| BACKGROUND:Global Tuberculosis Report 2014 showed that the incidence of tuberculosis(TB)and multidrug-resistant TB patients had increased worldwide.With the rising incidence of TB, the incidence of osteoarticular tuberculosis(OTB) also increased accordingly.Compared with TB,pain,dysfunction,deformities,paralysis and other symptoms caused by the invasive destruction of local bone from OTB more significantly affect the quality of work and life of the patients.What’s worse,as for the treatment,OTB is more complex than TB.Except for the standard therapy protocol by anti- tubercular drugs for a minimum of 8 and 20 months,a radical surgical debridement and the later reparation of bone defects are also involved.The following problems exist in the traditional treatment of OTB:1)the long-term medication results in patient noncompliance,side effects and drug resistance, 2)the inexhaustive debridement causes the recurrence of OTB, 3)the reparation of bone defects after the focal debridement tends to failure because of the recurrence of OTB,and so on.As to overcome these disadvantages, the scholars in the fields of medicine and bone tissue engineering have been opening a new stage of the treatment of OTB with biomaterials. Although the compositions of the biomaterials are different, the treatment mechanism of filling and repairing of bone defect after the debridement without autologous or allogeneic bone graft and releasing anti-TB drugs locally and sustainedly to effectively kill any remaining TB while maintaining the plasma concentration in the range of security and thus reducing drug side effects.OBJECTIVE:We prepared the cylindrical porous β-TCP scaffolds with the diameter of 5mm and the height of 5mm and the pore size of 400μm with 3D printing technology and prepared the PLGA microspheres with the diameter of 200±30μm,which co-encapsulated isoniazid and rifampicin,with the quality ratio of PLGA: INH: RFP = 60mg: 20mg: 15 mg and by the method of double emulsion solvent volatile.And then loaded the co-encapsulating-isoniazid-and-rifampicin PLGA microspheres into the porous β-TCP scaffolds by the centrifugation and gelatin encapsulation bonding technology to eventually get the composite materials of 3D printing porousβ-TCP scaffold loaded with anti-tuberculosis drugs control-release PLGA microspheres(CMs of S&M).They were used as the bone tissue engineering composite materials for filling bone defects to play the double roles of osteoconduction and local anti-tuberculosis treatment.METHODS AND RESULTS :The works are divided into three parts to study and evaluate theCMs of S&M:ExperimentⅠ:The preparation process,the scaffold porosity,the biomechanical property,and the degradation in vitro and the drug release of the composite material are researched and evaluated.The general process of a composite material is described above.Determined the scaffold porosity of the CMs of S&M(67.39±2.81)% by the weight and volume method.The maximum load of the CMs of S&M was 117.2±14.9N and the maximum strength 6.0±0.8MPa by the universal mechanical tester, relatively higher than 3D printing porous β-TCP scaffold with maximum load of 85.5±10.4N and the maximum strength of 4.4±0.5MPa(P < 0.05),and the two materials had met the requirements of the compressive strength of human cancellous bone(2-45MPa). The degradation time of the CMs of S&M was 20±1.3W by the quality of the composite material changing in SBF in vitro.Experiment Ⅱ:The biocompatibility of the CMs of S&M is evaluated in this part.The cytotoxicity in vitro of the CMs of S&M by the method of CCK-8 was classified as GradeⅠthat was non-cytotoxic. The blood test and the pathological examinations of heart,liver, spleen, lung and kidney were normal after the CMs of S&M were implanted in animals. ECT analysis showed that the uptake ratio of 99 m Tc-MDP(T/NT) is(98.2±7.6)% at 6 weeks after the implantation which indicated the good vascularization within the region of interest, consistent with the contralateral sides.The bone specimens pathology and micro-CT showed that bone grew well into the composite materials and gradually replaced the biodegrading materials. Through the above studies,it is confirmed that the CMs of S&M have the excellent biocompatibility, meeting the security requirements of the implant materials.Experiment Ⅲ:This part was about the research of the capacity of the CMs of S&M repairing the bone defect on femoral condyles in vivo animal experiment.ECT examination revealed the vascularization within a region of interest with the contralateral sides at 6 weeks after the implantation.The radiographic X-ray mammography, the Micro-CT images and the histological observation showed that the CMs of S&M within the bone defect gradually degraded and the bone mass increased so as to replace the composites in 1M, 3M and 5 M after the implantation. In 5M after the implantation,the bone volume fraction(BVF) was(90.56±6.43)%and the materialvolume fraction(MVF)(6.53±0.87)% by Micro-CT, with no significant difference from the scaffold group(P > 0.05).The bone density test showed the CMs of S&M group was(0.272±0.025)g/cm2 in 5M,not significantly different from the contralateral sides(P > 0.05).Under the NP70 Biological microscope,the bone sections of the composite material group by HE staining showed that the material had almost completely degraded and been replaced by new bone in 5M.As a result,apart from the mechanical strength significantly higher than the control group that was the scaffold group(P < 0.05),the experimental group that was the CMs of S&M group showed no significant difference(P > 0.05) in the terms of the biocompatibility, the material vascularization and degradation and the repair of bone defect.CONCLUSION:Therefore,the composite materials of a 3D printing porous β-TCP scaffold loaded with anti-tuberculosis drugs control-release PLGA microspheres(CMs of S&M) not reduces the biocompatibility and biological properties because of the load of anti-TB drugs PLGA microspheres.In conclusion,as the bone tissue engineering biomaterials, the CMs of S&M of excellent biocompatibility and biological properties shall have a tremendous potential in the reform process of the treatment of osteoarticular tuberculosis. |
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