Promotion of maturation of human pluripotent stem cell-derived cardiomyocytes via treatment with the peroxisome proliferator-activated receptor alpha agonist Fenofibrate.

As research on in vitro cardiotoxicity assessment and cardiac disease modeling becomes more important, the demand for human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) is increasing. However, it has been reported that differentiated hPSC-CMs are in a physiologically immature state compared to in vivo adult CMs. Since immaturity of hPSC-CMs can lead to poor drug response and loss of acquired heart disease modeling, various approaches have been attempted to promote maturation of CMs. Here, we confirm that peroxisome proliferator-activated receptor alpha (PPARα), one of the representative mechanisms of CM metabolism and cardioprotective effect also affects maturation of CMs. To upregulate PPARα expression, we treated hPSC-CMs with fenofibrate (Feno), a PPARα agonist used in clinical hyperlipidemia treatment, and demonstrated that the structure, mitochondria-mediated metabolism, and electrophysiology-based functions of hPSC-CMs were all mature. Furthermore, as a result of multi electrode array (MEA)-based cardiotoxicity evaluation between control and Feno groups according to treatment with arrhythmia-inducing drugs, drug response was similar in a dose-dependent manner. However, main parameters such as field potential duration, beat period, and spike amplitude were different between the 2 groups. Overall, these results emphasize that applying matured hPSC-CMs to the field of preclinical cardiotoxicity evaluation, which has become an essential procedure for new drug development, is necessary.

Subcutaneous Infliximab (CT-P13) as Maintenance Therapy for Inflammatory Bowel Disease: 2 Randomized Phase 3 Trials.

BACKGROUND & AIMS: CT-P13 subcutaneous (SC), an SC formulation of the intravenous (IV) infliximab biosimilar CT-P13 IV, creates a unique exposure profile. We aimed to demonstrate superiority of CT-P13 SC vs placebo as maintenance therapy in patients with Crohn's disease (CD) and ulcerative colitis (UC). METHODS: Two randomized, placebo-controlled, double-blind studies were conducted in patients with moderately to severely active CD or UC and inadequate response or intolerance to corticosteroids and immunomodulators. All patients received open-label CT-P13 IV 5 mg/kg at weeks 0, 2, and 6. At week 10, clinical responders were randomized (2:1) to CT-P13 SC 120 mg or placebo every 2 weeks until week 54 (maintenance phase) using prefilled syringes. Co-primary end points were clinical remission and endoscopic response (CD) and clinical remission (UC) at week 54 (all-randomized population). RESULTS: Overall, 396 patients with CD and 548 patients with UC received induction treatment. At week 54 in the CD study, statistically significant higher proportions of CT-P13 SC-treated patients vs placebo-treated patients achieved clinical remission (62.3% vs 32.1%; P < .0001) and endoscopic response (51.1% vs 17.9%; P < .0001). In the UC study, clinical remission rates at week 54 were statistically significantly higher with CT-P13 SC vs placebo (43.2% vs 20.8%; P < .0001). Achievement of key secondary end points was significantly higher with CT-P13 SC vs placebo across both studies. CT-P13 SC was well tolerated, with no new safety signals identified. CONCLUSIONS: CT-P13 SC was more effective than placebo as maintenance therapy and was well tolerated in patients with moderately to severely active CD or UC who responded to CT-P13 IV induction. CLINICALTRIALS: gov, Numbers: NCT03945019 (CD) and NCT04205643 (UC).

Emotional abuse mediated by negative automatic thoughts impacts functional connectivity during adolescence.

Background: Emotional abuse during childhood and adolescence is thought to be associated with the brain; however, the neural mechanism underlying the cognitive process remains unknown. Therefore, we aimed to investigate the mediating effect of negative automatic thoughts on the relationship between emotional abuse and resting-state functional connectivity (rsFC) during adolescence. Method: Our community sample included 54 adolescents aged 13-17 years in the statistical analysis. Resting-state functional and structural magnetic resonance imaging (MRI) was performed, while emotional abuse and negative automatic thoughts were assessed using self-reported scales. A mediation analysis was used to assess the contributions of early traumatic events and negative automatic thoughts to resting functional connectivity. Result: Higher negative automatic thoughts were associated with lower connectivity in the context of greater emotional abuse (i.e., suppression effect). Thus, the relationships between emotional abuse and connectivity in the precuneus (pCun)-medial prefrontal cortex, parahippocampal cortex-extrastriate cortex, and temporal cortex-temporal pole were decreased by negative automatic thoughts. In contrast, functional connections in the pCun-pCun, pCun-precuneus/posterior cingulate cortex, and nucleus accumbens-somatomotor areas were strongly mediated when emotionally abused adolescents reported a high tendency for negative automatic thoughts. Conclusion: Negative automatic thoughts strengthened the relationship between emotional abuse and rsFC. These findings highlight the underlying cognitive processing of the traumatic event-neural system, supporting the use of cognitive therapy for post-traumatic symptoms.

The fate of enterohemorrhagic Escherichia coli on alfalfa and fenugreek seeds and sprouts as affected by ascaroside #18 treatments.

Alfalfa and fenugreek sprouts are healthy foods, but they are occasionally contaminated with bacterial pathogens and serve as vehicles for transmitting foodborne illnesses. This study examined the efficacy of ascaroside (ascr)#18 treatment for the control of enterohemorrhagic E. coli (EHEC) growth on sprouts. Commercial alfalfa and fenugreek seeds were decontaminated with 20,000 ppm of NaClO, and residual chlorine was neutralized with Dey-Engley broth. Decontaminated seeds were treated with 1 mM or 1 µM ascr#18, a plant immunity modulator, before being dried and mixed with sandy soil inoculated with E. coli F4546 or BAA-2326 at 104-105 CFU/g. The inoculated seeds were sprouted on 1% water agar at 25ºC for 7 days in the dark. Seed or sprout samples were collected on days 0, 1, 3, 5, and 7 for enumeration of bacterial populations. Data was fit into the general linear model and analyzed using Fisher's least significant different test of the statistical analysis software. Treatment with ascr#18 significantly (P ≤ 0.05) reduced the cell population of EHEC on sprouts. The mean EHEC populations in the 1 mM or 1 µM treatment groups were 3.31 or 1.56 log CFU/g lower compared to the control groups. Besides treatment, sprout seed type and sprouting time were also significant independent variables influencing the growth of EHEC, according to the results of type III error analysis. However, EHEC strain type was not a significant independent variable. The study suggests that ascr#18 could be potentially used to control EHEC contamination and improve the microbial safety of sprouts.

Paintable Decellularized-ECM Hydrogel for Preventing Cardiac Tissue Damage.

The tissue-specific heart decellularized extracellular matrix (hdECM) demonstrates a variety of therapeutic advantages, including fibrosis reduction and angiogenesis. Consequently, recent research for myocardial infarction (MI) therapy has utilized hdECM with various delivery techniques, such as injection or patch implantation. In this study, a novel approach for hdECM delivery using a wet adhesive paintable hydrogel is proposed. The hdECM-containing paintable hydrogel (pdHA_t) is simply applied, with no theoretical limit to the size or shape, making it highly beneficial for scale-up. Additionally, pdHA_t exhibits robust adhesion to the epicardium, with a minimal swelling ratio and sufficient adhesion strength for MI treatment when applied to the rat MI model. Moreover, the adhesiveness of pdHA_t can be easily washed off to prevent undesired adhesion with nearby organs, such as the rib cages and lungs, which can result in stenosis. During the 28 days of in vivo analysis, the pdHA_t not only facilitates functional regeneration by reducing ventricular wall thinning but also promotes neo-vascularization in the MI region. In conclusion, the pdHA_t presents a promising strategy for MI treatment and cardiac tissue regeneration, offering the potential for improved patient outcomes and enhanced cardiac function post-MI.

Evaluation of the cardiotoxicity potential of bisphenol analogues in human induced pluripotent stem cells derived cardiomyocytes.

The importance of evaluating the cardiotoxicity potential of common chemicals as well as new drugs is increasing as a result of the development of animal alternative test methods using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM). Bisphenol A (BPA), which is used as a main material in plastics, is known as an endocrine-disrupting chemical, and recently reported to cause cardiotoxicity through inhibition of ion channels in CMs even with acute exposure. Accordingly, the need for the development of alternatives to BPA has been highlighted, and structural analogues including bisphenol AF, C, E, F, and S have been developed. However, cardiotoxicity data for analogues of bisphenol are not well known. In this study, in order to evaluate the cardiotoxicity potential of analogues, including BPA, a survival test of hiPSC-CMs and a dual-cardiotoxicity evaluation based on a multi-electrode array were performed. Acute exposure to all bisphenol analogues did not affect survival rate, but spike amplitude, beat period, and field potential duration were decreased in a dose-dependent manner in most of the bisphenols except bisphenol S. In addition, bisphenols, except for bisphenol S, reduced the contractile force of hiPSC-CMs and resulted in beating arrest at high doses. Taken together, it can be suggested that the developed bisphenol analogues could cause cardiotoxicity even with acute exposure, and it is considered that the application of the MEA-based dual-cardiotoxicity evaluation method can be an effective help in the development of safe alternatives.

PIBF1 regulates trophoblast syncytialization and promotes cardiovascular development.

Proper placental development in early pregnancy ensures a positive outcome later on. The developmental relationship between the placenta and embryonic organs, such as the heart, is crucial for a normal pregnancy. However, the mechanism through which the placenta influences the development of embryonic organs remains unclear. Trophoblasts fuse to form multinucleated syncytiotrophoblasts (SynT), which primarily make up the placental materno-fetal interface. We discovered that endogenous progesterone immunomodulatory binding factor 1 (PIBF1) is vital for trophoblast differentiation and fusion into SynT in humans and mice. PIBF1 facilitates communication between SynT and adjacent vascular cells, promoting vascular network development in the primary placenta. This process affected the early development of the embryonic cardiovascular system in mice. Moreover, in vitro experiments showed that PIBF1 promotes the development of cardiovascular characteristics in heart organoids. Our findings show how SynTs organize the barrier and imply their possible roles in supporting embryogenesis, including cardiovascular development. SynT-derived factors and SynT within the placenta may play critical roles in ensuring proper organogenesis of other organs in the embryo.

Affective dysfunction mediates the link between neuroimmune markers and the default mode network functional connectivity, and the somatic symptoms in somatic symptom disorder.

OBJECTIVE: Somatic symptom disorder (SSD) is characterized by physical symptoms and associated functional impairments that are often comorbid with depression and anxiety disorders. In this study, we explored relationships between affective symptoms and the functional connectivity of the default mode network (DMN) in SSD patients, as well as the impact of peripheral inflammation. We employed mediation analyses to investigate the potential pathways between these factors. METHODS: We recruited a total of 119 individuals (74 unmedicated SSD patients and 45 healthy controls), who were subjected to comprehensive psychiatric and clinical evaluations, blood tests, and resting-state functional magnetic resonance imaging scanning. We assessed neuroimmune markers (interleukin-6 (IL-6), high-sensitivity C-reactive protein (hs-CRP), tumor necrosis factor-α (TNF-α), tryptophan, serotonin, and 5-hydroxyindoleacetic acid (5-HIAA)), clinical indicators of somatic symptoms, depression, anxiety, anger, alexithymia, and functional connectivity (FC) within the DMN regions. Data were analyzed using correlation and mediation analysis, with a focus on exploring potential relations between clinical symptoms, blood indices, and DMN FCs. RESULTS: Patients with SSD had higher clinical scores as well as IL-6 and TNF-α levels compared with those in the control group (P < 0.05). The SSD group exhibited lower FC strength between the left inferior parietal lobule and left prefrontal cortex (Pfalse discovery rate (FDR) < 0.05). Exploratory correlation analysis revealed that somatic symptom scores were positively correlated with affective symptom scores, negatively correlated with the FC strength between the intra prefrontal cortex regions, and correlated with levels of IL-6, TNF- α, and tryptophan (uncorrected P < 0.01). Mediation analysis showed that levels of anxiety and trait anger significantly mediated the relations between DMN FC strength and somatic symptoms. In addition, the DMN FC mediated the level of trait anger with respect to somatic symptoms (all PFDR < 0.05). The levels of depression and trait anger exhibited significant mediating effects as suppressors of the relations between the level of 5-HIAA and somatic symptom score (all PFDR < 0.05). Further, the level of 5-HIAA had a mediating effect as a suppressor on the relation between DMN FC and state anger. Meanwhile, the levels of hs-CRP and IL-6 had full mediating effects as suppressors when explaining the relations of DMN FC strengths with the level of depression (all PFDR < 0.05). The patterns of valid mediation pathways were different in the control group. CONCLUSIONS: Affective symptoms may indirectly mediate the associations between DMN connectivity, somatic symptoms, and neuroimmune markers. Inflammatory markers may also mediate the impact of DMN connectivity on affective symptoms. These results emphasize the importance of affective dysregulation in understanding the mechanisms of SSD and have potential implications for the development of tailored therapeutic approaches for SSD patients with affective symptoms. Furthermore, in SSD research using DMN FC or neuroimmune markers, considering and incorporating such mediating effects of affective symptoms suggests the possibility of more accurate prediction and explanation.

Evaluation metric of smile classification by peri-oral tissue segmentation for the automation of digital smile design.

OBJECTIVES: This study aimed to develop and validate evaluation metric for an automated smile classification model termed the "smile index." This innovative model uses computational methods to numerically classify and analyze conventional smile types. METHODS: The datasets used in this study consisted of 300 images to verify, 150 images to validate, and 9 images to test the evaluation metric. Images were annotated using Labelme. Computational techniques were used to calculate smile index values for the study datasets, and the resulting values were evaluated in three stages. RESULTS: The smile index successfully classified smile types using cutoff values of 0.0285 and 0.193. High accuracy (0.933) was achieved, along with an F1 score greater than 0.09. The smile index successfully reclassified smiles into six types (low, low-to-medium, medium, medium-to-high, high, and extremely high smiles), thereby providing a clear distinction among different smile characteristics. CONCLUSION: The smile index is a novel dimensionless parameter for classifying smile types. The index acts as a robust evaluation tool for artificial intelligence models that automatically classify smile types, thereby providing a scientific basis for largely subjective aesthetic elements. CLINICAL SIGNIFICANCE: The computational approach employed by the smile index enables quantitative numerical classification of smile types. This fosters the application of computerized methods in quantifying and analyzing real smile characteristics observed in clinical practice, paving the way for a more objective evidence-based approach to aesthetic dentistry.

Comparative characteristics of fatigue in irritable bowel syndrome and inflammatory bowel disease: A systematic review and meta-analysis.

OBJECTIVE: Fatigue is a common symptom in both irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD), including Crohn's disease (CD) and ulcerative colitis (UC). This study aimed to distinguish fatigue characteristics in IBS and IBD, two functional and organic disorders. METHODS: We systematically searched the PubMed and Cochrane Library databases from inception to June 30, 2023, and conducted a meta-analysis to generate precise estimates and 95% confidence intervals. The analyses were stratified by fatigue type, severity, sex, disease phase, and comorbidities, and study quality was assessed using Newcastle-Ottawa Scale (NOS). RESULTS: Our analysis included 74 data (13 IBS, 31 CD, 30 UC) encompassing 16,689 participants (6484 males, 7402 females, and 2803 unknown). Overall, fatigue prevalence trended higher in IBS (54.5% [95%CI, 44.5-64.6]), followed by CD (49.8% [95%CI, 44.0-55.5]) and UC (43.6% [95%CI, 38.5-48.7]). This pattern persisted across sub-analyses, including general fatigue (63.4% vs. 51.3% vs. 45.3%) and moderate to severe fatigue (73.8% vs. 59.5% vs. 52.7%) for IBS, CD, and UC, respectively. Female predominance was observed in all three diseases (odds ratio: 1.5 in IBS and CD, 1.8 in UC). Fatigue prevalence significantly varied between disease phases (active vs. remission) in CD (61.3% vs. 36.3%) and UC (53.8% vs. 32.6%). Anemia, anxiety/depression, and/or IBS-like symptoms also contributed to fatigue in CD and UC. CONCLUSIONS: This study is the first extensive comparison of fatigue prevalence and features in IBS, CD, and UC. The findings offer valuable insights for treatment and management, aiding our understanding of functional and organic diseases.

Association between urinary arsenic concentration and genetic polymorphisms in Korean adults.

Arsenic (As) is a human carcinogen widely distributed in the environment. This study evaluated the association between the urinary As concentration and single nucleotide polymorphisms (SNPs) in Korean adults to determine the genetic factors related to As concentration. The study included 496 participants for the genome-wide association study (GWAS) and 1483 participants for the candidate gene approach study. Participants were 19 years and older. The concentrations of total As (Tot As) and total As metabolites (Tmet As, the sum of inorganic As and their metabolites; arsenite, arsenate, monomethylarsonic, and dimethylarsinic acid) in the urine were analyzed. The GWAS identified four SNPs (rs1432523, rs3776006, rs11171747, and rs807573) associated with urinary Tot As and four SNPs (rs117605537, rs3776006, rs11171747, and rs148103384) significantly associated with urinary Tmet As concentration (P < 1 × 10-4). The candidate gene study identified two SNPs (PRDX2 rs10427027 and GLRX rs3822751) in genes related to the reduction reaction associated with urinary Tot As and Tmet As. This study suggests that genetic factors may play a role in regulating As metabolism in the human body, affecting both exposure levels and its potential health risks in the general Korean population, even at low exposure levels. Supplementary Information: The online version contains supplementary material available at 10.1007/s43188-023-00216-x.

Development of heart organoid cryopreservation method through Fe3 O4 nanoparticles based nanowarming system.

Beyond single cell two-dimensional (2D) culture, research on organoids that can mimic human organs is rapidly developing. However, there are still problems in commercialization and joint research using organoids due to the lack of technology to safely store organoids. Since organoids are 3D complex structures with a certain size (0.1-5 mm) beyond the size of cells, the conventional cell-level cryopreservation method using cryoprotectant (CPA) cannot overcome the damage caused by volume change due to osmotic pressure difference and ice nucleation. Herein, we attempted to solve such limitations by applying a nanowarming system using CPA with high cell permeability and Fe3 O4 nanoparticles. By performing beat rate measurement, histological analysis, contractility analysis, and multi-electrode array, it was verified that the developed method could significantly improve functional recovery and survival of heart organoids after freezing and thawing. In this study, we demonstrated a successful organoid cryopreservation method based on a Fe3 O4 nanowarming system. The developed technology will provide clues to the field of tissue cryopreservation and spur the application of organoids.

Effect of high-speed sintering on the marginal and internal fit of CAD/CAM-fabricated monolithic zirconia crowns.

This study compared the marginal and internal fit of zirconia crowns fabricated using conventional and high-speed induction sintering. A typodont mandibular right first molar was prepared and 60 zirconia crowns were fabricated: 30 crowns using conventional sintering and 30 crowns using high-speed sintering. We presented a new evaluation methodology to measure the marginal and internal fit of restorations through digital scanning, aligning the two datasets, and measuring the distance between two arbitrary point sets of the datasets. For the marginal fit, we calculated the maximum values of the shortest distances between the marginal line of the prepared tooth and that of the crown. The calculated values ranged from 359 to 444 µm, with smaller values for the high-speed sintered crowns (P < 0.05). For the internal fit, we employed mesh sampling and computed the geodesic distances between the prepared tooth surface and the crown intaglio surface. The measured values ranged from 177 to 229 µm with smaller values for the high-speed sintered crowns, but no significant difference was found (P > 0.05). Based on our results, the high-speed sintering method can be considered a promising option for single-visit zirconia treatment in dental practice.

GIRK2 potassium channels expressed by the AgRP neurons decrease adiposity and body weight in mice.

It is well known that the neuropeptide Y (NPY)/agouti-related peptide (AgRP) neurons increase appetite and decrease thermogenesis. Previous studies demonstrated that optogenetic and/or chemogenetic manipulations of NPY/AgRP neuronal activity alter food intake and/or energy expenditure (EE). However, little is known about intrinsic molecules regulating NPY/AgRP neuronal excitability to affect long-term metabolic function. Here, we found that the G protein-gated inwardly rectifying K+ (GIRK) channels are key to stabilize NPY/AgRP neurons and that NPY/AgRP neuron-selective deletion of the GIRK2 subunit results in a persistently increased excitability of the NPY/AgRP neurons. Interestingly, increased body weight and adiposity observed in the NPY/AgRP neuron-selective GIRK2 knockout mice were due to decreased sympathetic activity and EE, while food intake remained unchanged. The conditional knockout mice also showed compromised adaptation to coldness. In summary, our study identified GIRK2 as a key determinant of NPY/AgRP neuronal excitability and driver of EE in physiological and stress conditions.

Safety and effectiveness of intravenous CT-P13 in inflammatory arthritis: post-marketing surveillance study in Thailand.

Background: The infliximab biosimilar CT-P13 was approved in Thailand in 2015. Methods: This open-label, multicenter, post-marketing surveillance study evaluated the safety (events of special interest [ESIs]; primary end point) and effectiveness of 46 weeks of CT-P13 treatment according to routine practice in patients with rheumatoid arthritis (RA), ankylosing spondylitis (AS), or psoriatic arthritis (PsA), with 1 year follow-up post-treatment. Results: 30 patients were enrolled (16 RA, 8 AS and 6 PsA). Infections were the most frequently reported study drug-related ESIs (2 RA and 2 AS). One patient with RA and one with PsA experienced infusion-related reactions. No cases of tuberculosis, malignancy (as expected, given 1 year follow-up), or drug-induced liver disease were reported. Disease activity improved across indications. Conclusion: CT-P13 was well tolerated and effective across indications.

Infliximab is one biological medicine used to treat inflammatory diseases, including rheumatoid arthritis (RA), ankylosing spondylitis (AS) and psoriatic arthritis (PsA). CT-P13 is a near-identical copy, called a biosimilar, of the original ('reference') version of infliximab. CT-P13 is the first biosimilar to receive regulatory approval for treatment of the same three diseases from the European Medicines Agency (EMA) and US Food and Drug Administration. Biosimilarity means that CT-P13 does not differ from the original version of infliximab in clinically important ways, such as how safe it is and how well it works. CT-P13 and reference infliximab provided similar symptom relief during previous clinical trials, and both drugs caused similar side effects. It is important to monitor the safety and performance of CT-P13 when given during routine clinical practice, and in different ethnic populations, such as through the study reported here. Following regulatory approval in Thailand, 30 patients prescribed CT-P13 during routine clinical practice participated in this study. The study included 16 patients with RA, eight with AS and six with PsA. The patients took CT-P13 for 46 weeks and were monitored for a further year. Side effects of CT-P13 were as expected based on previous experience and did not raise any safety concerns. Based on the known safety profile of CT-P13, the study looked at some side effects in particular: infections were the most common of these side effects, experienced by 16 patients overall (seven patients with RA, five patients with AS and four patients with PsA). CT-P13 improved symptoms for all of the diseases. The study suggests that CT-P13 can be given safely and reduces symptoms in Thai patients with AS, RA or PsA. Thai Clinical Trials Registry: TCTR20170817005 (www.thaiclinicaltrials.org/show/TCTR20170817005).

Food literacy and its relationship with food intake: a comparison between adults and older adults using 2021 Seoul Food Survey data.

OBJECTIVES: This study was conducted to examine the differences in food literacy between adults and older adults and the association of food literacy with food group intake. METHODS: In total, 4,039 participants from the 2021 Survey for Food Consumption in Seoul were included in this study. The intake of whole grains, high-protein foods, total vegetables, vegetables excluding kimchi and pickles, fresh fruits, and milk and dairy products was assessed using a simple food frequency questionnaire. Food literacy was measured using a food literacy measurement questionnaire. RESULTS: With the exception of milk and dairy, adults had a significantly higher proportion of insufficient food intake than older adults (p<0.001). Both adults and older adults with sufficient food group consumption had higher food literacy scores. Notably, the difference in food literacy scores by level of food intake was highest for fresh fruits (p<0.001). After adjusting for confounding factors affecting food literacy scores, a higher score was associated with a lower probability of having insufficient food intake in all food groups for both adults and older adults. Specifically, those in the highest food literacy score quartile were less likely to have insufficient intake of vegetables (odds ratio [OR], 0.35; 95% confidence interval [CI], 0.28 to 0.43) and fresh fruits (OR, 0.32; 95% CI, 0.24 to 0.43), compared to the lowest quartile. CONCLUSIONS: Improving food literacy is an important factor for promoting healthy food intake in older adults as well as adults. Therefore, it is necessary to develop intervention programs to work toward this goal.

Structural basis for proapoptotic activation of Bak by the noncanonical BH3-only protein Pxt1.

Bak is a critical executor of apoptosis belonging to the Bcl-2 protein family. Bak contains a hydrophobic groove where the BH3 domain of proapoptotic Bcl-2 family members can be accommodated, which initiates its activation. Once activated, Bak undergoes a conformational change to oligomerize, which leads to mitochondrial destabilization and the release of cytochrome c into the cytosol and eventual apoptotic cell death. In this study, we investigated the molecular aspects and functional consequences of the interaction between Bak and peroxisomal testis-specific 1 (Pxt1), a noncanonical BH3-only protein exclusively expressed in the testis. Together with various biochemical approaches, this interaction was verified and analyzed at the atomic level by determining the crystal structure of the Bak-Pxt1 BH3 complex. In-depth biochemical and cellular analyses demonstrated that Pxt1 functions as a Bak-activating proapoptotic factor, and its BH3 domain, which mediates direct intermolecular interaction with Bak, plays a critical role in triggering apoptosis. Therefore, this study provides a molecular basis for the Pxt1-mediated novel pathway for the activation of apoptosis and expands our understanding of the cell death signaling coordinated by diverse BH3 domain-containing proteins.

A Deep Learning Model for 3D Ground Reaction Force Estimation Using Shoes with Three Uniaxial Load Cells.

Ground reaction force (GRF) is essential for estimating muscle strength and joint torque in inverse dynamic analysis. Typically, it is measured using a force plate. However, force plates have spatial limitations, and studies of gaits involve numerous steps and thus require a large number of force plates, which is disadvantageous. To overcome these challenges, we developed a deep learning model for estimating three-axis GRF utilizing shoes with three uniaxial load cells. GRF data were collected from 81 people as they walked on two force plates while wearing shoes with three load cells. The three-axis GRF was calculated using a seq2seq approach based on long short-term memory (LSTM). To conduct the learning, validation, and testing, random selection was performed based on the subjects. The 60 selected participants were divided as follows: 37 were in the training set, 12 were in the validation set, and 11 were in the test set. The estimated GRF matched the force plate-measured GRF with correlation coefficients of 0.97, 0.96, and 0.90 and root mean square errors of 65.12 N, 15.50 N, and 9.83 N for the vertical, anterior-posterior, and medial-lateral directions, respectively, and there was a mid-stance timing error of 5.61% in the test dataset. A Bland-Altman analysis showed good agreement for the maximum vertical GRF. The proposed shoe with three uniaxial load cells and seq2seq LSTM can be utilized for estimating the 3D GRF in an outdoor environment with level ground and/or for gait research in which the subject takes several steps at their preferred walking speed, and hence can supply crucial data for a basic inverse dynamic analysis.

Verification of Therapeutic Effect through Accelerator Mass Spectrometry-Based Single Cell Level Quantification of hESC-Endothelial Cells Distributed into an Ischemic Model.

As the potential of pluripotent stem cell-derived differentiated cells has been demonstrated in regenerative medicine, differentiated vascular endothelial cells (ECs) are emerging as a therapeutic agent for the cardiovascular system. To verify the therapeutic efficacy of differentiated ECs in an ischemic model, human embryonic stem cells (hESCs) are induced as EC lineage and produce high-purity ECs through fluorescence-activated cell sorting (FACS). When hESC-ECs are transplanted into a hindlimb ischemic model, it is confirmed that blood flow and muscle regeneration are further improved by creating new blood vessels together with autologous ECs than the primary cell as cord blood endothelial progenitor cells (CB-EPCs). In addition, previously reported studies show the detection of transplanted cells engrafted in blood vessels through various tracking methods, but fail to provide accurate quantitative values over time. In this study, it is demonstrated that hESC-ECs are engrafted approximately sevenfold more than CB-EPCs by using an accelerator mass spectrometry (AMS)-based cell tracking technology that can perform quantification at the single cell level. An accurate quantification index is suggested. It has never been reported in in vivo kinetics of hESC-ECs that can act as therapeutic agents.

Development of Fluorescent Bacteria with Lux and Riboflavin Genes.

Lumazine protein from marine luminescent bacteria of Photobacterium species bind with very high affinity to the fluorescent chromophore 6,7-dimethyl-8-ribitylumazine. The light emission of bacterial luminescent systems is used as a sensitive, rapid, and safe assay for an ever-increasing number of biological systems. Plasmid pRFN4, containing the genes encoding riboflavin from the rib operon of Bacillus subtilis, was designed for the overproduction of lumazine. To construct fluorescent bacteria for use as microbial sensors, novel recombinant plasmids (pRFN4-Pp N-lumP and pRFN4-Pp luxLP N-lumP) were constructed by amplifying the DNA encoding the N-lumP gene (luxL) from P. phosphoreum and the promoter region (luxLP) present upstream of the lux operon of the gene by PCR and ligating into the pRFN4-Pp N-lumP plasmid. A new recombinant plasmid, pRFN4-Pp luxLP-N-lumP, was constructed with the expectation that the fluorescence intensity would be further increased when transformed into Escherichia coli. When this plasmid was transformed into E. coli 43R, the fluorescence intensity of transformants was 500 times greater than that of E. coli alone. As a result, the recombinant plasmid in which the gene encoding N-LumP and DNA containing the lux promoter exhibited expression that was so high as to show fluorescence in single E. coli cells. The fluorescent bacterial systems developed in the present study using lux and riboflavin genes can be utilized in the future as biosensors with high sensitivity and rapid analysis times.