The Role of Burn Tissue and Denatured Small Leucine Rich Proteoglycans in the Activation of the Toll-like Receptor 4 Pathway
Supervisor / Principle Investigator:
Dr. Edward Tredget
Dr. Jie Ding
Dr. Babita Agrawal
MD Class of 2021
Introduction: Hypertrophic scar, a common complication of burn injury to deep dermis with prolonged inflammation, causes reduced range of motion, intense pruritis, heat intolerance, and cosmetic problems. It does not respond well to current treatment options. Activation of toll-like receptor 4 (TLR4), a proinflammatory pathway, has been reported to be associated with hypertrophic scarring by responding to extracellular matrix (ECM) and endogenous cellular ligands to promote inflammation. Small leucine-rich proteoglycans (SLRPs) are molecules involved in wound healing that modify the ECM by altering fibroblast proliferation, collagen organization, and growth factors.
Hypothesis and Aim: We hypothesized that the endogenous molecules released from the damaged burn tissue could activate the TLR4 pathway in dermal cells. In this study, we determined the role of burn tissue and denatured SLRPs in the stimulation of the TLR4 pathway in vitro.
Methods: Burn tissues, including eschar and exudate, were collected from patients (n=8) at the University of Alberta Hospital. Total cells were isolated from part of the solid tissue by collagenase-digestion. HEK-Blue hTLR4 cells, the human embryonic kidney cells that were co-transfected with human TLR4, MD-2 and CD14 co-receptor genes and are used to determine the activation of TLK 4 pathway, were treated with the solid tissue, exudate, cells, denatured SLRPs decorin and biglycan, and LPS as a positive control. Secreted embryonic alkaline phosphatase (SEAP) assay was used to measure NF-kB activation as an indicator of TLR4 activity.
Results and Conclusion: We found that HEK-Blue hTLR4 cells treated with solid tissue, exudate, and burn tissue-isolated cells showed higher TLR4 activity compared to untreated cells, indicating that there may be endogenous molecules released from the burned tissue and cells that are able to stimulate the TLR4 pathway. We will further explore the mechanism, which will help in the development of future therapeutics, providing benefits to patients suffering with HTS and other fibroproliferative disorders.