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Aging, genetic mutations, and other pathological conditions cause impairment of skeletal growth and bone metabolism, which affect activities of daily living and quality of life in all life stages. Although several drugs have been used in clinical settings and new drugs have been developed for the treatment of skeletal degenerative disorders such as osteoporosis and genetic disorders such as osteogenesis imperfecta (OI), there is clear demand for development of new drugs, especially orally available anabolic drugs that are applicable for a wide range of skeletal disorders.To identify therapeutic candidates for skeletal disorders, peptide screening was performed. To validate the identified peptides, we performed a bone histomorphometric analysis with rat bone tissues andWe identified the egg yolk-derived functional peptide PF201. PF201 promotedD2 is likely to be a candidate for an orally available therapeutic for a range of skeletal disorders.

Cardiovascular diseases (CVDs) remain a leading cause of death globally. According to the American Heart Association, approximately 19.1 million deaths were attributed to CVDs in 2020, in particular, ischemic heart disease and stroke. Several known risk factors for CVDs include smoking, alcohol consumption, lack of regular physical activity, and diabetes. The last decade has been characterized by widespread diffusion in the use of wristband-style wearable devices which can monitor and collect heart rate data, among other information. Wearable devices allow the analysis and interpretation of physiological and activity data obtained from the wearer and can therefore be used to monitor and prevent potential CVDs. However, these data are often provided in a manner that does not allow the general user to immediately comprehend possible health risks, and often require further analytics to draw meaningful conclusions. In this paper, we propose a disentangled variational autoencoder (β-VAE) with a bidirectional long short-term memory network (BiLSTM) backend to detect in an unsupervised manner anomalies in heart rate data collected during sleep time with a wearable device from eight heterogeneous participants. Testing was performed on the mean heart rate sampled both at 30 s and 1 min intervals. We compared the performance of our model with other well-known anomaly detection algorithms, and we found that our model outperformed them in almost all considered scenarios and for all considered participants. We also suggest that wearable devices may benefit from the integration of anomaly detection algorithms, in an effort to provide users more processed and straightforward information.

Liquid–liquid phase separation (LLPS), an important phenomenon in the field of polymer science and material design, plays an essential role in cells and living bodies. Poly(vinyl alcohol) (PVA) is a popular semicrystalline polymer utilized in the synthesis of artificial biomaterials. The aqueous solutions of its derivatives with tuned degrees of saponification (DS) exhibit LLPS. However, the miscibility and LLPS behavior of PVA aqueous solution are still unclear. This study describes the miscibility diagram of the ternary mixture, where water and two types of poly(vinyl alcohol) (PVA) with different DSs [98 (PVA98), 88 (PVA88), 82 (PVA82), and 74 mol% (PVA74)] were blended. UV–Vis measurement was conducted to evaluate the miscibility. Immiscibility was more pronounced at elevated temperatures, exhibiting LLPS. The ternary immiscibility diagram, displaying miscible–immiscible behaviors in the aqueous mixtures of PVA74:PVA98, PVA82:PVA98, and PVA88:PVA98 (blended at a constant volume ratio), indicated that increasing the concentration, temperature, and blend ratio of PVAs at a lower DS increased immiscibility, suggesting that the free energy of mixing increases with increasing these parameters. The miscible–immiscible behaviors of PVAs/water systems provide fundamental knowledge about LLPS and the design of PVA-based materials.

Quantifying health status and identifying modifiable factors are essential for effective and individualized prevention of age-related conditions and for promoting health during aging. The ME-BYO concept from Kanagawa Prefecture, one of Japan’s largest prefectures, can be used to establish a healthy aging society. In disease etiology, ME-BYO considers the status of an individual’s body and mind as changing continuously from healthy to sick instead of being a dichotomy between the two. ME-BYO conceptualizes the entire process of this change. The ME-BYO index was developed in 2019 to comprehensively and numerically measure and visualize an individual’s current health status and future disease risk by quantifying data on the four domains of metabolic function, locomotor function, cognitive function, and mental resilience. The ME-BYO index has been implemented in the personal health management application “My ME-BYO.” However, scientific validation of this index and the development of a practical application using healthcare data remain to be completed. In 2020, our research team started a project to refine the ME-BYO index using data from the Kanagawa ME-BYO prospective cohort study, which is a large population-based genomic cohort study. This project will scientifically evaluate the ME-BYO index and develop a practical application for promoting healthy aging.

We demonstrate an experimental comparison of the bond lifetime, estimated using surface plasmon resonance (SPR), and the viscoelastic relaxation time of transient networks with well-controlled structures (dynamically cross-linked Tetra-PEG gel). SPR and viscoelastic measurements revealed that the temperature dependences of the two characteristic times are in agreement, while the viscoelastic response is delayed with respect to the lifetime by a factor of 2-3, dependent on the network strand length. Polymers cross-linked by temporary interactions form transient networks, which show fascinating viscoelasticity with a single relaxation mode. However, the molecular understanding of such simple viscoelasticity has remained incomplete because of the difficulty of experimentally evaluating bond lifetimes and heterogeneous structures in conventional transient networks. Our results suggest that bond dissociation and recombination both contribute to the macromechanical response. This report on direct bond-lifetime-viscoelastic-relaxation time comparison provides important information for the molecular design of transient network materials.

Cell-based therapy has been used to treat stroke related disorders, which have no treatment options available 4.5 hours after onset. Although the administration of tissue plasminogen activator and mechanical thrombectomy are potent treatments, their clinical implementation is limited within the available time. Here, we aimed to use induced pluripotent stem cell-derived neural progenitor cells (NPCs) for stroke treatment with higher delivery efficiency in stroke areas, which will improve the therapeutic effect. E-selectin binding oligopeptide (Esbp) was conjugated with poly(ethylene glycol)-conjugated-lipid (Esbp-PEG-lipid) with different molecular weights of PEG (5 and 40 kDa) for cell surface modification. Then, we optimized the cell surface modification of NPCs by studying cell-binding ability onto the model surfaces of stroke areas, such as recombinant E-selectin-immobilized surfaces and TNF-α activated endothelium. As a result, the cell surface modification of NPCs with Esbp-PEG-lipid was found to induce specific intercellular interactions with the activated endothelium through the binding of Esbp with E-selectin. Additionally, the shorter PEG spacer was suitable for intercellular interactions. Thus, our technique shows potential for use in cell therapy with enhanced cell accumulation in infarct areas.

PDF file - 57K, YY1 overexpression does not affect the expression of HIF-1α under hypoxia

PDF file - 142K, Contains Luciferase assay, Western blotting, ELISA, Immunoblotting, YY2 gene silencing, Cell viability, Quantitative PCR, and Immunoprecipitation data.

PDF file - 270K, The proliferation property of HeLa cells, wild type HCT116 (+/+) cells, and p53-null HCT116 (-/-) cells under hypoxic condition

PDF file - 60K, Primer pairs used in this study for quantitative Real-time RT-PCR

PDF file - 101K, YY1-silencing suppresses the colonization and growth of A549-luc cells in lungs

PDF file - 129K, The establishment of HCT116 p53-/-/shYY1 stable cells

PDF file - 127K, shYY1 vectors significantly silence YY1 expression

PDF file - 173K, The effect of YY2-silencing on HIF-1a target genes and the effect of YY1-silencing on hydroxyl HIF-1a under hypoxic condition

PDF file - 91K

Physiological time series are affected by many factors, making them highly nonlinear and nonstationary. As a consequence, heart rate time series are often considered difficult to predict and handle. However, heart rate behavior can indicate underlying cardiovascular and respiratory diseases as well as mood disorders. Given the importance of accurate modeling and reliable predictions of heart rate fluctuations for the prevention and control of certain diseases, it is paramount to identify models with the best performance in such tasks. The objectives of this study were to compare the results of three different forecasting models (Autoregressive Model, Long Short-Term Memory Network, and Convolutional Long Short-Term Memory Network) trained and tested on heart rate beats per minute data obtained from twelve heterogeneous participants and to identify the architecture with the best performance in terms of modeling and forecasting heart rate behavior. Heart rate beats per minute data were collected using a wearable device over a period of 10 days from twelve different participants who were heterogeneous in age, sex, medical history, and lifestyle behaviors. The goodness of the results produced by the models was measured using both the mean absolute error and the root mean square error as error metrics. Despite the three models showing similar performance, the Autoregressive Model gave the best results in all settings examined. For example, considering one of the participants, the Autoregressive Model gave a mean absolute error of 2.069 (compared to 2.173 of the Long Short-Term Memory Network and 2.138 of the Convolutional Long Short-Term Memory Network), achieving an improvement of 5.027% and 3.335%, respectively. Similar results can be observed for the other participants. The findings of the study suggest that regardless of an individual’s age, sex, and lifestyle behaviors, their heart rate largely depends on the pattern observed in the previous few minutes, suggesting that heart rate can be reasonably regarded as an autoregressive process. The findings also suggest that minute-by-minute heart rate prediction can be accurately performed using a linear model, at least in individuals without pathologies that cause heartbeat irregularities. The findings also suggest many possible applications for the Autoregressive Model, in principle in any context where minute-by-minute heart rate prediction is required (arrhythmia detection and analysis of the response to training, among others).

The mobility of sustained molecules is influenced by viscoelasticity, which is strongly correlated with the diffusional property in polymeric liquid. However, the study of transient networks formed by a reversible crosslink, which is the viscoelastic liquid, was insufficient due to the absence of a model system. We compare the viscoelastic and diffusional properties of the transient networks, using the model system with controlled network connectivity (Tetra-PEG slime). According to independent measurements of viscoelasticity and diffusion, the root-mean-square distance the polymer diffuses during the viscoelastic relaxation time shows a large deviation from the self-size of the polymer, which is contrary to the conventional understanding. This decoupling between viscoelasticity and diffusion is unique for transient networks, suggesting that the viscoelastic relaxation is not induced by the diffusion of one prepolymer, particularly in the network with low connectivity. These findings will provide a definite basis for discussion to understand the viscoelasticity in transient networks.

The effect of network connectivity on viscoelastic relaxation in transient networks with well-defined structures (Tetra-PEG slime) was experimentally evaluated and compared to bond dissociation kinetics. To control the connectivity and discuss the pure effect precisely, we mixed the precursors in off-stoichiometric ratio. With decreasing network connectivity, the viscoelastic relaxation time accelerated and became shorter than the bond dissociation time. With increasing polymer concentration, the connectivity at which the viscoelastic relaxation time matched the dissociation time shifted to the high-connectivity region. The dependence of viscoelastic relaxation on connectivity can be adequately explained within the framework of the lifetime of a backbone. The backbone has numerous breakage points in low-connectivity region nearby the gelation point, resulting in a shorter lifetime than the dissociation time. However, the Rubinstein-Semenov model based on backbone relaxation does not predict the concentration dependence, suggesting that the formation of the network in the dilute/semi-dilute region deviates from a random branching process. These findings provide a crucial foundation for the molecular comprehension of transient network materials.

Multinucleated giant cells like osteoclasts (OCs) often contain >10 nuclei. This raises the question of whether nuclei residing in the same cytoplasm are all regulated similarly or whether they are heterogeneous, expressing a different set of genes regulated by nucleus-specific epigenetic mechanisms. We combined imaging of living cells with confocal microscope, picking of intracellular components inside a single cell using the Single Cellome™ System SS2000 (Yokogawa, Japan), allowing next generation sequencing at high resolution for mRNA (iscRNA-seq) and for DNA methylation (iscWGBS; whole genome bisulfite sequencing) of any cellular components including a single OC nucleus. Our results reveal that individual nuclei within the same cell are heterogeneous in terms of gene expression and epigenetic regulation, possibly affecting regional cell function.

Injectable hydrogels are biomaterials that can be administered minimally invasively in liquid form and are considered promising artificial extracellular matrix (ECM) materials. However, ordinary injectable hydrogels are synthesized from water-soluble molecules to ensure injectability, resulting in non-phase-separated structures, making them structurally different from natural ECMs with phase-separated insoluble structural proteins, such as collagen and elastin. Here, we propose a simple material design approach to impart phase-separated structures to injectable hydrogels by adding inorganic salts. Injecting a gelling solution of mutually cross-linkable tetra-arm poly(ethylene glycol)s with potassium sulfate at optimal concentrations results in the formation of a hydrogel with phase-separated structures in situ. These phase-separated structures provide up to an 8-fold increase in fracture toughness while allowing the encapsulation of live mouse chondrogenic cells without compromising their proliferative activity. Our findings highlight that the concentration of inorganic salts is an important design parameter in injectable hydrogels for artificial ECMs.

We demonstrate that dissolved polymers can direct molecular crystallization behavior under dilute and crowded conditions. Larger poly(ethylene glycol)s (PEGs) accelerated caffeine crystal formation in the dilute regime of PEG solutions, which was attributed to the depletion attraction that promoted caffeine cluster aggregation into crystal nuclei. Alternatively, in the semidilute regime, the caffeine crystal formation rate was insensitive to the molecular weight (MW) of PEGs. PEGs with various MWs appeared to induce depletion attraction to a similar extent as the properties of polymer solutions in the semidilute regime described by blobs, which are constant in size at a given polymer concentration irrespective of MW. This study highlights differences between in vitro polymer solutions and crowded intracellular environments composed of folded biomacromolecules and contributes to developing the use of in vitro macromolecular crowding for the control of molecular self-assembly. Dissolved polymers directed molecular crystallization behavior under dilute and crowding conditions. Larger poly(ethylene glycol)s (PEGs) accelerated caffeine crystal formation in the dilute regime of PEG solutions, which was attributed to the depletion attraction that promoted caffeine cluster aggregation into crystal nuclei. In the semidilute regime, the crystal formation rate was insensitive to the molecular weight of PEGs. This observation was consistent with polymer blob properties, which govern depletion attraction and maintain a constant size at a given polymer concentration, irrespective of molecular weight in the semidilute regime.

Scaffold systems that can easily encapsulate cells and safely retrieve them at the desired time are important for the advancement of cell-based medicine. In this study, we designed and fabricated thioester-based poly(ethylene glycol) (PEG) hydrogels with injectability and on-demand degradability as new scaffold materials for cells. Hydrogels can be formed in situ within minutes via thioester cross-linking between PEG molecules and can be degraded under mild conditions in response to L-cysteine molecules through thiol exchange occurring at the thioester linkage. Various cell experiments, especially with sucrose, which enables the adjustment of the osmotic pressure around the cells, showed that the damage to the cells during encapsulation and degradation was minimal, indicating the capability of on-demand retrieval of intact cells. This hydrogel system is a versatile tool in the field of cell-based research and applications such as tissue regeneration and regenerative medicine.

Objective Cognitive decline is an important and well-documented health problem. The Mini-Cog, a simple cognitive function test, is recommended as a potential early cognitive screening tool. Kanagawa Prefecture, one of the largest prefectures in Japan, developed this self-testing application on a smartphone to enable a large number of residents to assess their cognitive function. This study aimed to verify the validity and reliability of the Mini-Cog. Results Twenty men and 20 women aged 65–85 years were enrolled. Criterion-related validity of the method tested by professional staff was found to have an acceptable positive association. The test–retest reliability was lower than the clinically expected intraclass correlation coefficient value because of the inclusion of learning and order effects. If the Mini-Cog score of this application is low, the system is equipped with a function that advises the users on preventing cognitive decline, directing them to the appropriate services, and recommending visits to a medical institution. Therefore, the system can be used continuously as a tool for health behaviors and promotions.

To investigate the effect of inhomogeneity in the first network on the enormously high toughness of double network (DN) gels, we fabricated DN gels with a nearly homogeneous first network structure (named St-TPEG/PAAm DN gels) based on tetra-PEG (TPEG) gels via a molecular stent method. The St-TPEG/PAAm DN gels also show excellent mechanical properties and yielding-like phenomena comparable to conventional DN gels. This result demonstrates that the inhomogeneity within the first network is not essential for the specific toughening mechanism of DN gels. On the other hand, the St-TPEG/PAAm DN gels and conventional DN gels undergo substantially different fracture processes before the yielding point. This suggests the importance of a “homogenization process” for the yielding of DN gels. Since the St-TPEG/PAAm DN gels consist of a well-defined first network, they may serve as model DN gels in the future for further studies on fracture processes of DN gels.

Human synovium-derived stem cells (hSSCs) are an attractive source of cells for cartilage repair. At present, the quality of tissue and techniques used for cartilage regeneration have scope for improvement. A small compound, TD-198946, was reported to enhance chondrogenic induction from hSSCs; however, other applications of TD-198946, such as priming the cell potential of hSSCs, remain unknown. Our study aimed to examine the effect of TD-198946 pretreatment on hSSCs. HSSCs were cultured with or without TD-198946 for 7 days during expansion culture and then converted into a three-dimensional pellet culture supplemented with bone morphogenetic protein-2 (BMP2) and/or transforming growth factor beta-3 (TGFβ3). Chondrogenesis in cultures was assessed based on the GAG content, histology, and expression levels of chondrogenic marker genes. Cell pellets derived from TD-198946-pretreated hSSCs showed enhanced chondrogenic potential when chondrogenesis was induced by both BMP2 and TGFβ3. Moreover, cartilaginous tissue was efficiently generated from TD-198946-pretreated hSSCs using a combination of BMP2 and TGFβ3. Microarray analysis revealed that NOTCH pathway-related genes and their target genes were significantly upregulated in TD-198946-treated hSSCs, although TD-198946 alone did not upregulate chondrogenesis related markers. The administration of the NOTCH signal inhibitor diminished the effect of TD-198946. Thus, TD-198946 enhances the chondrogenic potential of hSSCs via the NOTCH3 signaling pathway. This study is the first to demonstrate the gradual activation of NOTCH3 signaling during chondrogenesis in hSSCs. The priming of NOTCH3 using TD-198946 provides a novel insight regarding the regulation of the differentiation of hSSCs into chondrocytes.

Polyelectrolyte gels comprising fixed ions exhibit swelling behaviors because of external solution conditions. Such behaviors are usually explained by using the Flory–Rehner model that considers th...

Because the swelling of polymer gels is a diffusion process of the polymer network, the swelling rate is determined by the diffusion coefficient of a polymer network. The diffusion coefficient (D) includes the polymer–solvent mixing contribution (Dmix) and the elastic contribution from the polymer network (Del). In this study, we experimentally investigate D, Dmix, and Del, using tetra gels whose network structural parameters are systematically controllable. We measured D and the shear modulus (G) using dynamic light scattering and dynamic viscoelastic measurement, respectively. According to the Tanaka–Fillmore theory, we evaluate Dmix and Del using the linear relationship between D and G in the as-prepared state. Consequently, we demonstrate a relation of D = Dmix + Del in the as-prepared, swelling, and equilibrium swollen states.

Pluripotent stem cells (PSCs) are attractive regenerative therapy tools for skeletal tissues. However, a deep understanding of skeletal development is required in order to model this development with PSCs, and for the application of PSCs in clinical settings. Skeletal tissues originate from three types of cell populations: the paraxial mesoderm, lateral plate mesoderm, and neural crest. The paraxial mesoderm gives rise to the sclerotome mainly through somitogenesis. In this process, key developmental processes, including initiation of the segmentation clock, formation of the determination front, and the mesenchymal–epithelial transition, are sequentially coordinated. The sclerotome further forms vertebral columns and contributes to various other tissues, such as tendons, vessels (including the dorsal aorta), and even meninges. To understand the molecular mechanisms underlying these developmental processes, extensive studies have been conducted. These studies have demonstrated that a gradient of activities involving multiple signaling pathways specify the embryonic axis and induce cell-type-specific master transcription factors in a spatiotemporal manner. Moreover, applying the knowledge of mesoderm development, researchers have attempted to recapitulate the in vivo development processes in in vitro settings, using mouse and human PSCs. In this review, we summarize the state-of-the-art understanding of mesoderm development and in vitro modeling of mesoderm development using PSCs. We also discuss future perspectives on the use of PSCs to generate skeletal tissues for basic research and clinical applications., Regenerative medicine: the challenge of building skeletal tissue A deeper understanding of skeletal tissue development and improvements in tissue engineering will help pluripotent stem cell (PSC) therapies to reach their full potential for skeletal repair. The paraxial mesoderm, an embryonic germ layer, is crucial to the formation of healthy axial skeleton. Shoichiro Tani at the University of Tokyo, Japan, and co-workers reviewed current understanding of paraxial mesoderm development and studies involving in vitro PSC skeletal modeling. The formation of the paraxial mesoderm and associated connective tissues comprises multiple stages, and studies in vertebrate embryos have uncovered critical signaling pathways and cellular components important to PSC modeling. Although many individual cellular components can now be modeled, it remains challenging to recreate three-dimensional skeletal tissues. Such an achievement would facilitate a functioning model of bone metabolism, the next step in achieving skeletal regeneration.

Clinically relevant human induced pluripotent stem cell (hiPSC) derivatives require efficient protocols to differentiate hiPSCs into specific lineages. Here we developed a fully defined xeno-free strategy to direct hiPSCs toward osteoblasts within 21 days. The strategy successfully achieved the osteogenic induction of four independently derived hiPSC lines by a sequential use of combinations of small-molecule inducers. The induction first generated mesodermal cells, which subsequently recapitulated the developmental expression pattern of major osteoblast genes and proteins. Importantly, Col2.3-Cherry hiPSCs subjected to this strategy strongly expressed the cherry fluorescence that has been observed in bone-forming osteoblasts in vivo. Moreover, the protocol combined with a three-dimensional (3D) scaffold was suitable for the generation of a xeno-free 3D osteogenic system. Thus, our strategy offers a platform with significant advantages for bone biology studies and it will also contribute to clinical applications of hiPSCs to skeletal regenerative medicine.

Subarachnoid hemorrhage (SAH) frequently results in several serious complications, such as cerebral vasospasm. We previously reported the effect of trehalose on vasospasm, inflammatory responses, and lipid peroxidation induced by blood exposure. Herein, to further elucidate the mechanism of action of trehalose, we investigated whether or not post-administration of trehalose can directly influence blood clotting in the cistern. As a result of trehalose injection after the onset of experimental SAH, blood clotting around the basilar artery was clearly inhibited. We also found that trehalose positively impacted coagulation and fibrinolysis parameters in rat, rabbit and human plasma in vitro. These findings suggest that trehalose has suppressive effects on blood clotting in addition to vasospasm, inflammatory responses, and lipid peroxidation after SAH.

ObjectiveMaintaining or improving regular walking speed can help extend healthy life expectancy and prevent frailty. The evaluation of walking speed can help individuals meet their daily exercise goals; therefore, it may be beneficial as a public health policy for residents to measure and evaluate their walking speed easily. This study aimed to verify the validity and reliability of a smartphone application for the 10-m walk test, measured in the general population.MethodsThe study participants were men (n = 20) and women (n = 20) aged 65–85 years. The 10-m walk tests were performed at the usual walking speed, using the stopwatch function of a newly developed smartphone application. A total of three 10-m walk tests were performed simultaneously with the study participants and professional fitness instructors to evaluate the criterion-related validity and the test-retest reliability.ResultsA strong positive correlation was found in the criterion-related validity by the study participants and professional staff for the average of the three trials {r = 0.961 [95% confidence interval (CI) = 0.927, 0.979]}. The results revealed good reliability, with an intraclass correlation coefficient of 0.712 (95% CI = 0.571, 0.823).ConclusionThe smartphone application walking speed measurement method can be widely used by the general public and is useful for health promotion.

Injectable hydrogels are biomaterials that can be administered minimally invasively in liquid form, and are considered promising artificial extracellular matrix (ECM) materials. However, ordinary injectable hydrogels are synthesized from water-soluble molecules to ensure injectability, resulting in nonphase-separated structures, making them structurally different from natural ECMs with phase-separated insoluble structural proteins, such as collagen and elastin. Here, we propose a simple material design approach to impart phase-separated structures to injectable hydrogels by adding inorganic salts. Injecting a gelling solution of mutually crosslinkable tetra-arm poly(ethylene glycol)s with potassium sulfate at optimal concentrations results in the formation of a hydrogel with phase-separated structures in situ. These phase-separated structures provide up to an 8-fold increase in fracture toughness while allowing the encapsulation of live mouse chondrogenic cells without compromising their proliferative activity. Our findings highlight that the concentration of inorganic salts is an important design parameter in injectable hydrogels for artificial ECMs.

Activation of skeletal progenitors upon tissue injury and the subsequent cell fate specification are tightly coordinated in the bone repair process. Although known osteoimmunological signaling networks play important roles in the microenvironment of the bone defect sites, the molecular mechanism underlying the bone repair process has not been fully understood.To better understand the behavior of the skeletal progenitors and the heterogeneity of the cells during bone repair at the microenvironmental level, we performed a combinatorial analysis consisting of lineage tracing for skeletal progenitors using theLineage tracing analysis showed that skeletal progenitors marked by Sox9 were activated upon bone injury and contributed to bone repair by differentiating into osteoblasts. The scRNA-seq analysis characterized heterogeneous cell populations at the bone defect sites; the computational analysis predicted a bifurcated lineage from skeletal progenitors toward osteogenic and adipogenic lineages. Chemokine C-C motif ligand 9 (Ccl9) was identified as a signaling molecule that regulates bone regeneration in the mouse model, possibly through the regulation of adipogenic differentiation at the bone defect site.Multipotential skeletal progenitors and the direction of the cell differentiation were characterized at single cell resolution in a mouse bone repair model. The Ccl9 signaling pathway may be a key factor directing osteogenesis from the progenitors in the model and may be a therapeutic target for bone regeneration.

(1) Background: This study examined the cross-sectional association between metabolic syndrome (MetS) status classified into three groups and daily physical activity (PA; step count and active minutes) using a wearable device in Japanese office workers. (2) Methods: This secondary analysis used data from 179 participants in the intervention group of a randomized controlled trial for 3 months. Individuals who had received an annual health check-up and had MetS or were at a high risk of MetS based on Japanese guidelines were asked to use a wearable device and answer questionnaires regarding their daily life for the entire study period. Multilevel mixed-effects logistic regression models adjusted for covariates associated with MetS and PA were used to estimate associations. A sensitivity analysis investigated the associations between MetS status and PA level according to the day of the week. (3) Results: Compared to those with no MetS, those with MetS were not significantly associated with PA, while those with pre-MetS were inversely associated with PA [step count Model 3: OR = 0.60; 95% CI: 0.36, 0.99; active minutes Model 3: OR = 0.62; 95% CI: 0.40, 0.96]. In the sensitivity analysis, day of the week was an effect modifier for both PA (p < 0.001). (4) Conclusions: Compared to those with no MetS, those with pre-MetS, but not MetS, showed significantly lower odds of reaching their daily recommended PA level. Our findings suggest that the day of the week could be a modifier for the association between MetS and PA. Further research with longer study periods and larger sample sizes are needed to confirm our results.

Physiological time series are affected by many factors, making them highly nonlinear and nonstationary. As a consequence, heart rate time series are often considered difficult to predict and handle. However, heart rate behavior can indicate underlying cardiovascular and respiratory diseases as well as mood disorders. Given the importance of accurate modeling and reliable predictions of heart rate fluctuations for the prevention and control of certain diseases, it is paramount to identify models with the best performance in such tasks. The objectives of this study were to compare the results of three different forecasting models (Autoregressive Model, Long Short-Term Memory Network, and Convolutional Long Short-Term Memory Network) trained and tested on heart rate beats per minute data obtained from twelve heterogeneous participants and to identify the architecture with the best performance in terms of modeling and forecasting heart rate behavior. Heart rate beats per minute data were collected using a wearable device over a period of 10 days from twelve different participants who were heterogeneous in age, sex, medical history, and lifestyle behaviors. The goodness of the results produced by the models was measured using both the mean absolute error and the root mean square error as error metrics. Despite the three models showing similar performance, the Autoregressive Model gave the best results in all settings examined. For example, considering one of the participants, the Autoregressive Model gave a mean absolute error of 2.069 (compared to 2.173 of the Long Short-Term Memory Network and 2.138 of the Convolutional Long Short-Term Memory Network), achieving an improvement of 5.027% and 3.335%, respectively. Similar results can be observed for the other participants. The findings of the study suggest that regardless of an individual’s age, sex, and lifestyle behaviors, their heart rate largely depends on the pattern observed in the previous few minutes, suggesting that heart rate can be reasonably regarded as an autoregressive process. The findings also suggest that minute-by-minute heart rate prediction can be accurately performed using a linear model, at least in individuals without pathologies that cause heartbeat irregularities. The findings also suggest many possible applications for the Autoregressive Model, in principle in any context where minute-by-minute heart rate prediction is required (arrhythmia detection and analysis of the response to training, among others).

The transcriptional regulator Runx2 (runt-related transcription factor 2) has essential but distinct roles in osteoblasts and chondrocytes in skeletal development. However, Runx2-mediated regulatory mechanisms underlying the distinctive programming of osteoblasts and chondrocytes are not well understood. Here, we perform an integrative analysis to investigate Runx2-DNA binding and chromatin accessibility ex vivo using neonatal osteoblasts and chondrocytes. We find that Runx2 engages with cell-type-distinct chromatin-accessible regions, potentially interacting with different combinations of transcriptional regulators, forming cell-type-specific hotspots, and potentiating chromatin accessibility. Genetic analysis and direct cellular reprogramming studies suggest that Runx2 is essential for establishment of chromatin accessibility in osteoblasts. Functional enhancer studies identify an Sp7 distal enhancer driven by Runx2-dependent binding and osteoblast-specific chromatin accessibility, contributing to normal osteoblast differentiation. Our findings provide a framework for understanding the regulatory landscape encompassing Runx2-mediated and cell-type-distinct enhancer networks that underlie the specification of osteoblasts.

To determine whether the walking pattern of an organism is a Lévy walk or a Brownian walk, it has been compared whether the frequency distribution of linear step lengths follows a power law distribution or an exponential distribution. However, there are many cases where actual data cannot be classified into either of these categories. In this paper, we propose a general distribution that includes the power law and exponential distributions as special cases. This distribution has two parameters: One represents the exponent, similar to the power law and exponential distributions, and the other is a shape parameter representing the shape of the distribution. By introducing this distribution, an intermediate distribution model can be interpolated between the power law and exponential distributions. In this study, the proposed distribution was fitted to the frequency distribution of the step length calculated from the walking data of pill bugs. The autocorrelation coefficients were also calculated from the time-series data of the step length, and the relationship between the shape parameter and time dependency was investigated. The results showed that individuals whose step-length frequency distributions were closer to the power law distribution had stronger time dependence.

The Runt-related transcription factor (Runx) family plays various roles in the homeostasis of cartilage. Here, we examined the role of Runx2 and Runx3 for osteoarthritis (OA) development in vivo and in vitro. Runx3 knockout mice accelerated OA by surgical induction, accompanied with decreased expression of lubricin and aggrecan. Meanwhile, Runx2 conditional knockout mice showed biphasic phenotypes; OA was inhibited by hetero-knockout accompanied with decreased matrix metallopeptidase 13 (Mmp13) expression, but accelerated in homo-knockout of Runx2 accompanied with reduction of type II collagen (Col2a1) expression. Comprehensive transcriptional analyses revealed lubricin and aggrecan as transcriptional target genes of Runx3, and indicated that Runx2 sustained Col2a1 expression through an intron 6 enhancer when Sox9 was decreased. Intra-articular administration of Runx3 adenovirus ameliorated development of surgically induced OA. Runx3 protects adult articular cartilage through extracellular matrix protein production under the normal condition, while Runx2 exerts both catabolic and anabolic effects under the inflammatory condition.

Dynamically crosslinked gels are appealing materials for applications that require time-dependent mechanical responses. DNA duplexes are ideal crosslinkers for building such gels because of their excellent sequence addressability and flexible tunability in bond energy. However, the mechanical responses of most DNA gels are complicated and unpredictable despite the high potential of DNA. Here, we demonstrate a DNA gel with a highly homogeneous gel network and well-predictable mechanical behaviors by using a pair of star-polymer-DNA precursors with presimulated DNA sequences showing the two-state transition. The melting curve analysis of the DNA gels reveals the good correspondence between the thermodynamic potentials of the DNA crosslinkers and the presimulated values by DNA calculators. Stress-relaxation tests and dissociation kinetics measurements show that the macroscopic relaxation time of the DNA gels is approximately equal to the lifetime of the DNA crosslinkers over four orders of magnitude from 0.1-2,000 sec. Furthermore, a series of durability tests find the DNA gels are hysteresis-less and self-healable after the applications of repeated temperature and mechanical stimuli. These results demonstrate the great potential of star-polymer-DNA precursors for building gels with predictable and tunable viscoelastic properties, suitable for applications such as stress-response extracellular matrices, injectable solids, and soft robotics.

Objective The aim of the study was to investigate the associations of total sleep time (TST) and occupational stress based on the Brief Job Stress Questionnaire (BJSQ) with cholesterol levels in an occupational cohort of Japanese office workers. Methods The present study is a secondary analysis of a subset of participants from a randomized controlled trial. Participants were 179 employees from 5 companies in Tokyo who participated as the intervention group in a 3‐month lifestyle intervention study among office workers with metabolic syndrome or at risk of metabolic syndrome. All intervention‐group participants used a mobile app and a wearable device. The final population for analysis in the present study were 173 participants. Cholesterol measures were derived from participants' annual health check‐up data in the fiscal year preceding their inclusion in the study. Multiple linear regression models were used to determine the association between exposures and outcome. Results Overall, stress levels were significantly and inversely associated with LDL‐C (−7.12 mg/dl; 95% CI: −11.78, −2.45) and LDL‐C/HDL‐C ratio (−0.16 mg/dl; 95% CI: −0.27, −0.04) per standard deviation increase. Compared to average TST 5.9‐7.2 hours, average TST of 4.0‐5.3 hours (−4.82 mg/dl; 95% CI: −9.22, −0.43) was inversely associated with HDL‐C. Conclusion Incremental increases of stress were significantly and inversely associated with LDL‐C and LDL‐C/HDL‐C ratio. The shortest average TST was inversely associated with HDL‐C. The results should be interpreted with care given certain methodological limitations.

Dynamics of a polymer gel network is described by the theory proposed by Tanaka, Hocker, and Benedek (THB) that gives the diffusion coefficient of a polymer network (D=K+43Gf). Here, K is the osmotic bulk modulus, G is the shear modulus, and f is the friction coefficient between the polymer network and the solvent. Although several experimental studies investigated the dynamics of a polymer gel network, the THB theory has yet to be quantitatively validated. In this study, we quantitatively validate the THB theory by comparing D and G of Tetra-poly(ethylene glycol) gels measured by dynamic light scattering and dynamic viscoelastic measurement. The THB theory well described the linear increase in D with increasing G and gave K and f from the slope and the intercept. The gel network structural parameter dependences of the given K and f were well explained by the scaling laws. These results suggest that the THB theory is almost valid.

For the application of polymer gels, it is necessary to control independently and precisely their various physical properties. However, the heterogeneity of polymer gels hinders the precise control over the structure, as well as the verification of theories. To understand the structure-property relationship of polymer gels, many researchers have tried to develop a homogeneous model network. Most of the model networks were made from polymer melts that did not have a solvent and had many entanglements in the structure. Because the contribution of entanglements is much larger than that of chemical crosslinking, it was difficult to focus on the crosslinking structure, which is the structure considered in conventional theories. To overcome such a situation, we have developed a new model network system that contains much solvent. Specifically, we fabricated the polymer gel (Tetra-PEG gel) by mixing two types of solutions of tetra-armed poly(ethylene glycol) (Tetra-PEG) with mutually reactive end groups (amine (-PA) and activated ester (-HS)). Because the existence of a solvent strongly reduces the effect of entanglements, the effect of the crosslinking structure on the physical properties can be extracted. In this review, we present the structure-property relationship of Tetra-PEG gel. First, we show the structural homogeneity of Tetra-PEG gels. Then, we explain gelation reaction, elastic modulus, fracture energy and kinetics of swelling and shrinking of Tetra-PEG gels by comparing the theories and experimental results., Graphical abstract

The gelation process has been explained in the framework of percolation models. Although the percolation models accurately predict the gelation behaviors above the overlap concentration of prepolymers (c*0), they do not predict the gelation behaviors below c*0. We measured the osmotic pressure during the gelation of a series of model polymer gels, namely, tetra-polyethylene glycol (PEG) gels. The osmotic pressure decreased during the gelation reaction and was constant after the sol–gel transition. These results suggest that the clusters grow and fill the system at the sol–gel transition point. As a result, the gels behaved as semidilute systems regardless of the initial polymer concentration. This representation of the sol–gel transition at the overlap condition of the critical clusters corresponds well to the aggregation process predictrion. These results will help better understanding of general percolation problems in the dilute regime. The osmotic pressure during the gelation process below the overlapping concentration of prepolymers was investigated using a series of model polymer gels, namely, tetra-polyethylene glycol gels. The osmotic pressure decreased during the gelation reaction and was constant after the sol–gel transition, suggesting that the clusters grow and fill the system, at the sol–gel transition point. This representation of the sol–gel transition at the overlapping condition of the critical clusters corresponds well to the aggregation process prediction.

Summary A simple induction protocol to differentiate chondrocytes from pluripotent stem cells (PSCs) using small-molecule compounds is beneficial for cartilage regenerative medicine and mechanistic studies of chondrogenesis. Here, we demonstrate that chondrocytes are robustly induced from human PSCs by simple combination of two compounds, CHIR99021, a glycogen synthase kinase 3 inhibitor, and TTNPB, a retinoic acid receptor (RAR) agonist, under serum- and feeder-free conditions within 5–9 days. An excellent differentiation efficiency and potential to form hyaline cartilaginous tissues in vivo were demonstrated. Comprehensive gene expression and open chromatin analyses at each protocol stage revealed step-by-step differentiation toward chondrocytes. Genome-wide analysis of RAR and β-catenin association with DNA showed that retinoic acid and Wnt/β-catenin signaling collaboratively regulated the key marker genes at each differentiation stage. This method provides a promising cell source for regenerative medicine and, as an in vitro model, may facilitate elucidation of the molecular mechanisms underlying chondrocyte differentiation., Graphical Abstract, Highlights • Combination of a GSK3 inhibitor and an RAR agonist induces chondrocytes from hPSCs • The present protocol shows excellent differentiation efficiency • The potential to form hyaline cartilaginous tissues in vivo is demonstrated • RA and Wnt/β-catenin signals collaboratively regulate key genes for chondrogenesis, Saito and colleagues show that chondrocytes are robustly induced from human PSCs by simple combination of two compounds, a GSK3 inhibitor and an RAR agonist, within 5–9 days. Genome-wide analysis of RAR and β-catenin association with DNA for sequential samples at each protocol stage demonstrates that RA and Wnt/β-catenin signaling is collaboratively involved in direct regulation of chondrocyte differentiation.