Chemistry-dependent adsorption of serum proteins onto polyanhydride microparticles differentially influences dendritic cell uptake and activation
Introduction
The design of vaccine adjuvants capable of activating innate immunity is critical for the induction of protective immune responses [1], [2]. A key step in the activation of the innate immune system is the recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs) on the surface of antigen-presenting cells (APCs), including dendritic cells (DCs) [1], [2], [3]. DCs can internalize and process soluble antigens, resulting in interactions with other immune cells, such as naïve T cells [2], [4], [5]. The use of polymer particles to deliver antigen, either encapsulated or bound to the surface, has been shown to enhance antigen presentation compared with the administration of soluble antigen alone [6], [7], [8].
The interaction of antigen-loaded microparticles with DCs may benefit from engineering the microparticle surface by exploiting the material properties and introducing motifs that mimic pathogens [9]. For example, it has been demonstrated that cationic surfaces greatly enhance uptake [10]. On the other hand, the presence of certain ligands which bind to specific cellular receptors promotes internalization [3], [4], [11]. After contact with serum the particles undergo significant changes in their surface properties because of the rapid adsorption of serum proteins [12], [13].
Polyanhydride microparticles have been shown to possess immunomodulatory properties [14], [15] which, when combined with their ability to stabilize and provide sustained release of protein antigens [16], [17], [18], [19], [20], makes them excellent vaccine adjuvants. Our previous work has demonstrated that serum protein adsorption patterns on polyanhydride microparticles are correlated with their surface characteristics (i.e. hydrophobicity), suggesting that the adsorption of serum proteins can be tailored by controlling the particle surface chemistry [13]. Immunoglobulin G (IgG), complement factors, and other proteins (i.e. opsonins) that have been identified on the surface of microparticles likely influence particle uptake by APCs [13], [21], [22]. Indeed, pathogens such as Mycobacterium tuberculosis, Legionella pneumophila, and Mycobacterium leprae coat themselves with serum proteins [23], [24], [25], [26]. Opsonization of the pathogen facilitates host cell phagocytosis by promoting interactions with specific cell surface receptors, including complement, Fcγ, and mannose receptors [23], [24], [25], [26], [27], [28]. Therefore, understanding the biological consequences of serum protein adsorption onto particles and its effect on APC activation may provide vital insights for the rational design of improved biomaterial-based adjuvants.
This study was designed to investigate the differential adsorption of mouse serum proteins onto the surface of polyanhydride microparticles and to understand the effects of protein adsorption on uptake by and activation of DCs. Polyanhydrides based on sebacic acid (SA), 1,6-bis(p-carboxyphenoxy)hexane (CPH), and 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane (CPTEG) (Fig. 1) were evaluated in this work. The profile of serum proteins adsorbed onto the surface of the polyanhydride particles was indeed found to be influenced by the polymer chemistry and subsequently promoted differential effects on DC activation. Moreover, complement receptor 3 (CR3)-mediated pathways were determined to be critical for the internalization of polyanhydride particles by DCs.
Section snippets
Materials
The chemicals needed for monomer synthesis and polymerization, sebacic acid (99%), p-carboxy benzoic acid (99%), and 1-methyl-2-pyrrolidinone anhydrous (99%), were purchased from Aldrich (Milwaukee, WI); 4-p-hydroxybenzoic acid, 1,6-dibromohexane, 1-methyl-2 pyrrolidinone, and triethylene glycol were purchased from Sigma Aldrich (St Louis, MO); 4-p-fluorobenzonitrile was obtained from Apollo Scientific (Stockport, UK); potassium carbonate, dimethyl formamide, toluene, sulfuric acid, acetic
Chemistry-dependent adsorption of immunoglobulin G (IgG) and complement component C3 on polyanhydride particles
It is known that the surface charge of polymeric particles can influence their uptake by phagocytic cells [36]. Measurements of particle ζ-potential using quasi-elastic light scattering have resulted in similar values (−22 ± 5.5 mV) for all particles regardless of chemistry and these values are consistent with previous work [3]. The presence of deprotonated carboxylic groups may account for the negative surface charge of the polyanhydride particles. After incubation with mouse serum the average
Discussion
Biodegradable polymeric particles have been extensively studied as carriers for the delivery of antigens and drugs [1], [14], [15], [42], [43]. The interaction of the surface of these particles with membrane-bound receptors on APCs will initiate particle uptake and influence the magnitude of the resultant immune response [1], [2]. Adsorption of specific serum proteins onto the surface of polymeric particles alters their recognition and uptake by APCs [12], [13], [39]. The data presented herein
Conclusions
In this study the profile of serum proteins adsorbed onto the surface of polyanhydride particles was influenced by polymer chemistry and elicited differential effects on DC activation. We also observed that complement receptor C3-mediated pathways were involved in the internalization of polyanhydride microparticles by DCs regardless of the presence of serum proteins, highlighting the intrinsic pathogen-mimicking characteristics of these particles. The receptor-mediated internalization induced
Acknowledgements
The authors would like to acknowledge financial support by an ONR-MURI Award (NN00014-06-1-1176). The authors would also like to thank Shawn Rigby of the Iowa State Flow Cytometry Facility for his expertise in flow cytometry and Joel Nott of the Iowa State University Protein Facility for his assistance with the 2-D electrophoresis experiments and documentation. The authors are grateful to Dr. Mary Ann McDowell of the University of Notre Dame for generously providing the CR3−/− mice.
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