Tomonori Takagaki

We have previously studied the effects of chondrocyte sheets on the repair and regeneration of articular cartilage by using temperature-responsive culture inserts. On the basis of this work, we succeeded in rapid fabrication of chondrocyte sheets with the use of a coculture method in which inserts were placed between synoviocytes and chondrocytes. Treatment of cartilage defects using layered chondrocyte sheets promotes repair and regeneration; this method is compatible with in vivo osteoarthritis (OA) models that reproduce partial-thickness defects. In human stem cell clinical research guidelines, the Ministry of Health, Labour and Welfare (MHLW) approved several applications related to this technology. Indeed, its translation to a clinical setting is already yielding favorable results. Here, we evaluated the risk of tumorigenesis associated with this treatment and characterized the dynamics of biological processes associated with the post-transplantation cell sheets in vivo. Furthermore, we also confirmed the safety of the procedure by using array comparative genomic hybridization (array CGH) and G-band staining to screen for deleterious genetic aberrations during prolonged subculture of cells. The safety of chondrocytes that were cultured for longer than normal was confirmed by the array CGH and G-band staining results. Additionally, tumorigenicity testing confirmed that culture chondrocyte sheets are not tumorigenic. Furthermore, from the evaluation of bioluminescence imaging following implantation of the cell sheets, it was confirmed that the transplanted chondrocytes and synoviocytes remained in the knee joint and did not transfer elsewhere over time. We believe that the technique used in this study is a highly useful method for evaluating the safety of not only chondrocytes, but extensive subculturing in general.

T2 mapping is an MRI method particularly reflective of the collagen arrangement in the cartilage, and diffusion tensor (DT) imaging captures the diffusion of water molecules. Laser-induced photoacoustic measurement (LIPA) makes it possible to assess not only the thickness of the cartilage layer but also its viscoelastic properties. By assessing cartilage damage assessment using LIPA and 3.0 Tesla MRI (T2 mapping and DT imaging), this study investigates the usefulness of the various methods. The International Cartilage Repair Society (ICRS) classification was used to classify 29 bone cartilage pieces excised during surgical procedures. At the same time, LIPA was performed at sites matching the area of cartilage damage. MRI was performed preoperatively to measure the T2 and the apparent diffusion coefficient. In addition, tissue sections for histological assessment using the Mankin score were prepared for each ICRS grade, and the results with the various methods were compared. With DT imaging, significant differences were observed in all grades (P < 0.01). With T2 mapping, significant differences were observed in all grades except for grade 1 versus grade 2 (P < 0.01). With LIPA, significant differences were observed in ICRS grade 1 versus grade 3 (P < 0.05), grade 1 versus grade 4 (P < 0.01), grade 2 versus grade 4 (P < 0.01), and grade 3 versus grade 4 (P < 0.05). With the Mankin score, significant differences were observed in ICRS grade 1 versus grade 3 (P < 0.01), grade 1 versus grade 4 (P < 0.01), grade 2 versus grade 4 (P < 0.01), and grade 3 versus grade 4 (P < 0.01). Correlations were observed in all combinations of ICRS grade with DT imaging, T2 mapping, LIPA, and Mankin score. Correlations were observed between the degree of histological degeneration and DT imaging, T2 mapping, and ICRS grade, but LIPA had a weaker correlation than MRI. In the assessment of knee osteoarthritis, there are instances where it is difficult to assess the damaged cartilage site with MRI alone, and we believe that it is desirable to use a combination of LIPA and MRI.

Prevention and early detection of venous thromboembolism (VTE) is important after arthroplasty of the lower limb. The purpose of this study was to investigate the associations between VTE and hemostatic markers after minimally invasive total knee arthroplasty (MIS-TKA). We performed a retrospective study of 50 patients (55 knees) who underwent primary unilateral MIS-TKA with periodic determination of D-dimer and soluble fibrin monomer complex (SFMC) concentrations and with ultrasonography. The development of symptomatic and asymptomatic VTE, location of deep venous thrombosis (DVT; proximal or distal), changes in SFMC and D-dimer concentrations, and correlations between hemostatic markers and VTE onset were evaluated. Twenty-six patients (47 %) had an asymptomatic distal DVT, but none had proximal DVT, pulmonary embolism, or symptomatic DVT. DVT was detected at postoperative day 1 (POD1) in 16 patients, POD3 in six, and POD5 in three (excluding detections of the same DVT in the same position on different days). DVT onset correlated significantly with SFMC concentration on POD1 and with D-dimer concentration on POD3. The D-dimer concentration did not differ significantly between patients who developed DVT (DVT+) and those who did not (DVT−) at each postoperative time. SFMC concentration differed between DVT+ and DVT− patients only on POD1. Analysis of each hemostatic marker classified as either within or outside the normal concentration range showed no significant correlations between D-dimer concentration and DVT onset at each period. There were significant correlations between SFMC concentrations and DVT onset on POD1 and POD3. There were also significant correlations between D-dimer positive (+) findings and/or SFMC+ findings and DVT onset on POD1 and POD3. D-dimer+ and/or SFMC+ findings had better specificity on POD1 and a positive predictive value on POD1 and POD3 compared with SFMC+ alone. SFMC concentration is an effective hemostatic marker for early detection of DVT. D-dimer concentration alone has limited value as a hemostatic marker for early detection of DVT. Measurement of both D-dimer and SFMC concentrations might be a more sensitive diagnostic tool than measuring SFMC concentration alone.

In the present study, we measured damaged areas of cartilage with diffusion tensor (DT) imaging and T2 mapping, and investigated the extent to which cartilage damage could be determined using these techniques. Forty-one patients underwent arthroscopic knee surgery for osteoarthritis of the knee, a meniscus injury, or an anterior cruciate ligament injury. Preoperative magnetic resonance imaging of the knee was performed, including T2 mapping and diffusion tensor imaging. The presence of cartilage injury involving the medial and lateral femoral condyles and tibia plateau was assessed during surgery using the Outerbridge scale. The ADC, T2 values and fractional anisotropy of areas of cartilage injury were then retrospectively analysed. The ADC results identified significant differences between Outerbridge grades 0 and 2 (P = 0.041); 0 and 3 (P < 0.001); 1 and 2 (P = 0.045); 1 and 3 (P < 0.001); and 2 and 3 (P = 0.028). The FA results identified significant differences between grades 0 and 1 (P < 0.001); 0 and 2 (P < 0.001); and 0 and 3 (P < 0.001). T2 mapping identified significant differences between Outerbridge grades 0 and 2 (P = 0.032); 0 and 3 (P < 0.001); 1 and 3 (P < 0.001); and 2 and 3 (P < 0.001). Both the T2 mapping (R2 = 0.7883) and the ADC (R2 = 0.9184) correlated significantly with the Outerbridge grade. The FA (R2 = 0.6616) correlated slightly with the Outerbridge grade. T2 mapping can be useful for detecting moderate or severe cartilage damage, and the ADC can be used to detect early stage cartilage damage. The FA can also distinguish normal from damaged cartilage.