Disruption Of Focal Adhesion Kinase Mechanotransduction Signaling Regenerates Intradermal Adipose Tissue
Sun Hyung Kwon, PHD1, Britta A. Kuehlmann, MD2, Chikage Noishiki, MD, PHD2, Geoffrey C. Gurtner, MD2.
1Stanford University, Sunnyvale, CA, USA, 2Stanford University, Stanford, CA, USA.
PURPOSE: Focal adhesion kinase (FAK) plays a pivotal role in transducing mechanical signals to cells during wound repair resulting in increased fibrotic scar formation. Previously, we have shown that in small and large animal deep dermal wounds, pharmacological inhibition of FAK accelerates wound healing while attenuating fibrotic scar development and promoting regeneration of hair follicles and cutaneous glands. However, the molecular mechanisms by which FAK inhibition restores skin appendages leading to true tissue regeneration is far from completely understood. Here we reveal that loss of intradermal adipocytes is a hallmark of fibrotic scar lesions and blockade of FAK-mediated mechanotransduction signaling can regenerate intradermal adipocytes that may play a key role in regulation of hair follicle and skin appendage regrowth.
METHODS: Murine hypertrophic scar (HTS) model and porcine deep partial-thickness excisional wound model were studied to investigate the effects of FAK inhibition on regeneration of intradermal adipocytes. 1) Mouse model: For HTS model, full-thickness dorsal incisional wounds (2 cm) were immediately suture closed and left untreated until D4. On D4, sutures were removed and the healing tissue was subject to consistent mechanical loading until D14 to induce HTS. A small molecule FAK inhibitor (FAK-I; VS-6062, Verastem Oncology) was locally injected to the wound site daily until the animals were euthanized on D14. 2) Red Duroc model: Female red Duroc swine was used to create deep partial-thickness excisions (25 cm2 in size and 0.07 inches in depth). Wounds were immediately treated with FAK-I-releasing hydrogel for 90 days. FAK-I-releasing hydrogels were replenished twice a week until D90. Animals were euthanized on D180. 3) Tissue analyses: Specimens were collected at various time points after the initial injury. The presence of intradermal adipose tissue, hair follicles, and other skin appendages were visualized by histology and immunofluorescent techniques.
RESULTS: In the murine HTS model, healed scar lesions without FAK-I treatment displayed apparent fibrotic tissue lacking hair follicles and intradermal adipocytes positive for Perilipin A, a surface marker for fully differentiated adipocytes. Healed wounds treated with FAK-I showed an intact intradermal adipocyte layer positive for Perilipin A (N=3 each), similar to unwounded mouse skin. In the red Duroc model, intradermal adipocytes typically cluster around hair follicles and other cutaneous glands. With FAK-I treatment throughout early wound healing and remodeling phase, hair follicles, glands, and intradermal adipocytes positive for Perilipin A in the newly formed scar lesions began to reappear approximately at D28 (N=4 each) and increased in size thereafter. Regeneration of intradermal adipocytes and skin appendages were not found in untreated scar lesions at these time points examined.
CONCLUSION: The mechanisms by which inhibition of FAK-mediated mechanotransduction signaling results in wound tissue regeneration has not been identified previously. We used two distinct animal models to demonstrate that disruption of FAK leads to regeneration of intradermal adipocytes that may play a key role in regrowth of hair follicles and other skin appendages upon deep dermal injury. Our therapy, therefore, holds great potential as a unique therapeutic strategy for management of post-injury scar formation.
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