Biodegradable chitosan microparticles induce delayed STAT-1 activation and lead to distinct cytokine responses in differentially polarized human macrophages in vitro

[Display omitted] Current data suggest that chitosan activates wound macrophages to release endogenous factors that guide mesenchymal stem cell (MSC) to bone fractures. We tested the hypothesis that chitosan, a polymer containing glucosamine and N-acetyl glucosamine, stimulates macrophages in different polarization states to release functional MSC chemokines and mainly anabolic factors. Low-serum conditioned medium was collected from M0, M1 and M2a U937 macrophages previously differentiated with phorbol myristate acetate (PMA) and exposed or not for 24h to chitosan microparticles (80% degree of deacetylation, DDA, 130kDa). Chitosan particles were highly phagocytosed. Chitosan enhanced anabolic factor release from M0 and M2a macrophages (MCP-1, IP-10, MIP-1beta, IL-1ra, IL-10, PDGF), and IL-1beta release, with 25- to 400-fold excess IL-1ra over IL-1beta. In M1 macrophages, chitosan enhanced IL-1beta without enhancing or suppressing inflammatory factor release (IL-6, IP-10, IL-8). M0 and M2a macrophages, with or without chitosan stimulation, produced conditioned medium that promoted 2-fold more MSC chemotaxis than low-serum control medium, while M1-conditioned medium failed to induce MSC chemotaxis. Acetylated chitosan induced U937 macrophages to release IL-1ra without STAT-6 activation, and also induced a delayed STAT-1 activation/IP-10 release response that was not observed using non-biodegradable chitosan (98% DDA, 130kDa). In primary human macrophages, acetylated chitosan enhanced IL-1ra release without inducing IL-1beta, and required PMA priming to elicit STAT-1 activation and IP-10 release. We conclude that biodegradable chitosan particles enhance M0 and M2a macrophage anabolic responses independent of the IL4/STAT-6 axis, by inducing excess IL-1ra over IL-1beta and more chemokine release, without altering their inherent capacity to attract MSCs.