Are there significant gradients of PO2 in cells?
Grinberg OY, James PE, Swartz, HM
EPR Center for the Study of Viable Systems, Department of Diagnostic
Radiology, Dartmouth Medical
School, Hanover, NH 03755, USA
INTRODUCTION: It is widely recognized that the intracellular oxygen
tension (pO2) plays an
important role in cellular function. In experimental and theoretical
considerations involving the role
of the pO2 the values that are used usually are those for the pO2 at the
exterior of cells, because
these values can be more readily measured. Such an approach is based on the
assumption that the
intracellular pO2 is relatively uniform because oxygen freely diffuses
across cell membranes and
within cells. For the past several years we have been developing and
applying electron paramagnetic
resonance (EPR) techniques to test directly whether there are intracellular
gradients of pO2 and if so,
where they occur and what factors determine them. We previously reported
significant gradients
between the average pO2 in the intracellular and extracellular compartments
in cell suspensions.
More recently we have developed techniques that enabled us to measure
simultaneously the
concentration of oxygen within a specific compartment, the phagosomes of
activated specific cells,
macrophages, and the extracellular compartment. The precise mechanism of the
intracellular -
extracellular oxygen gradient remains uncertain. The possibilities include
that the diffusion of
oxygen is not as free as assumed (e.g. that the cell membrane can act as a
barrier) and active
transport of O2 out of the cells.
METHOD: A simple theoretical approach was used to evaluate the values
of three key parameters
which might account for the observed intracellular - extracellular oxygen
measurements: 1. the
diffusion coefficient of oxygen in the membrane; 2. the diffusion
coefficient of oxygen within the
cell; 3. the solubility of oxygen within the cell. We also used two
different models for the
relationship between the oxygen consuming compartment (assumed to be
primarily the
mitochondria) and the intracellular compartment in which the measurements
were made (especially
phagosomes): uniform and non-uniform distribution of the mitochondria. Using
a spherical model of
the cell with mitochondria homogeneously distributed within cell, and using
consensus values from
the literature, we were unable to account for the observed differences in
pO2 between the
intracellular-extracellular compartments. We then varied the parameters
one-by-one from the
accepted values, to values which would result in the observed gradient.
RESULTS: We found that with the variation of any one parameter we
could not plausibly account for
the measurements made in the phagosomal and extracellular compartments.
DISCUSSION AND CONCLUSIONS: There are at least three logical
possibilities to account for these
results: 1) this methodology is erroneous and/or produces artifacts in the
system (such as effecting the
cell membrane directly) resulting in invalid results; 2) the observation of
a gradient in oxygen
concentration between these two compartments arises from significant
variations of more than one of the
critical parameters which are used conventionally to calculate potential
gradients in pO2; 3) there is
another factor not considered in the model which accounts for the
observation (e.g. active transport;
significantly higher than expected barriers to oxygen diffusion in the
membrane; clustering of
phagosomes with mitochondria).
IMPLICATIONS FOR BIOMEDICINE: The occurrence of significant gradients
of oxygen within
the cell under physiological control is a logical means to maximize the
value of oxygen to cellular
metabolism while minimizing its potential deleterious effects. It seems
likely that changes in the
efficacy of the control mechanisms would lead to significant pathophysiology
and therefore may
account for some currently poorly understood pathological states.
REFERENCE: Grinberg OY, James PE, Swartz, HM, Are there significant
gradients of pO
2 in cells? Adv. Exp. Med. Biol. 454:415-423 (1998).