Abstract
Oxygen on its transport route from lung to tissue mitochondria has to cross several cell membranes. The permeability value of membranes for O2 (PO2), although of fundamental importance, is controversial. Previous studies by mostly indirect methods diverge between 0.6 and 125 cm/s. Here, we use a most direct approach by observing transmembrane O2 fluxes out of 100 nm liposomes at defined transmembrane O2 gradients in a stopped-flow system. Due to the small size of the liposomes intra- as well as extraliposomal diffusion processes do not affect the overall kinetics of the O2 release process. We find, for cholesterol-free liposomes, the unexpectedly low PO2 value of 0.03 cm/s at 35 °C. This PO2 would present a serious obstacle to O2 entering or leaving the erythrocyte. Cholesterol turns out to be a novel major modifier of PO2, able to increase PO2 by an order of magnitude. With a membrane cholesterol of 45 mol% as it occurs in erythrocytes, PO2 rises to 0.2 cm/s at 35 °C. This PO2 is just sufficient to ensure complete O2 loading during passage of erythrocytes through the lung’s capillary bed under the conditions of rest as well as maximal exercise.
Highlights
IntroductionIt has long been thought that the respiratory gases permeate all cell membranes without noticeable resistance
It has long been thought that the respiratory gases permeate all cell membranes without noticeable resistance. This assumption has been rebutted by several studies, at least for carbon dioxide, for which it has been shown that membranes can present a significant diffusion
The only reasonably sensitive PO2 determination based on an O2 flux observed across a cell membrane to our knowledge is that of Holland et al [15], who report an orders of magnitude lower P O2
Summary
It has long been thought that the respiratory gases permeate all cell membranes without noticeable resistance. Value of 0.6–0.8 cm/s for the red cell membrane using the stopped-flow technique. This discrepancy prompted us to readdress the question of membrane permeability to O2, following the strategy of using—instead of cells— the much simpler model of unilamellar lipid vesicles of 50 nm radius loaded with oxyhemoglobin. We measured the efflux of O 2 across the lipid membrane of this artificial but modifiable system This was achieved by mixing the suspension of oxyhemoglobin-loaded liposomes in a rapid reaction stopped-flow apparatus with a 50 mM dithionite solution that within 1 ms consumes dissolved O2 in the extraliposomal space, and practically instantaneously after mixing has established a zero partial pressure of O 2 in this space. We prepared liposomes from the most common lipids found in mammalian cell membranes, namely phosphatidylcholine (PC), phosphatidylethanolamine (PE) at a ratio of 8:2 (moles/mole) and different cholesterol (Chol) concentrations ranging from 0 up to 70 mol%
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