Mechanisms of amphipath-induced stomatocytosis in human erythrocytes.
We studied stomatocytosis induced in human red blood cells (RBC) by vinblastine and chlorpromazine, monitoring the movements of spin-labeled phosphatidylcholine (PC*) and sphingomyelin (SM*) by electron spin resonance (ESR) spectroscopy. This technique allows determination of the fraction of labeled lipids, respectively, on the external leaflet, on the cytosol face, or trapped in endocytic vacuoles. Both vinblastine and chlorpromazine produce a time- and concentration-dependent stomatocytic shape change, which is paralleled by a shift of approximately 10% to 33% of outer leaflet SM* and PC* inward. Of this amount, 8% to 12% was trapped in endocytic vacuoles and 8% to 19% had flipped to the inner leaflet. Vanadate, while inhibiting the stomatocytosis, did not block the flip of either SM* or PC* to the inner leaflet. To explain the inhibiting effect of vanadate, as well as the adenosine triphosphate (ATP) requirement for drug-induced stomatocytosis, we propose the following model: (1) addition of amphipath partially scrambles the bilayer; and (2) the flop of phosphatidylserine (PS) and phosphatidylethanolamine (PE) to the outer leaflet provides substrate for the aminophospholipid translocase (APLT), which flips back PS and PE inward faster than PC or SM can diffuse outward--thereby producing inner layer expansion or stomatocytosis. This role of APLT accounts for the vanadate inhibition of amphipath stomatocytosis. However, the vanadate effect can be overcome by increasing the amphipath concentration, which at such levels probably passively expands the inner leaflet.
Schrier
S L
SL
Division of Hematology, Stanford University School of Medicine, CA.
Zachowski
A
A
Devaux
P F
PF
D002352:Carrier Proteins / Q000502:physiology*; D002746:Chlorpromazine / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000187:drug effects* / Q000648:ultrastructure; D004913:Erythrocytes, Abnormal / Q000737:chemistry / Q000648:ultrastructure; D006801:Humans; D066298:In Vitro Techniques; D008051:Lipid Bilayers / Q000737:chemistry; D008563:Membrane Lipids / Q000097:blood*; D008565:Membrane Proteins / Q000502:physiology*; D010713:Phosphatidylcholines / Q000737:chemistry; D049294:Phospholipid Transfer Proteins; D013109:Sphingomyelins / Q000737:chemistry; D013113:Spin Labels; D014638:Vanadates / Q000494:pharmacology; D014747:Vinblastine / Q000494:pharmacology
D002352:Carrier Proteins; D008051:Lipid Bilayers; D008563:Membrane Lipids; D008565:Membrane Proteins; D010713:Phosphatidylcholines; D049294:Phospholipid Transfer Proteins; D013109:Sphingomyelins; D013113:Spin Labels; D014638:Vanadates; D014747:Vinblastine; D002746:Chlorpromazine