Experimental Study Of Hydroxyl Butyl Chitosan Membrane On Repairing Dural Defect

Part 1:The research on the production,characterization and properties of hydroxyl butyl chitosan membraneObjective:To study the production,characterization and properties of hydroxyl butyl chitosan membrane:Produce hydroxybutyl chitosan membrane and investigate its microstructure,mechanical properties,biological safety,biocompatibility,biodegradability,ability in the prevention of adhesion,and effects on fibroblasts.To evaluate the advantages and disadvantages of the use of hydroxyl butyl chitosan membrane as a dural substitute.Methods:Produce hydroxyl butyl chitosan membrane by means of gelatin phase change coating and observe its microstructure under electron microscope.Detect the tensile strength and tensile toughness of hydroxyl butyl chitosan membrane and compare it with normal dura.Evaluate the biological safety,biocompatibility,and effects on fibroblasts of hydroxyl butyl chitosan membrane as a dural substitute.assessing whether it has the ability of biodegradability and ability in the prevention of adhesion.Results:Under the dry state,the hydroxyl butyl chitosan membrane showed high tensile strength,and good tensile toughne under wet condition.The tensile toughne of hydroxyl butyl chitosan membrane was better than that of the human dura mater.Observed by electron microscope,the cross and longitudinal secion of hydroxyl butyl chitosan membrane presents a net-like structure.Thus,it can be concluded that hydroxyl butyl chitosan membrane has a three-dimensional structure like honeycomb.Hydroxyl butyl chitosan membrane is a safe biological material for its good biological safety and biocompatibility.It has the function of preventing adhesion and inhibition of fibrous tissue hyperplasia like chitosan and its derivatives.The results of fibroblast culture shows that it can promote the proliferation of fibroblasts,and some of them are inhibited.The biodegradability of hydroxyl butyl chitosan membrane has been proved,whether hydroxyl butyl chitosan membrane can be absorbed needs to be tested in third parts.Conclusion: The hydroxyl butyl chitosan membrane showed good mechanical properties,biological safety,biocompatibility and good properties of preventing adhesion.Part 2:The research on the effects of hydroxyl butyl chitosan membrane on the cerebrospinal fluid leakage.Objective:The research on using hydroxyl butyl chitosan membrane preventing immediate cerebrospinal fluid(CSF):Establish an animal model of cerebrospinal fluid leakage caused by dural defect,and observe the effects of hydroxyl butyl chitosan membrane on the cerebrospinal fluid leakage.Methods:Five healthy adult New Zealand rabbits were selected to prepare the skull defect(1.2 cm ×1.2 cm in size)via midline incision of head.Cutting 0.8 cm X 0.8cm cerebral dura mater symmetrically and put suitably sized hydroxyl butyl chitosan membrane onto the experimental side of dural defect area;simultaneously,on the other side,only cut and do nothing as control group.Drill a small hole at a distance of about3 CM from the dura mater defect in the midline of skull,put a transparent tube through the hole into the subdural and injected 10 ML methylene blue solution with syringe(20ML)slowly.Repeating it for 5 times and observate whether cerebrospinal fluid flows out.Results:Hydroxyl butyl chitosan membrane behaves well in covering the dural defect.In the experiment,when methylene blue solution was injected into the subarachnoid space,there is no cerebrospinal fluid flowing out from the edge of hydroxyl butyl chitosan membrane,while a lot of methylene blue solution leaking from the control group.Repeating the experiment for 5 times and shows consistent resultsConclusion: Hydroxyl butyl chitosan membrane can effectively prevent cerebrospinal fluid leakage caused by dural defectPart 3:The research on the long-term effect of hydroxyl butyl chitosan membrane as a dural substitute to repair dural defectsObjective:Do research on the long-term effect of hydroxyl butyl chitosan membrane as a dural substitute to repair dural defects: Establish the animal model of dural defects,and compare the long-term effects of hydroxyl butyl chitosan membran and biological membran consisted of bovine tendon type I collagen as dural substitutes.bilateral dural defect(0.8 cm ×0.8 cm in size)via midline incision of head.The left defect was repaired with hydroxyl butyl chitosan membrane as the experimental group,the right defect was repaired with biological membrane as the control group.Observating the general situtation of experimental animals.At 30?90 and 180 days after operation,5 rabbits were sacriiced to observe whether there is Adhesion and inflammation around the operation area and for sampling HE staining of the operation area.Counting the fibroblasts and compare it with normaldura of rabbit.Besides,observing the newborn membrane under hydroxyl butyl chitosan membrane of 90 days after operation by electron microscopy.Methods:Fifteen healthy adult New Zealand rabbits were selected to prepare theResults:(1)All the experimental animals were in good condition,no paralysis,incontinence and other neurological symptoms is observed.Wound infection was not observed.(2)30 days after the operation,most of the biological membrane was absorbed,and was completely absorbed 90 days after the operation,inflammatory reaction and adhesion were not observed in the biological membrane and its surrounding at the 3 time points.30 days,90 days and 180 days after operation,inflammatory reaction and adhesion were not observed in the hydroxyl butyl chitosan membrane and its surrounding.Under the hydroxyl butyl chitosan membrane,a newborn translucent membrane can be seen covering the brain tissue.No adhesion is seen between the neonatal membrane and brain tissue.There was no scar formation on the surface of brain tissue,and no abnormal color was found in brain tissue.180 days after operation,there was no evidence of absorption of hydroxyl butyl chitosan membrane.(3)The results of HE staining of 30 days after operation shows that a large number of fibroblasts and collagen fibers were observed in the newborn membrane under hydroxyl butyl chitosan membrane when observed by microscope.30 days after operation,fibroblasts,collagen fibers,and a lot of fat cells can be seen in biological membrane and its surroundings when observed by microscope.There were significant statistical differences in the number of fibroblasts counts between the two groups.180 days after operation,the number of fibroblasts observed by microscope in the newborn membrane significantly reduced,and a lot of collagen fibers orderly arranged can be observed,which is similar to normal rabbit dura mater.There was no statistical difference in the number of fibroblasts compared with normal rabbit dura mater in the newborn membrane.180 days after operation,a lot of fat cells and with few fibroblasts and little collagen fibers was observed by microscope in the biological membrane and its surrounding tissues,which is in low similarity to normal rabbit dura mater.There was significant difference in the number of fibroblasts in biological membrane and its surrounding with normal dura mater.30 days,90 days and 180 days after operation,there was no obvious inflammatory cell infiltration observed by microscope in experimental group and control group.90 days after operation,a large number of collagen fibers orderly arranged were observed by the electron microscope in the newborn membrane under hydroxyl butyl chitosan membrane.Conclusion:Repairing dural defects with hydroxyl butyl chitosan membrane does not lead to inflammation and adhesion.There was a new membrane similar to the normal rabbit dura mater formation under the hydroxyl butyl chitosan membrane in the long term.In conclusion,hydroxyl butyl chitosan membrane can meet the requirement of repairing dural defects.