Oxidation-induced changes in microrheologic properties of the red blood cell membrane

R. P. Hebbel, A. Leung, N. Mohandas

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160 Scopus citations


It has been hypothesized that some of the irreversible microrheologic abnormalities of sickle red blood cell (RBC) membranes could result from autoxidative perturbation. To model this possibility, we used micromechanical manipulation to examine the static extensional rigidity and inelastic or plastic behavior of normal RBCs exposed to phenazine methosulfate (PMS), an agent that generates superoxide from within the cell. In response to this stress, RBC membranes became stiff as evidenced by increasing extensional rigidity. At 50 μmol/L PMS they were as stiff as the membranes of most dense, dehydrated sickle RBCs; and at 25 μmol/L PMS the membranes were similar to somewhat less dense sickle RBCs. When examined for inelastic behavior, RBCs exposed to PMS even at 10 μmol/L showed hysteresis in loading and unloading phases of the curve relating aspiration length to suction pressure, and they developed membrane bumps that persisted after RBC release from the pipette. Examination of single cells in both isotonic and hypotonic buffers showed that the effect of PMS on RBC microrheology is not mediated by cellular dehydration. Independent confirmation of the membrane stiffening effect of PMS was obtained by ektacytometric analysis of resealed RBC ghosts, with sickle-like increases in membrane rigidity observed between 50 and 100 μmol/L PMS. The rigidity of these ghosts was partially ameliorated by exposure to a thiol reductant. In terms of biochemical abnormalities, treated RBCs became significantly different from control RBCs at 25 μmol/L PMS, at which point they just began to enter the sickle range for amounts of membrane thiol oxidation and membrane-associated heme. The sickle average was achieved at 50 μmol/L PMS (for thiol oxidation) to 100 μmol/L PMS (for membrane heme). Thus, micromolar concentrations of PMS induce abnormalities of membrane microrheology that closely mimic those of unmanipulated sickle RBCs while reproducing similar degrees of oxidative biochemical change. We conclude that membrane protein oxidation could explain existence of an irreversible component to the abnormal rheology of the sickle membrane.

Original languageEnglish (US)
Pages (from-to)1015-1020
Number of pages6
Issue number5
StatePublished - Sep 1 1990
Externally publishedYes


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