IL-10 Lentivirus Delivery for Induction and Retention of an Anti-inflammatory Microenvironment to Promote Spinal Cord Regeneration

Spinal cord injury causes permanent paralysis and loss of sensory function resulting in a reduced quality of life. The spinal cord cannot regenerate due to the creation of a growth-inhibitory microenvironment caused, in part, by neuroinflammation following injury. Neutrophils and macrophages are the primary instigators of the innate immune response causing cell death and production of inhibitory molecules. Macrophages are also responsible for transitioning to an anti-inflammatory (M2) phenotype and resolving the immune response; however, the immune response does not resolve after spinal cord injury leading to chronic inflammation. Therefore, sustained production of anti-inflammatory cytokines may be necessary to induce and sustain an M2 macrophage phenotype to resolve the immune response and promote regeneration. Sustained production of IL-10 by lentivirus gene delivery was investigated in vitro to overcome an inflammatory environment and induce and sustain an M2 macrophage phenotype. IL-10 lentivirus delivery was able to push established inflammatory (M1) macrophages towards an M2 phenotype and retain an M2 phenotype when challenged with M1 stimuli, while delivery of IL-10 protein was unable to accomplish either of these goals. Lentiviral gene delivery from poly(lactide-co-glycolide)/hydroxyapatite (PLG/HAp) scaffolds was employed in the intraperitoneal (IP) fat pad of mice to determine if long-term gene expression could be achieved. Scaffolds containing HAp retained lentivirus locally and preserved virus activity, which contributed to increased levels of transgene expression and increased duration up to 100 days in vivo compared to PLG scaffolds. Increased macrophage infiltration and transduction was responsible for increased early levels of transduction, but it was increased transduction of non-immune cells that persisted in the scaffold long-term. PLG scaffolds were then employed to induce an M2 macrophage phenotype in the IP fat pad. Delivery of IL-10 lentivirus from the scaffold resulted in increased IL-10 and decreased IFN-γ production from macrophages and decreased immune cell infiltration into the scaffold. IL-10 lentivirus delivery was subsequently employed from multiple-channel nerve bridges. Delivery of control lentivirus from the bridge increased neutrophil infiltration; however, addition of IL-10-encoding lentivirus was able to counteract this infiltration indicating that it will be a necessary component of future combinatorial lentivirus therapies in the spinal cord.