Macrophage Phenotype in Acute Porcine Vocal Fold Injuries

Abstract

Acute vocal fold (VF) injury results in diminished vocal quality because of changes to the composition of the extracellular matrix that disturb their natural viscoelastic properties. Macrophages (MQ) are located at the most superficial layer of the lamina propria where they can promote or resolve injury. Present study characterizes and compares MQ function in 2 models of acute injury. VF injuries were created bilaterally by either surgical biopsy or single injection of lipopolysaccharide (LPS; 2µg/kg). Pigs were sacrificed at 1d or 5d post LPS or 3d, 7d, or 23d post surgery (n= 3/time point/ injury). Flow cytometry was used to phenotype isolated VF cells and RT-PCR was used to quantify gene expression in VF tissue. Uninjured VF were used as controls. VF MQ were identified as hi SWC9+ lo SWC3+. MQ from surgical and LPS injuries identified as hi SLA-DR+ (p<0.05) compared to controls and exhibited similar temporal changes by expressing hi CD16+ at 1d and 3d respectively (p<0.05). In surgical injuries, MQ exhibited hi CD163+ (p<0.05) and VFs had high levels of TNFα and TGFβ1 mRNA at 3d compared to 23d (p<0.05). No changes to IL-12, IFNγ, IL-10, and IL-4 mRNA post surgery. In LPS injuries, VF had higher levels of TNFα, IL-12, IFNγ, IL-10, and IL-4 mRNA at 1d and 5d compared to uninjured VF (p<0.05). Higher levels of IL-10 mRNA were seen at 1d post LPS compared to 5d (p<0.05). No changes to CD163 or CD80/86 post LPS. Acute VF injuries can profoundly influence MQ phenotype, imparting context dependent polarizations. Surgical injury created a profibrotic state with a more dormant inflammatory response during mid-stages of healing when repair is needed. LPS injury created a more balanced inflammatory state through release of anti- and pro-inflammatory molecules that may help accelerate healing.