(93h) Induced M2 Macrophage Activation by IL10 Gene Delivery From PLG Scaffolds

The ability of injured tissue to functionally regenerate as opposed to developing into non-functional scar tissue is drastically impacted by the host response following injury. Many tissue engineering approaches involve the implantation of biomaterials and/or delivery of therapeutic factors. Thus, inflammation results from additive responses to the injury, the biomaterial, and the factors that are being delivered. Excessive inflammation can inhibit the regeneration of tissues such as the pancreas or spinal cord, where activated macrophages create an environment that induces cell death and matrix remodeling. Macrophage activation state may influence the extent of tissue regeneration. M1 (classical) activation is characterized by production of pro-inflammatory cytokines and toxic reactive oxygen species whereas M2 activation induces anti-inflammatory responses, debris scavenging, angiogenesis, and tissue repair. We have previously shown that macrophages are highly transfected after gene delivery from a scaffold implanted into the mouse intraperitoneal (IP) fat pad, and thus propose to investigate the utility of localized gene delivery to modulate macrophage phenotype in this model. A ubiquitin C (UbC) promoter-driven luciferase reporter gene produced expression within three days of implantation that persisted for at least nine weeks in the mouse IP fat, which contrasts with previous work in which the cytomegalovirus (CMV) promoter reporter gene had no expression after three weeks. This duration of expression is consistent with the presence of macrophages at an injury site, which arrive after 3 days and can persist for months. The impact of scaffold pore size on transgene expression was also investigated, with a pore size in the range of 63-106 microns improving expression relative to a pore size of 250-425 microns.

We subsequently investigated the ability of IL-10 gene delivery to influence the distribution of M1, M2 and total macrophages. Blank scaffolds with a reporter gene and empty scaffolds were used as control. Immunohistochemistry was performed on days three and seven with staining for M1 (CD80+), M2 (CD163+), and total macrophages (F4/80+). Preliminary results suggest that scaffolds delivering the IL10 plasmid had an increase in staining for M2 macrophages at both three and seven day time points. It was also shown that M2 macrophages increased with time while M1 macrophages decrease with time. Lastly, no apparent increase in M1 macrophage activation by delivery of naked plasmid or lipoplexes was observed, suggesting that DNA delivery does not increase the inflammatory response beyond that induced by the foreign body. These studies provide a step towards controlling the innate immune response to implanted biomaterials by gene delivery of an anti-inflammatory cytokine. The promotion of M2 macrophage phenotype by IL10 ultimately may enhance regeneration following injury.