Vancomycin Containing PDLLA and PLGA/ß-TCP Inhibit Biofilm Formation but Do Not Stimulate Osteogenic Transformation of Human Mesenchymal Stem Cells.

Aims: Chronic osteomyelitis, including implant-related prosthetic joint infection, is extremely difficult to cure. We develop vancomycin containing release systems from poly(d,l-lactide) (PDLLA) and poly(d,l-lactide-co-glycolide) (PLGA) composites with beta-tricalcium phosphate (ß-TCP) to treat methicillin-resistant Staphylococcus aureus osteomyelitis. We ask whether vancomycin containing PDLLA/ß-TCP and PLGA/ß-TCP composites will prevent early biofilm formation, allow cell proliferation and osteogenic differentiation, and stimulate osteogenic signaling molecules in the absence of an osteogenic medium. Methods: Composites were produced and characterized with scanning electron microscopy. In vitro vancomycin release was assessed for 6 weeks. Biofilm prevention was calculated by crystal violet staining. Human bone marrow-derived mesenchymal stem cells (hBM-MSCs) and osteosarcoma cell (SaOS-2) proliferation and differentiation were assessed with water soluble tetrazolium salt and alkaline phosphatase (ALP) staining. Real-time quantitative polymerase chain reaction defined osteogenic signaling molecules for hBM-MSCs. Results: Totally, 3.1 ± 0.2 mg and 3.4 ± 0.4 mg vancomycin released from PDLLA/ß-TCP and the PLGA/ß-TCP composites, respectively, and inhibited early biofilm formation. hBM-MSCs and SaOS-2 cells proliferated on the composites and stimulated ALP activity of cells. Runt-related transcription factor 2 (RUNX2) and SRY-Box transcription Factor 9 (SOX9) expressions were, however, lower with composites when compared with control. Conclusion: Vancomycin containing PDLLA/ß-TCP and PLGA/ß-TCP composites inhibited early biofilm formation and proliferated and differentiated hBM-MSCs and SaOS-2 cells, but osteogenesis-related RUNX2 and SOX9 transcription factors were not strongly expressed in the absence of an osteogenic medium for 14 days.

Biological Responses of Ceramic Bone Spacers Produced by Green Processing of Additively Manufactured Thin Meshes.

Bone spacers are exclusively used for replacing the tissue after trauma and/or diseases. Ceramic materials bring positive opportunities to enhance greater osteointegration and performance of implants, yet processing of porous geometries can be challenging. Additive Manufacturing (AM) opens opportunities to grade porosity levels in a part; however, its productivity may be low due to its batch processing approach. The paper studies the biological responses yielded by hydroxyapatite with ß-TCP (tricalcium phosphate) ceramic porous bone spacers manufactured by robocasting 2-layer meshes that are rolled in green and sintered. The implants are assessed in vitro and in vivo for their compatibility. Human bone marrow mesenchymal stem cells attached, proliferated and differentiated on the bone spacers produced. Cells on the spacers presented alkaline phosphatase staining, confirming osteogenic differentiation. They also expressed bone-specific COL1A1, BGAP, BSP, and SPP1 genes. The fold change of these genes ranged between 8 to 16 folds compared to controls. When implanted into the subcutaneous tissue of rabbits, they triggered collagen fibre formation and mild fibroblastic proliferation. In conclusion, rolled AM-meshes bone spacers stimulated bone formation in vitro and were biocompatible in vivo. This technology may give the advantage to custom produce spacers at high production rates if industrially upscaled.

In Vitro Assessment of Bioactive Glass Coatings on Alumina/Zirconia Composite Implants for Potential Use in Prosthetic Applications.

Achieving the stable osteointegration of prosthetic implants is one of the great challenges of modern orthopedic surgery. The fixation of ceramic acetabular cups of hip joint prostheses is usually achieved using a metal shell provided with screws or pegs that penetrate into the host pelvic bone. The deposition of bioactive coatings on the implant surface to be put in contact with bone could be a valuable strategy to promote a more "physiological" osteointegration. In this work, bioactive glass porous coatings were manufactured on the top of alumina/zirconia composite implants by two different methods, i.e., sponge replication and laser cladding. The coated samples underwent immersion studies in Kokubo's simulated body fluid (SBF) to assess in vitro bioactivity and were found to exhibit an excellent hydroxyapatite-forming ability, which is key to allow bonding to bone. Biological tests using mesenchymal stem and osteoblast-like cells revealed the good biocompatibility of both types of materials. Furthermore, a higher level of mineralization was induced by the sponge-replicated coatings at 10 days. Overall, these results are highly promising and encourage further research on these materials.

Stem Cell and Advanced Nano Bioceramic Interactions.

Bioceramics are type of biomaterials generally used for orthopaedic applications due to their similar structure with bone. Especially regarding to their osteoinductivity and osteoconductivity, they are used as biodegradable scaffolds for bone regeneration along with mesenchymal stem cells. Since chemical properties of bioceramics are important for regeneration of tissue, physical properties are also important for cell proliferation. In this respect, several different manufacturing methods are used for manufacturing nano scale bioceramics. These nano scale bioceramics are used for regeneration of bone and cartilage both alone or with other types of biomaterials. They can also act as carrier for the delivery of drugs in musculoskeletal infections without causing any systemic toxicity.

Treatment of implant-related methicillin-resistant Staphylococcus aureus osteomyelitis with vancomycin-loaded VK100 silicone cement: An experimental study in rats.

INTRODUCTION: The purpose of this present study is to investigate the efficacy of vancomycin-loaded VK100 silicone cement drug delivery system in the treatment of implant-related methicillin-resistant Staphylococcus aureus (MRSA) osteomyelitis in rats. MATERIALS AND METHODS: Thirty-six adult (18-20 weeks old) female Sprague-Dawley rats were included in the study. All rats underwent experimental osteomyelitis surgery via injecting 100 µL bacterial suspension of MRSA into the medullary canal. After a 2-week duration for the formation of osteomyelitis model, rats were assigned randomly into four groups: control (C), systemic vancomycin (V), local vancomycin-loaded VK100 silicone cement (vVK100), and systemic vancomycin and local vancomycin-loaded VK100 silicone cement (V+vVK100). The following treatment protocols were administered to each group for 4 weeks. For group C, 0.9% saline solution equivalent to the volume of vancomycin dose (approximately 1 ml/kg) was administered intraperitoneally twice daily (12-h intervals). For group V, 15 mg/kg of vancomycin was administered intraperitoneally twice daily (12-h intervals). For group vVK100, vVK100 polymer was included so that the intramedullary canal of the rats are affected. For group V+vVK100, vVK100 polymer was included so that the intramedullary canal of the rats are affected and 15 mg/kg of vancomycin was administered intraperitoneally twice daily (12-h intervals). After 4 weeks of treatment, clinical, radiologic, microbiologic, and histopathologic evaluations were performed for all groups. RESULTS: Results of this study revealed that all scores of the evaluation criteria for the treatment groups (groups V, vVK100, and V+vVK100) decreased due to the treatment protocols when compared to group C. These results show the effectiveness of all treatment protocols for the implant-related chronic MRSA osteomyelitis. However, there were no statistical difference between these three protocols. CONCLUSIONS: vVK100 polymer, as a local antibiotic delivery system, seems to be an effective method for the treatment of implant-related chronic MRSA osteomyelitis.

Mesenchymal Stem Cells and Nano-Bioceramics for Bone Regeneration.

Orthopedic disorders and trauma usually result in bone loss. Bone grafts are widely used to replace this tissue. Bone grafts excluding autografts unfortunately have disadvantages like evoking immune response, contamination and rejection. Autografts are of limited sources and optimum biomaterials that can replace bone have been searched for several decades. Bioceramics, which have the similar inorganic structure of natural bone, are widely used to regenerate bone or coat metallic implants. As people continuously look for a higher life quality, there are developments in technology almost everyday to meet their expectations. Nanotechnology is one of such technologies and it attracts everyone's attention in biomaterial science. Nano scale biomaterials have many advantages like larger surface area and higher biocompatibility and these properties make them more preferable than micro scale. Also, stem cells are used for bone regeneration besides nano-bioceramics due to their differentiation characteristics. This review covers current research on nano-bioceramics and mesenchymal stem cells and their role in bone regeneration.

Vancomycin containing PLLA/ß-TCP controls experimental osteomyelitis in vivo.

BACKGROUND: Implant-related osteomyelitis (IRO) is recently controlled with local antibiotic delivery systems to overcome conventional therapy disadvantages. In vivo evaluation of such systems is however too little. QUESTIONS/PURPOSES: We asked whether vancomycin (V)-containing poly-l-lactic acid/ß-tricalcium phosphate (PLLA/ß-TCP) composites control experimental IRO and promote bone healing in vivo. METHODS: Fifty-six rats were distributed to five groups in this longitudinal controlled study. Experimental IRO was established at tibiae by injecting methicillin-resistant Staphylococcus aureus (MRSA) suspensions with titanium particles in 32 rats. Vancomycin-free PLLA/ß-TCP composites were implanted into the normal and infected tibiae, whereas V-PLLA/ß-TCP composites and coated (C)-V-PLLA/ß-TCP composites were implanted into IRO sites. Sham-operated tibiae established the control group. Radiological and histological scores were quantified with microbiological findings on weeks 1 and 6. RESULTS: IRO is resolved in the CV- and the V-PLLA/ß-TCP groups but not in the PLLA/ß-TCP group. MRSA was not isolated in the CV- and the V-PLLA/ß-TCP groups at all times whereas the bacteria were present in the PLLA/ß-TCP group. Radiological signs secondary to infection are improved from 10.9 ± 0.9 to 3.0 ± 0.3 in the V-PLLA/ß-TCP group but remained constant in the PLLA/ß-TCP group. Histology scores are improved from 24.7 ± 6.5 to 17.6 ± 4.8 and from 27.6 ± 7.9 to 32.4 ± 8.9 in the CV-PLLA/ß-TCP and the V-PLLA/ß-TCP groups, respectively. New bone was formed in all the PLLA/ß-TCP group at weeks 1 and 6. CONCLUSIONS: CV- and V-PLLA/ß-TCP composites controlled experimental IRO and promoted bone healing. CLINICAL RELEVANCE: CV- and V-PLLA/ß-TCP composites have the potential of controlling experimental IRO and promoting bone healing.

Vancomycin containing PLLA/ß-TCP controls MRSA in vitro.

BACKGROUND: Osteomyelitis caused by Methicillin-resistant Staphylococcus aureus (MRSA) often requires surgery and prolonged systemic antibiotic treatment. Local antibiotic delivery systems of bioceramics or polymers have been developed to treat osteomyelitis. A disadvantage of biodegradable polymers is the initial burst of antibiotics into the environment; one advantage of bioceramics is its osteoconductivity. We therefore developed a vancomycin-containing poly-l-lactic acid/ß-tricalcium phosphate (PLLA/ß-TCP) composite to control antibiotic release and stimulate bone formation. QUESTIONS/PURPOSES: We (1) characterized these composites, (2) assessed vancomycin release in inhibitory doses, and (3) determined whether they would permit cell adhesion, proliferation, and mineralization in vitro. METHODS: We molded 250 vancomycin-containing (VC) and 125 vancomycin-free (VUC) composites using PLLA, ß-TCP, and chloroform. One hundred twenty-five VC composites were further dip-coated with PLLA (CVC) to delay antibiotic release. Composites were characterized according to their pore structure, size, volume, density, and surface area. Vancomycin release and bioactivity were determined. Adhesion, proliferation, and mineralization were assessed for two and three replicates on Days 3 and 7 with mesenchymal stem (MSC) and Saos type 2 cells. RESULTS: Pore size, volume, apparent density, and surface area of the CVC were 3.5 ± 1.9 µm, 0.005 ± 0.002 cm(3)/g, 1.18 g/cm(3) and 3.68 m(2)/g, respectively. CVC released 1.71 ± 0.13 mg (63.1%) and 2.49 ± 0.64 mg (91.9%) of its vancomycin on Day 1 and Week 6, respectively. MSC and Saos type 2 cells attached and proliferated on composites on Days 3 and 7. CONCLUSIONS: Vancomycin-containing PLLA/ß-TCP composites release antibiotics in inhibitory doses after dip coating and appeared biocompatible based on adhesion, proliferation, and mineralization. CLINICAL RELEVANCE: Vancomycin-containing PLLA/ß-TCP composites may be useful for controlling MRSA but will require in vivo confirmation.