Effect of hyperbaric oxygen therapy (HBO) on implant-associated osteitis in a femur fracture model in mice

<div><p>Hyperbaric oxygen therapy (HBO) is applied very successfully in treatment of various diseases such as chronic wounds. It has been already suggested as adjunctive treatment option for osteitis by immune- and fracture modulating effects. This study evaluates the importance of HBO in an early implant-associated localized osteitis caused by <i>Staphylococcus aureus</i> (SA) compared to the standard therapy. In a standardized murine model the left femur of 120 BALB/c mice were osteotomized and fixed by a titanium locking plate. Osteitis has been induced with a defined amount of SA into the fracture gap. Debridément and lavages were progressed on day 7, 14, 28 and 56 to determine the local bacterial growth and the immune reaction. Hyperbaric oxygen (2 ATA, 90%) was applied for 90 minutes on day 7 to 21 for those mice allocated to HBO therapy. To evaluate the effect of HBO therapy the following groups were analyzed: Two sham-groups (12 mice / group) with and without HBO therapy, two osteotomy groups (24 mice / group) with plate osteosynthesis of the femur with and without HBO therapy, and two osteotomy SA infection groups (24 mice / group) with and without HBO therapy. Fracture healing was also quantified on day 7, 14, 28 and 56 by a.p. x-ray and bone healing markers from blood samples. Progression of infection was assessed by estimation of colony-forming units (CFU) and immune response was analyzed by determination of polymorphonuclear neutrophils (PMN), Interleukin (IL) - 6, and the circulating free DNA (cfDNA) in lavage samples. Osteitis induced significantly higher IL-6, cfDNA- and PMN-levels in the lavage samples (on day 7 and 14, each p < 0.05). HBO-therapy did not have a significant influence on the CFU and immune response compared to the standard therapy (each p > 0.05). At the same time HBO-therapy was associated with a delayed bone healing assessed by x-ray radiography and a higher rate of non-union until day 28. In conclusion, osteitis led to significantly higher bacterial count and infection parameters. HBO-therapy neither had a beneficial influence on local infection nor on immune response or fracture healing compared to the standard therapy in an osteitis mouse model.</p></div

Radiographic analysis of fracture healing in infected mice.

<p>In contrast to the non-infected mice, mice allocated to both osteitis groups showed a higher frequency of nonunion. This mouse was allocated to the osteitis / HBO group and serial X-ray on day 0, 7, 14, and 28 shows development of a nonunion.</p

Quantification of circulating free DNA (cfDNA).

<p>Quantification of circulating free DNA (cfDNA).</p

Blood amino-terminal propeptide of type I collagen levels.

<p>Analysis of amino-terminal propeptide of type I collagen (PINP) levels in blood samples revealed only a significant influence of HBO therapy after osteitis 14 days after infection.</p

Blood alkaline phosphatase levels.

<p>Analysis of alkaline phosphatase (AP) in the blood serum revealed only significant differences between the osteotomy and the osteitis alone on day 7 and between osteotomy / HBO and the osteitis / HBO group on day 14.</p

Mean score values of fracture healing.

<p>The fracture gap was measured on a.-p. X-ray scans and fracture healing was classified by a score. Individual and mean values of the score are summarized. Animals of the osteotomy and osteotomy / HBO group showed a sufficient healing of the fracture within the observation period and subsequently decreasing mean values as assessed by the score. Mean score values also decreased for the osteitis group over time but the score showed a greater individual heterogeneity and nonunion in individual animals. In contrast, median bone healing score increased for the osteitis HBO group till day 28 and improved on day 56.</p