Tom Hodgkinson1,2, Ardeshir Bayat1,3. 1. Plastic and Reconstructive Surgery Research, Manchester Institute of Biotechnology, University of Manchester, Manchester - UK. 2. School of Chemical Engineering and Analytical Science, Manchester Institute of Biotechnology, University of Manchester, Manchester - UK. 3. University Hospital of South Manchester NHS Foundation Trust, Faculty of Medical and Human Sciences, Institute of Inflammation and Repair, University of Manchester, Manchester Academic Health Science Centre, Manchester - UK.
Abstract
PURPOSE: Widespread application of collagen-glycosaminoglycan dermal templates in the treatment of cutaneous defects has identified the interval between initial engraftment and skin graft application as important for improvement. The aim of this study was to evaluate the effect of hyaluronan supplementation of Integra® dermal template on human dermal fibroblasts and keratinocytes in both in vitro and ex vivo models. METHODS: This study utilized in vitro and ex vivo cell culture techniques to investigate supplementing Integra® Regeneration Template with hyaluronan (HA), as a strategy to decrease this interval. In vitro, Integra® was HA supplemented at 0.15, 1, 1.5 and 2 mg/mL-1. Primary human dermal fibroblast (PHDF) and keratinocyte proliferation, PHDF viability, migration and HA-induced signal transduction (phosphor-MAPK Array) were assessed. Ex vivo, wound models (wound diameter 4 mm) were created within 8 mm skin biopsies. Wounds were filled with Integra® or HA supplemented Integra®. Re-epithelialization was compared through hematoxylin and eosin-stained cross-sections at 7, 14 and 21 days in culture. Model viability was assessed through lactate dehydrogenase (LDH) assays. RESULTS: In vitro, PHDF and keratinocyte proliferation were enhanced significantly (p<0.001) when supplemented with HA. S-Phase and G2/M PHDFs in HA supplemented scaffolds increased. PHDF viability was enhanced to 72 hours culture with 1.5 mg/mL-1 HA (p = 0.016). PHDF migration was maximally enhanced at 1 mg/mL-1 and 1.5 mg/mL-1, whilst increased levels of phosphorylated Erk/MAPK proteins indicated increased metabolic activity. In ex vivo models, HA supplementation accelerated re-epithelialization at all concentrations. This ex vivo model provides a robust model for preclinical assessment of skin substitutes. CONCLUSIONS: HA supplementation to Integra® demonstrates increased in vitro growth, viability and migration. Whilst ex vivo data suggest HA supplementation of Integra® may increase rapidity of wound closure.
PURPOSE: Widespread application of collagen-glycosaminoglycan dermal templates in the treatment of cutaneous defects has identified the interval between initial engraftment and skin graft application as important for improvement. The aim of this study was to evaluate the effect of hyaluronan supplementation of Integra® dermal template on human dermal fibroblasts and keratinocytes in both in vitro and ex vivo models. METHODS: This study utilized in vitro and ex vivo cell culture techniques to investigate supplementing Integra® Regeneration Template with hyaluronan (HA), as a strategy to decrease this interval. In vitro, Integra® was HA supplemented at 0.15, 1, 1.5 and 2 mg/mL-1. Primary human dermal fibroblast (PHDF) and keratinocyte proliferation, PHDF viability, migration and HA-induced signal transduction (phosphor-MAPK Array) were assessed. Ex vivo, wound models (wound diameter 4 mm) were created within 8 mm skin biopsies. Wounds were filled with Integra® or HA supplemented Integra®. Re-epithelialization was compared through hematoxylin and eosin-stained cross-sections at 7, 14 and 21 days in culture. Model viability was assessed through lactate dehydrogenase (LDH) assays. RESULTS: In vitro, PHDF and keratinocyte proliferation were enhanced significantly (p<0.001) when supplemented with HA. S-Phase and G2/M PHDFs in HA supplemented scaffolds increased. PHDF viability was enhanced to 72 hours culture with 1.5 mg/mL-1 HA (p = 0.016). PHDF migration was maximally enhanced at 1 mg/mL-1 and 1.5 mg/mL-1, whilst increased levels of phosphorylated Erk/MAPK proteins indicated increased metabolic activity. In ex vivo models, HA supplementation accelerated re-epithelialization at all concentrations. This ex vivo model provides a robust model for preclinical assessment of skin substitutes. CONCLUSIONS:HA supplementation to Integra® demonstrates increased in vitro growth, viability and migration. Whilst ex vivo data suggest HA supplementation of Integra® may increase rapidity of wound closure.
Authors: Matthew J Smith; Sandi G Dempsey; Robert Wf Veale; Claudia G Duston-Fursman; Chloe A F Rayner; Chettha Javanapong; Dane Gerneke; Shane G Dowling; Brandon A Bosque; Tanvi Karnik; Michael J Jerram; Arun Nagarajan; Ravinder Rajam; Alister Jowsey; Samuel Cutajar; Isaac Mason; Roderick G Stanley; Andrew Campbell; Jenny Malmstrom; Chris H Miller; Barnaby C H May Journal: J Biomater Appl Date: 2021-11-07 Impact factor: 2.646
Authors: Giovanni Nicoletti; Marco Saler; Laura Villani; Agnese Rumolo; Marco Mario Tresoldi; Angela Faga Journal: Front Bioeng Biotechnol Date: 2019-01-22