Norimasa Nakamura

Autologous bone marrow stromal cell transplantation for repair of full-thickness articular cartilage defects in human patellae : two case reports

This study assessed the effectiveness of autologous bone marrow stromal cell transplantation for the repair of full-thickness articular cartilage defects in the patellae of a 26-year-old female and a 44-year-old male. These two patients presented in our clinic because their knee pain prevented them from walking normally. After thorough examination, we concluded that the knee pain was due to the injured articular cartilage and decided to repair the defect with bone marrow stromal cell transplantation. Three weeks before transplantation, bone marrow was aspirated from the iliac crest of each patient. After erythrocytes had been removed by use of dextran, the remaining nucleated cells were placed in culture. When the attached cells reached subconfluence, they were passaged to expand in culture. Adherent cells were subsequently collected, embedded in a collagen gel, transplanted into the articular cartilage defect in the patellae, and covered with autologous periosteum. Six months after transplantation, clinical symptoms (pain and walking ability) had improved significantly and the improvement has remained in effect (5 years and 9 months posttransplantation in one case, and 4 years in the other), and both patients have been satisfied with the outcome. As early as 2 months after transplantation, the defects were covered with tissue that showed slight metachromatic staining. Two years after the first and 1 year after the second transplantation, arthroscopy was performed and the defects were repaired with fibrocartilage. Results indicate autologous bone marrow stromal cell transplantation is an effective approach in promoting the repair of articular cartilage defects.

Direct anterior cruciate ligament insertion to the femur assessed by histology and 3-dimensional volume-rendered computed tomography.

PURPOSE The purpose of this study was to histologically identify the direct and indirect insertion of the femoral anterior cruciate ligament (ACL) insertion. Furthermore, we quantitatively measured the direct femoral insertion area by use of the 3-dimensional (3D) volume-rendered (VR) computed tomography (CT) model. METHODS By use of 8 intact cadaveric knees, the lateral femoral condyle including the ACL attachment was sectioned for histologic examination in 3 oblique-axial planes parallel to the roof of the intercondylar notch and in the sagittal planes. Before sectioning, these knees had been subjected to CT to obtain 3D VR images of the femur. Once the direct insertion of the ACL was identified on each histologic section, the corresponding image was superimposed on the corresponding CT image. RESULTS The direct ACL insertion, in which dense collagen fibers were connected to the bone by the fibrocartilaginous layer, was microscopically identified at the region between the posteromedial articular cartilage margin of the lateral femoral condyle and the linear bony ridge 7 to 10 mm anterior to the articular cartilage margin. Meticulous comparison of histologic analysis and the 3D VR CT model showed that the ACL direct insertion coincided with a crescent-shaped hollow just behind the linear bony ridge. The direct insertion measured 17.4 +/- 0.9 mm (mean +/- SD) in length, 8.0 +/- 0.5 mm in width, and 128.3 +/- 10.5 mm(2) in area. CONCLUSIONS The direct insertion of the ACL is located in the depression between the residents ridge and the articular cartilage margin on the lateral femoral condyle. It measured 17.4 +/- 0.9 mm in length, 8.0 +/- 0.5 mm in width, and 128.3 +/- 10.5 mm(2) in area. CLINICAL RELEVANCE Delineation of the ACL femoral direct insertion by 3D VR CT could be a useful tool for planning of accurate femoral tunnel positioning in anatomic ACL reconstruction.

Rectangular Tunnel Double-Bundle Anterior Cruciate Ligament Reconstruction with Bone-Patellar Tendon-Bone Graft to Mimic Natural Fiber Arrangement

We describe our current technique of anatomic, double-bundle (DB), rectangular tunnel anterior cruciate ligament (ACL) reconstruction with bone-patellar tendon-bone (BPTB) graft. This technique mimics the natural, or anatomic, arrangement of the native ACL fibers. This technique has the following advantages: (1) creation of a DB ACL reconstruction with a single BPTB graft; (2) maximization of graft-tunnel contact area; (3) containment of the tunnel apertures within the anatomic ACL attachment footprint; (4) rotational control of the graft within the tunnels during and after fixation; and (5) preservation of notch anatomy.

In Vitro Generation of a Scaffold-Free Tissue-Engineered Construct (TEC) Derived from Human Synovial Mesenchymal Stem Cells: Biological and Mechanical Properties and Further Chondrogenic Potential

The purpose of this study was to characterize a tissue-engineered construct (TEC) generated with human synovial mesenchymal stem cells (MSCs). MSCs were cultured in medium with ascorbic acid 2-phosphate (Asc-2P) and were subsequently detached from the substratum. The detached cell/matrix complex spontaneously contracted to develop a basic TEC. The volume of the TEC assessed by varying initial cell density showed that it was proportional to initial cell densities up to 4 x 10(5) cells/cm(2). Assessment of the mechanical properties of TEC using a custom device showed that the load at failure and stiffness of the constructs significantly increased with time of culture in the presence of Asc-2P, while in the absence of Asc-2P, the constructs were mechanically weak. Thus, the basic TEC possesses sufficiently self-supporting mechanical properties in spite of not containing artificial scaffolding. TEC further cultured in chondrogenic media exhibited positive alcian blue staining with elevated expression of chondrogenic marker genes. Based on these findings, such human TEC may be a promising method to promote cartilage repair for future clinical application.

Conservative treatment of isolated injuries to the posterior cruciate ligament in athletes

We assessed arthroscopically 22 young athletes with an isolated acute posterior cruciate ligament (PCL) injury. Four had significant damage to the articular cartilage of the medial femorotibial compartment and were advised not to resume sports. Three underwent PCL reconstruction because of a reparable meniscal tear or instability. The other 15 were treated conservatively and resumed sport. At an average follow-up of 51 months, one had developed arthritic symptoms due to newly-developed severe chondral damage to the medial femoral condyle, but none of the other 14 had developed arthritic symptoms and most remained athletically active. Severe chondral damage should be seen at an early arthroscopy. Knees with an isolated injury to the PCL with concomitant articular damage may be successfully managed by conservative treatment.

The influence of skeletal maturity on allogenic synovial mesenchymal stem cell-based repair of cartilage in a large animal model.

One of the potential factors that may affect the results of mesenchymal stem cell (MSC)-based therapy is the age of donors and recipients. However, there have been no controlled studies to investigate the influence of skeletal maturity on the MSC-based repair of cartilage. The purpose of this study was to compare the repair quality of damaged articular cartilage treated by a scaffold-free three-dimensional tissue-engineered construct (TEC) derived from synovial MSCs between immature and mature pigs. Synovial MSCs were isolated from immature and mature pigs and the proliferation and chondrogenic differentiation capacities were compared. The TEC derived from the synovial MSCs were then implanted into equivalent chondral defects in the medial femoral condyle of both immature and mature pigs, respectively. The implanted defects were morphologically and biomechanically evaluated at 6 months postoperatively. There was no skeletal maturity-dependent difference in proliferation or chondrogenic differentiation capacity of the porcine synovial MSCs. The TEC derived from synovial MSCs promoted the repair of chondral lesion in both immature and mature pigs without the evidence of immune reaction. The repaired tissue by the TEC also exhibited similar viscoelastic properties to normal cartilage regardless of the skeletal maturity. The results of the present study not only suggest the feasibility of allogenic MSC-based cartilage repair over generations but also may validate the use of immature porcine model as clinically relevant to test the feasibility of synovial MSC-based therapies in chondral lesions.

Results of isolated meniscal repair evaluated by second-look arthroscopy

Thirty-six isolated torn menisci in 35 patients (average age, 24 years) which had been repaired arthroscopically using an inside-out technique were evaluated by second-look arthroscopy. The time from meniscal repair to second-look arthroscopy ranged from 2 to 10 months with a mean of 5 months. The indications for meniscal repair were a longitudinal or oblique tear located at the outer half of the meniscus. Twenty (56%) were graded as excellent, 10 (28%) as good, and 6 (16%) were graded as poor. Neither age nor length of time between injury and repair affected meniscal healing. The medial meniscal repairs showed better results than the lateral repairs (rate of excellent results: medial, 82%; lateral, 44%; P < .01, chi-squared test). The rate of excellent results for those with normal meniscal bodies at the time of repair was 79%, which was significantly higher than that seen in the cases with deformed and/or superficial damage to the meniscal body (36%; P < .05, chi-squared test).

Second-look arthroscopy after meniscal repair. Review of 132 menisci repaired by an arthroscopic inside-out technique

From 1986 to 1993, we repaired 278 torn menisci in 264 patients using an arthroscopically assisted inside-out technique. A total of 132 meniscal repairs in 122 patients were evaluated by second-look arthroscopy. At review, only nine patients had meniscal symptoms, such as locking, swelling or pain. Ninety-seven menisci (73%) had healed completely at the repair site, but there were new tears in different areas of 21 menisci, some of which had complete healing at the repair site. Incomplete healing, seen in 23 menisci (17%), was frequently near the popliteus tendon, most commonly where there had been an associated anterior-cruciate-ligament injury. Arthroscopically-assisted meniscal repair seems to be a reliable procedure, but some clinically successful cases had incomplete healing at the repair site or a newly-formed tear in the meniscal body or both. These lesions may cause meniscal symptoms to appear at a later date.

Comparison of Human Serum with Fetal Bovine Serum for Expansion and Differentiation of Human Synovial MSC: Potential Feasibility for Clinical Applications:

The aim of this study was to evaluate the effect of human serum (HS) on growth and differentiation capacity of human synovium-derived mesenchymal stem cells (MSC) in comparison to cells grown in fetal bovine serum (FBS). Human MSCs were isolated from the synovium of knee joints of three donors and the cells were cultured individually in varying concentrations of allogenic HS or FBS. Bovine MSCs were isolated from synovium and cultured in the same manner. Cell proliferation was assessed by the tetrazolium assay after passage 3. The capacity for chondrogenic and osteogenic differentiation was investigated in specific media followed by 1,9-dimethylmethylene blue assay and alcian blue staining, or by alizarin red staining, respectively. Human MSCs proliferated significantly more rapidly in the presence of HS than with equivalent levels of FBS. Chondrogenic or osteogenic differentiation occurred to nearly identical levels in HS or FBS. The results of this study indicate that HS is superior for the culture of human MSCs compared with FBS in terms of cellular expandability, without losing chondrogenic or osteogenic differentiation capacity. Coupled with the advantage in eliminating the potential risk accompanied with the use of xeno-derived materials, pooled, well-characterized HS could be a useful reagent to promote cellular expansion for clinical synovial stem cell-based therapy.

Imatinib mesylate inhibits osteoclastogenesis and joint destruction in rats with collagen-induced arthritis (CIA).

Macrophage colony-stimulating factor (M-CSF) is a key factor for osteoclastogenesis at the bone–pannus interface in patients with rheumatoid arthritis as well as a receptor activator of NF-κB ligand (RANKL). Imatinib mesylate inhibits the phosphorylation of c-fms, a receptor for M-CSF. The present study investigates the effect of imatinib mesylate on joint destruction in rats with collagen-induced arthritis (CIA) and on osteoclastogenesis in vitro. Imatinib mesylate (50 or 150 mg/kg), dexamethasone, or vehicle was administered daily to CIA rats for 4 weeks from the onset of arthritis. Hind-paw swelling and body weight were measured weekly. At weeks 2 and 4, the metatarsophalangeal (MTP) joints and the ankle and subtalar joints were radiographically and histologically assessed. The effect of imatinib mesylate on osteoclast formation from rat bone marrow cells with M-CSF and soluble RANKL (sRANKL) in vitro was also examined. Radiographic assessment showed that 150 mg/kg imatinib mesylate suppressed the destruction of the MTP and the ankle and subtalar joints at week 2, and MTP joint destruction at week 4 in CIA rats, although hind-paw swelling was not suppressed. The number of TRAP-positive cells at the bone–pannus interface was significantly reduced in the group administered with 150 mg/kg imatinib mesylate compared with that given vehicle at week 4. Imatinib mesylate dose-dependently inhibited the proliferation of M-CSF-dependent osteoclast precursor cells in vitro as well as osteoclast formation induced by M-CSF and sRANKL. These findings suggest that imatinib mesylate could prevent joint destruction in patients with rheumatoid arthritis.