GATA-1 Deficiency Rescues Trabecular but not Cortical Bone in OPG Deficient Mice

Authors

Tomas E. Meijome, R. Adam Hooker, Ying-Hua Cheng M.D., Ph.D., Whitney Walker, Mark C. Horowitz, Robyn K. Fuchs Ph.D. and Melissa A. Kacena Ph.D.

Abstract

GATA-1low/low mice have an increase in megakaryocytes (MKs) and trabecular bone. The latter is thought to result from MKs directly stimulating osteoblastic bone formation while simultaneously inhibiting osteoclastogenesis. Osteoprotegerin (OPG) is known to inhibit osteoclastogenesis and OPG-/- mice have reduced trabecular and cortical bone due to increased osteoclastogenesis. Interestingly, GATA-1low/low mice have increased OPG levels. Here we sought to determine whether GATA-1 knockdown in OPG-/- mice could rescue the observed osteoporotic bone phenotype. GATA-1low/low mice were bred with OPG-/- mice and bone phenotype assessed. GATA-1low/low X OPG-/- mice have increased cortical bone porosity, similar to OPG-/- mice. Both OPG-/- and GATA-1low/low X OPG-/- mice, were found to have increased osteoclasts localized to cortical bone, possibly producing the observed elevated porosity. Biomechanical assessment indicates that OPG-/- and GATA-1low/low X OPG-/- femurs are weaker and less stiff than C57BL/6 or GATA-1low/low femurs. Notably, GATA-1low/low X OPG-/- mice had trabecular bone parameters that were not different from C57BL/6 values, suggesting that GATA-1 deficiency can partially rescue the trabecular bone loss observed with OPG deficiency. The fact that GATA-1 deficiency appears to be able to partially rescue the trabecular, but not the cortical bone phenotype suggests that MKs can locally enhance trabecular bone volume, but that MK secreted factors cannot access cortical bone sufficiently to inhibit osteoclastogenesis or that OPG itself is required to inhibit osteoclastogenesis in cortical bone

Link To Article

http://dx.doi.org/10.1002/jcp.24803

A Novel Splice-Mutation in PLS3 Causes X-Linked Early-Onset Low-Turnover Osteoporosis

Authors

Christine M. Laine MD PhD, Maija Wessman PhD, Sanna Toiviainen-Salo MD PhD, Mari A. Kaunisto PhD, Mervi K. Mäyränpää MD PhD, Tero Laine MD PhD, Minna Pekkinen PhD, Heikki Kröger MD PhD, Ville-Valtteri Välimäki MD PhD, Matti J. Välimäki MD PhD, Anna-Elina Lehesjoki MD PhD, and Outi Mäkitie MD PhD

Abstract

Genetic factors play an important role in the development of osteoporosis. Several monogenic forms of osteoporosis have been recognized; most recently an X-chromosomal form resulting from mutations in the gene encoding plastin 3 (PLS3). PLS3 is a protein involved in actin bundle formation in the cytoskeleton. We present a large family with early-onset osteoporosis and X-linked inheritance. Phenotyping was performed on 19 family members and whole-exome sequencing on seven family members; five with a diagnosis of early-onset osteoporosis and two with normal bone parameters. Osteoporosis had its onset in childhood and was characterized by recurrent peripheral fractures, low BMD, vertebral compression fractures, and significant height loss in adulthood. Males were in general more severely affected than females. Bone histomorphometry findings in four males and one female showed severe trabecular osteoporosis, low amount of osteoid and decreased mineral apposition rate indicating impaired bone formation; resorption parameters were increased in some. All affected subjects shared a single base substitution (c.73-24T > A) in intron 2 of PLS3 on Xq23. The mutation, confirmed by Sanger sequencing, segregated according to the skeletal phenotype. The mutation introduces a new acceptor splice site with a predicted splice score of 0.99 and thereby, as confirmed by cDNA sequencing, induces the insertion of 22 bases between exons 2 and 3, causing a frameshift and premature termination of mRNA translation (p.Asp25Alafs*17). The mutation affects the first N-terminal calcium-binding EF-hand domain and abolishes all calcium- and actin-binding domains of the protein. Our results confirm the role of PLS3 mutations in early-onset osteoporosis. The mechanism whereby PLS3 affects bone health is unclear, but it may be linked to osteocyte dendrite function and skeletal mechanosensing. Future studies are needed to elucidate the role of PLS3 in osteoporosis and to define optimal treatment.

Link To Article

http://dx.doi.org/10.1002/jbmr.2355

Identical subchondral bone microarchitecture pattern with increased bone reporption observed in rheumatoid arthritis as compared to osteoarthritis

Authors

Guangyi Li, Yuanchen Ma, Tak S. Cheng, Euphemie Landao-Bassonga, An Qin, Nathan J. Pavlos, Changqing Zhang, Qiujian Zheng, Ming H. Zheng

Abstract

Objectives To analyze the differences in microarchitecture and bone remodeling of subchondral bone in femoral heads from patients with rheumatoid arthritis (RA) and osteoarthritis (OA).

Designs Peri-articualr bone samples, including subchondral trabecular bone (STB) and deeper trabecular bone (DTB) were extracted from the load-bearing region of femoral heads from 20 patients with RA and 40 patients with OA during hip replacement surgery. Micro-CT, histomorphometry and backscatter scanning electron microscopy were performed to assess microarchitecture and bone histology parameters.

Results In both RA and OA, STB showed more sclerotic microarchitecture and more active bone remodeling, compared to DTB. RA and OA showed similar microarchitecture characteristics in both STB and DTB, despite STB in RA exhibiting higher bone resorption. In addition, there was no difference in the frequency of bone cysts in STB between RA and OA. In STB, the trabecular bone surrounding subchondral bone cysts (Cys-Tb) was more sclerotic than the trabecular bone found distant to cysts (Peri-Tb), with a higher level of bone remodeling. Both Cys-Tb region and Peri-Tb region were detected to have similar microarchitectural and bone remodeling characteristics in RA and OA.

Conclusions Apart from higher bone resorption in the general subchondral bone of RA samples, the peri-articular bone exhibited similar microarchitectural and bone remodeling characteristics in RA and OA.

Link To Article

http://dx.doi.org/10.1016/j.joca.2014.08.015

The Kynurenine Pathway of Tryptophan Degradation is Activated During Osteoblastogenesis

Authors

Christopher Vidal, Wei Li, Brigitte Santner-Nanan, Chai K. Lim,4, Gilles J. Guillemin, Helen J. Ball, Nicholas H. Hunt, Ralph Nanan and Gustavo Duque

Abstract

The mechanisms involved in the anabolic effect of IFNγ on bone have not been carefully examined. Using microarray expression analysis, we found that IFNγ upregulates a set of genes associated with a tryptophan degradation pathway, known as the kynurenine pathway, in osteogenic differentiating human mesenchymal stem cells. We therefore hypothesized that activation of the kynurenine pathway plays a role in osteoblastogenesis even in the absence of IFNγ. Initially, we observed a strong increase in tryptophan degradation during osteoblastogenesis with and without IFNγ in the media. We next blocked indoleamine 2,3-dyoxygenase-1 (IDO1), the most important enzyme in the kynurenine pathway, using a siRNA and pharmacological approach and observed a strong inhibition of osteoblastogenesis with a concomitant decrease in osteogenic factors. We next examined the bone phenotype of Ido1 knock-out (Ido1-/-) mice. Compared to their wild-type littermates, Ido1-/- mice exhibited osteopenia associated with low osteoblast and high osteoclast numbers. Finally, we tested whether the end-products of the kynurenine pathway have an osteogenic effect on hMSC. We identified that picolinic acid had a strong and dose-dependent osteogenic effect in vitro. In summary, we demonstrate that the activation of the kynurenine pathway plays an important role during the commitment of hMSC into the osteoblast lineage in vitro, and that this process can be accelerated by exogenous addition of IFNγ. In addition, we found that mice lacking IDO1 activity are osteopenic. These data therefore support a new role for the kynurenine pathway and picolinic acid as essential regulators of osteoblastogenesis and as potential new targets of bone-forming cells in vivo.

Link To Article

http://dx.doi.org/10.1002/stem.1836

mTORC2 signaling promotes skeletal growth and bone formation in mice

Authors

Jianquan Chen, Nilsson Holguin, Yu Shi, Matthew J. Silva and Fanxin Long

Abstract

Mammalian target of rapamycin (mTOR) is an evolutionarily conserved serine/threonine kinase controlling many physiological processes in mammals. mTOR functions in two distinct protein complexes, namely mTORC1 and mTORC2. Compared to mTORC1, the specific roles of mTORC2 are less well understood. To investigate the potential contribution of mTORC2 to skeletal development and homeostasis, we have genetically deleted rictor, an essential component of mTORC2, in the limb skeletogenic mesenchyme of the mouse embryo. Loss of rictor leads to shorter and narrower skeletal elements in both embryos and postnatal mice. In the embryo, rictor deletion reduces the width but not the length of the initial cartilage anlage. Subsequently, the embryonic skeletal elements are shortened due to a delay in chondrocyte hypertrophy, with no change in proliferation, apoptosis, cell size or matrix production. Postnatally, rictor-deficient mice exhibit impaired bone formation, resulting in thinner cortical bone, but the trabecular bone mass is relatively normal thanks to a concurrent decrease in bone resorption. Moreover, rictor-deficient bones exhibit a lesser anabolic response to mechanical loading. Thus, mTORC2 signaling is necessary for optimal skeletal growth and bone anabolism.

Link To Article

http://dx.doi.org/10.1002/jbmr.2348

Registration of 2D histological sections with 3D micro-CT datasets from small animal vertebrae and tibiae

Authors

Oleg Museykoa, Robert Percy Marshall, Jing Lu, Andreas Hess, Georg Schett, Michael Amling, Willi A. Kalender & Klaus Engelk

Abstract

The aim of this study was the registration of digitized thin 2D sections of mouse vertebrae and tibiae used for histomorphometry of trabecular bone structure into 3D micro computed tomography (μCT) datasets of the samples from which the sections were prepared. Intensity-based and segmentation-based registrations (SegRegs) of 2D sections and 3D μCT datasets were applied. As the 2D sections were deformed during their preparation, affine registration for the vertebrae was used instead of rigid registration. Tibiae sections were additionally cut on the distal end, which subsequently undergone more deformation so that elastic registration was necessary. The Jaccard distance was used as registration quality measure. The quality of intensity-based registrations and SegRegs was practically equal, although precision errors of the elastic registration of segmentation masks in tibiae were lower, while those in vertebrae were lower for the intensity-based registration. Results of SegReg significantly depended on the segmentation of the μCT datasets. Accuracy errors were reduced from approximately 64% to 42% when applying affine instead of rigid transformations for the vertebrae and from about 43% to 24% when using B-spline instead of rigid transformations for the tibiae. Accuracy errors can also be caused by the difference in spatial resolution between the thin sections (pixel size: 7.25 μm) and the μCT data (voxel size: 15 μm). In the vertebrae, average deformations amounted to a 6.7% shortening along the direction of sectioning and a 4% extension along the perpendicular direction corresponding to 0.13–0.17 mm. Maximum offsets in the mouse tibiae were 0.16 mm on average.

Link To Article

http://dx.doi.org/10.1080/10255842.2014.941824