Pirfenidone-exosomes as innovative strategies for scarless tissue repair in wound healing.

To facilitate scarless wound healing, developing an anti-scarring treatment that modulates dermal fibroblast activity is a promising strategy, with pirfenidone (PFD) showing potential due to its anti-fibrotic properties by targeting intracellular pathways that regulate collagen disposition. PFD, particularly when delivered via dermal fibroblast-derived exosomes, may further enhance therapeutic effectiveness and promote scarless healing. Two common isolation methods-PEG precipitation and affinity-based techniques-were compared to identify the most efficient approach for obtaining high-purity and relatively homogenous exosomes derived from human dermal fibroblasts. Characterization techniques, including transmission electron microscopy (TEM), atomic force microscopy (AFM), antibody arrays, and enzyme-linked immunosorbent assays (ELISA), confirmed the successful isolation of high-purity exosomes. The affinity-based method demonstrated superior performance, yielding well-dispersed and highly pure exosomes. We optimized the encapsulation and formulation of the antifibrotic compound PFD by exploiting exosomes as a drug delivery platform, employing an active loading method via sonication to enhance encapsulation efficiency (EE%) and loading efficiency (LE%), while carefully controlling the sonication process to preserve exosome integrity. The optimal formulation of PFD-exosomes achieved an EE% of 11.14% ± 1.27% and an LE of 10.01% ± 1.03%, with a particle recovery rate of exosomes at 64.21% ± 8.49% using sonication technique. Then, we investigated how to harness exosomes and PFD-exosomes as innovative strategies for achieving scarless tissue repair in wound healing. Our findings showed that exosomes enhanced fibroblast migration and proliferation, highlighting their potential as a stand-alone cell-free therapy for wound healing. Additionally, this study was ground-breaking in demonstrating that exosomes can improve the efficacy of PFD as a drug carrier, amplifying its anti-fibrotic effects in both in vitro and in vivo models. The in vivo results indicated that PFD-exosomes accelerated wound healing while organizing the extracellular matrix (ECM) by reducing excessive collagen deposition. Overall, PFD-exosomes present an innovative strategy for pre-scarring interventions, offering benefits of enhanced wound healing outcomes while minimizing scarring.