C-Jun Induction Leads to Increased Scar Formation and Fibrosis in Mice
Bryan Duoto, Bachelor's of Science1, Alessandra Moore, MD1,2, Deshka Foster, MD1, Ruth E. Jones, MD1,3, Shamik Mascharak, Bachelor's of Science1, Gerlinde Wernig, MD1, Michael T. Longaker, MD, MBA1.
1Stanford University, Stanford, CA, USA, 2Brigham and Women's Hospital, Boston, MA, USA, 3University of Texas Southwestern Medical Center, Dallas, TX, USA.
Fibrosis and scar formation are major clinical issues which result in disfigurement and permanent functional loss. In both adults and children, excessive fibrosis after surgery or injury can result in complications that are difficult to treat, often recur, and have few effective therapeutic options. Additionally, the only animal models that exist to simulate these processes in humans include the red Duroc pig and rabbit ear, which are expensive, difficult to use, and do not provide transgenic modeling. Recently, a paper describing a transgenic mouse strain that utilizes over-expression of c-Jun, an AP-1 transcription factor, to induce global tissue fibrosis was published. We hypothesize that local induction of c-Jun in the same transgenic mouse would result in increased scarring after wound healing, simulating hypertrophic scarring and keloid formation.
Stented excisional dorsal wounding was performed on c-JuntetO; R26-M2rtTA mice. Mice received either c-Jun inducing agent doxycycline at 0.1mg/mL, and 2mg/mL concentrations or injections of phosphate buffered saline (PBS). Induction and dressing changes were performed every other day until wounds were completely healed. At each time point, photos were taken to quantify wound closure rate. Wounds were then harvested and stained with hematoxylin and eosin for scar thickness, trichrome for collagen deposition, and picrosirius red to quantify collagen subtypes. These assays quantify stain-specific wound characteristics. Healed wounds were also harvested and digested for flow cytometric analysis that identified key dermal lineage fibroblast precursors and terminal cells that contribute to wound healing.
Preliminary histological staining data reveals a significantly increased scar thickness in the c-JuntetO R26-M2rtTA mice as compared to the C57BL/6J PBS control mice (*p<0.0001, n=5) and the C57BL/6J 2mg/mL doxycycline control mice (*p<0.0001, n=5) (figure 1). Comparatively, in regards to the amount of scar collagen deposition, no significance exists between the c-JuntetO R26-M2rtTA mice and the C57BL/6J PBS control mice (p=0.0832) as well as the C57BL/6J 2mg/mL doxycycline control mice (p=0.1692) (figure 1). Flow cytometric data shows dermal progenitors which contribute to wound healing such as dermal papilla, papillar dermal fibroblasts, and reticular dermal fibroblasts exist in increased numbers in induced c-JuntetO R26-M2rtTA wounds (*p<0.0001, n=3).
Histological stains of c-JuntetO R26-M2rtTA dorsal wounds support that there are distinct differences in scar formation in the c-Jun transgenic model as compared to controls. These data support that this novel mouse model can be developed to study the molecular pathways which lead to, and inhibit, fibrosis. In future studies we will investigate novel inhibitors of fibrosis in this animal model, study the scar forming fibroblasts in greater detail, and compare our results to human specimens.
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