In polytrauma patients invasive surgeries can potentiate the posttraumatic systemic inflammation thus increasing the risk of multi organ dysfunction. Therefore, fractures are initially treated by external fixators, which later are replaced by intramedullary nails. We showed that a severe trauma impaired the healing of fractures stabilized by external fixation. Here we studied, whether the conversion to an intramedullary nail increases posttraumatic inflammation and leads to further impairment of healing.

44 rats received a femur osteotomy stabilized by an external fixator (FixEx). Half of the rats underwent a thoracic trauma (TXT) at the same time. After 4 days the fixator was replaced by an intramedullary nail (IMN) in half of the rats of each group. The rats were killed after 40 and 47 days. C5a serum levels were measured 0, 6, 24, and 72h after the 1st as well as the 2nd surgery. The calli were evaluated by three-point-bending test, μCT and histomorphometry.

The TXT significantly increased serum C5a levels after the 2nd surgical intervention. After 40 days the switch from FixEx to IMN significantly decreased bending stiffness in rats with and without TXT. After 47 days flexural rigidity in rats subjected to conversion was significantly decreased compared to rats treated only with a FixEx, particularly in combination with TXT.

This study showed that after a severe trauma the conversion of the fixation could provoke a second hit and contribute to delayed fracture healing.

There is evidence that fracture healing is delayed in severely injured patients. We recently demonstrated that a blunt chest trauma, which induced posttraumatic systemic inflammation, considerably impaired fracture healing in rats. Because the complement anaphylatoxin C5a is an important trigger of systemic inflammation, we tested the hypothesis, whether the impairment of fracture healing observed after a severe trauma resulted from systemically activated complement.

16 male Wistar rats received a thoracic trauma and a femur osteotomy stabilized by an external fixator. Immediately and 12 h after the trauma, half of the animals received a C5aR-antagonist to prevent the C5a-dependent systemic inflammation. Control rats received a nonsense peptide, which does not provoke any biological effect. The animals were killed after 35 days and the calli were analyzed by three point bending testing, μCT and histomorphometry. Statistics: Mann-Whitney U test, level of significance to p<0.05.

The treatment with the C5aR-antagonist increased flexural rigidity significantly by 55%, improved bony bridging of the fracture gap and led to a slightly larger and qualitatively improved callus as evaluated by μCT and histological measurements.

This study shows, that the immunomodulation by a C5aR-antagonist significantly reduced the deleterious effects of a thoracic trauma on fracture healing. C5a could possibly represent a target to prevent delayed bone healing in patients with severe trauma.

Secondary fracture healing processes are strongly influenced by interfragmentary motion. Shear movement is assumed to be more critical than axial movement, however experimental results are controversial. Numerical fracture healing models allow to simulate the fracture healing process with variation of single input parameters and under comparable normalized mechanical conditions. Therefore, a direct comparison of different in vivo scenarios is possible. The aim of this study was to simulate fracture healing under several axial and shear movement scenarios and compare their respective time to heal. We hypothesize that shear movement is always more critical than axial loading. For the presented study, we used a corroborated numerical model for fracture healing in sheep. Numerous variations of the movement amplitude, the fracture gap size and the musculoskeletal loads were simulated for comparable axial compressive and shear load cases. In all simulated cases, axial compressive load had less inhibitory influences on the healing process than shear load. Therefore, shear loading is more critical for the fracture healing outcome in general. Thus, our findings suggest osteosynthesis implants to be optimized to limit shear movements under musculoskeletal loading.

Introduction

The fracture healing outcome is often evaluated via ex vivo testing of the fracture callus. However, there is only a small time window, where the callus stiffness is significantly different, i.e. a delayed fracture healing might be undetected if the time point of sacrifice is improper. The aim of this study was to develop an in vivo monitoring concept, which allows determining the fracture callus stiffness in vivo over the whole healing time in rats.

Dynamization of fracture fixation is used clinically to improve the bone healing process. This study evaluated the effect of late dynamization on callus stiffness and size in a rat diaphyseal femoral osteotomy. The external unilateral fixator was dynamized by removal of the inner fixator bar, at three weeks (D3-group: n=8) or four weeks (D4-group: n=9) post-operation. Published data of a five week rigid (R-group: n=8) and flexible fixation group (F-group: n=8) were included for comparison. Preoperative and postoperative movements of the rats were measured using a motion detection system. After 5 weeks the rats were sacrificed and healing was evaluated by biomechanical and densitometric methods. By 34 days post-operation, rats from the four fixation groups had similar activity levels. There was no significant difference in flexural rigidity, callus volume or callus mineral density between the D3 and D4-groups. Both the D3-group and D4-group had significantly greater flexural rigidity (p< 0.01) and significantly lower callus total volume (p< 0.03) and callus bone volume (p< 0.03) compared to the F-group. There was no significant difference in flexural rigidity or callus mineral density between the dynamized groups compared to the R-group. However, the D3-group had less callus bone volume (p=0.06) compared to the R-group. The D4-group had significantly less callus bone volume (p=0.02) and less callus total volume (p=0.05) compared to the R-group. Late dynamization led to a stiffer callus with a smaller callus volume compared to continuously flexible fixation. The late dynamized groups had less callus volume than the continuously rigid group, but the stiffness and calcification and of the callus were similar. The late dynamized groups had undergone resorption processes, indicative of more advanced healing. Late dynamization enhanced fracture healing compared to the continuously rigid or flexible fixation.

Introduction: Dynamization is used to improve the healing process. The optimal time for dynamization however remains unknown. In this study we proved the hypothesis that an early dynamization will improve the fracture healing.

Material and Methods: Twenty-four rats underwent a diaphyseal femoral osteotomy, with a 1mm gap. The osteotomy was stabilized by either rigid (R-group; n=8) or flexible (F-group; n=8) external fixation. The dynamized group (D-group: n=8) had a rigid fixation for 1 week, and then a flexible fixation for the remaining 4 weeks. The flexible fixation design resulted in an axial stiffness of 10N/mm and the rigid fixation in 74N/mm. After 5 weeks, healing was evaluated by biomechanical, densitometric, and histological methods.

Results: The flexural rigidity was 47% higher in the R-group than in the F-group (p< 0.01). Also, the flexural rigidity was 45% higher in the R-group than in the D-group (p< 0.01) (Table 1). Mineralized callus tissue volume was 37% lower in the R-group than the D-group (p=0.002).

Conclusion: The hypothesis could not be supported, in that early dynamization did not improve healing compared to rigid or flexible fixation. The rigid fixation had a stiffer callus with smaller callus volume, and more calcified tissue in the whole callus. The rigid fixation had bridging in the gap more often, which explains the increased flexural rigidity measured. Dynamization utilized in previous studies allowed closure of the fracture gap and thereby enhanced the rate of healing, which was not the case in the present investigation.

Introduction: Lateral callus distraction can be used to treat bone defects and increase bone diameter. However, this requires longitudinal splitting of the bone, which can be avoided by a new method.

Material and Methods: Twenty sheep were operated at the medial site of the tibia. After drilling 1 mm holes into the medullary cavity, a titanium plate with a hydroxyapatite coating was attached to the bone surface using a custom-made device

After 10 days this device was distracted 0.3 mm, twice a day until a distance of 6 mm was achieved (n=10). In the control group (n=10), the titanium plate was adjusted to a distance of 6 mm. All sheep received fluorescence labeling. 10 weeks p. o., bone formation underneath the titanium plate was investigated using pQCT, x-ray and histomorphology.

Results: In the distraction group there was 5 times more newly formed bone. > Bone columns bridging the space between the cortex and titanium plate were found. In the first 30 days, the bone formation was significantly enhanced in the distraction group as indicated by the fluorescent labeling

Conclusion: A strain induced bone formation is not only possible between two bony surfaces created by an osteotomy, but also between a bony surface and an appropriately designed implant. This new method allows for bone apposition in a large number of bones, which could not be properly treated in the past.

Introduction: Metaphyseal fracture healing is seldom studied and models used so far do not control the biomechanical conditions in the healing area. The aim of this study was to develop a new standardized and biomechanically adjustable osteotomy gap in an animal model.

Material and Methods: A pilot study with two sheep was performed. An incomplete osteotomy was created in the retro patellar region of the distal femur. The compression forces of the patella lead to a cyclic reversible reduction of the 3 mm osteotomy gap size. This inter-fragmentary movement (IM) can be limited by a steel implant fitted into the proximal osteotomy gap. After 8 weeks the sheep were sacrificed and the healing studied by x-ray and histology.

Results: The osteotomy gap, which allowed 0.3 mm IM showed woven bone bridging in the whole gap. The sheep with 1 mm IM showed only partial bone formation but mainly fibrous tissue and fibrous cartilage. There was no external callus formation.

Discussion: This new model allows the study of metaphyseal bone healing under various biomechanical conditions. Even though metaphyseal healing is different from diaphysical callus healing, IM appears to have a similar effect, leading to direct bone formation under stable condition and enchondral ossification under high IM.

Deficiencies of folate and vitamin B6 and B12 as well as increased methionine serum concentrations have been indicated to disturb bone metabolism, most probably due to an induction of hyperhomocysteinemia (HHCY). However, there is a complete lack of information on whether these metabolic changes affect fracture healing.

Therefore, the aim of this study was to analyze the impact of a methionine-enriched (n=13) and a B vitamin-deficient diet (n=14) on bone repair in mice. Controls were fed by the accordant standard diet (n=12 and n=13). Four weeks after stable fixation of a closed femoral fracture, animals were sacrificed to prepare bones for histomorphometric and biomechanical analyses. In addition, blood samples were obtained to evaluate serum concentrations of homocysteine (HCY), folate, and vitamin B12.

Quantitative analysis of blood samples revealed significantly increased serum concentrations of HCY associated with significantly decreased serum concentrations of folate and vitamin B12 in animals fed with the methionine-enriched diet or the B vitamin-deficient diet when compared to controls. Biomechanical evaluation showed no significant differences in bending stiffness between bones of the experimental and those of the control groups. In accordance, the histomorphometric analysis demonstrated a comparable size and tissue composition of the callus in all groups analyzed.

We conclude that a methionine-enriched and a B vitamin-deficient diet leads to HHCY, however, without affecting bone repair in mice.

Although IL-6 mRNA expression in rat is restricted to the first day post-fracture, the inflammatory phase, the protein has been observed later in the healing process, indicating additional roles. The importance of IL-6 was demonstrated by delayed healing in knockout mice through diminished osteoclast numbers, formation thereof being stimulated by IL-6. The aim of our study was to investigate with which cells this cytokine is associated and when during fracture healing.

A closed fracture of the lower right limb was created in rats. The tibia was obtained from six animals at each of 1, 3, 7, 14 and 28 days post-fracture, decalcified and prepared for standard immunohistochemistry with an IL-6-specific polyclonal antibody. The number and types of cells positively stained for IL-6 along the whole length of the periosteal callus on one surface and in the fracture was evaluated.

Mostly inflammatory cells were initially stained, becoming virtually absent by day 7 when this phase has normally ended. Within the immediate vicinity of the fracture where endochondrial ossification occurred, staining of chondrocytes was significant (69%) by day 7 when this cell was laying down cartilaginous tissue that was also calcified. Distally to the fracture where direct bone formation occurred through intra-membranous ossification by osteoblasts, staining of these cells was observed, peaking at day 14 (56%). As this bone started to take on the appearance of cortex and surviving embedded osteoblasts differentiated to osteocytes, the latter cells were stained, suggesting a role in remodelling. At the fracture as bone replaced the cartilaginous tissue and union occurred, staining of chondrocytes decreased, whereas local osteoblasts were positive.

IL-6 appears to play a role throughout fracture healing, in endochondrial and intra-membranous ossification. The level of staining of each cell type reflected the degree of their activity with respect to production of related tissue.

Kinematic evaluation of the knee after total joint arthroplasty plays an important role to analyze and understand the post operative outcome of the surgical procedure. The objective of the study was to quantify in vivo kinematics of two different knee designs (dual radius, single radius) by combining video fluoroscopy and helical axis of motion analysis.

3D position of the finite helical axis (FHA) of the displacement of the tibial component of the prosthesis relative the femoral component during a knee extension from 55° to 20° flexion underweight bearing conditions was computed. The motion data were extracted from in vivo fluoroscopy measurement. Angular deviations as angles between each FHA and the mediolateral axis of the femoral component of the prosthesis, and the localization deviation as the distance between each FHA and the center of the femoral component of the prosthesis were calculated. The median and the interquartile range (IQR) of the angular deviation and the localization deviation were computed. Non-parametric Wilcoxon test compared the values of the two designs.

The angular and localization deviations of the dual radius design were bigger than of the single radius design. Median localization deviation, IQR Angle deviation, IQR localization deviation showed highly significant differences between the two designs (p< 0.01).

Compared to the dual radius design the single radius design modified the knee kinematics in vivo. Since it is asingle axis design FHA is therefore concentrated near this unique single axis. On the contrary the dual radius design has two axes, and the FHA floated between these two axes.

In vitro experiments have shown, that stabilisation of the fibula in complete fractures of the lower leg give more stability compared to a single stabilisation of the tibia. However it is not known how this biomechanical conditions influence the bone healing process. To investigate the effect of fibula stability in tibia fracture healing tibial osteotomies in rats with and without fibula fractures were compared.

Male wistar rats (n=18) were operated by a transverse osteotomy of the proximal tibia of the left leg. Fracture was stabilised by intramedullary nailing. In 8 cases an additional closed fibula fracture was performed. The healing period was 21 days.

Each whole leg was examined by x-ray. After explantation of the tibia and removing of the nail and the fibula, the tibia was examined by CT-Scan, three-point-bending and histological evaluation.

Animals, who had a fibula fracture along with the tibia fracture presented with delayed healing. Density in CT-scan was 30% lower (p=0,0002) in animals with a fibula fracture (405mg/ccm, SD:64) compared to those without a fibula fracture (mean=577mg/ccm, SD:17). In three point bending the bending stiffness was 79% lower (p=0,0006) in animals with a fibula fracture (mean=252Nmm/mm, SD:118) compared to animals without a fibula fracture (mean=1219Nmm/mm, SD:478). The breaking force was 59% lower (p=0,0004) in animals with a fibula fracture (mean=17,5N, SD:6) compared to animals without a fibula fracture (mean=42,4N, SD:14).

Complete fractures of the lower leg healed considerably worse than solitary fractures of the tibia. We conclude that the missing of rotational stability of our k-wire fixation of the tibia with a unfixed fibula fracture is one of the reasons for the delay in fracture repair. The results support the in vitro findings of the biomechanical importance of the fibula for the stability of tibia fractures.

Non-steroidal anti-rheumatics (NSAR) are often used in patients with fractured bones for analgetic reasons. This animal experiment was performed to determine the influence of NSAR on the process of fracture healing. As an alternative central acting analgetic without peripheral effect Tramadol was included in this experiment.

Wistar rats were operated by a transverse osteotomy of the proximal tibia of the left leg, fracture was stabilized by intramedullary nailing (healing period 21 days). All therapeutics were applied orally, twice a day. The animals were divided in 4 groups, 10 rats each: Group 1 was treated with placebo, group2 with tramadol (20mg/kg bodyweight/day), group3 with Diclofenac-Colestyramin (5mg/kg/bw./day) over 7 days followed by 14 days placebo, group4 with Diclofenac-Colestyramin (5mg/kg/bw./day) over 21 days. On day 21 the rats were sacrificed and each leg was examined by x-ray, than the tibia was examined by CT-Scan, three-point-bending and histological evaluation.

There were no significant differences between group1 and 2 and between group3 and 4, respectively. Therefore the data of group1 and 2 as well as group3 and 4 are put together.

The results of CT and 3-point-bending showed, that rats treated by Diclofenac presented with delayed fracture healing compared to those treated by placebo or Tramadol. Bone density was 30% lower (p = 0,0001) in animals treated with Diclofenac (mean = 577mg/ccm, SD:53,1 in group1 and 2 vs. mean = 404,3mg/ccm, SD:27,3 in group3 and 4).

The breaking force was 45% (p = 0,0009) lower (mean = 42,4N, SD:14,2 vs. mean = 23,3N, SD:8,2) and the bending stiffness 56% (p = 0,0039) lower (mean = 1218,9Nmm/mm, SD:477,9 vs. mean = 532,6Nmm/mm, SD:389,9) in animals, treated with diclofenac. Diclofenacserumlevels on day 21 in rats with longtime diclofenac application (mean = 242ng/ml, SD:47,7) were comparable to those in humans.

Oral application of Diclofenac significantly delayed fracture healing in rats. This effect might be comparable to other NSAR and fracture healing in humans.

The treatment of large bony defects by callus distraction is well accepted, but the duration of treatment is long and the rate of complications increases accordingly. We have examined the effect of the stiffness of the axial fixator on reducing the time for maturation of callus.

We created a mid-diaphyseal defect of 15 mm in the metatarsal bone in sheep and stabilised it with a ring fixator. After four days a bony segment was transported for 16 days at 1 mm per day. After 64 days the animals were divided into four groups, three with axial interfragmentary movement (IFM) of 0.5, 1.2 and 3.0 mm, respectively, and a control group.

The 3.0 mm IFM group had the smallest bone density (p = 0.001) and area of callus and the largest IFM after 12 weeks; it also had typical clinical signs of hypertrophic nonunion. The most rapid stiffening of the callus was in the 0.5 mm group which had the smallest IFM (p = 0.04) after 12 weeks and radiological signs of bridging of the defect. These results indicate that suitable dynamic axial stimulation can enhance maturation of distraction callus when the initial amplitude is small, but that a large IFM can lead to delayed union.