ATP6AP2, a regulator of LRP6/ß-catenin protein trafficking, promotes Wnt/ß-catenin signaling and bone formation in a cell type dependent manner.

Wnt/ß-catenin signaling is critical for various cellular processes in multiple cell types, including osteoblast (OB) differentiation and function. Exactly how Wnt/ß-catenin signaling is regulated in OBs remain elusive. ATP6AP2, an accessory subunit of V-ATPase, plays important roles in multiple cell types/organs and multiple signaling pathways. However, little is known whether and how ATP6AP2 in OBs regulates Wnt/ß-catenin signaling and bone formation. Here we provide evidence for ATP6AP2 in the OB-lineage cells to promote OB-mediated bone formation and bone homeostasis selectively in the trabecular bone regions. Conditionally knocking out (CKO) ATP6AP2 in the OB-lineage cells (Atp6ap2Ocn-Cre) reduced trabecular, but not cortical, bone formation and bone mass. Proteomic and cellular biochemical studies revealed that LRP6 and N-cadherin were reduced in ATP6AP2-KO BMSCs and OBs, but not osteocytes. Additional in vitro and in vivo studies revealed impaired ß-catenin signaling in ATP6AP2-KO BMSCs and OBs, but not osteocytes, under both basal and Wnt stimulated conditions, although LRP5 was decreased in ATP6AP2-KO osteocytes, but not BMSCs. Further cell biological studies uncovered that osteoblastic ATP6AP2 is not required for Wnt3a suppression of ß-catenin phosphorylation, but necessary for LRP6/ß-catenin and N-cadherin/ß-catenin protein complex distribution at the cell membrane, thus preventing their degradation. Expression of active ß-catenin diminished the OB differentiation deficit in ATP6AP2-KO BMSCs. Taken together, these results support the view for ATP6AP2 as a critical regulator of both LRP6 and N-cadherin protein trafficking and stability, and thus regulating ß-catenin levels, demonstrating an un-recognized function of osteoblastic ATP6AP2 in promoting Wnt/LRP6/ß-catenin signaling and trabecular bone formation.

Constraints-related microscopic fatigue crack propagation behaviour of polycrystalline alloys.

Based on the microscopic polycrystalline fatigue crack propagation (MPFCP) model, the MPFCP behaviours of GH4169 alloy under different micro-notch depths and lengths (constraints) were studied from aspects of MPFCP path, MPFCP rate and stress distribution. The influences of the initial crack angle on MPFCP behaviour were further explored. It was observed that the grain boundary, the grain size and the stress state were different during crack propagation under different constraints, resulting in different MPFCP paths. The MPFCP path was straighter under high constraints, and the MPFCP rate was related to the micro-notch size and the loading direction. The crack tip needed more stress accumulation at low constraints than under high constraints to ensure smooth MPFCP behaviour. The influence of the initial crack angle on the MPFCP path was mainly reflected in the grain interior where the initial crack was located. The initial crack angle had a greater influence on the MPFCP rate than on the MPFCP path.

Inhibition of UFM1 expression suppresses cancer progression and is linked to the dismal prognosis and immune infiltration in oral squamous cell carcinoma.

BACKGROUND: Ubiquitin fold modifier 1 (UFM1) overexpression is associated with cancer cell proliferation, migration and invasion. However, the roles and pathways of UFM1 in oral squamous cell carcinoma (OSCC) has remained undefined. METHODS: The expression of UFM1 and the relationship between UFM1 expression and prognosis were investigated using data of OSCC patients from The Cancer Genome Atlas (TCGA) database. The UFM1 co-expressed genes, and the association between the UFM1 expression and immune cells and ubiquitination were explored. The effects of UFM1 expression on the growth and migration of OSCC cells were investigated by siRNA interference, Cell Counting Kit-8 (CCK-8), Transwell, Western blotting, and wound healing experiments. RESULTS: UFM1 was highly expressed in OSCC. UFM1 overexpression was associated with short overall survival, disease-specific survival, and progression-free interval, and was an adverse factor for prognosis in OSCC. UFM1-related nomograms were significantly associated with poor prognosis in OSCC patients. Decreased UFM1 expression could inhibit the proliferation, migration, and invasion of OSCC cells. UFM1 was associated with the immune cells (such as the Th17 cells, T helper cells, and cytotoxic cells) and ubiquitination. CONCLUSION: Elevated UFM1 expression was associated with poor prognosis, ubiquitination and immune infiltration in OSCC, and inhibition of UFM1 expression delayed OSCC progression, showing that UFM1 could be a biomarker for prognosis and treating OSCC patients.

Distributed optical fiber biosensor based on optical frequency domain reflectometry.

In situ acquisition of spatial distribution of biochemical substances is important in cell analysis, cancer detection and other fields. Optical fiber biosensors can achieve label-free, fast and accurate measurements. However, current optical fiber biosensors only acquire single-point of biochemical substance content. In this paper, we present a distributed optical fiber biosensor based on tapered fiber in optical frequency domain reflectometry (OFDR) for the first time. To enhance evanescent field at a relative long sensing range, we fabricate a tapered fiber with a taper waist diameter of 6 µm and a total stretching length of 140 mm. Then the human IgG layer is coated on the entire tapered region by polydopamine (PDA) -assisted immobilization as the sensing element to achieve to sense anti-human IgG. We measure shifts of the local Rayleigh backscattering spectra (RBS) caused by the refractive index (RI) change of an external medium surrounding a tapered fiber after immunoaffinity interactions by using OFDR. The measurable concentration of anti-human IgG and RBS shift has an excellent linearity in a range from 0 ng/ml to 14 ng/ml with an effective sensing range of 50 mm. The concentration measurement limit of the proposed distributed biosensor is 2 ng/ml for anti-human IgG. Distributed biosensing based on OFDR can locate a concentration change of anti-human IgG with an ultra-high sensing spatial resolution of 680 µm. The proposed sensor has a potential to realize a micron-level localization of biochemical substances such as cancer cells, which will open a door to transform single-point biosensor to distributed biosensor.

Reconstruction error model of distributed shape sensing based on the reentered frame in OFDR.

At present, the reconstruction error of optical fiber shape sensing is commonly represented by Euclidean distance error. However, the Euclidian error of shape reconstruction will be dependent on the shape complexity, which depends on length, curvature and torsion. In this paper, we establish a reconstruction error model of distributed shape sensing in optical frequency domain reflectometry (OFDR) based on the Frenet-Serret frame and the error delivering theory, which illustrates the relationship between the reconstruction error and parameters such as curvature, torsion, fiber length and strain measurement error. We experimentally verify the feasibility and applicability of the proposed reconstruction error model by distributed optical fiber shape sensing system based on OFDR. The proposed reconstruction error model can provide a prediction of the maximal reconstruction error when the estimated range of curvature, torsion, fiber length of a shape needs to be reconstructed and strain measurement errors of OFDR system are known. It is very useful to judge whether the shape reconstruction error meets the requirement according to the shape to be reconstructed.

Muscular Swedish mutant APP-to-Brain axis in the development of Alzheimer's disease.

Alzheimer's disease (AD) is the most common form of dementia. Notably, patients with AD often suffer from severe sarcopenia. However, their direct link and relationship remain poorly understood. Here, we generated a mouse line, TgAPPsweHSA, by crossing LSL (LoxP-STOP-LoxP)-APPswe with HSA-Cre mice, which express APPswe (Swedish mutant APP) selectively in skeletal muscles. Examining phenotypes in TgAPPsweHSA mice showed not only sarcopenia-like deficit, but also AD-relevant hippocampal inflammation, impairments in adult hippocampal neurogenesis and blood brain barrier (BBB), and depression-like behaviors. Further studies suggest that APPswe expression in skeletal muscles induces senescence and expressions of senescence-associated secretory phenotypes (SASPs), which include inflammatory cytokines and chemokines; but decreases growth factors, such as PDGF-BB and BDNF. These changes likely contribute to the systemic and hippocampal inflammation, deficits in neurogenesis and BBB, and depression-like behaviors, revealing a link of sarcopenia with AD, and uncovering an axis of muscular APPswe to brain in AD development.

A comparative study of audiology and cone beam computed tomography in TMD patients with otological symptoms through occlusal splint therapy.

INTRODUCTION: To investigate the changes of audiological tests and the cone beam computed tomography (CBCT) measurements of temporomandibular joint (TMJ) and middle-inner ear structure after occlusal splint therapy in temporomandibular disorders (TMD) patients with otological symptoms, and explore the etiological mechanism between TMD and otological symptoms. METHODS: The 25 subjects aged 18 to 40 years who diagnosed with TMD combined the otological symptoms enrolled in the study.They all had received orthodontic treatment in the outpatient clinic of the orthodontic department in Beijing Stomatological Hospital. All the subjects underwent the audiological tests of pure tone audiometry (PTA) and CBCT before and after the occlusal splint therapy. RESULTS: After the stabilization occlusal splint therapy, subjects with improvement or complete remission in TMD and otological symptoms accounted for 84% and 80% in all subjects respectively. There were statistically differences in the distances between condylar center (CoC) and sella (S) in sagittal and vertical directions before and after treatment, and statistically difference between ATM and S in sagittal direction. The threshold of PTA at 8000Hz were negatively correlated with the sagittal displacement of condyle and positively correlated with the coronal displacement of condyle. The thickness of top 1/3 of anterior wall of tympanum in sagittal were positively correlated with the threshold of PTA at 4000Hz. CONCLUSION: The changes in the TMJ position through occlusal splint therapy might cause the changes in structure of middle-inner ear, which might be one of the reasons for the improvement in otological symptoms. KEY WORDS: Audiology, CBCT, Otological symptoms, TMD.

Parkinson's in the bone.

Patients with Parkinson's disease (PD) exhibit systemic deficits, including arthritis and osteoporosis-like symptoms. However, the questions, how the deficits in periphery organs or tissues occur in PD patients, and what are the relationship (s) of the periphery tissue deficits with the brain pathology (e.g., dopamine neuron loss), are at the beginning stage to be investigated. Notice that both PD and osteoporosis are the products of a complex interaction of genetic and environmental risk factors. Genetic mutations in numerous genes have been identified in patients either with recessive or autosomal dominant PD. Most of these PD risk genes are ubiquitously expressed; and many of them are involved in regulation of bone metabolism. Here, we review the functions of the PD risk genes in regulating bone remodeling and homeostasis. The knowledge gaps in our understanding of the bone-to-brain axis in PD development are also outlined.

Osteoblastic Swedish mutant APP expedites brain deficits by inducing endoplasmic reticulum stress-driven senescence.

Patients with Alzheimer's disease (AD) often have osteoporosis or osteopenia. However, their direct link and relationship remain largely unclear. Previous studies have detected osteoporotic deficits in young adult Tg2576 and TgAPPsweOCN mice, which express APPswe (Swedish mutant) ubiquitously and selectively in osteoblast (OB)-lineage cells. This raises the question, whether osteoblastic APPswe contributes to AD development. Here, we provide evidence that TgAPPsweOCN mice also exhibit AD-relevant brain pathologies and behavior phenotypes. Some brain pathologies include age-dependent and regional-selective increases in glial activation and pro-inflammatory cytokines, which are accompanied by behavioral phenotypes such as anxiety, depression, and altered learning and memory. Further cellular studies suggest that APPswe, but not APPwt or APPlon (London mutant), in OB-lineage cells induces endoplasmic reticulum-stress driven senescence, driving systemic and cortex inflammation as well as behavioral changes in 6-month-old TgAPPsweOCN mice. These results therefore reveal an unrecognized function of osteoblastic APPswe to brain axis in AD development.

Hepcidin contributes to Swedish mutant APP-induced osteoclastogenesis and trabecular bone loss.

Patients with Alzheimer's disease (AD) often have lower bone mass than healthy individuals. However, the mechanisms underlying this change remain elusive. Previously, we found that Tg2576 mice, an AD animal model that ubiquitously expresses Swedish mutant amyloid precursor protein (APPswe), shows osteoporotic changes, reduced bone formation, and increased bone resorption. To understand how bone deficits develop in Tg2576 mice, we used a multiplex antibody array to screen for serum proteins that are altered in Tg2576 mice and identified hepcidin, a master regulator of iron homeostasis. We further investigated hepcidin's function in bone homeostasis and found that hepcidin levels were increased not only in the serum but also in the liver, muscle, and osteoblast (OB) lineage cells in Tg2576 mice at both the mRNA and protein levels. We then generated mice selectively expressing hepcidin in hepatocytes or OB lineage cells, which showed trabecular bone loss and increased osteoclast (OC)-mediated bone resorption. Further cell studies suggested that hepcidin increased OC precursor proliferation and differentiation by downregulating ferroportin (FPN) expression and increasing intracellular iron levels. In OB lineage cells, APPswe enhanced hepcidin expression by inducing ER stress and increasing OC formation, in part through hepcidin. Together, these results suggest that increased hepcidin expression in hepatocytes and OB lineage cells in Tg2576 mice contributes to enhanced osteoclastogenesis and trabecular bone loss, identifying the hepcidin-FPN-iron axis as a potential therapeutic target to prevent AD-associated bone loss.

Linking skeletal muscle aging with osteoporosis by lamin A/C deficiency.

The nuclear lamina protein lamin A/C is a key component of the nuclear envelope. Mutations in the lamin A/C gene (LMNA) are identified in patients with various types of laminopathy-containing diseases, which have features of accelerated aging and osteoporosis. However, the underlying mechanisms for laminopathy-associated osteoporosis remain largely unclear. Here, we provide evidence that loss of lamin A/C in skeletal muscles, but not osteoblast (OB)-lineage cells, results in not only muscle aging-like deficit but also trabecular bone loss, a feature of osteoporosis. The latter is due in large part to elevated bone resorption. Further cellular studies show an increase of osteoclast (OC) differentiation in cocultures of bone marrow macrophages/monocytes (BMMs) and OBs after treatment with the conditioned medium (CM) from lamin A/C-deficient muscle cells. Antibody array screening analysis of the CM proteins identifies interleukin (IL)-6, whose expression is markedly increased in lamin A/C-deficient muscles. Inhibition of IL-6 by its blocking antibody in BMM-OB cocultures diminishes the increase of osteoclastogenesis. Knockout (KO) of IL-6 in muscle lamin A/C-KO mice diminishes the deficits in trabecular bone mass but not muscle. Further mechanistic studies reveal an elevation of cellular senescence marked by senescence-associated beta-galactosidase (SA-ß-gal), p16Ink4a, and p53 in lamin A/C-deficient muscles and C2C12 muscle cells, and the p16Ink4a may induce senescence-associated secretory phenotype (SASP) and IL-6 expression. Taken together, these results suggest a critical role for skeletal muscle lamin A/C to prevent cellular senescence, IL-6 expression, hyperosteoclastogenesis, and trabecular bone loss, uncovering a pathological mechanism underlying the link between muscle aging/senescence and osteoporosis.

Lack of Myosin X Enhances Osteoclastogenesis and Increases Cell Surface Unc5b in Osteoclast-Lineage Cells.

Normal bone mass is maintained by balanced bone formation and resorption. Myosin X (Myo10), an unconventional "myosin tail homology 4-band 4.1, ezrin, radixin, moesin" (MyTH4-FERM) domain containing myosin, is implicated in regulating osteoclast (OC) adhesion, podosome positioning, and differentiation in vitro. However, evidence is lacking for Myo10 in vivo function. Here we show that mice with Myo10 loss of function, Myo10m/m , exhibit osteoporotic deficits, which are likely due to the increased OC genesis and bone resorption because bone formation is unchanged. Similar deficits are detected in OC-selective Myo10 conditional knockout (cko) mice, indicating a cell autonomous function of Myo10. Further mechanistic studies suggest that Unc-5 Netrin receptor B (Unc5b) protein levels, in particular its cell surface level, are higher in the mutant OCs, but lower in RAW264.7 cells or HEK293 cells expressing Myo10. Suppressing Unc5b expression in bone marrow macrophages (BMMs) from Myo10m/m mice by infection with lentivirus of Unc5b shRNA markedly impaired RANKL-induced OC genesis. Netrin-1, a ligand of Unc5b, increased RANKL-induced OC formation in BMMs from both wild-type and Myo10m/m mice. Taken together, these results suggest that Myo10 plays a negative role in OC formation, likely by inhibiting Unc5b cell-surface targeting, and suppressing Netrin-1 promoted OC genesis. © 2019 American Society for Bone and Mineral Research.

APP promotes osteoblast survival and bone formation by regulating mitochondrial function and preventing oxidative stress.

Amyloid precursor protein (APP) is ubiquitously expressed in various types of cells including bone cells. Mutations in App gene result in early-onset Alzheimer's disease (AD). However, little is known about its physiological function in bone homeostasis. Here, we provide evidence for APP's role in promoting bone formation. Mice that knocked out App gene (APP-/-) exhibit osteoporotic-like deficit, including reduced trabecular and cortical bone mass. Such a deficit is likely due in large to a decrease in osteoblast (OB)-mediated bone formation, as little change in bone resorption was detected in the mutant mice. Further mechanical studies of APP-/- OBs showed an impairment in mitochondrial function, accompanied with increased reactive oxygen species (ROS) and apoptosis. Intriguingly, these deficits, resemble to those in Tg2576 animal model of AD that expresses Swedish mutant APP (APPswe), were diminished by treatment with an anti-oxidant NAC (n-acetyl-l-cysteine), uncovering ROS as a critical underlying mechanism. Taken together, these results identify an unrecognized physiological function of APP in promoting OB survival and bone formation, implicate APPswe acting as a dominant negative factor, and reveal a potential clinical value of NAC in treatment of AD-associated osteoporotic deficits.

YAP promotes osteogenesis and suppresses adipogenic differentiation by regulating ß-catenin signaling.

YAP (yes-associated protein) is a transcriptional factor that is negatively regulated by Hippo pathway, a conserved pathway for the development and size control of multiple organs. The exact function of YAP in bone homeostasis remains controversial. Here we provide evidence for YAP's function in promoting osteogenesis, suppressing adipogenesis, and thus maintaining bone homeostasis. YAP is selectively expressed in osteoblast (OB)-lineage cells. Conditionally knocking out Yap in the OB lineage in mice reduces cell proliferation and OB differentiation and increases adipocyte formation, resulting in a trabecular bone loss. Mechanistically, YAP interacts with ß-catenin and is necessary for maintenance of nuclear ß-catenin level and Wnt/ß-catenin signaling. Expression of ß-catenin in YAP-deficient BMSCs (bone marrow stromal cells) diminishes the osteogenesis deficit. These results thus identify YAP-ß-catenin as an important pathway for osteogenesis during adult bone remodeling and uncover a mechanism underlying YAP regulation of bone homeostasis.

Osteoblastic Lrp4 promotes osteoclastogenesis by regulating ATP release and adenosine-A2AR signaling.

Bone homeostasis depends on the functional balance of osteoblasts (OBs) and osteoclasts (OCs). Lrp4 is a transmembrane protein that is mutated in patients with high bone mass. Loss of Lrp4 in OB-lineage cells increases bone mass by elevating bone formation by OBs and reducing bone resorption by OCs. However, it is unclear how Lrp4 deficiency in OBs impairs osteoclastogenesis. Here, we provide evidence that loss of Lrp4 in the OB lineage stabilizes the prorenin receptor (PRR) and increases PRR/V-ATPase-driven ATP release, thereby enhancing the production of the ATP derivative adenosine. Both pharmacological and genetic inhibition of adenosine-2A receptor (A2AR) in culture and Lrp4 mutant mice diminishes the osteoclastogenic deficit and reduces trabecular bone mass. Furthermore, elevated adenosine-A2AR signaling reduces receptor activator of nuclear factor κB (RANK)-mediated osteoclastogenesis. Collectively, these results identify a mechanism by which osteoblastic Lrp4 controls osteoclastogenesis, reveal a cross talk between A2AR and RANK signaling in osteoclastogenesis, and uncover an unrecognized pathophysiological mechanism of high-bone-mass disorders.