Synovium and blood capillaries at the femoral attachment of anterior cruciate ligament are significantly abundant than those at the tibial attachment / Sinovial y capilares sanguíneos en la inserción femoral del ligamento cruzado anterior son significativamente más abundantes que los de la inserción tibial

SUMMARY: The anterior cruciate ligament (ACL) is a ligament that mainly controls the anterior and rotational mobility of the knee joint, and its surface is covered by a synovial membrane with large number of blood vessels. In general, nutritional supply to the ligament is from many capillaries in the adjacent synovium. However, statistical studies of the capillaries distributed to the ACL are insufficient. In this study, we examined cross-sectional histological images of the femoral attachment (femoral level), middle level of the tendon (middle level), and tibial attachment (tibial level) of the ACL and statistically analyzed blood capillary distribution among the three levels. The ACLs of 10 cadavers were divided into 5 equal sections, and 4mm-thick paraffin sections were made at the femoral level, middle level, and tibial level, and then hematoxylin-eosin (HE) staining were performed. The area of each transverse section was measured using Image-J 1.51n (U. S. National Institutes of Health, Bethesda, MD, USA). Fiber bundles of the ACL were relatively small and sparse in cross-sectional area at the femoral level and became larger and denser toward the tibial level. Many blood levels. The synovium at the attachment of ACL covered the surface of the fiber bundle and also penetrated deeply between the fiber bundles. In particular, the blood capillaries were densely distributed in the synovium at the femoral attachment rather than another two levels. Indeed, the number of capillaries were also most abundant in the femoral level. The cross-sectional ACL area at the femoral level is significantly small, however, the blood capillaries were most abundant. Therefore, when the ACL is injured, its reconstruction with preservation of the femoral ligamentous remnant may be clinically useful for remodeling of the grafted tendon.

El ligamento cruzado anterior (LCA) es un ligamento que controla principalmente la movilidad anterior y rotacional de la articulación de la rodilla, y su superficie está cubierta por una membrana sinovial con gran cantidad de vasos sanguíneos. En general, el suministro de nutrientes al ligamento proviene de muchos capilares en la sinovial adyacente. Sin embargo, los estudios estadísticos de los capilares distribuidos en el LCA son insuficientes. En este estudio, examinamos imágenes histológicas trans- versales de la inserción femoral (nivel femoral), el nivel medio del tendón (nivel medio) y la inserción tibial (nivel tibial) del LCA y analizamos estadísticamente la distribución de los capilares sanguíneos entre los tres niveles. Los LCA de 10 cadáveres se dividieron en 5 secciones iguales y se realizaron cortes en parafina de 4 µm de espesor a nivel femoral, medio y tibial, y luego se realizó tinción con hematoxilina-eosina (HE). El área de cada sección transversal se midió utilizando Image-J 1.51n (Institutos Nacionales de Salud de EE. UU., Bethesda, MD, EE. UU.). Los haces de fibras del LCA eran relativamente pequeños y escasos en el área de la sección transversal a nivel femoral y se hicieron más grandes y más densos hacia el nivel tibial. La membrana sinovial en la unión del LCA cubría la superficie del haz de fibras y también penetraba profundamente entre entre los haces de fibras. En particular, los capilares sanguíneos estaban densamente distribuidos en la unión femoral de la sinovial respecto a los otros dos niveles. De hecho, el número de capilares también fue más abundante a nivel femoral. El área transversal del LCA a nivel femoral era significativamente pequeña, sin embargo, los capilares sanguíneos fueron los más abundantes. Por lo tanto, cuando hay una lesión del LCA su reconstrucción con preservación del ligamento femoral remanente puede ser clínicamente útil para remodelar el tendón injertado.

Histological and cellular evaluation of anterior cruciate ligament.

BACKGROUND: We hypothesized that the torn anterior cruciate ligament (ACL) demonstrates a great healing response after initial trauma and has competent cells leading to the healing but differs in its response based on the type of tear and duration of injury. This study aimed to evaluate the histological and cellular responses to the injured ACL. METHODS: Fifty-two tissue samples from the ACL were harvested from patients undergoing arthroscopy. Detailed histological and cellular examinations were performed for ligament angiogenesis, fibrocytes, and synovial tissue infiltration. We compared the cellular response to injury in partially and completely ruptured ACLs. The duration of ACL injury and its response to cellular characteristics were also examined. Immunohistochemical studies using cluster of differentiation 34 (CD34) staining was used to evaluate endothelial cells and fibrocytes. RESULTS: We found a significantly higher density of synovial and ligament angiogenesis and fibrocytes at the torn end of ACL (Mann-Whitney, P < 0.050). Numerous fibrocytes were identified in complete ACL tears versus partial tears (Mann-Whitney = 0.020). Increased cellular proliferation was identified at the ruptured end of ACL remnant (Kruskal-Wallis, P < 0.050). The cellular proliferation of ruptured ACL decreased after 12 months. CONCLUSIONS: Based on our findings of the time-dependent decrease in the cellular response at the torn ends of the ACL, we recommend early intervention, preservation of the ACL remnant, and primary ACL repair or augmented reconstruction.

Fibrocartilaginous metaplasia and neovascularization of the anterior cruciate ligament in patients with osteoarthritis.

INTRODUCTION: The anterior cruciate ligament (ACL) prevents the anterior translocation and medial rotation of the tibia against the femur. It is typically composed of dense regular connective tissue (DRCT), small amount of loose connective tissue, little vasculature, and few nerve endings. The objective of the current study was to evaluate the details of histological changes in ACLs of patients with clinically diagnosed osteoarthritis (OA). MATERIALS AND METHODS: The ACLs of six patients undergoing total knee replacement because of OA (OA group) were compared with 16 normal ACLs from cadavers (control). The ACLs were analyzed for tissue composition and number of blood vessels across the full length and thickness of the ligament. Percentages for areas of DRCT, fibrocartilage, degenerative tissue, and vasculature were calculated. Tissue composition and relative number of blood vessels were compared between groups. RESULTS: The proportion of DRCT to non-DRCT was significantly smaller in the OA group than the control group (p < .001); non-DRCT included degenerative connective tissue and fibrocartilage. The number of blood vessels to area was greater in the OA group than the control group (p = .002). Six of control (37.5%) and five of OA ACLs (83%) showed areas of calcification. CONCLUSIONS: These results indicate that inflammatory processes contributing to OA in the knee cause changes in the composition of the ACL that lead to destruction of collagen bundles, increased vascularization, calcification, and formation of fibrocartilage-like tissue inside the ligament. These changes make ligament-retaining total knee arthroplasty a less beneficial option for knee repair.

Cruciate ligament healing and injury prevention in the age of regenerative medicine and technostress: homeostasis revisited.

PURPOSE: This clinical concepts paper discusses the essential elements of cruciate ligament recuperation, micro-trauma repair, and remodeling. METHODS: Cruciate ligament mechanobiology and tissue heterogeneity, anatomy and vascularity, and synovial membrane and fluid functions are discussed in relationship to deficiency-induced inflammatory responses, nervous and immune system function, recuperation, repair and remodeling, and modern threats to homeostasis. RESULTS: Cruciate ligament surgical procedures do not appreciate the vital linked functions of the central, peripheral, and autonomic nervous systems and immune system function on knee ligament injury recuperation, micro-trauma repair, and remodeling. Enhanced knowledge of these systems could provide innovative ways to decrease primary non-contact knee injury rates and improve outcomes following reconstruction or primary repair. CONCLUSIONS: Restoration of knee joint homeostasis is essential to cruciate ligament recuperation, micro-trauma repair, and remodeling. The nervous and immune systems are intricately involved in this process. Varying combinations of high-intensity training, under-recovery, technostress, and environmental pollutants (including noise) regularly expose many athletically active individuals to factors that abrogate the environment needed for cruciate ligament recuperation, micro-trauma repair, and remodeling. Current sports training practice, lifestyle psychobehaviors, and environmental factors combine to increase both primary non-contact knee injury risk and the nervous and immune system dysregulation that lead to poor sleep, increased anxiety, and poorly regulated hormone and cytokine levels. These factors may create a worst-case scenario leading to poor ligament recuperation, micro-trauma repair, and remodeling. Early recognition and modification of these factors may decrease knee ligament injury rates and improve cruciate ligament repair or reconstruction outcomes. LEVEL OF EVIDENCE: V.

Histologic assessment of ligament vascularity and synovitis in dogs with cranial cruciate ligament disease.

OBJECTIVE: To assess the relationship between histologic degeneration of cranial cruciate ligaments (CCLs) and severity of synovitis and ligament vascularity. SAMPLE: CCL and synovium from 59 stifle joints (53 dogs). PROCEDURES: CCL and synovium specimens were obtained from stifle joints of juvenile (15 joints; 12 dogs) and adult (25 joints; 22 dogs) dogs with intact CCLs and dogs with CCL rupture (rCCL; 19 joints; 19 dogs). Vascular density and degenerative changes of the CCL core region and severity of synovitis were semiquantitatively evaluated. Relationships were analyzed by use of a random effects model to account for correlated specimens. RESULTS: Mean ± SD modified Bonar scores (scale, 0 to 9) of adults (4.85 ± 0.44) and dogs with rCCL (5.69 ± 0.49) were significantly higher than scores of juveniles (1.13 ± 0.55). Vascularity scores (scale, 0 to 3) were significantly higher for juveniles (3.00 ± 0.24) than for adults (1.53 ± 0.27) and dogs with rCCL (0.78 ± 0.23). Synovitis scores were not significantly different among groups. There was a significant negative relationship between modified Bonar scores and vascularity scores for juveniles and adults and for adults and dogs with rCCL when controlling for age, but there was not a significant relationship between modified Bonar scores and synovitis scores. There was a significant relationship between modified Bonar scores and body weight of adults. CONCLUSIONS AND CLINICAL RELEVANCE: Poor blood supply to the core region could be an important underlying condition for spontaneous degeneration of the CCL in at-risk dogs.

The ACL injury response: A collagen-based analysis.

BACKGROUND: Anterior cruciate ligament (ACL) injuries do not effectively heal. Tendon graft tissue after reconstruction shows rapid tissue turnover and 'ligamentization.' It is unknown whether native torn ACL tissue undergoes significant collagen turnover after injury or is arrested by the intraarticular environment. It is also unknown whether injury mechanism or chronicity affect torn ligament tissue turnover. METHODS: Thirty-three mid-substance ACL biopsies were obtained during primary arthroscopic ACL reconstruction (n=31; nine contact injuries, 22 non-contact injuries, 22 males, 11 females; mean age 28.5 years; median injury to surgery time 12 weeks), or from cadavers as uninjured ACL (n=2). As a marker for collagen turnover, immature collagen cross-link content was determined by ninhydrin reagent assays. The immature cross-link content was assessed against injury mechanism, patient age, and injury to surgery time. Histochemical analysis was conducted on two uninjured ACL cadaveric controls, a four-week-old ACL tear, and a four-year-old ACL tear. RESULTS: Contact and non-contact groups were not demographically different with respect to sex, patient age, injury to surgery time, and activity involvement prior to injury, which ranged from basketball to logging. Collagen crosslink content was very low across all samples, suggesting high tissue turnover between injury and surgery regardless of injury mechanism (non-contact: 1.68ng/mol, CI 0.48-2.89; contact: 1.50ng/mol, CI 0.14-2.86; p=0.842). CONCLUSION: Collagen turnover occurs rapidly after ACL injury regardless of contact or non-contact mechanism. Robust tissue turnover starts within the first several weeks after injury and persists to some extent throughout the life of the torn ACL.

Permanent knee sensorimotor system changes following ACL injury and surgery.

The cruciate ligaments are components of the knee capsuloligamentous system providing vital neurosensory and biomechanical function. Since most historical primary ACL repair attempts were unsuccessful, reconstruction has become the preferred surgery. However, an increased understanding of the efficacy of lesion-site scaffolding, innovative suturing methods and materials, and evolving use of biological healing mediators such as platelet-rich plasma and stem cells has prompted reconsideration of what was once believed to be impossible. A growing number of in vivo animal studies and prospective clinical studies are providing increasing support for this intervention. The significance of ACL repair rather than reconstruction is that it more likely preserves the native neurosensory system, entheses, and ACL footprints. Tissue preservation combined with restored biomechanical function increases the likelihood for premorbid neuromuscular control system and dynamic knee stability recovery. This recovery should increase the potential for more patients to safely return to sports at their desired intensity and frequency. This current concepts paper revisits cruciate ligament neurosensory and neurovascular anatomy from the perspective of knee capsuloligamentous system function. Peripheral and central nerve pathways and central cortical representation mapping are also discussed. Surgical restoration of a more physiologically sound knee joint may be essential to solving the osteoarthritis dilemma. Innovative rehabilitative strategies and outcome measurement methodologies using more holistic and clinically relevant measurements that closely link biomechanical and neurosensory characteristics of physiological ACL function are discussed. Greater consideration of task-specific patient physical function and psychobehavioral links should better delineate the true efficacy of all ACL surgical and non-surgical interventions. Level of evidence IV.

Reduced Anterior Cruciate Ligament Vascularization Is Associated With Chondral Knee Lesions.

This study tested the association between periligamentous vascularization of the anterior cruciate ligament (ACL) and the presence of chondral knee lesions via retrospective analysis of prospectively collected data from 702 consecutive knee arthroscopic procedures. In each case, the ACL periligamentous envelope was documented as follows: (1) vascular, where the ACL was covered with blood vessels along its entire length; (2) centrally avascular, where the central third of the ACL was not covered but peripheral vascularized coverage was present; and (3) avascular, where there was no blood vessel coverage of the ACL. Inclusion criteria for the study were as follows: (1) age older than 18 years and (2) normal knee ligament laxity. Univariate analysis and multiple logistic regression were used to test the association between chondral lesions and each of the variables: sex, age, meniscus tear, decreased ACL vascularity, and concomitant chondral lesion in another knee compartment. The cohort included 516 knees. In the univariate analysis, all variables were associated with a chondral lesion, but only older age and decreased ACL vascularity were associated with chondral lesions in each knee compartment. In the regression model, only decreased ACL vascularity was associated with chondral lesions in each knee compartment. For avascular knees, the odds ratio was 2.84 for medial femoral condyle lesions (95% confidence interval, 1.73-4.68; P=.000), 2.44 for lateral femoral condyle lesions (95% confidence interval, 1.19-5.03; P=.015), and 2.48 for patellofemoral lesions (95% confidence interval, 1.55-3.97; P=.000). The findings showed that decreased ACL periligamentous vascularization is associated with chondral lesions of the femoral condyles in knees with preserved ACL laxity. [Orthopedics. 2016; 39(4):e737-e743.].

Healing Potential of the Anterior Cruciate Ligament Remnant Stump.

AIM: The aim of this study was to analyze the microstructural architecture and cellular differentiation of the anterior cruciate ligament (ACL) stumps in different stages after injury, as this could augment graft biointegration. MATERIALS AND METHODS: The histological appearance and immunoreaction for cluster of differentiation 34 antigen (CD34) of 54 biopsies from 27 remnants were compared to 10 biopsies from 5 normal cruciate ligaments. RESULTS: CD34 reaction in endothelial cells, fibroblasts and fibrocytes was consistently positive in small synovial vessels. Remnants also exhibited CD34(+) cells among collagen fibers. Blood vessel density varied between specimens. The mean vascular microdensity was 43 per ×200 field in remnants compared to 15.2 in controls. A total of 94.44% of remnant ACL samples had significant hyperplasia of stellate and fusiform stromal cells, CD34(+); 22.4% had developed capillary vessels inside the ligament; 33% exhibited ongoing angiogenesis. CONCLUSION: Significant differences exist between torn and intact ACL regarding microvascularization. The remnants contain stellate stromal cells and CD34(+) fibrocytes, and display angiogenesis both at synovia as well as in the ligament itself. These findings underline the potential contribution to neoligament healing when remnants are preserved.

Central defect type partial ACL injury model on goat knees: the effect of infrapatellar fat pad excision.

BACKGROUND: The mid-substance central defect injury has been used to investigate the primary healing capacity of the anterior cruciate ligament (ACL) in a goat model. The sagittal plane stability on this model has not been confirmed, and possible effects of fat pad excision on healing have not been evaluated. We hypothesize that excising the fat pad tissue results in poorer ligament healing as assessed histologically and decreased tensile strength of the healing ligament. We further hypothesize that the creation of a central defect does not affect sagittal plane knee stability. METHODS: A mid-substance central defect was created with a 4-mm arthroscopic punch in the ACLs of right knees of all the subjects through a medial mini-arthrotomy. Goats were assigned to groups based on whether the fat pad was preserved (group 1, n = 5) or excised completely (group 2, n = 5). The left knees served as controls in each goat. Histopathology of the defect area along with measurement of type I collagen in one goat from each group were performed at 10th week postoperatively. The remaining knees were evaluated biomechanically at the 12th week, by measuring anterior tibial translation (ATT) of the knee joints at 90° of flexion and testing tensile properties (ultimate tensile load (UTL), ultimate elongation (UE), stiffness (S), failure mode (FM)) of the femur-ACL-tibia complex. RESULTS AND DISCUSSION: Histopathology analysis revealed that the central defect area was fully filled macroscopically and microscopically. However, myxoid degeneration and fibrosis were observed in group 2 and increased collagen type I content was noted in group 2. There were no significant differences within and between groups in terms of ATT values (p = 0.715 and p = 0.149, respectively). There were no significance between or within groups in terms of ultimate tensile load and ultimate elongation; however, group 2 demonstrated greater stiffness than group 1 that was correlated with the fibrotic changes detected microscopically (p = 0.043). CONCLUSIONS: The central defect type injury model was confirmed to be biomechanically stable in a goat model. Resection of the fat pad was noted to negatively affect defect healing and increase ligament stiffness in the central defect injury model.

The effect of blocking angiogenesis on anterior cruciate ligament healing following stem cell transplantation.

Ruptured human anterior cruciate ligaments (ACL) contain vascular stem cells capable of enhancing the healing of tendon grafts. In the current study we explored the role that neo-angiogenesis plays in ACL healing. ACL-derived CD34+ cells were isolated via Fluorescence Activated Cell Sorting (FACS) from the rupture sites of human ACLs. The cells were then virally transduced to express either vascular endothelial growth factor (VEGF) or soluble FLT-1 (sFLT-1), which is an antagonist of VEGF. We established five groups: CD34+VEGF(100%), where 100% of the cells were transduced with VEGF, CD34+VEGF(25%), where only 25% of the cells were transduced with VEGF, CD34+, CD34+sFLT-1, and a No cells group. The CD34+sFLT1 group had a significant reduction in biomechanical strength compared to the CD34+ group at 4 and 8 weeks; whereas the biomechanical strength of the CD34+VEGF(25%) group was significantly greater than the CD34+ group at week 4; however, no difference was observed by week 8. Immunohistochemical staining demonstrated a significantly lower number of isolectin B4 and hCD31 positive cells, markers associated with angiogenesis, in the CD34+sFLT1 group, and a higher number of isolectin B4 and hCD31 positive cells in the CD34+VEGF(100%) and CD34+VEGF(25%) groups compared to the CD34+ group. Graft maturation was significantly delayed in the CD34+sFLT1 group and accelerated in the CD34+VEGF(25%) group compared to the CD34+ group. In conclusion, blocking VEGF reduced angiogenesis, graft maturation and biomechanical strength following ACL reconstruction. Native expression of VEGF by the CD34+ cells improved tendon graft maturation and biomechanical strength; however, over-expression of VEGF impeded improvements in biomechanical strength.

A Cadaveric Study of the Branching Pattern and Diameter of the Genicular Arteries: A Focus on the Middle Genicular Artery.

The aim of the study was to gain an appreciation of the variation in the branching pattern and diameter of the genicular arteries arising directly from the popliteal artery (PA), namely, the superior medial genicular artery (SMGA), superior lateral genicular artery (SLGA), inferior medial genicular artery (IMGA), inferior lateral genicular artery (ILGA), and middle genicular artery (MGA). Twenty cadaveric knees aged between 62 and 92 years were dissected. A posterior midline vertical incision was used to gain access to the PA. The diameter of the PA, the sequence of branching, and subsequent diameter of genicular vessels and common trunks were recorded. PA average diameter was 7.9 mm. The SMGA (1.6-mm diameter) was the first branch in 45% and the second branch in 20%, and in seven limbs it arose from a common trunk with the SLGA. The SLGA was the second branch in 30% and the first branch in 25%, and it branched from a common trunk in 45%. The MGA (1.1-mm diameter) arose as the second branch in four knees and as the third branch in eight knees. It arose from a common trunk in eight knees, either with a superior genicular (three knees) or with an inferior genicular (five knees). The IMGA (1.5-mm diameter) was the third branch in 25%, the fourth branch in 35%, and the fifth branch in 15%. In five cases, it arose from a common trunk (25%). The ILGA (1.4-mm diameter) was the third branch in 15%, the fourth branch in 30%, and the fifth branch in 25%. It arose from a common trunk in six knees. These trunks also gave rise to the IMGA in all cases. There is extensive variation in the branching pattern and diameter of the genicular arteries differing from textbook descriptions. An awareness of vascular variation is imperative for preservation of the blood supply to the knee, which may promote recovery after anterior cruciate ligament reconstruction and popliteal aneurysm repair.

Vascular structure and function in the medial collateral ligament of anterior cruciate ligament transected rabbit knees.

To determine if decreased vascular responsiveness in the medial collateral ligament (MCL) of anterior cruciate ligament transected (ACL-t) rabbit knees is due to pericyte deficiency associated with angiogenesis. Vascular responses to potassium chloride (KCl), phenylephrine, acetylcholine, and sodium nitroprusside (SNP) were evaluated in ACL-t rabbit knees (n = 6) and control knees (n = 5) using laser speckle perfusion imaging. Ligament degeneration was determined by ultrasound imaging. Vascular and pericyte volume were measured using quantitative immunohistochemical volumetric analysis using CD31 and α-smooth muscle actin antibodies with co-localization analysis. Perfusion was increased in the ACL-t rabbits 2.5-fold. Responsiveness to phenylephrine, SNP, and acetylcholine was significantly decreased in the ACL knee while no change in KCl responses was seen. MCL ultrasound imaging revealed decreased collagen organization, increased ligament thickness, and increased water content in the ACL-t MCL. Vascular Volume was increased fourfold in ACL deficient knees, while pericyte volume to endothelial volume was not changed. No difference in CD31 and α-SMA co-localization was found. Blood vessels in the MCL of ACL-t knees do not lack smooth muscle. The MCL vasculature can undergo constrictive response to KCl, but have impaired receptor mediated responses and impaired nitric oxide signaling.

Immunohistochemical study on androgen receptors in the anterior cruciate ligament in dogs.

Androgen is closely involved as the cause of rupture of anterior cruciate ligament (ACL) in human. In dogs, however, factors contributing to rupture of ACL remain unknown. In this study, expression of androgen receptor (AR) and histological distribution of blood vessels in ACL, and serum testosterone concentration were investigated in relation with age and sex to confirm whether canine ACL is an androgen-responsive tissue. Materials of ACL were obtained from 26 dogs: 12 young female Beagles, 2 old female mixed breeds, 9 young male Beagles, and 3 old male mixed breeds. In all canine ACL, positive AR expression was recognized in the nuclei of the fibrocytes, fibroblasts, synovial cells, and vascular endothelial cells of ACL. Expressions of AR were lesser in old males compared to the young males; however, females had no age difference in expression. Distributions of blood vessels in the synovial membrane of the ligament were fewer in old dogs both of males and females than youngs. Although distributions of vessels in the interstitium were apparently fewer in young females. Serum testosterone concentration was significantly higher in young males. Females had no age difference in the levels. From these results, it is suggested that canine ACL is an androgen-responsive tissue, and this consideration seems to closely relate to the epidemiological background that the incidence of rupture of ACL of dogs is higher in females than in males.

Biological and biomechanical evaluation of the ligament advanced reinforcement system (LARS AC) in a sheep model of anterior cruciate ligament replacement: a 3-month and 12-month study.

PURPOSE: The purposes of this study were to assess tissue ingrowth within the Ligament Advanced Reinforcement System (LARS) artificial ligament (LARS AC; LARS, Arc sur Tille, France) and to study the biomechanical characteristics of the reconstructed knees in a sheep model of anterior cruciate ligament (ACL) replacement. METHODS: Twenty-five female sheep underwent excision of the proximal third of the left ACL and intra-articular joint stabilization with a 44-strand polyethylene terephthalate ligament (mean ultimate tensile failure load, 2,500 N). Animals were killed either 3 or 12 months after surgery. Explanted knees were processed for histology (n = 10) or mechanical tests including tests of laxity and loading to failure in tension (n = 15). RESULTS: Well-vascularized tissue ingrowth within the artificial ligament was only observed in the portions of the ligament in contact with the host's tissues (native ligament and bone tunnels). Ligament wear was observed in 40% of explanted knees. The ultimate tensile failure loads of the operated knees at both time points were inferior to those of the contralateral, intact knees (144 ± 69 N at 3 months and 260 ± 126 N at 12 months versus 1,241 ± 270 N and 1,218 ± 189 N, respectively) (P < .01). In specimens with intact artificial ligaments, failure occurred by slippage from the bone tunnels in all specimens explanted 3 months postoperatively and in half of the specimens explanted 12 months postoperatively. CONCLUSIONS: This study provides evidence that the LARS AC has a satisfactory biointegration but that it is not suitable for ACL replacement if uniform tissue ingrowth is contemplated. Despite good clinical performance up to 1 year after implantation, none of the reconstructions approached the mechanical performance of the normal ACL in the ovine model. Partial tearing of the artificial ligament, which led to a significant decrease in ultimate tensile strength, was observed in 40% of cases in the ovine model. CLINICAL RELEVANCE: The LARS is not a suitable scaffold for ACL replacement. Further animal studies are needed to evaluate its potential for augmentation of ligament repair.

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.

The effect of remnant preservation on patterns of gene expression in a rabbit model of anterior cruciate ligament reconstruction.

BACKGROUND: Anterior cruciate ligament (ACL) reconstruction with remnant preservation technique had been thought to be a more favorable milieu for graft reinnervation, revascularization, and ligamentization. However, the influence of preserving tibial residual fibers on mRNA expression during the graft remodeling process has never been investigated. MATERIALS AND METHODS: Healthy mature New Zealand white rabbits were randomly assigned to one of four groups: remnant dissected, remnant preserved, sham operated, and normal control. Ligament tissue was dissected at 2, 6, and 12 wk after surgery, and real-time PCR was performed using primers for VEGF, TGF-ß1, COLlAl, COL3A1, GAP-43, and NT-3. RESULTS: In the remnant preservation group, mRNA levels for matrix components COL l Al, COL3A1, growth factor TGF-ß1, and nerve-related genesGAP-43 all increased 6 wk after surgery, compared with the remnant dissection group (P < 0.05). An increased level of VEGF mRNA was also detected in the remnant preservation group 12 wk after operation (P < 0.05). An increased level of NT-3 mRNA was also observed in the remnant preservation group 2 and 12 wk after operation (P < 0.05). CONCLUSIONS: Our results suggest that there is a time dependent alteration of angiogenesis-promoting, repair-related, and nerve-related gene expression after ACL reconstruction during the process of graft remodeling. Furthermore, they demonstrate that remnant preservation in ACL reconstruction determines the different molecular profiles of these target genes, especially during the early stages of graft remodeling, which perhaps explains the potential role in promoting revascularization, reinnervation, and ligamentization.

Isolation and characterization of human anterior cruciate ligament-derived vascular stem cells.

The anterior cruciate ligament (ACL) usually fails to heal after rupture mainly due to the inability of the cells within the ACL tissue to establish an adequate healing process, making graft reconstruction surgery a necessity. However, some reports have shown that there is a healing potential of ACL with primary suture repair. Although some reports showed the existence of mesenchymal stem cell-like cells in human ACL tissues, their origin still remains unclear. Recently, blood vessels have been reported to represent a rich supply of stem/progenitor cells with a characteristic expression of CD34 and CD146. In this study, we attempted to validate the hypothesis that CD34- and CD146-expressing vascular cells exist in hACL tissues, have a potential for multi-lineage differentiation, and are recruited to the rupture site to participate in the intrinsic healing of injured ACL. Immunohistochemistry and flow cytometry analysis of hACL tissues demonstrated that it contains significantly more CD34 and CD146-positive cells in the ACL ruptured site compared with the noninjured midsubstance. CD34+CD45- cells isolated from ACL ruptured site showed higher expansionary potentials than CD146+CD45- and CD34-CD146-CD45- cells, and displayed higher differentiation potentials into osteogenic, adipogenic, and angiogenic lineages than the other cell populations. Immunohistochemistry of fetal and adult hACL tissues demonstrated a higher number of CD34 and CD146-positive cells in the ACL septum region compared with the midsubstance. In conclusion, our findings suggest that the ACL septum region contains a population of vascular-derived stem cells that may contribute to ligament regeneration and repair at the site of rupture.

Immunohistochemical and histomorphometric evaluation of vascular distribution in intact canine cranial cruciate ligament.

OBJECTIVES: To (1) describe vascular distribution in the grossly intact canine cranial cruciate ligament (CCL) using immunohistochemical techniques specific to 2 components of blood vessels (factor VIII for endothelial cells, laminin for basement membrane); and (2) compare the vascularity in different areas of interest (craniomedial versus caudolateral bands; core versus epiligamentous regions; and proximal versus middle versus distal portions) in the intact normal canine CCL. STUDY DESIGN: In vitro study. ANIMALS: Large, mature dogs (n=7) of breeds prone to CCL disease that were euthanatized for nonorthopedic conditions. METHODS: Intact CCL were collected from fresh canine cadavers free from stifle pathology. CCL tissue was processed for immunohistochemistry and stained for factor VIII and laminin. Vascular density was determined by histomorphometric analysis. RESULTS: Specific vascular staining was sparsely identified throughout the CCL; however, the proximal portion of the CCL appears to have a greater number of vessels than the middle or distal portion of the ligament. CONCLUSIONS: The CCL is a hypovascular tissue and its vascular distribution is not homogeneous.

Vascular distribution in ruptured canine cranial cruciate ligament.

OBJECTIVE: To (1) determine the microanatomic vascular distribution in ruptured canine cranial cruciate ligaments (CCL) using specific vascular immunohistochemical techniques, and (2) compare vessel density between ruptured and intact canine CCL and between different areas of interest in ruptured CCL using histomorphometric analysis. STUDY DESIGN: In vitro study. ANIMALS: Dogs (n=41) admitted for surgical treatment of ruptured CCL and 19 dogs euthanatized for nonorthopedic conditions. METHODS: Diseased (variable CCL rupture) and intact (normal control) CCL were processed for immunohistochemical staining specific to vessels (factor VIII, laminin). Mean vascular density was assessed and compared in areas of interest (torn end versus remaining core regions of CCL, proximal femoral versus distal tibial core CCL regions). RESULTS: Ruptured CCL was more vascular than intact CCL; however there was no difference in vascular density between the torn end and the remaining core area of the ruptured CCL. Ruptured CCL was vascularized to a greater degree at the proximal portion than the distal portion of the CCL. Partially ruptured CCLs had a higher vessel density than completely ruptured CCLs. CONCLUSIONS: Vascular density is increased in diseased CCL compared with intact CCL. It remains to be determined whether this finding is associated with the cause of CCL rupture or is a result of CCL degeneration and rupture.