Nobuaki Chinzei

Oxidative stress-induced apoptosis and matrix loss of chondrocytes is inhibited by eicosapentaenoic acid

Eicosapentaenoic acid (EPA) is an antioxidant and n‐3 polyunsaturated fatty acid that reduces the production of inflammatory cytokines. We evaluated the role of EPA in chondrocyte apoptosis and degeneration. Normal human chondrocytes were treated with EPA and sodium nitroprusside (SNP). Expression of metalloproteinases (MMPs) was detected by real‐time polymerase chain reaction (PCR) and that of apoptosis‐related proteins was detected by western blotting. Chondrocyte apoptosis was detected by flow cytometry. C57BL/6J mice were used for the detection of MMP expression by immunohistochemistry and for investigation of chondrocyte apoptosis. EPA inhibited SNP‐induced chondrocyte apoptosis, caspase 3 and poly(ADP‐ribose) polymerase cleavage, phosphorylation of p38 MAPK and p53, and expression of MMP3 and MMP13. Intra‐articular injection of EPA prevented the progression of osteoarthritis (OA) by inhibiting MMP13 expression and chondrocyte apoptosis. EPA treatment can control oxidative stress‐induced OA progression, and thus may be a new approach for OA therapy.

Satisfactory results at 8 years mean follow-up after ADVANCE® medial-pivot total knee arthroplasty.

BACKGROUND Although good overall results have been reported with TKA, certain problems and limitations remain, primarily due to postoperative differences in joint kinematics, when compared with the normal knee. ADVANCE® Medial-Pivot TKA involves replicating the medial pivoting behavior observed in normal knees. Here, we aimed to investigate the clinical and radiological results and complications of TKA using this implant, at mid-term follow-up. METHODS From January 2001 to March 2012, we retrospectively selected 76 patients (85 knees; mean age at operation, 70.2±8.1 years; range, 51-88 years) with a mean follow-up period of 93.1±14.3 months (range, 72-132 months). Indications for TKA included primary degenerative osteoarthritis (60 knees), rheumatoid arthritis (22 knees), osteonecrosis (two knees), and osteoarthritis following high tibial osteotomy (one knee). The clinical and radiographic results were evaluated. RESULTS Kaplan-Meier survivorship analysis indicated a success rate of 98.3% (95% confidence interval, 96.6-99.9%). Comparison of pre- and postoperative knee extension angles and ranges of motion showed significant improvement postoperatively, in both the Knee Society Scores (KSS) and Knee Society Functional Scores (KSFS) (p<0.05). In one case, radiographic assessment indicated implant loosening due to infection; however, despite this complication, significant improvement of postoperative varus or valgus deformity angles were noted in all cases (p<0.05). CONCLUSION Patients undergoing ADVANCE® Medial-Pivot TKA achieved excellent clinical and radiographic results without any implant-related failures at mid-term follow-up. LEVEL OF EVIDENCE Level IV.

Evaluation of patellofemoral joint in ADVANCE® Medial-pivot total knee arthroplasty

PurposeADVANCE® Medial Pivot (MP) (Wright Medical) total knee arthroplasty (TKA) was established to replicate normal tibio-femoral knee joint kinematics, however, its influence on the patello-femoral (PF) joint is unclear. The purpose in this study was to assess the PF joint conditions in Advance MP TKA, via radiography and three-dimensional image-matching software.MethodsTen subjects with osteoarthritis were treated with the ADVANCE MP TKA. Pre-operatively and one month after surgery, skyline views at 30, 60, and 90° of flexion were taken, and patella shift and tilt were measured. With 2D–3D registration techniques using software, implant orientations were matched with the pre-operative CT and changes in the anterior part of the femoral prosthesis, condylar twist angle (CTA) for femoral rotation, and tibial rotation were evaluated. The relationships between morphological and rotational changes were evaluated.ResultsThere were significant differences in patella tilt at 60° and patella shift at all angles between pre- and post-operation (p < 0.05). No correlation was found between morphological changes in the anterior femur with patella tilt and shift. A positive correlation between postoperative CTA and patella shift at 90° was found (p < 0.05); however, no correlation was found between rotational alignment of the tibial component and patella tilt and shift.ConclusionsADVANCE MP TKA changed patello-femoral joint kinematics, compared to that found before surgery. The kinematic features were mainly due to the design concepts for tibio-femoral joint motion, indicating the difficulty to reproduce normal patello-femoral joint kinematics after TKA.

P21 Deficiency Delays Regeneration of Skeletal Muscular Tissue

The potential relationship between cell cycle checkpoint control and tissue regeneration has been indicated. Despite considerable research being focused on the relationship between p21 and myogenesis, p21 function in skeletal muscle regeneration remains unclear. To clarify this, muscle injury model was recreated by intramuscular injection of bupivacaine hydrochloride in the soleus of p21 knockout (KO) mice and wild type (WT) mice. The mice were sacrificed at 3, 14, and 28 days post-operation. The results of hematoxylin-eosin staining and immunofluorescence of muscle membrane indicated that muscle regeneration was delayed in p21 KO mice. Cyclin D1 mRNA expression and both Ki-67 and PCNA immunohistochemistry suggested that p21 deficiency increased cell cycle and muscle cell proliferation. F4/80 immunohistochemistry also suggested the increase of immune response in p21 KO mice. On the other hand, both the mRNA expression and western blot analysis of MyoD, myogenin, and Pax7 indicated that muscular differentiation was delayed in p21KO mice. Considering these results, we confirmed that muscle injury causes an increase in cell proliferation. However, muscle differentiation in p21 KO mice was inhibited due to the low expression of muscular synthesis genes, leading to a delay in the muscular regeneration. Thus, we conclude that p21 plays an important role in the in vivo healing process in muscular injury.

PTEN regulates matrix synthesis in adult human chondrocytes under oxidative stress.

Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) was identified as an important tumor suppressor gene. PTEN functions as a negative regulator of phosphoinositol‐3‐kinase (PI3K)‐Akt and MEK/ERK signaling. The PI3K‐Akt pathway is critical for cell survival, differentiation, and matrix synthesis. Oxidative stress is considered a critical factor in the onset and progression of osteoarthritis (OA). Therefore, we investigated the function of PTEN in OA chondrocytes under oxidative stress. Chondrocytes were treated with insulin‐like growth factor‐1 (IGF‐1) and/or tert‐butyl hydroperoxide (tBHP), which causes oxidative stress. The expression levels of type2 collagen (Col2a1) and aggrecan were analyzed by real‐time PCR, and phosphorylation of Akt and ERK1/2 was analyzed by Western blotting. Chondrocytes were treated with PTEN‐specific small interfering RNA (siRNA), as well as IGF‐1 and/or tBHP. PTEN and IGF‐1 expressions in OA chondrocytes were increased. The downregulation of PTEN expression increased the expression levels of Col2a1 and aggrecan, and increased proteoglycan synthesis under oxidative stress. Oxidative stress decreased the phosphorylation of Akt and increased that of ERK1/2. The downregulation of PTEN expression increased Akt phosphorylation, but did not increase that of ERK 1/2. Our results suggest that PTEN regulates matrix synthesis via the PI3K‐Akt pathway under oxidative stress.

Inflammation and Degeneration in Cartilage Samples from Patients with Femoroacetabular Impingement

BACKGROUND Femoroacetabular impingement (FAI) has been reported as a cause of hip pain in young patients and is suggested as the trigger for hip osteoarthritis (OA). The goal of this study was to quantify the metabolic profiles of articular tissues (cartilage, synovium, and labrum) harvested from patients with FAI and with end-stage OA. In addition, we sought to investigate the development of secondary OA in hips with FAI. METHODS Tissue samples were obtained from thirty hips undergoing arthroscopic surgery for FAI with or without labral tear and thirty hips undergoing total hip arthroplasty for OA. Quantitative real-time polymerase chain reaction (PCR) was performed to determine the gene expression of inflammatory cytokines and metabolic (anabolic and catabolic) enzymes. The differences in gene expression in articular tissues from the patients with FAI were also evaluated on the basis of clinical parameters (age range and alpha angle). RESULTS The messenger RNA (mRNA) expression of the inflammatory cytokines interleukin-1 beta (IL-1β) and IL-8 and of matrix metalloproteinase (MMP)-3 (a catabolic gene) in both the synovium and the labrum, and the expression of collagen type I alpha 1 (an anabolic gene) in the labrum, was higher in the samples from hips with OA than in those from hips with FAI (p < 0.05). In cartilage, however, the mRNA expression of the inflammatory cytokines and the catabolic genes MMP-13 and ADAMTS-4 (a disintegrin and metalloproteinase with thrombospondin motifs-4) was higher in the FAI samples compared with the OA samples (p < 0.01). When the expression of inflammatory cytokines was evaluated among the three types of tissues within each disease group, the expression levels were the highest in cartilage within the FAI samples (p < 0.01). In FAI cartilage, we found higher gene expression of aggrecan (ACAN) and ADAMTS-4 in the samples from patients with larger alpha angles (≥60°) (p < 0.01). CONCLUSIONS Our results indicate that the metabolic conditions of articular cartilage in FAI and OA are different and that the expression of genes associated with inflammation is greater in the articular cartilage of patients with FAI compared with the synovium and the labrum. The metabolic changes were heightened by mechanical impingement. CLINICAL RELEVANCE The articular cartilage from the impingement lesion in patients with FAI showed biologically higher inflammation and degeneration, supporting the concept that FAI may be a trigger for joint degeneration.

Post-Traumatic Osteoarthritis in Mice Following Mechanical Injury to the Synovial Joint

We investigated the spectrum of lesions characteristic of post-traumatic osteoarthritis (PTOA) across the knee joint in response to mechanical injury. We hypothesized that alteration in knee joint stability in mice reproduces molecular and structural features of PTOA that would suggest potential therapeutic targets in humans. The right knees of eight-week old male mice from two recombinant inbred lines (LGXSM-6 and LGXSM-33) were subjected to axial tibial compression. Three separate loading magnitudes were applied: 6N, 9N, and 12N. Left knees served as non-loaded controls. Mice were sacrificed at 5, 9, 14, 28, and 56 days post-loading and whole knee joint changes were assessed by histology, immunostaining, micro-CT, and magnetic resonance imaging. We observed that tibial compression disrupted joint stability by rupturing the anterior cruciate ligament (except for 6N) and instigated a cascade of temporal and topographical features of PTOA. These features included cartilage extracellular matrix loss without proteoglycan replacement, chondrocyte apoptosis at day 5, synovitis present at day 14, osteophytes, ectopic calcification, and meniscus pathology. These findings provide a plausible model and a whole-joint approach for how joint injury in humans leads to PTOA. Chondrocyte apoptosis, synovitis, and ectopic calcification appear to be targets for potential therapeutic intervention.

p21 deficiency is susceptible to osteoarthritis through STAT3 phosphorylation

IntroductionOsteoarthritis (OA) is a multifactorial disease, and recent studies have suggested that cell cycle–related proteins play a role in OA pathology. p21 was initially identified as a potent inhibitor of cell cycle progression. However, it has been proposed that p21 is a regulator of transcription factor activity. In this study, we evaluated the role of p21 in response to biomechanical stress.MethodsHuman chondrocytes were treated with p21-specific small interfering RNA (siRNA), and cyclic tensile strain was introduced in the presence or absence of a signal transducer and activator of transcription 3 (STAT3)-specific inhibitor. Further, we developed an in vivo OA model in a p21-knockout background for in vivo experiments.ResultsThe expression of matrix metalloproteinase (MMP13) mRNA increased in response to cyclic tensile strain following transfection with p21 siRNA, whereas the expression of aggrecan was decreased. Phospho-STAT3 and MMP-13 protein levels increased following downregulation of p21, and this was reversed by treatment with a STAT3 inhibitor. p21-deficient mice were susceptible to OA, and this was associated with increased STAT3 phosphorylation, elevated MMP-13 expression, and elevation of synovial inflammation. The expression of p21 mRNA was decreased and phosphorylation of STAT3 was elevated in human OA chondrocytes.ConclusionsThe lack of p21 has catabolic effects by regulation of aggrecan and MMP-13 expression through STAT3 phosphorylation in the cartilage tissue. p21 may function as a regulator of transcriptional factors other than the inhibitor of cell cycle progression in the cartilage tissue. Thus, the regulation of p21 may be a therapeutic strategy for the treatment of OA.

Comparison of the Sliding and Femoral Head Rotation among Three Different Femoral Head Fixation Devices for Trochanteric Fractures

Background Recently, various femoral head fixation devices (HFDs) for trochanteric fractures have become available. However, there are some cases in which femoral head rotation with excessive sliding of the HFD is observed and it is often followed by cutout. The purpose of this study is to compare the ability of the three types of HFDs to prevent femoral head rotation. Methods Between July 2005 and December 2009, 206 patients aged over 60 years with trochanteric fractures who had undergone surgical treatment using a short femoral nail in our institution were enrolled into the study. We used the gamma 3 nail (GMN) as the screw-type HFD in 66 cases, the gliding nail (GLN) as a non-cylindrical blade in 76 cases, and the proximal femoral nail antirotation (PFNA) as a cylindrical blade in 64 cases. The sliding length of HFDs and the occurrence of femoral head rotation were evaluated by assessing radiographs as the main outcome, and the results were compared among these devices. Results A comparison of the degree of sliding in the GMN group showed that femoral head rotation was observed significantly more frequently in cases with rotation. Further, it appeared that femoral head rotation occurred more frequently in comminuted fractures. However, no significant differences between the sliding lengths of the different HFDs were observed among three groups. Femoral head rotation was observed in 15 cases of GMN (22.7%), 0 case of GLN, and 5 case of PFNA (7.8%). Significant differences with regard to the occurrence of femoral head rotation were observed among the three groups. Furthermore, significant differences were also observed between GLN and PFNA with respect to the occurrence of femoral head rotation. Conclusions The ability to stabilize femoral head appears to be greater with blade-type materials than with screw-type materials. Furthermore, we believe that a non-cylindrical blade is preferable to a cylindrical blade for the surgical treatment of comminuted, unstable trochanteric fractures in order to prevent femoral head rotation and cut-out.

Accurate and Easy Measurement of Sliding Distance of Intramedullary Nail in Trochanteric Fracture

Background In daily clinical practice, it is essential to properly evaluate the postoperative sliding distance of various femoral head fixation devices (HFD) for trochanteric fractures. Although it is necessary to develop an accurate and reproducible method that is unaffected by inconsistent postoperative limb position on radiography, few studies have examined which method is optimal. Therefore, the purpose of this study is to prospectively compare the accuracy and reproducibility of our four original methods in the measurement of sliding distance of the HFD. Methods Radiographs of plastic simulated bone implanted with Japanese proximal femoral nail antirotation were taken in five limb postures: neutral, flexion, minute internal rotation, greater external rotation, and flexion with external rotation. Orthopedic surgeons performed five measurements of the sliding distance of the HFD in each of the flowing four methods: nail axis reference (NAR), modified NAR, inner edge reference, and nail tip reference. We also assessed two clinical cases by using these methods and evaluated the intraclass correlation coefficients. Results The measured values were consistent in the NAR method regardless of limb posture, with an even smaller error when using the modified NAR method. The standard deviation (SD) was high in the nail tip reference method and extremely low in the modified NAR method. In the two clinical cases, the SD was the lowest in the modified NAR method, similar to the results using plastic simulated bone. The intraclass correlation coefficients showed the highest value in the modified NAR method. Conclusions We conclude that the modified NAR method should be the most recommended based on its accuracy, reproducibility, and usefulness.