Treatment with Vancomycin Loaded Calcium Sulphate and Autogenous Bone in an Improved Rabbit Model of Bone Infection
Summary
This study presents an improved rabbit model infected with Staphylococcus aureus by blocking the same amount of bacteria in bone marrow. Vancomycin loaded calcium sulphate and autogenous bone are used for antibiotic and bone repair treatment. The protocol could be helpful for studying bone infection and regeneration.
Abstract
Bone infection results from bacterial invasion, which is extremely difficult to treat in clinical, orthopedic, and traumatic surgery. The bone infection may result in sustained inflammation, osteomyelitis, and eventual bone non-union. Establishment of a feasible, reproducible animal model is important to bone infection research and antibiotic treatment. As an in vivo model, the rabbit model is widely used in bone infection research. However, previous studies on rabbit bone infection models showed that the infection status was inconsistent, as the amount of bacteria was variable. This study presents an improved surgical method for inducing bone infection on a rabbit, by blocking the bacteria in the bone marrow. Then, multi-level evaluations can be carried out to verify the modelling method.
In general, debriding necrotic tissue and implantation of vancomycin-loaded calcium sulphate (VCS) are predominant in antibiotic treatment. Although calcium sulphate in VCS benefits osteocyte crawling and new bone growth, massive bone defects occur after debriding. Autogenous bone (AB) is an appealing strategy to overcome bone defects for the treatment of massive bone defects after debriding necrotic bone.
In this study, we used the tail bone as an autogenous bone implanted in the bone defect. Bone repair was measured using micro-computed-tomography (micro-CT) and histological analysis after animal sacrifice. As a result, in the VCS group, bone non-union was consistently obtained. In contrast, the bone defect areas in the VCS-AB group were decreased significantly. The present modeling method described a reproducible, feasible, stable method to prepare a bone infection model. The VCS-AB treatment resulted in lower bone non-union rates after antibiotic treatment. The improved bone infection model and the combination treatment of VCS and autogenous bone could be helpful in studying the underlying mechanisms in bone infection and bone regeneration pertinent to traumatology orthopedic applications.
Introduction
Bone infection usually results from bacteria or other microorganism invasion after trauma, bone fracture, or other bone diseases1. Bone infection may induce a high level of inflammation and bone tissue destruction. In the clinic, Staphylococcus aureus (S. aureus) is the predominant causative agent of bone infection2,3. The bone infection is painful, debilitating, and often takes a chronic course that is extremely difficult to treat4. At present, debridement of necrotic tissue and implanting of vancomycin-loaded calcium (VCS) beads have been confirmed as an efficient strategy for controlling local infection5,6. However, 10% to 15% of patients experienced a prolonged bone repair process, delayed union, or non-union after anti-infection treatment7. The large segment of a bone defect is the most difficult issue for orthopedic surgeons. An autologous bone graft is considered the optimal bone replacement in bone non-union treatment8,9.
To date, most of the studies on bone infection and autologous bone implantation have been conducted in various kinds of animal models, such as rats, rabbits, dogs, pigs and sheep10,11. Rabbit models are most commonly used for bone infection studies, as first performed by Norden and Kennedy in 197012,13. In our previous study, we used rabbit models following Norden's method, and we found that the quantity of S. aureus injected into bone marrow could not be quantified accurately, as the blood leaking out of bone marrow led to bacteria solution overflow.
This article presents an improved surgical method for inducing bone infection on rabbits. At the end of the procedure, a blood biochemistry test, a bacteriological examination, and a histopathologic examination were performed to verify the bone infection model. Then, VCS was implanted to inhibit infection, and autogenous bone was implanted to promote bone regeneration.
Protocol
Representative Results
Discussion
In the previous studies, various kinds of animal models were constructed to study both acute and chronic bone infection; however, the search for the ideal model still persists17,18. In addition, the ideal bone infection model is expected to simulate the pathological characteristics of bone infection in clinical setting, while the modelling periods, remain low cost and easy to carry out. So far, the rabbit bone infection model is the most common model in inflammat…
Divulgations
The authors have nothing to disclose.
Acknowledgements
This work was supported by the National Natural Science Foundation of China (81803808, 81873062), Zhejiang Provincial Medical and Health Science and Technology Fund (2017KY271) and the Science and Technology Project of Zhejiang Province (2017C37181).
Materials
| absorbable surgical suture | Jinghuan | 18S0604A | |
| asepsis injector | Jinglong | 20170501 | |
| bone wax | ETHICON | JH5CQLM | |
| CCD camera | Olympus | DP72 | |
| EDTA-K2 anticoagulant blood vessel | XINGE | 20170802 | |
| Electric bone drill unit | Bao Kang | BKZ-1 | |
| Electric shaver | Codos | 3800 | |
| flexible silica gel mold | WRIGHT | 1527745 | |
| Hematoxylin and Eosin Staining Kit | Beyotime | 20170523 | |
| Luria-Bertani culture medium | Baisi Biothchnology | 20170306 | |
| Medical-grade calcium sulphate | WRIGHT | 1527745 | |
| microcomputed tomography (micro-CT) | Bruker | SkyScan 1172 | |
| Microscope | Olympus | CX41 | |
| New Zealand white rabbits | Zhejiang Experimental Animal Center | SCXK 2014-0047 | |
| No. 11 scalpel | Yuanlikang | 20170604 | |
| normal saline | Mingsheng | 20170903 | |
| PBS | TBD(Jingyi) | 20170703-0592 | |
| pentobarbital sodium | Merk | 2070124 | |
| povidone-iodinesolution | Lierkang | 20170114 | |
| S. aureus freeze drying powder | China General Microbiological Culture Collection Center | ATCC 6538 | |
| sheep blood agar | HuanKai Microbial | 3103210 | |
| tryptic soy agar plates | HuanKai Microbial | 3105697 | |
| tryptic soy broth tubes | HuanKai Microbial | 3104260 | |
| Vancomycin | Lilly | C599180 |
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