Induction of chondrogenesis with a RANKL-binding peptide, WP9QY, in vitro and in vivo in a rabbit model.

WP9QY (W9) is a receptor activator of nuclear factor-κB ligand (RANKL)-binding peptide that inhibits osteoclastogenesis by blunting the RANKL-RANK interaction, and also increases osteoblastogenesis via RANKL reverse signaling. W9 has dual effects on osteoclasts and osteoblasts; however, it is unknown whether the peptide has an effect on chondrocytes. Here, we report that W9 induces proliferation and differentiation of chondrocytes in vitro and repairs full-thickness articular cartilage defects in vivo. W9 stimulated chondrocyte differentiation in a two-dimensional (2D) culture of human mesenchymal stem cells (hMSCs), and transforming growth factor ß3 (TGF-ß3) showed synergistic effects with W9 on chondrogenesis. W9 enlarged the size of 3D pellet cultures of hMSCs and produced chondrocyte-specific matrices, especially in combined treatment with TGF-ß3. The peptide also stimulated proliferation of hMSCs with induction of expression of chondrogenesis-related genes. Several RANKL inhibitors had no effect on chondrocytic differentiation. RANKL-knockdown experiments showed that W9 did not induce chondrogenesis through RANKL, but did induce osteoblastogenesis through RANKL. Intraarticular injection of W9 resulted in significant repair of full-thickness articular cartilage defects in rabbits. Taken together, these results suggest that W9 ameliorates the articular cartilage defects by increasing the volume of cartilaginous matrices with accompanying induction of proliferation and differentiation of chondrocytes via mechanisms independent of RANKL inhibition and RANKL reverse signaling. Since no pharmaceuticals are clinically available for treatment of cartilage damage such as osteoarthritis, our findings demonstrate the potential of W9 to address the unmet medical needs.

Assessment of Meniscal Healing Status by Magnetic Resonance Imaging T2 Mapping After Meniscal Repair.

BACKGROUND: Although the sensitivity and specificity of magnetic resonance imaging (MRI) for the diagnosis of primary meniscal tears are high, these values are lower for the assessment of healing status of repaired menisci. PURPOSE: To compare the accuracy of MRI T2 mapping and conventional MRI in assessing meniscal healing after repair. STUDY DESIGN: Cohort study (diagnosis); Level of evidence, 2. METHODS: Patients who underwent meniscal repair with concurrent anterior cruciate ligament reconstruction between 2012 and 2016 and had a follow-up second-look arthroscopy were enrolled. The patients were divided into healed and incompletely/not healed groups based on the second-look arthroscopy findings. For the repaired menisci, the following were compared between the groups, (1) Stoller and Crues classification on conventional MRI with a proton density-weighted fat-saturated sequence and (2) the remaining colored meniscal tear line on T2 mapping coincident with the high signal line showing the primary tear on conventional MRI were compared. The change of T2 relaxation time (ΔT2) of the colored meniscal tear line pre- to postoperatively was compared between the groups. The mean T2 relaxation time of the whole area of the postoperative meniscus at each slice was also compared with that of control menisci to assess the whole quality of the repaired meniscus. RESULTS: A total of 26 menisci from 24 knees were assessed (16 healed menisci, 10 incompletely/not healed menisci). According to the Crues classification on conventional MRI, 8 of 16 healed menisci and 3 of 10 incompletely/not healed menisci improved from grade 3 to 2, with there being no significant difference between the groups (P = .43). However, the colored meniscal tear line remained in only 3 of the 16 healed menisci as compared with 9 of the 10 incompletely/not healed menisci, and the presence of this colored line allowed differentiation between healed menisci and incompletely/not healed menisci (sensitivity, 81.3%; specificity, 90.0%; odds ratio, 39.0; P = .001). The mean (SD) ΔT2 was -31.1 ± 3.2 and -19.9 ± 4.4 ms in the healed and incompletely/not healed groups, respectively (P < .001). Receiver operating characteristic curve analysis showed a cutoff ΔT2 value of -22.3 ms for separation of meniscal healing (P < .001). The T2 relaxation times of the whole area of the repaired menisci were 31.7 ± 3.4 and 32.8 ± 3.8 ms in the healed and incompletely/unhealed groups, respectively (P = .69), with these values being significantly longer than the 26.9 ± 2.2 ms in the controls (P < .001). CONCLUSION: MRI T2 mapping allowed the differentiation of healing status after meniscal repair, with high sensitivity and specificity as compared with conventional MRI.

Prevalence, development, and factors associated with cyst formation after meniscal repair with the all-inside suture device.

PURPOSE: To investigate the prevalence of cyst formation after using all-inside meniscal repair device and analysed the risk factors associated with it. METHODS: Between August 2008 and September 2013, 51 menisci of 46 patients were included in the study, 46 menisci of which had concomitant anterior cruciate ligament (ACL) ruptures and had an ACL reconstruction. Magnetic resonance imaging (MRI) of the knee was performed at 3, 6, 12 and 24 months after meniscal surgery. The MRIs were assessed to detect the development of cysts encasing the suture anchors and to evaluate meniscal healing. Statistical analysis was performed using multiple regression analysis. RESULTS: Out of the 51 menisci examined, MRI revealed cysts in 15 menisci. Cysts were detected in 3 menisci at 6 months, in 9 menisci at 12 months, and in 3 menisci at 24 months after surgery. Only 3 patients (6.5%) were symptomatic, and cystectomy was performed in 2 of these patients and arthroscopic debridement in the other. Compared with using both the suture device and an inside-out suture repair, using the suture device alone was more likely to be associated with cyst development [odds ratio (OR), 12.04]. The medial meniscus was also significantly more likely to develop a cyst compared with the lateral meniscus (OR, 12.48). There was an increased outcome for the number of device use (P = 0.033). Though it was not statistically significant, the patients with anterior knee laxity (side-to-side difference > 3 mm using a knee arthrometer) were more likely to develop cysts than those without anterior knee laxity (P = 0.06). There were no significant differences between the remaining variables. CONCLUSIONS: The prevalence of cyst formation around the suture implant was 29%, but most cases were not symptomatic. Significant risk factors for cyst formation included the use of a suture device alone, and a location in the medial meniscus. LEVEL OF EVIDENCE: III.

Incidence and Risk Factors for Meniscal Cyst After Meniscal Repair.

PURPOSE: To investigate the incidence of magnetic resonance imaging-confirmed cyst formation after meniscal repair and to analyze associated risk factors. METHODS: This retrospective study included cases repaired arthroscopically with the all-inside (AI) technique (using suture anchors) and/or the inside-out (IO) technique between October 2008 and December 2014. A meniscal cyst was detected on T2 fat-suppressed magnetic resonance images. All cases were divided into 3 groups according to the repair method (AI, IO, and combined technique). The incidence of radiographically confirmed meniscal cyst formation in each group and the associated risk factors (age, sex, AI device, medial meniscus, Tegner activity scale preinjury) were analyzed. RESULTS: A total of 102 menisci in 96 knees were evaluated. The mean follow-up period was 3.8 (range, 2-8) years. The mean patient age was 21.0 (range, 6-53) years. Thirty cases were in the AI group, 60 in the IO group, and 12 in the combined group. Demographically, there were significant differences among groups regarding the number of medial, lateral, and discoid tears; concomitant anterior cruciate ligament tears; Tegner scale; and suture number. Meniscal cysts developed in 14 of 102 cases. Two of the 14 cysts were symptomatic, requiring open cystectomy. The incidence of meniscal cyst was significantly higher in the AI group (12 of 30, 40%) than in the IO group (1 of 60, 1.7%) or the combined-technique group (1 of 12, 8.3%) (P < .001). Both symptomatic cysts were in the AI group and were in continuity with the anchors. Medial meniscus tear (odds ratio = 6.92) and the use of AI suture anchors (odds ratio = 15.03) significantly increased the risk of cyst formation. CONCLUSIONS: The incidence of meniscal cysts after arthroscopic meniscal repair was 1.7% to 40.0%, depending on the surgical method. Medial meniscus tears and use of an AI device are suggested as risk factors for cyst formation in this retrospective study. LEVEL OF EVIDENCE: Level Ⅲ, retrospective comparative study.

Risk Factors Associated With Knee Joint Degeneration After Arthroscopic Reshaping for Juvenile Discoid Lateral Meniscus.

BACKGROUND: Although arthroscopic meniscal reshaping for discoid lateral meniscus (DLM) has better outcomes than total or subtotal meniscectomy, degenerative changes on radiographs are still seen in some patients with meniscal reshaping. PURPOSE: To assess the risk factors associated with knee joint degeneration after reshaping surgery for juvenile DLM. STUDY DESIGN: Case-control study; Level of evidence, 3. METHODS: Forty patients (45 knees) with a mean age of 12.0 years who underwent arthroscopic meniscal reshaping for DLM were enrolled at a mean of 39.6 months after surgery. For all patients, meniscal saucerization was performed first. Then, if the residual meniscus was unstable, stabilization was provided with suture fixation. At final follow-up, we obtained radiographs to assess degenerative changes to the knee joint using the classification by Tapper and Hoover. Residual meniscal width and meniscal extrusion (defined as a relative percentage of extrusion [RPE]) were measured with magnetic resonance imaging (MRI). Then, the correlation between radiographic evidence of degenerative changes (Tapper and Hoover grade), residual meniscal width, and RPE were assessed with Pearson and Spearman correlation analyses. Logistic regression analysis was used to examine whether preoperative characteristics correlated with degeneration and residual meniscal width. RESULTS: The mean residual meniscal width was 4.6 mm (range, 3.8-6.0 mm), and the mean ± SD RPE was 25.5% ± 21.8% at the final follow-up. There were 28 knees with Tapper and Hoover grade 0, 10 knees with grade 1, and 7 knees with grade 2. The residual meniscal width and RPE were significantly correlated with Tapper and Hoover grade (ρ = -0.489, P = .0007; ρ = 0.414, P = .005, respectively). The residual meniscal width was also significantly correlated with RPE ( r = -0.416, P = .004). The receiver operating characteristic curve showed that a 5.0-mm residual meniscal width was the cutoff value leading to evidence of degeneration. Multiple logistic regression analysis showed that an anterocentral shift on preoperative MRI was a risk factor for degeneration (odds ratio, 27.2; 95% CI, 1.1-360.5; P = .012) and residual meniscal width less than 5.0 mm (odds ratio, 20.9; 95% CI, 1.5-281; P = .022). CONCLUSION: Smaller meniscal width and greater severity of meniscal extrusion correlated with knee joint degeneration. An anterocentral shift on preoperative MRI was a risk factor for degenerative changes and smaller residual meniscal width.

The origin and distribution of CD68, CD163, and αSMA+ cells in the early phase after meniscal resection in a parabiotic rat model.

We previously reported that circulating peripheral blood-borne cells (PBCs) contribute to early-phase meniscal reparative change. Because macrophages and myofibroblasts are important contributors of tissue regeneration, we examined their origin and distribution in the reparative meniscus. Reparative menisci were evaluated at 1, 2, and 4 weeks post-meniscectomy by immunohistochemistry to locate monocytes and macrophages (stained positive for CD68 and CD163), and myofibroblasts (stained positive for αSMA). Of the total number of cells, 13% were CD68+ at 1 week post-meniscectomy, which decreased to 1% by 4 weeks post-meniscectomy; of these, almost half of CD68+ cells (49.4%: 98.8% as PBCs) were green fluorescent protein (GFP)-positive post-meniscectomy (1, 2, and 4 weeks), indicating that the majority of CD68+ cells were derived from PBCs. Of the total cells, 6% were CD163+ at 1 week post-meniscectomy, which decreased to 1% by week 4. Of the CD163+ cells, the majority were GFP-positive (42.5%: 85.0% as PBCs) after 1 week; however, this decreased significantly over time, which indicates that the majority of CD163+ cells are derived from PBCs during the early phase of meniscal reparative change, but are derived from resident cells at later time points. Of the total cells, 38% were αSMA+ at 1 week post-meniscectomy, which decreased to 3% by 4 weeks. The proportion of GFP-positive αSMA+ cells was 2.8% after 1 week, with no significant change over time, which indicates that the majority of αSMA+ cells originated from resident cells. Here, we describe the origin and distribution of macrophages and myofibroblasts during meniscal reparative change.

Circulating nucleated peripheral blood cells contribute to early-phase meniscal healing.

The purpose of this study was to assess how peripheral blood cells (PBCs) contribute to meniscus repair, using a parabiotic rat model. Wild-type (WT) and green fluorescent protein (GFP) transgenic rats were conjoined at the torso. After 4 weeks, the anterior part of the medial meniscus of both groups of rats was removed. At 1, 2, 4, 8 and 12 weeks post-meniscectomy, repaired tissue was evaluated using stereomicroscopy, histology with toluidine blue staining, and immunofluorescence microscopy. Stereomicroscopic observations and confocal laser microscopy revealed that a high number of GFP-positive cells were present in the repaired meniscus of WT rats 1 week post-meniscectomy, and the number of GFP-positive cells decreased over time. Based on blood chimerism, the ratios of PBCs in the repaired meniscus were 20.5 ± 2.3% at 1 week, 8.3 ± 0.9% at 2 weeks, 4.4 ± 0.9% at 4 weeks, 2.1 ± 0.9% at 8 weeks, and 0.5 ± 0.4% at 12 weeks, post-meniscectomy. Histologically, fibrochondrocytes were observed in the repaired meniscus of WT rats after 4 weeks, some of which were GFP-positive. The chondrogenic marker, type II collagen, was merged within the PBCs in the repaired tissue. However, type-II-collagen-positive cell ratio and metachromasia in the repaired meniscus were not equivalent in normal meniscal tissue. This indicated that PBCs were present within the repaired meniscus at an early phase, replacing the excised meniscal cells, suggesting PBCs contributed to meniscal healing. The tissue repair contribution by these cells decreased at later phases. Copyright © 2014 John Wiley & Sons, Ltd.

A case of asymptomatic bilateral massive pulmonary embolism after arthroscopic multiple knee ligament reconstruction.

Venous thromboembolism, which includes deep venous thrombosis (DVT) and pulmonary embolism (PE), is a serious complication after operations involving the lower extremities, and it can be fatal. However, few reports have described the incidence of PE and its associated risk factors after arthroscopic knee surgery. We present a case of bilateral massive PE of the main pulmonary arteries and DVT detected on multi-detector row computed tomography after arthroscopic multiple knee ligament reconstruction. Our patient was asymptomatic despite having several risk factors for thromboembolic events (43 years of age, a long operation time, obesity, and diabetes mellitus) and receiving no pharmacologic thromboembolic prophylaxis. Although fatality due to PE is relatively uncommon, when a patient has several risk factors for PE, perioperative thromboprophylaxis should be considered. Level of evidence IV.

Direct bone-to-bone integration between recombinant human bone morphogenetic protein-2-injected tendon graft and tunnel wall in an anterior cruciate ligament reconstruction model.

PURPOSE: This study was performed to evaluate one-stage anterior cruciate ligament (ACL) reconstruction using a semitendinosus tendon graft injected with bone morphogenetic protein 2 (BMP-2) in a rabbit model. METHODS: We injected recombinant human BMP-2 (rhBMP-2) in the experimental group and phosphate-buffered saline in the control group at two sites of the semitendinosus tendon (15 µg in each site) to replace tendon with bone in the bone tunnel. Twenty minutes later, the injected tendon graft was transplanted for ACL reconstruction by passing the graft through the bone tunnel. The animals were harvested at four, eight, or 12 weeks postoperatively and examined by histological and biomechanical methods. RESULTS: Histological analysis revealed that the tendon graft was replaced with new bone in the tunnel of the experimental group. Characteristic features identical to the regenerated direct insertion morphology at the bone-tendon junction were acquired at eight or 12 weeks in the experimental group. Biomechanical pull-out testing revealed greater stiffness in the experimental than control group at 12 weeks, although the maximum load to failure showed no significant difference between the two groups at four, eight, or 12 weeks. CONCLUSION: These results indicate the potential for ACL reconstruction with regenerated direct insertion morphology.

Proposed Referential Index to Resect Femoroacetabular Cam-Type Impingement During Arthroscopy Using a Cadaveric Hip Model.

PURPOSE: To establish a reference index for the simple identification of the optimum resection point for cam-type impingement on arthroscopy. METHODS: Twelve cadaveric left hips with a 20° to 40° center-edge angle, without osteoarthritis, were examined (mean age, 85 ± 10.1 years). The pelvis was fixed such that the anterior pelvic plane and femur were parallel to the table. The resection line for impingement was first defined on the femoral head surface 5 mm distal to the acetabular labrum, from the 9-o'clock (anterior) to 12-o'clock (superior) position. Next, we measured the hip flexion angle necessary for the head-neck junction to reach the resection line. After positioning the wire on the femoral head surface along the resection line from the 9- to 12-o'clock area of the femoral head, we measured the target alpha angle on radiographs at 0°, 15°, 30°, 45°, and 60° of hip flexion using the frog-leg 45/45/30 view (45° of flexion, 45° of abduction, and 30° of external rotation) and Dunn 45 view (45° of flexion, 20° of abduction, and neutral rotation). RESULTS: The mean hip flexion angle at which the head-neck junction reached the resection line was 31° ± 4.6°. For 0°, 15°, 30°, 45°, and 60° of hip flexion, the mean target alpha angle was 75.5° ± 5.5°, 65.3° ± 5.6°, 56.3° ± 5.8°, 49.0° ± 6.6°, and 42.6° ± 5.8°, respectively, using the frog-leg 45/45/30 view and 75.0° ± 6.0°, 65.8° ± 6.2°, 57.2° ± 7.3°, 50.7° ± 6.9°, and 44.2° ± 5.8°, respectively, using the Dunn 45 view. There were no significant differences between the 2 radiographic techniques (P = .82, P = .84, P = .76, P = .57, and P = .52, respectively). CONCLUSIONS: A description of the degree of hip flexion during cam resection can affect the final alpha angle when using the labrum as a reference for resection. CLINICAL RELEVANCE: The described index allows systematic navigation of cam lesions during arthroscopy for femoroacetabular impingement patients using the hip flexion angle.

Effect of the direct injection of bone marrow mesenchymal stem cells in hyaluronic acid and bone marrow stimulation to treat chondral defects in the canine model.

INTRODUCTION: The purpose of this study was to assess the direct injection of bone marrow-derived mesenchymal stem cells (BMSCs) suspended in hyaluronic acid (HA) combined with drilling as a treatment for chondral defects in a canine model. METHODS: Tibial bone marrow was aspirated, and BMSCs were isolated and cultured. One 8.0-mm diameter chondral defect was created in the femoral groove, and nine 0.9-mm diameter holes were drilled into the defect. BMSCs (2.14 × 107 cells) suspended in HA were injected into the defect. HA alone was injected into a similar defect on the contralateral knee as a control. Animals were sacrificed at 3 and 6 months. RESULTS: Although the percentage of coverage assessed macroscopically was significantly better at 6 months than at 3 months in both the BMSC (p = 0.02) and control (p = 0.001) groups, there were no significant differences in the International Cartilage Repair Society grades. The Wakitani histological score was significantly better at 6 months than at 3 months in the BMSC and control groups. While the control defects were mostly filled with fibrocartilage, several of the defects in the BMSC group contained hyaline-like cartilage. The mean Wakitani scores of the BMSC group improved from 7.0 ± 1.0 at 3 months to 4.6 ± 0.9 at 6 months, and those of the control group improved from 9.4 ± 1.2 to 6.0 ± 0.6. The BMSC group showed significantly better regeneration than the control group at 3 months (p = 0.04), but the difference at 6 months was not significant (p = 0.06). CONCLUSIONS: The direct injection of BMSCs in HA combined with drilling enhanced cartilage regeneration.