Double-network hydrogel enhanced by SS31-loaded mesoporous polydopamine nanoparticles: Symphonic collaboration of near-infrared photothermal antibacterial effect and mitochondrial maintenance for full-thickness wound healing in diabetes mellitus.

Diabetic wound healing has become a serious healthcare challenge. The high-glucose environment leads to persistent bacterial infection and mitochondrial dysfunction, resulting in chronic inflammation, abnormal vascular function, and tissue necrosis. To solve these issues, we developed a double-network hydrogel, constructed with pluronic F127 diacrylate (F127DA) and hyaluronic acid methacrylate (HAMA), and enhanced by SS31-loaded mesoporous polydopamine nanoparticles (MPDA NPs). As components, SS31, a mitochondria-targeted peptide, maintains mitochondrial function, reduces mitochondrial reactive oxygen species (ROS) and thus regulates macrophage polarization, as well as promoting cell proliferation and migration, while MPDA NPs not only scavenge ROS and exert an anti-bacterial effect by photothermal treatment under near-infrared light irradiation, but also control release of SS31 in response to ROS. This F127DA/HAMA-MPDA@SS31 (FH-M@S) hydrogel has characteristics of adhesion, superior biocompatibility and mechanical properties which can adapt to irregular wounds at different body sites and provide sustained release of MPDA@SS31 (M@S) NPs. In addition, in a diabetic rat full thickness skin defect model, the FH-M@S hydrogel promoted macrophage M2 polarization, collagen deposition, neovascularization and wound healing. Therefore, the FH-M@S hydrogel exhibits promising therapeutic potential for skin regeneration.

Identifying therapeutic biomarkers of zoledronic acid by metabolomics.

Zoledronic acid (ZOL) is a potent antiresorptive agent that increases bone mineral density (BMD) and reduces fracture risk in postmenopausal osteoporosis (PMOP). The anti-osteoporotic effect of ZOL is determined by annual BMD measurement. In most cases, bone turnover markers function as early indicators of therapeutic response, but they fail to reflect long-term effects. We used untargeted metabolomics to characterize time-dependent metabolic shifts in response to ZOL and to screen potential therapeutic markers. In addition, bone marrow RNA-seq was performed to support plasma metabolic profiling. Sixty rats were assigned to sham-operated group (SHAM, n = 21) and ovariectomy group (OVX, n = 39) and received sham operation or bilateral ovariectomy, respectively. After modeling and verification, rats in the OVX group were further divided into normal saline group (NS, n = 15) and ZOL group (ZA, n = 18). Three doses of 100 µg/kg ZOL were administrated to the ZA group every 2 weeks to simulate 3-year ZOL therapy in PMOP. An equal volume of saline was administered to the SHAM and NS groups. Plasma samples were collected at five time points for metabolic profiling. At the end of the study, selected rats were euthanatized for bone marrow RNA-seq. A total number of 163 compound were identified as differential metabolites between the ZA and NS groups, including mevalonate, a critical molecule in target pathway of ZOL. In addition, prolyl hydroxyproline (PHP), leucyl hydroxyproline (LHP), 4-vinylphenol sulfate (4-VPS) were identified as differential metabolites throughout the study. Moreover, 4-VPS negatively correlated with increased vertebral BMD after ZOL administration as time-series analysis revealed. Bone marrow RNA-seq showed that the PI3K-AKT signaling pathway was significantly associated with ZOL-mediated changes in expression (adjusted-p = 0.018). In conclusion, mevalonate, PHP, LHP, and 4-VPS are candidate therapeutic markers of ZOL. The pharmacological effect of ZOL likely occurs through inhibition of the PI3K-AKT signaling pathway.

Prrx1 marks stem cells for bone, white adipose tissue and dermis in adult mice.

Specialized connective tissues, including bone and adipose tissues, control various physiological activities, including mineral and energy homeostasis. However, the identity of stem cells maintaining these tissues throughout adulthood remains elusive. By conducting genetic lineage tracing and cell depletion experiments in newly generated knock-in Cre/CreERT2 lines, we show here that rare Prrx1-expressing cells act as stem cells for bone, white adipose tissue and dermis in adult mice, which are indispensable for the homeostasis and repair of these tissues. Single-cell profiling reveals the cycling and multipotent nature of Prrx1-expressing cells and the stemness of these cells is further validated by transplantation assays. Moreover, we identify the cell surface markers for Prrx1-expressing stem cells and show that the activities of these stem cells are regulated by Wnt signaling. These findings expand our knowledge of connective tissue homeostasis/regeneration and may help improve stem-cell-based therapies.

Clinical characteristics of 10 Chinese patients with melorheostosis and identification of a somatic MAP2K1 variant in one case.

BACKGROUND: Melorheostosis (MEL) is an exceptionally rare sclerosing bone dysplasia with asymmetrically exuberant bone formation and soft tissue lesions in a segmental distribution. We aimed to summarize the clinical characteristics of Chinese MEL patients and identify their pathogenic cause. METHODS: In total, 10 Chinese MEL patients were recruited, and clinical manifestations and radiological characteristics were recorded. Sanger sequencing of the LEMD3 gene was performed on peripheral blood samples of all patients, while the exome sequencing of matched peripheral blood, melorheostotic bone, and skin lesion samples was conducted on one patient who provided affected bone and skin tissues. Micro-computed tomography (micro-CT) was also used to scan the melorheostotic bone tissue. RESULTS: We found the average age of the 10 MEL patients was 29.5 years (range 11-40 years), and the major symptoms were bone pain, restricted movement, and bone deformity. The lesions sites were mainly located in femur (8/10), tibia (8/10), fibula (6/10), and foot (7/10), the next was pelvis (4/10), and the last were patella (1/10), hand (1/10) and spine (1/10). Radiological examinations showed a mixture of hyperostosis consisting of classic "dripping candle wax," "osteoma-like," or "myositis ossificans-like" patterns in most patients. No germline pathogenic variants in the LEMD3 gene were found in all patients, but a disease-causing somatic variant of MAP2K1 (c.167A > C, p.Gln56Pro) was detected in melorheostotic bone from one patient. Moreover, the micro-CT analysis showed increased porosity in the melorheostotic bone with somatic MAP2K1 variant. CONCLUSION: This is a summary of the clinical characteristics of Chinese MEL patients and we first identify the somatic MAP2K1 variant in Chinese patients. Our findings validate the molecular genetic mechanism of MEL and broaden its phenotype spectrum in the Chinese population.

Toll-like receptor 9 deficiency induces osteoclastic bone loss via gut microbiota-associated systemic chronic inflammation.

Toll-like receptors (TLRs) play pivotal roles in inflammation and provide important links between the immune and skeletal systems. Although the activation of TLRs may affect osteoclast differentiation and bone metabolism, whether and how TLRs are required for normal bone remodeling remains to be fully explored. In the current study, we show for the first time that TLR9-/- mice exhibit a low bone mass and low-grade systemic chronic inflammation, which is characterized by the expansion of CD4+ T cells and increased levels of inflammatory cytokines, including TNFα, RANKL, and IL1ß. The increased levels of these cytokines significantly promote osteoclastogenesis and induce bone loss. Importantly, TLR9 deletion alters the gut microbiota, and this dysbiosis is the basis of the systemic inflammation and bone loss observed in TLR9-/- mice. Furthermore, through single-cell RNA sequencing, we identified myeloid-biased hematopoiesis in the bone marrow of TLR9-/- mice and determined that the increase in myelopoiesis, likely caused by the adaptation of hematopoietic stem cells to systemic inflammation, also contributes to inflammation-induced osteoclastogenesis and subsequent bone loss in TLR9-/- mice. Thus, our study provides novel evidence that TLR9 signaling connects the gut microbiota, immune system, and bone and is critical in maintaining the homeostasis of inflammation, hematopoiesis, and bone metabolism under normal conditions.

Polymeric coating on ß-TCP scaffolds provides immobilization of small extracellular vesicles with surface-functionalization and ZEB1-Loading for bone defect repair in diabetes mellitus.

Repair of critical-size bone defects in patients with diabetes mellitus (DM) has always been a challenge in clinical treatment. The process of bone defect regeneration can be impaired by underlying diseases including DM, but the mechanism remains unclear. In bone tissue engineering, the integration of bionic coatings and bioactive components into basic scaffolds are common function-enhancing strategies. Small extracellular vesicles (sEVs) have been applied for cell-free tissue regeneration in the last few years. We previously reported that sEVs have flexible and easily-extensible potential, through modular design and engineering modification. The impairment of CD31hiendomucinhi endothelial cells (ECs) whose function is coupling of osteogenesis and angiogenesis, is considered an important contributor to diabetic bone osteopathy, and ZEB1, which is highly expressed in CD31hiendomucinhi ECs, promotes angiogenesis-dependent bone formation. Thus we believe these ECs hold much promise for use in bone regeneration. In addition, c(RGDfC) has been reported to be a highly-effective peptide targeting αvß3, which is highly expressed in the bone microenvironment. In this study, we developed a hyaluronic acid (HA)/poly-L-lysine (PLL) layer-by-layer (LbL) self-assembly coating on ß-TCP (ß-tricalcium phosphate) scaffolds providing immobilization of modularized engineered sEVs (with c(RGDfC) surface functionalization and ZEB1 loading) to facilitate bone defect regeneration under DM conditions. RNA-seq was used to explore possible molecular mechanisms, and the therapeutic effects of bone regeneration were systematically evaluated in vitro and in vivo. Our data demonstrated that this strategy could be very effective in promoting the repair of diabetic bone defects, by enhancing angiogenesis, promoting osteogenesis and inhibiting osteoclast formation.

Genetics Evaluation of Targeted Exome Sequencing in 223 Chinese Probands With Genetic Skeletal Dysplasias.

Genetic skeletal dysplasias (GSDs) are a type of disease with complex phenotype and high heterogeneity, characterized by cartilage and bone growth abnormalities. The variable phenotypes of GSD make clinical diagnosis difficult. To explore the clinical utility of targeted exome sequencing (TES) in the diagnosis of GSD, 223 probands with suspected GSD were enrolled for TES with a panel of 322 known disease-causing genes. After bioinformatics analysis, all candidate variants were prioritized by pathogenicity. Sanger sequencing was used to verify candidate variants in the probands and parents and to trace the source of variants in family members. We identified the molecular diagnoses for 110/223 probands from 24 skeletal disorder groups and confirmed 129 pathogenic/likely pathogenic variants in 48 genes. The overall diagnostic rate was 49%. The molecular diagnostic results modified the diagnosis in 25% of the probands, among which mucopolysaccharidosis and spondylo-epi-metaphyseal dysplasias were more likely to be misdiagnosed. The clinical management of 33% of the probands also improved; 21 families received genetic counseling; 4 families accepted prenatal genetic diagnosis, 1 of which was detected to carry pathogenic variants. The results showed that TES achieved a high diagnostic rate for GSD, helping clinicians confirm patients' molecular diagnoses, formulate treatment directions, and carry out genetic counseling. TES could be an economical diagnostic method for patients with GSD.

Small extracellular vesicles in combination with sleep-related circRNA3503: A targeted therapeutic agent with injectable thermosensitive hydrogel to prevent osteoarthritis.

Osteoarthritis (OA), characterized by chondrocyte apoptosis and disturbance of the balance between catabolism and anabolism of the extracellular matrix (ECM), is the most common age-related degenerative joint disease worldwide. As sleep has been found to be beneficial for cartilage repair, and circular RNAs (circRNAs) have been demonstrated to be involved in the pathogenesis of OA, we performed RNA sequencing (RNA-seq), and found circRNA3503 was significantly increased after melatonin (MT)-induced cell sleep. Upregulation of circRNA3503 expression completely rescued the effects of interleukin-1ß (IL-1ß), which was used to simulate OA, on apoptosis, ECM degradation- and synthesis-related genes. Mechanistically, circRNA3503 acted as a sponge of hsa-miR-181c-3p and hsa-let-7b-3p. Moreover, as we previously showed that small extracellular vesicles (sEVs) derived from synovium mesenchymal stem cells (SMSCs) can not only successfully deliver nucleic acids to chondrocytes, but also effectively promote chondrocyte proliferation and migration, we assessed the feasibility of sEVs in combination with sleep-related circRNA3503 as an OA therapy. We successfully produced and isolated circRNA3503-loaded sEVs (circRNA3503-OE-sEVs) from SMSCs. Then, poly(D,l-lactide)-b-poly(ethylene glycol)-b-poly(D,l-lactide) (PDLLA-PEG-PDLLA, PLEL) triblock copolymer gels were used as carriers of sEVs. Through in vivo and in vitro experiments, PLEL@circRNA3503-OE-sEVs were shown to be a highly-effective therapeutic strategy to prevent OA progression. Through multiple pathways, circRNA3503-OE-sEVs alleviated inflammation-induced apoptosis and the imbalance between ECM synthesis and ECM degradation by acting as a sponge of hsa-miR-181c-3p and hsa-let-7b-3p. In addition, circRNA3503-OE-sEVs promoted chondrocyte renewal to alleviate the progressive loss of chondrocytes. Our results highlight the potential of PLEL@circRNA3503-OE-sEVs for preventing OA progression.

Mutations in Profilin 1 Cause Early-Onset Paget's Disease of Bone With Giant Cell Tumors.

Paget's disease of bone (PDB) is a late-onset chronic progressive bone disease characterized by abnormal activation of osteoclasts that results in bone pain, deformities, and fractures. PDB is very rare in Asia. A subset of PDB patients have early onset and can develop malignant giant cell tumors (GCTs) of the bone (PDB/GCTs), which arise within Paget bone lesions; the result is a significantly higher mortality rate. SQSTM1, TNFRSF11A, OPG, VCP, and HNRNPA2B1 have been identified as pathogenic genes of PDB, and ZNF687 is the only confirmed gene to date known to cause PDB/GCT. However, the molecular mechanism underlying PDB/GCT has not been fully elucidated. Here, we investigate an extended Chinese pedigree with eight individuals affected by early-onset and polyostotic PDB, two of whom developed GCTs. We identified a heterozygous 4-bp deletion in the Profilin 1 (PFN1) gene (c.318_321delTGAC) by genetic linkage analysis and exome sequencing for the family. Sanger sequencing revealed another heterozygous 1-bp deletion in PFN1 (c.324_324delG) in a sporadic early-onset PDB/GCT patient, further proving its causative role. Interestingly, a heterozygous missense mutation of PFN1 (c.335 T > C) was identified in another PDB/GCT family, revealing that not only deletion but also missense mutations in PFN1 can cause PDB/GCT. Furthermore, we established a Pfn1-mutated mouse model (C57BL/6J mice) and successfully obtained Pagetic phenotypes in heterozygous mice, verifying loss of function of PFN1 as the cause of PDB/GCT development. In conclusion, our findings reveal mutations in PFN1 as the pathological mechanism in PDB/GCT, and we successfully established Pfn1-mutated mice as a suitable animal model for studying PDB-associated pathological mechanisms. The identification of PFN1 mutations has great diagnostic value for identifying PDB individuals predisposed toward developing GCTs. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Mesoporous Bioactive Glass Scaffold Delivers Salvianolic Acid B to Promote Bone Regeneration in a Rat Cranial Defect Model.

BACKGROUND: Mesoporous Bioactive Glass (MBG) has been widely studied because of its excellent histocompatibility and degradability. However, due to the lack of good osteoinductive activity, the pure MBG scaffold is not effective in repairing large-scale bone defects. OBJECTIVE: To observe the repair effect of MBG scaffolds delivering Salvianolic acid B (SB) on critical bone defects in rats. METHODS: In this study, MBG scaffolds were used as delivery vehicle. SB, a small molecular active drug with good osteogenic differentiation ability, was loaded into the MBG scaffolds at low, medium and high doses. The effect of SB released from the MBG scaffolds on osteogenic differentiation of rat Bone Marrow Mesenchymal Stem Cells (rBMSCs) was investigated using alkaline phosphatase staining, alizarin red staining and real-time quantitative polymerase chain reaction. Moreover, 8 weeks after implantation of the scaffolds, the bone regeneration was evaluated by micro- CT, sequential fluorescence labeling and histological staining analysis. RESULTS: The in vitro results showed that different doses of SB had similar release rate from scaffolds and could be released from scaffolds continuously. The middle dose (MBG/MSB) and high dose (MBG/HSB) groups significantly promoted the osteogenic differentiation of rBMSCs when compared with a low dose (MBG/LSB) group. Moreover, SB produced significant increases in newly formed bone of calvarial bone defects in rats. CONCLUSION: It is concluded that the use of MBG scffolds delivering SB is an effective strategy for the treatment of bone defects.

EWSAT1 Acts in Concert with Exosomes in Osteosarcoma Progression and Tumor-Induced Angiogenesis: The "Double Stacking Effect".

The prognosis for osteosarcoma (OS) continues to be unsatisfactory due to tumor recurrence as a result of metastasis and drug resistance. Several studies have shown that Ewing sarcoma associated transcript 1 (EWSAT1) plays an important role in the progression of OS. Exosomes (Exos) act as important carriers in intercellular communication and play an important role in the tumor microenvironment, especially in tumor-induced angiogenesis. Nonetheless, the specific mechanism via which EWSAT1 and Exos regulate OS progression is unknown, and whether they can be effective therapeutic targets also requires verification. Hence, in this study, it is aimed to investigate the mechanisms of action of EWSAT1 and Exos. EWSAT1 significantly promotes proliferation, migration, colony formation, and survival of OS. EWSAT1 regulates OS-induced angiogenesis via two mechanisms, called the "double stacking effect," which is a combination of the increase in sensitivity/reactivity of vascular endothelial cells triggered by Exos-carrying EWSAT1, and the EWSAT1-induced increase in angiogenic factor secretion. In vivo experiments further validates the "double stacking effect" and shows that EWSAT1-KD effectively inhibits tumor growth in OS. The above observations indicate that EWSAT1 can be used as not only a potential diagnostic and prognostic marker, but also as a precise therapeutic target for OS.

Atorvastatin promotes bone formation in aged apoE-/- mice through the Sirt1-Runx2 axis.

BACKGROUND: Statins are the most widely used drugs in elderly patients; the most common clinical application of statins is in aged hyperlipemia patients. There are few studies on the effects and mechanisms of statins on bone in elderly mice with hyperlipemia. The study is to examine the effects of atorvastatin on bone phenotypes and metabolism in aged apolipoprotein E-deficient (apoE-/-) mice, and the possible mechanisms involved in these changes. METHODS: Twenty-four 60-week-old apoE-/- mice were randomly allocated to two groups. Twelve mice were orally gavaged with atorvastatin (10 mg/kg body weight/day) for 12 weeks; the others served as the control group. Bone mass and skeletal microarchitecture were determined using micro-CT. Bone metabolism was assessed by serum analyses, qRT-PCR, and Western blot. Bone marrow-derived mesenchymal stem cells (BMSCs) from apoE-/- mice were differentiated into osteoblasts and treated with atorvastatin and silent information regulator 1 (Sirt1) inhibitor EX-527. RESULTS: The results showed that long-term administration of atorvastatin increases bone mass and improves bone microarchitecture in trabecular bone but not in cortical bone. Furthermore, the serum bone formation marker osteocalcin (OCN) was ameliorated by atorvastatin, whereas the bone resorption marker tartrate-resistant acid phosphatase 5b (Trap5b) did not appear obviously changes after the treatment of atorvastatin. The mRNA expression of Sirt1, runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), and OCN in bone tissue were increased after atorvastatin administration. Western blot showed same trend in Sirt1 and Runx2. The in vitro study showed that when BMSCs from apoE-/- mice were pretreated with EX527, the higher expression of Runx2, ALP, and OCN activated by atorvastatin decreased significantly or showed no difference compared with the control. The protein expression of Runx2 showed same trend. CONCLUSIONS: Accordingly, the current study validates the hypothesis that atorvastatin can increase bone mass and promote osteogenesis in aged apoE-/- mice by regulating the Sirt1-Runx2 axis.

Parathyroid Hormone Derivative with Reduced Osteoclastic Activity Promoted Bone Regeneration via Synergistic Bone Remodeling and Angiogenesis.

Osteogenesis, osteoclastogenesis, and angiogenesis are the most important processes in bone repair. Parathyroid hormone (PTH) has pro-osteogenic, pro-osteoclastogenic, and proangiogenic effects and may be a candidate for use in bone defect repair. However, the local application of PTH to bone defects is counterproductive due to its excessive osteoclastic and bone resorptive effects. In this study, a PTH derivative, PTHrP-2, is developed that can be applied to local bone defects. First, a modified peptide with a calcium-binding repeat glutamine tail undergoes controlled local release from a ceramic material and is shown to be a better fit for the repair process than the unmodified peptide. Second, the modified peptide is shown to have strong pro-osteogenic activity due to mineralization and its facilitation of serine (Ser) phosphorylation. Third, the modified peptide is shown to maintain the pro-osteoclastogenic and proangiogenic properties of the unmodified peptide, but its pro-osteoclastogenic activity is reduced compared to that of the unmodified peptide. The reduced pro-osteoclastogenic and increased pro-osteogenic properties of the modified peptide reverse the imbalance between osteoblasts and osteoclasts with local PTH application and shift bone resorption to bone regeneration.

Activation of hedgehog signaling in mesenchymal stem cells induces cartilage and bone tumor formation via Wnt/ß-Catenin.

Indian Hedgehog (IHH) signaling, a key regulator of skeletal development, is highly activated in cartilage and bone tumors. Yet deletion of Ptch1, encoding an inhibitor of IHH receptor Smoothened (SMO), in chondrocyte or osteoblasts does not cause tumorigenesis. Here, we show that Ptch1 deletion in mice Prrx1+mesenchymal stem/stromal cells (MSCs) promotes MSC proliferation and osteogenic and chondrogenic differentiation but inhibits adipogenic differentiation. Moreover, Ptch1 deletion led to development of osteoarthritis-like phenotypes, exostoses, enchondroma, and osteosarcoma in Smo-Gli1/2-dependent manners. The cartilage and bone tumors are originated from Prrx1+ lineage cells and express low levels of osteoblast and chondrocyte markers, respectively. Mechanistically, Ptch1 deletion increases the expression of Wnt5a/6 and leads to enhanced ß-Catenin activation. Inhibiting Wnt/ß-Catenin pathway suppresses development of skeletal anomalies including enchondroma and osteosarcoma. These findings suggest that cartilage/bone tumors arise from their early progenitor cells and identify the Wnt/ß-Catenin pathway as a pharmacological target for cartilage/bone neoplasms.

Customized Mobile LiDAR System for Manhole Cover Detection and Identification.

Manhole covers, which are a key element of urban infrastructure management, have a direct impact on travel safety. At present, there is no automatic, safe, and efficient system specially used for the intelligent detection, identification, and assessment of manhole covers. In this work, we developed an automatic detection, identification, and assessment system for manhole covers. First, we developed a sequential exposure system via the addition of multiple cameras in a symmetrical arrangement to realize the joint acquisition of high-precision laser data and ultra-high-resolution ground images. Second, we proposed an improved histogram of an oriented gradient with symmetry features and a support vector machine method to detect manhole covers effectively and accurately, by using the intensity images and ground orthophotos that are derived from the laser points and images, respectively, and apply the graph segmentation and statistical analysis to achieve the detection, identification, and assessment of manhole covers. Qualitative and quantitative analyses are performed using large experimental datasets that were acquired with the modified manhole-cover detection system. The detected results yield an average accuracy of 96.18%, completeness of 94.27%, and F-measure value of 95.22% in manhole cover detection. Defective manhole-cover monitoring and manhole-cover ownership information are achieved from these detection results. The results not only provide strong support for road administration works, such as data acquisition, manhole cover inquiry and inspection, and statistical analysis of resources, but also demonstrate the feasibility and effectiveness of the proposed method, which reduces the risk involved in performing manual inspections, improves the manhole-cover detection accuracy, and serves as a powerful tool in intelligent road administration.

Lkb1 deletion in periosteal mesenchymal progenitors induces osteogenic tumors through mTORC1 activation.

Bone osteogenic sarcoma has a poor prognosis as the exact cell of origin and the signaling pathways underling tumor formation remain undefined. Here, we report an osteogenic tumor mouse model based on the conditional knockout of liver kinase b1 (Lkb1; also known as Stk11) in Cathepsin K (Ctsk)-Cre expressing cells. Lineage tracing studies demonstrated that Ctsk-Cre could label a population of periosteal cells. The cells functioned as mesenchymal progenitors with regard to markers and functional properties. LKB1 deficiency increased proliferation and osteoblast differentiation of Ctsk+ periosteal cells, while downregulation of mTORC1 activity, using Raptor genetic mouse model or mTORC1 inhibitor treatment, ameliorated tumor progression of Ctsk-Cre Lkb1fllfl mice. Xenograft mouse models, using human osteosarcoma cell lines, also demonstrated that LKB1 deficiency promoted tumor formation, while mTOR inhibition suppressed xenograft tumor growth. In summary, we identified periosteum-derived Ctsk-Cre expressing cells as a cell of origin for osteogenic tumor and suggested the LKB1-mTORC1 pathway as a promising target for treatment of osteogenic tumor.

Roles for HB-EGF in Mesenchymal Stromal Cell Proliferation and Differentiation During Skeletal Growth.

HB-EGF, a member of the EGF superfamily, plays important roles in development and tissue regeneration. However, its functions in skeletal stem cells and skeleton development and growth remain poorly understood. Here, we used the Cre/LoxP system to ablate or express HB-EGF in Dermo1+ mesenchymal stromal cells and their progenies, including chondrocytes and osteoblast lineage cells, and bone marrow stromal cells (BMSCs). Dermo1-Cre; HB-EGFf/f mice only showed a modest increase in bone mass, whereas Dermo1-HB-EGF mice developed progressive chondrodysplasia, chondroma, osteoarthritis-like joint defects, and loss of bone mass and density, which were alleviated by treatment with EGFR inhibitor AG1478. The cartilage defects were recapitulated in chondrocyte-specific HB-EGF overexpression (Col2-HB-EGF) mice with a lesser severity. Dermo1-HB-EGF mice showed an increase in proliferation but defects in differentiation of chondrocytes and osteoblasts. HB-EGF promoted BMSC proliferation via the Akt1 and Erk pathways but inhibited BMSC differentiation via restraining Smad1/5/8 activation. However, Dermo1-HB-EGF mice showed normal osteoclastogenesis and bone resorption. These results reveal an important function of autocrine or paracrine HB-EGF in mesenchymal stromal cell proliferation and differentiation and suggest that EGF signaling needs to be tightly controlled to maintain bone and articular cartilage integrity. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc.

Foxp2 regulates anatomical features that may be relevant for vocal behaviors and bipedal locomotion.

Fundamental human traits, such as language and bipedalism, are associated with a range of anatomical adaptations in craniofacial shaping and skeletal remodeling. However, it is unclear how such morphological features arose during hominin evolution. FOXP2 is a brain-expressed transcription factor implicated in a rare disorder involving speech apraxia and language impairments. Analysis of its evolutionary history suggests that this gene may have contributed to the emergence of proficient spoken language. In the present study, through analyses of skeleton-specific knockout mice, we identified roles of Foxp2 in skull shaping and bone remodeling. Selective ablation of Foxp2 in cartilage disrupted pup vocalizations in a similar way to that of global Foxp2 mutants, which may be due to pleiotropic effects on craniofacial morphogenesis. Our findings also indicate that Foxp2 helps to regulate strength and length of hind limbs and maintenance of joint cartilage and intervertebral discs, which are all anatomical features that are susceptible to adaptations for bipedal locomotion. In light of the known roles of Foxp2 in brain circuits that are important for motor skills and spoken language, we suggest that this gene may have been well placed to contribute to coevolution of neural and anatomical adaptations related to speech and bipedal locomotion.

Valproic acid prevents glucocorticoid­induced osteonecrosis of the femoral head of rats.

Glucocorticoids (GCs) are the most common cause of atraumatic osteonecrosis of the femoral head (ONFH) because their effect compromises the osteogenic capability of bone marrow­derived mesenchymal stem cells (BMSCs). Valproic acid (VPA) is a widely used anti­epileptic and anti­convulsant drug. Previous studies have reported that VPA promotes osteogenic differentiation of MSCs in vitro and osteogenesis in vivo as a histone deacetylase (HDAC) inhibitor. The purpose of the present study was to investigate the efficacy of VPA as a precautionary treatment of ONFH after GC treatment in rats. In vitro, the effect of VPA, dexamethasone or a combination treatment of the two on the proliferation and osteogenic differentiation of human BMSCs was assessed using a Cell Counting Kit­8 and apoptosis assays, and by measuring the expression of proteins associated with osteogenesis. In vivo, a GC­induced ONFH model was established in rats and VPA was added during GC treatment to investigate the preventive effect of VPA against ONFH. Rat BMSCs were also extracted to investigate the osteogenic capacity. The results of micro­computed tomography scanning, angiography of the femoral head and histological and immunohistochemical analyses indicated that 11 of 15 rats induced with methylprednisolone (MP) presented with ONFH, while only 2 of 15 rats treated with a combination of MP and VPA developed ONFH. VPA produced beneficial effects on subchondral bone trabeculae in the femoral head with significant preservation of bone volume and blood supply, as well as improved osteogenic capability of BMSCs compared with those in rats treated with GC alone. In conclusion, VPA attenuated the inhibitory effect of GC on BMSC proliferation and osteogenesis by inhibiting apoptosis and elevating the expression of proteins associated with osteogenesis, which may contribute to the prevention of GC­induced ONFH in rats.