Correlating the Uptake and Dendritic Cell Activation by MDP-loaded Microparticles
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Correlating the Uptake and Dendritic Cell Activation by MDP-loaded Microparticles Stefanie Schmidt, Toralf Roch, Simi Mathew, Nan Ma, Christian Wischke, and Andreas Lendlein Institute of Biomaterial Science and Berlin-Brandenburg Center for Regenerative Therapies, Helmholtz-Zentrum Geesthacht, Kantstraße 55, 14513 Teltow, Germany ABSTRACT Polymer-based, degradable microparticles (MP) are attractive delivery vehicles for vaccines as the polymer properties can be specifically tailored and the carrier can be loaded with adjuvant. For all newly developed carrier systems it is important to analyze cellular uptake efficiency and the specific effects mediated by the encapsulated agent when phagocytosed by the cells, which is barely reported so far. By the encapsulation of N-acetylmuramyl-L-alanyl-Disoglutamine (MDP) labeled with fluoresceinisothiocyanat (FITC) in poly[(rac-lactide)-coglycolide] (PLGA) MP, the MP was fluorescent and used to visualize the phagocytic uptake. Since encapsulated MDP can activate dendritic cells (DC) via the cytosolic nucleotide-binding oligomerization domain receptors (NOD), it can be investigated whether only cells that have phagocytosed the MP are activated or whether bystander effects occur, resulting in activation of cells, which did not take up MDP-FITC loaded MP. Here, it is demonstrated that increasing MP concentrations in the culture medium had no impact on the viability of DC and that the MP uptake efficiency was dose dependent. Interestingly, it could be shown by the CD86 expression, that only DC, which had engulfed MP, were significantly stronger activated than DC, which had not phagocytosed MDP-FITC loaded MP. On the one hand these results indicate that sufficient amounts of MDP were released from the PLGA carriers into the cytosol of the DC. On the other hand, based on the correlation of uptake and activation on the single cell level, minimal MP induced bystander effects may be expected for in vivo applications. INTRODUCTION Even with high hygienic standards, infections for example with Vibrio cholerae, Mycobacterium tuberculosis or with influenza viruses are still challenging in their treatment and have a big impact on health economics. Effective vaccine strategies could help to improve the protection against these pathogens. Carrier systems are evaluated for vaccine delivery combining different advantages, such as protection of encapsulated antigens or adjuvants from microenvironment, which could lead to instability, degradation or unwanted in vivo effects. In this regard, polymeric PLGA MP are of interest as the degradation rate and hydrophobicity can be influenced by the lactide/glycolide ratio. PLGA has been shown to be an immunologically inert carrier as it was not activating different immune cells [1, 2]. To achieve a long-term protective immunological memory by vaccination, it is of high importance that the antigen/adjuvants are delivered to professional antigen-presenting cells like DC, efficiently processed and subsequently presented to T cells to initiate adaptive immune
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