Optimizing the safety of intradiscal platelet-rich plasma: an in vitro study with Cutibacterium acnes.

Aim: The most common risk associated with intradiscal injection of platelet-rich plasma (PRP) is discitis with Cutibacterium acnes. It is hypothesized that antimicrobial activity of PRP can be enhanced through inclusion of leukocytes or antibiotics in the injectate. Materials & methods: Multiple PRP preparations of varying platelet and leukocyte counts were co-cultured with C. acnes with or without cefazolin, with viable bacterial colony counts being recovered at 0, 4, 24 and 48 hours post-inoculation. Results: A direct correlation between C. acnes recovery and granulocyte counts were observed. Conclusion: We observed the greatest antimicrobial activity with the leukocyte-rich, high platelet PRP preparation combined with an antibiotic in the injectate. However, cefazolin did not completely clear the bacteria in this assay.

Osteoconductivity and osteoinductivity of Puros(R) DBM putty.

Bone graft substitutes have been developed due to the limited supply and morbidity associated with using autogenous graft material. Allogeneic demineralized bone matrix (DBM) has been used extensively as a clinical graft material because of its inherent osteoinductive and osteoconductive properties. Differential enhancement of these properties may optimize the performance of these products for various orthopedic and craniofacial applications. Commercially available bone paste products consist of formulations that combine DBM with a carrier to facilitate handling and containment. In the present study, we present results of a comprehensive in vitro and in vivo characterization of a 100% human DBM putty product, Puros DBM Putty. Results indicate the DBM particles are completely dispersed in the putty. Data are presented showing the porosity of and cell attachment to Puros DBM Putty, thereby demonstrating the osteoconductive properties of this DBM. Puros DBM Putty was also shown to be osteoinductive in the rat ectopic pouch model. We demonstrate here for the first time that Puros DBM Putty maintains its activity to markedly stimulate or induce bone formation over the entire period of its shelf life. Taken together, these data demonstrate that the 100% human allograft derived Puros DBM Putty could be an effective bone graft substitute.

Demineralized bone matrix as an osteoinductive biomaterial and in vitro predictors of its biological potential.

The osteoinductivity of demineralized bone matrix (DBM) varies from donor to donor as a result of varying levels of multiple growth factors, matrix integrity, and artifacts from material processing. Many in vitro assays are currently used for screening the osteoinductivity of DBM. The objectives of this study were to determine the correlation of specific growth factors and in vitro mitotic stimulation to in vivo ectopic bone formation capacity with a large number of DBM samples. Samples were assayed using ELISA methods for BMP-2/4 and TGF-beta1 (n = 304) and cell proliferation using SAOS-2 osteoblasts (n = 239). All samples were then implanted intramuscularly in the abdomen of nude rats. All in vitro assays showed significant variability for any particular level of ostoinductivity determined by in vivo model. A significant, but only very weak, positive correlation to in vivo results was found for TGF-beta1 (r(2) = 0.016), BMP 2/4 (r(2) = 0.065), and SAOS-2 cell proliferation (r(2) = 0.053). The results of this study amplify the notion that a multitude of factors and their relative interplay, rather than a single factor are likely to determine the potency of any particular lot of DBM.

Factors that influence transgene expression and cell viability on DNA-PEI-seeded collagen films.

Gene delivery from tissue-engineering devices has the potential to improve healing, but better regulation of the level and duration of gene expression is needed. We hypothesized that transgene expression could be controlled by varying the fabrication and soaking parameters used in making collagen- based gene delivery scaffolds. Collagen films were made from acid-insoluble type I bovine dermal collagen and seeded with plasmid DNA encoding firefly luciferase, complexed with polyethylenimine. By varying the thickness of the films, the volume of the DNA soak solution, and the pH of the DNA soak solution, and by cross-linking the films, we identified variable combinations that produce significantly different levels of cell number and transgene expression in L-929 cells in vitro. Increasing film thickness or soak volume increased overall reporter gene expression. Decreasing film thickness or soak volume decreased cell number but did not significantly change reporter gene expression per cell. Cross-linking by ultraviolet irradiation (before adding the DNA) significantly decreased transgene expression, probably because of decreased swelling of the collagen film. These results suggest that collagen-based biomaterials may be designed and fabricated to induce, in a controlled fashion, various levels of cellularity and transgene expression.