M. Granoski1, K. Fischer1, H. Kussie1, W. Hahn1, D. Sivaraj2, E. McKenna1, A. Hostler1, J. Yasmeh1, R. Erickson1, M. Witte1, K. Chen1, G. Gurtner1 1University Of Arizona, Surgery, Tucson, AZ, USA 2Stanford University, Palo Alto, CA, USA
Introduction: The FOXC2 transcription factor has been importantly tied to a wide range of disease states, serving as a promising prognostic biomarker clinically associated with highly aggressive basal-like human breast cancers (increased cancer invasion and metastasis). Systemic dysregulation of FOXC2 expression has also been found to promote defects in lymphatic remodeling and the rare familial syndrome of hyperplastic lymphedema-distichiasis (LD). Since chronic lymphedema is a forerunner of several malignancies and cancers have been known to arise from chronic wounds, we examined the effect of Foxc2 dysfunction on skin wound healing.
Methods: We used our splinted excisional wounding model that mimics human-like wound healing on wildtype and Foxc2 +/- mice (n=4), which demonstrate incomplete lymphatic vasculature, lymphatic dysfunction, and enhanced cancer metastasis. Wound size was measured over the course of 18 days. Tissue was explanted from both groups at post-operative day (POD) 14 and 18 and stained with Masson’s Trichrome to assess scar formation, Picrosirius Red for dermal integrity, or immunofluorescence to assess lymphatic (LYVE1) cell populations.
Results: Wildtype mice healed completely by POD 14, while Foxc2+/- mice did not completely heal until POD18. On PODs 8 (p<0.0001), 10 (p<0.0001), 12 (p<0.0001), and 14 (p=0.0002), the average wound size of the Foxc2+/- mice was significantly larger than that of the wild-type mice. Scar area of recently healed Foxc2+/- mice (POD 18) was significantly larger than that of recently healed wild-type mice (POD 14; p=0.0294). At POD 14, lymphatic dysfunction caused collagen fibers in the healing scar to be narrower (p=0.0117) and more highly aligned (p=0.0110), indicating significantly more fibrosis in Foxc2+/- mice compared to wildtype mice. Collagen fibers in both groups became significantly longer (p=0.0116) and wider (p=0.0020) over time (from POD 14 to 18), indicating a temporal evolution of fibrosis even during the typical remodeling phase of wound healing. Foxc2+/- mice also had lower numbers of LYVE1 cells compared to wildtype mice, indicating poor lymphatic vessel infiltration at the wound site.
Conclusion: Individuals over 65 years old are more likely to develop cancer and are highly susceptible to developing chronic wounds, which contribute to morbidity and mortality. Here, we found that FOXC2, which is tied to cancer metastasis and lymphatic dysregulation, also impairs wound healing and promotes fibrotic tissue architecture, linking these disease states together. With FOXC2 proposed as a potential therapeutic target for cancer metastasis, its pleiomorphic downstream systemic effects should be considered and carefully weighed against the increased chance of developing nonhealing wounds. Further delineation of the microenvironment, cellular events, and molecular signals during normal and Foxc2-associated abnormal wound healing will significantly improve clinical therapies targeting FOXC2.