Abstract
Purpose: :
We have previously demonstrated sex-specific delay in corneal wound healing. Inflammation and reparative responses are intimately linked; however, which inflammatory cells drive sex-specific differences in the corneal response remains to be determined. Macrophages (MØ) are critical regulators of all aspects of wound healing and lymphocyte differentiation. MØ polarize into two phenotypic states: the classically activated pro-inflammatory M1 and alternatively activated pro-resolving M2. We investigated whether estrogen regulates MØ polarization and function, which may drive sex-specific differences in corneal injury responses.
Methods: :
Mice were subjected to recurrent corneal epithelial injury to induce an amplified acute inflammatory response. To directly assess estrogen’s affects on MØ polarization and function, MØ were isolated from bone marrow of female mice and differentiated into M1 or M2. Apoptotic neutrophils (PMN) were coincubated with MØ and PMN myeloperoxidase activity in MØ was used as a phagocytosis index. PPT and DPN were used to quantify ERα- and ERβ-specific actions, respectively. QPCR was used to quantify gene expression and LC/MS/MS used to measure lipid autacoid formation.
Results: :
Expression of M2 markers was significantly lower in healing female corneas. M2 phagocytic capacity to remove apoptotic PMN was significantly lower (50±5%) than M1. Both ERα and ERβ decreased the phagocytic index of MØ by 34±4% and 18±4%, respectively, while LXA4 increased it by 58±16%, demonstrating that estrogen and LXA4 have opposing bioactions in MØ. This correlated with ERα (73%) and ERβ (55%) inhibition of 5-LOX and 15-LOX expression and function, essential enzymes in MØ driven formation of LXA4.
Conclusions: :
These findings provide evidence for inherent sex-specific differences in MØ polarization and function during acute corneal injury responses. Specifically, injury response in females triggers a reduced M2 response compared to males. Both ERα and ERβ regulate MØ function and downregulate the protective LXA4 circuit, which drives M2 polarization and promotes phagocytosis of apoptotic PMN. Taken together, these findings have important ramifications for sex-specific inflammatory diseases, where wound healing MØ are essential for restoring tissue homeostasis and regulating lymphocyte activation.
Keywords: inflammation • phagocytosis and killing • eicosanoids