Angiogenic Steroids Induce Pathologic Scarring in a Preclinical Swine Model Via Dysfunctional Extracellular Matrix Deposition

Background Hypertrophic scarring is a major source of morbidity for surgery patients. An increasing number of transgender patients undergo surgery while on exogenous hormones. Based on clinical observations of increased frequency of post-op hypertrophic scarring in transgender males, we hypothesized that androgenic steroids lead to abnormal scarring and developed a preclinical swine model to characterize these observed effects. Methods A total of six male (XY) and female (XX) Hanford mini-swine underwent castration (ovariectomy/orchiectomy) and were randomly assigned to no testosterone (noT) or biweekly testosterone therapy (+T). Ten 3.5cm dorsal excisional wounds were created on each pig. To mimic a chronic wound, some wounds were re-excised at two weeks. Scars (POD42) and chronic wounds (POD28) were harvested six weeks after initial wounding and underwent analysis via histology, RNA seq, and tensile strength testing. Freshly excised tissue allocated for tensile testing was measured to obtain a cross-sectional area for normalization, loaded onto a 100N force-displacement apparatus, and subjected to a constant strain rate until burst failure. Results Histologic analysis of chronic POD28 wounds showed increased granulation tissue area (0.52cm2 for XXnoT, 0.66cm2 for XX+T, 0.88cm2 for XY+T, p=0.039) as well as increased epithelial thickness in the testosterone-treated groups (0.45cm2 for XXnoT, 0.62cm2 for XX+T, 0.68cm2 for XY+T, p=0.05). POD42 scars from the +T swine showed increased mean fibrosis area compared to XXnoT swine (0.157cm2 noT, 0.290cm2 XX+T, 0.268cm2 XY+T; p=0.007). Mean fibrosis thickness was also increased in the scars from T-treated swine (0.246cm2 noT, 0.389cm2 XX+T; 0.423cm2 XY+T; p<0.001). Scar tissue analyzed with mass spectrometry showed samples from noT swine had lower testosterone levels compared to the +T swine (p< 0.001). Scars in XX+T and XY+T pigs had greater tensile burst strength (p=0.024 and p=0.013 respectively) compared to scars in noT swine. Conclusion We developed a novel preclinical model to study the effects of the sex hormone testosterone on scarring. Testosterone induces early proliferation of excessive granulation tissue, which eventually leads to increased scar tissue. T appears to increase the physical strength of scars via supraphysiologic deposition of collagen and other ECM factors. The increase in burst strength observed for both XX and XY suggests that hormonal administration is a stronger influence on mechanical properties than karyotype. Anti-androgen topical therapies are a promising future area of research. ### Competing Interest Statement The authors have declared no competing interest.