Mitochondrial Bioenergetics of Metastatic Breast Cancer Cells in Response to Dynamic Changes in Oxygen Tension: Effects of HIF-1α

Anne R Diers,Corinne E Griguer,Praveen K Vayalil,Claudia R Oliva,Douglas R Hurst,Danny R Welch,Aimee Landar,Victor Darley-Usmar,Janine Santos

doi:10.1371/journal.pone.0068348

Abstract

Solid tumors are characterized by regions of low oxygen tension (OT), which play a central role in tumor progression and resistance to therapy. Low OT affects mitochondrial function and for the cells to survive, mitochondria must functionally adapt to low OT to maintain the cellular bioenergetics. In this study, a novel experimental approach was developed to examine the real-time bioenergetic changes in breast cancer cells (BCCs) during adaptation to OT (from 20% to <1% oxygen) using sensitive extracellular flux technology. Oxygen was gradually removed from the medium, and the bioenergetics of metastatic BCCs (MDA-MB-231 and MCF10CA clones) was compared with non-tumorigenic (MCF10A) cells. BCCs, but not MCF10A, rapidly responded to low OT by stabilizing HIF-1α and increasing HIF-1α responsive gene expression and glucose uptake. BCCs also increased extracellular acidification rate (ECAR), which was markedly lower in MCF10A. Interestingly, BCCs exhibited a biphasic response in basal respiration as the OT was reduced from 20% to <1%. The initial stimulation of oxygen consumption is found to be due to increased mitochondrial respiration. This effect was HIF-1α-dependent, as silencing HIF-1α abolished the biphasic response. During hypoxia and reoxygenation, BCCs also maintained oxygen consumption rates at specific OT; however, HIF-1α silenced BCC were less responsive to changes in OT. Our results suggest that HIF-1α provides a high degree of bioenergetic flexibility under different OT which may confer an adaptive advantage for BCC survival in the tumor microenvironment and during invasion and metastasis. This study thus provides direct evidence for the cross-talk between HIF-1α and mitochondria during adaptation to low OT by BCCs and may be useful in identifying novel therapeutic agents that target the bioenergetics of BCCs in response to low OT.

Highlights

Partial pressure of oxygen is a micro-environmental factor that plays key roles in tumor progression and responses to treatment
Exposure of tumor cells to either chronic or acute hypoxia initiates signaling events which promote spontaneous metastasis [6,7]; some studies suggest that intermittent cycles of acute hypoxia are more significant in driving the metastatic process than chronic hypoxia [8,9]
Since HIF-1a is a hallmark of the cellular response to low oxygen tension (OT), stabilization and nuclear translocation of HIF-1a was determined from cells exposed to low OT in the XF24 instrument

Summary

Introduction

Partial pressure of oxygen (pO2) is a micro-environmental factor that plays key roles in tumor progression and responses to treatment. Two types of tumor cell hypoxia (defined as OT ,1%) have been observed [3]. Chronic, prolonged hypoxia, termed ‘diffusion-limited hypoxia,’ develops in regions distal from blood vessels. These areas of the tumor are characteristically adjacent to necrotic regions and occur as a consequence of permanent limitations in oxygen diffusion [4]. Intermittent hypoxia results from acute and fluctuating changes in OT which may last for less than a minute to several hours in tumors. Intermittent hypoxia can develop as a result of temporary obstruction or cessation of tumor blood flow [5]. Exposure of tumor cells to either chronic or acute hypoxia initiates signaling events which promote spontaneous metastasis [6,7]; some studies suggest that intermittent cycles of acute hypoxia are more significant in driving the metastatic process than chronic hypoxia [8,9]

Methods

Results

Conclusion