Effects of in vivo cyclic compressive loading on the distribution of local Col2 and superficial lubricin in rat knee cartilage

This study aimed to examine the effects of an episode of in vivo cyclic loading on rat knee articular cartilage (AC) under medium-term observation, while also investigating relevant factors associated with the progression of post-traumatic osteoarthritis (PTOA). Twelve-week-old Wistar rats underwent one episode comprising 60 cycles of 20 N or 50 N dynamic compression on the right knee joint. Spatiotemporal changes in the AC after loading were evaluated using histology and immunohistochemistry at 3 days and 1, 2, 4, and 8 weeks after loading (n = 6 for each condition). Chondrocyte vitality was assessed at 1, 3, 6, and 12 hours after loading (n = 2 for each condition). A localized AC lesion on the lateral femoral condyle was confirmed in all subjects. The surface and intermediate cartilage in the affected area degenerated after loading, but the calcified cartilage remained intact. Expression of type II collagen in the lesion cartilage was upregulated after loading, whereas the superficial lubricin layer was eroded in response to cyclic compression. However, the distribution of superficial lubricin gradually recovered to the normal level 4 weeks after loading-induced injury. We confirmed that 60 repetitions of cyclic loading exceeding 20 N could result in cartilage damage in the rat knee. Endogenous repairs in well-structured joints work well to rebuild protective layers on the lesion cartilage surface, which may be the latent factor delaying the progression of PTOA

A Non-Invasive Method for Generating the Cyclic Loading-Induced Intra-Articular Cartilage Lesion Model of the Rat Knee

The pathophysiology of primary osteoarthritis (OA) remains unclear. However, a specific subclassification of OA in relatively younger age groups is likely correlated with a history of articular cartilage damage and ligament avulsion. Surgical animal models of OA of the knee play an important role in understanding the onset and progression of post-traumatic OA and aid in the development of novel therapies for this disease. However, non-surgical models have been recently considered to avoid traumatic inflammation that could affect the evaluation of the intervention. In this study, an intra-articular cartilage lesion rat model induced by in vivo cyclic compressive loading was developed, which allowed researchers to (1) determine the optimal magnitude, speed, and duration of load that could cause focal cartilage damage; (2) assess post-traumatic spatiotemporal pathological changes in chondrocyte vitality; and (3) evaluate the histological expression of destructive or protective molecules that are involved in the adaptation and repair mechanisms against joint compressive loads. This report describes the experimental protocol for this novel cartilage lesion in a rat model

SMN promotes mitochondrial metabolic maturation during myogenesis by regulating the MYOD-miRNA axis

脊髄性筋萎縮症における骨格筋病変の発症メカニズムの一部を解明. 京都大学プレスリリース. 2023-01-17.Pathogenesis of skeletal muscle lesions in spinal muscular atrophy. 京都大学プレスリリース. 2023-02-17.Spinal muscular atrophy (SMA) is a congenital neuromuscular disease caused by the mutation or deletion of the survival motor neuron 1 (SMN1) gene. Although the primary cause of progressive muscle atrophy in SMA has classically been considered the degeneration of motor neurons, recent studies have indicated a skeletal muscle–specific pathological phenotype such as impaired mitochondrial function and enhanced cell death. Here, we found that the down-regulation of SMN causes mitochondrial dysfunction and subsequent cell death in in vitro models of skeletal myogenesis with both a murine C2C12 cell line and human induced pluripotent stem cells. During myogenesis, SMN binds to the upstream genomic regions of MYOD1 and microRNA (miR)-1 and miR-206. Accordingly, the loss of SMN down-regulates these miRs, whereas supplementation of the miRs recovers the mitochondrial function, cell survival, and myotube formation of SMN-deficient C2C12, indicating the SMN-miR axis is essential for myogenic metabolic maturation. In addition, the introduction of the miRs into ex vivo muscle stem cells derived from Δ7-SMA mice caused myotube formation and muscle contraction. In conclusion, our data revealed novel transcriptional roles of SMN during myogenesis, providing an alternative muscle-oriented therapeutic strategy for SMA patients

One session of 20 ​N cyclic compression induces chronic knee osteoarthritis in rats: A long-term study

OBJECTIVE: Mechanical stimulation is a risk factor for knee osteoarthritis. Non-surgical compression has been used to study the effects of mechanical stimulation in vivo. However, the long-term effects of low-force compression on knee joint had not been studied. Therefore, we sought to identify the long-term effects of low-force cyclic compression on the rat knee joint. DESIGN: In this study, we applied one session cyclic compression with a peak load of 20 ​N for 60 cycles to the rat knee joint in an approximately 140-degree flexion position (Wistar, male, 12 weeks old), followed by 1 year of observation (including data from 1 week, 2 weeks, 4 weeks, 8 weeks, 6 months, and 1 year after compression), and then performed a sub-regional analysis with hematoxylin-eosin, Safranin O and Fast Green, and MMP13 immunohistochemical staining. RESULTS: We observed osteoarthritis-like cartilage damage, synovial inflammation, and high expression of MMP13 within 1 year after compression. However, these changes progressed slowly, with obvious matrix cracks that did not appear until 1 year after compression. In the regional analysis, we found that low-force compression caused a much slower development of injury at the compression contact site, and no significant structural cartilage damage was observed after 1 year of compression. In contrast, the non-contact site during compression at tibial cartilage in the same joint was the first to show significant structural damage. CONCLUSION: This study demonstrates that one session of 20 ​N cyclic compression induces a chronic osteoarthritis-like phenotype in the rat knee in the long term

Grafting of iPS cell-derived tenocytes promotes motor function recovery after Achilles tendon rupture

ヒトのiPS細胞から腱の細胞を作製する --アキレス腱断裂のラットに移植し、機能回復を確認--. 京都大学プレスリリース. 2021-08-31.Repairing tendon injuries with stem cells. 京都大学プレスリリース. 2021-08-31.Tendon self-renewal is a rare occurrence because of the poor vascularization of this tissue; therefore, reconstructive surgery using autologous tendon is often performed in severe injury cases. However, the post-surgery re-injury rate is relatively high, and the collection of autologous tendons leads to muscle weakness, resulting in prolonged rehabilitation. Here, we introduce an induced pluripotent stem cell (iPSC)-based technology to develop a therapeutic option for tendon injury. First, we derived tenocytes from human iPSCs by recapitulating the normal progression of step-wise narrowing fate decisions in vertebrate embryos. We used single-cell RNA sequencing to analyze the developmental trajectory of iPSC-derived tenocytes. We demonstrated that iPSC-tenocyte grafting contributed to motor function recovery after Achilles tendon injury in rats via engraftment and paracrine effects. The biomechanical strength of regenerated tendons was comparable to that of healthy tendons. We suggest that iPSC-tenocytes will provide a therapeutic option for tendon injury

Successful treatment of unresectable advanced rectal cancer with liver metastases by hemostasis re-irradiation of the rectal cancer and palliative low-dose whole-liver radiation therapy: a case report.

A 72-year-old man was admitted to the hospital with fatigue. Colonoscopy revealed a 50 × 50 mm rectal tumor with bleeding. Based on close inspection, he was diagnosed with unresectable advanced rectal cancer with multiple liver metastases. Chemotherapy was administered as 10 cycles of bevacizumab + mFOLFOX6 and 7 cycles of bevacizumab + FOLFIRI. Nine months later, he presented with hematochezia and progression of anemia. It was difficult to stop the bleeding via endoscopy. He underwent radiation therapy (39 Gy in 13 fractions), and hemostasis was confirmed. Then, further chemotherapy was performed with 3 cycles of bevacizumab + FOLFIRI and 2 cycles of TAS102. However 14 months after the initial visit, he presented with right hypochondralgia and abdominal fullness due to the progression of multiple liver metastases. Palliative low-dose whole-liver radiation therapy (WLRT) (30 Gy in 10 fractions) was performed. He developed Grade 2 nausea, but his right hypochondralgia reduced, liver dysfunction improved, and he successfully completed radiotherapy. At approximately the same time his anemia progressed, and colonoscopy revealed recurrent bleeding from the tumor. Re-irradiation (15 Gy in 5 fractions) of the rectal tumor was carried out and a blood transfusion was performed for the bleeding. He was discharged after confirmation the anemia had not progressed. Few reports have been published on the use of both palliative re-irradiation to stop bleeding from rectal cancer and palliative low-dose WLRT. Based on our experience with this case, we believe that palliative radiotherapy can be useful in treating patients with a poor prognosis

Development of a novel model for intraarticular adhesion in rat knee joint

In this study, a novel rat model of knee joint adhesion was developed, and its formation was analyzed quantitatively over time. Thirty-nine Wistar rats were randomly divided into intact control (n = 3) and experimental (n = 36) groups. The latter was equally divided into three groups according to the experimental intervention: fixed with deep bending of the knee joint (group I), fixed after incision of the capsule (group II), and fixed after exposure of the patellofemoral joint to artificial patellar subluxation (group III). All rats were subdivided according to their joint immobilization period (1, 2, or 4 weeks). Thereafter, the limited range of motion of the knee joint with (limited knee range of motion) and without (limited knee joint intrinsic range of motion) skin and muscles were measured. The lengths of adhesions of the anterior knee joint and posterior capsules were evaluated histologically. The limited intrinsic range of motion of the knee joint was found to be increased in groups II and III compared to that in group I 4 weeks after immobilization. Adhesions were confirmed within 1 week after immobilization in groups II and III. The length of the adhesions in group III was significantly longer than in other groups at 2 weeks and remained longer than in group I at 4 weeks. This model may contribute to the assessment of the adhesion process and development of new therapeutic avenues following trauma or surgical invasion

N-Acetylcysteine prevents amyloid-β secretion in neurons derived from human pluripotent stem cells with trisomy 21

ダウン症患者さん由来の神経細胞からのアミロイドβ分泌は抗酸化剤で抑止される. 京都大学プレスリリース. 2021-08-31.Stopping dementia in Down syndrome patients. 京都大学プレスリリース. 2021-08-31.Down syndrome (DS) is caused by the trisomy of chromosome 21. Among the many disabilities found in individuals with DS is an increased risk of early-onset Alzheimer's disease (AD). Although higher oxidative stress and an upregulation of amyloid β (Aβ) peptides from an extra copy of the APP gene are attributed to the AD susceptibility, the relationship between the two factors is unclear. To address this issue, we established an in vitro cellular model using neurons differentiated from DS patient-derived induced pluripotent stem cells (iPSCs) and isogenic euploid iPSCs. Neurons differentiated from DS patient-derived iPSCs secreted more Aβ compared to those differentiated from the euploid iPSCs. Treatment of the neurons with an antioxidant, N-acetylcysteine, significantly suppressed the Aβ secretion. These findings suggest that oxidative stress has an important role in controlling the Aβ level in neurons differentiated from DS patient-derived iPSCs and that N-acetylcysteine can be a potential therapeutic option to ameliorate the Aβ secretion

A VCP modulator, KUS121, as a promising therapeutic agent for post-traumatic osteoarthritis

京大開発の薬剤「KUS121」の変形性膝関節症への効果を確認 --外傷性変形性関節症の治療薬として臨床応用へ--. 京都大学プレスリリース. 2020-12-17.Post-traumatic osteoarthritis (PTOA) is a major cause which hinders patients from the recovery after intra-articular injuries or surgeries. Currently, no effective treatment is available. In this study, we showed that inhibition of the acute stage chondrocyte death is a promising strategy to mitigate the development of PTOA. Namely, we examined efficacies of Kyoto University Substance (KUS) 121, a valosin-containing protein modulator, for PTOA as well as its therapeutic mechanisms. In vivo, in a rat PTOA model by cyclic compressive loading, intra-articular treatments of KUS121 significantly improved the modified Mankin scores and reduced damaged-cartilage volumes, as compared to vehicle treatment. Moreover, KUS121 markedly reduced the numbers of TUNEL-, CHOP-, MMP-13-, and ADAMTS-5-positive chondrocytes in the damaged knees. In vitro, KUS121 rescued human articular chondrocytes from tunicamycin-induced cell death, in both monolayer culture and cartilage explants. It also significantly downregulated the protein or gene expression of ER stress markers, proinflammatory cytokines, and extracellular-matrix-degrading enzymes induced by tunicamycin or IL-1β. Collectively, these results demonstrated that KUS121 protected chondrocytes from cell death through the inhibition of excessive ER stress. Therefore, KUS121 would be a new, promising therapeutic agent with a protective effect on the progression of PTOA

Reproducibility and validity of food group intake in a short food frequency questionnaire for the middle-aged Japanese population

Purpose: The purpose of this study was to evaluate the reproducibility and validity of a short food frequency questionnaire (FFQ) for food group intake in Japan, the reproducibility and partial validity of which were previously confirmed for nutrients. Methods: A total of 288 middle-aged healthy volunteers from 11 different areas of Japan provided nonconsecutive 3-day weighed dietary records (DRs) at 3-month intervals over four seasons. We evaluated reproducibility based on the first (FFQ1) and second (FFQ2) questionnaires and their validity against the DRs by comparing the intake of 20 food groups. Spearman’s rank correlation coefficients (SRs) were calculated between energy-adjusted intake from the FFQs and that from the DRs. Results: The intake of 20 food groups estimated from the two FFQs was mostly equivalent. The median energy-adjusted SRs between the FFQ1 and FFQ2 were 0.61 (range 0.38–0.86) for men and 0.66 (0.45–0.84) for women. For validity, the median de-attenuated SRs between DRs and the FFQ1 were 0.51 (0.17–0.76) for men and 0.47 (0.23–0.77) for women. Compared with the DRs, the proportion of cross-classification into exact plus adjacent quintiles with the FFQ1 ranged from 58 to 86% in men and from 57 to 86% in women. According to the robust Z scores and the Bland–Altman plot graphs, the underestimation errors in the FFQ1 tended to be greater in individuals with high mean levels of consumption for meat for men and for other vegetables for both men and women. Conclusion: The FFQ demonstrated high reproducibility and reasonable validity for food group intake. This questionnaire is short and remains appropriate for identifying associations between diet and health/disease among adults in Japan