Reaction time and brake pedal force after total knee replacement: timeframe for return to car driving

Purpose: This prospective cohort study aimed to examine objective and subjective parameters in patients who underwent total knee replacement (TKR) to assess from when on driving a car can be deemed safe again. Methods: Thirty patients (16 women, 14 men, age 66±11 years) who received TKR of the right knee and 45 healthy controls (26 women, 19 men, age 32±9 years) were asked to perform an emergency braking manoeuvre using a driving simulator. Brake pedal force (BPF), neuronal reaction time (NRT), brake reaction time (BRT), and subjective parameters (pain, subjective driving ability) were measured preoperatively as well as 5 days, 3–4, and 6 weeks after TKR. Results: Preoperative NRT was 506±162 ms, BRT 985±356 ms, and BPF 614±292 N. NRT increased to 561±218 ms, BRT to 1091±404 ms and BPF decreased to 411±191 N 5 days after TKR. Three weeks after surgery, NRT was 581±164 ms and BRT 1013±260 ms, while BPF increased to 555±200 N. Only BPF showed signifcant diferences (p<0.01). In week 6, all parameters were restored to baseline levels; patients showed signifcant pain decrease and evaluated their driving abil ity as “good” again. Conclusion: BPF was the only parameter displaying a signifcant postoperative decrease. However, preoperative patients’ baseline levels and subjective confdence in driving ability were only reached 6 weeks after the operation. These results indicate that a minimum waiting period of 6 weeks should be considered before patients can safely participate in road traffic at their individual preoperative safety level again. Level of evidence II

BMP-2 Dependent Increase of Soft Tissue Density in Arthrofibrotic TKA

Arthrofibrosis after total knee arthroplasty (TKA) is difficult to treat, as its aetiology remains unclear. In a previous study, we established a connection between the BMP-2 concentration in the synovial fluid and arthrofibrosis after TKA. The hypothesis of the present study was, therefore, that the limited range of motion in arthrofibrosis is caused by BMP-2 induced heterotopic ossifications, the quantity of which is dependent on the BMP-2 concentration in the synovial fluid

Strategies to increase patient satisfaction by preventing complications in total knee arthroplasty

Ziel dieser Arbeit war es, die relevanten Komplikationen in der Knieendoprothetik bezüglich ihrer Entstehung und Vermeidbarkeit zu analysieren. Aufbauend auf diesen Ergebnissen sollten Strategien zur Prävention dieser Komplikationen und damit zur Steigerung der Patientenzufriedenheit entwickelt werden. Als relevante Komplikationen wurden die Fehlpositionierung der Implantate, die frühzeitige aseptische Lockerung, die Infektion und die Arthrofibrose identifiziert. In der konventionellen Operationstechnik konnte gezeigt werden, dass Durchführung der Resektionen fehleranfällig ist.Hierfür wurde ein neues Verfahren zur intraoperativen Überprüfung und Korrektur der Resektion entwickelt. Im Vergleich zur konventionellen Technik ermöglichen Patienten-spezifische Schnittblöcke zwar eine Verbesserung des Alignments, zeigen jedoch Abweichungen untereinander. Hier konnte gezeigt werden, dass die Magnetresonanztomographie (MRT) der Computertomographie (CT) in der Planung überlegen ist. Neben der Fehlpositionierung kann eine frühzeitige aseptische Lockerung durch eine unzureichende Fixation des Implantates verursacht sein. Im Bereich der tibialen Komponente erfolgt diese überwiegend zementiert. Die Einflussfaktoren der Zementierqualität sind folglich mögliche Ursachen einer frühzeitigen aseptischen Lockerung. Die Benutzung der Blutsperre konnte hier erstmals eine Verbesserung der Zementierqualität nachweisen. Die optimierte Zementiertechnik stellt daher die wichtigste Präventionsstrategie der unzureichenden Fixation und frühzeitigen aseptischen Lockerung dar. Die Ursache der septischen Lockerung ist die periprothetische Infektion. Operations- und patientenbezogene Faktoren stellen die wichtigsten Modulatoren des Risikos dar. Die durchgeführte Analyse zielte auf die Bedeutung dieser Risikofaktoren für die periprothetische Infektion. Eine präoperative Erhöhung des C-reaktiven Proteins (CrP) > 0,5 mg/dl war mit einem erhöhten postoperativen Infektionsrisiko verbunden. Die notwendige Änderung der Handlungsweise zur Prävention besteht eindeutig in der Verschiebung der elektiven Operation bei präoperativer CrP-Erhöhung > 0,5 mg/dl. Da die Ursachen der primären Arthrofibrose weitestgehend unbekannt sind, steht die Erforschung möglicher Einflussfaktoren und Regulationsmechanismen im Vordergrund. Aufgrund der bereits nachgewiesenen pathologischen Inflammation wurden Genexpressionsmuster und Zytokine der Inflammation bevorzugt betrachtet. Das proinflammatorische Zytokin Bone morphogenetic protein 2 (BMP-2) konnte als Einflussfaktor in der Gelenkflüssigkeit und der Genregulation von Synovialzel- len bei Arthrofibrose identifiziert werden. Die Konzentration des BMP-2 in der Gelenkflüssigkeit zeigte darüber hinaus eine lineare Korrelation mit der Zunahme der Fibriosierung der Gelenkkapsel. Im Gegensatz zu den vorherigen Komplikationen ist die Arthrofibrose mit dem gegenwärtigen Wissensstand als unvermeidbar anzusehen. Eine sinnvolle Präventionsstrategie der Arthrofibrose ist daher nicht vorhanden. Durch systematische Strategien zur Komplikationsvermeidung in der Knieendoprothetik lassen sich meißten der untersuchten Komplikationen vermeiden oder zumindest vermindern.The aim of this work was to analyze the relevant complications in total knee arthroplasty regarding their origin and preventability. Strategies for prevention of these complications and therefore improvement in patient satisfaction should be developed based on these findings. Malalignment, early aseptic loosening, infection, and arthrofibrosis were identified as the relevant complications. Conventional instrumentation is sensitive for implant malpositioning. A new surgical technique for control and adaptation of the distal femoral resection was established. Patient-specific instrumentation (PSI) leads to improvement in implant alignment in comparison to the conventional technique. It was shown that magnetic resonance imaging (MRI)-based were more accurate than computer-tomography (CT)-based PSI. Alongside malalignment, early aseptic loosening due to insufficient implant fixation can be a cause of failure. The gold standard for tibial implant fixation is cementing. Cementing technique is therefore a possible cause of failure. The use of a tourniquet was shown to improve cementing quality and might therefore prevent early loosening. Loosening can also be caused by infection. Surgical- and patient-side risk factors are the most important influencing factors of risk for infection. An increase of the C-reactive protein (CRP) above 0.5 mg/dl was found to be an independent risk factor for periprosthetic infection. Therefore surgery should be delayed in case of elevated CRP. The pathogenesis of arthrofibrosis is mostly unknown. Therefore the regulation and influencing factors are in the focus. As a pathological inflammation was described before, we analyzed the gene expression profile of the joint fluid of patients with arthrofibrosis after TKA. The pro inflammatory Bone morphogenetic protein 2 (BMP-2) was found as influencing factor in the joint fluid and a synovial cell culture. Moreover the concentration of BMP-2 correlated with the fibrosis of the periarticular soft- tissue. With the recent knowledge arthrofibrosis is still unpreventable. With systematic strategies for prevention of complications in total knee arthroplasty most of the analyzed complications can be decreased or avoided

Impact of antagonistic muscle co-contraction on in vivo knee contact forces

Abstract Background The onset and progression of osteoarthritis, but also the wear and loosening of the components of an artificial joint, are commonly associated with mechanical overloading of the structures. Knowledge of the mechanical forces acting at the joints, together with an understanding of the key factors that can alter them, are critical to develop effective treatments for restoring joint function. While static anatomy is usually the clinical focus, less is known about the impact of dynamic factors, such as individual muscle recruitment, on joint contact forces. Methods In this study, instrumented knee implants provided accurate in vivo tibio-femoral contact forces in a unique cohort of 9 patients, which were used as input for subject specific musculoskeletal models, to quantify the individual muscle forces during walking and stair negotiation. Results Even between patients with a very similar self-selected gait speed, the total tibio-femoral peak forces varied 1.7-fold, but had only weak correlation with static alignment (varus/valgus). In some patients, muscle co-contraction of quadriceps and gastrocnemii during walking added up to 1 bodyweight (~ 50%) to the peak tibio-femoral contact force during late stance. The greatest impact of co-contraction was observed in the late stance phase of stair ascent, with an increase of the peak tibio-femoral contact force by up to 1.7 bodyweight (66%). Conclusions Treatment of diseased and failed joints should therefore not only be restricted to anatomical reconstruction of static limb axes alignment. The dynamic activation of muscles, as a key modifier of lower limb biomechanics, should also be taken into account and thus also represents a promising target for restoring function, patient mobility, and preventing future joint failure. Trial registration German Clinical Trials Register: ID: DRKS00000606, date: 05.11.2010

Functional Comparison of Chronological and <em>In Vitro</em> Aging: Differential Role of the Cytoskeleton and Mitochondria in Mesenchymal Stromal Cells

<div><p>Mesenchymal stromal cells (MSCs) are of high relevance for the regeneration of mesenchymal tissues such as bone and cartilage. The promising role of MSCs in cell-based therapies and tissue engineering appears to be limited due to a decline of their regenerative potential with increasing donor age, their limited availability in human tissues and the need of <em>in vitro</em> expansion prior to treatment. We therefore aimed to determine to which degree <em>in vitro</em> aging and chronological aging may be similar processes or if <em>in vitro</em> culture-related changes at the cellular and molecular level are at least altered as a function of donor age. For that purpose we established MSCs cultures from young (yMSCs) and aged (aMSCs) rats that were cultured for more than 100 passages. These long-term MSCs cultures were non-tumorigenic and exhibited similar surface marker patterns as primary MSCs of passage 2. During <em>in vitro</em> expansion, but not during chronological aging, MSCs progressively lose their progenitor characteristics, e.g., complete loss of osteogenic differentiation potential, diminished adipogenic differentiation, altered cell morphology and increased susceptibility towards senescence. Transcriptome analysis revealed that long-term <em>in vitro</em> MSCs cultivation leads to down-regulation of genes involved in cell differentiation, focal adhesion organization, cytoskeleton turnover and mitochondria function. Accordingly, functional analysis demonstrated altered mitochondrial morphology, decreased antioxidant capacities and elevated ROS levels in long-term cultivated yMSCs as well as aMSCs. Notably, only the MSC migration potential and their antioxidative capacity were altered by <em>in vitro</em> as well as chronological aging. Based on specific differences observed between the impact of chronological and <em>in vitro</em> MSC aging we conclude that both are distinct processes.</p> </div

Long-term cultivation of MSCs alters their mitochondrial function.

<p>(<b>A</b>): Fluorescence microscopy was used to investigate the morphology of the mitochondrial network within long-term cultivated and primary aMSCs and yMSCs. Upon <i>in vitro</i> aging mitochondrial network appeared to be altered. Images show immunofluorescence of mitochondria and the actin cytoskeleton stained with a specific antibody recognizing cytochrome C and Alexa 594-conjugated phalloidin, respectively. Nuclei were counterstained with DAPI. (<b>B</b>): During <i>in vitro</i> aging the relative mitochondrial area per cell area increases in aMSCs and yMSCs of passage P30 and P100 compared to P2. The mitochondrial network and the cellular area were quantified after staining with MitoTracker™ Red and phalloidin, respectively. Diagram values represent ratio of the mitochondria network area relative to the cell area. (<b>C</b>): The total antioxidant capacity decreases with increasing passage number. Moreover, yMSCs of P2 and P100 exhibited significant higher antioxidant activities than aMSCs of the same passage. The Trolox® equivalent antioxidant assay kit was used to determine the total antioxidant capacity of whole MSC lysates and quantified against a Trolox® standard row. (<b>D</b>): Intracellular ATP levels decline significantly in long-term cultivated aMSCs and yMSCs of passage P30 and P100. Cellular ATP was determined using ATPLite™ bioluminescence luciferase-based assay and normalized to total DNA content determined by CyQuant®. (<b>E</b>): Long-term cultivated yMSCs and aMSCs of P30 and P100 displayed higher ROS production than primary MSCs of P2. After treatment with pyocyanin, which increases ROS levels, the observed difference between P2 and P30/P100 remained but the absolute value amplified about 2-fold. Intracellular ROS level were determined using CM-H2-DCFDA and normalized to total DNA content. (<b>F</b>): Measurement of the mitochondrial membrane potential (ΔΨm) revealed a progressive increase during <i>in vitro</i> aging with highest values in aMSCs of P100. Upon treatment with valomycin, an inhibitor of the mitochondrial respiratory chain, ΔΨm declined in aMSCs and yMSCs of all passages. The mitochondrial ΔΨm was determined with the MitoProbe® JC-1. * indicates statistical significance (p<0.05).</p

Long-term cultivation negatively influences the differentiation and migration potential of aMSCs and yMSCs.

<p>(<b>A</b>): In contrast to primary MSCs of passage 2, <i>in vitro</i> aged aMSCs and yMSCs of P30 and P100 show no matrix mineralization. Osteogenic differentiation was initiated with dexamethason and determined by matrix mineralization (Alizarin Red, AR) and normalized to cell number (alamarBlue®, AB). Dashed lines indicate differentiation potential of the negative control cultured in EM. (<b>B</b>): Under stimulation with BMP2, aMSCs and yMSCs of P2 show strong osteogenic differentiation, while again no matrix mineralization was observed in long-term MSC cultures of P30 and P100. (<b>C</b>): Adipogenic differentiation of aMSCs and yMSCs of P30 and P100, induced by adipogenic medium, was diminished by 50% compared to aMSCs and yMSCs of P2. In reference to the negative control maintained in EM (dashed line), aMSCs and yMSCs of P30 and P100 retained a potential for adipogenic differentiation. Differentiation was determined by using Oil red O (OR) staining and normalized to cell number. Diagram shows values normalized to negative control. (<b>D</b>): The number of migrated cells declined with increased <i>in vitro</i> passage. Moreover, aMSCs of each passage demonstrated significantly lower migratory potential compared to yMSCs. Migration rates were measured with a modified Boyden chamber assay. At least five independent experiments were carried out for all assays. Abbreviations: OD, optical density. * indicates statistical significance (p<0.05).</p

Transcriptional profiling of aMSCs and yMSCs at P2, P30 and P100.

<p>(<b>A</b>): The absolute number of genes detected after thresholding diminished during advanced <i>in vitro</i> culture independent from donor age (second column). The correlation coefficient (r²) was significantly reduced between aMSCs and yMSCs of P30 and P100 compared to P2. Only minor differences in gene expression were detected between aMSCs and yMSCs of each passage. (<b>B</b>): Functional annotation clustering of genes exclusively expressed either in primary MSC of P2 or <i>in vitro</i> aged MSCs of P30 and P100 revealed 431 and 124 differentially regulated genes, respectively. At P2 genes were mainly associated with chemokine signaling, apoptosis, cell migration, and calcium homeostasis. Whereas at P30 and P100 exclusively expressed genes are involved in Notch signaling, cell cycle progression and receptor signaling. (<b>C</b>): Analysis of pathways down-regulated after long-term <i>in vitro</i> culture revealed involvement of mitochondria, focal adhesions, cytoskeleton organization, TGF-β/BMP, WNT, and PPARγ signaling. Pathways up-regulated upon long-term <i>in vitro</i> culture were associated with cell cycle progression, DNA replication, p53, MAPK, and insulin signaling. (<b>D</b>): Differential statistical analysis summarizes all pathways and genes significantly up- and down-regulated during <i>in vitro</i> culture. The most numerous genes down-regulated during <i>in vitro</i> aging of aMSCs and yMSCs were associated with focal adhesions, actin cytoskeleton organization and mitochondrial function.</p

Generation and characterization of <i>in vitro</i> aged MSCs.

<p>(<b>A</b>): Cumulative population doublings of aMSCs and yMSCs during the first 80 days of culture are shown (n = 5). (<b>B</b>): Long-term cultivation has no influence on short-term proliferation rate of aMSCs and yMSCs of passage 30 and 100. Proliferation assay was performed using CyQuant®. (<b>C</b>): Graphs illustrate quantified signal intensities of p21^WAF1/CIP1 and p16^INK4A relative to GAPDH. (<b>D</b>): Representative Western blots showing increased p21^WAF1/CIP1 and p16^INK4A expression during <i>in vitro</i> aging. GAPDH served as endogenous control. (<b>E</b>): In anchorage-independent growth assays <i>in vitro</i> aged MSCs_P100 did not form colonies, while the breast carcinoma cell line MDA-MB-231, which served as positive control, produced numerous colonies (n = 3). Abbreviations: aMSCs, mesenchymal stromal cells from aged donors; yMSCs, mesenchymal stromal cells from young donors; P: passage. * indicates statistical significance (p<0.05).</p

Long-term <i>in vitro</i> culture alters MSC morphology independent from the donor age.

<p>(<b>A</b>): Cell diameter of aMSCs and yMSCs decreases during the course of long-term cultivation. Diagram shows the cell size distribution of MSCs measured by CASY® TT cell analyzer system at indicated passages after trypsinization. (<b>B</b>): Cellular area of attached aMSCs and yMSCs significantly decreases during <i>in vitro</i> aging. Measurements were performed from fluorescence images of identical exposure conditions. (<b>C</b>): Representative images of phalloidin labeled MSCs highlight reduction of cellular expansion. Additionally, <i>in vitro</i> aged aMSCs and yMSCs exhibited less filopodia, lamellipodia and cell spreading (white arrows). * indicates statistical significance (p<0.05).</p