Experimental

31. Double-dose cefuroxime concentrations in bone, knee joint, and subcutaneous adipose tissue – A randomized porcine microdialysis study
Andrea René Jørgensen , Pelle Hanberg , Mats Bue, Maja Thomassen, Nis Pedersen Jørgensen , Maiken Stilling
Orthopaedic Research Unit , Aarhus University Hospital ; Department of Orthopaedic Surgery , Horsens Regional Hospital ; Department of Orthopaedic Surgery , Aarhus University Hospital ; Orthopaedic Research Unit , Aarhus University Hospital ; Department of Infectious Diseases, Aarhus University Hospital ; Department of Orthopaedic Surgery , Aarhus University Hospital


Background: Surgical site infection is a severe complication to orthopaedic surgery, which can prolong admission and increase cost. Optimal perioperative antimicrobial prophylactic treatment is a key factor in preventing surgically related infections.
Purpose / Aim of Study: To evaluate the time with concentrations above relevant minimal inhibitory concentrations (fT>MIC) of 4 μg/mL in bone, knee joint, and subcutaneous adipose tissue after double dose of cefuroxime given as either one bolus administration (1x3,000 mg) or two single doses with a four-hour interval (2x1,500 mg)
Materials and Methods: Sixteen female pigs (Danish Landrace breed, weight 73-77 kg) were randomized into two groups of eight: Group 1 received a bolus administration of 3,000 mg cefuroxime. Group 2 received two single doses of 1,500 mg administered with a four- hour interval. Microdialysis was applied for sampling in cortical and cancellous bone, knee joint, and subcutaneous adipose tissue. Plasma samples were collected as reference. Sampling was performed for eight hours.
Findings / Results: During an 8 h sampling interval, the mean percentage fT>MIC (4 μg/mL) across compartments was longer for Group 2 (298 – 422 min) compared to Group 1 (221 – 269 min) (p<0.01). In cortical bone, there was a tendency towards longer fT>MIC (4 μg/mL) in Group 2 (298 min) compared to Group 1 (221 min) (p=0.053). Within 50 min after administration, the mean concentration of 4 μg/mL was reached for both groups in all compartments. In tissues the mean concentrations decreased below 4 μg/mL after approximately 4 h (Group 1) and 3 h (Group 2) from initiation of administration (time zero).
Conclusions: A delayed tissue penetration was found in all tissues, where a mean concentration of 4 μg/mL was reached within 50 min for both groups in all compartments. During an 8 h interval, double dose cefuroxime administered as bolus 2x1,500 mg with a 4 h interval provides longer time above MIC breakpoint for S. aureus (4 μg/mL) than a single bolus of 3,000 mg cefuroxim. However, to maintain sufficient tissue concentrations during longer surgeries, re- administration of cefuroxime (1,500 mg) should be considered already 3 h after the first administration.

32. What is so special about the myotendinous junction ?– a RNA-sequencing study
Jens Rithamer Jakobsen, Peter Schjerling, Michael Kjær, Abigail Mackey, Michael Rindom Krogsgaard
Department of sports traumatology M51, Copenhagen University Hospital, Bispebjerg and Frederiksberg; Institute of Sports Medicine, Department of Orthopaedic Surgery M, Copenhagen University Hospital, Bispebjerg and Frederiksberg; Institute of Sports Medicine, Department of Orthopaedic Surgery M, Copenhagen University Hospital, Bispebjerg and Frederiksberg; Center for Healthy Aging, Department of Biomedical Sciences, Faculty of Health and Medical Science, University of Copenhagen; Department of sports traumatology M51, Copenhagen University Hospital, Bispebjerg and Frederiksberg


Background: The connection between the muscle fibers and the tendon, name the myotendinous junction (MTJ), is architecturally constructed to transmit force between muscle and tendon, but at the same time it is vulnerable to strain injury. In order to explain why these injuries occur and suggest how they can be prevented, a better understanding of the composition and cellular components of the MTJ is needed. Previous studies have shown the presence of an unique collagen type at the MTJ, Collagen XXII, which is not demonstrated elsewhere in the skeletal muscle system.
Purpose / Aim of Study: The purpose was to evaluate the gene expression of the MTJ and compare it to the adjacent muscle and tendon. We aimed to find new targets that are unique to the MTJ and of importance for the strength or recovery of the tissue. In addition, we wanted to identify targets that are higher expressed at the MTJ compared to the neighboring muscle and tendon.
Materials and Methods: Samples were collected from the superficial digitorum flexor muscle from 20 horses, frozen and sliced into sections containing muscle, MTJ and tendon tissue before preparation for RT- PCR. Based on the mRNA results a t-stochastic neighboring embedded plot (t-SNE) was made and sets of samples from 5 horses with the clearest separation between tissues were chosen for RNA sequencing. An expected contribution of muscle and tendon was calculated for all targets based on the known expression of 2-300 of the most selective muscle and tendon genes. Any variation between the expected and measured gene expression was regarded as expressed by the MTJ.
Findings / Results: No targets were found to be uniquely expressed at the MTJ. Collagen XXIIα1 was expressed 17-fold higher compared to the expected value. Generally, genes involved in remodeling and reformation of skeletal muscle fibers and extracellular matrix were expressed to a larger extent at the MTJ.
Conclusions: Despite the MTJ being a region specialized in force transmission with a highly specialized morphology no genes could be demonstrated as being unique to this region. The genes expressed higher in the MTJ compared to muscle and tendon were related to remodeling activities, and this confirms the previous finding of high rates of remodeling at the MTJ.

33. ELECTRICAL IMPEDANCE CHARACTERIZATION OF BONE FRACTURE
Markus Frost, Ming Shen, Stanislav Zhekov, Ben Krøyer, Gert Frølund Pedersen, Søren Kold
Dept. of Orthopaedics and Interdisciplinary Orthopaedics, Aalborg University Hospital, Denmark; Dept. of Antennas, Propagation and Millimetre-wave Systems , Aalborg University, Denmark; Dept. of Antennas, Propagation and Millimetre-wave Systems , Aalborg University, Denmark; Dept. of Antennas, Propagation and Millimetre-wave Systems , Aalborg University, Denmark; Dept. of Antennas, Propagation and Millimetre-wave Systems , Aalborg University, Denmark; Dept. of Orthopaedics and Interdisciplinary Orthopaedics, Aalborg University Hospital, Denmark


Background: Differences in electrical characteristics of different tissues can provide information of the tissue composition at the fracture region. Real-time monitoring of bone healing with electrical impedance spectroscopy might provide information for an individualized treatment of fracture patients. However, electrodes must be placed at a distance to the fracture site in order not to interfere with bone healing.
Purpose / Aim of Study: We investigated whether electrical impedance measurements from electrodes placed at a distance to a bone defect can detect differences between intact bone and bone defects in vivo.
Materials and Methods: Approval was granted from the Inspectorate of the Animal Experimentation of Danish Ministry of Justice. 6 rabbits were anaesthetized, and a fracture protocol was subjected to both tibias. Electrical impedance was measured in frequencies from 10 Hz to 1 MHz at each step: 1) intact bone, 2) medial defect, 3) medial and lateral defect, 4) complete 2 mm bone defect. One electrode was placed in the medullary canal and two electrodes extracortical (lateral and posterior) 2 mm from the defect. For each rabbit, one tibia had measurements with a free inner electrode and the other tibia had measurements both with a nail and an isolated nail.
Findings / Results: For all tibias, the intact bone resulted in higher impedance compared with the complete defect, and this difference was most pronounced in the frequency range of 1kHz to 100 kHz. This applied for all types of electrodes including electrode, nail, isolated nail. The isolated nail showed the biggest impedance difference between the intact bone and the complete defect. Incomplete bone defects had lower impedance compared with intact bone, but no consistent pattern for differences in impedance was observed between the different applied defects.
Conclusions: Consistent impedance differences between intact bone and complete defects were detected in-vivo in rabbits. Further research is needed to explore whether the presented method can be used to characterize bone healing over time.

34. The ability of comorbidity indices to predict mortality in an orthopedic setting: A systematic review and meta-analysis
Mari Jørstad, Per Gundtoft, Julie Erichsen, Hagen Schmal, Bjarke Viberg
Orthopaedic Surgery and Traumatology, Lillebaelt Hospital, University Hospital of Southern Denmark; Orthopaedic Surgery and Traumatology, Lillebaelt Hospital, University Hospital of Southern Denmark; Orthopaedic Surgery and Traumatology, Lillebaelt Hospital, University Hospital of Southern Denmark; Clinic of Orthopaedic Surgery Medical Center, Faculty of Medicine, University of Freiburg, Germany; Orthopaedic Surgery and Traumatology, Lillebaelt Hospital, University Hospital of Southern Denmark


Background: Several comorbidity indices have been created to measure and adjust for the estimated burden of comorbidity.
Purpose / Aim of Study: The objective of this systematic review was to evaluate and compare the ability of different comorbidity indices to predict mortality in an orthopedic setting.
Materials and Methods: A search string was developed in collaboration with a scientific librarian. The search string was used to extract possible studies from Embase, Medline, and the Cochrane Library. Two reviewers independently screened the studies using Covidence. This study was registered in Prospero and ROBINS-I assessment tool was used to assess the risk of bias. The area under the curve (AUC) was chosen as the primary effect estimate. An exploratory meta-analysis was made comparing the ability of the Elixhauser Comorbidity Index (ECI) and the Charlson Comorbidity Index (CCI) to predict in-hospital and 1-year mortality.
Findings / Results: Of the 5338 studies identified, 16 met the eligibility criteria. Most studies included patients with either hip fractures (7 studies) or arthroplasties (5 studies). The risk of bias was serious for 2 studies and moderate for the remaining. Overall the predictive ability of the different comorbidity indices ranged from poor (i.e. AUC <0.70) to excellent (AUC >0.89). The majority of the included studies only compared the ECI and CCI. In-hospital mortality for ECI and CCI was reported in 8 studies yielding an overall effect size of 0.84 (CI 0.81- 0.88) for ECI and 0.83 (CI 0.79-0.86) for CCI. The AUC values were generally lower for all other time points ranging from 0.67 to 0.78. For 1-year mortality the overall effect size was 0.69 (CI 0.66-0.72) for ECI and 0.70 (CI 0.67-0.74) for CCI.
Conclusions: ECI and CCI can equally be used to adjust for comorbidities when analyzing in-hospital mortality in an orthopedic setting. However, in general, both indices have poor to fair AUC values for 30-day and 1-year mortality, where other indices might perform better.

35. Sampling of the myotendinous junction – how can we do it?
Jens Rithamer Jakobsen, Peter Schjerling, Michael Kjær, Abigail Mackey, Michael Rindom Krogsgaard
Department of sports traumatology M51, Copenhagen University Hospital, Bispebjerg and Frederiksberg; Institute of Sports Medicine, Department of Orthopaedic Surgery M, , Copenhagen University Hospital, Bispebjerg and Frederiksberg; Institute of Sports Medicine, Department of Orthopaedic Surgery M, , Copenhagen University Hospital, Bispebjerg and Frederiksberg; Center for Healthy Aging, Department of Biomedical Sciences, Faculty of Health and Medical Sciences, University of Copenhagen; Department of sports traumatology M51, Copenhagen University Hospital, Bispebjerg and Frederiksberg


Background: The myotendinous junction (MTJ) is the region where strain injuries most often occur. Clinically, the risk for these injuries can be reduced through specific resistance training. This positive effect may be caused by changes in the concentration of structural proteins, leading to a strengthening of the MTJ. However, specific knowledge about the effect on the structure and tissue composition of the MTJ of resistance exercise is sparse. In order to study changes in protein content or gene expression at the MTJ it is necessary to isolate MTJ from the skeletal muscle and tendon to avoid that results from different tissues are mixed.
Purpose / Aim of Study: We aimed to develop a method to divide a sample taken from the MTJ into its three components: muscle, tendon and MTJ.
Materials and Methods: Samples were collected from the superficial digitorum flexor muscle from 20 horses and frozen routinely for immunohistochemistry. In frozen form each specimen was manually sliced parallel to MTJ into 10 µm thick sections and sampled for further processing. By controlling every 20th section visually it was noted whether the section contained muscle, MTJ or tendon. RT-PCR was performed on the collected sections identifying mRNA targets regularly used in the study of skeletal muscle and tendon. A Principle Component Analysis (PCA) and a t- distributed stochastic neighboring plot (t-SNE) were made on all the results to evaluate how well the different tissue regions had been isolated.
Findings / Results: It was possible visually to group the samples according to the three tissues. The t-SNE plot confirmed that the MTJ samples grouped specifically and were very similar in relation to their expression of the mRNA targets.
Conclusions: It was possible by this method to divide a specimen from the MTJ into muscle-, tendon- and MTJ-tissue. It is a cheap and specific method which is useful in studies looking into changes introduced at the MTJ following resistance exercise and experimental set- ups.

36. Manipulating the journal impact factor? The use of Journal-Self-Citations among orthopedic journals to boost journal rankings
Dorte Drongstrup, Søren Overgaard, David Minguillo
Research and Analysis Department, University Library of Southern Denmark; Orthopaedic research unit, Department of Orthopaedic Surgery and Traumatology, Odense University Hospital, Department of Clinical Research, University of Southern Denmark; , KTH Royal Institute of Technology


Background: The Journal Impact Factor (JIF) is often used as an indicator of research quality by tenure, promotion, and funding assessment committees. Thus, a higher JIF could lead to increased visibility for journals and more publication submissions. This provides incentives for journal editors to optimize in accordance with the JIF formula; the number of citations received in a given year to a journal’s publications from previous two years divided by the number of only articles and reviews from previous two years. However, the use of JIF to assess research quality is highly problematic, since it can easily be manipulated. A strategy to boost the JIF-score is by increasing the rate of Journal-Self-Citations (JSC) to the two previous years (JIF-years), which increases the number of citations (size of the numerator).
Purpose / Aim of Study: The aim is to investigate to what extent Orthopedic journals might use different strategies to influence and increase their JIF- scores.
Materials and Methods: All journals indexed in the subject category Orthopedics by the Journal Citation Report between 1997 and 2018 were analyzed. The data source was the in-house database version of Web of Science owned by the Royal Institute of Technology (KTH). The study covers 95 journals, 210,528 publications, and 3,990,809 citations. We analyze the publishing and citation patterns of these journals and apply different measures to identify which strategies might be the most frequent in the field to optimize the impact factors and which journals might take most advantage of these strategies to boost their JIF and ranking.
Findings / Results: Our first results show that the rate of JSC to JIF- years tend to be as almost double as high than usual. Still, there are large variance in the JSC intensity among journals. If the JSC to the JIF- years are excluded, the impact factor on average decreases 15%. For the 2018 JIF ranking, four journals in the top10 changes position when JCS are excluded.
Conclusions: The study finds a strong tendency for JSC in the JIF-years. It suggests that the inclusion of JSC in the calculation influences the JIF-scores and ranking of journals.

37. Bacteriophage treatment against Staphyloccus aureus periprosthetic osteomyelitis in rats
Britt Siesing-Mejer, Mogens Kilstrup, Steen Sejer Poulsen, Kristine Stub Rønø, Ulla Rydahl Johansen, Niels Henrik Søe
Department of Orthopaedic Surgery Handsection, Hillerød University Hospital; DTU Bioengineering, Technical University Lyngby; Endocrinology and metabolism, The Panum Institute, KU; DTU Bioengineering, Technical University Lyngby; Department of Microbiology, , Rigshospitalet, KU; Handsection, Department of Orthopaedic, , Herlev and Gentofte university Hospital, Denmark


Background: Bacteriophages (phages) are virus-like entities that only target bacteria and are composed of protein-encapsulated phage chromosomes. Using recognition proteins attached to the protein capsule, phages bind to the surface of specific bacteria and inject their chromosome into the cytoplasm. Inside the cell the bacterial gene expression machinery is hijacked by the phage chromosome, to produce a large litter of phage progeny, eventually killing the bacterium and releasing the phages into the environment.
Purpose / Aim of Study: To investigate the ability of Bacteriophages to eradicate S. aureus, in a knee prosthesis model of osteomyelitis in rats.
Materials and Methods: Ten Sprague-Dawley rats had prosthesis inserted in their left knee, and were divided into three groups. 103 S. aureus MN8, ica+ was inserted into the femoral and tibial bone marrow of the knee, before insertion of the prosthesis. The study included two bacteriophage groups with 4 rats in each group and one control group with two rats: One Bacteriophage group was given high dose (106), and one low dose (103) of bacteriophages, before insertion of the prosthesis. Control rats were given sterile saline (0.1 ml) and S. aureus bacteria. After two weeks the rats were sacrificed, and all specimens were analyzed clinically, radiographically, microbiologically and histologically.
Findings / Results: In the group with high dose of bacteriophage, two rats died of allergic reaction or cytokine storm. The remaining two rats showed no reaction to treatment. In the group with low dose of bacteriophage, one rat died. One rat had a nearly total clearing of infection. The other two rats, showed a significate reduction of infection in nearly all parameters. One rat in the control group died of unknown causes.
Conclusions: Bacteriophage treatment against S. aureus osteomyelitis, reduced the infection in the low dose group, but no effect was seen in the group given high dose of bacteriophage in any parameters. Two rats died in the group given high dose of bacteriophages because of cytokine storm. Lower dose than 103 bacteriophages might reduce the infection around the prosthesis, as judged by all parameters.