Wound Stratification Elucidates Novel Genomic Perspective Of Negative Pressure Therapy In A Porcine Model
Jacob Hodge, MS1, Ashley Pistorio, MD2, Richard Korentager, MD1, David Zamierowski, MD1, Jennifer Robinson, PhD3, Adam Mellott, PhD1.
1University of Kansas Medical Center, Kansas City, KS, USA, 2University of Nevada Las Vegas, Las Vegas, NV, USA, 3University of Kansas, Lawrence, KS, USA.
PURPOSE: Negative Pressure Wound Therapy (NPWT) has been shown to modulate the behavior of wound tissue. Yet, the dynamic mechanism(s) of how NPWT exerts its effects at a molecular and cellular level remains unresolved; specifically, the key early signaling pathways initiated and the roles of different cellular populations upon exposure to negative pressure and different dressing materials. To date, wound analyses have only evaluated gene expressional profiles of whole wound isolates. In this study we provide a novel perspective that compares the expressional profile of "Full" (undissected skin layers) wounds relative to each separate, individual layer of wounded skin (epidermis/dermis/subcutaneous). Additionally, we investigate how different dressing materials modulate the early wound environment in the setting of NPWT.
METHODS: An acute incisional porcine wound model was developed by creating 2cm (long) x 2cm (deep) incisions on the flanks of two female Yucatan pigs (n=2). Four different wound dressings were applied within the incisional wounds, in addition to one control group. Dressing materials included Owen’s Rayon™, Granufoam™, Polycaprolactone (PCL) Mesh, and Fibrin Glue. Wounds were then treated with/without NPWT and evaluated over 8-hours. Tissue samples were collected at 0-, 2-, 4-, and 8-hours post-wounding and processed for Immunocytochemistry/Immunofluorescence (ICC/IF), histological staining, and RNA sequencing. Specimens, transverse to the incision, were bisected along the incision, with one side representing "Full" wounds and the opposing side further dissected into the epidermis/dermis/subcutaneous layers.
RESULTS: RNA sequencing provided spatial localization of expressional changes in key wound signaling pathways to specific tissue layers, including angiogenesis, inflammation, proliferation, and remodeling. Different expressional profiles were observed between individual tissue layers relative to each other within a single wound group and between each individual layer relative to "Full" wounds. For instance, upon stratifying the inflammatory expressional profile of "Full" wounds, identification of key expressional changes were seen in the epidermis and dermis for wounds treated with Granufoam™ and NPWT. Additionally, significant differences were seen between separate wound groups within a single tissue layer (epidermis vs epidermis) that were not always seen when comparing "Full" wounds. Similarly, different dressing materials with/without NPWT demonstrated varying abilities to modulate early wound signaling. As early as 8-hours post-wounding, notable differences in inflammation and re-epithelialization were observed histologically; specifically, the PCL mesh exhibited the lowest level of immune cell infiltration with evidence of early re-epithelialization occurring at the leading edge of the epidermal-dermal junction, whereas Fibrin Glue appeared to exhibit the highest level of inflammatory cell infiltration.
CONCLUSION: This study provides valuable information on medical material interactions with wound tissue and/or NPWT and provides insight into early genomic changes of wound tissue. The unique wound stratification and spatial transcriptomic approach utilized in this study provides a new methodology to observe expressional patterns more precisely within tissue and identify significant drivers of wound healing that may have otherwise not been detectable. Together this experimental data offers novel insight into early expressional patterns and genomic profiles, within and between tissue layers, in wound healing pathways that could potentially help guide clinical decisions and improve wound outcomes.
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