Published papers

BACKGROUND: The membrane components of cardiomyocytes are rich in polyunsaturated fatty acids, which are easily oxidized. Thus, an efficient glutathione-based lipid redox system is essential for maintaining cellular functions. However, the relationship between disruption of the redox system during ischemia-reperfusion (IR), oxidized lipid production, and consequent cell death (ferroptosis) remains unclear. We investigated the mechanisms underlying the disruption of the glutathione-mediated reduction system related to ferroptosis during IR and developed intervention strategies to suppress ferroptosis. METHODS: In vivo fluctuations of both intra- and extracellular metabolite levels during IR were explored via microdialysis and tissue metabolome analysis. Oxidized phosphatidylcholines were assessed using liquid chromatography high-resolution mass spectrometry. The areas at risk following IR were assessed using triphenyl-tetrazolium chloride/Evans blue stain. RESULTS: Metabolomic analysis combined with microdialysis revealed a significant release of glutathione from the ischemic region into extracellular spaces during ischemia and after reperfusion. The release of glutathione into extracellular spaces and a concomitant decrease in intracellular glutathione concentrations were also observed during anoxia-reperfusion in an in vitro cardiomyocyte model. This extracellular glutathione release was prevented by chemical inhibition or genetic suppression of glutathione transporters, mainly MRP1 (multidrug resistance protein 1). Treatment with MRP1 inhibitor reduced the intracellular reactive oxygen species levels and lipid peroxidation, thereby inhibiting cell death. Subsequent in vivo evaluation of endogenously oxidized phospholipids following IR demonstrated the involvement of ferroptosis, as levels of multiple oxidized phosphatidylcholines were significantly elevated in the ischemic region 12 hours after reperfusion. Inhibition of the MRP1 transporter also alleviated intracellular glutathione depletion in vivo and significantly reduced the generation of oxidized phosphatidylcholines. Administration of MRP1 inhibitors significantly attenuated infarct size after IR injury. CONCLUSIONS: Glutathione was released continuously during IR, primarily in an MRP1-dependent manner, and induced ferroptosis. Suppression of glutathione release attenuated ferroptosis and reduced myocardial infarct size following IR.

Background: Atrial septal defect (ASD) increases the risk of adverse cardiovascular outcomes. Despite the potential for risk mitigation through minimally invasive percutaneous closure, ASD remains underdiagnosed due to subtle symptoms and examination findings. To bridge this diagnostic gap, we propose a novel screening strategy aimed at early detection and enhanced diagnosis through the implementation of a convolutional neural network (CNN) to identify ASD from 12-lead electrocardiography (ECG). Methods: ECGs were collected from patients with at least one recorded echocardiogram at 3 hospitals from 2 continents (Keio University Hospital from July 2011 to December 2020, Brigham and Women's Hospital from January 2015 to December 2020, and Dokkyo Medical University Saitama Medical Center from January 2010 and December 2021). ECGs from patients with a diagnosis of ASD were labeled as positive cases while the remainder were labeled as negative. ECGs after the closure of ASD were excluded. After randomly splitting the ECGs into 3 datasets (50% derivation, 20% validation, and 30% test) with no patient overlap, a CNN-based model was trained using the derivation datasets from 2 hospitals and was tested on held-out datasets along with an external validation on the 3rd hospital. All eligible ECGs were used for derivation and validation whereas the earliest ECG for each patient was used for the test and external validation. The discrimination of ASD was assessed by the area under the receiver operating characteristic curve (AUROC). Multiple subgroups were examined to identify any heterogeneity. Findings: A total of 671,201 ECGs from 80,947 patients were collected from the 3 institutions. The AUROC for detecting ASD was 0.85–0.90 across the 3 hospitals. The subgroup analysis showed excellent performance across various characteristics Screening simulation using the model greatly increased sensitivity from 80.6% to 93.7% at specificity 33.6% when compared to using overt ECG abnormalities. Interpretation: A CNN-based model using 12-lead ECG successfully identified the presence of ASD with excellent generalizability across institutions from 2 separate continents. Funding: This work was supported by research grants from JST ( JPMJPF2101), JSR corporation, Taiju Life Social Welfare Foundation, Kondou Kinen Medical Foundation, Research fund of Mitsukoshi health and welfare foundation, Tokai University School of Medicine Project Research and Internal Medicine Project Research, Secom Science and Technology Foundation, and Grants from AMED ( JP23hma922012 and JP23ym0126813). This work was partially supported by One Brave Idea, co-funded by the American Heart Association and Verily with significant support from AstraZeneca and pillar support from Quest Diagnostics.

The efficacy of extensive balloon pulmonary angioplasty (BPA) beyond hemodynamic improvement in chronic thromboembolic pulmonary hypertension (CTEPH) patients has been verified. However, the relationship between extensive BPA in CTEPH patients after partial hemodynamic improvement and exercise tolerance or quality of life (QOL) remains unclear. We prospectively enrolled 22 CTEPH patients (66±10 years, females: 59%) when their mean pulmonary artery pressure initially decreased to <30 mmHg during BPA sessions. Hemodynamic and echocardiographic data, cardiopulmonary exercise testing, and QOL scores using the 36-item short form questionnaire (SF-36) were evaluated at enrollment (entry), just after the final BPA session (finish), and at the 6-month follow-up (followup). We analyzed whether extensive BPA improves exercise capacity and QOL scores over time. Moreover, the clinical characteristics leading to improvement were elucidated. The peak oxygen uptake (VO2) showed significant improvement at entry, finish, and follow-up (17.3±5.5, 18.4±5.9, and 18.9±5.3 mL/kg/min, respectively; P<0.001). Regarding the QOL, the physical component summary (PCS) scores significantly improved (32±11, 38±13, and 43±13, respectively; P<0.001), but the mental component summary scores remained unchanged. Linear regression analysis revealed that age and a low peak VO2 at entry were predictors of improvement in peak VO2, while low PCS scores and low TAPSE at entry were predictors of improvement in PCS scores. In conclusion, extensive BPA led to improved exercise tolerance and physical QOL scores, even in CTEPH patients with partially improved hemodynamics.

Objectives: The minimum body mass index (BMI) required to menarche and the relationship between the training onset age of sports and menarche is not fully understood. The aim of this study is to elucidate the minimum BMI required to attain menarche in female adolescent athletes competing at a national level, and to determine how the occurrence of menarche is associated with training onset age in track-and-field athletes. Methods: Overall, 134 sprinters and 44 long-distance (LD) runners of ninth-grade females at a national level were enrolled and an anonymous questionnaire was administered. Results: As BMI increased, the proportion of athletes who had attained menarche increased. The BMI cutoff values for menarche were 17.3 and 17.1 kg/m2 for sprinters and LD runners, respectively. Menarche had not occurred in almost 50% of the LD runners who began training at elementary school, and among LD runners, those who began training at elementary school had 18.4 higher odds of not attaining menarche until the age of 15 years as opposed to those who started training after elementary school. Conclusions: The BMI cutoff values could be an indicator for menarche in sprinters and LD runners. For LD runners, starting to compete at elementary school could be a risk factor for delayed menarche.

Low energy availability in female athletes often causes hypothalamic amenorrhea and osteoporosis, in turn promoting stress fractures. Mechanisms underlying these conditions remain unclear. Here we show that model mice subjected to food restriction (FR) or FR-plus-voluntary running exercise exhibit significantly reduced bone mineral density, cortical bone parameters and uterine weight than do control mice, and that these parameters worsen in the FR-plus-exercise group. Relative to controls, FR and FR-plus-exercise groups showed significantly lower mineral apposition rate and osteoclast number and significantly reduced serum insulin-like growth factor-1 (IGF1) levels. Outcomes were rescued by ED71 or 1.25(OH)2D3 treatment. Thus, we conclude that administration of active vitamin D analogues represents a possible treatment to prevent these conditions.

Definitive diagnosis of familial hypercholesterolemia (FH) is paramount for the risk management of patients and their relatives. The present study aimed to investigate the frequency of gene variants contributing to low-density lipoprotein cholesterol (LDL-C) metabolism and their clinical relevance in patients with early-onset coronary artery disease (EOCAD). Among 63 consecutive patients with EOCAD (men <55 years or women <65 years) who underwent percutaneous coronary intervention (PCI) from 2013 to 2019 at Keio University Hospital, 52 consented to participate in this retrospective study. Targeted sequencing of LDLR, PCSK9, APOB, and LDLRAP1 was performed. Of the 52 patients enrolled (42 men; mean age: 50 ± 6 years), one (LDLR, c.1221_1222delCGinsT) harbored a pathogenic mutation, and one (APOB, c.10591A>G) harbored variants of uncertain significance. Both the patients harboring the variants were male, showing no history of diabetes mellitus or chronic kidney disease, no family history of EOCAD, and no physical findings of FH (i.e., tendon xanthomas or Achilles tendon thickening). Patients harboring the LDLR variant had three-vessel disease, were on a statin prescription at baseline, and had stable LDL-C levels; however, the case showed a poor response to the intensification of medication after PCI. Approximately 3.8% of patients with EOCAD harbored variants of gene related to LDL-C metabolism; there were no notable indicators in the patients' background or clinical course to diagnose FH. Given the difficulty in diagnosing FH based on clinical manifestations and family history, genetic testing could enable the identification of hidden risk factors and provide early warnings to their relatives.

Objective: CT imaging precisely and quantitatively analyzes the kinematics of the carpal bones to evaluate the etiology of related osteoarthritis. Previous studies have investigated the kinematics of the trapeziometacarpal joint using static CT scans of various postures including the pinch position. This study analyzed the in-vivo kinematics of the trapeziometacarpal joint during dynamic pinch motion in young healthy volunteers using four-dimensional CT. Materials and methods: Twelve healthy young volunteers participated in this study. Each participant held the pinch meter between their thumb and index finger and pinched it with maximum force for a period of 6 s. This series of movement was recorded using a four-dimensional CT. The surface data of the trapezium and first metacarpal of all frames were reconstructed, and bone movement at the trapeziometacarpal joint was calculated using sequential three-dimensional registration. The instantaneous pinch force of each frame was measured using a pointer on a pinch meter that was reconstructed from the CT data. Results: The first metacarpal was abducted (15.9 ± 8.3°) and flexed (12.2 ± 7.1°) relative to the trapezium, and significantly translated to the volar　(0.8 ± 0.6 mm) and ulnar directions (0.9 ± 0.8 mm) with maximum pinch force. This movement consistently increased with the pinch force. Conclusion: This study successfully employed 4D-CT to precisely demonstrate changes in rotation and translation at the trapeziometacarpal joint during pinch motion for various instantaneous forces.

Background: The association between Patient-Rated Elbow Evaluation: Japanese version (PREE-J) and Japanese Orthopaedic Association-Japan Elbow Society Elbow Function score (JOA-JES score) is unclear. This study evaluated the association between PREE-J and JOA-JES scores. Methods: The patients with elbow disorders were divided into two groups: Group A (conservative treatment, n = 97) and Group B (surgical treatment, n = 156). The patients were also divided into four disease subgroups according to the JOA-JES classification (rheumatoid arthritis, trauma, sports, and epicondylitis groups), and the association between PREE-J and JOA-JES scores in each disease category was examined. In group B, associations between PREE-J and JOA-JES scores were examined pre-and postoperatively. Results: In group A, there were significant associations between PREE-J and JOA-JES scores. In group B, a strong association between preoperative PREE-J and JOA-JES scores was observed in all disease categories. There was also a significant association between postoperative PREE-J and JOA-JES scores. Additionally, group B showed significant postoperative improvements in PREE-J and JOA-JES scores. Conclusions: The PREE-J score correlates well with the JOA-JES score and reflects treatment response before and after treatment.

Particulate generation occurs during exercise-induced exhalation, and research on this topic is scarce. Moreover, infection-control measures are inadequately implemented to avoid particulate generation. A laminar airflow ventilation system (LFVS) was developed to remove respiratory droplets released during treadmill exercise. This study aimed to investigate the relationship between the number of aerosols during training on a treadmill and exercise intensity and to elucidate the effect of the LFVS on aerosol removal during anaerobic exercise. In this single-center observational study, the exercise tests were performed on a treadmill at Running Science Lab in Japan on 20 healthy subjects (age: 29±12 years, men: 80%). The subjects had a broad spectrum of aerobic capacities and fitness levels, including athletes, and had no comorbidities. All of them received no medication. The exercise intensity was increased by 1-km/h increments until the heart rate reached 85% of the expected maximum rate and then maintained for 10 min. The first 10 subjects were analyzed to examine whether exercise increased the concentration of airborne particulates in the exhaled air. For the remaining 10 subjects, the LFVS was activated during constant-load exercise to compare the number of respiratory droplets before and after LFVS use. During exercise, a steady amount of particulates before the lactate threshold (LT) was followed by a significant and gradual increase in respiratory droplets after the LT, particularly during anaerobic exercise. Furthermore, respiratory droplets ≥0.3 μm significantly decreased after using LFVS (2120800±759700 vs. 560 ± 170, p<0.001). The amount of respiratory droplets significantly increased after LT. The LFVS enabled a significant decrease in respiratory droplets during anaerobic exercise in healthy subjects. This study's findings will aid in exercising safely during this pandemic.

Introduction: In women, the female athlete triad, marked by low energy availability, functional hypothalamic amenorrhea and osteoporosis, is a recognized risk for stress fractures. Stress injuries also occur in men, but by contrast risks and mechanisms underlying them are less characterized. Materials and methods: 5 week-old wild-type male mice were fed ad libitum (ad) or subjected to 60% food restriction (FR) for five weeks. In both groups, some mice were allowed access to an exercise wheel in cages to allow voluntary wheel running (ex) and/or treated with active vitamin D analogues. Mice were sacrificed and analyzed at 10 weeks of age. Result: Male FR mice exhibited significantly reduced testicle weight, serum testosterone levels and bone mass. Such bone losses in FR male mice were enhanced by exercise. Histological analysis revealed that both bone-resorbing and -forming activities were significantly reduced in FR or FR plus exercise (FR + ex) mice, mimicking a state of low bone turnover. Significantly reduced bone mass in FR or FR + ex male mice was significantly rescued by treatment with active vitamin D analogues, with significant restoration of osteoblastic activities. Serum levels of insulin-like growth factor I (IGF-I), which is critical for bone remodeling, were significantly lower in FR versus control male mice. Conclusions: Low energy availability puts men at risk for stress injuries as well, and low energy availability is upstream of gonadal dysfunction and osteoporosis in males. Active vitamin D analogues could serve as therapeutic or preventive options for stress injuries in men.

Background: The midcarpal joint and the radiocarpal joint contribute to the extension and flexion of the wrist. Little is known about the contribution of the distal radioulnar joint (DRUJ) to the extension and flexion of the wrist. This study evaluated the ulnar motion in extension and flexion of the wrist using computed tomography (CT) imaging. Methods: A total of 30 wrists of healthy volunteers were enrolled. CT images of the axial sections of the DRUJ were obtained with 3 different positions of the wrist: 0° of extension (straight position), maximum active extension, and maximum active flexion. Each wrist motion was performed with 3 different forearm positions: neutral, pronation, and supination. Ulnar position at the DRUJ level was measured and ulnar position with the wrist in straight position was defined as baseline. The ulnar position was recorded as positive value when the position of the ulnar head was volar side and negative value when the position of the ulnar head was dorsal side. The difference from baseline in a position of maximum extension and flexion was evaluated. Results: In forearm neutral position and pronation, a value of ulnar position in maximum wrist flexion is significantly negative compared to that in the wrist straight position: the ulnar head moved dorsally from the wrist straight position to wrist flexion. In forearm supination, a value of ulnar position in maximum wrist extension is significantly positive compared to that in the wrist straight position: the ulnar head moved to the volar side from the wrist straight position to wrist extension. Conclusions: The ulnar head moves during extension and flexion of the wrist. The direction of the ulnar motion was different according to the wrist and forearm position.

The lateral capitello-humeral angle (LCHA), which is an index of sagittal alignment of the elbow, has gradually been adopted for the postoperative assessment of radiographic results. However, the normal values and ranges of the LCHA remain unclear. A retrospective cohort study was performed to evaluate the normal values and ranges of the LCHA in a sample of healthy children with even distributions of age, sex and laterality. A total of 168 radiographs of the elbows of healthy children (age range, 0–11 years) with even distributions of age, sex and laterality were reviewed. The primary aim was to analyze the normal values and ranges of the LCHA categorized by age, sex and laterality. The secondary aim was to assess the association of the LCHA with increasing age. The LCHA between sex or laterality in each age category was also compared. The mean LCHA of the 168 patients was 47.1º(range, 27º–63º). There was a weak association between the LCHA and increasing age (r = 0.41). The mean LCHA in females (49.1º) was significantly larger than that in males (45.1º). Significant sex-related differences were observed in age categories between 2 and 7 years. Results of this study will be useful in the postoperative radiographic assessment of sagittal alignment of the elbow in children.

In the human body, skeletal muscle microstructures have been evaluated only by biopsy. Noninvasive examination of the microstructure of muscles would be useful for research and clinical practice in sports and musculoskeletal areas. The study is aimed at determining if q-space imaging (QSI) can reveal the microstructure of muscles in humans. Forty-three Japanese subjects (controls, distance runners, powerlifting athletes, and teenage runners) were included in this cross-sectional study. Magnetic resonance imaging of the lower leg was performed. On each leg muscle, full width at half maximum (FWHM) which indicated the muscle cell diameters and pennation angle (PA) were measured and compared. FWHM showed significant positive correlations with PA, which is related to muscle strength. In addition, FWHM was higher for powerlifting, control, distance running, and teenager, in that order, suggesting that it may be directing the diameter of each muscle cell. Type 1 and type 2 fibers are enlarged by growth, so the fact that the FWHM of the control group was larger than that of the teenagers in this study may indicate that the muscle fibers were enlarged by growth. Also, FWHM has the possibility to increase with increased muscle fibers caused by training. We showed that QSI had the possibility to depict noninvasively the microstructure like muscle fiber type and subtle changes caused by growth and sports characteristics, which previously could only be assessed by biopsy.