Kenji Serigano

Transplantation of mesenchymal stem cells in a canine disc degeneration model

Transplantation of mesenchymal stem cells (MSCs) is effective in decelerating disc degeneration in small animals; much remains unknown about this new therapy in larger animals or humans. Fas‐ligand (FasL), which is only found in tissues with isolated immune privilege, is expressed in IVDs, particularly in the nucleus pulposus (NP). Maintaining the FasL level is important for IVD function. This study evaluated whether MSC transplantation has an effect on the suppression of disc degeneration and preservation of immune privilege in a canine model of disc degeneration. Mature beagles were separated into a normal control group (NC), a MSC group, and the disc degeneration (nucleotomy‐only) group. In the MSC group, 4 weeks after nucleotomy, MSCs were transplanted into the degeneration‐induced discs. The animals were followed for 12 weeks after the initial operation. Subsequently, radiological, histological, biochemical, immunohistochemical, and RT‐PCR analyses were performed. MSC transplantation effectively led to the regeneration of degenerated discs. FACS and RT‐PCR analyses of MSCs before transplantation demonstrated that the MSCs expressed FasL at the genetic level, not at the protein level. GFP‐positive MSCs detected in the NP region 8 weeks after transplantation expressed FasL protein. The results of this study suggest that MSC transplantation may contribute to the maintenance of IVD immune privilege by the differentiation of transplanted MSCs into cells expressing FasL.

Effect of cell number on mesenchymal stem cell transplantation in a canine disc degeneration model

Transplantation of mesenchymal stem cells (MSCs) inhibits the progression of disc degeneration in animal models. We know of no study to determine the optimal number of cells to transplant into the degenerated intervertebral disc (IVD). To determine the optimal donor cell number for maximum benefit, we conducted an in vivo study using a canine disc degeneration model. Autologous MSCs were transplanted into degenerative discs at 105, 106, or 107 cells per disc. The MSC‐transplanted discs were evaluated for 12 weeks using plain radiography, magnetic resonance imaging, and gross and microscopic evaluation. Preservation of the disc height, annular structure was seen in MSC‐transplantation groups compared to the operated control group with no MSC transplantation. Result of the number of remaining transplanted MSCs, the survival rate of NP cells, and apoptosis of NP cells in transplanted discs showed both structural microenvironment and abundant extracellular matrix maintained in 106 MSCs transplanted disc, while less viable cells were detected in 105 MSCs transplanted and more apoptotic cells in 107 MSCs transplanted discs. The results of this study demonstrate that the number of cells transplanted affects the regenerative capability of MSC transplants in experimentally induced degenerating canine discs. It is suggested that maintenance of extracellular matrix by its production from transplanted cells and/or resident cells is important for checking the progression of structural disruption that leads to disc degeneration. Published by Wiley Periodicals, Inc. J Orthop Res 28:1267–1275, 2010

Human nucleus pulposus cells significantly enhanced biological properties in a coculture system with direct cell-to-cell contact with autologous mesenchymal stem cells

Activated nucleus pulposus (NP) cells can be reinserted into the disc to inhibit intervertebral disc degeneration. Experimental studies in animals showed that using a coculture system with direct cell‐to‐cell contact with mesenchymal stem cells (MSCs) significantly upregulated the biological activity of NP cells. The purpose of this study is to determine whether this activation of NP cells by autologous MSCs is applicable to human cells in vitro. Human NP tissue was obtained from surgical specimens and MSCs from bone marrow of 10 subjects. Six‐well culture plates and inserts were used for culture; 1.0 × 104 NP cells were seeded onto each insert and incubated alone, in standard coculture with 1.0 × 104 MSCs, or cocultured with direct cell‐to‐cell contact. NP cell proliferation, DNA synthesis, and proteoglycan (PG) synthesis were evaluated. Chromosome abnormalities in the activated NP cells and tumorigenesis of the cells were evaluated in an additional 10 patients by microscopic examination for segmented cells and histological assessment of activated cells transplanted into nude mice. Cell proliferation, DNA synthesis, and PG synthesis were significantly upregulated. The positive effects of the coculture system with direct cell‐to‐cell contact seen in animal studies were also confirmed in human cells. Chromosome abnormalities and tumorigenesis were not observed in the activated NP cells. In conclusion, a coculture system with direct cell‐to‐cell contact demonstrated a significant positive effect, enhancing the biological properties of human NP cells, as it did in animal models. These results should prove useful for conducting trials leading to the clinical use of activated NP cell transplantation.

Low-intensity pulsed ultrasound stimulation enhances TIMP-1 in nucleus pulposus cells and MCP-1 in macrophages in the rat.

Recent studies have reported that low‐intensity pulsed ultrasound (LIPUS) stimulates cell proliferation and proteoglycan production in rabbit intervertebral disc cells, and moreover promotes the secretion of MCP‐1 (monocyte chemotaxis protein‐1) from macrophages in a disc organ culture model. These findings suggest the possible application of LIPUS for biological repair of disc degeneration and herniation. Although the mechanisms involved are not well understood, several cytokine pathways may play a role. Therefore, in order to evaluate the effect of LIPUS stimulation on cytokine production by nucleus pulposus cells and macrophages, in vitro culture studies were designed. Nucleus pulposus cells and macrophages were collected from Sprague‐Dawley rats, cultured separately in a monolayer, and stimulated with LIPUS for 7 days. After culture, the culture medium and the cells were analyzed by cytokine array, RT‐PCR, and ELISA. Cytokine array showed that LIPUS stimulation significantly upregulated TIMP‐1 (tissue inhibitor of metalloproteinase‐1) in the nucleus pulposus and MCP‐1 in macrophages in comparison with the control. This was confirmed at the gene level by RT‐PCR in nucleus pulposus cells and macrophages after stimulation with LIPUS. Quantitative evaluation of these proteins by ELISA showed higher levels in nucleus pulposus cells and macrophages stimulated by LIPUS than in controls. These results showed that LIPUS stimulation significantly activated TIMP‐1 and MCP‐1 in nucleus pulposus cells and macrophages at both the protein and gene levels, suggesting that LIPUS may be a promising supplemental treatment for intervertebral disc herniation.

Diffusion tensor imaging can detect the early stages of cartilage damage: a comparison study

BackgroundIn 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.MethodsForty-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.ResultsThe 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.ConclusionsT2 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.

Cross talk between Smad transcription factors and TNF-α in intervertebral disc degeneration

The transforming growth factor-beta (TGF-beta) and the tumor necrosis factor-alpha (TNF-alpha) families are known to play important roles in intervertebral disc degeneration (IVD). However, molecular interactions between the TGF-beta and TNF-alpha signaling pathways have yet to be elucidated. The purpose of this study was to analyze the expression patterns of Smad transcription factor signaling associated with IVDs with aging and to examine the modulation of Smad signaling by TNF-alpha in IVD cells using SD rats. According to these experimental results, BMP signals in the TGF-beta family were more likely to be a key factor in IVD degeneration by aging, and it was predicted that besides the involvement of catabolic factors like MMPs and ADAMS-TS, there may be a decrease in expression of anabolic factors through cross talk of signaling between TNF-alpha and TGF-beta pathway in pathogenesis of disc degeneration.

Modified Elmslie–Trillat Procedure for Recurrent Dislocation of the Patella

Abstract Many groups have reported good to excellent clinical outcomes of the Elmslie‐Trillat (ET) procedure for recurrent dislocation of the patella. However, the mean follow‐up period of these studies was no more than 5 years. We hypothesized that the long‐term postoperative outcomes of the modified ET procedure for recurrent dislocation of the patella would be acceptable compared with those of other procedures. The long‐term postoperative outcomes of patients treated with the modified ET procedure were examined. A total of 31 knees in 27 patients with recurrent dislocation of the patella who underwent a modified ET procedure (without medial capsular plication) were evaluated. The mean follow‐up period was 13.0 years. Pre‐ and postoperative radiographs were examined to determine the Q‐angle, tilting angle, lateral shift ratio, sulcus angle, congruence angle, and Insall‐Salvati index. Clinical outcomes were evaluated based on the Fulkerson patellofemoral joint evaluation score, Kujala score, and the presence of the apprehension sign. Radiological evaluation revealed significant postoperative improvements in the Q‐angle, tilting angle, lateral shift ratio, congruence angle, Kujala score, and Fulkerson score (p < 0.001). Sixty‐eight percent of all patients had a “good” or “excellent” Fulkerson score. The apprehension sign continued to be shown for six knees. Progression of osteoarthritic changes was observed in three knees, and one knee was symptomatic. The modified ET procedure provided satisfactory outcomes based on radiological and clinical evaluations. These results show the long‐term effectiveness and safety of the modified ET procedure for recurrent patellar dislocation.

Clinical Outcomes of Patients with Valgus Deformity Undergoing Minimally Invasive Total Knee Arthroplasty Through the Medial Approach

Objective: The purpose of this study was to compare the clinical outcomes between patients with a valgus or varus deformity undergoing minimally invasive total knee arthroplasty through the medial approach. Methods: The patients were classified into 2 groups according to the preoperative femorotibial angle measured on an anteroposterior long leg roentgenogram. The valgus group comprised of 26 knees in 21 patients with a femorotibial angle <170° (163.5 ± 5.7), and the varus group comprised of 24 knees in 21 patients with a femorotibial angle >190° (195.9 ± 5.5). The following background variables were compared between the groups: age at the time of the operation, sex, causative disease, preoperative femoral mechanical–anatomical angle, and postoperative knee range of motion, Knee Society score, femorotibial angle, and implant position. Results: There were significant differences between the valgus and varus groups in the age (68.0 ± 6.9 vs 75.8 ± 6.2 years), percentage of males (23.8% vs 0%), percentage with rheumatoid arthritis (61.9% vs 4.8%), and preoperative femoral mechanical–anatomical angle (6.2 ± 1.0° vs 7.4 ± 2.1°). Clinical outcome variables of postoperative femorotibial angle (173.1 ± 3.9° vs 175.2 ± 1.6°) and α angle (96.6 ± 3.1° vs 95.0 ± 1.9°) also differed. Conclusion: It was assumed that over-valgus resection of the femur is a contributory factor to residual valgus alignment. However, knee range of motion and Knee Society score did not differ between the groups. We suggest that minimally invasive total knee arthroplasty through the medial approach is one of the treatment options for patients with valgus deformity.

Tibia Rotational Technique to Drill Femoral Bone Tunnel in Anatomic Double-Bundle Anterior Cruciate Ligament Reconstruction

In anatomic anterior cruciate ligament (ACL) reconstruction, several pitfalls in creating the femoral bone tunnels at the correct position are of great concern. Our new method, the tibia rotational (TR) technique, may contribute to resolving these. The purpose of this study is to describe further details about the TR technique in anatomic double-bundle ACL reconstruction. Both anteromedial and posterolateral femoral bone tunnels were drilled through a posterolateral tibial bone tunnel using tibial rotation without deep knee flexion. When it is difficult to reach the mark with the rigid guide pin, the narrow curved TR technique guide and the flexible drill system allow drilling femoral bone tunnels in the correct position. The TR technique offers the technical ease required for widespread acceptance while prioritizing the fundamental goals of an anatomic double-bundle ACL reconstruction.