Co-Culture of S. epidermidis and Human Osteoblasts on Implant Surfaces: An Advanced In Vitro Model for Implant-Associated Infections

• Published in: PloS one 2016-03, Vol.11 (3), p.e0151534-e0151534
• Main Authors: Zaatreh, Sarah, Wegner, Katharina, Strauß, Madlen, Pasold, Juliane, Mittelmeier, Wolfram, Podbielski, Andreas, Kreikemeyer, Bernd, Bader, Rainer
• Format: Article
• Language: English
• Subjects: Adhesion tests; Aerobic conditions; Antibiotics; Antiinfectives and antibacterials; Arthroplasty; Bacteria; Bacterial infections; Biocompatibility; Biofilms; Biology and Life Sciences; Biomechanics; Biomedical materials; Bone implants; Bone surgery; Cell growth; Cement; Centrifugation; Coatings; Coculture Techniques; Complications and side effects; Culture; Electron microscopy; Health aspects; Humans; In Vitro Techniques; Infections; Joint surgery; Laboratories; Medical equipment; Medicine; Medicine and Health Sciences; Models, Biological; Multiplicity of infection; Orthopedics; Osteoblasts; Osteoblasts - cytology; Physical Sciences; Polystyrene; Polystyrene resins; Prostheses and implants; Prosthesis-Related Infections - prevention & control; Research and Analysis Methods; Risk factors; Scanning electron microscopy; Sonication; Staphylococcal infections; Staphylococcus epidermidis; Staphylococcus epidermidis - physiology; Studies; Surface Properties; Surgery; Surgical implants; Titanium alloys; Titanium base alloys; Transplants & implants; Virology
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0151534

Total joint arthroplasty is one of the most frequent and effective surgeries today. However, despite improved surgical techniques, a significant number of implant-associated infections still occur. Suitable in vitro models are needed to test potential approaches to prevent infection. In the present study, we aimed to establish an in vitro co-culture setup of human primary osteoblasts and S. epidermidis to model the onset of implant-associated infections, and to analyze antimicrobial implant surfaces and coatings. For initial surface adhesion, human primary osteoblasts (hOB) were grown for 24 hours on test sample discs made of polystyrene, titanium alloy Ti6Al4V, bone cement PALACOS R®, and PALACOS R® loaded with antibiotics. Co-cultures were performed as a single-species infection on the osteoblasts with S. epidermidis (multiplicity of infection of 0.04), and were incubated for 2 and 7 days under aerobic conditions. Planktonic S. epidermidis was quantified by centrifugation and determination of colony-forming units (CFU). The quantification of biofilm-bound S. epidermidis on the test samples was performed by sonication and CFU counting. Quantification of adherent and vital primary osteoblasts on the test samples was performed by trypan-blue staining and counting. Scanning electron microscopy was used for evaluation of topography and composition of the species on the sample surfaces. After 2 days, we observed approximately 104 CFU/ml biofilm-bound S. epidermidis (103 CFU/ml initial population) on the antibiotics-loaded bone cement samples in the presence of hOB, while no bacteria were detected without hOB. No biofilm-bound bacteria were detectable after 7 days in either case. Similar levels of planktonic bacteria were observed on day 2 with and without hOB. After 7 days, about 105 CFU/ml planktonic bacteria were present, but only in the absence of hOB. Further, no bacteria were observed within the biofilm, while the number of hOB was decreased to 10% of its initial value compared to 150% in the mono-culture of hOB. We developed a co-culture setup that serves as a more comprehensive in vitro model for the onset of implant-associated infections and provides a test method for antimicrobial implant materials and coatings. We demonstrate that observations can be made that are unavailable from mono-culture experiments.