| Implant-associated osteomyelitis is a serious complication after orthopedic surgery,but the mechanism is not clear.The animal model of implant-associated osteomyelitis reported previously have their limitations and are difficult to meet the research needs.Here,we intend to establish a new mouse model of implant-associated osteomyelitis,and study its key genes through transcriptome analysis,providing new targret for further research.Chapter 1 establishment of a mouse model of implant-associated osteomyelitisObjective:To establish a mouse model of implant-associated osteomyelitis and evaluate its effectivenessMethods:Male C57BL/6 mice aged 10-12 weeks were divided into two groups(control group and S.aureus infection group).A 27-gauge needle was used to make a unicortical channel in the middle part of the right dorsal femur of the mouse.A 2mm stainless steel needle with a diameter of 0.3mm was inserted into the medullary cavity.Then,2μl PBS containing S.aureus(1X106 CFU/ml)was injected into the medullary cavity.X-ray imaging was performed on the 3rd,14th and 28th day after the operation to evaluate the radiographic changes.Bacterial burden was detected by bone tissue homogenization and the PBS washing implant by sonicated.SEM was used to detect the biofilms on the surface of implant.The localization of S.aureus in bone tissue was detected by immunohistochemical staining.Bone histological morphology was analyzed by HE staining and Goldner’s trichome staining.The number of osteoclasts was analyzed by TRAP stainingResults:The X-ray results showed that the mild periosteal reaction around the implant in the infection group occurred 14 days after the operation,and severe periosteal reaction and necrosis of bone were observed 28 days after the operation After bone homogenization,the burden of S.aureus in the day after 14 days and 28 days were significantly higher than 3 days after surgery.The results of implant bacteria culture and SEM showed that S.aureus could be colonized to the surface of implant Further immunohistochemical staining of S.aureus showed that S.aureus in the infection group was located near the implant in the medullary cavity and gradually spread to the distal femur with the extension of infection time.HE staining showed that with the extension of infection time,the pathological changes of bone tissue changed from acute inflammatory infiltration to gradual bone destruction,osteolysis and sequestrum formation,which gradually involved the distal femur.The results of Goldner’s trichome staining and TRAP staining indicated that the osteogenesis reaction outside the periosteal was intense and osteoclast activity was enhanced at 28 days after infectionConclusion:we successfully established a repearable,mouse model of implant-associated osteomyelitis with repearable,convenient and the can mimic the clinical disease development.Chapter 2:The bioinformatics analysis of the key genes of implant-associated osteomyelitisObjective:To explore the key genes of implant-associated osteomyelitisMethods:on the 3rd and 14th day after the operation,transcriptome sequencing and DEGs analysis were performed on the bone tissues of mice infected with S.aureus and control group,and the corresponding function,pathway information and interaction were further analyzed by GO,KEGG and PPIResults:On the 3rd day after surgery,101 DEGs were found,and most of these genes were involved in response to bacteria.On the 14th day after the operation,there were 1,702 DEGs in total.These genes not only focus on immunity of S.aureus,but also on angiogenesis and other tissue regeneration.Furthermore,Cxcl10 and Cxc19 induced by IFN-y may play a key role in the development of the disease.Conclusion:Transcriptometric analysis of bone tissue in mice with implant-associated osteomyelitis indicates a new target for further studies. |
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