Decellularized Dermal Hydrogel Mediated Wound Healing

Cutaneous wound healing is an intricate process in which the skin repairs itself after injury.Delayed wound healing or non-healing eventually creates additional costs and deterioration to the patient’s quality of life,even leading to disability and death.Epidemiological studies suggest that the prevalence of chronic trauma among all hospitalized patients in China is 1.7%,placing a serious burden on the health system and socio-economics of China.In the United States,approximately 6.5 million people suffer from trauma,and the annual health economic expenditure for the treatment of trauma and complications is $50 billion.In clinic,with the development of diagnosis and treatment technology,flap and skin graft transplantation,as the main treatment for cutaneous wounds,can make most patients get effective treatment.However,the limited source of autologous donor skin and the loss of donor tissue function seriously limit the application of this treatment.With the rapid development of tissue engineering and regenerative medicine,engineered biomaterials transplantation,as a potential treatment method,is increasingly used in the treatment of skin defects and wounds.Among them,acellular biological scaffold is a natural biological material that removes the cellular components in tissues and organs,retains the unique spatial structure of extracellular matrix in tissues and organs,preserves growth factors,cryptopeptides and other active components.The decellularized biomaterial is notable for its outstanding biological activity,fine biocompatibility,non-immunogenicity,and as a comprehensive raw material source.Vast researches reveal that acellular biological scaffolds provide physical and chemical support for cell survival and activities,and can effectively promote tissue and organ remodeling and regeneration.These advantages make acellular scaffold become one of the most promising biomaterials in the field of translational medicine.However,the geometric and mechanical characteristics of acellular biological scaffolds greatly restrict their clinical application.Tunable physical and mechanical properties,protecting wounds from microbial infection,and biodegradability properties make hydrogels ideal for treating skin wounds.Injectable hydrogels can change their original shape to meet the needs of wound size,and can be delivered to deep target areas through minimally invasive surgery.Thus,in the study conducted herein,we applied the accessible porcine dermal-derived decellularized scaffolds(ddECM)to fabricate a covalently crosslinked dermal extracellular matrix hydrogel(ddECMMA)through the amide bond reaction between methacryloyl groups and amino functional groups under the ultraviolet light,and evaluated the properties of the ddECMMA hydrogels from three aspects: material characterization,in vitro experiments and in vivo experiments.We explored the manufacture method and defined the preparation process and detailed parameters of ddECM scaffolds and ddECMMA hydrogels.Histological and immunofluorescent staining results showed that the cell components in the ddECM scaffold were removed,and the physical microstructure and main components of the dermal extracellular matrix were retained.Scanning electron microscope(SEM)was used to observe the microscopic morphology of native dermal tissue,the ddECM scaffold and the ddECMMA hydrogel.The results showed that the fibers in the ddECMMA hydrogel interlaced to form a stable topological structure.The micropore structure and size were conducive to nutrient exchange and cell infiltration.Growth factor protein chip analysis was used to compare the expression profile difference of growth factors among dermal tissue,the ddECM scaffold and the ddECMMA hydrogel.The results showed that the ddECM scaffold and the ddECMMA hydrogel retained most of the growth factors in normal dermal tissue,which laid a foundation for the biological activity of the ddECM scaffold and the ddECMMA hydrogel.The cytotoxicity of the ddECMMA hydrogel was detected by live/dead staining.The results showed that the ddECMMA hydrogel had good biocompatibilities.The tube-forming activity of HUVECs was evaluated by a tube formation assay,the results showed that the ddECMMA hydrogel showed high angiogenic activity.By constructing the rat back skin defect model,the ddECMMA hydrogel was implanted to repair the defect area,and the wound healing and skin regeneration effects were evaluated within 2 weeks.At0,3,7,14 days after operation,the growth of skin appendages such as hair follicles and sweat glands was analyzed by H&E staining and SOX9 immunofluorescence staining.The difference of angiogenesis in the wound was analyzed by micro-CT scanning and immunofluorescence staining.Masson’s trichrome staining and western blot analysis were used to evaluate the distribution and expression of collagen deposition in the wound.The results showed that the ddECMMA hydrogel significantly promoted the efficiency of wound healing,activated SOX9 positive hair follicle stem cells,promoted hair follicle development,and significantly improved angiogenesis.The time trajectory of various immune cells in the wound tissue was analyzed by flow cytometry.The results showed that the ddECMMA bioactive hydrogel activated the immune microenvironment in the wound,and induced the rapid transformation of M1 phenotype to M2 macrophages.Consequently,the decellularized dermal matrix-derived bioactive hydrogel orchestrates the entire skin healing microenvironment to promote wound healing and will be of high value in treatment of cutaneous wound healing.As such,this biomimetic ddECMMA hydrogel provides a promising versatile opinion for the clinical translation.