Particle-Assisted Control over Macrophage-Neutrophil interactions (Pac-Man) - Project Summary
Volumetric muscle loss (VML) is a debilitating injury caused by trauma or disease to skeletal muscle that
leads to incapacitating fibrosis and loss of limb function. We and others have identified delayed clearance of
apoptotic neutrophils as a primary mediator of fibrosis via detrimental effects on satellite cell and macrophage
behavior. Because macrophages are critical regulators of wound healing and also the primary cell type that clear
apoptotic neutrophils in a process called efferocytosis, macrophage cell therapy is a promising therapeutic
approach, but is limited by two main challenges: 1) Macrophages are highly plastic cells that rapidly shift
phenotype in response to microenvironmental cues. Therefore, a strategy is needed to control their phenotype
in situ following administration, to prevent them from changing phenotype in response to pro-inflammatory cues
at the site of injury. 2) High manufacturing costs and regulatory hurdles prevent the use of autologous (patient-
derived) macrophages, especially because very high numbers are required, but allogeneic (donor-derived)
macrophages elicit a strong T cell-mediated adverse immune response. We developed an innovative
biomaterial-mediated macrophage cell therapy strategy that simultaneously addresses both of these challenges.
In this strategy, referred to as Particle-Assisted Control over Macrophage-Neutrophil interactions (Pac-Man), the
macrophages are first loaded ex vivo with polymeric biodegradable microparticles that slowly release
dexamethasone intracellularly, thus controlling macrophage phenotype from the inside out following their
administration to sites of injury. Dexamethasone was selected because it causes an anti-inflammatory/pro-
regenerative phenotype in macrophages, increases their efferocytosis of apoptotic cells, and suppresses their
ability to activate T cells. Thus, the intracellular release of dexamethasone following administration of the
macrophages to sites of injury is expected to make them clear detrimental neutrophils, resolve inflammation, and
suppress T cell activation to prevent rejection of allogeneic cells. In Aim 1, the functional phenotype of the
adoptively transferred Pac-Man macrophages will be rigorously characterized over time in vitro and in vivo using
flow cytometry, gene expression profiling, and analysis of their interactions with apoptotic neutrophils. Effects on
muscle repair will be assessed using histology and functional mechanical testing. In Aim 2, the potential to use
an allogeneic cell source will be tested in vitro and in vivo using primary human immune cells from mixed donors
and mice from different strains. This project will advance an innovative off-the-shelf, translational cell therapy to
enhance clearance of apoptotic cells, which would be transformative for the treatment of fibrotic injuries such as
VML and many others.
