Aging effects on osteoclast progenitor dynamics affect variability in bone turnover via feedback regulation.

Bone turnover markers (BTMs) are commonly used in osteoporosis treatment as indicators of cell activities of bone-resorbing osteoclasts and bone-forming osteoblasts. The wide variability in their values due to multiple factors, such as aging and diseases, makes it difficult for physicians to utilize them for clinical decision-making. The progenitors of osteoclasts and osteoblasts are indispensable for a comprehensive interpretation of the variability in BTM values because these upstream progenitors strongly regulate the downstream cell activities of bone turnover. However, understanding the complex interactions among the multiple populations of bone cells is challenging. In this study, we aimed to gain a fundamental understanding of the mechanism by which the progenitor dynamics affect the variability in bone turnover through in silico experiments by exploring the cell dynamics with aging effects on osteoporosis. Negative feedback control driven by the consumptive loss of progenitors prevents rapid bone loss due to excessive bone turnover, and through feedback regulation, aging effects on osteoclast differentiation and osteoclast progenitor proliferation cause variability in the osteoclast and osteoblast activity balance and its temporal transition. By expressing the variability in the bone turnover status, our model describes the individualities of patients based on their clinical backgrounds. Therefore, our model could play a powerful role in assisting tailored treatment and has the potential to resolve the various health problems associated with osteoporosis worldwide.

Spatial cell fate manipulation of human pluripotent stem cells by controlling the microenvironment using photocurable hydrogel.

Human pluripotent stem cells (hPSCs) dynamically respond to their chemical and physical microenvironment, dictating their behavior. However, conventional in vitro studies predominantly employ plastic culture wares, which offer a simplified representation of the in vivo microenvironment. Emerging evidence underscores the pivotal role of mechanical and topological cues in hPSC differentiation and maintenance. In this study, we cultured hPSCs on hydrogel substrates with spatially controlled stiffness. The use of culture substrates that enable precise manipulation of spatial mechanical properties holds promise for better mimicking in vivo conditions and advancing tissue engineering techniques. We designed a photocurable polyethylene glycol-polyvinyl alcohol (PVA-PEG) hydrogel, allowing the spatial control of surface stiffness and geometry at a micrometer scale. This versatile hydrogel can be functionalized with various extracellular matrix proteins. Laminin 511-functionalized PVA-PEG gel effectively supports the growth and differentiation of hPSCs. Moreover, by spatially modulating the stiffness of the patterned gel, we achieved spatially selective cell differentiation, resulting in the generation of intricate patterned structures.

Mathematical expansion and clinical application of chronic kidney disease stage as vector field.

There are cases in which CKD progression is difficult to evaluate, because the changes in estimated glomerular filtration rate (eGFR) and proteinuria sometimes show opposite directions as CKD progresses. Indices and models that enable the easy and accurate risk prediction of end-stage-kidney disease (ESKD) are indispensable to CKD therapy. In this study, we investigated whether a CKD stage coordinate transformed into a vector field (CKD potential model) accurately predicts ESKD risk. Meta-analysis of large-scale cohort studies of CKD patients in PubMed was conducted to develop the model. The distance from CKD stage G2 A1 to a patient's data on eGFR and proteinuria was defined as r. We developed the CKD potential model on the basis of the data from the meta-analysis of three previous cohort studies: ESKD risk = exp(r). Then, the model was validated using data from a cohort study of CKD patients in Japan followed up for three years (n = 1,564). Moreover, the directional derivative of the model was developed as an index of CKD progression velocity. For ESKD prediction in three years, areas under the receiver operating characteristic curves (AUCs) were adjusted for baseline characteristics. Cox proportional hazards models with spline terms showed the exponential association between r and ESKD risk (p<0.0001). The CKD potential model more accurately predicted ESKD with an adjusted AUC of 0.81 (95% CI 0.76, 0.87) than eGFR (p<0.0001). Moreover, the directional derivative of the model showed a larger adjusted AUC for the prediction of ESKD than the percent eGFR change and eGFR slope (p<0.0001). Then, a chart of the transformed CKD stage was developed for implementation in clinical settings. This study indicated that the transformed CKD stage as a vector field enables the easy and accurate estimation of ESKD risk and CKD progression and suggested that vector analysis is a useful tool for clinical studies of CKD and its related diseases.

Chondrocyte hypertrophy in the growth plate promotes stress anisotropy affecting long bone development through chondrocyte column formation.

The length of long bones is determined by column formation of proliferative chondrocytes and subsequent chondrocyte hypertrophy in the growth plate during bone development. Despite the importance of mechanical loading in long bone development, the mechanical conditions of the cells within the growth plate, such as the stress field, remain unclear owing to the difficulty in investigating spatiotemporal changes within dynamically growing tissues. In this study, the mechanisms of longitudinal bone growth were investigated from a mechanical perspective through column formation of proliferative chondrocytes within the growth plate before secondary ossification center formation using continuum-based particle models (CbPMs). A one-factor model, which simply describes essential aspects of a biological signaling cascade regulating cell activities within the growth plate, was developed and incorporated into CbPM. Subsequently, the developmental process and maintenance of the growth plate structure and resulting bone morphogenesis were simulated. Thus, stress anisotropy in the proliferative zone that affects bone elongation through chondrocyte column formation was identified and found to be promoted by chondrocyte hypertrophy. These results provide further insights into the mechanical regulation of multicellular dynamics during bone development.

New marker for chronic kidney disease progression and mortality in medical-word virtual space.

A new marker reflecting the pathophysiology of chronic kidney disease (CKD) has been desired for its therapy. In this study, we developed a virtual space where data in medical words and those of actual CKD patients were unified by natural language processing and category theory. A virtual space of medical words was constructed from the CKD-related literature (n = 165,271) using Word2Vec, in which 106,612 words composed a network. The network satisfied vector calculations, and retained the meanings of medical words. The data of CKD patients of a cohort study for 3 years (n = 26,433) were transformed into the network as medical-word vectors. We let the relationship between vectors of patient data and the outcome (dialysis or death) be a marker (inner product). Then, the inner product accurately predicted the outcomes: C-statistics of 0.911 (95% CI 0.897, 0.924). Cox proportional hazards models showed that the risk of the outcomes in the high-inner-product group was 21.92 (95% CI 14.77, 32.51) times higher than that in the low-inner-product group. This study showed that CKD patients can be treated as a network of medical words that reflect the pathophysiological condition of CKD and the risks of CKD progression and mortality.

The nucleolar shell provides anchoring sites for DNA untwisting.

DNA underwinding (untwisting) is a crucial step in transcriptional activation. DNA underwinding occurs between the site where torque is generated by RNA polymerase (RNAP) and the site where the axial rotation of DNA is constrained. However, what constrains DNA axial rotation in the nucleus is yet unknown. Here, we show that the anchorage to the nuclear protein condensates constrains DNA axial rotation for DNA underwinding in the nucleolus. In situ super-resolution imaging of underwound DNA reveal that underwound DNA accumulates in the nucleolus, a nuclear condensate with a core-shell structure. Specifically, underwound DNA is distributed in the nucleolar core owing to RNA polymerase I (RNAPI) activities. Furthermore, underwound DNA in the core decreases when nucleolar shell components are prevented from binding to their recognition structure, G-quadruplex (G4). Taken together, these results suggest that the nucleolar shell provides anchoring sites that constrain DNA axial rotation for RNAPI-driven DNA underwinding in the core. Our findings will contribute to understanding how nuclear protein condensates make up constraints for the site-specific regulation of DNA underwinding and transcription.

Cyclic Stretching Enhances Angiocrine Signals at Liver Bud Stage from Human Pluripotent Stem Cells in Two-Dimensional Culture.

Angiocrine signals during the development and growth of organs, including the liver, intestine, lung, and bone, are essential components of intercellular communication. The signals elicited during the liver bud stage are critical for vascularization and enhanced during the intercellular communication between the cells negative for kinase insert domain receptor (KDR) (KDR- cells) and the cells positive for KDR (KDR+ cells), which constitute the liver bud. However, the use of a human pluripotent stem cell (hPSC)-derived system has not facilitated the generation of a perfusable vascularized liver organoid that allows elucidation of liver development and has great potential for liver transplantation. This is largely owing to the lack of fundamental understanding to induce angiocrine signals in KDR- and KDR+ cells during the liver bud stage. We hypothesized that mechanical stimuli of cyclic stretching/pushing by the fetal heart adjacent to the liver bud could be the main contributor to promoting angiocrine signals in KDR- and KDR+ cells during the liver bud stage. In this study, we show that an organ-on-a-chip platform allows the emulation of an in vivo-like mechanical environment for the liver bud stage in vitro and investigate the role of cyclic mechanical stretching (cMS) to angiocrine signals in KDR- and KDR+ cells derived from hPSCs. RNA sequencing revealed that the expression of genes associated with epithelial-to-mesenchymal transition, including angiocrine signals, such as hepatocyte growth factor (HGF) and matrix metallopeptidase 9 (MMP9), were increased by cMS in cocultured KDR- and KDR+ cells. The expression and secretions of HGF and MMP9 were increased by 1.98- and 1.69-fold and 3.23- and 3.72-fold with cMS in the cocultured KDR- and KDR+ cells but were not increased by cMS in the monocultured KDR- and KDR+ cells, respectively. Finally, cMS during the liver bud stage did not lead to the dedifferentiation of hepatocytes, as the cells with cMS showed hepatic maker expression (CYP3A4, CYP3A7, ALB, and AAT) and 1.71-fold higher CYP3A activity than the cells without cMS, during 12 day-hepatocyte maturation after halting cMS. Our findings provide new insights into the mechanical factors during the liver bud stage and directions for future improvements in the engineered liver tissue.

Profiling to Probing: Atomic force microscopy to characterize nano-engineered implants.

Surface modification of implants in the nanoscale or implant nano-engineering has been recognized as a strategy for augmenting implant bioactivity and achieving long-term implant success. Characterizing and optimizing implant characteristics is crucial to achieving desirable effects post-implantation. Modified implant enables tailored, guided and accelerated tissue integration; however, our understanding is limited to multicellular (bulk) interactions. Finding the nanoscale forces experienced by a single cell on nano-engineered implants will aid in predicting implants' bioactivity and engineering the next generation of bioactive implants. Atomic force microscope (AFM) is a unique tool that enables surface characterization and understanding of the interactions between implant surface and biological tissues. The characterization of surface topography using AFM to gauge nano-engineered implants' characteristics (topographical, mechanical, chemical, electrical and magnetic) and bioactivity (adhesion of cells) is presented. A special focus of the review is to discuss the use of single-cell force spectroscopy (SCFS) employing AFM to investigate the minute forces involved with the adhesion of a single cell (resident tissue cell or bacterium) to the surface of nano-engineered implants. Finally, the research gaps and future perspectives relating to AFM-characterized current and emerging nano-engineered implants are discussed towards achieving desirable bioactivity performances. This review highlights the use of advanced AFM-based characterization of nano-engineered implant surfaces via profiling (investigating implant topography) or probing (using a single cell as a probe to study precise adhesive forces with the implant surface). STATEMENT OF SIGNIFICANCE: Nano-engineering is emerging as a surface modification platform for implants to augment their bioactivity and achieve favourable treatment outcomes. In this extensive review, we closely examine the use of Atomic Force Microscopy (AFM) to characterize the properties of nano-engineered implant surfaces (topography, mechanical, chemical, electrical and magnetic). Next, we discuss Single-Cell Force Spectroscopy (SCFS) via AFM towards precise force quantification encompassing a single cell's interaction with the implant surface. This interdisciplinary review will appeal to researchers from the broader scientific community interested in implants and cell adhesion to implants and provide an improved understanding of the surface characterization of nano-engineered implants.

Age-associated reduction of nuclear shape dynamics in excitatory neurons of the visual cortex.

Neurons decline in their functionality over time, and age-related neuronal alterations are associated with phenotypes of neurodegenerative diseases. In nonneural tissues, an infolded nuclear shape has been proposed as a hallmark of aged cells and neurons with infolded nuclei have also been reported to be associated with neuronal activity. Here, we performed time-lapse imaging in the visual cortex of Nex-Cre;SUN1-GFP mice. Nuclear infolding was observed within 10 min of stimulation in young nuclei, while the aged nuclei were already infolded pre-stimulation and showed reduced dynamics of the morphology. In young nuclei, the depletion of the stimuli restored the nucleus to a spherical shape and reduced the dynamic behavior, suggesting that nuclear infolding is a reversible process. We also found the aged nucleus to be stiffer than the young one, further relating to the age-associated loss of nuclear shape dynamics. We reveal temporal changes in the nuclear shape upon external stimulation and observe that these morphological dynamics decrease with age.

Directionality of developing skeletal muscles is set by mechanical forces.

Formation of oriented myofibrils is a key event in musculoskeletal development. However, the mechanisms that drive myocyte orientation and fusion to control muscle directionality in adults remain enigmatic. Here, we demonstrate that the developing skeleton instructs the directional outgrowth of skeletal muscle and other soft tissues during limb and facial morphogenesis in zebrafish and mouse. Time-lapse live imaging reveals that during early craniofacial development, myoblasts condense into round clusters corresponding to future muscle groups. These clusters undergo oriented stretch and alignment during embryonic growth. Genetic perturbation of cartilage patterning or size disrupts the directionality and number of myofibrils in vivo. Laser ablation of musculoskeletal attachment points reveals tension imposed by cartilage expansion on the forming myofibers. Application of continuous tension using artificial attachment points, or stretchable membrane substrates, is sufficient to drive polarization of myocyte populations in vitro. Overall, this work outlines a biomechanical guidance mechanism that is potentially useful for engineering functional skeletal muscle.

Development of continuum-based particle models of cell growth and proliferation for simulating tissue morphogenesis.

Biological tissues acquire various characteristic shapes through morphogenesis. Tissue shapes result from the spatiotemporally heterogeneous cellular activities influenced by mechanical and biochemical environments. To investigate multicellular tissue morphogenesis, this study aimed to develop a novel multiscale method that can connect each cellular activity to the mechanical behaviors of the whole tissue by constructing continuum-based particle models of cellular activities. This study proposed mechanical models of cell growth and proliferation that are expressed as volume expansion and cell division by extending the material point method. By simulating cell hypertrophy and proliferation under both free and constraint conditions, the proposed models demonstrated potential for evaluating the mechanical state and tracing cells throughout tissue morphogenesis. Moreover, the effect of a cell size checkpoint was incorporated into the cell proliferation model to investigate the mechanical behaviors of the whole tissue depending on the condition of cellular activities. Consequently, the accumulation of strain energy density was suppressed because of the influence of the checkpoint. In addition, the whole tissues acquired different shapes depending on the influence of the checkpoint. Thus, the models constructed herein enabled us to investigate the change in the mechanical behaviors of the whole tissue according to each cellular activity depending on the mechanical state of the cells during morphogenesis.

Machine-learning-based Web system for the prediction of chronic kidney disease progression and mortality.

Chronic kidney disease (CKD) patients have high risks of end-stage kidney disease (ESKD) and pre-ESKD death. Therefore, accurately predicting these outcomes is useful among CKD patients, especially in those who are at high risk. Thus, we evaluated whether a machine-learning system can predict accurately these risks in CKD patients and attempted its application by developing a Web-based risk-prediction system. We developed 16 risk-prediction machine-learning models using Random Forest (RF), Gradient Boosting Decision Tree, and eXtreme Gradient Boosting with 22 variables or selected variables for the prediction of the primary outcome (ESKD or death) on the basis of repeatedly measured data of CKD patients (n = 3,714; repeatedly measured data, n = 66,981) in their electronic-medical records. The performances of the models were evaluated using data from a cohort study of CKD patients carried out over 3 years (n = 26,906). One RF model with 22 variables and another RF model with 8 variables of time-series data showed high accuracies of the prediction of the outcomes and were selected for use in a risk-prediction system. In the validation, the 22- and 8-variable RF models showed high C-statistics for the prediction of the outcomes: 0.932 (95% CI 0.916, 0.948) and 0.93 (0.915, 0.945), respectively. Cox proportional hazards models using splines showed a highly significant relationship between the high probability and high risk of an outcome (p<0.0001). Moreover, the risks of patients with high probabilities were higher than those with low probabilities: 22-variable model, hazard ratio of 104.9 (95% CI 70.81, 155.3); 8-variable model, 90.9 (95% CI 62.29, 132.7). Then, a Web-based risk-prediction system was actually developed for the implementation of the models in clinical practice. This study showed that a machine-learning-based Web system is a useful tool for the risk prediction and treatment of CKD patients.

3D osteogenic differentiation of human iPSCs reveals the role of TGFß signal in the transition from progenitors to osteoblasts and osteoblasts to osteocytes.

Although the formation of bone-like nodules is regarded as the differentiation process from stem cells to osteogenic cells, including osteoblasts and osteocytes, the precise biological events during nodule formation are unknown. Here we performed the osteogenic induction of human induced pluripotent stem cells using a three-dimensional (3D) culture system using type I collagen gel and a rapid induction method with retinoic acid. Confocal and time-lapse imaging revealed the osteogenic differentiation was initiated with vigorous focal proliferation followed by aggregation, from which cells invaded the gel. Invading cells changed their morphology and expressed osteocyte marker genes, suggesting the transition from osteoblasts to osteocytes. Single-cell RNA sequencing analysis revealed that 3D culture-induced cells with features of periosteal skeletal stem cells, some of which expressed TGFß-regulated osteoblast-related molecules. The role of TGFß signal was further analyzed in the transition from osteoblasts to osteocytes, which revealed that modulation of the TGFß signal changed the morphology and motility of cells isolated from the 3D culture, suggesting that the TGFß signal maintains the osteoblastic phenotype and the transition into osteocytes requires down-regulation of the TGFß signal.

Spheroid culture for chondrocytes triggers the initial stage of endochondral ossification.

Endochondral ossification is the process of bone formation derived from growing cartilage duringskeletal development. In previous studies, we provoked the osteocyte differentiation of osteoblast precursor cells under a three-dimensional (3D) culture model. To recapitulate the endochondral ossification, the present study utilized the self-organized scaffold-free spheroid model reconstructed by pre-chondrocyte cells. Within 2-day cultivation in the absence of the chemically induced chondrogenesis supplements, the chondrocyte marker was greatly expressed in the inner region of the spheroid, whereas the hypertrophic chondrocyte marker was strongly detected in the surface region of the spheroid. Notably, we found out that the gene expression levels of osteocyte markers were also greatly upregulated compared to the conventional 2D monolayer. Moreover, after long-term cultivation for 28 days, it induced morphological changes in the spheroid, such as cellular hypertrophy and death. In this study, in order to recapitulate the initial stage of the endochondral ossification, we highlighted the potentials of the 3D culture method to drive the hypertrophic chondrocyte differentiation of the pre-chondrocyte cells.

Effect of chemically induced osteogenesis supplements on multicellular behavior of osteocytic spheroids.

Understanding in multicellular behaviors in three-dimensional (3D) culture models such as organoids is important to help us better comprehend the mechanisms of the morphogenesis and functions of diverse organs in vivo cellular environment. In this study, we elucidated the multicellular behaviors of the osteocytic spheroids in response to the chemically induced osteogenesis supplements (OS). Particularly, we conducted 1) size change measurement, 2) fusion experiment, and 3) collagen embedding experiment of spheroids, in response to the OS. We found out that the OS alters the multicellular behaviors of the spheroid by greater reduction in the size change measurement and slowing down the speed of fusion experiment and collagen embedding experiment of the spheroids. We also highlighted that the driving force of these changes was the tight actin filaments generated on the surface of the spheroids. Hence, the results altogether indicate that the spheroid model exerted the different multicellular behaviors against the differentiation capability. This study will contribute to understanding the multicellular behaviors of the 3D culture model reconstructed by the cells with greater cell-cell interaction force.

Pluripotency state of mouse ES cells determines their contribution to self-organized layer formation by mesh closure on microstructured adhesion-limiting substrates.

Assembly of pluripotent stem cells to initiate self-organized tissue formation on engineered scaffolds is an important process in stem cell engineering. Pluripotent stem cells are known to exist in diverse pluripotency states, with heterogeneous subpopulations exhibiting differential gene expression levels, but how such diverse pluripotency states orchestrate tissue formation is still an unrevealed question. In this study, using microstructured adhesion-limiting substrates, we aimed to clarify the contribution to self-organized layer formation by mouse embryonic stem cells in different pluripotency states: ground and naïve state. We found that while ground state cells as well as sorted REX1-high expression cells formed discontinuous cell layers with limited lateral spread, naïve state cells could successfully self-organize to form a continuous layer by progressive mesh closure within 3 days. Using sequential immunofluorescence microscopy to examine the mesh closure process, we found that KRT8+ cells were particularly localized around unfilled holes, occasionally bridging the holes in a manner suggestive of their role in the closure process. These results highlight that compared with ground state cells, naïve state cells possess a higher capability to contribute to self-organized layer formation by mesh closure. Thus, this study provides insights with implications for the application of stem cells in scaffold-based tissue engineering.

Wolverine cutting balloon in the treatment of stent underexpansion in heavy coronary calcification: bench test using a three-dimensional printer and computer simulation with the finite-element method.

Heavy coronary calcification hinders successful stent implantation, and cutting balloons can be used for post-dilation after stent deployment. However, evidence regarding its use is limited to case reports. Therefore, this study aimed to investigate in-stent dilation in circumferential coronary calcifications using Wolverine cutting balloons, compared with conventional non-compliance (NC) balloons. Circumferential coronary calcification models were designed based on the patient's intravascular ultrasound images. Three-dimensional printed models were subjected to bench tests and software analysis was performed using the finite-element method (FEM). As a result, the bench test showed that higher balloon pressure was needed to dilate the models with stent implantation, either using Wolverine (17.1 ± 2.7 atm) or NC Emerge (18.9 ± 1.8 atm), while lower pressure was needed in models without stents using Wolverine [11.7 ± 2.9 atm, analysis of variance (ANOVA) p < 0.001]. Furthermore, models without stents were all successfully cracked by Wolverine at the first dilation, while models with stent implantation needed more dilations (ANOVA p = 0.0132). The FEM showed similar results that the first principal stress was the highest in Wolverine-dilated models without stents. In conclusion, implanted stents significantly increase the difficulty of balloon dilation and adequate pretreatment is critical for successful coronary stenting.

Efficacy of the Wolverine cutting balloon on a circumferential calcified coronary lesion: Bench test using a three-dimensional printer and computer simulation with the finite element method.

Heavy calcification is one of the factors that hinder the success of coronary angioplasty, and a cutting balloon is used for such lesions. This study aimed to explore the optimal method of dilation of highly calcified lesions using a cutting balloon. Calcification models were developed from patient computed tomography and intravascular ultrasound data, and were constructed using three-dimensional printers. The lesions were dilated using a Wolverine™ cutting balloon and NC Emerge™ noncompliant balloon catheter, and the success rate of dilation and maximum dilation pressure were compared. The maximum first principal stresses in calcified lesions were also evaluated by computer simulation using the finite element method. In the bench test, the dilation success rate of the Wolverine™ cutting balloon was higher and the maximum dilation pressure required was lower (p < 0.01 in all analyses), compared with that of the NC Emerge™ balloon catheter. Finite element analysis showed that the cutting blade increased the maximum first principal stresses in calcified lesions, thus allowing for successful dilation at low pressures. The highest stress was obtained when the cutting blade was positioned at the thinnest part of the calcification. The cutting balloon allows for efficient calcification expansion by concentrating the stresses in the blade. When a cutting balloon is used, if the calcified lesion cannot be expanded in a single dilation, dilation success may be achieved after the balloon is rotated and the position of the blade is changed.

Computational framework for analyzing flow-induced strain on osteocyte as modulated by microenvironment.

Osteocytes buried in bone matrix are major mechanosensory cells that regulate bone remodeling in response to interstitial fluid flow in a lacuno-canalicular porosity. To gain an understanding of the mechanism of osteocyte mechanosensing, it is important to be able to evaluate the local strain on the osteocyte process membrane induced by the interstitial fluid flow. The microenvironment of the osteocytes, including the pericellular matrix (PCM) and canalicular ultrastructure, is a key modulator of the flow-induced strain on the osteocyte process membrane because it produces heterogeneous flow patterns in the pericellular space. To investigate the effect of changes in the microenvironment of osteocytes on the flow-induced strain, we developed a novel computational framework for analyzing the fluid-structure interaction. Computer simulations based on the proposed framework enabled evaluation of the spatial distribution of flow-induced strain on the osteocyte process membrane according to changes in the PCM density and canalicular curvature. The simulation results reveal that a decrease in PCM density and an increase in canalicular curvature, each of which is associated with aging and bone disease, have the notable effect of enhancing local flow-induced strain on the osteocyte process membrane. We believe that the proposed computational framework is a promising framework for investigating cell-specific mechanical stimuli and that it has the potential to accelerate the mechanobiological study of osteocytes by providing a deeper understanding of their mechanical environment in living bone tissue.

Vasculature atrophy causes a stiffened microenvironment that augments epidermal stem cell differentiation in aged skin.

Stem cell loss causes tissue deterioration associated with aging. The accumulation of genomic and oxidative stress-induced DNA damage is an intrinsic cue for stem cell loss1,2; however, whether there is an external microenvironmental cue that triggers stem cell loss remains unclear. Here we report that the involution of skin vasculature causes dermal stiffening that augments the differentiation and hemidesmosome fragility of interfollicular epidermal stem cells (IFESCs) in aged mouse skin. Aging-related IFESC dysregulation occurs in plantar and tail skin, and is correlated with prolonged calcium influx, which is contributed by the mechanoresponsive ion channel Piezo1 (ref. 3). Epidermal deletion of Piezo1 ameliorated IFESC dysregulation in aged skin, whereas Piezo1 activation augmented IFESC differentiation and hemidesmosome fragility in young mice. The dermis stiffened with age, which was accompanied by dermal vasculature atrophy. Conversely, induction of the dermal vasculature softened the dermis and ameliorated IFESC dysregulation in aged skin. Single-cell RNA sequencing of dermal fibroblasts identified an aging-associated anti-angiogenetic secretory molecule, pentraxin 3 (ref. 4), which caused dermal sclerotization and IFESC dysregulation in aged skin. Our findings show that the vasculature softens the microenvironment for stem cell maintenance and provide a potential mechanobiology-based therapeutic strategy against skin disorders in aging.