Experimental Study On Osteoarticular Tuberculosis

As one of the oldest communicable diseases accompanied with the history of human being, tuberculosis had been controlled for a time. But for some reasons such as people's neglect of prevention, unreasonable medication and the emerging of drug-resistance Mycobacterium tuberculosis, during the recent 20 years, the morbidity of tuberculosis has increased. According to the report of WHO at the eighth annual meeting of tuberculosis in 2005, Geneva, the number of emerging TB patients has increased to 8,800,000 and about 1,700,000 persons die of TB every year in the world since 2003【1】. China has taken the second place with respect to TB incidence and one of the 22 high-burden countries of TB in the world, and according to the notifiable disease report of the Ministry of Health, TB has become the first killer of communicable disease in China【2】. Account for 3-5 %【3】incidence among general TB and 35-50% of extrapulmonary TB, osteoarticular tuberculosis is on the first place of extrapulmonary TB【4】. The characteristics of osteoarticular TB are atypism and invasiveness, and together with destruction of bone, sequestration is the main pathological feature. As the results of the destruction of normal bone structure and formation of bone defects, limb deformity and paraplegia will severely affect the quality of the patients` life. Without any doubts, bone grafting is one of the optimal methods for the repair of bone defects, however the existence of residual Mycobacterium tuberculosis in the local tissue usually cause the failure of bone grafting. Based on the investigation of the TB pathogenesis and creation of osteoarticular tuberculosis animal model, we have prepared a novel bone graft material with the ability of anti- Mycobacterium tuberculosis and osteoinduction, which can be grafted at the first stage for repairing bone defects and killing mycobacterium at the same time, may be one of effective method for clinical treatment of osteoarticular TB.Part I: Study on the pathogenesis of osteoarticular tuberculosisObjective: To investigate the pathogeny and pathomechanism of bone destruction in osteoarticular tuberculosis. Methods: 1. Tuberculosis factors prepared by different methods were added to the osteoclast culture system in vitro to observe their effects on the inducing and activing osteoclasts. 2. Tuberculosis factors prepared by different methods were added to the osteoblast culture system in vitro to observe their effects on the quantity and activity of osteoblasts. 3. Tuberculosis factor prepared by sonic spallation(Mt sonicate)was injected subcutaneouly to the mice head to observe the changing of skull bone. Results: 1. Mt sonicate could increase the secretion of tumor necrosis factor(TNF-α)and induce the formation and activity of osteoclasts; 2. Mt sonicate could not inhibit the formation and activity of osteoblasts markedly; 3. Local injection of Mt sonicate could cause lower grade bone resorption and increasing of calcium ion in vivo. Conclusion: The pathological change of osteoarticular tuberculosis may be associated with that Mt sonicate activates the immune factor in vivo, induces the formation and activity of osteoclasts and causes the bone destruction locally.Part II: The creation of immune osteoarticular TB modelObjective: To create an osteoarticular TB model based on the pathogenesis of TB. Methods: Rabbits with negative reaction to purified protein derivative (PPD)of tuberculin were chosen as the model animal. Low dose of mycobacterium antigen was used to allergize the rabbits in advance, then high dose of Mt sonicate was injected locally to induce the delayed type hypersensitivity (DTH), causing bone destruction and immune osteoarticular TB model was created. The effects of this model were evaluated by gross observation, radiology and histology. Results: Caseification, osteoclast hyperplasia and bone destruction were found in the location of Mt sonicate injection. Conclusion: This animal model can simulate the pathological characteristics and can be used for research in depth.Part III: The preparation of anti-TB boneObjective: To prepare a novel anti-TB bone with both ability of anti-TB and osteoinduction. Method: Anti-TB bone was made of reconstituted bone xenograft (RBX)combined with 4 antituberculosis drugs ( Isoniazide, Rifampicin, Pyrazinamide, Ethambutol ) at certain concentration and encapsulated by gelatin and polycaprolactone. Mice muscle pouch test was used to detect the osteoinductivity of anti-TB bone and drug release tests in vitro and in vivo were used to detect the release characteristics of anti-TB bone. Results: The anti-TB bone could induce ectopic cartilage and bone formation in the muscle pouch of mice. The release of anti-TB bone was shown as rapid release at initial phase, slow release at middle phase and effective drug concentration lasting for long time. The local concentration at the 28d still could be effective for killing Mycobacterium tuberculosis. Conclusion: This anti-TB bone can be used as bone graft that could kill Mycobacterium tuberculosis and keeping good osteoinductivity at the same time.Part IV: The anti-TB bone used for treatment in animal modelObjective: To observe the healing effects of anti-TB bone on the animal models. Method: 1. Focal clearance combined with bone grafting of anti-TB bone at first stage was used to treat the osteoarticular tuberculosis model and the curative effects were evaluated. 2. The osteoinductivity of anti-TB bone was evaluated by using it to repair a critical radius defect model. Results: 1. The anti-TB bone could repair the defects by focal clearance and the local drug concentration could also kill the Mycobacterium tuberculosis effectively and the blood drug concentration is safe. 2. The anti-TB bone could repair critical radius defect and keep safe blood drug concentration at the same time. Conclusion: This anti-TB bone graft can be used for the treatment of osteoarticular tuberculosis model locally with systemic drug concentration within safety limits.Part V: The effects of antituberculosis drugs on the bone cells Objective: To observe the effects of antituberculosis drugs in anti-TB bone on the bone cells. Method: 1. Anti-TB drugs were added to the osteoblast culture system in vitro to observe their effects on the induction and activation of osteoblasts. 2. Anti-TB drugs or Anti-TB drugs combined with Tuberculosis factors were added to the osteoclast culture system in vitro to observe their effects on the quantity and activity of osteoblasts. Results: 1.A certain concentration of the antituberculosis drugs could decrease the quantity and activity of osteoblast. 2. Anti-TB drugs combined with Tuberculosis factors could decrease the quantity and activity of osteoclast. Conclusion: Combined with the antituberculosis drugs this anti-TB bone can affect the quantity and activity of the cell therefore cation is recommended when using it.With the upper five experiments, we believe that this anti-TB bone has both perfect osteoinductivity and effective anti-TB ability. For the general safety, it has promised a wonderful use for clinical treatment of osteoarticular tuberculosis.