title: The mechanism and modulation of complement activation on polymer grafted cells.
abstract: Cell surface engineering using polymers is a promising approach to address unmet needs and adverse immune reactions in the fields of transfusion, transplantation, and cell-based therapies. Furthermore, cell surface modification may minimize or prevent adverse immune reactions to homologous incompatible cells as the interface between the host immune system and the cell surface is modified. In this report, we investigate the immune system reaction, precisely the complement binding and activation on cell surfaces modified with a functional polymer, hyperbranched polyglycerol (HPG). We used red blood cells (RBCs) as a model system to investigate the mechanism of complement activation on cell surfaces modified with various forms of HPG. Using a battery of in vitro assays including: traditional diagnostic hemolytic assays involving sheep and rabbit erythrocytes, ELISAs and flow cytometry, we show that HPG modified RBCs at certain concentrations and molecular weights activate complement via the alternative pathway. We show that by varying the grafting concentration, molecular weight and the number of cell surface reactive groups of HPG, the complement activity on the cell surface can be modulated. HPGs with molecular weights greater than 28kDa and grafting concentrations greater than 1.0mM, as well as a high degree of HPG functionalization with cell surface reactive groups result in the activation of the complement system via the alternative pathway. No complement activation observed when these threshold levels are not exceeded. These insights may have an impact on devising key strategies in developing novel next generation cell-surface engineered therapeutic products for applications in the fields of cell therapy, transfusion and drug delivery.
  
  
  CONCLUSIONS
  Cell-surface engineering using functional polymers is a fast emerging area of research. Importantly modified cells are used in many experimental therapeutics, transplantation and in transfusion. The success of such therapies depend on the ability of modified products to avoid immune detection and subsequent rejection or removal. Polymer grafting has been shown to modulate immune response, however, there is limited knowledge available. Thus in this manuscript, we investigated the interaction of human complement, part of our innate immune system, by polymer modified cells. Our results provide important evidences on the mechanism of complement activation by the modified cells and also found ways to modulate the innate immune response. These results will have implications in development of next generation cell-based therapies.
authors.affiliation: Centre for Blood Research, University of British Columbia, Vancouver, BC V6T 1Z3, Canada; Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BC V6T 1Z3, Canada.
authors.forename: Vincent L
authors.initials: VL
authors.lastname: Leung
authors.affiliation: Centre for Blood Research, University of British Columbia, Vancouver, BC V6T 1Z3, Canada; Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BC V6T 1Z3, Canada; Department of Chemistry, University of British Columbia, Vancouver, BC V6T 1Z3, Canada. Electronic address: jay@pathology.ubc.ca.
authors.forename: Jayachandran N
authors.initials: JN
authors.lastname: Kizhakkedathu
mesh_terms: D000017:ABO Blood-Group System; D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry; D002462:Cell Membrane / Q000378:metabolism; D003167:Complement Activation; D003176:Complement C3 / Q000737:chemistry; D015934:Complement C4a / Q000737:chemistry; D011415:Complement Factor B / Q000737:chemistry; D015938:Complement Membrane Attack Complex / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000166:cytology; D005434:Flow Cytometry; D005990:Glycerol / Q000737:chemistry; D006461:Hemolysis; D006801:Humans; D007113:Immunity, Innate; D008970:Molecular Weight; D011108:Polymers / Q000737:chemistry*; D011817:Rabbits; D012756:Sheep; D013696:Temperature
chemical_list: D000017:ABO Blood-Group System; D001672:Biocompatible Materials; D003176:Complement C3; D015938:Complement Membrane Attack Complex; D011108:Polymers; C047610:SC5b-9 protein complex; C043941:polyglycerol; D015934:Complement C4a; D011415:Complement Factor B; D005990:Glycerol
keywords: Biocompatibility; Cell surface engineering; Complement system; Immunomodulation; Innate immune system; Red blood cells