Generation of two multipotent mesenchymal progenitor cell lines capable of osteogenic, mature osteocyte, adipogenic, and chondrogenic differentiation.

Mesenchymal progenitors differentiate into several tissues including bone, cartilage, and adipose. Targeting these cells in vivo is challenging, making mesenchymal progenitor cell lines valuable tools to study tissue development. Mesenchymal stem cells (MSCs) can be isolated from humans and animals; however, obtaining homogenous, responsive cells in a reproducible fashion is challenging.

Sclerostin Directly Stimulates Osteocyte Synthesis of Fibroblast Growth Factor-23.

Osteocyte produced fibroblast growth factor 23 (FGF23) is the key regulator of serum phosphate (Pi) homeostasis. The interplay between parathyroid hormone (PTH), FGF23 and other proteins that regulate FGF23 production and serum Pi levels is complex and incompletely characterised. Evidence suggests that the protein product of the SOST gene, sclerostin (SCL), also a PTH target and also produced by osteocytes, plays a role in FGF23 expression, however the mechanism for this effect is unclear.

Advancing of Additive-Manufactured Titanium Implants with Bioinspired Micro- to Nanotopographies.

There is an increasing demand for low-cost and more efficient titanium (Ti) medical implants that will provide improved osseointegration and at the same time reduce the likelihood of infection. In the past decade, additive manufacturing (AM) using metal selective laser melting (SLM) or three-dimensional (3D) printing techniques has emerged to enable novel implant geometries or properties to overcome such potential challenges. This study presents a new surface engineering approach to create bioinspired multistructured surfaces on SLM-printed Ti alloy (Ti6Al4V) implants by combining SLM technology, electrochemical anodization, and hydrothermal (HT) processes.

Novel Insights into Staphylococcus aureus Deep Bone Infections: the Involvement of Osteocytes.

Periprosthetic joint infection (PJI) is a potentially devastating complication of orthopedic joint replacement surgery. PJI with associated osteomyelitis is particularly problematic and difficult to cure. Whether viable osteocytes, the predominant cell type in mineralized bone tissue, have a role in these infections is not clear, although their involvement might contribute to the difficulty in detecting and clearing PJI.

Both ligand and VDR expression levels critically determine the effect of 1α,25-dihydroxyvitamin-D on osteoblast differentiation.

Previous studies have shown that 1α,25-dihydroxyvitamin D (1,25D) through vitamin D receptor (VDR) signalling has both catabolic and anabolic effects on osteoblast differentiation. However, the mechanism of these differential effects by 1,25D is not fully understood. In this study, mice with three different genetic backgrounds, representing a normal VDR level (wild-type, WT), VDR over-expression specifically in mature osteoblasts (ObVDR-B6) and global VDR knockout (VDRKO), were utilised to generate primary osteoblast-like cultures to further elucidate the effects of 1,25D on osteoblast differentiation.

Isolation of osteocytes from human trabecular bone.

Osteocytes are essential regulators of bone homeostasis. However, they are difficult to study due to their location within the bone mineralised matrix. Although several techniques have been published for the isolation of osteocytes from mouse bone, no such technique has been described for human osteocytes.

Early response of the human SOST gene to stimulation by 1α,25-dihydroxyvitamin D.

The osteocyte expressed gene SOST encodes sclerostin, a potent negative regulator of bone formation and inducer of bone resorption. We have recently demonstrated that the human SOST gene is positively regulated in response to 1α,25-dihydroxyvitamin D (1,25D). Responsiveness may be mediated at least in part by a single classical DR3-type vitamin D response element (VDRE).

1α,25-dihydroxyvitamin D3 stimulates human SOST gene expression and sclerostin secretion.

Sclerostin, the SOST gene product, is a negative regulator of bone formation and a positive regulator of bone resorption. In this study, treatment of human primary osteoblasts, including cells differentiated to an osteocyte-like stage, with 1α,25-dihydroxyvitaminD3 (1,25D) resulted in the dose-dependent increased expression of SOST mRNA. A similar effect was observed in human trabecular bone samples cultured ex vivo, and in osteocyte-like cultures of differentiated SAOS2 cells.

1,25-Dihydroxyvitamin D3 and extracellular calcium promote mineral deposition via NPP1 activity in a mature osteoblast cell line MLO-A5.

While vitamin D supplementation is common, the anabolic mechanisms that improve bone status are poorly understood. Under standard mineralising conditions including media ionised calcium of 1.1 mM, 1,25-dihydroxyvitamin D3 (1,25D) enhanced differentiation and mineral deposition by the mature osteoblast/pre-osteocyte cell line, MLO-A5.

Regulation of FGF23 expression in IDG-SW3 osteocytes and human bone by pro-inflammatory stimuli.

Fibroblast growth factor-23 (FGF23), produced by osteocytes, is the key physiological regulator of phosphate homeostasis. Sepsis patients often experience transient hypophosphataemia, suggesting the regulation of FGF23 levels by pro-inflammatory factors. Here, we used the osteocyte-like cell line IDG-SW3 to investigate the effect of pro-inflammatory stimuli on FGF23 production.

SaOS2 Osteosarcoma cells as an in vitro model for studying the transition of human osteoblasts to osteocytes.

The central importance of osteocytes in regulating bone homeostasis is becoming increasingly apparent. However, the study of these cells has been restricted by the relative paucity of cell line models, especially those of human origin. Therefore, we investigated the extent to which SaOS2 human osteosarcoma cells can differentiate into osteocyte-like cells.

Sclerostin regulates release of bone mineral by osteocytes by induction of carbonic anhydrase 2.

The osteocyte product sclerostin is emerging as an important paracrine regulator of bone mass. It has recently been shown that osteocyte production of receptor activator of NF-κB ligand (RANKL) is important in osteoclastic bone resorption, and we reported that exogenous treatment of osteocytes with sclerostin can increase RANKL-mediated osteoclast activity. There is good evidence that osteocytes can themselves liberate mineral from bone in a process known as osteocytic osteolysis.

Sclerostin stimulates osteocyte support of osteoclast activity by a RANKL-dependent pathway.

Sclerostin is a product of mature osteocytes embedded in mineralised bone and is a negative regulator of bone mass and osteoblast differentiation. While evidence suggests that sclerostin has an anti-anabolic role, the possibility also exists that sclerostin has catabolic activity. To test this we treated human primary pre-osteocyte cultures, cells we have found are exquisitely sensitive to sclerostin, or mouse osteocyte-like MLO-Y4 cells, with recombinant human sclerostin (rhSCL) and measured effects on pro-catabolic gene expression.

Sclerostin is a locally acting regulator of late-osteoblast/preosteocyte differentiation and regulates mineralization through a MEPE-ASARM-dependent mechanism.

The identity of the cell type responsive to sclerostin, a negative regulator of bone mass, is unknown. Since sclerostin is expressed in vivo by mineral-embedded osteocytes, we tested the hypothesis that sclerostin would regulate the behavior of cells actively involved in mineralization in adult bone, the preosteocyte. Differentiating cultures of human primary osteoblasts exposed to recombinant human sclerostin (rhSCL) for 35 days displayed dose- and time-dependent inhibition of in vitro mineralization, with late cultures being most responsive in terms of mineralization and gene expression.

Vitamin K promotes mineralization, osteoblast-to-osteocyte transition, and an anticatabolic phenotype by {gamma}-carboxylation-dependent and -independent mechanisms.

The vitamin K family members phylloquinone (vitamin K1) and the menaquinones (vitamin K2) are under study for their roles in bone metabolism and as potential therapeutic agents for skeletal diseases. We have investigated the effects of two naturally occurring homologs, phytonadione (vitamin K1) and menatetrenone (vitamin K2), and those of the synthetic vitamin K, menadione (vitamin K3), on human primary osteoblasts. All homologs promoted in vitro mineralization by these cells.

Pro-inflammatory cytokines TNF-related weak inducer of apoptosis (TWEAK) and TNFalpha induce the mitogen-activated protein kinase (MAPK)-dependent expression of sclerostin in human osteoblasts.

We have recently shown that TNF-related weak inducer of apoptosis (TWEAK) is a mediator of inflammatory bone remodeling. The aim of this study was to investigate the role of TWEAK in modulating human osteoblast activity, and how TWEAK and TNFalpha might interact in this context. Recombinant TWEAK and TNF were both mitogenic for human primary osteoblasts (NHBC).