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In Vitro Osteoblast Model for Bone Wound Infections and Antimicrobial Therapy

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Technical Report,01 Oct 2011,30 Sep 2012

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The Geneva Foundation Tacoma United States

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We proposed to determine the changes in osteoblast differentiation in vitro when infected with multidrug resistant bacteria and the therapeutic effects of two well characterized antimicrobial peptides. For the first part of the study, an in vitro 3 dimensional3D model of primary human osteoblasts in a collagen scaffold was developed for infection. The 3D osteoblast cultures were infected with clinical isolates of multidrug resistant Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. Custom PCR arrays with genes associated with osteoblast differentiationmaturation were used to study gene expression in primary osteoblasts at several time points 4, 24, 48 and 72 hours post infection. The temporal gene expression patterns of infected osteoblasts in the 3D model were studied by real time PCR. Initial work suggested changes in the expression of osteoblast marker genes, several transcription factors, cell adhesion related genes and proinflammatory cytokines may contribute to an impediment for osteoblast differentiation and maturation when osteoblasts harbor any of these 3bacteria as intracellular pathogens. Antimicrobial therapy was tested with KSL-W peptide on osteoblasts in 3-D scaffolds and was effective against extracellular Acinetobacter baumannii and Klebsiella pneumoniae compared to uninfected controls. KSLW was not effective against Pseudomonas aeruginosa. KSL-W was not effective against intracellular bacteria. The outcome of our findings may contribute to accelerate treatment options and management of osteomyelitis due to drug resistant bacteria in the Wounded Warrior.

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