LXA₄ Upregulates Pro-Resolving Mediators to Limit Scarring After Cleft-Lip Repair

Objectives: Scarring is a frequent postoperative complication after cleft lip (CL) repair surgery that requires multiple revision surgeries to improve soft tissue form, function/movement. Dysregulated inflammation in the wound microenvironment favors fibrosis. In this rabbit study, we test the hypothesis that treatment with lipoxin A₄ (LXA₄), reduces scarring after CL repair surgery by decreasing excess inflammation via alterations of the wound lipid mediator profile.
Methods: A lip defect was surgically created to simulate a unilateral CL and then repaired in 8 six-week-old New Zealand white rabbits. An equal number of rabbits was randomly assigned to topical application of PBS (Control) or 1 ug of LXA₄ delivered as a methyl ester (ME) salt in PBS (treatment). Wound fluid was collected with filter paper at five timepoints up to 23 days after CL repair (n=40 samples). Filter papers were stored at -80°C until liquid chromatography–mass spectrometry (LC-MS/MS) quantification. Photographs of the cleft lip area defect and histologic specimens 42 days after the surgery were used to assess scarring between treatment groups. We used MetaboAnalyst v5.0.for lipidomic analysis.
Results: Animals treated with LXA₄-ME showed reduced scarring after CL repair. Scarring scores (Visual Assessment, Histology) were positively correlated with 13-HDoHE (pro-inflammatory) and negatively correlated with 5(S),15(S)-DiHEPE (pro-resolving). Linear Mixed Effects Models with covariate adjustments showed that treatment with LXA₄-ME resulted in an increase of pro-resolving mediators (LXB₄, RVE1, 5(S),15(S)-DiHEPE, MaR1(n-3,DPA)) and a decrease in pro-inflammatory mediators (5-HETE, 11-HETE, PGE₂, LTB₄) in wound fluid compared to treatment with PBS (p<0.05).
Conclusions: Topical application of LXA₄-ME reduces scarring after CL repair by shifting the wound lipid mediator profile from pro-inflammatory to pro-resolving. Our data elucidate the clinical translational potential of lipid mediators as biomarkers of wound healing after CL repair and offer a mechanistic rationale for future clinical trials of LXA₄-ME to limit scarring.