Low sirtuin 1 levels in human osteoarthritis subchondral osteoblasts lead to abnormal sclerostin expression which decreases Wnt/β-catenin activity

IntroductionWnt/β-catenin (cWnt) signaling plays a key role in osteogenesis by promoting the differentiation and mineralization of osteoblasts, activities altered in human osteoarthritic subchondral osteoblast (OA Ob). Sclerostin (SOST) has been shown to alter cWnt signaling. Sirtuin 1 (SIRT1) acts as a novel bone regulator and represses SOST levels in Ob. However the role of SIRT1 and SOST in OA Ob remains unknown. Herein, we explored the role played by SIRT1 and SOST on the abnormal mineralization and cWnt signaling in OA Ob.MethodsPrimary human normal and OA Ob were prepared from tibial plateaus. SOST levels were evaluated by immunohistochemistry, the expression and production of genes by qRT-PCR and WB analysis. Their inhibitions were performed using siRNA. cWnt signaling was measured by the TOPflash TCF/lef luciferase reporter assay. Mineralization was determined by alizarin red staining.ResultsSOST levels were significantly increased in OA Ob compared to normal and were linked with elevated TGF-β1 levels in these cells. SIRT1 expression was significantly reduced in OA Ob compared to normal yet not modified by TGF-β1. Specific inhibition of SIRT1 increased TGF-β1 and SOST expressions in OA Ob, while stimulating SIRT1 activity with β-Nicotinamide mononucleotide reduced the expression of TGF-β1 and SOST, and increased mineralization in OA Ob. Resveratrol also reduced SOST expression in OA Ob. Reduced cWnt signaling, β-catenin levels, and mineralization in OA Ob were all corrected via reducing SOST expression.ConclusionThese data indicate that high level of SOST is responsible, in part, for the reduced cWnt and mineralization of human OA Ob, which in turn is linked with abnormal SIRT1 levels in these pathological cells.     

Regulation of Wnt/β-catenin signaling within and from osteocytes

Bone has long been known to be responsive to mechanical loading. For at least 25 years it has been known that osteocytes sense mechanical load, and because of their response to mechanical loading, osteocytes are believed to be the mechanosensory cell. The Wnt/β-catenin signaling pathway has been shown to be crucial in bone development. Mutations in LRP5 and SOST, which cause high bone mass, have increased interest in the Wnt pathway as a potential target for osteoporosis therapy and have helped link Wnt/β-catenin signaling to bone's response to mechanical loading. Because of its specificity to osteocytes, the Wnt inhibitor sclerostin is a target for anabolic bone therapies. The response of bone to mechanical loading is critically regulated by osteocytes secreting sclerostin, which binds to Lrp5.This article is part of a Special Issue entitled "The Osteocyte".     

Comparison of gene expression of the oncogenic Wnt/β-catenin signaling pathway components in the mouse and human epididymis

β-catenin is an integral part of the Wnt signaling pathway and has been linked to tumorigenesis and multiple developmental processes. The high β-catenin expression with low tumor incidence in the human epididymis is thus intriguing. In the present study, the β-catenin gene and protein was found to be highly expressed in the murine caput epididymidis, and the protein mainly localized along the lateral plasma membranes of adjacent epithelial cells throughout both human and mouse epididymides. Furthermore, the adult mouse epididymis was found to express almost all the Wnt/β-catenin signaling pathway genes that were determined previously by our group in the human organ. Despite the differences in epididymal structure, the similar location of β-catenin and the high concordance of this pathway''s components’ gene expression in both the adult human and mouse epididymides make the mouse a suitable animal model for studying the anti-tumor mechanism of the epididymis. In addition, both the mRNA and protein expression of β-catenin shared a similar spatial expression as the mRNA of Ros1, a proto-oncogene and a key developmental regulator of the initial segment of the mouse epididymis. The observations on the parallel temporal expression of β-catenin and Ros1 during postnatal development raise the possibility that the canonical Wnt signaling pathway has an additional role in the postnatal development of mouse epididymis.     

Lgr4 promotes aerobic glycolysis and differentiation in osteoblasts via the canonical Wnt/β-catenin pathway

Lgr4, a G-protein-coupled receptor, is associated with various physiological and pathological processes including oncogenesis, energy metabolism, and bone remodeling. However, whether Lgr4 is involved in osteoblasts' metabolism is not clear. Here we uncover that in preosteoblast cell line, lacking Lgr4 results in decreased osteogenic function along with reduced glucose consumption, glucose uptake, and lactate production in the presence of abundant oxygen, which is referred to as aerobic glycolysis. Activating canonical Wnt/β-catenin signaling rescued the glycolytic dysfunction. Lgr4 promotes the expression of pyruvate dehydrogenase kinase 1 (pdk1) and is abolished by interfering canonical Wnt/β-catenin signaling. Mice lacking Lgr4 specifically in osteoblasts (Lgr4^osb−/−) exhibit decreased bone mass and strength due to reduced bone formation. Additionally, glycolysis of osteoblasts is impaired in Lgr4^osb−/− mice. Our study reveals a novel function of Lgr4 in regulating the cellular metabolism of osteoblasts. © 2021 American Society for Bone and Mineral Research (ASBMR).     

WNT/β-catenin signaling in polycystic kidney disease

Cystic kidney diseases have been linked to defective WNT signal transduction. Perturbations of cystic disease genes cause activation of canonical WNT/β-catenin/TCF/Lef1 signaling in model organisms and cultured cells. Inappropriate levels of WNT/β-catenin signaling cause renal cyst formation in mice. These observations have prompted the idea that an activation of WNT/β-catenin signaling may constitute a common causative event in cyst formation. Now this view is challenged by key genetic mouse models of cystic kidney disease that do not display WNT/β-catenin activity in cyst-lining epithelia.     

High-dose bone morphogenetic protein–induced ectopic abdomen bone growth

Background contextInfuse [bone morphogenetic protein (BMP)] is increasingly used in spinal fusion surgery.PurposeThe authors report a rare complication of BMP use.Study designThis is a case report.Patient sampleA 55-year-old male underwent a thoracic T8 to the pelvis fusion for degenerative lumbar disc disease and pseudarthrosis at another institution. The procedure involved an anterior and posterior approach with the use of multiple units of BMP.MethodsThe patient presented to our institution with complaints of weight loss, pain, tenderness, and increasing solid growth in the left lower quadrant several months after his surgery. A computed tomography revealed ectopic bone growth in the retroperitoneal area and pelvis contiguous to the anterior lumbar exposure.ResultsThe anterior wound was reexplored, and a large sheet of ectopic bone was removed from the retroperitoneal space.ConclusionsWe report a rare case of extraspinal ectopic bone growth because of the use of multiple packages of BMP.     

Attenuated Wnt/β-catenin signalling mediates methotrexate chemotherapy-induced bone loss and marrow adiposity in rats

Cancer chemotherapy often causes significant bone loss, marrow adiposity and haematopoietic defects, yet the underlying mechanisms and recovery potential remain unclear. Wnt/β-catenin signalling is integral to the regulation of osteogenesis, adipogenesis and haematopoiesis; using a rat model, the current study investigated roles of this signalling pathway in changes to bone marrow stromal and haematopoietic cell differentiation after chemotherapy with methotrexate (MTX), a commonly used antimetabolite. MTX treatment in rats (5 daily administrations at 0.75 mg/kg) has previously been found to decrease bone volume and increase marrow fat, which was associated with increased osteoclastogenesis in haematopoietic cells and with an osteogenesis to adipogenesis switch in bone marrow stromal cells of treated rats. In the current study, on day 6 after the first MTX dose we found that accompanying these changes as well as a suppressed haematopoietic cellularity but increased granulocyte/macrophage differentiation potential, there was an increase in mRNA expression of Wnt antagonists sFRP-1 and Dkk-1 in bone, a reduction in nuclear β-catenin protein in bone marrow stromal cells, and decreased mRNA levels of β-catenin target genes lef-1, cyclin D1 and survivin, suggesting reduced activation of Wnt/β-catenin signalling in the bone during MTX-induced damage. Concurrent administration of BIO, a GSK-3β inhibitor that stabilises β-catenin, partially abrogated the MTX-induced transient changes in osteogenic/adipogenic commitment, granulocyte/macrophage lineage differentiation and osteoclast number. These findings demonstrate a potentially important role of Wnt/β-catenin signalling in MTX chemotherapy-induced cellular changes to the bone marrow microenvironment.     

Acidic preparations of platelet concentrates release bone morphogenetic protein‐2

Background and purpose?Growth factors released from platelets have potent effects on fracture and wound healing. The acidic tide of wound healing, i.e. the pH within wounds and fractures, changes from acidic pH to neutral and alkaline pH as the healing process progresses. We investigated the influence of pH on lysed platelet concentrates regarding the release of growth factors.

Material and methods?Platelet concentrates free of leukocyte components were lysed and incubated in buffers with pH between 4.3 and 8.6. Bone morphogenetic protein-2 (BMP-2), platelet‐derived growth factor (PDGF), transforming growth factor-β (TGF-β), and vascular endothelial growth factor (VEGF) were measured by quantitative enzyme‐linked immunosorbent assays.

Results?PDGF, TGF-β, and VEGF were present in all platelet preparations but the levels varied in a pH‐dependent fashion. BMP-2 was only detected in the most acidic preparation (pH 4.3), which is interesting since BMP-2 has been reported to be an endogenous mediator of fracture repair and to be responsible for the initiation of fracture healing.

Interpretation?Our findings indicate that platelets release substantial amounts of BMP-2 only under conditions of low pH, the milieu associated with the critical initial stage of fracture healing.     

Repression of osteocyte Wnt/β-catenin signaling is an early event in the progression of renal osteodystrophy

Chronic kidney disease-mineral bone disorder (CKD-MBD) is defined by abnormalities in mineral and hormone metabolism, bone histomorphometric changes, and/or the presence of soft-tissue calcification. Emerging evidence suggests that features of CKD-MBD may occur early in disease progression and are associated with changes in osteocyte function. To identify early changes in bone, we utilized the jck mouse, a genetic model of polycystic kidney disease that exhibits progressive renal disease. At 6 weeks of age, jck mice have normal renal function and no evidence of bone disease but exhibit continual decline in renal function and death by 20 weeks of age, when approximately 40% to 60% of them have vascular calcification. Temporal changes in serum parameters were identified in jck relative to wild-type mice from 6 through 18 weeks of age and were subsequently shown to largely mirror serum changes commonly associated with clinical CKD-MBD. Bone histomorphometry revealed progressive changes associated with increased osteoclast activity and elevated bone formation relative to wild-type mice. To capture the early molecular and cellular events in the progression of CKD-MBD we examined cell-specific pathways associated with bone remodeling at the protein and/or gene expression level. Importantly, a steady increase in the number of cells expressing phosphor-Ser33/37-β-catenin was observed both in mouse and human bones. Overall repression of Wnt/β-catenin signaling within osteocytes occurred in conjunction with increased expression of Wnt antagonists (SOST and sFRP4) and genes associated with osteoclast activity, including receptor activator of NF-κB ligand (RANKL). The resulting increase in the RANKL/osteoprotegerin (OPG) ratio correlated with increased osteoclast activity. In late-stage disease, an apparent repression of genes associated with osteoblast function was observed. These data confirm that jck mice develop progressive biochemical changes in CKD-MBD and suggest that repression of the Wnt/β-catenin pathway is involved in the pathogenesis of renal osteodystrophy.     

Wnt/β-catenin信号通路与骨关节炎

近年大量研究证实Wnt/β-catenin信号通路通过调节胚胎期软骨发育及出生后软骨发生、成骨细胞和破骨细胞生成、软骨内成骨、骨重塑、骨折修复等过程,调控骨骼系统相关疾病和骨代谢,对骨关节炎(OA)发生发展、骨软骨组织诱导和修复起着至关重要的作用。基因敲除小鼠模型常用于Wnt/β-catenin信号通路与OA关系研究,全基因组扫描已发现一些与OA密切相关的单核苷酸多态性位点。目前在多个水平进行Wnt/β-catenin信号通路靶向治疗OA研究,重点考虑药物安全性。该文就Wnt/β-catenin与OA关系的研究进展作一综述。     

Canonical Wnt/β-catenin signaling mediates transforming growth factor-β1-driven podocyte injury and proteinuria

Transforming growth factor-β1 (TGF-β1) upregulation occurs in virtually all chronic kidney diseases and is associated with podocyte injury and proteinuria; however, the mechanisms contributing to this in vivo are ambiguous. In vitro, incubation of podocytes with TGF-β1 induced Wnt1 expression, β-catenin activation, and stimulated the expression of Wnt/β-catenin downstream target genes. Ectopic expression of Wnt1 or β-catenin mimicked TGF-β1, induced Snail1, and suppressed nephrin expression. The Wnt antagonist, Dickkopf-1, blocked TGF-β1-induced β-catenin activation, Snail1 induction, and nephrin suppression. In vivo, ectopic expression of TGF-β1 induced Wnt1 expression, activated β-catenin, and upregulated Wnt target genes such as Snail1, MMP-7, MMP-9, desmin, Fsp1, and PAI-1 in mouse glomeruli, leading to podocyte injury and albuminuria. Consistently, concomitant expression of Dickkopf-1 gene abolished β-catenin activation, inhibited TGF-β1-triggered Wnt target gene expression, and mitigated albuminuria. Thus, canonical Wnt/β-catenin signaling mediates TGF-β1-driven podocyte injury and proteinuria. These studies suggest that Wnt/β-catenin signaling may be exploited as a therapeutic target for the treatment of proteinuric kidney diseases.