Effect of testosterone on the female anterior cruciate ligament

Injuries to the anterior cruciate ligament (ACL) result in immediate and long-term morbidity and expense. Young women are more likely to sustain ACL injuries than men who participate in similar athletic and military activities. Although significant attention has focused on the role that female sex hormones may play in this disparity, it is still unclear whether the female ACL also responds to androgens. The purpose of this study was to determine whether the female ACL was an androgen-responsive tissue. To identify and localize androgen receptors in the female ACL, we used Western blotting and immunofluorescent labeling, respectively, of ACL tissue harvested during surgery from young women (n = 3). We then measured ACL stiffness and assessed total testosterone (T) and free [free androgen index (FAI)] testosterone concentrations, as well as relative estradiol to testosterone ratios (E(2)/T and E(2)/FAI) at three consecutive menstrual stages (n = 20). There were significant rank-order correlations between T (0.48, P = 0.031), FAI (0.44, P = 0.053), E(2)/T (-0.71, P < 0.001), E(2)/FAI (-0.63, P = 0.003), and ACL stiffness near ovulation. With the influences of the other variables controlled, there were significant negative partial rank-order correlations between ACL stiffness and E(2)/T (-0.72, P < 0.001) and E(2)/FAI (-0.59, P = 0.012). The partial order residuals for T and FAI were not significant. These findings suggest that the female ACL is an androgen-responsive tissue but that T and FAI are not independent predictors of ACL stiffness near ovulation. Instead, the relationship between T, FAI, and ACL stiffness was likely influenced by another hormone or sex hormone binding globulin.     

Effect of knee musculature on anterior cruciate ligament strain in vivo

Squatting is a commonly prescribed exercise following reconstruction of the anterior cruciate ligament (ACL). The objective of this paper was to measure the in vivo strain patterns of the normal ACL and the load at the knee for the simple squat and for squatting with a “sport cord”. A sport cord is a large elastic rubber tube used for added resistance. Strain patterns were deduced using displacement data from a Hall Effect Strain Transducer (HEST), while joint loads were determined by a mathematical model with inputs from a force plate and electrogoniometers. ACL strain for the free squat in one subject had a maximum of 2% at a knee angle of 10° and was slack for knee angles >17°. In squatting with a sport cord, peak strain was 1% at 10° and was slack at knee angles >14°. Since these peak strains are low, squatting appears to be a safe exercise for conservative rehabilitation of ACL reconstruction patients. In addition, the sport cord is a recommended augmentation to the activity. We believe that the decrease in strain with the sport cord results from added joint stiffness due to greater compressive forces at the tibiofemoral joint. This greater compressive force results from the approximately 10% increase in quadriceps activity. From shear force data predicted by the mathematical model, the maximum anterior drawer force for free squatting (50 N) was considerably less than for sport cord squatting (430 N). Therefore, the value of shear force at the tibiofemoral joint only partially determines the load placed on the ACL.     

Results of the surgical reconstruction of the anterior cruciate ligament

Results of the surgical reconstruction of the anterior cruciate ligament (ACL), using as a graft fourfold hamstring tendons (gracilis and semitendinosus) and middle third of the patellar ligament, were compared. In all patients that were participating in this study clinical examination and magnetic resonance showed ACL rupture, and apart from the choice of the graft, surgical technique was identical. We evaluated 112 patients with implemented patellar ligament graft and fourfold hamstring tendons graft six months after the procedure. Both groups were similar according to age, sex, activity level, knee instability level and rehabilitation program. The results showed that there was no significant difference between groups regarding Lysholm Knee score, IKDC 2000 score, activity level, musculature hypotrophy, and knee joint stability 6 months after the surgery. Anterior knee pain incidence is significantly higher in the group with patellar ligament graft (44% vs. 21%). Both groups had a significant musculature hypotrophy of the upper leg of the knee joint that was surgically treated, six months after the procedure. Both grafts showed good subjective and objective results.     

Side-to-side differences in anterior cruciate ligament volume in healthy control subjects

Examination of anterior cruciate ligament (ACL) anatomy is of great interest both in studying injury mechanisms and surgical reconstruction. However, after a typical acute ACL rupture it is not possible to measure the dimensions of the ACL itself due to concomitant or subsequent degeneration of the remaining ligamentous tissue. The contralateral ACL may be an appropriate surrogate for measuring anatomical dimensions, but it remains unknown whether side-to-side differences preclude using the contralateral as a valid surrogate for the ruptured ACL. This study examined whether the ACL volume is significantly different between the left and right knees of uninjured subjects. ACL volumes were calculated for the left and right sides of 28 individuals using a previously validated MRI-based method. The mean ACL volume was not significantly different (p=0.2331) between the two sides in this population. Side-to-side ACL volume was also well correlated (correlation=0.91, p<0.0001). The results of this study show that the volume of the contralateral ACL is a valid surrogate measure for a missing ACL on the injured side. This non-invasive, in vivo technique for measuring ACL volume may prove useful in future large-scale comprehensive studies of potential risk factors for ACL rupture, in quantifying potential loading effects on ACL size as a prophylactic measure against ACL rupture, and in the use of ACL volume as a screening tool for assessing risk of injury.     

Measurement of in vivo anterior cruciate ligament strain during dynamic jump landing

Despite recent attention in the literature, anterior cruciate ligament (ACL) injury mechanisms are controversial and incidence rates remain high. One explanation is limited data on in vivo ACL strain during high-risk, dynamic movements. The objective of this study was to quantify ACL strain during jump landing. Marker-based motion analysis techniques were integrated with fluoroscopic and magnetic resonance (MR) imaging techniques to measure dynamic ACL strain non-invasively. First, eight subjects' knees were imaged using MR. From these images, the cortical bone and ACL attachment sites of the tibia and femur were outlined to create 3D models. Subjects underwent motion analysis while jump landing using reflective markers placed directly on the skin around the knee. Next, biplanar fluoroscopic images were taken with the markers in place so that the relative positions of each marker to the underlying bone could be quantified. Numerical optimization allowed jumping kinematics to be superimposed on the knee model, thus reproducing the dynamic in vivo joint motion. ACL length, knee flexion, and ground reaction force were measured. During jump landing, average ACL strain peaked 55±14 ms (mean and 95% confidence interval) prior to ground impact, when knee flexion angles were lowest. The peak ACL strain, measured relative to its length during MR imaging, was 12±7%. The observed trends were consistent with previously described neuromuscular patterns. Unrestricted by field of view or low sampling rate, this novel approach provides a means to measure kinematic patterns that elevate ACL strains and that provide new insights into ACL injury mechanisms.     

Forward-dynamics simulation of anterior cruciate ligament forces developed during isokinetic dynamometry

A mathematical (computer) model was developed and used to study the mechanics of the human knee during extension exercises employing an isokinetic dynamometer. All parts of the body were fixed to ground, except for the right shank and foot, which were free to move in the parasagittal plane. A linkage attached the dynamometer to the shank; tibiofemoral articulation consisted of single-point contact, allowing both sliding and rolling to occur. Physiologically based representations of ligaments and muscles imparted forces to the shank. A forward dynamics simulation was performed to calculate the forces developed in the knee for isokinetic speeds ranging from 0 (isometric exercise) to 300 degrees /s. Simulations were conducted for a constant-speed phase during isokinetic knee extension exercise. It was assumed for the duration of each simulated exercise that the quadriceps were fully activated and the other muscles were fully deactivated. The force in the anterior cruciate ligament was found to be governed by the force-velocity properties of the quadriceps; the model predicts that 300 deg/sec isokinetic exercise can reduce the force transmitted to the ACL by almost a factor of two compared with that present during isometric knee extension.     

An examination of possible quadriceps force at the time of anterior cruciate ligament injury during landing: A simulation study

Anterior cruciate ligament (ACL) rupture is a common and traumatic injury. Although, identifying the mechanism of ACL injury has received considerable research attention, there are still many unanswered questions. One proposed mechanism asserts that the ACL is injured due to an aggressive quadriceps muscle contraction. However, recently it has been questioned if the magnitude of quadriceps force needed to tear the ACL is physiologically realistic under the conditions where injury occurs during landing (e.g. near full knee extension and within 50ms after impact). To answer this question, a simple simulation model was developed to examine the upper bounds of quadriceps force that can be developed under these conditions. The model included force-length, and force-velocity properties as well as activation dynamics. Model parameters were chosen to provide a high estimate for possible quadriceps force in a young healthy man. The effects of varying quadriceps pre-activation levels were also examined. When using realistic pre-activation levels, the simulated quadriceps force was less than half of what has been shown to cause ACL injury. Even when using maximum pre-activation, the quadriceps force still did not reach close to the level shown to cause injury. Therefore, we conclude that quadriceps force alone seems to be an unlikely mechanism for ACL injury.     

Association between matrix metalloproteinase-3 polymorphism and anterior cruciate ligament ruptures

Anterior cruciate ligament (ACL) ruptures are considered to be the most severe joint injury in sports. However, the precise etiologies of ACL injuries are not fully understood. Recently, the gene encoding the matrix metalloproteinase-3 (MMP-3, stromelysin-1) was shown to be associated with anterior cruciate ligament ruptures. The 5A/6A polymorphism in the promoter of the MMP-3 gene affects the regulation of MMP-3 gene expression. We examined the association between polymorphism within -1612 of the MMP-3 gene and ACL rupture in an independent population. Eighty-six participants between 20 and 40 years of age with surgically diagnosed ACL ruptures and 100 healthy controls between 18 and 28 years of age without history of ligament or tendon injuries were recruited for the study. All participants were genotyped for the MMP-3 polymorphism (-1612 5A/6A). Statistical analyses of genotype frequencies between patients and healthy controls were performed by the chi-square test. A significant difference was found between ACL rupture subgroups in terms of genotype association (5A+ (5A/5A, 5A/6A): 37.5% in contact sports vs 20% in non-contact sports; P = 0.02). In allelic association, there were significant differences (6A: 81.2% in contact sports vs 89.1% in non-contact sports, 5A: 18.8% in contact sports vs 10.9% in non-contact sports, P = 0.01). The 5A+ genotype of MMP-3 was represented in ACL ruptures in contact sport participants. We propose that this sequence variant is a specific genetic element that should be included in a multifactorial model to understand the etiologies and risk factors for ACL rupture.     

Age-dependent expression of VEGF isoforms and receptors in the rabbit anterior cruciate ligament

Vascular endothelial growth factor (VEGF) gene gives rise to several distinct isoforms of VEGF. Those isoforms differ in biochemical and biological properties, and it has been reported that their expression patterns are tissue and age specific as well. We investigated the expression levels of VEGF isoforms (VEGF121, VEGF165, VEGF183, VEGF189) and its receptors (VEGFR-1, flt-1 and VEGFR-2, flk-1/KDR) in the anterior cruciate ligament (ACL) of 2- to 3-week-, 2-month-, and 18-month-old New Zealand White rabbits using Sybr green Real-Time RT-PCR. VEGF isoforms and both receptors were expressed in the ACL at all investigated ages. VEGF121 was found to be the most abundant isoform at the ages under investigation, followed by VEGF165, VEGF189 and VEGF183. All isoforms showed decreased expression levels with age, however the larger membrane bound isoforms, VEGF183 and VEGF189, showed the most striking age-associated decrease in expression level. VEGFR-1 expression levels increased with age, while the expression level of VEGFR-2 expression was highest at 2-3 weeks and was significantly lower at 2 and 18 months of age. Distinct age-associated differences in the expression level of VEGF isoforms as well as their receptors suggest differential physiological functions during development, maturation and ageing of the ACL.     

Quantitation of estrogen receptors and relaxin binding in human anterior cruciate ligament fibroblasts

Summary The significantly higher incidence of anterior cruciate ligament (ACL) injuries in collegiate women compared with men may result from relative ligament laxity. Differences in estrogen and relaxin activity, similar to that seen in pregnancy, may account for this. We quantified estrogen receptors by flow cytometry and relaxin receptors by radioligand binding assay in human ACL cells and compared the presence of these receptors in males and females. ACL stumps were harvested from seven males and eight females with acute ACL injuries. The tissue was placed in M199 cell culture medium. Outgrowth cultures were obtained, and passage 2 cells were used for all studies. Estrogen receptor determination was performed using flow cytometry. Relaxin binding was performed in ACL cells derived from five female and male patients using I¹²⁵-labeled relaxin. Estrogen receptors were identified by flow cytometry in 4 to 10% of ACL cells. Mean fluorescence of cells expressing estrogen receptors was approximately twice that of controls, with no significant differences between males and females. Relaxin studies showed low-level binding of I¹²⁵-relaxin-labeled ACL cells. Relaxin binding was present in four out of five female ACL cells versus one out of five male ACL cells.     

Sensitivity of insertion locations on length patterns of anterior cruciate ligament fibers

A technique is demonstrated, employing an instrumented spatial linkage, for the determination of the length patterns of discrete fiber bundles within a ligament under controlled loading conditions. The instrumented spatial linkage was used to measure the three-dimensional joint motion. The linkage was also used as a three-dimensional coordinate digitizer to determine the spatial location of bony landmarks and the ligament's insertion areas. The length of pseudo fiber bundles was determined as the straight line distance between bone attachments. A comparison is presented, showing good agreement, between elongation patterns obtained from this method and those measured using an instrumented fine wire cable fiber. A sensitivity analysis was performed to evaluate the influence of tibial and femoral attachment location on the length pattern of fiber bundles of the anterior cruciate ligament. It was found that the relationship between fiber elongation and knee flexion depended strongly on the fibers femoral attachment location but not on its tibial attachment location.     

The effect of weightbearing and external loading on anterior cruciate ligament strain

A force balance between the ligaments, articular contact, muscles and body weight maintains knee joint stability. Thus, it is important to study anterior cruciate ligament (ACL) biomechanics, in vivo, under weightbearing conditions. Our objective was to compare the ACL strain response under weightbearing and non-weightbearing conditions and in combination with three externally applied loadings: (1) anterior-posterior shear forces, (2) internal-external torques, and (3) varus-valgus moments. A strain transducer was implanted on the ACL of 11 subjects. All joint loadings were performed with the knee at 20 degrees of flexion. A significant increase in ACL strain was observed as the knee made the transition from non-weightbearing to weightbearing. During anterior shear loading, the strain values produced during weightbearing were greater than those of the non-weightbearing knee (shear loads <40N). At higher shear loads, the strain values became equal. During axial torsion, an internal torque of 10Nm strained the ACL when the knee was non-weightbearing while an equivalent external torque did not. Weightbearing significantly increased ACL strain values in comparison to non-weightbearing with the application of external torques and low internal torques (<3Nm). The strains became equal for higher internal torques. For V-V loading, the ACL was not strained in the non-weightbearing knee. However, weightbearing increased the ACL strain values over the range of moments tested. These data have important clinical ramifications in the development of rehabilitation protocols following ACL reconstruction since weightbearing has been previously thought to provide a protective mechanism to the healing graft.     

Muscle co-contraction after anterior cruciate ligament reconstruction: Influence of functional level

Background

ACL reconstruction is recommended to improve function in subjects with ligament injuries. However, after surgery, some individuals are not able to return to their pre-injury functional level. The mechanisms related to this incapacity are not well understood.

Study design

Cross-sectional study.

Methods

Co-contraction levels were assessed in individuals who returned to their pre-injury functional level and in 10 individuals who were not able to return to full activity after unilateral ACL reconstruction. Electromyography of the vastus lateralis and biceps femoris muscles before and after sudden perturbations applied during the stance phase of walking was used to calculate co-contraction.

Results

The involved limb had lower co-contraction pre-perturbation than the non-involved limb in both groups (p = 0.049). The co-contraction level post-perturbation was significantly higher in the limited return group than in the full return group (p = 0.03).

Conclusion

Decreased co-contraction in the involved limb before perturbation may be caused by sensorial changes resulting from surgery or injury. Increased co-contraction levels observed in the limited return group after perturbation may be a compensatory mechanism to make up for possible decreased intrinsic stability of the knee joint.

Clinical relevance

Increased co-contraction after perturbation does not contribute to knee stability.     

Tissue engineering of the anterior cruciate ligament using a braid-twist scaffold design

The anterior cruciate ligament (ACL) is the most commonly injured intra-articular ligament of the knee. The insufficient vascularization of this tissue prevents it from healing completely after extreme tearing or rupture, creating a need for ACL grafts for reconstruction. The limitations of existing grafts have motivated the investigation of tissue-engineered ACL grafts. A successful tissue-engineered graft must possess mechanical properties similar to the ACL; to date no commercially available synthetic graft has achieved this. To accomplish this goal we have combined the techniques of polymer fiber braiding and twisting to design a novel poly L-lactic acid (PLLA) braid-twist scaffold for ACL tissue engineering. The scaffold is designed to accurately mimic the biomechanical profile and mechanical properties of the ACL. In this study, braid-twist scaffolds were constructed and compared to braided scaffolds and twisted fiber scaffolds. The addition of fiber twisting to the braided scaffold resulted in a significant increase in the ultimate tensile strength, an increase in ultimate strain, and an increase in the length of the toe region in these constructs over scaffolds that were braided. Based on the findings of this study, the braid-twist scaffold studied was found to be a promising construct for tissue engineering of the ACL.     

Morphological characterization of a novel scaffold for anterior cruciate ligament tissue engineering

Tissue engineering offers an interesting alternative to current anterior cruciate ligament (ACL) surgeries. Indeed, a tissue-engineered solution could ideally overcome the long-term complications due to actual ACL reconstruction by being gradually replaced by biological tissue. Key requirements concerning the ideal scaffold for ligament tissue engineering are numerous and concern its mechanical properties, biochemical nature, and morphology. This study is aimed at predicting the morphology of a novel scaffold for ligament tissue engineering, based on multilayer braided biodegradable copoly(lactic acid-co-(e-caprolactone)) (PLCL) fibers The process used to create the scaffold is briefly presented, and the degradations of the material before and after the scaffold processing are compared. The process offers varying parameters, such as the number of layers in the scaffold, the pitch length of the braid, and the fibers' diameter. The prediction of the morphology in terms of pore size distribution and pores interconnectivity as a function of these parameters is performed numerically using an original method based on a virtual scaffold. The virtual scaffold geometry and the prediction of pore size distribution are evaluated by comparison with experimental results. The presented process permits creation of a tailorable scaffold for ligament tissue engineering using basic equipment and from minimum amounts of raw material. The virtual scaffold geometry closely mimics the geometry of real scaffolds, and the prediction of the pore size distribution is found to be in good accordance with measurements on real scaffolds. The scaffold offers an interconnected network of pores the sizes of which are adjustable by playing on the process parameters and are able to match the ideal pore size reported for tissue ingrowth. The adjustability of the presented scaffold could permit its application in both classical ACL reconstructions and anatomical double-bundle reconstructions. The precise knowledge of the scaffold morphology using the virtual scaffold will be useful to interpret the activity of cells once it will be seeded into the scaffold. An interesting perspective of the present work is to perform a similar study aiming at predicting the mechanical response of the scaffold according to the same process parameters, by implanting the virtual scaffold into a finite element algorithm.     

Lower-limb dominance as a possible etiologic factor in noncontact anterior cruciate ligament tears

The purpose of this study was to determine if lower-limb dominance is a potential etiologic factor in noncontact anterior cruciate ligament (ACL) tears. A multicenter retrospective case analysis was performed. In each of the participating centers, patients were questioned to confirm a noncontact ACL injury and to determine lower-limb dominance. Three hundred and two subjects (149 males and 153 females) who presented with unilateral noncontact ACL tears participated in the study. The relationships between limb dominance, side of injury, and gender were analyzed. There was no significant correlation between the side of injury and the dominant limb for kicking (p = 0.30). There was no significant gender effect of the relationship between side of injury and dominant limb (p = 0.36). When assessing gender types and side of ACL tears, females showed a strong trend toward tearing the left ACL more frequently than the right (p = 0.06). No such trend existed for males. The results of this study indicate that there is no significant relationship between lower-limb dominance and the likelihood of sustaining a noncontact ACL tear. However, the strong trend toward females tearing their left ACLs more often than their right ACLs warrants further investigation to determine what neuromuscular asymmetries may exist between the right and left lower limbs.     

An implantable transducer for measuring tension in an anterior cruciate ligament graft.

The goal of this study was to develop a new implantable transducer for measuring anterior cruciate ligament (ACL) graft tension postoperatively in patients who have undergone ACL reconstructive surgery. A unique approach was taken of integrating the transducer into a femoral fixation device. To devise a practical in vivo calibration protocol for the fixation device transducer (FDT), several hypotheses were investigated: (1) The use of a cable versus the actual graft as the means for applying load to the FDT during calibration has no significant effect on the accuracy of the FDT tension measurements; (2) the number of flexion angles at which the device is calibrated has no significant effect on the accuracy of the FDT measurements; (3) the friction between the graft and femoral tunnel has no significant effect on measurement accuracy. To provide data for testing these hypotheses, the FDT was first calibrated with both a cable and a graft over the full range of flexion. Then graft tension was measured simultaneously with both the FDT on the femoral side and load cells, which were connected to the graft on the tibial side, as five cadaver knees were loaded externally. Measurements were made with both standard and overdrilled tunnels. The error in the FDT tension measurements was the difference between the graft tension measured by the FDT and the load cells. Results of the statistical analyses showed that neither the means of applying the calibration load, the number of flexion angles used for calibration, nor the tunnel size had a significant effect on the accuracy of the FDT. Thus a cable may be used instead of the graft to transmit loads to the FDT during calibration, thus simplifying the procedure. Accurate calibration requires data from just three flexion angles of 0, 45, and 90 deg and a curve fit to obtain a calibration curve over a continuous range of flexion within the limits of this angle group. Since friction did not adversely affect the measurement accuracy of the FDT, the femoral tunnel can be drilled to match the diameter of the graft and does not need to be overdrilled. Following these procedures, the error in measuring graft tension with the FDT averages less than 10 percent relative to a full-scale load of 257 N.