A Novel Long Noncoding RNA in Osteocytes Regulates Bone Formation through the Wnt/ß-Catenin Signaling Pathway.

The vast majority of transcribed RNAs are noncoding RNAs. Among noncoding RNAs, long noncoding RNAs (lncRNAs), which contain hundreds to thousands of bases, have received attention in many fields. The vast majority of the constituent cells in bone tissue are osteocytes, but their regulatory mechanisms are incompletely understood. Considering the wide range of potential contributions of lncRNAs to physiological processes and pathological conditions, we hypothesized that lncRNAs in osteocytes, which have not been reported, could be involved in bone metabolism. Here, we first isolated osteocytes from femurs of mice with osteocyte-specific GFP expression. Then, through RNA-sequencing, we identified osteocyte-specific lncRNAs and focused on a novel lncRNA, 9530026P05Rik (lncRNA953Rik), which strongly suppressed osteogenic differentiation. In the IDG-SW3 osteocyte line with lncRNA953Rik overexpression, the expression of Osterix and its downstream genes was reduced. RNA pull-down and subsequent LC-MS/MS analysis revealed that lncRNA953Rik bound the nuclear protein CCAR2. We demonstrated that CCAR2 promoted Wnt/ß-catenin signaling and that lncRNA953Rik inhibited this pathway. lncRNA953Rik sequestered CCAR2 from HDAC1, leading to deacetylation of H3K27 in the Osterix promoter and consequent transcriptional downregulation of Osterix. This research is the first to clarify the role of a lncRNA in osteocytes. Our findings can pave the way for novel therapeutic options targeting lncRNAs in osteocytes to treat bone metabolic diseases such as osteoporosis.

Regulation of Osteoblast to Osteocyte Differentiation by Cyclin-Dependent Kinase-1.

Osteocytes have recently been identified as a new regulator of bone remodeling, but the detailed mechanism of their differentiation from osteoblasts remains unclear. The purpose of this study is to identify cell cycle regulators involved in the differentiation of osteoblasts into osteocytes and determine their physiological significance. The study uses IDG-SW3 cells as a model for the differentiation from osteoblasts to osteocytes. Among the major cyclin-dependent kinases (Cdks), Cdk1 is most abundantly expressed in IDG-SW3 cells, and its expression is down-regulated during differentiation into osteocytes. Inhibition of CDK1 activity reduces IDG-SW3 cell proliferation and differentiation into osteocytes. Osteocyte and Osteoblast-specific Cdk1 knockout in mice (Dmp1-Cdk1KO ) results in trabecular bone loss. Pthlh expression increases during differentiation, but inhibiting CDK1 activity reduces Pthlh expression. Parathyroid hormone-related protein concentration is reduced in the bone marrow of Dmp1-Cdk1KO mice. Four weeks of Parathyroid hormone administration partially recovers the trabecular bone loss in Dmp1-Cdk1KO mice. These results demonstrate that Cdk1 plays an essential role in the differentiation from osteoblast to osteocyte and the acquisition and maintenance of bone mass. The findings contribute to a better understanding of the mechanisms of bone mass regulation and can help develop efficient therapeutic strategies for osteoporosis treatment.

Interstitial-fluid shear stresses induced by vertically oscillating head motion lower blood pressure in hypertensive rats and humans.

The mechanisms by which physical exercise benefits brain functions are not fully understood. Here, we show that vertically oscillating head motions mimicking mechanical accelerations experienced during fast walking, light jogging or treadmill running at a moderate velocity reduce the blood pressure of rats and human adults with hypertension. In hypertensive rats, shear stresses of less than 1 Pa resulting from interstitial-fluid flow induced by such passive head motions reduced the expression of the angiotensin II type-1 receptor in astrocytes in the rostral ventrolateral medulla, and the resulting antihypertensive effects were abrogated by hydrogel introduction that inhibited interstitial-fluid movement in the medulla. Our findings suggest that oscillatory mechanical interventions could be used to elicit antihypertensive effects.

SIRT6-PAI-1 axis is a promising therapeutic target in aging-related bone metabolic disruption.

The mechanistic regulation of bone mass in aged animals is poorly understood. In this study, we examined the role of SIRT6, a longevity-associated factor, in osteocytes, using mice lacking Sirt6 in Dmp-1-expressing cells (cKO mice) and the MLO-Y4 osteocyte-like cell line. cKO mice exhibited increased osteocytic expression of Sost, Fgf23 and senescence inducing gene Pai-1 and the senescence markers p16 and Il-6, decreased serum phosphate levels, and low-turnover osteopenia. The cKO phenotype was reversed in mice that were a cross of PAI-1-null mice with cKO mice. Furthermore, senescence induction in MLO-Y4 cells increased the Fgf23 and Sost mRNA expression. Sirt6 knockout and senescence induction increased HIF-1α binding to the Fgf23 enhancer sequence. Bone mass and serum phosphate levels were higher in PAI-1-null aged mice than in wild-type mice. Therefore, SIRT6 agonists or PAI-1 inhibitors may be promising therapeutic options for aging-related bone metabolism disruptions.

l-Type amino acid transporter 1 in hypothalamic neurons in mice maintains energy and bone homeostasis.

Hypothalamic neurons regulate body homeostasis by sensing and integrating changes in the levels of key hormones and primary nutrients (amino acids, glucose, and lipids). However, the molecular mechanisms that enable hypothalamic neurons to detect primary nutrients remain elusive. Here, we identified l-type amino acid transporter 1 (LAT1) in hypothalamic leptin receptor-expressing (LepR-expressing) neurons as being important for systemic energy and bone homeostasis. We observed LAT1-dependent amino acid uptake in the hypothalamus, which was compromised in a mouse model of obesity and diabetes. Mice lacking LAT1 (encoded by solute carrier transporter 7a5, Slc7a5) in LepR-expressing neurons exhibited obesity-related phenotypes and higher bone mass. Slc7a5 deficiency caused sympathetic dysfunction and leptin insensitivity in LepR-expressing neurons before obesity onset. Importantly, restoring Slc7a5 expression selectively in LepR-expressing ventromedial hypothalamus neurons rescued energy and bone homeostasis in mice deficient for Slc7a5 in LepR-expressing cells. Mechanistic target of rapamycin complex-1 (mTORC1) was found to be a crucial mediator of LAT1-dependent regulation of energy and bone homeostasis. These results suggest that the LAT1/mTORC1 axis in LepR-expressing neurons controls energy and bone homeostasis by fine-tuning sympathetic outflow, thus providing in vivo evidence of the implications of amino acid sensing by hypothalamic neurons in body homeostasis.

Three-dimensional visualization of neural networks inside bone by Osteo-DISCO protocol and alteration of bone remodeling by surgical nerve ablation.

Bone is one of the largest organ systems in humans and is considered to regulate whole-body homeostasis in cooperation with other organs. We have previously reported that a sympathetic or sensory nervous system inside bone regulates bone homeostasis. However, the detailed regulatory mechanism, including the distribution of nerves inside bone, remains unknown. Although a two-dimensional histological analysis has been widely used to evaluate the structure of nerves or blood vessels, the actual structure is more complex, suggesting that it should be evaluated three-dimensionally. Here, we established a novel bone tissue clearing technique (Osteo-DISCO) for murine bones which enabled us to visualize the detailed distribution of nerves or blood vessels inside bone. Interestingly, we found that there is a specific nerve entry site in each long bone and that surgical ablation of the specific nerve fibers entering bone tissue led to decreased bone formation and impaired bone regeneration. Furthermore, we revealed that the administration of calcitonin gene-related peptide (CGRP), which is primarily released from sensory nerves, suppressed the bone loss caused by surgical nerve ablation. An in vitro study also indicated that CGRP directly promotes osteoblast activity, suggesting that sensory nerves inside bone can regulate osteogenesis via the secretion of CGRP.

Conditional inactivation of the L-type amino acid transporter LAT1 in chondrocytes models idiopathic scoliosis in mice.

Scoliosis, usually diagnosed in childhood and early adolescence, is an abnormal lateral curvature of the spine. L-type amino acid transporter 1 (LAT1), encoded by solute carrier transporter 7a5 (Slc7a5), plays a crucial role in amino acid sensing and signaling in specific cell types. We previously demonstrated the pivotal role of LAT1 on bone homeostasis in mice, and the expression of LAT1/SLC7A5 in vertebral cartilage of pediatric scoliosis patients; however, its role in chondrocytes on spinal homeostasis and implications regarding the underlying mechanisms during the onset and progression of scoliosis, remain unknown. Here, we identified LAT1 in mouse chondrocytes as an important regulator of postnatal spinal homeostasis. Conditional inactivation of LAT1 in chondrocytes resulted in a postnatal-onset severe thoracic scoliosis at the early adolescent stage with normal embryonic spinal development. Histological analyses revealed that Slc7a5 deletion in chondrocytes led to general disorganization of chondrocytes in the vertebral growth plate, along with an increase in apoptosis and a decrease in cell proliferation. Furthermore, loss of Slc7a5 in chondrocytes activated the general amino acid control (GAAC) pathway but inactivated the mechanistic target of rapamycin complex 1 (mTORC1) pathway in the vertebrae. The spinal deformity in Slc7a5-deficient mice was corrected by genetic inactivation of the GAAC pathway, but not by genetic activation of the mTORC1 pathway. These findings suggest that the LAT1-GAAC pathway in chondrocytes plays a critical role in the maintenance of proper spinal homeostasis by modulating cell proliferation and survivability.

Transcriptional coregulator Ess2 controls survival of post-thymic CD4+ T cells through the Myc and IL-7 signaling pathways.

Ess2, also known as Dgcr14, is a transcriptional co-regulator of CD4+ T cells. Ess2 is located in a chromosomal region, the loss of which has been associated with 22q11.2 deletion syndrome (22q11DS), which causes heart defects, skeletal abnormalities, and immunodeficiency. However, the specific association of Ess2 with 22q11DS remains unclear. To elucidate the role of Ess2 in T-cell development, we generated Ess2 floxed (Ess2fl/fl) and CD4+ T cell-specific Ess2 KO (Ess2ΔCD4/ΔCD4) mice using the Cre/loxP system. Interestingly, Ess2ΔCD4/ΔCD4 mice exhibited reduced naïve T-cell numbers in the spleen, while the number of thymocytes (CD4-CD8-, CD4+CD8+, CD4+CD8-, and CD4-CD8+) in the thymus remained unchanged. Furthermore, Ess2ΔCD4/ΔCD4 mice had decreased NKT cells and increased γδT cells in the thymus and spleen. A genome-wide expression analysis using RNA-seq revealed that Ess2 deletion alters the expression of many genes in CD4 single-positive thymocytes, including genes related to the immune system and Myc target genes. In addition, Ess2 enhanced the transcriptional activity of c-Myc. Some genes identified as Ess2 targets in mice show expressional correlation with ESS2 in human immune cells. Moreover, Ess2ΔCD4/ΔCD4 naïve CD4+ T cells did not maintain survival in response to IL-7. Our results suggest that Ess2 plays a critical role in post-thymic T-cell survival through the Myc and IL-7 signaling pathways.

The role of CDK8 in mesenchymal stem cells in controlling osteoclastogenesis and bone homeostasis.

Bone marrow mesenchymal stem cells (MSCs) are critical regulators of postnatal bone homeostasis. Osteoporosis is characterized by bone volume and strength deterioration, partly due to MSC dysfunction. Cyclin-dependent kinase 8 (CDK8) belongs to the transcription-related CDK family. Here, CDK8 in MSCs was identified as important for bone homeostasis. CDK8 level was increased in aged MSCs along with the association with aging-related signals. Mouse genetic studies revealed that CDK8 in MSCs plays a crucial role in bone resorption and homeostasis. Mechanistically, CDK8 in MSCs extrinsically controls osteoclastogenesis through the signal transducer and transcription 1 (STAT1)-receptor activator of the nuclear factor κ Β ligand (RANKL) axis. Moreover, aged MSCs have high osteoclastogenesis-supporting activity, partly through a CDK8-dependent manner. Finally, pharmacological inhibition of CDK8 effectively repressed MSC-dependent osteoclastogenesis and prevented ovariectomy-induced osteoclastic activation and bone loss. These findings highlight that the CDK8-STAT1-RANKL axis in MSCs could play a crucial role in bone resorption and homeostasis.

Foxf2 represses bone formation via Wnt2b/ß-catenin signaling.

Differentiation of mesenchymal stem cells (MSCs) into osteoblasts is a critical process for proper skeletal development and acquisition/maintenance of bone mass. However, since this regulatory mechanism has not yet been fully elucidated, the treatment of severe osteoporosis and fractures is a challenge. Here, through a comprehensive analysis of gene expression during the differentiation of MSCs into osteoblasts, we show that the forkhead transcription factor Foxf2 is a crucial regulator of this process. Foxf2 expression transiently increased during MSC osteoblastic differentiation. Overexpression of Foxf2 in MSCs inhibited osteoblastic differentiation, and conversely, knockdown of Foxf2 expression promoted this process. Osteoprogenitor-specific Foxf2 knockout mice developed a high bone mass phenotype due to increased bone formation. RNA-seq analysis and molecular experiments revealed that Foxf2 regulation of bone formation is mediated by Wnt2b. Knockdown of Foxf2 in mouse femurs enhanced bone regeneration in vivo. FOXF2 expression was correlated with hip bone mineral density in postmenopausal women with low bone mass. Finally, inhibition of FOXF2 promoted osteoblastic differentiation of human MSCs. This study uncovers a critical role of Foxf2 in the differentiation of MSCs into osteoblasts and provides insight into the pathogenesis associated with bone-related diseases such as osteoporosis and nonunion after fracture.

Erk5 in Bone Marrow Mesenchymal Stem Cells Regulates Bone Homeostasis by Preventing Osteogenesis in Adulthood.

Extracellular signal-regulated kinase 5 (Erk5) belongs to the mitogen-activated protein kinase (MAPK) family. Previously, we demonstrated that Erk5 directly phosphorylates Smad-specific E3 ubiquitin protein ligase 2 (Smurf2) at Thr249 (Smurf2Thr249) to activate its E3 ubiquitin ligase activity. Although we have clarified the importance of Erk5 in embryonic mesenchymal stem cells (MSCs) on skeletogenesis, its role in adult bone marrow (BM)-MSCs on bone homeostasis remains unknown. Leptin receptor-positive (LepR+) BM-MSCs represent a major source of bone in adult bone marrow and are critical regulators of postnatal bone homeostasis. Here, we identified Erk5 in BM-MSCs as an important regulator of bone homeostasis in adulthood. Bone marrow tissue was progressively osteosclerotic in mice lacking Erk5 in LepR+ BM-MSCs with age, accompanied by increased bone formation and normal bone resorption in vivo. Erk5 deficiency increased the osteogenic differentiation of BM-MSCs along with a higher expression of Runx2 and Osterix, essential transcription factors for osteogenic differentiation, without affecting their stemness in vitro. Erk5 deficiency decreased Smurf2Thr249 phosphorylation and subsequently increased Smad1/5/8-dependent signaling in BM-MSCs. The genetic introduction of the Smurf2T249E mutant (a phosphomimetic mutant) suppressed the osteosclerotic phenotype in Erk5-deficient mice. These findings suggest that the Erk5-Smurf2Thr249 axis in BM-MSCs plays a critical role in the maintenance of proper bone homeostasis by preventing excessive osteogenesis in adult bone marrow.

Developing an iOS application that uses machine learning for the automated diagnosis of blepharoptosis.

PURPOSE: To assess the performance of artificial intelligence in the automated classification of images taken with a tablet device of patients with blepharoptosis and subjects with normal eyelid. METHODS: This is a prospective and observational study. A total of 1276 eyelid images (624 images from 347 blepharoptosis cases and 652 images from 367 normal controls) from 606 participants were analyzed. In order to obtain a sufficient number of images for analysis, 1 to 4 eyelid images were obtained from each participant. We developed a model by fully retraining the pre-trained MobileNetV2 convolutional neural network. Subsequently, we verified whether the automatic diagnosis of blepharoptosis was possible using the images. In addition, we visualized how the model captured the features of the test data with Score-CAM. k-fold cross-validation (k = 5) was adopted for splitting the training and validation. Sensitivity, specificity, and the area under the curve (AUC) of the receiver operating characteristic curve for detecting blepharoptosis were examined. RESULTS: We found the model had a sensitivity of 83.0% (95% confidence interval [CI], 79.8-85.9) and a specificity of 82.5% (95% CI, 79.4-85.4). The accuracy of the validation data was 82.8%, and the AUC was 0.900 (95% CI, 0.882-0.917). CONCLUSION: Artificial intelligence was able to classify with high accuracy images of blepharoptosis and normal eyelids taken using a tablet device. Thus, the diagnosis of blepharoptosis with a tablet device is possible at a high level of accuracy. TRIAL REGISTRATION: Date of registration: 2021-06-25. TRIAL REGISTRATION NUMBER: UMIN000044660. Registration site: https://upload.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000051004.

Mechanical load regulates bone growth via periosteal Osteocrin.

Mechanical stimuli including loading after birth promote bone growth. However, little is known about how mechanical force triggers biochemical signals to regulate bone growth. Here, we identified a periosteal-osteoblast-derived secretory peptide, Osteocrin (OSTN), as a mechanotransducer involved in load-induced long bone growth. OSTN produced by periosteal osteoblasts regulates growth plate growth by enhancing C-type natriuretic peptide (CNP)-dependent proliferation and maturation of chondrocytes, leading to elongation of long bones. Additionally, OSTN cooperates with CNP to regulate bone formation. CNP stimulates osteogenic differentiation of periosteal osteoprogenitors to induce bone formation. OSTN binds to natriuretic peptide receptor 3 (NPR3) in periosteal osteoprogenitors, thereby preventing NPR3-mediated clearance of CNP and consequently facilitating CNP-signal-mediated bone growth. Importantly, physiological loading induces Ostn expression in periosteal osteoblasts by suppressing Forkhead box protein O1 (FoxO1) transcription factor. Thus, this study reveals a crucial role of OSTN as a mechanotransducer converting mechanical loading to CNP-dependent bone formation.

Effects of long-term plate fixation with different fixation modes on the radial cortical bone in dogs.

The aim of this study was to examine the effect of long-term locking plate fixation on the cortical bone of the canine radius. Locking compression plates were fixed to the left and right radius in dogs (n = 3). The left radius was fixed with a locking head screw (Locking Plate group, LP). The locking compression plate was compressed periosteally in the right radius using a cortex screw (Compression Plate group, CP). Radial bones from dogs that were euthanized for other purposes were collected as an untreated control group (Control group). After euthanasia at 36 weeks following plate fixation, radial bones were evaluated for bone mineral density and underwent histological analysis. Bone metabolic markers were analyzed by quantitative polymerase chain reaction (qPCR). Statistical analyses were performed for comparisons between groups. The LP group showed no significant difference in bone mineral density after plate fixation, whereas the CP group showed significantly lower bone mineral density. Histological analysis indicated that the number of osteoclasts and rate of empty lacunae increased significantly in the CP group relative to the Control and LP groups. qPCR analysis indicated increased expression of inflammatory cytokines, such as tumor necrosis factor-alpha, interleukin-6, and tumor necrosis factor ligand superfamily member 11 in the CP group, whereas Runt-related transcription factor 2, an osteoblast marker, was similar in all groups. The expression of hypoxia-inducible factor-1α in the CP group was also increased relative to that in the Control and LP groups. Thus, locking plate fixation is a biologically superior fixation method that does not cause implant-induced osteoporosis in the bone in the long term.

A novel PAI-1 inhibitor prevents ageing-related muscle fiber atrophy.

Sarcopenia is among the most common medical problems of the aging population worldwide and a major social concern. Here, we explored the therapeutic potential of TM5484, a novel orally available PAI-1 inhibitor, to prevent sarcopenia. The sarcopenic phenotypes of the calf muscle of 12- and 6-month-old middle-aged mice were compared. Although significant decline of isometric gastrocnemius muscle force was detected in the older untreated mice, those administered TM5484 had significantly greater calf muscle force, as determined using isometric measurements by electrical stimulation. Histological analysis indicated that cross-sectional gastrocnemius muscle fibers in untreated older mice were thinner than those in younger mice; however, TM5484-treated group showed thicker fibers than younger mice. Treatment with TM5484 for 6 months enhanced Igf1, Atrogin-1, Mt-Co1, and Chrna1 mRNA expression in the mice gastrocnemius muscle, with increased serum IGF-1 concentration. TM5484 induced dose-dependent Igf1, Atrogin-1, and Chrna1 expression in C2C12 myoblastic cells, confirming cell autonomous effect. Further, the presence of plasmin for 72 h caused significantly increased Igf1 expression in C2C12 cells. These findings suggest that oral PAI-1 inhibitors represent a promising therapeutic candidate for preventing sarcopenia progression in humans.

Cyclin-Dependent Kinase 1 Is Essential for Muscle Regeneration and Overload Muscle Fiber Hypertrophy.

Satellite cell proliferation is an essential step in proper skeletal muscle development and muscle regeneration. However, the mechanisms regulating satellite cell proliferation are relatively unknown compared to the knowledge associated with the differentiation of satellite cells. Moreover, it is still unclear whether overload muscle fiber hypertrophy is dependent on satellite cell proliferation. In general, cell proliferation is regulated by the activity of cell cycle regulators, such as cyclins and cyclin-dependent kinases (CDKs). Despite recent reports on the function of CDKs and CDK inhibitors in satellite cells, the physiological role of Cdk1 in satellite cell proliferation remains unknown. Herein, we demonstrate that Cdk1 regulates satellite cell proliferation, muscle regeneration, and muscle fiber hypertrophy. Cdk1 is highly expressed in myoblasts and is downregulated upon myoblast differentiation. Inhibition of CDK1 activity inhibits myoblast proliferation. Deletion of Cdk1 in satellite cells leads to inhibition of muscle recovery after muscle injury due to reduced satellite cell proliferation in vivo. Finally, we provide direct evidence that Cdk1 expression in satellite cells is essential for overload muscle fiber hypertrophy in vivo. Collectively, our results demonstrate that Cdk1 is essential for myoblast proliferation, muscle regeneration, and muscle fiber hypertrophy. These findings could help to develop treatments for refractory muscle injuries and muscle atrophy, such as sarcopenia.

The long noncoding RNA Crnde regulates osteoblast proliferation through the Wnt/ß-catenin signaling pathway in mice.

In the past decade, a growing importance has been placed on understanding the significance of long noncoding RNAs (lncRNAs) in regulating development, metabolism, and homeostasis. Osteoblast proliferation and differentiation are essential elements in skeletal development, bone metabolism, and homeostasis. However, the underlying mechanisms of lncRNAs in the process of osteoblast proliferation and differentiation remain largely unknown. Through comprehensive analysis of lncRNAs during bone formation, we show that colorectal neoplasia differentially expressed (Crnde), previously viewed as a cancer-related lncRNA, is an important regulator of osteoblast proliferation and differentiation. Crnde was found to be expressed in osteoblasts, and its expression was induced by parathyroid hormone. Furthermore, Crnde knockout mice developed a low bone mass phenotype due to impaired osteoblast proliferation and differentiation. Overexpression of Crnde in osteoblasts promoted their proliferation, and conversely, reduced Crnde expression inhibited osteoblast proliferation. Although ablation of Crnde inhibited osteoblast differentiation, overexpression of Crnde restored it. Finally, we provided evidence that Crnde modulates bone formation through Wnt/ß-catenin signaling. Therefore, our data suggest that Crnde is a novel regulator of bone metabolism.

The L-type amino acid transporter LAT1 inhibits osteoclastogenesis and maintains bone homeostasis through the mTORC1 pathway.

L-type amino acid transporter 1 (LAT1), which is encoded by solute carrier transporter 7a5 (Slc7a5), plays a crucial role in amino acid sensing and signaling in specific cell types, contributing to the pathogenesis of cancer and neurological disorders. Amino acid substrates of LAT1 have a beneficial effect on bone health directly and indirectly, suggesting a potential role for LAT1 in bone homeostasis. Here, we identified LAT1 in osteoclasts as important for bone homeostasis. Slc7a5 expression was substantially reduced in osteoclasts in a mouse model of ovariectomy-induced osteoporosis. The osteoclast-specific deletion of Slc7a5 in mice led to osteoclast activation and bone loss in vivo, and Slc7a5 deficiency increased osteoclastogenesis in vitro. Loss of Slc7a5 impaired activation of the mechanistic target of rapamycin complex 1 (mTORC1) pathway in osteoclasts, whereas genetic activation of mTORC1 corrected the enhanced osteoclastogenesis and bone loss in Slc7a5-deficient mice. Last, Slc7a5 deficiency increased the expression of nuclear factor of activated T cells, cytoplasmic 1 (Nfatc1) and the nuclear accumulation of NFATc1, a master regulator of osteoclast function, possibly through the canonical nuclear factor κB pathway and the Akt-glycogen synthase kinase 3ß signaling axis, respectively. These findings suggest that the LAT1-mTORC1 axis plays a pivotal role in bone resorption and bone homeostasis by modulating NFATc1 in osteoclasts, thereby providing a molecular connection between amino acid intake and skeletal integrity.

Pericytes as a Source of Osteogenic Cells in Bone Fracture Healing.

Pericytes are mesenchymal cells that surround the endothelial cells of small vessels in various organs. These cells express several markers, such as NG2, CD146, and PDGFRß, and play an important role in the stabilization and maturation of blood vessels. It was also recently revealed that like mesenchymal stem cells (MSCs), pericytes possess multilineage differentiation capacity, especially myogenic, adipogenic, and fibrogenic differentiation capacities. Although some previous studies have reported that pericytes also have osteogenic potential, the osteogenesis of pericytes can still be further elucidated. In the present study, we established novel methods for isolating and culturing primary murine pericytes. An immortalized pericyte line was also established. Multilineage induction of the pericyte line induced osteogenesis, adipogenesis, and chondrogenesis of the cells in vitro. In addition, pericytes that were injected into the fracture site of a bone fracture mouse model contributed to callus formation. Furthermore, in vivo pericyte-lineage-tracing studies demonstrated that endogenous pericytes also differentiate into osteoblasts and osteocytes and contribute to bone fracture healing as a cellular source of osteogenic cells. Pericytes can be a promising therapeutic candidate for treating bone fractures with a delayed union or nonunion as well as bone diseases causing bone defects.