The Effect Of58S Bioactive Glass Coating On Polyethylene Terephthalates On Graft-bone Healing

The Anterior cruciate ligament (ACL) is the most commonly injured ligament in the body, for which surgery is frequently performed. In the young and active population with an ACL tear, reconstruction is often the best therapeutic option.Due to the limited intrinsic healing capabilities and the problems induced by autografts and allografts, such as donor site-related problems, disease transfusion, immunological rejection, several researchers have developed different ACL substitutes to meet clinical needs. The ACL reconstructions using artificial ligament was prevalent in the1980s and early1990s. The LARS artificial ligaments (Ligament Advanced Reinforcement System; Surgical Implants and Devices, Arc-sur-Tille, France), made of polyethylene terephthalate (PET), is one of the artificial options.Nowadays, more and more surgeon had accepted the new generation artificial ligament---LARS artificial ligaments. By virtue of its excellent mechanical property, using LARS has shortened the patients’back to sports time in the early postoperative period.However, in long-term follow-up the disadvantages were found that degenerative osteoarthritis had developed in most of the patients. It has been observed that there was fibrous scar tissue between the artificial ligament and the bone tunnel, which may hamper graft-to-bone healing, result in graft failure. The lack of bioactivity and biocompatibility of the artificial ligament might be the major cause of failure. Much effort has been made to improve the biocompatibility of the PET, such as surface modification by grafting functionalized chemical groups like carboxylic [8], amid, sulfonate and phosphate groups.Bioactive glasses are a group of synthetic silica-based bioactive materials with bone bonding properties first discovered by Larry Hench in the early1970s. The critical feature for bioactivity is a SiO2content<60%in weight. Bioactive glasses have been extensively investigated for applications as bone grafts owing to their ability to form a chemical bond to living bone. The most rapid bonding is achieved with bioactive glasses containing45-52%in weight of SiO2. Glasses with a SiO2content55-60%in weight react more slowly, show long-lasting bioactivity. The clinical applications of bioactive glass initiated in the1980s. Since then, bioactive glasses have been used in many medical and dental applications.58S is one of the bioactive glasses families. As to its high ratio of Ca/P, it is often used for bone frame.Therefore, we supposed that coating bioactive glasses (58S) on the PET fibers could enhance the graft-bone healing in ACL reconstructions. MC3T3-E1osteoblasts were used to investigate the coating effect in vitro. The tibia-articular model was designed to investigate the coating effect on graft-bone healing in vivo. Part One The Construction and Identification of bioactive glass composition materialObjective To construct and identify the bioactive glass coating materialMethods PET sheets which were taken from a LARS ligament or PET film were modified with a scissor into round slices (15.5mm in diameter) to fit for the24-well culture plate. The sheets were immersed in75%ethanol solution for4h, washed with a large amount of deionized water. Suction pressure drying system was adopted for drying the sheets at37℃for24h. The cleaned PET sheets were modified through plasma surface modification and subsequently immersed in a differential concentration coating solution (12,1:3,1:4, and1:5). The sheets were fully stirred in the solution by magnetic stirring apparatus for5minutes at37℃, then rinsed with deionized water and dried in air for48h. Both the coated and uncoated sheets were sterilized through a conventional gas sterilization technique by ethylene oxide gas. The samples were vacuum-coated with gold palladium and examined in an SEM at20-kV accelerating voltage to observe the surface morphology. The surface chemical composition was investigated by an X-ray photoelectron spectrometer.Results The uncoated PET film and fibers remained smooth and uniform. The best choice for coating solution was made with0.2g of bioactive glass powder,0.8g of gelatin powder and50ml phosphate-buffered saline (PBS). The particle size of58S glass ranges from90to710ppm. After modification, many particles could be found adhering onto the surfaces of the modified PET film and fibers. To further determine the chemical nature of these particles, the X-ray photoelectron spectrometry was adopted to investigate the chemical composition of the sheets. The control group was composed with C and O, while Si, Ca, P could also be found in the experimental group, which perfectly matched the58S bioglass composition (58%SiO2,36%CaO,6%P205).Conclusion using coating technique, composition material (58S-PET) was successful constructed at the appropriated coating solution. Part Two The effect of Polyethylene terephthalate (PET) coated with bioactive glass on osteoblastic proliferation and cytoactivity in vitroObjective To observe the different effects on osteoblastic proliferation and cytoactivity the different material in vitroMethods MC3T3-E1osteoblast cells were seeded onto the PET and the coating PET. Inverted microscope was used to observe the morphology of the osteoblast cells on both film groups. The numbers of osteoblast cells was counted by using Automatic Cell Counter. Alkaline phosphatase activity and MTT test were also taken to test the osteoblastic proliferation and cytoactivity at1,3,5days.Results In vitro study, the morphology of the osteoblast cells were as normal after72h culture. Furthermore, the density of the osteoblast cells in experiment group had seemed to be higher than the control group. The number of osteoblast cells was significantly higher in the coating PET than the PET by using Automatic Cell Counter at the5th day (6.250±0.27,8.92±0.17;P<0.05). Alkaline phosphatase activity and MTT test also showed that the coated fibrous scaffold induced significant osteogenesis and cell proliferation compared with PET without coating at1,3,5d.(p<0.05)Conclusion The58S coating showed to be nontoxic to osteoblasts. Both the quantity and the activity of the MT3T3-E1enhanced significantly in the experimental group than that of the control. Part Three The effect of Polyethylene terephthalate coated with bioactive glass on Mechanical examination and Histological assay in vivoObjective To observe the different effects on Mechanical examination and histological assay in vivoMethods Twenty-four skeletally mature male New Zealand white rabbits (mean weight2.7±0.3kg) underwent a surgical procedure to establish a tibia-articular tendon-bone healing model. The tibia-articular model was designed to mimic the real ACL reconstruction. The rabbits were anaesthetized with3%pentobarbital (30mg/kg). A medial, Para-patellar arthrotomy was performed, and the ACL was identified and horizontally transected in the midsection, creating a gap between the ends of the cut tissue. A3-mm diameter tunnel was drilled from the proximal tibia metaphysis to the keen joint cavity by a Keith needle (3.0mm diameter). The graft, with a size of4×1cm, was rolled into4cm in length and3mm in diameter.The graft was pulled into the tunnel of the tibia through the joint cavity and the graft ends were sutured with the adjacent periosteum and soft tissue. At about0.5cm length of graft was left out of the tunnel entrance for later biomechanical testing. The joint was irrigated, and the synovium and capsule were closed in a single layer with interrupted0-Vicryl suture. The skin was closed with interrupted nylon suture. To minimize the possible cross-effect of the two grafts (such as ions releasing from the bioglass into the blood circulation), the same grafts were implanted into both the lower limbs of the same rabbit. The animals were allowed full cage activity postoperatively until sacrifice at6and12weeks after surgery.Results The maximum load increased by time in both groups. At week6, the Load-to-failure was significantly higher in the58S-PET group (61.70±6.95N) than that of the control group (65.21±9.78N, p=0.03). At week12, the Load-to-failure was also significantly higher in the58S-PET group(89.25±9.50N) than that of the control group (71.38±6.26N, p=0.02). In the histological assay we have found that there was new bone formation in the indistinct interface between the graft and the host bone in both groups at6,12weeks, and a stronger binding was seen in the58S-PET group than in the control groupConclusion The58S-coating on PET could promote the new bone formation and subsequently leads to a positive effect on tendon-bone healing.