Variation in mitochondrial function in hypoxia-sensitive and hypoxia-tolerant human glioma cells

Abstract

We have shown previously that human glioblastoma multiforme cells vary in their ability to survive under hypoxic conditions. Under oxygen limiting conditions, hypoxia-tolerant cells decrease their oxygen consumption rate whereas hypoxia-sensitive cells continue to consume oxygen at a relatively steady rate until the oxygen supply becomes exhausted. We now show that hypoxia-tolerant and hypoxia-sensitive cells exhibit distinct patterns of mitochondrial function in response to hypoxic challenge. Hypoxia-tolerant cell lines retain stable mitochondrial membrane potential and ATP concentration when incubated under oxygen limiting conditions. In addition, hypoxia-tolerant cell lines are consistently more sensitive to a wide spectrum of inhibitors of mitochondrial function than are hypoxia-sensitive cells. In contrast, the hypoxia-sensitive cells are unable to maintain stable mitochondrial membrane potential and ATP levels when incubated at reduced oxygen tension. These results demonstrate significant differences in the mitochondrial function between these two phenotypes and reinforce previous data that suggest a regulatory role for mitochondria in the development of hypoxia tolerance.

Figure 1

Changes in mitochondrial membrane potential (MMP) (JC-1 fluorescence) and mitochondrial mass (Mitofluor Green fluorescence) in hypoxia-sensitive (M010b) vs hypoxia-tolerant (M006x, M059K) cell lines. Cells were grown under normoxic (20% O₂) or hypoxic conditions (0.6% O₂) for 1–3 days for MMP determinations, and 1–2 days for mitochondrial mass measurements. The per cent change observed in hypoxic cultures, relative to aerobic controls, is shown. Mean (±s.e.) values calculated for three separate determinations are shown. Where not displayed, error bars were too small to plot.

Figure 2

Cellular [ATP] (pmol cell⁻¹) in human glioma cell lines maintained under aerobic or hypoxic (0.6% O₂) conditions for 24 or 72 h. Mean (±s.e.) values calculated for three or more experiments are shown. Where not displayed, error bars were too small to plot.