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Abstract
The ideal wound closure device should restore the original integrity of the tissue, offer easy application and seamless, fluid-tight seal. Commonly used wound closure devices including sutures, staples and tissue adhesives do not offer effective sealing of the wound and possess a range of associated disadvantages including dehiscence, infection, toxicity and iatrogenic related trauma. Laser tissue welding has been proposed as an alternative and is becoming increasingly popular. This technique uses laser irradiation to initiate chemical reactions in a target material, thereby activating chemical components within the material to form an immediate seal. However, laser tissue welding is reliant upon temperatures exceeding the collagen denaturation point and is associated with some tissue damage.
SurgiLux is a chitosan-based thin film surgical adhesive that relies upon laser irradiation to increase the strength of chitosan binding to tissue without thermal damage to tissue and avoiding many of the disadvantages of current wound closure devices. Previous in vitro and in vivo studies of SurgiLux has demonstrated the potential of SurgiLux in sutureless repair of peripheral nerves. Recent approaches to regenerative medicine and tissue engineering involve the use of decellularised extracellular matrix as biological scaffolds to augment the formation of new functional tissue and facilitate successful tissue reconstruction. The aim of the work reported here was to combine SurgiLux with an extracellular matrix scaffold derived from porcine urinary bladder matrix to potentially improve the capacity of the SurgiLux technology to enhance wound healing by promoting functional tissue regeneration, for potential applications in peripheral nerve repair.
The bio-scaffold was incorporated into the SurgiLux film in a variety of ways; bio-scaffold embedded into SurgiLux had a greater tensile strength (32.4 ± 5.2 MPa), crystallinity (12.1 ± 1.3 %) and hydrophilicity (75.0 ± 2.0)° than the chitosan adhesive alone (8.5 ± 3.1 MPa, 10.7 ± 1.2% crystallinity, Ө = 98.1 ± 2.03°). Tissue adhesion strengths using these hybrid biomaterials were maintained at ~15 kPa compared to 3 kPa for fibrin glue. Furthermore, histological analysis demonstrated that laser irradiation of the UBM-SurgiLux adhesive caused no thermal damage to tissue.
In vitro biocompatibility of the composite films was assessed by examining their influence on the proliferation and health of olfactory ensheathing cells and human monocyte-derived macrophages. Incorporation of the bio-scaffold into the SurgiLux films increased the attachment and proliferation of olfactory ensheathing cells and decreased the cytotoxicity of the films. Similarly, while chitosan films induced a cell population to undergo early apoptotic activation, the composite films apparently increased biocompatibility, preventing the cells from undergoing necrosis. Similarly, while SurgiLux showed a significantly reduced macrophage response compared to chitosan film, introduction of the bio-scaffold into the SurgiLux reduced their response further. A quantitative real time PCR approach was undertaken to identify polarised macrophage phenotype, M1 (pro-inflammatory) and M2 (anti-inflammatory) through the detection of specific cytokines expressed by the macrophages. Reduced cell spread (6.2e3 ± 7.0e2 μm2) and lack of foreign body giant cell formation lead to significantly reduced expression of M1 markers, IL-23p19, IL-12p40 and IL-12p35; thereby suggesting the presence of an alternative anti-inflammatory, tissue remodelling pathway.
A protein expression profile of the UBM scaffold was generated to identify novel proteins within the UBM via an advanced mass spectrometry methodology. A total of 129 proteins were identified with the majority of these revealing a role in maintaining cell structure (19%) and adhesion (13%), while the smallest groups (1 %) had remodelling and stimulatory roles. A number of growth promoting proteins including galectins 1 and 7, obscurin, fibulin and have been identified that may have enhanced cellular proliferation on the UBM-SurgiLux composite scaffolds compared to chitosan films alone. UBM also contains proteins that have neurotrophic, anti-angiogenic, tumour suppressor activity and proteins known to promote tissue remodelling and morphogenesis.
Therefore, coalescence of the bio-scaffold within SurgiLux matrix resulted in a surgical adhesive with enhanced biocompatibility and reduced cytotoxicity compared to chitosan films. The results suggest that the unique combination of extracellular matrix bio-scaffold with SurgiLux technology has the potential to promote functional tissue regeneration leading to enhanced sutureless nerve repair.