Application Of Photochemical Tissue Bonding Technique In Nerve Repair In Rat Model Of Sciatic Nerve Injury

With the development of modern industry and transportation industry, peripheral nerve injury has become a common disease. For short nerve gap lesions, direct end-to-end nerve sutures are proposed as treatments. For long-lesion gaps, transplanting autologous nerve grafts (autografts) from uninjured sites to the injured site or using tissue engineering grafts to form a bridge between the two nerve stumps are proposed as treatments. However, none of these treatments have achieved satisfactory restoration of function. According to the literatures, the effect of peripheral nerve repair in between 60-85%.Local nerve tissue repair response is a kind of tissue regeneration; a variety of cells, inflammatory neurotransmitters may participate in and influence the effect of repair. Most previous studies focus on how to improve nerve regeneration; however, few experimental studies have focus on reducing the inhibition factors of nerve regeneration.The most common inhibit factor is scar formation at the repair sites. Scar formation is a very important aspect for normal wound healing, however, once the scars occurred at the repair site, the scar formation, as a physical barrier, might impede the extension of regenerated axons. The nerve tethering to the adjacent tissue might restrict the mobility of the nerve and thus cause traction injury and ischemia.In recent years, people realized that sacr formation is still inevitable even using careful microsurgical techniques and satisfactory restoration of function is still difficult to achieve. With the studies of nerve regeneration chamber, people gradually realized that avoiding the scar formation during the nerve repair progress through the intervention of external environment is also a very effective approach in promoting nerve regeneration.Photochemical tissue bonding (PTB) is a developingtissue repair technique that utilizes a photosensitizingdye and visible laser light to produce an immediate, water-tight seal between tissue surfaces. Due to the characteristics of suture-free, the adverse effects such as suture admage and suture reactions can be eliminated. Therefore, if the scar formation at the repair sites can be decreased, the blocking effect to the axons will be reduced, it is possible to better promote the recovery of function.In the current study, we successfully fabricated a collagen-chitoson (CCH) scaffold with longitudinally oriented channels using freeze-drying method. Then this scaffold was used to bridging a 10 mm long sciatic nerve defect in rats. Meanwhile, in order to reduce scar formation, we use photochemical nerve warps wrapping the scaffold. Finally, a combination of morphological and functional techniques was used to evaluate its efficacy. Part one: Fabrication and evaluation of collagen-chitoson scaffolds[Objectives] To prepare and evaluate the scaffold resembling the normal nerve.[Methods] The Nerve guidance scaffolds were fabricated by freeze-drying technique. Scanning electronic microscope was used to evaluate the inner structureof the scaffolds. The pore sizes, swelling ratio, biodegradability and biomechanical properties were also examined.[Results] We successfully fabricated a collagen-chitoson (CCH) scaffold with longitudinally oriented channels resembling the basal lamina micro-channels of normal nerves in the current study. All the scaffolds were circular cylinder, the microscopic channels were arranged in parallel manners and the pore sizes of the channels were uniform (27μm-68μm). And the results showed that the swelling ratio, biodegradability and biomechanical properties were markly improved cross-linking treatment. Part two: Assessment of processed human amniotic membrane as a protective barrier in rat model of sciatic nerve injury[Objectives] To investigate the effect of application of HAM as a nerve wrap to a 10 mm segment of transected and repaired nerve in rats.[Methods] The conglutinations deglee of muscle, fascia or nerve tissue and peripheral tissue was evaluated according to Petersen clasoification. And the picrosirius red staining was used to measure the thickness of the scar and nerve tissue. Meanwhile, the efficacy of the nerve regeneration was investigated by using a combination of morphological and functional techniques, including transmission electron microscope, electrophysiology, retrograde-labelling and sciatic function index. [Results] Nerves wrapped with HAM had significantly fewer adhesions and less scar formation than controls. Although the final outcome, both functionally and morphologically, was not significantly improved by wrapping the nerve with HAM, the observed decrease in adhesions and scar formation might help the nerve retain its mobility and thus prevent traction injury and ischemia, which are caused by nerve tethering to the adjacent tissue during the healing process. Part three: Peripheral nerve repair using photochemical tissue bonding technique[Objectives] To investigate the efficacy of PTB technique in bridging a long nerve gap in rats.[Methods] Application of PTB technique combined with HAM wraps to the 10 mm autograft and the 10 mm scaffold. The conglutinations deglee of muscle, fascia or nerve tissue and peripheral tissue was evaluated according to Petersen clasoification. Meanwhile, the efficacy of the nerve regeneration was investigated by using a combination of morphological and functional techniques, including transmission electron microscope, electrophysiology, retrograde- labelling and sciatic function index.[Results] Nerves treated with HAM wraps and sealed with PTB technique showed a statistically significant improvement across both functional and histologic evaluation. Nerves wrapped with HAM had significantly fewer adhesions and less scar formation than controls.