Nonlinear Local Deformations of Red Blood Cell Membranes: Effects of Toxins and Pharmaceuticals (Part 2)

Modifiers of membranes cause local defects on the cell surface. Measurement of the rigidity at the sites of local defects can provide further information about the structure of defects and mechanical properties of altered membranes.

The purpose of the study: a step-by-step study of the process of a nonlinear deformation of red blood cells membranes under the effect of modifiers of different physico-chemical nature.

Materials and methods. The membrane deformation of a viscoelastic composite erythrocyte construction inside a cell was studied by the atomic force spectroscopy. Nonlinear deformations formed under the effect of hemin, Zn2+ ions, and verapamil were studied.

Results. The process of elastic deformation of the membrane with the indentation of a probe at the sites of local defects caused by modifiers was demonstrated. The probe was inserted during the same step of the piezo scanner z displacement; the probe indentation occured at the different discrete values of h, which are the functions of the membrane structure. At the sites of domains, under the effect of the hemin, tension areas and plasticity areas appeared. A mathematical model of probe indentation at the site of membrane defects is presented.

Conclusion. The molecular mechanisms of various types of nonlinear deformations occurring under the effect of toxins are discussed. The results of the study may be of interest both for fundamental researchers of the blood cell properties and for practical reanimatology and rehabilitology. 

1. Khromova V.S., Myshkin A.E. Coagulation of zinc-modified hemoglobin. Rus. J. Gen. Chem. 2002; 72 (10): 1645–1649. DOI: 10.1023/A:1023356221708. [In Russ., In Engl.]

2. Gudkova O.Ye., Bushueva A.V., Kozlov A.P., Chernysh A.M. Nanostructure and local rigidity of red blood cells (RBC) under influence of membrane modificators and ionizing radiation. Conf. Proc. CLINAM. 2013; 6: 172-173.

3. Moroz V.V., Chernysh A.M., Kozlova E.K., Borshegovskaya P.Y., Bliznjuk U.A., Rysaeva R.M., Gudkova O.Y. Comparison of red blood cell membrane microstructure after different physicochemical influences: atomic force microscope research. J. Crit. Care. 2010; 25 (3): 539.e1-539.e12. DOI: 10.1016/j.jcrc.2010.02.007. PMID: 20381299

4. Perepelitsa S.A., Sergunova V.A., Gudkova O.E. The effect of perinatal hypoxia on red blood cell morphology in newborns. Obshchaya Reanimatologiya = General Reanimatology. 2017; 13 (2): 14-23. DOI: 10.15360/1813-9779-2017-2-14-23. [In Russ., In Engl.]

5. Moroz V.V., Novoderzhkina I.S., Afanasyev A.V., Zarzhetsky Y.V., Ryzhkov I.A., Kozlova E.K., Chernysh A.M. Effect of perftoran on membrane nanostructure of discocyte and stomatocyte after acute blood loss. Obshchaya Reanimatologiya = General Reanimatology. 2017; 13 (2): 32-39. DOI: 10.15360/1813-9779-2017-2-32-39. [In Russ., In Engl.]

6. Kozlova E., Chernysh A., Moroz V., Sergunova V., Gudkova O., Kuzovlev A. Nanodefects of membranes cause destruction of packed red blood cells during long-term storage. Exp. Cell. Res. 2015; 337 (2): 192-201. DOI: 10.1016/j.yexcr.2015.07.009. PMID: 26169694

7. Chernysh A.M., Kozlova E.K., Moroz V.V., Sergunova V.A., Gudkova O.E., Kozlov A.P., Manchenko E.F. Nonlinear lokal deformations of erythrocyte membranes: normal erythrocytes (part 1). Obshchaya Reanimatologiya = General Reanimatology. 2017; 13 (5): 58-68. DOI: 10.15360/1813- 9779-2017-5-58-68. [In Russ., In Engl.]

8. Roduit C., van der Goot F.G., De Los Rios P., Yersin A., Steiner P., Dietler G., Catsicas S., Lafont F., Kasas S. Elastic membrane heterogeneity of living cells revealed by stiff nanoscale membrane domains. Biophys. J. 2008; 94 (4): 1521-1532. DOI: 10.1529/biophysj.107.112862. PMID: 17981897

9. Voïtchovsky K., Antoranz Contera S., Kamihira M., Watts A., Ryan J.F. Differential stiffness and lipid mobility in the leaflets of purple membranes. Biophys. J. 2006; 90 (6): 2075-2085. DOI: 10.1529/biophysj.105.072405. PMID: 16387758

10. Buys A.V., Van Rooy M.J., Soma P., Van Papendorp D., Lipinski B., Pretorius E. Changes in red blood cell membrane structure in type 2 diabetes: a scanning electron and atomic force microscopy study. Сardiovasc. Diabetol. 2013; 12: 25. DOI: 10.1186/1475-2840-12-25. PMID: 23356738

11. Kuznetsova T.G., Starodubtseva M.N., Yegorenkov N. I., Chizhik S.A., Zhdanov R.I. Atomic force microscopy probing of cell elasticity. Micron. 2007; 38 (8): 824–833. DOI: 10.1016/j.micron.2007.06.011. PMID: 17709250

12. Li M., Liu L., Xi N., Wang Y., Dong Z., Xiao X., Zhang W. Atomic force microscopy imaging and mechanical properties measurement of red blood cells and aggressive cancer cells. Sci. China Life Sci. 2012; 55 (11): 968- 973. DOI: 10.1007/s11427-012-4399-3. PMID: 23160828

13. Yu M., Wang J., Wang H., Dong S. Calculation of the intracellular elastic modulus based on an atomic force microscope micro-cutting system. Chin. Sci. Bull. 2012; 57 (15): 1868-1872. DOI: 10.1007/s11434-012-5053-y

14. Sirghi L., Ponti J., Broggi F., Rossi F. Probing elasticity and adhesion of live cells by atomic force microscopy indentation. Eur. Biophys. J. 2008; 37 (6): 935-945. DOI: 10.1007/s00249-008-0311-2. PMID: 18365186

15. Lekka M., Fornal M., Pyka-Fos´ciak G., Lebed K., Wizner B., Grodzicki T., Styczen´ J. Erythrocyte stiffness probed using atomic force microscope. Biorheology. 2005; 42 (4): 307-317. PMID: 16227658

16. Sergunova V.A., Kozlova E.K., Myagkova E.A., Chernysh A.M. Chernysh A.M. In vitro measurement of the elastic properties of the native red blood cell membrane. Obshchaya Reanimatologiya = General Reanimatology. 2015; 11 (3): 39-44. DOI: 10.15360/1813-9779-2015-3-39-44. [In Russ., In Engl.]

17. Shi X., Zhang X., Xia T., Fang X. Living cell study at the single-molecule and single-cell levels by atomic force microscopy. Nanomedicine (Lond.). 2012; 7 (10): 1625- 1637. DOI: 10.2217/nnm.12.130. PMID: 23148543

18. Bremmell K.E., Evans A., Prestidge C.A. Deformation and nano-rheology of red blood cells: an AFM investigation. Colloids Surf. B. Biointerfaces. 2006; 50 (1): 43-48. DOI: 10.1016/j.colsurfb.2006.03.002. PMID: 16701986

19. Fisseha D., Katiyar V.K. Analysis of mechanical behavior of red cell membrane in sickle cell disease. Appl. Mathematics. 2012; 2 (2): 40-46. DOI: 10.5923/j.am.20120202.08

20. Sen S., Subramanian S., Discher D.E. Indentation and adhesive probing of a cell membrane with AFM: theoretical model and experiments. Biophys. J. 2005; 89 (5): 3203–3213. DOI: 10.1529/biophysj.105.063826. PMID: 16113121

21. Dupres V., Verbelen C., Dufrêne Y.F. Probing molecular recognition sites on biosurfaces using AFM. Biomaterials. 2007; 28 (15): 2393-2402. DOI: 10.1016/j.biomaterials.2006.11.011. PMID: 17126394

22. Liu S.C., Zhai S., Lawler J., Palek J. Hemin-mediated dissociation of erythrocyte membrane skeletal proteins. J. Biol. Chem. 1985; 260 (22): 12234-12239. PMID: 4044594

23. Vadillo-Rodriguez V., Beveridge T.J., Dutcher J.R. Surface viscoelasticity of individual gram-negative bacterial cells measured using atomic force microscopy. J. Bacteriol. 2008; 190 (12): 4225-4232. DOI: 10.1128/JB.00132- 08. PMID: 18408030

24. Kozlova E.K., Chernysh A.M., Moroz V.V., Kuzovlev A.N. Analysis of nanostructure of red blood cells membranes by space Fourier transform of AFM images. Micron. 2013; 44: 218-227. DOI: 10.1016/j.micron.2012.06.012. PMID: 22854216

25. Kozlova E., Chernysh A., Moroz V., Gudkova O., Sergunova V., Kuzovlev A. Transformation of membrane nanosurface of red blood cells under hemin action. Sci. Rep. 2014; 4: 6033. DOI: 10.1038/srep06033. PMID: 25112597

26. Chernysh A.M., Kozlova E.K., Moroz V.V., Sergunova V.A., Gudkova O.Ye., Fedorova M.S. Reversible zinc-induced injuries to erythrocyte membrane nanostructure. Bull. Exp. Biol. Med. 2012; 154 (1): 84-88. PMID: 23330097. [In Russ.]

27. Kozlova E., Chernysh A., Moroz V., Sergunova V., Gudkova O., Fedorova M., Kuzovlev A. Opposite effects of electroporation of red blood cell membranes under the influence of zinc ions. Acta Bioeng. Biomech. 2012; 14 (1): 3-13. PMID: 22741531

28. Hekele O., Goesselsberger C.G., Gebeshuber I.C. Nanodiagnostics performed on human red blood cells with atomic force microscopy. Mater. Sci. Technol. 2008; 24 (9): 1162-1165. DOI: 10.1179/174328408X341834

29. Hartmann D. A multiscale model for red blood cell mechanics. Biomech. Model. Mechanobiol. 92010; (1): 1-17. DOI: 10.1007/s10237-009-0154- 5. PMID: 19440743

30. Fedosov D.A., Lei H., Caswell B., Suresh S., Karniadakis G.E. Multiscale modeling of red blood cell mechanics and blood flow in malaria. PLoS Comput. Biol. 2011. 7 (12): e1002270. DOI: 10.1371/journal.pcbi.1002270. PMID: 22144878

31. Mirijanian D.T., Voth G.A. Unique elastic properties of the spectrin tetramer as revealed by multiscale coarse-grained modeling. Proc. Natl. Acad. Sci. USA. 2008; 105 (4): 1204-1208. DOI: 10.1073/pnas.0707500105. PMID: 18202182

32. Shaklai N., Avissar N., Rabizadeh E., Shaklai M. Disintegration of red cell membrane cytoskeleton by hemin. Biochem. Int. 1986; 13 (3): 467-477. PMID: 3790141

33. Nakamura M., Bessho S., Wada S. Spring-network-based model of a red blood cell for simulating mesoscopic blood flow. Int. J. Numer. Method Biomed. Eng. 2013; 29 (1): 114-128. DOI: 10.1002/cnm.2501. PMID: 23293072

34. Parshina E.Yu., Sarycheva A.S., Yusipovich A.I., Brazhe N.A., Goodilin E.A., Maksimov G.V. Combined Raman and atomic force microscopy study of hemoglobin distribution inside erythrocytes and nanoparticle localization on the erythrocyte surface. Laser Physics Letters. 2013; 10 (7): 1-6. DOI: 10.1088/1612-2011/10/7/075607

35. Kim Y., Kim K., Park Y.K. Blood cell - an overview of studies in hematology. In: Moschandreou T.E. (ed.). Measurement techniques for red blood cell deformability: recent advances. Rijeka, Croatia; 2012: 167-195. DOI: 10.5772/50698

36. Gov N., Cluitmans J., Sens P., Bosman G.J.C.G.M. Chapter 4. Cytoskeletal control of red blood cell shape: theory and practice of vesicle formation. In: Advances in planar lipid bilayers and liposomes. v.10: 95-119. DOI: 10.1016/S1554-4516(09)10004-2

37. Hosseini S.M., Feng J.J. How malaria parasites reduce the deformability of infected red blood cells. Biophys. J. 2012; 103 (1): 1-10. DOI: 10.1016/j.bpj.2012.05.026. PMID: 22828326