.3 μM in hypoxia). Significant changes in mRNA levels of some purinergic receptors in response to hypoxia were observed, however, these were often different from those previously observed with EGF-induced EMT.

Discussion: Hypoxia-induced EMT alters ATP-mediated calcium signalling and mRNA levels of purinergic receptors in MDA-MB-468 breast cancer cells. Further studies are required to assess the significance of altered purinegic receptor-mediated Ca²⁺ signalling associated with the acquisition of EMT.