UBC Theses and Dissertations
Immunocamouflage : the biophysical and biological basis of immunoprotection by grafted methoxypoly(ethylene glycol) Le, Yevgeniya
Development of novel approaches for the direct immunomodulation of allogeneic donor cells would have significant utility in tissue transplantation. Immunocamouflage of cell surfaces by covalently grafted methoxypoly(ethylene glycol) (mPEG; PEGylation) has emerged as a promising approach. While previous studies demonstrated the in vitro and in vivo efficacy of immunocamouflaged allogeneic cells and viruses, the biophysical mechanisms of immunoprotection have not been well-defined due to the labile nature of biological samples. To overcome this limitation, polystyrene latex particles (1.2 and 8.0 µm) were used to elucidate the biophysical mechanisms of immunocamouflage via the effects of the chemical and physical properties of the polymer, as well as the consequences of target size. These findings were correlated with the biological studies utilizing human red blood cells and lymphocytes. It was demonstrated that the two biophysical mechanisms were responsible for the immunocamouflage of PEGylated surfaces: 1) hydrodynamic shielding of surface charge; and 2) steric exclusion of macromolecules from the surface. Surface charge camouflage of latex particles and erythrocytes was best achieved with long polymer chains, regardless of the target size. However, inhibition of surface-macromolecule interactions indicated a target size dependence. The biophysical latex model demonstrated that short chain polymers (2 kDa) were more effective at preventing protein adsorption to small beads (1.2 µm), while long chain polymers (20 kDa) exhibited increased efficacy on large particles (8.0 µm). Consistent with the biophysical model, immunocamouflage of lymphocytes (~10 µm) was best achieved using long chain polymers as measured by: 1) inhibition of antigen-antibody binding (CD3, CD4 and CD28); and 2) allorecognition in a 2-way mixed lymphocyte reaction. The biological model also demonstrated that cell surface topography and antigen localization were critical in selecting the optimal polymer size. Importantly, PEGylation did not result in any cellular toxicity at immunoprotective levels that rendered modified surfaces more biocompatible. Thus, these studies delineated the biophysical mechanisms of immunocamouflage defined by the chemical and physical parameters of the polymer and influenced by target size and surface complexity. Cell or tissue specific optimization of these factors will be critical for the efficient immunocamouflage of allogeneic cells in transfusion and transplantation medicine.
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