(WHS-K3.04) A Novel Ex Vivo Human Fascio cutaneous Flap Perfusion Model to Investigate Skin Injuries
Thursday, May 16, 2024
9:15 AM – 10:15 AM East Coast USA Time
Skin is the first line of defense against burns, chemicals, radiation, and trauma injuries. Recent research discovered a wide range of pathways and agents to treat skin injuries but still, there is a wide gap in the knowledge due to the complex nature of the injuries. Often, animal models are used for testing new agents, yet they lack anatomical feature resemblance to human tissue. Human tissue-based models are ideal; however, maintaining complex tissue ex vivo is challenging. Here we describe a novel, optimized, and well-characterized model of a full-thickness human skin perfusion system that utilizes surgical waste skin to cultivate flaps ex vivo. Abdominal panniculectomy samples were collected as surgical waste. Under sterile conditions, we isolated and cannulated perforators of the superficial and deep inferior epigastric systems. We perfused the cannulated tissue using a bioreactor system capable of real-time monitoring of pressure, flow rate, fluidic temperature, and tissue temperature. Albumin-supplemented culture media at 60mmHg pressure with 6ml/min inflow was perfused throughout the run time of approximately three weeks. Angiosome distribution was confirmed by fluorescein angiography and infrared imaging. Flow rate measurements, vascular reactivity, daily tissue biopsy samples for histology and electron microscopy, cell viability, lactate production, and gene expression levels were measured to assess the viability of the flap. Utilization of the skin perfusion model for chemical and burn injuries was assessed by induction of chemical (Nitrogen Mustard) and burn wounds. Samples were collected from the wounded and control tissue for histology, protein, and gene expression analysis. Angiography verified that the SIEA to SIEV flap system successfully fed ~ 90% surface area of a large flap for two-three weeks. Flow rate, temperature, and pressure remained steady throughout ex vivo cultivation. The vascular reactivity test showed a physiological response of the vasculature upon application of a vasoconstrictor (epinephrine) and vasodilator (papaverine). H&E staining and TUNEL immunofluorescence staining revealed healthy and viable cells during the perfusion run. Isolated adipose stem cells and dermal fibroblasts at different time points during perfusion showed viability and proliferation dynamics compared to fresh tissue isolates. We observed a decrease in circulatory glucose levels and increased lactate levels upon insulin challenge. Nitrogen mustard wounds showed a gradual increase in the dead TUNEL-positive cells. We observed the epithelium layer disruption and damage upon burn injuries. Our results suggest that this novel model system can keep the tissue viable for an extended period (app. 3 weeks) ex vivo. This viable system can help us understand the pathways and be used as a subclinical drug testing model.