Early microvascular coronary endothelial dysfunction precedes pembrolizumab-induced cardiotoxicity. Preventive role of high dose of atorvastatin.

Immune checkpoint inhibitors (ICIs) exhibit remarkable antitumor activity and immune-related cardiotoxicity of unknown pathomechanism. The aim of the study was to investigate the ICI class-dependent cardiotoxicity in vitro and pembrolizumab's (Pem's) cardiotoxicity in vivo, seeking for translational prevention means. Cytotoxicity was investigated in primary cardiomyocytes and splenocytes, incubated with ipilimumab, Pem and avelumab. Pem's cross-reactivity was assessed by circular dichroism (CD) on biotechnologically produced human and murine PD-1 and in silico. C57BL6/J male mice received IgG4 or Pem for 2 and 5 weeks. Echocardiography, histology, and molecular analyses were performed. Coronary blood flow velocity mapping and cardiac magnetic resonance imaging were conducted at 2 weeks. Human EA.hy926 endothelial cells were incubated with Pem-conditioned media from human mononuclear cells, in presence and absence of statins and viability and molecular signaling were assessed. Atorvastatin (20 mg/kg, daily) was administered in vivo, as prophylaxis. Only Pem exerted immune-related cytotoxicity in vitro. Pem's cross-reactivity with the murine PD-1 was confirmed by CD and docking. In vivo, Pem initiated coronary endothelial and diastolic dysfunction at 2 weeks and systolic dysfunction at 5 weeks. At 2 weeks, Pem induced ICAM-1 and iNOS expression and intracardiac leukocyte infiltration. At 5 weeks, Pem exacerbated endothelial activation and triggered cardiac inflammation. Pem led to immune-related cytotoxicity in EA.hy926 cells, which was prevented by atorvastatin. Atorvastatin mitigated functional deficits, by inhibiting endothelial dysfunction in vivo. We established for the first time an in vivo model of Pem-induced cardiotoxicity. Coronary endothelial dysfunction precedes Pem-induced cardiotoxicity, whereas atorvastatin emerges as a novel prophylactic therapy.

A computer tablet software can quantify the deviation of scapula medial border from the thoracic wall during clinical assessment of scapula dyskinesis.

PURPOSE: Aim of this study is to establish an objective and easily applicable method that will allow clinicians to quantitatively assess scapular dyskinesis during clinical examination using a computer tablet software. Hypothesis is that dyskinetic scapulae present greater motion-deviation from the thoracic wall-compared to the non-dyskinetic ones and that the software will be able to record those differences. METHODS: Twenty-five patients and 19 healthy individuals were clinically evaluated for the presence of dyskinesis or not. According to the clinical diagnosis, the observations were divided into three groups; A. Dyskinetic scapulae with symptoms (n = 25), B. Contralateral non-dyskinetic scapulae without symptoms (n = 25), C. Healthy control scapulae (n = 38). Then, all individuals were tested using a tablet with the PIVOT™ image-based analysis software (PIVOT, Impellia, Pittsburgh, PA, USA). The motion produced by the scapula medial border and inferior angle deviation from the thoracic wall was recorded. RESULTS: The deviation of the medial border and inferior angle of the scapula from the thoracic wall was 24.6 ± 7.3 mm in Group A, 14.7 ± 4.9 mm in Group B, and 12.4 ± 5.2 mm in Group C. The motion recorded in the dyskinetic scapulae group was significantly greater than both the contralateral non-dyskinetic scapulae group (p < 0.01) and the healthy control scapulae group (p < 0.01). CONCLUSION: The PIVOT™ software was efficient to detect significant differences in the motion between dyskinetic and non-dyskinetic scapulae. This system can support the clinical diagnosis of dyskinesis with a numeric value, which not only contributes to scapula dyskinesis grading but also to the evaluation of the progress and efficacy of the applied treatment, thus providing a feedback to the clinician and the patient. LEVEL OF EVIDENCE: IV, laboratory study.

Consensus protein engineering on the thermostable histone-like bacterial protein HUs significantly improves stability and DNA binding affinity.

Consensus-based protein engineering strategy has been applied to various proteins and it can lead to the design of proteins with enhanced biological performance. Histone-like HUs comprise a protein family with sequence variety within a highly conserved 3D-fold. HU function includes compacting and regulating bacterial DNA in a wide range of biological conditions in bacteria. To explore the possible impact of consensus-based design in the thermodynamic stability of HU proteins, the approach was applied using a dataset of sequences derived from a group of 40 mesostable, thermostable, and hyperthermostable HUs. The consensus-derived HU protein was named HUBest, since it is expected to perform best. The synthetic HU gene was overexpressed in E. coli and the recombinant protein was purified. Subsequently, HUBest was characterized concerning its correct folding and thermodynamic stability, as well as its ability to interact with plasmid DNA. A substantial increase in HUBest stability at high temperatures is observed. HUBest has significantly improved biological performance at ambience temperature, presenting very low Kd values for binding plasmid DNA as indicated from the Gibbs energy profile of HUBest. This Kd may be associated to conformational changes leading to decreased thermodynamic stability and, therefore, higher flexibility at ambient temperature.

Screening of Heteroaromatic Scaffolds against Cystathionine Beta-Synthase Enables Identification of Substituted Pyrazolo[3,4-c]Pyridines as Potent and Selective Orthosteric Inhibitors.

Cystathionine ß-synthase (CBS) is a key enzyme in the production of the signaling molecule hydrogen sulfide, deregulation of which is known to contribute to a range of serious pathological states. Involvement of hydrogen sulfide in pathways of paramount importance for cellular homeostasis renders CBS a promising drug target. An in-house focused library of heteroaromatic compounds was screened for CBS modulators by the methylene blue assay and a pyrazolopyridine derivative with a promising CBS inhibitory potential was discovered. The compound activity was readily comparable to the most potent CBS inhibitor currently known, aminoacetic acid, while a promising specificity over the related cystathionine γ-lyase was identified. To rule out any possibility that the inhibitor may bind the enzyme regulatory domain due to its high structural similarity with cofactor s-adenosylmethionine, differential scanning fluorimetry was employed. A sub-scaffold search guided follow-up screening of related compounds, providing preliminary structure-activity relationships with respect to requisites for efficient CBS inhibition by this group of heterocycles. Subsequently, a hypothesis regarding the exact binding mode of the inhibitor was devised on the basis of the available structure-activity relationships (SAR) and a deep neural networks analysis and further supported by induced-fit docking calculations.

Restoring tibiofemoral alignment during ACL reconstruction results in better knee biomechanics.

PURPOSE: Anterior cruciate ligament (ACL) reconstruction (ACLR) aims to restore normal knee joint function, stability and biomechanics and in the long term avoid joint degeneration. The purpose of this study is to present the anatomic single bundle (SB) ACLR that emphasizes intraoperative correction of tibiofemoral subluxation that occurs after ACL injury. It was hypothesized that this technique leads to optimal outcomes and better restoration of pathological tibiofemoral joint movement that results from ACL deficiency (ACLD). METHODS: Thirteen men with unilateral ACLD were prospectively evaluated before and at a mean follow-up of 14.9 (SD = 1.8) months after anatomic SB ACLR with bone patellar tendon bone autograft. The anatomic ACLR replicated the native ACL attachment site anatomy and graft orientation. Emphasis was placed on intraoperative correction of tibiofemoral subluxation by reducing anterior tibial translation (ATT) and internal tibial rotation. Function was measured with IKDC, Lysholm and the Tegner activity scale, ATT was measured with the KT-1000 arthrometer and tibial rotation (TR) kinematics were measured with 3Dmotion analysis during a high-demand pivoting task. RESULTS: The results showed significantly higher TR of the ACL-deficient knee when compared to the intact knee prior to surgery (12.2° ± 3.7° and 10.7° ± 2.6° respectively, P = 0.014). Postoperatively, the ACLR knee showed significantly lower TR as compared to the ACL-deficient knee (9.6°±3.1°, P = 0.001) but no difference as compared to the control knee (n.s.). All functional scores were significantly improved and ATT was restored within normal values (P < 0.001). CONCLUSIONS: Intraoperative correction of tibiofemoral subluxation that results after ACL injury is an important step during anatomic SB ACLR. The intraoperative correction of tibiofemoral subluxation along with the replication of native ACL anatomy results in restoration of rotational kinematics of ACLD patients to normal levels that are comparable to the control knee. These results indicate that the reestablishment of tibiofemoral alignment during ACLR may be an important step that facilitates normal knee kinematics postoperatively. LEVEL OF EVIDENCE: Level II, prospective cohort study.

Peroneal electromechanical delay and fatigue in patients with chronic ankle instability.

PURPOSE: The purpose of this study was to investigate the effect of chronic ankle instability (CAI) on electromechanical delay times (EMD) before and after fatigue. Understanding the mechanisms that contribute to CAI is essential for the development of effective rehabilitation programmes. It was hypothesized that patients with CAI will demonstrate prolonged EMD times compared to healthy subjects and that fatigue will cause greater increases in EMD times in the CAI group. METHODS: Twenty-one male volunteers participated in the study providing data on 16 ankles with CAI and 26 with no history of ankle injury. EMD was measured on an isokinetic dynamometer. Measurements were taken with the ankle in neutral (0°) and at 30° of inversion. All subjects followed an isokinetic fatigue protocol until eversion torque fell below 50 % of initial torque for three consecutive repetitions. A 2 × 2 × 2 ANOVA was used to calculate the effect of ankle status (CAI vs. healthy), fatigue, angle (0° vs. 30°) and their interactions on EMD. RESULTS: Fatigue caused a significant increase on EMD [non-fatigued: 122(29)ms vs. fatigue 155(54)ms; p < 0.001]. EMD times were shorter at 30° of inversion compared to neutral [neutral: 145(39)ms vs. 30° of inversion: 132(40)ms, p = 0.015]. An interaction effect for ankle status and angle was found (p = 0.026) with CAI ankles demonstrating longer EMD [CAI: 156(45)ms vs. healthy: 133(40)ms] in neutral but not at 30° of inversion [CAI: 133(46)ms vs. 132(33)ms]. CONCLUSIONS: Patients with CAI had longer EMD times in neutral, but not when the ankle was placed in inversion. This suggests that rehabilitation programmes may be more effective when retraining occurs with the ankle in neutral position. It is likely that low EMD times prevent ankle acceleration at the beginning of the mechanism of injury, but they are less important when the ankle has already inverted at 30°. Both CAI and healthy subjects demonstrated longer EMD after fatigue, emphasizing the importance of proper conditioning in the prevention of delayed peroneal response and subsequent ankle injury. Improving resistance to fatigue of the peroneals may prove to be an effective prevention tool of ankle sprain recurrence in patients with CAI. LEVEL OF EVIDENCE: III.

Genome Instability and γH2AX.

γH2AX has emerged in the last 20 years as a central player in the DDR (DNA damage response), with specificity for DSBs (double-strand breaks). Upon the generation of DSBs, γ-phosphorylation extends along megabase-long domains in chromatin, both sides of the damage. The significance of this mechanism is of great importance; it depicts a biological amplification mechanism where one DSB induces the γ-phosphorylation of thousands of H2AX molecules along megabaselong domains of chromatin, that are adjusted to the sites of DSBs. A sequential recruitment of signal transduction factors that interact to each other and become activated to further amplify the signal that will travel to the cytoplasm take place on the γ-phosphorylated chromatin. γ-phosphorylation is an early event in the DSB damage response, induced in all phases of the cell cycle, and participates in both DSB repair pathways, the HR (homologous recombination) and NHEJ (non-homologous end joining). Today, numerous studies support the notion that γH2AX functions as a guardian of the genome by preventing misrepaired DSB that increase the mutation load of the cells and may further lead to genome instability and carcinogenesis.

Analysis of the complete genome sequence of the archaeon Pyrococcus chitonophagus DSM 10152 (formerly Thermococcus chitonophagus).

Here we analyze the first complete genome sequence of Pyrococcus chitonophagus. The archaeon was previously suggested to belong to the Thermococcus rather than the Pyrococcus genus. Whole genome phylogeny as well as whole proteome comparisons using all available complete genomes in Thermococcales clearly showed that the species belongs to the Pyrococcus genus. P. chitonophagus was originally isolated from a hydrothermal vent site and it has been described to effectively degrade chitin debris, and therefore is considered to play a major role in the sea water ecology and metabolic activity of microbial consortia within hot sea water ecosystems. Indeed, an obvious feature of the P. chitonophagus genome is that it carries proteins showing complementary activities for chitin degradation, i.e. endo- and exo-chitinase, diacetylchitobiose deacetylase and exo-ß-D glucosaminidase activities. This finding supports the hypothesis that compared to other Thermococcales species P. chitonophagus is adapted to chitin degradation.

Insights into the functionality and stability of designer cellulosomes at elevated temperatures.

Enzymatic breakdown of lignocellulose is a major limiting step in second generation biorefineries. Assembly of the necessary activities into designer cellulosomes increases the productivity of this step by enhancing enzyme synergy through the proximity effect. However, most cellulosomal components are obtained from mesophilic microorganisms, limiting the applications to temperatures up to 50 °C. We hypothesized that a scaffoldin, comprising modular components of mainly mesophilic origin, can function at higher temperatures when combined with thermophilic enzymes, and the resulting designer cellulosomes could be employed in higher temperature reactions. For this purpose, we used a tetravalent scaffoldin constituted of three cohesins of mesophilic origin as well as a cohesin and cellulose-binding module derived from the thermophilic bacterium Clostridium thermocellum. The scaffoldin was combined with four thermophilic enzymes from Geobacillus and Caldicellulosiruptor species, each fused with a dockerin whose specificity matched one of the cohesins. We initially verified that the biochemical properties and thermal stability of the resulting chimeric enzymes were not affected by the presence of the mesophilic dockerins. Then we examined the stability of the individual single-enzyme-scaffoldin complexes and the full tetravalent cellulosome showing that all complexes are stable and functional for at least 6 h at 60 °C. Finally, within this time frame and conditions, the full complex appeared over 50 % more efficient in the hydrolysis of corn stover compared to the free enzymes. Overall, the results support the utilization of scaffoldin components of mesophilic origin at relatively high temperatures and provide a framework for the production of designer cellulosomes suitable for high temperature biorefinery applications.

Effectiveness of aspiration in knee joint effusion management: a prospective randomized controlled study.

PURPOSE: Knee effusion is a common symptom in various knee disorders of both traumatic and non-traumatic aetiology. Although intra-articular aspiration is a widespread treatment approach, its beneficial effect has not been confirmed by a randomized controlled study. The purpose was to evaluate the effectiveness and safety of joint aspiration in acute knee effusion, in relation to traumatic or non-traumatic aetiology. METHODS: One hundred and sixty-seven consecutive patients with acute knee joint effusion were allocated in a randomized controlled fashion into two groups. In the first group, joint aspiration was performed, while in the second group, no aspiration was performed. Range of motion, pain relief, use of analgesics and oedema were evaluated post management. The Knee Society Score and the International Knee Documentation Committee Subjective Evaluation Form were also obtained. In addition, a subgroup analysis of our results in relation to the presence of trauma or not was performed. RESULTS: Aspiration exhibited a temporary improvement in all clinical parameters evaluated, especially in the post-traumatic effusion. However, this improvement lasted only for the first week, due to the early re-accumulation of the effusion. There was no difference between the different groups regarding the clinical outcome in neither trauma or non-trauma patients at the end of the follow-up period. Aspiration aided in earlier establishment of the diagnosis in the non-trauma cases of effusion. CONCLUSIONS: Aspiration resulted in only temporary improvement of the outcome in the treatment of traumatic or not traumatic knee effusion. Aspiration is suggested in effusions of unknown origin in order to establish the diagnosis and for immediate clinical relief. However, aspiration should be performed with consideration in the presence of trauma.

Management of proximal interphalangeal joint hyperextension injuries: a randomized controlled trial.

PURPOSE: To compare the effectiveness of buddy strapping and aluminum orthosis for treatment of proximal interphalangeal (PIP) joint hyperextension injuries. We also evaluated the effect of age on the outcome by comparing our results in adults and children. METHODS: One hundred twenty-one consecutive patients with a PIP joint hyperextension injury of the index, middle, ring, or little finger and without fracture were evaluated. Patients were randomly assigned into 2 groups. In the first group, treatment included buddy strapping of the injured finger to its neighbor noninjured finger for a week. In the second group, immobilization was secured with an aluminum extension blocking orthosis for a week in 15° flexion. Assessment of motion, edema, pain, and strength were performed weekly for the first month and then at 3, 6, and 12 months after injury. RESULTS: The patients treated with buddy strapping exhibited similar outcomes compared with those treated with aluminum orthoses. In patients with full recovery, buddy strapping allowed earlier recovery of motion and resolution of edema and pain compared with aluminum orthosis immobilization. Furthermore, PIP injuries appear to have better outcomes in children than in adults. CONCLUSIONS: Buddy strapping is easy and effective treatment for PIP joint hyperextension injuries. TYPE OF STUDY/LEVEL OF EVIDENCE: Therapeutic I.

Investigation of the Microbiome of Industrial PDO Sfela Cheese and Its Artisanal Variants Using 16S rDNA Amplicon Sequencing and Shotgun Metagenomics.

Sfela is a white brined Greek cheese of protected designation of origin (PDO) produced in the Peloponnese region from ovine, caprine milk, or a mixture of the two. Despite the PDO status of Sfela, very few studies have addressed its properties, including its microbiology. For this reason, we decided to investigate the microbiome of two PDO industrial Sfela cheese samples along with two non-PDO variants, namely Sfela touloumotiri and Xerosfeli. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), 16S rDNA amplicon sequencing and shotgun metagenomics analysis were used to identify the microbiome of these traditional cheeses. Cultured-based analysis showed that the most frequent species that could be isolated from Sfela cheese were Enterococcus faecium, Lactiplantibacillus plantarum, Levilactobacillus brevis, Pediococcus pentosaceus and Streptococcus thermophilus. Shotgun analysis suggested that in industrial Sfela 1, Str. thermophilus dominated, while industrial Sfela 2 contained high levels of Lactococcus lactis. The two artisanal samples, Sfela touloumotiri and Xerosfeli, were dominated by Tetragenococcus halophilus and Str. thermophilus, respectively. Debaryomyces hansenii was the only yeast species with abundance > 1% present exclusively in the Sfela touloumotiri sample. Identifying additional yeast species in the shotgun data was challenging, possibly due to their low abundance. Sfela cheese appears to contain a rather complex microbial ecosystem and thus needs to be further studied and understood. This might be crucial for improving and standardizing both its production and safety measures.

Evaluation with contrast-enhanced magnetic resonance imaging of the anterior cruciate ligament graft during its healing process: a two-year prospective study.

OBJECTIVE: The aim of this study was to evaluate, with contrast-enhanced-magnetic resonance imaging (MRI), the changing imaging appearance of an anterior cruciate ligament (ACL) graft during the revascularization phase by quantitatively assessing the morphological and signal intensity changes taking place at its cross-sectional surface over time. MATERIALS AND METHODS: Fifty patients underwent contrast-enhanced-MRI on the third postoperative day and at a mean of 6, 12, and 24 months time interval after surgery. Proton-density images were obtained to evaluate morphological and signal intensity characteristics. Oblique-axial T1-weighted images obtained before and after intravenous gadolinium administration were used for quantitative analysis. Enhancement index (EI: signal-to-noise quotient(after gadolinium)÷signal-to-noise quotient(before gadolinium)) and cross-sectional area (CSA) were calculated for two regions of interest: the transplanted graft and its surrounding hypervascular tissue, and at three distinct graft sites (intra-articular, intraosseous tibial tunnel, and intraosseous juxta screw sites). Comparisons of EI and CSA at every site and time interval were performed using analysis of variance. RESULTS: A variable MRI appearance of the graft during the different time intervals was attributed to the varying amount of the hypervascular tissue gradually surrounding the graft. Graft EI and peripheral tissue CSA progress in a parallel, time- and site-related pattern along the graft course. The initial heterogeneity with intermediate signal intensity at the intra-articular graft site reflected intense revascularization. A slower revascularization progress was noticed at the other two intraosseously enclosed sites. CONCLUSION: During the healing process the amount of revascularization tissue influences the MR imaging characteristics of the graft according to the examined site and the time interval after surgery. By 2 years postoperatively, revascularization completion coincides with the homogeneously low signal intensity of the graft, closely resembling native ACL.

Effects of knee bracing on tibial rotation during high loading activities in anterior cruciate ligament-reconstructed knees.

PURPOSE: To test whether knee bracing restores normal rotational knee kinematics in anterior cruciate ligament (ACL)-reconstructed knees during high-demand, athletic activities. METHODS: Twenty male patients who had undergone unilateral ACL reconstruction with a bone-patellar tendon-bone autograft were assessed in vivo. The mean time from surgery to data collection was 26 months (range, 25 to 28 months). An 8-camera optoelectronic system was used to collect kinematic data while each patient performed 2 demanding tasks: (1) immediate pivoting after descending from a stair and (2) immediate pivoting after landing from a platform. Each task was performed under 3 conditions for the reconstructed knee: (1) wearing a prophylactic brace (braced condition), (2) wearing a patellofemoral brace (sleeved condition), and (3) without a brace (non-braced condition). As a control group, patients with intact ACLs were tested without any bracing. This study protocol was identical to the protocol of a previous study that investigated the effect of bracing on ACL-deficient athletes. RESULTS: For both tasks, the range of motion of tibial rotation was significantly lower in the intact knee compared with all 3 conditions of the ACL-reconstructed knee (P ≤ .014). Placing a brace or a sleeve on the ACL-reconstructed knee resulted in lower rotation than the non-braced condition (P ≤ .022), whereas no significant differences were found between the sleeved and the braced conditions (P ≥ .110). CONCLUSIONS: Bracing limited the excessive tibial rotation in ACL-reconstructed knees during pivoting that occurs under high-demand activities. However, full restoration to normative values was not achieved. Thereby, braces have the potential to decrease rotational knee instability that still remains after ACL reconstruction.

The shape and the thickness of the anterior cruciate ligament along its length in relation to the posterior cruciate ligament: a cadaveric study.

PURPOSE: The purpose of this study was to evaluate the shape of the native anterior cruciate ligament (ACL) along its length in relation to the posterior cruciate ligament (PCL) and compare it with the size of the 3 commonly used autografts (bone-patellar tendon-bone [BPTB], single-bundle hamstring, and double-bundle hamstring). METHODS: With the knee in extension, we filled the intercondylar notch with paraffin, fixing the cruciate ligaments in their natural position, in 8 cadaveric specimens. The ACL-PCL tissue specimen, embedded in paraffin, was removed en bloc. Gross sections were prepared in the coronal plane and were evaluated histologically. The width, thickness, and cross-sectional area of both the ACL and PCL were determined. The dimensions of the semitendinosus tendon (ST), gracilis tendon (GT), and BPTB grafts were measured and compared with those of the native ACL. RESULTS: The PCL occupies the largest part of the intercondylar area, leaving only a small space for the ACL in knee extension. The ACL midsubstance has a width of 5 mm, resembling a band shape. Only before its tibial insertion does the ACL fan out and take the form of its tibial attachment. The BPTB graft has a thickness of 5.8 mm, whereas the ST and GT grafts have a thickness of 6.25 mm and 4.5 mm, respectively, and are comparable to the midsubstance of the ACL but undersized in the tibial insertion (P = .0016 for BPTB graft, P = .002 for ST graft, and P = .0003 for GT graft). A quadruple-looped ST-GT graft, with a diameter of 8 mm, is oversized in the midsubstance (P = .0002) but fits better in the tibial attachment. CONCLUSIONS: The ACL midsubstance has a width of 5 mm, resembling a band shape. Before its tibial insertion, the ACL fans out like a trumpet, taking the form of its wide tibial attachment. CLINICAL RELEVANCE: The dimensions of the native ACL have to be considered in graft selection for anatomic ACL reconstruction.

The effect of knee braces on tibial rotation in anterior cruciate ligament-deficient knees during high-demand athletic activities.

OBJECTIVE: To examine if bracing can restrict tibial rotation in anterior cruciate ligament (ACL)-deficient patients during high loading activities. DESIGN: Repeated measures. Kinematic data were collected with an 8-camera Vicon system while each patient performed 2 tasks that are known to cause increased rotational and translational loads on the knee: (1) descending from a stair and subsequent pivoting, and (2) landing from a platform and subsequent pivoting. The tasks were repeated under 3 brace conditions for the ACL-deficient knee: (1) wearing a prophylactic brace (braced condition), (2) wearing a patellofemoral brace (sleeved condition), and (3) without brace (unbraced condition). SETTING: Biomechanical laboratory study. PATIENTS: Twenty-one male subjects with a confirmed unilateral ACL rupture were assessed in vivo. MAIN OUTCOME MEASURES: Tibial internal rotation. Two repeated measures ANOVAs tested for differences in tibial internal rotation among the 3 conditions of the ACL-deficient knee and the unbraced condition of the intact knee. RESULTS: In both tasks, tibial rotation was significantly lower in the intact knee compared with all 3 conditions of the ACL-deficient knee (P ≤ 0.031). Bracing the ACL-deficient knee resulted in lower rotation than the unbraced (P ≤ 0.001) and sleeved (P ≤ 0.033) conditions. The sleeved condition resulted in lower tibial rotation in the drop landing and pivoting task compared with the unbraced condition (P = 0.019) but not in the stair descending and pivoting task (P = 0.256). CONCLUSIONS: Bracing decreased the excessive tibial rotation in ACL-deficient patients during high-demand activities but failed to fully restore normative values. If knee braces can enhance rotational knee stability in ACL-deficient patients, then they could possibly play an important role in preventing further knee pathology in such patients.

Lessons learned from the last 20 years of ACL-related in vivo-biomechanics research of the knee joint.

PURPOSE: Technological advances in recent years have allowed the easy and accurate assessment of knee motion during athletic activities. Subsequently, thousands of studies have been published that greatly improved our understanding of the aetiology, surgical reconstruction techniques and prevention of anterior cruciate ligament (ACL) injuries. The purpose of this review is to summarize the evidence from biomechanical studies on ACL-related research. METHODS: High-impact articles that enhanced understanding of ACL injury aetiology, rehabilitation, prevention and adaptations after reconstruction were selected. RESULTS: The importance of restoring internal tibial rotation after ACL reconstruction has emerged in several studies. Criteria-based, individualized rehabilitation protocols have replaced the traditional time-based protocols. Excessive knee valgus, poor trunk control, excessive quadriceps forces and leg asymmetries have been identified as potential high risk biomechanical factors for ACL tear. Injury prevention programmes have emerged as low cost and effective means of preventing ACL injuries, particularly in female athletes. CONCLUSION: As a result of biomechanical research, clinicians have a better understanding of ACL injury aetiology, prevention and rehabilitation. Athletes exhibiting neuromuscular deficits predisposing them to ACL injury can be identified and enrolled into prevention programmes. Clinicians should assess ACL-reconstructed patients for excessive internal tibial rotation that may lead to poor outcomes.

Hamstring-dominant strategy of the bone-patellar tendon-bone graft anterior cruciate ligament-reconstructed leg versus quadriceps-dominant strategy of the contralateral intact leg during high-intensity exercise in male athletes.

PURPOSE: The purpose of this study was to investigate the effect of anterior cruciate ligament (ACL) reconstruction on the quadriceps-dominant strategy as a parameter associated with the neuromuscular control of the knee joint. METHODS: In this study 14 competitive soccer players who had undergone ACL reconstruction with bone-patellar tendon-bone autograft and 14 healthy competitive soccer players performed two 10-minute treadmill runs, 1 at moderate intensity and 1 at high intensity. Electromyographic recordings were acquired by use of a telemetric system at the third, fifth, seventh, and tenth minute of the runs from the vastus lateralis and the biceps femoris bilaterally. The dependent variable examined was the peak electromyographic amplitude during the stance phase. Analyses of variance were used to examine significant main effects and interactions. RESULTS: Vastus lateralis electromyographic activity during high-intensity running increased for both the control leg and intact leg (F = 4.48, P < .01), whereas it remained unchanged for the reconstructed leg (P > .05). Biceps femoris electromyographic activity during high-intensity running increased for the reconstructed leg only compared with both the control leg (F = 3.03, P < .05) and intact leg (F = 3.36, P < .03). CONCLUSIONS: There is no presence of the quadriceps-dominant strategy in ACL-reconstructed athletes during moderate-intensity exercise. During high-intensity exercise, the intact contralateral leg develops the quadriceps-dominant strategy whereas the reconstructed leg does not. The reconstructed leg instead increases biceps femoris activity, developing a "hamstring-dominant" strategy, and this "asymmetry" may theoretically be in favor of the reconstructed knee. LEVEL OF EVIDENCE: Level III, retrospective comparative study.

Correlation between anterior cruciate ligament graft obliquity and tibial rotation during dynamic pivoting activities in patients with anatomic anterior cruciate ligament reconstruction: an in vivo examination.

PURPOSE: To investigate the effect of coronal- and sagittal-plane anterior cruciate ligament (ACL) graft obliquity on tibial rotation (TR) range of motion (ROM) during dynamic pivoting activities after ACL reconstruction with bone-patellar tendon-bone (BPTB) autograft. METHODS: We evaluated 19 ACL-reconstructed patients (mean age, 29 years; age range, 18 to 38 years; mean time interval postoperatively, 19.9 months) and 19 matched control subjects (mean age, 30.6 years; age range, 24 to 37 years) using motion analysis during (1) descending a stairway and pivoting and (2) landing from a jump and pivoting. Magnetic resonance imaging was used to measure the coronal and sagittal ACL graft angle. The dependent variables were TR ROM during pivoting and the side-to-side difference (SSD) in TR ROM between the reconstructed knee and the contralateral intact knee. RESULTS: TR ROM of the ACL-reconstructed knee was significantly increased compared with both the contralateral intact knee and the healthy control knee (P < .05). A significant positive correlation was observed between TR ROM and coronal ACL graft angle (r = 0.727, P = .0006 for descending and pivoting; r = 0.795, P = .0001 for landing and pivoting) as well as between SSD of TR ROM and coronal ACL graft angle (r = 0.789, P < .0001 for descending and pivoting; r = 0.799, P < .0001 for landing and pivoting). No correlation was found with the sagittal ACL graft angle. CONCLUSIONS: After ACL reconstruction with a BPTB graft, patients' knees showed higher TR values than their uninjured knees and the knees of uninjured control volunteers during dynamic pivoting activities. The findings of this study show that TR was better restored in ACL-reconstructed patients with a more oblique graft in the coronal plane. A similar relation was not observed for graft orientation in the sagittal plane. Although these data do not imply a cause-and-effect relation between the 2 variables, they may indicate that a more oblique placement of a single BPTB ACL graft in the coronal plane is correlated with better control of TR. LEVEL OF EVIDENCE: Level IV, case series.

RGD binding to integrin alphavbeta3 affects cell motility and adhesion in primary human breast cancer cultures.

Integrins mediate cell adhesion to the extracellular matrix. Integrin alphavbeta3 recognizes the RGD motif as a ligand-binding site and has been associated with high malignant potential in breast cancer cells, signaling the onset of widespread metastasis. In recent years, several antagonists of integrin alphavbeta3, including RGD peptides, have been used as potential anti-cancer agents. In the present work, the effect of the linear RGD hexapeptide GRGDSP was studied, for the first time, on breast tumor explants, as well as on well-spread human breast cancer cells from primary cultures, using the explant technique, to clarify the role of this peptide in the suppression of breast cancer cell migration. The results showed that incubation of breast tumor explants with RGD peptide at the beginning of culture development inhibited completely the migration of cancer cells out of the tissue fragment as revealed by electron microscopy. RGD incubation of well-spread breast cancer cells from primary culture resulted in rounding and shrinkage of the cells accompanied by altered distribution of integrin alphavbeta3 and concomitant F-actin cytoskeletal disorganization, as revealed by immunofluorescence. Electron immunocytochemistry showed aggregation of integrin alphavbeta3 at the cell periphery and its detection in noncoated vesicles. However, Western immunoblotting showed no change in beta3 subunit expression, despite the altered distribution of the integrin alphavbeta3. In light of the above, it appears that the RGD peptide plays an important role in the modulation of cell motility and in the perturbation of cell attachment affecting the malignant potential of breast cancer cells in primary cultures.