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. 1999 Dec;77(6):3085–3095. doi: 10.1016/S0006-3495(99)77139-0

Elasticity of the red cell membrane and its relation to hemolytic disorders: an optical tweezers study.

J Sleep 1, D Wilson 1, R Simmons 1, W Gratzer 1
PMCID: PMC1300579  PMID: 10585930

Abstract

We have used optical tweezers to study the elasticity of red cell membranes; force was applied to a bead attached to a permeabilized spherical ghost and the force-extension relation was obtained from the response of a second bead bound at a diametrically opposite position. Interruption of the skeletal network by dissociation of spectrin tetramers or extraction of the actin junctions engendered a fourfold reduction in stiffness at low applied force, but only a twofold change at larger extensions. Proteolytic scission of the ankyrin, which links the membrane skeleton to the integral membrane protein, band 3, induced a similar effect. The modified, unlike the native membranes, showed plastic relaxation under a prolonged stretch. Flaccid giant liposomes showed no measurable elasticity. Our observations indicate that the elastic character is at least as much a consequence of the attachment of spectrin as of a continuous membrane-bound network, and they offer a rationale for formation of elliptocytes in genetic conditions associated with membrane-skeletal perturbations. The theory of Parker and Winlove for elastic deformation of axisymmetric shells (accompanying paper) allows us to determine the function BH(2) for the spherical saponin-permeabilized ghost membranes (where B is the bending modulus and H the shear modulus); taking the literature value of 2 x 10(-19) Nm for B, H then emerges as 2 x 10(-6) Nm(-1). This is an order of magnitude higher than the value reported for intact cells from micropipette aspiration. Reasons for the difference are discussed.

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Selected References

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  1. Boal D. H. Computer simulation of a model network for the erythrocyte cytoskeleton. Biophys J. 1994 Aug;67(2):521–529. doi: 10.1016/S0006-3495(94)80511-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bronkhorst P. J., Streekstra G. J., Grimbergen J., Nijhof E. J., Sixma J. J., Brakenhoff G. J. A new method to study shape recovery of red blood cells using multiple optical trapping. Biophys J. 1995 Nov;69(5):1666–1673. doi: 10.1016/S0006-3495(95)80084-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chabanel A., Flamm M., Sung K. L., Lee M. M., Schachter D., Chien S. Influence of cholesterol content on red cell membrane viscoelasticity and fluidity. Biophys J. 1983 Nov;44(2):171–176. doi: 10.1016/S0006-3495(83)84288-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chabanel A., Sung K. L., Rapiejko J., Prchal J. T., Palek J., Liu S. C., Chien S. Viscoelastic properties of red cell membrane in hereditary elliptocytosis. Blood. 1989 Feb;73(2):592–595. [PubMed] [Google Scholar]
  5. Chasis J. A., Mohandas N. Erythrocyte membrane deformability and stability: two distinct membrane properties that are independently regulated by skeletal protein associations. J Cell Biol. 1986 Aug;103(2):343–350. doi: 10.1083/jcb.103.2.343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Discher D. E., Mohandas N. Kinematics of red cell aspiration by fluorescence-imaged microdeformation. Biophys J. 1996 Oct;71(4):1680–1694. doi: 10.1016/S0006-3495(96)79424-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Elias P. M., Goerke J., Friend D. S., Brown B. E. Freeze-fracture identification of sterol-digitonin complexes in cell and liposome membranes. J Cell Biol. 1978 Aug;78(2):577–596. doi: 10.1083/jcb.78.2.577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Evans E. A. Bending elastic modulus of red blood cell membrane derived from buckling instability in micropipet aspiration tests. Biophys J. 1983 Jul;43(1):27–30. doi: 10.1016/S0006-3495(83)84319-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Evans E. A. New membrane concept applied to the analysis of fluid shear- and micropipette-deformed red blood cells. Biophys J. 1973 Sep;13(9):941–954. doi: 10.1016/S0006-3495(73)86036-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Evans E, Rawicz W. Entropy-driven tension and bending elasticity in condensed-fluid membranes. Phys Rev Lett. 1990 Apr 23;64(17):2094–2097. doi: 10.1103/PhysRevLett.64.2094. [DOI] [PubMed] [Google Scholar]
  11. Fairbanks G., Steck T. L., Wallach D. F. Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry. 1971 Jun 22;10(13):2606–2617. doi: 10.1021/bi00789a030. [DOI] [PubMed] [Google Scholar]
  12. Fischer T. M., Haest C. W., Stöhr M., Kamp D., Deuticke B. Selective alteration of erythrocyte deformabiliby by SH-reagents: evidence for an involvement of spectrin in membrane shear elasticity. Biochim Biophys Acta. 1978 Jul 4;510(2):270–282. doi: 10.1016/0005-2736(78)90027-5. [DOI] [PubMed] [Google Scholar]
  13. Gordon S., Ralston G. B. Solubilization and denaturation of monomeric actin from erythrocyte membranes by p-mercuribenzenesulfonate. Biochim Biophys Acta. 1990 Jun 11;1025(1):43–48. doi: 10.1016/0005-2736(90)90188-t. [DOI] [PubMed] [Google Scholar]
  14. Hall T. G., Bennett V. Regulatory domains of erythrocyte ankyrin. J Biol Chem. 1987 Aug 5;262(22):10537–10545. [PubMed] [Google Scholar]
  15. Hansen J. C., Skalak R., Chien S., Hoger A. An elastic network model based on the structure of the red blood cell membrane skeleton. Biophys J. 1996 Jan;70(1):146–166. doi: 10.1016/S0006-3495(96)79556-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hansen J. C., Skalak R., Chien S., Hoger A. Influence of network topology on the elasticity of the red blood cell membrane skeleton. Biophys J. 1997 May;72(5):2369–2381. doi: 10.1016/S0006-3495(97)78882-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hansen J., Skalak R., Chien S., Hoger A. Spectrin properties and the elasticity of the red blood cell membrane skeleton. Biorheology. 1997 Jul-Oct;34(4-5):327–348. doi: 10.1016/s0006-355x(98)00008-0. [DOI] [PubMed] [Google Scholar]
  18. Hemming N. J., Anstee D. J., Staricoff M. A., Tanner M. J., Mohandas N. Identification of the membrane attachment sites for protein 4.1 in the human erythrocyte. J Biol Chem. 1995 Mar 10;270(10):5360–5366. doi: 10.1074/jbc.270.10.5360. [DOI] [PubMed] [Google Scholar]
  19. Hénon S., Lenormand G., Richert A., Gallet F. A new determination of the shear modulus of the human erythrocyte membrane using optical tweezers. Biophys J. 1999 Feb;76(2):1145–1151. doi: 10.1016/S0006-3495(99)77279-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Jay A. W. Geometry of the human erythrocyte. I. Effect of albumin on cell geometry. Biophys J. 1975 Mar;15(3):205–222. doi: 10.1016/S0006-3495(75)85812-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Jinbu Y., Sato S., Nakao T., Nakao M., Tsukita S., Tsukita S., Ishikawa H. The role of ankyrin in shape and deformability change of human erythrocyte ghosts. Biochim Biophys Acta. 1984 Jun 27;773(2):237–245. doi: 10.1016/0005-2736(84)90087-7. [DOI] [PubMed] [Google Scholar]
  22. Kozlov M. M., Markin V. S. Model of red blood cell membrane skeleton: electrical and mechanical properties. J Theor Biol. 1987 Dec 21;129(4):439–452. doi: 10.1016/s0022-5193(87)80023-1. [DOI] [PubMed] [Google Scholar]
  23. Kuypers F. A., Lewis R. A., Hua M., Schott M. A., Discher D., Ernst J. D., Lubin B. H. Detection of altered membrane phospholipid asymmetry in subpopulations of human red blood cells using fluorescently labeled annexin V. Blood. 1996 Feb 1;87(3):1179–1187. [PubMed] [Google Scholar]
  24. Käs J., Sackmann E. Shape transitions and shape stability of giant phospholipid vesicles in pure water induced by area-to-volume changes. Biophys J. 1991 Oct;60(4):825–844. doi: 10.1016/S0006-3495(91)82117-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  26. Liu S. C., Derick L. H., Agre P., Palek J. Alteration of the erythrocyte membrane skeletal ultrastructure in hereditary spherocytosis, hereditary elliptocytosis, and pyropoikilocytosis. Blood. 1990 Jul 1;76(1):198–205. [PubMed] [Google Scholar]
  27. Liu S. C., Palek J. Spectrin tetramer-dimer equilibrium and the stability of erythrocyte membrane skeletons. Nature. 1980 Jun 19;285(5766):586–588. doi: 10.1038/285586a0. [DOI] [PubMed] [Google Scholar]
  28. Maksymiw R., Sui S. F., Gaub H., Sackmann E. Electrostatic coupling of spectrin dimers to phosphatidylserine containing lipid lamellae. Biochemistry. 1987 Jun 2;26(11):2983–2990. doi: 10.1021/bi00385a005. [DOI] [PubMed] [Google Scholar]
  29. McGough A. M., Josephs R. On the structure of erythrocyte spectrin in partially expanded membrane skeletons. Proc Natl Acad Sci U S A. 1990 Jul;87(13):5208–5212. doi: 10.1073/pnas.87.13.5208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Meiselman H. J., Evans E. A., Hochmuth R. M. Membrane mechanical properties of ATP-depleted human erythrocytes. Blood. 1978 Sep;52(3):499–504. [PubMed] [Google Scholar]
  31. Mohandas N., Evans E. Mechanical properties of the red cell membrane in relation to molecular structure and genetic defects. Annu Rev Biophys Biomol Struct. 1994;23:787–818. doi: 10.1146/annurev.bb.23.060194.004035. [DOI] [PubMed] [Google Scholar]
  32. Needham D., Nunn R. S. Elastic deformation and failure of lipid bilayer membranes containing cholesterol. Biophys J. 1990 Oct;58(4):997–1009. doi: 10.1016/S0006-3495(90)82444-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Parker K. H., Winlove C. P. The deformation of spherical vesicles with permeable, constant-area membranes: application to the red blood cell. Biophys J. 1999 Dec;77(6):3096–3107. doi: 10.1016/S0006-3495(99)77140-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Peters L. L., Shivdasani R. A., Liu S. C., Hanspal M., John K. M., Gonzalez J. M., Brugnara C., Gwynn B., Mohandas N., Alper S. L. Anion exchanger 1 (band 3) is required to prevent erythrocyte membrane surface loss but not to form the membrane skeleton. Cell. 1996 Sep 20;86(6):917–927. doi: 10.1016/s0092-8674(00)80167-1. [DOI] [PubMed] [Google Scholar]
  35. Pinder J. C., Pekrun A., Maggs A. M., Brain A. P., Gratzer W. B. Association state of human red blood cell band 3 and its interaction with ankyrin. Blood. 1995 May 15;85(10):2951–2961. [PubMed] [Google Scholar]
  36. Pinder J. C., Weeds A. G., Gratzer W. B. Study of actin filament ends in the human red cell membrane. J Mol Biol. 1986 Oct 5;191(3):461–468. doi: 10.1016/0022-2836(86)90141-5. [DOI] [PubMed] [Google Scholar]
  37. Rangachari K., Beaven G. H., Nash G. B., Clough B., Dluzewski A. R., Myint-Oo, Wilson R. J., Gratzer W. B. A study of red cell membrane properties in relation to malarial invasion. Mol Biochem Parasitol. 1989 Apr;34(1):63–74. doi: 10.1016/0166-6851(89)90020-0. [DOI] [PubMed] [Google Scholar]
  38. Seeman P., Cheng D., Iles G. H. Structure of membrane holes in osmotic and saponin hemolysis. J Cell Biol. 1973 Feb;56(2):519–527. doi: 10.1083/jcb.56.2.519. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Seeman P. Transient holes in the erythrocyte membrane during hypotonic hemolysis and stable holes in the membrane after lysis by saponin and lysolecithin. J Cell Biol. 1967 Jan;32(1):55–70. doi: 10.1083/jcb.32.1.55. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Stokke B. T., Mikkelsen A., Elgsaeter A. The human erythrocyte membrane skeleton may be an ionic gel. I. Membrane mechanochemical properties. Eur Biophys J. 1986;13(4):203–218. doi: 10.1007/BF00260368. [DOI] [PubMed] [Google Scholar]
  41. Stokke B. T., Mikkelsen A., Elgsaeter A. The human erythrocyte membrane skeleton may be an ionic gel. III. Micropipette aspiration of unswollen erythrocytes. J Theor Biol. 1986 Nov 21;123(2):205–211. doi: 10.1016/s0022-5193(86)80154-0. [DOI] [PubMed] [Google Scholar]
  42. Strey H., Peterson M., Sackmann E. Measurement of erythrocyte membrane elasticity by flicker eigenmode decomposition. Biophys J. 1995 Aug;69(2):478–488. doi: 10.1016/S0006-3495(95)79921-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Svoboda K., Schmidt C. F., Branton D., Block S. M. Conformation and elasticity of the isolated red blood cell membrane skeleton. Biophys J. 1992 Sep;63(3):784–793. doi: 10.1016/S0006-3495(92)81644-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Tyler J. M., Reinhardt B. N., Branton D. Associations of erythrocyte membrane proteins. Binding of purified bands 2.1 and 4.1 to spectrin. J Biol Chem. 1980 Jul 25;255(14):7034–7039. [PubMed] [Google Scholar]
  45. Vertessy B. G., Steck T. L. Elasticity of the human red cell membrane skeleton. Effects of temperature and denaturants. Biophys J. 1989 Feb;55(2):255–262. doi: 10.1016/S0006-3495(89)82800-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Waugh R. E., Agre P. Reductions of erythrocyte membrane viscoelastic coefficients reflect spectrin deficiencies in hereditary spherocytosis. J Clin Invest. 1988 Jan;81(1):133–141. doi: 10.1172/JCI113284. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Yi S. J., Liu S. C., Derick L. H., Murray J., Barker J. E., Cho M. R., Palek J., Golan D. E. Red cell membranes of ankyrin-deficient nb/nb mice lack band 3 tetramers but contain normal membrane skeletons. Biochemistry. 1997 Aug 5;36(31):9596–9604. doi: 10.1021/bi9704966. [DOI] [PubMed] [Google Scholar]

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