Biomechanical properties of bone in a mouse model of Rett syndrome

PubMed Central

Kamal, Bushra; Russell, David; Payne, Anthony; Constante, Diogo; Tanner, K. Elizabeth; Isaksson, Hanna; Mathavan, Neashan; Cobb, Stuart R.

2015-01-01

Rett syndrome (RTT) is an X-linked genetic disorder and a major cause of intellectual disability in girls. Mutations in the methyl-CpG binding protein 2 (MECP2) gene are the primary cause of the disorder. Despite the dominant neurological phenotypes, MECP2 is expressed ubiquitously throughout the body and a number of peripheral phenotypes such as scoliosis, reduced bone mineral density and skeletal fractures are also common and important clinical features of the disorder. In order to explore whether MeCP2 protein deficiency results in altered structural and functional properties of bone and to test the potential reversibility of any defects, we have conducted a series of histological, imaging and biomechanical tests of bone in a functional knockout mouse model of RTT. Both hemizygous Mecp2stop/y male mice in which Mecp2 is silenced in all cells and female Mecp2stop/+ mice in which Mecp2 is silenced in ~ 50% of cells as a consequence of random X-chromosome inactivation, revealed significant reductions in cortical bone stiffness, microhardness and tensile modulus. Microstructural analysis also revealed alterations in both cortical and cancellous femoral bone between wild-type and MeCP2-deficient mice. Furthermore, unsilencing of Mecp2 in adult mice cre-mediated stop cassette deletion resulted in a restoration of biomechanical properties (stiffness, microhardness) towards wild-type levels. These results show that MeCP2-deficiency results in overt, but potentially reversible, alterations in the biomechanical integrity of bone and highlights the importance of targeting skeletal phenotypes in considering the development of pharmacological and gene-based therapies. PMID:25445449

Anti-osteoporotic activity of harpagide by regulation of bone formation in osteoblast cell culture and ovariectomy-induced bone loss mouse models.

PubMed

Chung, Hwa-Jin; Kyung Kim, Won; Joo Park, Hyen; Cho, Lan; Kim, Me-Riong; Kim, Min Jeong; Shin, Joon-Shik; Ho Lee, Jin; Ha, In-Hyuk; Kook Lee, Sang

2016-02-17

Harpagide, an iridoid glucoside, is a constituent of the root of Harpagophytum procumbens var. sublobatum (Engl.) Stapf, Devil's claw which has been used in patients with osteoarthritis (OA). In the present study, we investigated the anti-osteoporotic potential of harpagide and its underlying mechanism of action in in vitro cell culture and in vivo bone loss animal models. Harpagide was obtained from the alkalic hydrolysis of harpagoside, a major constituent of H. procumbens var. sublobatum Analysis of biomarkers for bone formation in osteoblastic MC3T3-E1 cells and bone resorption in osteoclast cells derived from mouse bone marrow cells was performed to evaluate the mechanism of action. The protective activity of harpagide against bone loss was also evaluated in ovariectomized (OVX) mouse model. Harpagide improved bone properties by stimulating the process of differentiation and maturation of osteoblast cells and suppressing the process of RANKL-induced differentiation of osteoclast cells. In OVX-induced bone loss mouse model, oral administration of harpagide significantly improved recovery of bone mineral density, trabecular bone volume, and trabecular number in the femur. Harpagide also prevented increase of trabecular separation and structure model index induced by OVX. Harpagide effectively inhibited the serum levels of biochemical markers of bone loss, including alkaline phosphatase, osteocalcin, C-terminal telopeptide, and tartrate-resistant acid phosphatase. Taken together, the present study demonstrates that harpagide has a potential for prevention of bone loss in OVX mice by regulating the stimulation of osteoblast differentiation and the suppression of osteoclast formation. Therefore, these findings suggest that harpagide might serve as a bioactive compound derived from H. procumbens var. sublobatum for improvement of age-dependent bone destruction disease. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Primary Mouse Myoblast Purification using Magnetic Cell Separation.

PubMed

Sincennes, Marie Claude; Wang, Yu Xin; Rudnicki, Michael A

2017-01-01

Primary myoblasts can be isolated from mouse muscle cell extracts and cultured in vitro. Muscle cells are usually dissociated manually by mincing with razor blades or scissors in a collagenase/dispase solution. Primary myoblasts are then gradually enriched by pre-plating on collagen-coated plates, based on the observation that mouse fibroblasts attach quickly to collagen-coated plates, and are less adherent. Here, we describe an automated muscle dissociation protocol. We also propose an alternative to pre-plating using magnetic bead separation of primary myoblasts, which improve myoblast purity by minimizing fibroblast contamination.

Automated Cell Detection and Morphometry on Growth Plate Images of Mouse Bone

PubMed Central

Ascenzi, Maria-Grazia; Du, Xia; Harding, James I; Beylerian, Emily N; de Silva, Brian M; Gross, Ben J; Kastein, Hannah K; Wang, Weiguang; Lyons, Karen M; Schaeffer, Hayden

2014-01-01

Microscopy imaging of mouse growth plates is extensively used in biology to understand the effect of specific molecules on various stages of normal bone development and on bone disease. Until now, such image analysis has been conducted by manual detection. In fact, when existing automated detection techniques were applied, morphological variations across the growth plate and heterogeneity of image background color, including the faint presence of cells (chondrocytes) located deeper in tissue away from the image’s plane of focus, and lack of cell-specific features, interfered with identification of cell. We propose the first method of automated detection and morphometry applicable to images of cells in the growth plate of long bone. Through ad hoc sequential application of the Retinex method, anisotropic diffusion and thresholding, our new cell detection algorithm (CDA) addresses these challenges on bright-field microscopy images of mouse growth plates. Five parameters, chosen by the user in respect of image characteristics, regulate our CDA. Our results demonstrate effectiveness of the proposed numerical method relative to manual methods. Our CDA confirms previously established results regarding chondrocytes’ number, area, orientation, height and shape of normal growth plates. Our CDA also confirms differences previously found between the genetic mutated mouse Smad1/5CKO and its control mouse on fluorescence images. The CDA aims to aid biomedical research by increasing efficiency and consistency of data collection regarding arrangement and characteristics of chondrocytes. Our results suggest that automated extraction of data from microscopy imaging of growth plates can assist in unlocking information on normal and pathological development, key to the underlying biological mechanisms of bone growth. PMID:25525552

Myricetin Prevents Alveolar Bone Loss in an Experimental Ovariectomized Mouse Model of Periodontitis

PubMed Central

Huang, Jialiang; Wu, Chuanlong; Tian, Bo; Zhou, Xiao; Ma, Nian; Qian, Yufen

2016-01-01

Periodontitis is a common chronic inflammatory disease, which leads to alveolar bone resorption. Healthy and functional alveolar bone, which can support the teeth and enable their movement, is very important for orthodontic treatment. Myricetin inhibited osteoclastogenesis by suppressing the expression of some genes, signaling pathways, and cytokines. This study aimed to investigate the effects of myricetin on alveolar bone loss in an ovariectomized (OVX) mouse model of periodontitis as well as in vitro osteoclast formation and bone resorption. Twenty-four healthy eight-week-old C57BL/J6 female mice were assigned randomly to four groups: phosphate-buffered saline (PBS) control (sham) OVX + ligature + PBS (vehicle), and OVX + ligature + low or high (2 or 5 mg∙kg−1∙day−1, respectively) doses of myricetin. Myricetin or PBS was injected intraperitoneally (i.p.) every other day for 30 days. The maxillae were collected and subjected to further examination, including micro-computed tomography (micro-CT), hematoxylin and eosin (H&E) staining, and tartrate-resistant acid phosphatase (TRAP) staining; a resorption pit assay was also performed in vitro to evaluate the effects of myricetin on receptor activator of nuclear factor κ-B ligand (RANKL)-induced osteoclastogenesis. Myricetin, at both high and low doses, prevented alveolar bone resorption and increased alveolar crest height in the mouse model and inhibited osteoclast formation and bone resorption in vitro. However, myricetin was more effective at high dose than at low dose. Our study demonstrated that myricetin had a positive effect on alveolar bone resorption in an OVX mouse model of periodontitis and, therefore, may be a potential agent for the treatment of periodontitis and osteoporosis. PMID:27011174

Mutagenicity testing with transgenic mice. Part I: Comparison with the mouse bone marrow micronucleus test

PubMed Central

Wahnschaffe, U; Bitsch, A; Kielhorn, J; Mangelsdorf, I

2005-01-01

As part of a larger literature study on transgenic animals in mutagenicity testing, test results from the transgenic mutagenicity assays (lacI model; commercially available as the Big Blue® mouse, and the lacZ model; commercially available as the Muta™Mouse), were compared with the results on the same substances in the more traditional mouse bone marrow micronucleus test. 39 substances were found which had been tested in the micronucleus assay and in the above transgenic mouse systems. Although, the transgenic animal mutation assay is not directly comparable with the micronucleus test, because different genetic endpoints are examined: chromosome aberration versus gene mutation, the results for the majority of substances were in agreement. Both test systems, the transgenic mouse assay and the mouse bone marrow micronucleus test, have advantages and they complement each other. However, the transgenic animal assay has some distinct advantages over the micronucleus test: it is not restricted to one target organ and detects systemic as well as local mutagenic effects. PMID:15655069

The terminator mouse: salvation for primary cell culture.

PubMed

Kabgani, Nazanin; Moeller, Marcus J

2013-11-01

The Terminator had to come back from the future already several times in an effort to bring salvation to mankind. In the present issue of Kidney International, Guo et al. brought us a novel transgenic mouse model: the terminator mouse. This highly elegant mouse may facilitate significantly the derivation of primary cultures of a specific cell type from a tissue containing multiple cell populations.

The bone diagnostic instrument III: Testing mouse femora

NASA Astrophysics Data System (ADS)

Randall, Connor; Mathews, Phillip; Yurtsev, Eugene; Sahar, Nadder; Kohn, David; Hansma, Paul

2009-06-01

Here we describe modifications that allow the bone diagnostic instrument (BDI) [P. Hansma et al., Rev. Sci. Instrum. 79, 064303 (2008); Rev. Sci. Instrum. 77, 075105 (2006)], developed to test human bone, to test the femora of mice. These modifications include reducing the effective weight of the instrument on the bone, designing and fabricating new probe assemblies to minimize damage to the small bone, developing new testing protocols that involve smaller testing forces, and fabricating a jig for securing the smaller bones for testing. With these modifications, the BDI was used to test the hypothesis that short-term running has greater benefit on the mechanical properties of the femur for young growing mice compared to older, skeletally mature mice. We measured elastic modulus, hardness, and indentation distance increase (IDI), which had previously been shown to be the best discriminators in model systems known to exhibit differences in mechanical properties at the whole bone level. In the young exercised murine femora, the IDI was significantly lower than in young control femora. Since IDI has a relation to postyield properties, these results suggest that exercise during bone development increases post yield mechanical competence. We were also able to measure effects of aging on bone properties with the BDI. There was a significant increase in the IDI, and a significant decrease in the elastic modulus and hardness between the young and old groups. Thus, with the modifications described here, the BDI can take measurements on mouse bones and obtain statistically significant results.

Generation and Identification of GM-CSF Derived Alveolar-like Macrophages and Dendritic Cells From Mouse Bone Marrow

PubMed Central

Dong, Yifei; Arif, Arif A.; Poon, Grace F. T.; Hardman, Blair; Dosanjh, Manisha; Johnson, Pauline

2016-01-01

Macrophages and dendritic cells (DCs) are innate immune cells found in tissues and lymphoid organs that play a key role in the defense against pathogens. However, they are difficult to isolate in sufficient numbers to study them in detail, therefore, in vitro models have been developed. In vitro cultures of bone marrow-derived macrophages and dendritic cells are well-established and valuable methods for immunological studies. Here, a method for culturing and identifying both DCs and macrophages from a single culture of primary mouse bone marrow cells using the cytokine granulocyte macrophage colony-stimulating factor (GM-CSF) is described. This protocol is based on the established procedure first developed by Lutz et al. in 1999 for bone marrow-derived DCs. The culture is heterogeneous, and MHCII and fluoresceinated hyaluronan (FL-HA) are used to distinguish macrophages from immature and mature DCs. These GM-CSF derived macrophages provide a convenient source of in vitro derived macrophages that closely resemble alveolar macrophages in both phenotype and function. PMID:27404290

Bone disease in primary hyperparathyroidism

PubMed Central

Bandeira, Francisco; Cusano, Natalie E.; Silva, Barbara C.; Cassibba, Sara; Almeida, Clarissa Beatriz; Machado, Vanessa Caroline Costa; Bilezikian, John P.

2015-01-01

Bone disease in severe primary hyperparathyroidism (PHPT) is described classically as osteitis fibrosa cystica (OFC). Bone pain, skeletal deformities and pathological fractures are features of OFC. Bone mineral density is usually extremely low in OFC, but it is reversible after surgical cure. The signs and symptoms of severe bone disease include bone pain, pathologic fractures, proximal muscle weakness with hyperreflexia. Bone involvement is typically characterized as salt-and-pepper appearance in the skull, bone erosions and bone resorption of the phalanges, brown tumors and cysts. In the radiography, diffuse demineralization is observed, along with pathological fractures, particularly in the long bones of the extremities. In severe, symptomatic PHPT, marked elevation of the serum calcium and PTH concentrations are seen and renal involvement is manifested by nephrolithiasis and nephrocalcinosis. A new technology, recently approved for clinical use in the United States and Europe, is likely to become more widely available because it is an adaptation of the lumbar spine DXA image. Trabecular bone score (TBS) is a gray-level textural analysis that provides an indirect index of trabecular microarchitecture. Newer technologies, such as high-resolution peripheral quantitative computed tomography (HR-pQCT), have provided further understanding of the microstructural skeletal features in PHPT. PMID:25166047

Choroidal metastasis from primary bone leiomyosarcoma.

PubMed

Cristina, Nieto Gómez; Francisco, Escudero Domínguez; Vanesa, Rivero Gutiérrez; Fernando, Cruz González; Luis, Cacharro Moras; Emiliano, Hernández Galilea

2015-10-01

Choroidal metastases, the most common form of intraocular malignancies, are principally caused by primary tumors from breast, lung, and gastrointestinal tract. These lesions are mostly symptomatic and rarely detected incidentally in the extension study of a previously diagnosed tumor. Leiomyosarcoma is a neoplasm of mesenchymal cells with smooth muscle differentiation and represents the most prevalent soft-tissue sarcoma. Leiomyosarcoma is a notably rare tumor in ophthalmic region. We report a case of primary bone leiomyosarcoma metastatic to the choroid that was treated with chemotherapy and surgery. Although three cases of choroidal metastasis from leiomyosarcomas have been already reported, to our knowledge this is the first case of choroidal metastasis from primary bone leiomyosarcoma.

Remnant Woven Bone and Calcified Cartilage in Mouse Bone: Differences between Ages/Sex and Effects on Bone Strength

PubMed Central

Ip, Victoria; Toth, Zacharie; Chibnall, John; McBride-Gagyi, Sarah

2016-01-01

Introduction Mouse models are used frequently to study effects of bone diseases and genetic determinates of bone strength. Murine bones have an intracortical band of woven bone that is not present in human bones. This band is not obvious under brightfield imaging and not typically analyzed. Due to the band’s morphology and location it has been theorized to be remnant bone from early in life. Furthermore, lamellar and woven bone are well known to have differing mechanical strengths. The purpose of this study was to determine (i) if the band is from early life and (ii) if the woven bone or calcified cartilage contained within the band affect whole bone strength. Woven Bone Origin Studies In twelve to fourteen week old mice, doxycycline was used to label bone formed prior to 3 weeks old. Doxycycline labeling and woven bone patterns on contralateral femora matched well and encompassed an almost identical cross-sectional area. Also, we highlight for the first time in mice the presence of calcified cartilage exclusively within the band. However, calcified cartilage could not be identified on high resolution cone-beam microCT scans when examined visually or by thresholding methods. Mechanical Strength Studies Subsequently, three-point bending was used to analyze the effects of woven bone and calcified cartilage on whole bone mechanics in a cohort of male and female six and 13 week old Balb/C mice. Three-point bending outcomes were correlated with structural and compositional measures using multivariate linear regression. Woven bone composed a higher percent of young bones than older bones. However, calcified cartilage in older bones was twice that of younger bones, which was similar when normalized by area. Area and/or tissue mineral density accounted for >75% of variation for most strength outcomes. Percent calcified cartilage added significant predictive power to maximal force and bending stress. Calcified cartilage and woven bone could have more influence in genetic

Establishment of a bilateral femoral large segmental bone defect mouse model potentially applicable to basic research in bone tissue engineering.

PubMed

Xing, Junchao; Jin, Huiyong; Hou, Tianyong; Chang, Zhengqi; Luo, Fei; Wang, Pinpin; Li, Zhiqiang; Xie, Zhao; Xu, Jianzhong

2014-12-01

To understand the cellular mechanism underlying bone defect healing in the context of tissue engineering, a reliable, reproducible, and standardized load-bearing large segmental bone defect model in small animals is indispensable. The aim of this study was to establish and evaluate a bilateral femoral defect model in mice. Donor mouse bone marrow mesenchymal stem cells (mBMSCs) were obtained from six mice (FVB/N) and incorporated into partially demineralized bone matrix scaffolds to construct tissue-engineered bones. In total, 36 GFP(+) mice were used for modeling. Titanium fixation plates with locking steel wires were attached to the femurs for stabilization, and 2-mm-long segmental bone defects were created in the bilateral femoral midshafts. The defects in the left and right femurs were transplanted with tissue-engineered bones and control scaffolds, respectively. The healing process was monitored by x-ray radiography, microcomputed tomography, and histology. The capacity of the transplanted mBMSCs to recruit host CD31(+) cells was investigated by immunofluorescence and real-time polymerase chain reaction. Postoperatively, no complication was observed, except that two mice died of unknown causes. Stable fixation of femurs and implants with full load bearing was achieved in all animals. The process of bone defect repair was significantly accelerated due to the introduction of mBMSCs. Moreover, the transplanted mBMSCs attracted more host CD31(+) endothelial progenitors into the grafts. The present study established a feasible, reproducible, and clinically relevant bilateral femoral large segmental bone defect mouse model. This model is potentially suitable for basic research in the field of bone tissue engineering. Copyright © 2014 Elsevier Inc. All rights reserved.

Absence of bone sialoprotein (BSP) impairs cortical defect repair in mouse long bone.

PubMed

Malaval, Luc; Monfoulet, Laurent; Fabre, Thierry; Pothuaud, Laurent; Bareille, Reine; Miraux, Sylvain; Thiaudiere, Eric; Raffard, Gerard; Franconi, Jean-Michel; Lafage-Proust, Marie-Hélène; Aubin, Jane E; Vico, Laurence; Amédée, Joëlle

2009-11-01

Matrix proteins of the SIBLING family interact with bone cells and with bone mineral and are thus in a key position to regulate bone development, remodeling and repair. Within this family, bone sialoprotein (BSP) is highly expressed by osteoblasts, hypertrophic chondrocytes and osteoclasts. We recently reported that mice lacking BSP (BSP-/-) have very low trabecular bone turnover. In the present study, we set up an experimental model of bone repair by drilling a 1 mm diameter hole in the cortical bone of femurs in both BSP-/- and +/+ mice. A non-invasive MRI imaging and bone quantification procedure was designed to follow bone regeneration, and these data were extended by microCT imaging and histomorphometry on undecalcified sections for analysis at cellular level. These combined approaches revealed that the repair process as reflected in defect-refilling in the cortical area was significantly delayed in BSP-/- mice compared to +/+ mice. Concomitantly, histomorphometry showed that formation, mineralization and remodeling of repair (primary) bone in the medulla were delayed in BSP-/- mice, with lower osteoid and osteoclast surfaces at day 15. In conclusion, the absence of BSP delays bone repair at least in part by impairing both new bone formation and osteoclast activity.

The Influence of Primary Microenvironment on Prostate Cancer Osteoblastic Bone Lesion Development

DTIC Science & Technology

2015-09-01

for inhibiting PCa bone lesion development: 3a. Basic fibroblast growth factor (bFGF) in PC3 bone metastasis: bFGF was identified by cytokine...II receptor (TβRII) knockout (Tgfbr2 KO) mouse models. Col1creERT/Tgfbr2 KO (Col/Tgfbr2 KO), which have TGF-β signaling specific KO in fibroblasts ... fibroblasts and osteoblasts in the bone by Colcre/Tgfbr2 KO, or in the myeloid lineage cells, including osteoclasts in the bone by LysMcre/Tgfbr2 KO

Optimisation of the differing conditions required for bone formation in vitro by primary osteoblasts from mice and rats

PubMed Central

ORRISS, ISABEL R.; HAJJAWI, MARK O.R.; HUESA, CARMEN; MACRAE, VICKY E.; ARNETT, TIMOTHY R.

2014-01-01

The in vitro culture of calvarial osteoblasts from neonatal rodents remains an important method for studying the regulation of bone formation. The widespread use of transgenic mice has created a particular need for a reliable, simple method that allows the differentiation and bone-forming activity of murine osteoblasts to be studied. In the present study, we established such a method and identified key differences in optimal culture conditions between mouse and rat osteoblasts. Cells isolated from neonatal rodent calvariae by collagenase digestion were cultured for 14–28 days before staining for tissue non-specific alkaline phosphatase (TNAP) and bone mineralisation (alizarin red). The reliable differentiation of mouse osteoblasts, resulting in abundant TNAP expression and the formation of mineralised ‘trabecular-shaped’ bone nodules, occurred only following culture in α minimum essential medium (αMEM) and took 21–28 days. Dexamethasone (10 nM) inhibited bone mineralisation in the mouse osteoblasts. By contrast, TNAP expression and bone formation by rat osteoblasts were observed following culture in both αMEM and Dulbecco’s modified Eagle’s medium (DMEM) after approximately 14 days (although ~3-fold more effectively in αMEM) and was strongly dependent on dexamethasone. Both the mouse and rat osteoblasts required ascorbate (50 μg/ml) for osteogenic differentiation and β-glycerophosphate (2 mM) for mineralisation. The rat and mouse osteoblasts showed similar sensitivity to the well-established inhibitors of mineralisation, inorganic pyrophosphate (PPi) and adenosine triphosphate (ATP; 1–100 μM). The high efficiency of osteogenic differentiation observed following culture in αMEM, compared with culture in DMEM possibly reflects the richer formulation of the former. These findings offer a reliable technique for inducing mouse osteoblasts to form bone in vitro and a more effective method for culturing bone-forming rat osteoblasts. PMID:25200658

How tough is Brittle Bone? Investigating Osteogenesis Imperfecta in Mouse Bone††

PubMed Central

Carriero, A.; Zimmermann, E. A.; Paluszny, A.; Tang, S. Y.; Bale, H.; Busse, B.; Alliston, T.; Kazakia, G.

2015-01-01

The multiscale hierarchical structure of bone is naturally optimized to resist fractures. In osteogenesis imperfecta, or brittle bone disease, genetic mutations affect the quality and/or quantity of collagen, dramatically increasing bone fracture risk. Here we reveal how the collagen defect results in bone fragility in a mouse model of osteogenesis imperfecta (oim), which has homotrimeric α1(I) collagen. At the molecular level we attribute the loss in toughness to a decrease in the stabilizing enzymatic crosslinks and an increase in non-enzymatic crosslinks, which may break prematurely inhibiting plasticity. At the tissue level, high vascular canal density reduces the stable crack growth, and extensive woven bone limits the crack-deflection toughening during crack growth. This demonstrates how modifications at the bone molecular level have ramifications at larger length scales affecting the overall mechanical integrity of the bone; thus, treatment strategies have to address multiscale properties in order to regain bone toughness. In this regard, findings from the heterozygous oim bone, where defective as well as normal collagen are present, suggest that increasing the quantity of healthy collagen in these bones helps to recover toughness at the multiple length scales. PMID:24420672

Chronic CCl4 intoxication causes liver and bone damage similar to the human pathology of hepatic osteodystrophy: a mouse model to analyse the liver-bone axis.

PubMed

Nussler, Andreas K; Wildemann, Britt; Freude, Thomas; Litzka, Christian; Soldo, Petra; Friess, Helmut; Hammad, Seddik; Hengstler, Jan G; Braun, Karl F; Trak-Smayra, Viviane; Godoy, Patricio; Ehnert, Sabrina

2014-04-01

Patients with chronic liver diseases frequently exhibit decreased bone mineral densities (BMD), which is defined as hepatic osteodystrophy (HOD). HOD is a multifactorial disease whose regulatory mechanisms are barely understood. Thus, an early diagnosis and therapy is hardly possible. Therefore, the aim of our study consisted in characterizing a mouse model reflecting the human pathomechanism. Serum samples were collected from patients with chronic liver diseases and 12-week old C57Bl6/N mice after 6-week treatment with carbon tetrachloride (CCl4). Repetitive injections of CCl4 induced liver damage in mice, resembling liver fibrosis in patients, as assessed by serum analysis and histological staining. Although CCl4 did not affect primary osteoblast cultures, μCT analysis revealed significantly decreased BMD, bone volume, trabecular number and thickness in CCl4-treated mice. In both HOD patients and CCl4-treated mice, an altered vitamin D metabolism with decreased CYP27A1, CYP2R1, vitamin D-binding protein GC and increased 7-dehydrocholesterol reductase hepatic gene expression, results in decreased 25-OH vitamin D serum levels. Moreover, both groups exhibit excessively high active transforming growth factor-beta (TGF-β) serum levels, inhibiting osteoblast function in vitro. Summarizing, our mouse model presents possible mediators of HOD, e.g. altered vitamin D metabolism and increased active TGF-β. Liver damage and significant changes in bone structure and mineralization are already visible by μCT analysis after 6 weeks of CCl4 treatment. This fast response and easy transferability makes it an ideal model to investigate specific gene functions in HOD.

Primary bone marrow oedema syndromes.

PubMed

Patel, Sanjeev

2014-05-01

MRI scanning in patients with rheumatological conditions often shows bone marrow oedema, which can be secondary to inflammatory, degenerative, infective or malignant conditions but can also be primary. The latter condition is of uncertain aetiology and it is also uncertain whether it represents a stage in the progression to osteonecrosis in some patients. Patients with primary bone marrow oedema usually have lower limb pain, commonly the hip, knee, ankle or feet. The diagnosis is one of exclusion with the presence of typical MRI findings. Treatment is usually conservative and includes analgesics and staying off the affected limb. The natural history is that of gradual resolution of symptoms over a number of months. Evidence for medical treatment is limited, but open-label studies suggest bisphosphonates may help in the resolution of pain and improve radiological findings. Surgical decompression is usually used as a last resort.

Mineral metabolism in isolated mouse long bones: Opposite effects of microgravity on mineralization and resorption

NASA Technical Reports Server (NTRS)

Veldhuijzen, Jean Paul; Vanloon, Jack J. W. A.

1994-01-01

An experiment using isolated skeletal tissues under microgravity, is reported. Fetal mouse long bones (metatarsals) were cultured for 4 days in the Biorack facility of Spacelab during the IML-1 (International Microgravity Laboratory) mission of the Space Shuttle. Overall growth was not affected, however glucose consumption was significantly reduced under microgravity. Mineralization of the diaphysis was also strongly reduced under microgravity as compared to the on-board 1 g group. In contrast, mineral resorption by osteoclasts was signficantly increased. These results indicate that these fetal mouse long bones are a sensitive and useful model to further study the cellular mechanisms involved in the changed mineral metabolism of skeletal tissues under microgravity.

Assessment of bone dysplasia by micro-CT and glycosaminoglycan levels in mouse models for mucopolysaccharidosis type I, IIIA, IVA, and VII

PubMed Central

Rowan, Daniel J.; Tomatsu, Shunji; Grubb, Jeffrey H.; Montaño, Adriana M.; Sly, William S.

2012-01-01

Summary Mucopolysaccharidoses (MPS) are a group of lysosomal storage diseases caused by mutations in lysosomal enzymes involved in degradation of glycosaminoglycans (GAGs). Patients with MPS grow poorly and become physically disabled due to systemic bone disease. While many of the major skeletal effects in mouse models for MPS have been described, no detailed analysis that compares GAGs levels and characteristics of bone by micro-CT has been done. The aims of this study were to assess severity of bone dysplasia among four MPS mouse models (MPS I, IIIA, IVA and VII), to determine the relationship between severity of bone dysplasia and serum keratan sulfate (KS) and heparan sulfate (HS) levels in those models, and to explore the mechanism of KS elevation in MPS I, IIIA, and VII mouse models. Clinically, MPS VII mice had the most severe bone pathology; however, MPS I and IVA mice also showed skeletal pathology. MPS I and VII mice showed severe bone dysplasia, higher bone mineral density, narrowed spinal canal, and shorter sclerotic bones by micro-CT and radiographs. Serum KS and HS levels were elevated in MPS I, IIIA, and VII mice. Severity of skeletal disease displayed by micro-CT, radiographs and histopathology correlated with the level of KS elevation. We showed that elevated HS levels in MPS mouse models could inhibit N-acetylgalactosamine-6-sulfate sulfatase enzyme. These studies suggest that KS could be released from chondrocytes affected by accumulation of other GAGs and that KS could be useful as a biomarker for severity of bone dysplasia in MPS disorders. PMID:22971960

Polarization in Raman spectroscopy helps explain bone brittleness in genetic mouse models

NASA Astrophysics Data System (ADS)

Makowski, Alexander J.; Pence, Isaac J.; Uppuganti, Sasidhar; Zein-Sabatto, Ahbid; Huszagh, Meredith C.; Mahadevan-Jansen, Anita; Nyman, Jeffry S.

2014-11-01

Raman spectroscopy (RS) has been extensively used to characterize bone composition. However, the link between bone biomechanics and RS measures is not well established. Here, we leveraged the sensitivity of RS polarization to organization, thereby assessing whether RS can explain differences in bone toughness in genetic mouse models for which traditional RS peak ratios are not informative. In the selected mutant mice-activating transcription factor 4 (ATF4) or matrix metalloproteinase 9 (MMP9) knock-outs-toughness is reduced but differences in bone strength do not exist between knock-out and corresponding wild-type controls. To incorporate differences in the RS of bone occurring at peak shoulders, a multivariate approach was used. Full spectrum principal components analysis of two paired, orthogonal bone orientations (relative to laser polarization) improved genotype classification and correlation to bone toughness when compared to traditional peak ratios. When applied to femurs from wild-type mice at 8 and 20 weeks of age, the principal components of orthogonal bone orientations improved age classification but not the explanation of the maturation-related increase in strength. Overall, increasing polarization information by collecting spectra from two bone orientations improves the ability of multivariate RS to explain variance in bone toughness, likely due to polarization sensitivity to organizational changes in both mineral and collagen.

Diagnosing and discriminating between primary and secondary aneurysmal bone cysts

PubMed Central

Sasaki, Hiromi; Nagano, Satoshi; Shimada, Hirofumi; Yokouchi, Masahiro; Setoguchi, Takao; Ishidou, Yasuhiro; Kunigou, Osamu; Maehara, Kosuke; Komiya, Setsuro

2017-01-01

Aneurysmal bone cysts (ABCs) are benign bony lesions frequently accompanied by multiple cystic lesions and aggressive bone destruction. They are relatively rare lesions, representing only 1% of bone tumors. The pathogenesis of ABCs has yet to be elucidated. In the present study, a series of 22 cases of primary and secondary ABC from patients treated in Department of Orthopedic Surgery, Kagoshima University Hospital (Kagoshima, Japan) from 2001–2015 were retrospectively analyzed. The average age at the time of diagnosis of primary ABC was 17.9 years. Intralesional curettage and artificial bone grafting were performed in the majority of the patients with primary ABC. The local recurrence rate following curettage for primary ABC was 18%, and the cause of local recurrence was considered to be insufficient curettage. Although no adjuvant therapy was administered during the surgeries, it may assist the prevention of local recurrence in certain cases. The cases of secondary ABC were preceded by benign bone tumors, including fibrous dysplasia, giant cell tumors, chondroblastoma and non-ossifying fibroma. The features of the secondary ABC typically reflected those of the preceding bone tumor. In the majority of cases, distinguishing the primary ABC from the secondary ABC was possible based on characteristic features, including age of the patient at diagnosis and the tumor location. In cases that exhibit ambiguous features, including a soft tissue mass or a thick septal enhancement on the preoperative magnetic resonance images, a biopsy must be obtained in order to exclude other types of aggressive bone tumors, including giant cell tumor, osteosarcoma and telangiectatic osteosarcoma. PMID:28454393

Neogambogic Acid Suppresses Receptor Activator of Nuclear Factor κB Ligand (RANKL)-Induced Osteoclastogenesis by Inhibiting the JNK and NF-κB Pathways in Mouse Bone Marrow-Derived Monocyte/Macrophages.

PubMed

Jin, Gu; Wang, Fang-Fang; Li, Tao; Jia, Dong-Dong; Shen, Yong; Xu, Hai-Chao

2018-04-26

BACKGROUND Neogambogic acid (NGA) is used in traditional Chinese medicine. The aim of this study was to investigate the effects of NGA on gene signaling pathways involved in osteoclastogenesis in mouse bone marrow-derived monocyte/macrophages (BMMs) and on bone resorption in vitro. MATERIAL AND METHODS Primary mouse BMMs were cultured with increasing concentrations of NGA. Real-time polymerase chain reaction was used to study the expression of mRNAs corresponding to gene products specific to receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation, including tartrate-resistant acid phosphatase (TRAP), calcitonin receptor (CTR), cathepsin K (CTSK), and nuclear factor of activated T cells c1 (NFATc1). A cell counting kit-8 assay was used to evaluate cell proliferation. Western blotting and confocal immunofluorescence microscopy were used to investigate the signaling pathways. A bone resorption model was used to quantify bone resorption. RESULTS An NGA dose of ≤0.4 μg/ml had no significant effect on the proliferation of mouse BMMs in vitro (P>0.05); concentrations of between 0.1-0.4 μg/ml significantly inhibited RANKL-induced osteoclastogenesis (P<0.01) in a dose-dependent manner. Compared with the control group, NGA significantly reduced RANKL-induced bone resorption in vitro (P <0.01), and downregulated the expression of osteoclast-related mRNAs of TRAP, CTR, CTSK, and NFATc1. NGA suppressed the activation of JNK but not the p38 signaling pathway and significantly reduced NF-κB p65 phosphorylation and the nuclear transport of NF-κB molecules, which inhibited NFATc1 expression. CONCLUSIONS NGA suppressed RANKL-induced osteoclastogenesis by inhibiting the JNK and NF-κB pathways in mouse BMMs in vitro and reduced osteoclastic bone resorption.

Primary Ewing's Sarcoma of the temporal bone in an infant.

PubMed

Goudarzipour, Kourosh; Shamsian, Shahin; Alavi, Samin; Nourbakhsh, Kazem; Aghakhani, Roxana; Eydian, Zahra; Arzanian, Mohammad Taghi

2015-04-01

Introduction : Ewing's sarcoma is the second most common primary malignant tumor of bone found in children after Osteosarcoma. It accounts for 4-9% of primary malignant bone tumors and it affects bones of the skull or face in only 1-4% of cases. Hence it rarely affects the head and neck. Subject and Method : In this case report, we describe a case of primary Ewing's sarcoma occurring in the temporal bone. The tumor was surgically excised, and the patient underwent chemotherapy for ten months. Results : Neither recurrence nor distant metastasis was noted in these 10 months after surgery but about 18 months after surgery our patient was expired. Conclusion : Although the prognosis of Ewing's sarcoma is generally poor because of early metastasis to the lungs and to other bones, a review of the article suggested that Ewing's sarcoma occurring in the skull can often be successfully managed by intensive therapy with radical excision and chemotherapy. This result was supported by the case reported here.

Hajdu Cheney Mouse Mutants Exhibit Osteopenia, Increased Osteoclastogenesis, and Bone Resorption.

PubMed

Canalis, Ernesto; Schilling, Lauren; Yee, Siu-Pok; Lee, Sun-Kyeong; Zanotti, Stefano

2016-01-22

Notch receptors are determinants of cell fate and function and play a central role in skeletal development and bone remodeling. Hajdu Cheney syndrome, a disease characterized by osteoporosis and fractures, is associated with NOTCH2 mutations resulting in a truncated stable protein and gain-of-function. We created a mouse model reproducing the Hajdu Cheney syndrome by introducing a 6955C→T mutation in the Notch2 locus leading to a Q2319X change at the amino acid level. Notch2(Q2319X) heterozygous mutants were smaller and had shorter femurs than controls; and at 1 month of age they exhibited cancellous and cortical bone osteopenia. As the mice matured, cancellous bone volume was restored partially in male but not female mice, whereas cortical osteopenia persisted in both sexes. Cancellous bone histomorphometry revealed an increased number of osteoclasts and bone resorption, without a decrease in osteoblast number or bone formation. Osteoblast differentiation and function were not affected in Notch2(Q2319X) cells. The pre-osteoclast cell pool, osteoclast differentiation, and bone resorption in response to receptor activator of nuclear factor κB ligand in vitro were increased in Notch2(Q2319X) mutants. These effects were suppressed by the γ-secretase inhibitor LY450139. In conclusion, Notch2(Q2319X) mice exhibit cancellous and cortical bone osteopenia, enhanced osteoclastogenesis, and increased bone resorption. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Bone sialoprotein, but not osteopontin, deficiency impairs the mineralization of regenerating bone during cortical defect healing.

PubMed

Monfoulet, Laurent; Malaval, Luc; Aubin, Jane E; Rittling, Susan R; Gadeau, Alain P; Fricain, Jean-Christophe; Chassande, Olivier

2010-02-01

Bone healing is a complex multi-step process, which depends on the position and size of the lesion, and on the mechanical stability of the wounded area. To address more specifically the mechanisms involved in cortical bone healing, we created drill-hole defects in the cortex of mouse femur, a lesion that triggers intramembranous repair, and compared the roles of bone sialoprotein (BSP) and osteopontin (OPN), two proteins of the extracellular matrix, in the repair process. Bone regeneration was analyzed by ex vivo microcomputerized X-ray tomography and histomorphometry of bones of BSP-deficient, OPN-deficient and wild-type mice. In all mouse strains, the cortical gap was bridged with woven bone within 2 weeks and no mineralized tissue was observed in the marrow. Within 3 weeks, lamellar cortical bone filled the gap. The amount and degree of mineralization of the woven bone was not affected by OPN deficiency, but cortical bone healing was delayed in BSP-deficient mice due to delayed mineralization. Gene expression studies showed a higher amount of BSP transcripts in the repair bone of OPN-deficient mice, suggesting a possible compensation of OPN function by BSP in OPN-null mice. Our data suggest that BSP, but not OPN, plays a role in primary bone formation and mineralization of newly formed bone during the process of cortical bone healing. (c) 2009 Elsevier Inc. All rights reserved.

PULSED FOCUSED ULTRASOUND TREATMENT OF MUSCLE MITIGATES PARALYSIS-INDUCED BONE LOSS IN THE ADJACENT BONE: A STUDY IN A MOUSE MODEL

PubMed Central

Poliachik, Sandra L.; Khokhlova, Tatiana D.; Wang, Yak-Nam; Simon, Julianna C.; Bailey, Michael R.

2015-01-01

Bone loss can result from bed rest, space flight, spinal cord injury or age-related hormonal changes. Current bone loss mitigation techniques include pharmaceutical interventions, exercise, pulsed ultrasound targeted to bone and whole body vibration. In this study, we attempted to mitigate paralysis-induced bone loss by applying focused ultrasound to the midbelly of a paralyzed muscle. We employed a mouse model of disuse that uses onabotulinumtoxinA-induced paralysis, which causes rapid bone loss in 5 d. A focused 2 MHz transducer applied pulsed exposures with pulse repetition frequency mimicking that of motor neuron firing during walking (80 Hz), standing (20 Hz), or the standard pulsed ultrasound frequency used in fracture healing (1 kHz). Exposures were applied daily to calf muscle for 4 consecutive d. Trabecular bone changes were characterized using micro-computed tomography. Our results indicated that application of certain focused pulsed ultrasound parameters was able to mitigate some of the paralysis-induced bone loss. PMID:24857416

Primary pericranial Ewing's sarcoma on the temporal bone: A case report.

PubMed

Kawano, Hiroto; Nitta, Naoki; Ishida, Mitsuaki; Fukami, Tadateru; Nozaki, Kazuhiko

2016-01-01

Primary Ewing's sarcoma originating in the pericranium is an extremely rare disease entity. A 9-year-old female patient was admitted to our department due to a left temporal subcutaneous mass. The mass was localized under the left temporal muscle and attached to the surface of the temporal bone. Head computed tomography revealed a mass with bony spicule formation on the temporal bone, however, it did not show bone destruction or intracranial invasion. F-18 fluorodeoxyglucose positron emission tomography showed no lesions other than the mass on the temporal bone. Magnetic resonance imaging showed that the mass was located between the temporal bone and the pericranium. The mass was completely resected with the underlying temporal bone and the overlying deep layer of temporal muscle, and was diagnosed as primary Ewing's sarcoma. Because the tumor was located in the subpericranium, we created a new classification, "pericranial Ewing's sarcoma," and diagnosed the present tumor as pericranial Ewing's sarcoma. We herein present an extremely rare case of primary pericranial Ewing's sarcoma that developed on the temporal bone.

The effects of cortical bone thickness and trabecular bone strength on noninvasive measures of the implant primary stability using synthetic bone models.

PubMed

Hsu, Jui-Ting; Fuh, Lih-Jyh; Tu, Ming-Gene; Li, Yu-Fen; Chen, Kuan-Ting; Huang, Heng-Li

2013-04-01

This study investigated how the primary stability of a dental implant as measured by the insertion torque value (ITV), Periotest value (PTV), and implant stability quotient (ISQ) is affected by varying thicknesses of cortical bone and strengths of trabecular bone using synthetic bone models. Four synthetic cortical shells (with thicknesses of 0, 1, 2, and 3 mm) were attached to four cellular rigid polyurethane foams (with elastic moduli of 137, 47.5, 23, and 12.4 MPa) and one open-cell rigid polyurethane foam which mimic the osteoporotic bone (with an elastic modulus 6.5 MPa), to represent the jawbones with various cortical bone thicknesses and strengths of trabecular bone. A total of 60 bone specimens accompanied with implants was examined by a torque meter, Osstell resonance frequency analyzer, and Periotest electronic device. All data were statistically analyzed by two-way analysis of variance. In addition, second-order nonlinear regression was utilized to assess the correlations of the primary implant stability with the four cortex thicknesses and five strengths of trabecular bone. ITV, ISQ, and PTV differed significantly (p < .05) and were strongly correlated with the thickness of cortical bone (R(2) > 0.9) and the elastic modulus of trabecular bone (R(2) = 0.74-0.99). The initial stability at the time of implant placement is influenced by both the cortical bone thickness and the strength of trabecular bone; however, these factors are mostly nonlinearly correlated with ITV, PTV, and ISQ. Using ITV and PTV seems more suitable for identifying the primary implant stability in osteoporotic bone with a thin cortex. © 2011 Wiley Periodicals, Inc.

Mobilization of bone marrow mesenchymal stem cells in vivo augments bone healing in a mouse model of segmental bone defect.

PubMed

Kumar, Sanjay; Ponnazhagan, Selvarangan

2012-04-01

Although the number of mesenchymal stem cells (MSC) in the bone marrow is sufficient to maintain skeletal homeostasis, in osteopenic pathology, aggravated osteoclast activity or insufficient osteoblast numbers ensue, affecting normal bone remodeling. Most of the currently available therapies are anti-resorptive with limited osteogenic potential. Since mobilization of stem/progenitors from the BM is a prerequisite for their participation in tissue repair, amplification of endogenous stem cells may provide an alternative approach in these conditions. The present study determined the potential of MSC mobilization in vivo, using combinations of different growth factors with the CXCR4 antagonist, AMD3100, in a mouse model of segmental bone defect. Results indicated that among several factors tested IGF1 had maximum proliferative ability of MSC in vitro. Results of the in vivo studies indicated that the combination of IGF1 and AMD3100 provided significant augmentation of bone growth as determined by DXA, micro-CT and histomorphometry in mice bearing segmental fractures. Further, characterization of MSC isolated from mice treated with IGF1 and AMD3100 indicated Akt/PI3K, MEK1/2-Erk1/2 and smad2/3 as key signaling pathways mediating this effect. These data indicate the potential of in vivo stem cell mobilization as a novel alternative for bone healing. Copyright © 2012 Elsevier Inc. All rights reserved.

Defective Endochondral Ossification-Derived Matrix and Bone Cells Alter the Lymphopoietic Niche in Collagen X Mouse Models

PubMed Central

Sweeney, Elizabeth; Roberts, Douglas; Lin, Angela; Guldberg, Robert

2013-01-01

Despite the appreciated interdependence of skeletal and hematopoietic development, the cell and matrix components of the hematopoietic niche remain to be fully defined. Utilizing mice with disrupted function of collagen X (ColX), a major hypertrophic cartilage matrix protein associated with endochondral ossification, our data identified a cytokine defect in trabecular bone cells at the chondro-osseous hematopoietic niche as a cause for aberrant B lymphopoiesis in these mice. Specifically, analysis of ColX transgenic and null mouse chondro-osseous regions via micro-computed tomography revealed an altered trabecular bone environment. Additionally, cocultures with hematopoietic and chondro-osseous cell types highlighted impaired hematopoietic support by ColX transgenic and null mouse derived trabecular bone cells. Further, cytokine arrays with conditioned media from the trabecular osteoblast cocultures suggested an aberrant hematopoietic cytokine milieu within the chondro-osseous niche of the ColX deficient mice. Accordingly, B lymphopoiesis was rescued in the ColX mouse derived trabecular osteoblast cocultures with interlukin-7, stem cell factor, and stromal derived factor-1 supplementation. Moreover, B cell development was restored in vivo after injections of interlukin-7. These data support our hypothesis that endrochondrally-derived trabecular bone cells and matrix constituents provide cytokine-rich niches for hematopoiesis. Furthermore, this study contributes to the emerging concept that niche defects may underlie certain immuno-osseous and hematopoietic disorders. PMID:23656481

Defective endochondral ossification-derived matrix and bone cells alter the lymphopoietic niche in collagen X mouse models.

PubMed

Sweeney, Elizabeth; Roberts, Douglas; Lin, Angela; Guldberg, Robert; Jacenko, Olena

2013-10-01

Despite the appreciated interdependence of skeletal and hematopoietic development, the cell and matrix components of the hematopoietic niche remain to be fully defined. Utilizing mice with disrupted function of collagen X (ColX), a major hypertrophic cartilage matrix protein associated with endochondral ossification, our data identified a cytokine defect in trabecular bone cells at the chondro-osseous hematopoietic niche as a cause for aberrant B lymphopoiesis in these mice. Specifically, analysis of ColX transgenic and null mouse chondro-osseous regions via micro-computed tomography revealed an altered trabecular bone environment. Additionally, cocultures with hematopoietic and chondro-osseous cell types highlighted impaired hematopoietic support by ColX transgenic and null mouse derived trabecular bone cells. Further, cytokine arrays with conditioned media from the trabecular osteoblast cocultures suggested an aberrant hematopoietic cytokine milieu within the chondro-osseous niche of the ColX deficient mice. Accordingly, B lymphopoiesis was rescued in the ColX mouse derived trabecular osteoblast cocultures with interlukin-7, stem cell factor, and stromal derived factor-1 supplementation. Moreover, B cell development was restored in vivo after injections of interlukin-7. These data support our hypothesis that endrochondrally-derived trabecular bone cells and matrix constituents provide cytokine-rich niches for hematopoiesis. Furthermore, this study contributes to the emerging concept that niche defects may underlie certain immuno-osseous and hematopoietic disorders.

Primary Hyperparathyroidism: The Influence of Bone Marrow Adipose Tissue on Bone Loss and of Osteocalcin on Insulin Resistance

PubMed Central

Mendonça, Maira L.; Batista, Sérgio L.; Nogueira-Barbosa, Marcello H.; Salmon, Carlos E.G.; de Paula, Francisco J.A.

2016-01-01

OBJECTIVES: Bone marrow adipose tissue has been associated with low bone mineral density. However, no data exist regarding marrow adipose tissue in primary hyperparathyroidism, a disorder associated with bone loss in conditions of high bone turnover. The objective of the present study was to investigate the relationship between marrow adipose tissue, bone mass and parathyroid hormone. The influence of osteocalcin on the homeostasis model assessment of insulin resistance was also evaluated. METHODS: This was a cross-sectional study conducted at a university hospital, involving 18 patients with primary hyperparathyroidism (PHPT) and 21 controls (CG). Bone mass was assessed by dual-energy x-ray absorptiometry and marrow adipose tissue was assessed by 1H magnetic resonance spectroscopy. The biochemical evaluation included the determination of parathyroid hormone, osteocalcin, glucose and insulin levels. RESULTS: A negative association was found between the bone mass at the 1/3 radius and parathyroid hormone levels (r = -0.69; p<0.01). Marrow adipose tissue was not significantly increased in patients (CG = 32.8±11.2% vs PHPT = 38.6±12%). The serum levels of osteocalcin were higher in patients (CG = 8.6±3.6 ng/mL vs PHPT = 36.5±38.4 ng/mL; p<0.005), but no associations were observed between osteocalcin and insulin or between insulin and both marrow adipose tissue and bone mass. CONCLUSION: These results suggest that the increment of adipogenesis in the bone marrow microenvironment under conditions of high bone turnover due to primary hyperparathyroidism is limited. Despite the increased serum levels of osteocalcin due to primary hyperparathyroidism, these patients tend to have impaired insulin sensitivity. PMID:27626477

The influence of bone graft procedures on primary stability and bone change of implants placed in fresh extraction sockets.

PubMed

Jun, Sang Ho; Park, Chang-Joo; Hwang, Suk-Hyun; Lee, Youn Ki; Zhou, Cong; Jang, Hyon-Seok; Ryu, Jae-Jun

2018-12-01

This study was to evaluate the effect of bone graft procedure on the primary stability of implants installed in fresh sockets and assess the vertical alteration of peri-implant bone radiographically. Twenty-three implants were inserted in 18 patients immediately after tooth extraction. The horizontal gap between the implant and bony walls of the extraction socket was grafted with xenografts. The implant stability before and after graft procedure was measured by Osstell Mentor as implant stability quotient before bone graft (ISQ bbg) and implant stability quotient after bone graft (ISQ abg). Peri-apical radiographs were taken to measure peri-implant bone change immediately after implant surgery and 12 months after implant placement. Data were analyzed by independent t test; the relationships between stability parameters (insertion torque value (ITV), ISQ abg, and ISQ bbg) and peri-implant bone changes were analyzed according to Pearson correlation coefficients. The increase of ISQ in low primary stability group (LPSG) was 6.87 ± 3.62, which was significantly higher than the increase in high primary stability group (HPSG). A significant correlation between ITV and ISQ bbg ( R  = 0.606, P  = 0.002) was found; however, age and peri-implant bone change were not found significantly related to implant stability parameters. It was presented that there were no significant peri-implant bone changes at 1 year after bone graft surgery. Bone graft procedure is beneficial for increasing the primary stability of immediately placed implants, especially when the ISQ of implants is below 65 and that bone grafts have some effects on peri-implant bone maintenance.

Systemic Mesenchymal Stromal Cell Transplantation Prevents Functional Bone Loss in a Mouse Model of Age-Related Osteoporosis.

PubMed

Kiernan, Jeffrey; Hu, Sally; Grynpas, Marc D; Davies, John E; Stanford, William L

2016-05-01

Age-related osteoporosis is driven by defects in the tissue-resident mesenchymal stromal cells (MSCs), a heterogeneous population of musculoskeletal progenitors that includes skeletal stem cells. MSC decline leads to reduced bone formation, causing loss of bone volume and the breakdown of bony microarchitecture crucial to trabecular strength. Furthermore, the low-turnover state precipitated by MSC loss leads to low-quality bone that is unable to perform remodeling-mediated maintenance--replacing old damaged bone with new healthy tissue. Using minimally expanded exogenous MSCs injected systemically into a mouse model of human age-related osteoporosis, we show long-term engraftment and markedly increased bone formation. This led to improved bone quality and turnover and, importantly, sustained microarchitectural competence. These data establish proof of concept that MSC transplantation may be used to prevent or treat human age-related osteoporosis. This study shows that a single dose of minimally expanded mesenchymal stromal cells (MSCs) injected systemically into a mouse model of human age-related osteoporosis display long-term engraftment and prevent the decline in bone formation, bone quality, and microarchitectural competence. This work adds to a growing body of evidence suggesting that the decline of MSCs associated with age-related osteoporosis is a major transformative event in the progression of the disease. Furthermore, it establishes proof of concept that MSC transplantation may be a viable therapeutic strategy to treat or prevent human age-related osteoporosis. ©AlphaMed Press.

Systemic Mesenchymal Stromal Cell Transplantation Prevents Functional Bone Loss in a Mouse Model of Age-Related Osteoporosis

PubMed Central

Kiernan, Jeffrey; Hu, Sally; Grynpas, Marc D.

2016-01-01

Age-related osteoporosis is driven by defects in the tissue-resident mesenchymal stromal cells (MSCs), a heterogeneous population of musculoskeletal progenitors that includes skeletal stem cells. MSC decline leads to reduced bone formation, causing loss of bone volume and the breakdown of bony microarchitecture crucial to trabecular strength. Furthermore, the low-turnover state precipitated by MSC loss leads to low-quality bone that is unable to perform remodeling-mediated maintenance—replacing old damaged bone with new healthy tissue. Using minimally expanded exogenous MSCs injected systemically into a mouse model of human age-related osteoporosis, we show long-term engraftment and markedly increased bone formation. This led to improved bone quality and turnover and, importantly, sustained microarchitectural competence. These data establish proof of concept that MSC transplantation may be used to prevent or treat human age-related osteoporosis. Significance This study shows that a single dose of minimally expanded mesenchymal stromal cells (MSCs) injected systemically into a mouse model of human age-related osteoporosis display long-term engraftment and prevent the decline in bone formation, bone quality, and microarchitectural competence. This work adds to a growing body of evidence suggesting that the decline of MSCs associated with age-related osteoporosis is a major transformative event in the progression of the disease. Furthermore, it establishes proof of concept that MSC transplantation may be a viable therapeutic strategy to treat or prevent human age-related osteoporosis. PMID:26987353

Neuronal Representation of Ultraviolet Visual Stimuli in Mouse Primary Visual Cortex

PubMed Central

Tan, Zhongchao; Sun, Wenzhi; Chen, Tsai-Wen; Kim, Douglas; Ji, Na

2015-01-01

The mouse has become an important model for understanding the neural basis of visual perception. Although it has long been known that mouse lens transmits ultraviolet (UV) light and mouse opsins have absorption in the UV band, little is known about how UV visual information is processed in the mouse brain. Using a custom UV stimulation system and in vivo calcium imaging, we characterized the feature selectivity of layer 2/3 neurons in mouse primary visual cortex (V1). In adult mice, a comparable percentage of the neuronal population responds to UV and visible stimuli, with similar pattern selectivity and receptive field properties. In young mice, the orientation selectivity for UV stimuli increased steadily during development, but not direction selectivity. Our results suggest that, by expanding the spectral window through which the mouse can acquire visual information, UV sensitivity provides an important component for mouse vision. PMID:26219604

Positive effects of bisphosphonates on bone and muscle in a mouse model of Duchenne muscular dystrophy.

PubMed

Yoon, Sung-Hee; Sugamori, Kim S; Grynpas, Marc D; Mitchell, Jane

2016-01-01

Patients with Duchenne muscular dystrophy are at increased risk of decreased bone mineral density and bone fracture as a result of inactivity. To determine if antiresorptive bisphosphonates could improve bone quality and their effects on muscle we studied the Mdx mouse, treated with pamidronate during peak bone growth at 5 and 6 weeks of age, and examined the outcome at 13 weeks of age. Pamidronate increased cortical bone architecture and strength in femurs with increased resistance to fracture. While overall long bone growth was not affected by pamidronate, there was significant inhibition of remodeling in metaphyseal trabecular bone with evidence of residual calcified cartilage. Pamidronate treatment had positive effects on skeletal muscle in the Mdx mice with decreased serum and muscle creatine kinase and evidence of improved muscle histology and grip strength. Copyright © 2015 Elsevier B.V. All rights reserved.

Effects of Loading Duration and Short Rest Insertion on Cancellous and Cortical Bone Adaptation in the Mouse Tibia

PubMed Central

Yang, Haisheng; Embry, Rachel E.; Main, Russell P.

2017-01-01

The skeleton’s osteogenic response to mechanical loading can be affected by loading duration and rest insertion during a series of loading events. Prior animal loading studies have shown that the cortical bone response saturates quickly and short rest insertions between load cycles can enhance cortical bone formation. However, it remains unknown how loading duration and short rest insertion affect load-induced osteogenesis in the mouse tibial compressive loading model, and particularly in cancellous bone. To address this issue, we applied cyclic loading (-9 N peak load; 4 Hz) to the tibiae of three groups of 16 week-old female C57BL/6 mice for two weeks, with a different number of continuous load cycles applied daily to each group (36, 216 and 1200). A fourth group was loaded under 216 daily load cycles with a 10 s rest insertion after every fourth cycle. We found that as few as 36 load cycles per day were able to induce osteogenic responses in both cancellous and cortical bone. Furthermore, while cortical bone area and thickness continued to increase through 1200 cycles, the incremental increase in the osteogenic response decreased as load number increased, indicating a reduced benefit of the increasing number of load cycles. In the proximal metaphyseal cancellous bone, trabecular thickness increased with load up to 216 cycles. We also found that insertion of a 10 s rest between load cycles did not improve the osteogenic response of the cortical or cancellous tissues compared to continuous loading in this model given the age and sex of the mice and the loading parameters used here. These results suggest that relatively few load cycles (e.g. 36) are sufficient to induce osteogenic responses in both cortical and cancellous bone in the mouse tibial loading model. Mechanistic studies using the mouse tibial loading model to examine bone formation and skeletal mechanobiology could be accomplished with relatively few load cycles. PMID:28076363

Raman spectroscopy detects deterioration in biomechanical properties of bone in a glucocorticoid-treated mouse model of rheumatoid arthritis

NASA Astrophysics Data System (ADS)

Maher, Jason R.; Takahata, Masahiko; Awad, Hani A.; Berger, Andrew J.

2011-08-01

Although glucocorticoids are frequently prescribed for the symptomatic management of inflammatory disorders such as rheumatoid arthritis, extended glucocorticoid exposure is the leading cause of physician-induced osteoporosis and leaves patients at a high risk of fracture. To study the biochemical effects of glucocorticoid exposure and how they might affect biomechanical properties of the bone, Raman spectra were acquired from ex vivo tibiae of glucocorticoid- and placebo-treated wild-type mice and a transgenic mouse model of rheumatoid arthritis. Statistically significant spectral differences were observed due to both treatment regimen and mouse genotype. These differences are attributed to changes in the overall bone mineral composition, as well as the degree of phosphate mineralization in tibial cortical bone. In addition, partial least squares regression was used to generate a Raman-based prediction of each tibia's biomechanical strength as quantified by a torsion test. The Raman-based predictions were as accurate as those produced by microcomputed tomography derived parameters, and more accurate than the clinically-used parameter of bone mineral density. These results suggest that Raman spectroscopy could be a valuable tool for monitoring bone biochemistry in studies of bone diseases such as osteoporosis, including tests of drugs being developed to combat these diseases.

A rare case of primary bone lymphoma mimicking a pelvic abscess

PubMed Central

Al Wattar, BH; Mohanty, K

2011-01-01

Primary bone lymphoma (PBL) is a rare, malignant, neoplastic disorder of the skeleton that accounts for less than 5% of all primary bone tumours. We present an extremely rare case of PBL mimicking a pelvic abscess around the sacroiliac joint, which has never been reported in the medical literature, and discuss learning points highlighted from this case. PMID:22004625

Establishing Biomechanical Mechanisms in Mouse Models: Practical Guidelines for Systematically Evaluating Phenotypic Changes in the Diaphyses of Long Bones

PubMed Central

Jepsen, Karl J; Silva, Matthew J; Vashishth, Deepak; Guo, X Edward; van der Meulen, Marjolein CH

2016-01-01

Mice are widely used in studies of skeletal biology, and assessment of their bones by mechanical testing is a critical step when evaluating the functional effects of an experimental perturbation. For example, a gene knockout may target a pathway important in bone formation and result in a “low bone mass” phenotype. But how well does the skeleton bear functional loads; eg, how much do bones deform during loading and how resistant are bones to fracture? By systematic evaluation of bone morphological, densitometric, and mechanical properties, investigators can establish the “biomechanical mechanisms” whereby an experimental perturbation alters whole-bone mechanical function. The goal of this review is to clarify these biomechanical mechanisms and to make recommendations for systematically evaluating phenotypic changes in mouse bones, with a focus on long-bone diaphyses and cortical bone. Further, minimum reportable standards for testing conditions and outcome variables are suggested that will improve the comparison of data across studies. Basic biomechanical principles are reviewed, followed by a description of the cross-sectional morphological properties that best inform the net cellular effects of a given experimental perturbation and are most relevant to biomechanical function. Although morphology is critical, whole-bone mechanical properties can only be determined accurately by a mechanical test. The functional importance of stiffness, maximum load, postyield displacement, and work-to-fracture are reviewed. Because bone and body size are often strongly related, strategies to adjust whole-bone properties for body mass are detailed. Finally, a comprehensive framework is presented using real data, and several examples from the literature are reviewed to illustrate how to synthesize morphological, tissue-level, and whole-bone mechanical properties of mouse long bones. PMID:25917136

R-Spondin 1 promotes vibration-induced bone formation in mouse models of osteoporosis

PubMed Central

Wang, Haitao; Brennan, Tracy A.; Russell, Elizabeth; Kim, Jung-Hoon; Egan, Kevin P.; Chen, Qijun; Israelite, Craig; Schultz, David C.; Johnson, Frederick B.; Pignolo, Robert J.

2013-01-01

Bone tissue adapts to its functional environment by optimizing its morphology for mechanical demand. Among the mechanosensitive cells that recognize and respond to forces in the skeleton are osteocytes, osteoblasts, and mesenchymal progenitor cells (MPCs). Therefore, the ability to use mechanical signals to improve bone health through exercise and devices that deliver mechanical signals is an attractive approach to age-related bone loss; however, the extracellular or circulating mediators of such signals are largely unknown. Using SDS-PAGE separation of proteins secreted by MPCs in response to low magnitude mechanical signals and in-gel trypsin digestion followed by HPLC and mass spectroscopy, we identified secreted proteins up-regulated by vibratory stimulation. We exploited a cell senescence-associated secretory phenotype screen, and reasoned that a subset of vibration-induced proteins with diminished secretion by senescent MPCs will have the capacity to promote bone formation in vivo. We identified one such vibration-induced bone-enhancing (vibe) gene as R-Spondin 1, a Wnt pathway modulator, and demonstrated that it has the capacity to promote bone formation in three mouse models of age-related bone loss. By virtue of their secretory status, some vibe proteins may be candidates for pre-clinical development as anabolic agents for the treatment of osteoporosis. PMID:23974989

Bone Marrow Stromal Cells Contribute to Bone Formation Following Infusion into Femoral Cavities of a Mouse Model of Osteogenesis Imperfecta

PubMed Central

Li, Feng; Wang, Xujun; Niyibizi, Christopher

2010-01-01

Currently, there are conflicting data in literature regarding contribution of bone marrow stromal cells (BMSCs) to bone formation when the cells are systemically delivered in recipient animals. To understand if BMSCs contribute to bone cell phenotype and bone formation in osteogenesis imperfecta bones (OI), MSCs marked with GFP were directly infused into the femurs of a mouse model of OI (oim). The contribution of the cells to the cell phenotype and bone formation was assessed by histology, immunohistochemistry and biomechanical loading of recipient bones. Two weeks following infusion of BMSCs, histological examination of the recipient femurs demonstrated presence of new bone when compared to femurs injected with saline which showed little or no bone formation. The new bone contained few donor cells as demonstrated by GFP fluorescence. At six weeks following cell injection, new bone was still detectable in the recipient femurs but was enhanced by injection of the cells suspended in pepsin solublized type I collagen. Immunofluorescence and immunohistochemical staining showed that donor GFP positive cells in the new bone were localized with osteocalcin expressing cells suggesting that the cells differentiated into osteoblasts in vivo. Biomechanical loading to failure in thee point bending, revealed that, femurs infused with BMSCs in PBS or in soluble type I collagen were biomechanically stronger than those injected with PBS or type I collagen alone. Taken together, the results indicate that transplanted cells differentiated into osteoblasts in vivo and contributed to bone formation in vivo; we also speculate that donor cells induced differentiation or recruitment of endogenous cells to initiate reparative process at early stages following transplantation. PMID:20570757

Glenoid bone grafting in primary reverse total shoulder arthroplasty.

PubMed

Ernstbrunner, Lukas; Werthel, Jean-David; Wagner, Eric; Hatta, Taku; Sperling, John W; Cofield, Robert H

2017-08-01

Severe glenoid bone loss remains a challenge in patients requiring shoulder arthroplasty and may necessitate glenoid bone grafting. The purpose of this study was to determine results, complications, and rates of failure of glenoid bone grafting in primary reverse shoulder arthroplasty. Forty-one shoulders that underwent primary reverse arthroplasty between 2006 and 2013 with a minimum follow-up of 2 years (mean, 2.8 years; range, 2-6 years) were reviewed. Thirty-four (83%) received corticocancellous grafts and 7 (17%) structural grafts. Active range of motion and pain levels were significantly improved (P < .001), with mean American Shoulder and Elbow Surgeons score of 77, Simple Shoulder Test score of 9, and patient satisfaction of 93% at the most recent follow-up. Preoperative severe glenoid erosion and increasing body mass index were significantly associated with worse American Shoulder and Elbow Surgeons scores (P = .04). On radiographic evaluation, 7 patients (18%) had grade 1 or grade 2 glenoid lucency. Glenoid bone graft incorporation was observed in 31 patients (78%). Twelve patients (30%) suffered from grade 1 or grade 2 scapular notching. All of the patients with structural grafts showed graft incorporation and no signs of glenoid lucency. Although glenoid lucency, glenoid graft resorption, and scapular notching were present at short-term to midterm follow-up, none of the patients needed revision surgery. Primary reverse shoulder arthroplasty with glenoid reconstruction using bone graft relieved pain and restored shoulder function and stability. Copyright © 2017 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.

Phenotypic integration among trabecular and cortical bone traits establishes mechanical functionality of inbred mouse vertebrae.

PubMed

Tommasini, Steven M; Hu, Bin; Nadeau, Joseph H; Jepsen, Karl J

2009-04-01

Conventional approaches to identifying quantitative trait loci (QTLs) regulating bone mass and fragility are limited because they examine cortical and trabecular traits independently. Prior work examining long bones from young adult mice and humans indicated that skeletal traits are functionally related and that compensatory interactions among morphological and compositional traits are critical for establishing mechanical function. However, it is not known whether trait covariation (i.e., phenotypic integration) also is important for establishing mechanical function in more complex, corticocancellous structures. Covariation among trabecular, cortical, and compositional bone traits was examined in the context of mechanical functionality for L(4) vertebral bodies across a panel of 16-wk-old female AXB/BXA recombinant inbred (RI) mouse strains. The unique pattern of randomization of the A/J and C57BL/6J (B6) genome among the RI panel provides a powerful tool that can be used to measure the tendency for different traits to covary and to study the biology of complex traits. We tested the hypothesis that genetic variants affecting vertebral size and mass are buffered by changes in the relative amounts of cortical and trabecular bone and overall mineralization. Despite inheriting random sets of A/J and B6 genomes, the RI strains inherited nonrandom sets of cortical and trabecular bone traits. Path analysis, which is a multivariate analysis that shows how multiple traits covary simultaneously when confounding variables like body size are taken into consideration, showed that RI strains that tended to have smaller vertebrae relative to body size achieved mechanical functionality by increasing mineralization and the relative amounts of cortical and trabecular bone. The interdependence among corticocancellous traits in the vertebral body indicated that variation in trabecular bone traits among inbred mouse strains, which is often thought to arise from genetic factors, is also

An investigation of the mineral in ductile and brittle cortical mouse bone.

PubMed

Rodriguez-Florez, Naiara; Garcia-Tunon, Esther; Mukadam, Quresh; Saiz, Eduardo; Oldknow, Karla J; Farquharson, Colin; Millán, José Luis; Boyde, Alan; Shefelbine, Sandra J

2015-05-01

Bone is a strong and tough material composed of apatite mineral, organic matter, and water. Changes in composition and organization of these building blocks affect bone's mechanical integrity. Skeletal disorders often affect bone's mineral phase, either by variations in the collagen or directly altering mineralization. The aim of the current study was to explore the differences in the mineral of brittle and ductile cortical bone at the mineral (nm) and tissue (µm) levels using two mouse phenotypes. Osteogenesis imperfecta model, oim(-/-) , mice have a defect in the collagen, which leads to brittle bone; PHOSPHO1 mutants, Phospho1(-/-) , have ductile bone resulting from altered mineralization. Oim(-/-) and Phospho1(-/-) were compared with their respective wild-type controls. Femora were defatted and ground to powder to measure average mineral crystal size using X-ray diffraction (XRD) and to monitor the bulk mineral to matrix ratio via thermogravimetric analysis (TGA). XRD scans were run after TGA for phase identification to assess the fractions of hydroxyapatite and β-tricalcium phosphate. Tibiae were embedded to measure elastic properties with nanoindentation and the extent of mineralization with backscattered electron microscopy (BSE SEM). Results revealed that although both pathology models had extremely different whole-bone mechanics, they both had smaller apatite crystals, lower bulk mineral to matrix ratio, and showed more thermal conversion to β-tricalcium phosphate than their wild types, indicating deviations from stoichiometric hydroxyapatite in the original mineral. In contrast, the degree of mineralization of bone matrix was different for each strain: brittle oim(-/-) were hypermineralized, whereas ductile Phospho1(-/-) were hypomineralized. Despite differences in the mineralization, nanoscale alterations in the mineral were associated with reduced tissue elastic moduli in both pathologies. Results indicated that alterations from normal crystal size

Early Changes of Articular Cartilage and Subchondral Bone in The DMM Mouse Model of Osteoarthritis.

PubMed

Fang, Hang; Huang, Lisi; Welch, Ian; Norley, Chris; Holdsworth, David W; Beier, Frank; Cai, Daozhang

2018-02-12

To examine the early changes of articular cartilage and subchondral bone in the DMM mouse model of osteoarthritis, mice were subjected to DMM or SHAM surgery and sacrificed at 2-, 5- and 10-week post-surgery. Catwalk gait analyses, Micro-Computed Tomography, Toluidine Blue, Picrosirius Red and Tartrate-Resistant Acid Phosphatase (TRAP) staining were used to investigate gait patterns, joint morphology, subchondral bone, cartilage, collagen organization and osteoclasts activity, respectively. Results showed OA progressed over 10-week time-course. Gait disparity occurred only at 10-week post-surgery. Osteophyte formed at 2-week post-surgery. BMDs of DMM showed no statistical differences comparing to SHAM at 2 weeks, but BV/TV is much higher in DMM mice. Increased BMD was clearly found at 5- and 10-week post-surgery in DMM mice. TRAP staining showed increased osteoclast activity at the site of osteophyte formation of DMM joints at 5- and 10-week time points. These results showed that subchondral bone turnover might occurred earlier than 2 weeks in this mouse DMM model. Gait disparity only occurred at later stage of OA in DMM mice. Notably, patella dislocation could occur in some of the DMM mice and cause a different pattern of OA in affected knee.

Effect of oral calcium and calcium + fluoride treatments on mouse bone properties during suspension

NASA Technical Reports Server (NTRS)

Simske, S. J.; Luttges, M. W.; Allen, K. A.; Spooner, B. S. (Principal Investigator)

1992-01-01

The bone effects of oral dosages of calcium chloride with or without supplementary sodium fluoride were assessed in antiorthostatically suspended mice. Two calcium dosages were used to replace half (3.1 mM) or all(6.3 mM) of the dietary calcium lost due to reduced food intake by the suspended mice. Two groups of 6.3 mM CaCl2-treated mice were additionally treated with 0.25 or 2.5 mM NaF. The results indicate that supplementation of the mouse drinking water with calcium salts prevents bone changes induced by short-term suspension, while calcium salts in combination with fluoride are less effective as fluoride dosage increases. However, the calcium supplements change the relationship between the femur mechanical properties and the mineral composition of the bone. Because of this, it appears that oral calcium supplements are effective through a mechanism other than simple dietary supplementation and may indicate a dependence of bone consistency on systemic and local fluid conditions.

In situ intracellular calcium oscillations in osteocytes in intact mouse long bones under dynamic mechanical loading

PubMed Central

Jing, Da; Baik, Andrew D.; Lu, X. Lucas; Zhou, Bin; Lai, Xiaohan; Wang, Liyun; Luo, Erping; Guo, X. Edward

2014-01-01

Osteocytes have been hypothesized to be the major mechanosensors in bone. How in situ osteocytes respond to mechanical stimuli is still unclear because of technical difficulties. In vitro studies have shown that osteocytes exhibited unique calcium (Ca2+) oscillations to fluid shear. However, whether this mechanotransduction phenomenon holds for in situ osteocytes embedded within a mineralized bone matrix under dynamic loading remains unknown. Using a novel synchronized loading/imaging technique, we successfully visualized in real time and quantified Ca2+ responses in osteocytes and bone surface cells in situ under controlled dynamic loading on intact mouse tibia. The resultant fluid-induced shear stress on the osteocyte in the lacunocanalicular system (LCS) was also quantified. Osteocytes, but not surface cells, displayed repetitive Ca2+ spikes in response to dynamic loading, with spike frequency and magnitude dependent on load magnitude, tissue strain, and shear stress in the LCS. The Ca2+ oscillations were significantly reduced by endoplasmic reticulum (ER) depletion and P2 purinergic receptor (P2R)/phospholipase C (PLC) inhibition. This study provides direct evidence that osteocytes respond to in situ mechanical loading by Ca2+ oscillations, which are dependent on the P2R/PLC/inositol trisphosphate/ER pathway. This study develops a novel approach in skeletal mechanobiology and also advances our fundamental knowledge of bone mechanotransduction.—Jing, D., Baik, A. D., Lu, X. L., Zhou, B., Lai, X., Wang, L., Luo, E., Guo, X. E. In situ intracellular calcium oscillations in osteocytes in intact mouse long bones under dynamic mechanical loading. PMID:24347610

THE EFFECT OF ANTISERUM, ALONE AND WITH HYDROCORTISONE, ON FOETAL MOUSE BONES IN CULTURE

PubMed Central

Fell, Honor B.; Weiss, L.

1965-01-01

1. The effects of normal rabbit serum and of rabbit antiserum to whole foetal mouse tissues, on the isolated limb bones of late foetal mice were studied in organ culture, and the influence of hydrocortisone on these effects was investigated. 2. Unheated normal serum caused slight loss of metachromatic material from the cartilage matrix, and some resorption of both cartilage and bone. 3. In unheated antiserum to foetal mouse tissues, the terminal cartilage was smaller and less metachromatic than in paired controls in normal serum, while osteoclasis was so intense that in many explants the bone had almost disappeared. The amount of necrosis varied with different batches of antiserum. 4. The changes produced by normal serum and antiserum could be largely prevented by heating the sera to 57°C for 45 minutes. 5. The effects could also be inhibited by the addition of hydrocortisone to the unheated sera; as little as 0.1 µg hydrocortisone per ml of medium had a well marked protective action. 6. It is suggested that (a) unheated antiserum causes a release of lysosomal enzymes with consequent breakdown of intercellular material, (b) this release is due to an indirect action on the lysosome via an increased permeability of the cell membrane, (c) hydrocortisone does not affect the antigen-antibody reaction, but inhibits the autolytic changes that normally follow this reaction, possibly by stabilising both the lysosomal and cell membranes. PMID:14276776

Mouse bone marrow-derived mesenchymal stem cells inhibit leukemia/lymphoma cell proliferation in vitro and in a mouse model of allogeneic bone marrow transplant

PubMed Central

SONG, NINGXIA; GAO, LEI; QIU, HUIYING; HUANG, CHONGMEI; CHENG, HUI; ZHOU, HONG; LV, SHUQING; CHEN, LI; WANG, JIANMIN

2015-01-01

The allogeneic hematopoietic stem cell (HSC) transplantation of mesenchymal stem cells (MSCs) contributes to the reconstitution of hematopoiesis by ameliorating acute graft-versus-host disease (aGVHD). However, the role of MSCs in graft-versus-leukemia remains to be determined. In the present study, we co-cultured C57BL/6 mouse bone marrow (BM)-derived MSCs with A20 murine B lymphoma, FBL3 murine erythroleukemia and P388 murine acute lymphocytic leukemia cells. Cell proliferation, apoptosis, cell cycle progression and the amount of cytokine secretion were then measured using a Cell Counting kit-8, Annexin V/propidium iodide staining, flow cytometry and ELISA, respectively. We also established a model of allogeneic bone marrow transplantation (BMT) using BALB/c mice. Following the administration of A20 cells and MSCs, we recorded the symptoms and the survival of the mice for 4 weeks, assessed the T cell subsets present in peripheral blood, and, after the mice were sacrifice, we determined the infiltration of MSCs into the organs by histological staining. Our results revealed that the MSCs inhibited the proliferation of the mouse lymphoma and leukemia cells in vitro, leading to cell cycle arrest and reducing the secretion of interleukin (IL)-10. In our model of allogeneic BMT, the intravenous injection of MSCs into the mice injected wth A20 cells decreased the incidence of lymphoma, improved survival, increased the fraction of CD3+CD8+ T cells, decreased the fraction of CD3+CD4+ T cells and CD4+CD25+ T cells in peripheral blood, and ameliorated the manifestation of aGVHD. The results from the present study indicate that MSCs may be safe and effective when used in allogeneic BMT for the treatment of hemotological malignancies. PMID:25901937

Mouse bone marrow-derived mesenchymal stem cells inhibit leukemia/lymphoma cell proliferation in vitro and in a mouse model of allogeneic bone marrow transplant.

PubMed

Song, Ningxia; Gao, Lei; Qiu, Huiying; Huang, Chongmei; Cheng, Hui; Zhou, Hong; Lv, Shuqing; Chen, Li; Wang, Jianmin

2015-07-01

The allogeneic hematopoietic stem cell (HSC) transplantation of mesenchymal stem cells (MSCs) contributes to the reconstitution of hematopoiesis by ameliorating acute graft‑versus‑host disease (aGVHD). However, the role of MSCs in graft‑versus‑leukemia remains to be determined. In the present study, we co‑cultured C57BL/6 mouse bone marrow (BM)‑derived MSCs with A20 murine B lymphoma, FBL3 murine erythroleukemia and P388 murine acute lymphocytic leukemia cells. Cell proliferation, apoptosis, cell cycle progression and the amount of cytokine secretion were then measured using a Cell Counting kit‑8, Annexin V/propidium iodide staining, flow cytometry and ELISA, respectively. We also established a model of allogeneic bone marrow transplantation (BMT) using BALB/c mice. Following the administration of A20 cells and MSCs, we recorded the symptoms and the survival of the mice for 4 weeks, assessed the T cell subsets present in peripheral blood, and, after the mice were sacrifice, we determined the infiltration of MSCs into the organs by histological staining. Our results revealed that the MSCs inhibited the proliferation of the mouse lymphoma and leukemia cells in vitro, leading to cell cycle arrest and reducing the secretion of interleukin (IL)‑10. In our model of allogeneic BMT, the intravenous injection of MSCs into the mice injected wth A20 cells decreased the incidence of lymphoma, improved survival, increased the fraction of CD3+CD8+ T cells, decreased the fraction of CD3+CD4+ T cells and CD4+CD25+ T cells in peripheral blood, and ameliorated the manifestation of aGVHD. The results from the present study indicate that MSCs may be safe and effective when used in allogeneic BMT for the treatment of hemotological malignancies.

Culturing primary mouse pancreatic ductal cells.

PubMed

Reichert, Maximilian; Rhim, Andrew D; Rustgi, Anil K

2015-06-01

The most common subtype of pancreatic cancer is pancreatic ductal adenocarcinoma (PDAC). PDAC resembles ductal cells morphologically. To study pancreatic ductal cell (PDC) and pancreatic intraepithelial neoplasia (PanIN)/PDAC biology, it is essential to have reliable in vitro culture conditions. Here we describe a methodology to isolate, culture, and passage PDCs and duct-like cells from the mouse pancreas. It can be used to isolate cells from genetically engineered mouse models (GEMMs), providing a valuable tool to study disease models in vitro to complement in vivo findings. The culture conditions allow epithelial cells to outgrow fibroblast and other "contaminating" cell types within a few passages. However, the resulting cultures, although mostly epithelial, are not completely devoid of fibroblasts. Regardless, this protocol provides guidelines for a robust in vitro culture system to isolate, maintain, and expand primary pancreatic ductal epithelial cells. It can be applied to virtually all GEMMs of pancreatic disease and other diseases and cancers that arise from ductal structures. Because most carcinomas resemble ductal structures, this protocol has utility in the study of other cancers in addition to PDAC, such as breast and prostate cancers. © 2015 Cold Spring Harbor Laboratory Press.

The bcl-2 knockout mouse exhibits marked changes in osteoblast phenotype and collagen deposition in bone as well as a mild growth plate phenotype

PubMed Central

BOOT-HANDFORD, R. P.; MICHAELIDIS, T. M.; HILLARBY, M. C.; ZAMBELLI, A.; DENTON, J.; HOYLAND, J. A.; FREEMONT, A. J.; GRANT, M. E.; WALLIS, G. A.

1998-01-01

Histological examination of long bones from 1-day-old bcl-2 knockout and age-matched control mice revealed no obvious differences in length of bone, growth plate architecture or stage of endochondral ossification. In 35-day-old bcl-2 knockout mice that are growth retarded or ‘dwarfed’, the proliferative zone of the growth plate appeared slightly thinner and the secondary centres of ossification less well developed than their age-matched wild-type controls. The most marked histological effects of bcl-2 ablation were on osteoblasts and bone. 35-day-old knockout mouse bones exhibited far greater numbers of osteoblasts than controls and the osteoblasts had a cuboidal phenotype in comparison with the normal flattened cell appearance. In addition, the collagen deposited by the osteoblasts in the bcl-2 knockout mouse bone was disorganized in comparison with control tissue and had a pseudo-woven appearance. The results suggest an important role for Bcl-2 in controlling osteoblast phenotype and bone deposition in vivo. PMID:10193316

Methods and theory in bone modeling drift: comparing spatial analyses of primary bone distributions in the human humerus.

PubMed

Maggiano, Corey M; Maggiano, Isabel S; Tiesler, Vera G; Chi-Keb, Julio R; Stout, Sam D

2016-01-01

This study compares two novel methods quantifying bone shaft tissue distributions, and relates observations on human humeral growth patterns for applications in anthropological and anatomical research. Microstructural variation in compact bone occurs due to developmental and mechanically adaptive circumstances that are 'recorded' by forming bone and are important for interpretations of growth, health, physical activity, adaptation, and identity in the past and present. Those interpretations hinge on a detailed understanding of the modeling process by which bones achieve their diametric shape, diaphyseal curvature, and general position relative to other elements. Bone modeling is a complex aspect of growth, potentially causing the shaft to drift transversely through formation and resorption on opposing cortices. Unfortunately, the specifics of modeling drift are largely unknown for most skeletal elements. Moreover, bone modeling has seen little quantitative methodological development compared with secondary bone processes, such as intracortical remodeling. The techniques proposed here, starburst point-count and 45° cross-polarization hand-drawn histomorphometry, permit the statistical and populational analysis of human primary tissue distributions and provide similar results despite being suitable for different applications. This analysis of a pooled archaeological and modern skeletal sample confirms the importance of extreme asymmetry in bone modeling as a major determinant of microstructural variation in diaphyses. Specifically, humeral drift is posteromedial in the human humerus, accompanied by a significant rotational trend. In general, results encourage the usage of endocortical primary bone distributions as an indicator and summary of bone modeling drift, enabling quantitative analysis by direction and proportion in other elements and populations. © 2015 Anatomical Society.

Spatial integration in mouse primary visual cortex.

PubMed

Vaiceliunaite, Agne; Erisken, Sinem; Franzen, Florian; Katzner, Steffen; Busse, Laura

2013-08-01

Responses of many neurons in primary visual cortex (V1) are suppressed by stimuli exceeding the classical receptive field (RF), an important property that might underlie the computation of visual saliency. Traditionally, it has proven difficult to disentangle the underlying neural circuits, including feedforward, horizontal intracortical, and feedback connectivity. Since circuit-level analysis is particularly feasible in the mouse, we asked whether neural signatures of spatial integration in mouse V1 are similar to those of higher-order mammals and investigated the role of parvalbumin-expressing (PV+) inhibitory interneurons. Analogous to what is known from primates and carnivores, we demonstrate that, in awake mice, surround suppression is present in the majority of V1 neurons and is strongest in superficial cortical layers. Anesthesia with isoflurane-urethane, however, profoundly affects spatial integration: it reduces the laminar dependency, decreases overall suppression strength, and alters the temporal dynamics of responses. We show that these effects of brain state can be parsimoniously explained by assuming that anesthesia affects contrast normalization. Hence, the full impact of suppressive influences in mouse V1 cannot be studied under anesthesia with isoflurane-urethane. To assess the neural circuits of spatial integration, we targeted PV+ interneurons using optogenetics. Optogenetic depolarization of PV+ interneurons was associated with increased RF size and decreased suppression in the recorded population, similar to effects of lowering stimulus contrast, suggesting that PV+ interneurons contribute to spatial integration by affecting overall stimulus drive. We conclude that the mouse is a promising model for circuit-level mechanisms of spatial integration, which relies on the combined activity of different types of inhibitory interneurons.

Systemically Transplanted Bone Marrow-derived Cells Contribute to Dental Pulp Regeneration in a Chimeric Mouse Model.

PubMed

Xu, Wenan; Jiang, Shan; Chen, Qiuyue; Ye, Yanyan; Chen, Jiajing; Heng, Boon Chin; Jiang, Qianli; Wu, Buling; Ding, Zihai; Zhang, Chengfei

2016-02-01

Migratory cells via blood circulation or cells adjacent to the root apex may potentially participate in dental pulp tissue regeneration or renewal. This study investigated whether systemically transplanted bone marrow cells can contribute to pulp regeneration in a chimeric mouse model. A chimeric mouse model was created through the injection of bone marrow cells from green fluorescent protein (GFP) transgenic C57BL/6 mice into the tail veins of recipient wild-type C57BL/6 mice that had been irradiated with a lethal dose of 8.5 Gy from a high-frequency linear accelerator. These mice were subjected to pulpectomy and pulp revascularization. At 1, 4, and 8 weeks after surgery, in vivo animal imaging and histologic analyses were conducted. In vivo animal imaging showed that the green biofluorescence signal from the transplanted GFP+ cells increased significantly and was maintained at a high level during the first 4 weeks after surgery. Immunofluorescence analyses of tooth specimens collected at 8 weeks postsurgery showed the presence of nestin+/GFP+, α smooth muscle actin (α-SMA)/GFP+, and NeuN/GFP+ cells within the regenerated pulplike tissue. These data confirm that transplanted bone marrow-derived cells can contribute to dental pulp regeneration. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Variable Bone Fragility Associated With an Amish COL1A2 Variant and a Knock-in Mouse Model

PubMed Central

Daley, Ethan; Streeten, Elizabeth A; Sorkin, John D; Kuznetsova, Natalia; Shapses, Sue A; Carleton, Stephanie M; Shuldiner, Alan R; Marini, Joan C; Phillips, Charlotte L; Goldstein, Steven A; Leikin, Sergey; McBride, Daniel J

2010-01-01

Osteogenesis imperfecta (OI) is a heritable form of bone fragility typically associated with a dominant COL1A1 or COL1A2 mutation. Variable phenotype for OI patients with identical collagen mutations is well established, but phenotype variability is described using the qualitative Sillence classification. Patterning a new OI mouse model on a specific collagen mutation therefore has been hindered by the absence of an appropriate kindred with extensive quantitative phenotype data. We benefited from the large sibships of the Old Order Amish (OOA) to define a wide range of OI phenotypes in 64 individuals with the identical COL1A2 mutation. Stratification of carrier spine (L1–4) areal bone mineral density (aBMD) Z-scores demonstrated that 73% had moderate to severe disease (less than −2), 23% had mild disease (−1 to −2), and 4% were in the unaffected range (greater than −1). A line of knock-in mice was patterned on the OOA mutation. Bone phenotype was evaluated in four F1 lines of knock-in mice that each shared approximately 50% of their genetic background. Consistent with the human pedigree, these mice had reduced body mass, aBMD, and bone strength. Whole-bone fracture susceptibility was influenced by individual genomic factors that were reflected in size, shape, and possibly bone metabolic regulation. The results indicate that the G610C OI (Amish) knock-in mouse is a novel translational model to identify modifying genes that influence phenotype and for testing potential therapies for OI. © 2010 American Society for Bone and Mineral Research PMID:19594296

Engineering a humanized bone organ model in mice to study bone metastases.

PubMed

Martine, Laure C; Holzapfel, Boris M; McGovern, Jacqui A; Wagner, Ferdinand; Quent, Verena M; Hesami, Parisa; Wunner, Felix M; Vaquette, Cedryck; De-Juan-Pardo, Elena M; Brown, Toby D; Nowlan, Bianca; Wu, Dan Jing; Hutmacher, Cosmo Orlando; Moi, Davide; Oussenko, Tatiana; Piccinini, Elia; Zandstra, Peter W; Mazzieri, Roberta; Lévesque, Jean-Pierre; Dalton, Paul D; Taubenberger, Anna V; Hutmacher, Dietmar W

2017-04-01

Current in vivo models for investigating human primary bone tumors and cancer metastasis to the bone rely on the injection of human cancer cells into the mouse skeleton. This approach does not mimic species-specific mechanisms occurring in human diseases and may preclude successful clinical translation. We have developed a protocol to engineer humanized bone within immunodeficient hosts, which can be adapted to study the interactions between human cancer cells and a humanized bone microenvironment in vivo. A researcher trained in the principles of tissue engineering will be able to execute the protocol and yield study results within 4-6 months. Additive biomanufactured scaffolds seeded and cultured with human bone-forming cells are implanted ectopically in combination with osteogenic factors into mice to generate a physiological bone 'organ', which is partially humanized. The model comprises human bone cells and secreted extracellular matrix (ECM); however, other components of the engineered tissue, such as the vasculature, are of murine origin. The model can be further humanized through the engraftment of human hematopoietic stem cells (HSCs) that can lead to human hematopoiesis within the murine host. The humanized organ bone model has been well characterized and validated and allows dissection of some of the mechanisms of the bone metastatic processes in prostate and breast cancer.

Analysis of imaging characteristics of primary malignant bone tumors in children

PubMed Central

Sun, Yingwei; Liu, Xueyong; Pan, Shinong; Deng, Chunbo; Li, Xiaohan; Guo, Qiyong

2017-01-01

The present study aimed to investigate the imaging characteristics of primary malignant bone tumors in children. The imaging results of 34 children with primary malignant bone tumors confirmed by histopathological diagnosis between March 2008 and January 2014 were retrospectively analyzed. In total, 25 patients had osteosarcoma, with radiography and computed tomography (CT) showing osteolytic bone destruction or/and osteoblastic bone sclerosis, an aggressive periosteal reaction, a soft-tissue mass and cancerous bone. The tumors appeared as mixed magnetic resonance imaging (MRI) signals that were inhomogeneously enhanced. A total of 5 patients presented with Ewing sarcoma, with radiography and CT showing invasive bone destruction and a soft-tissue mass. Of the 5 cases, 2 showed a laminar periosteal reaction. The tumors were shown to have mixed low signal on T1-weighted images (T1WI) and high signal on T2-weighted images (T2WI); 1 case showed marked inhomogeneous enhancement. Another 3 patients exhibited chondrosarcoma. Of these cases, 1 was adjacent to the cortex of the proximal tibia, and presented with local cortical bone destruction and a soft-tissue mass containing scattered punctate and amorphous calcifications. MRI revealed mixed low T1 signal and high T2 signals. Another case was located in the medullary cavity of the distal femur, with radiography revealing a localized periosteal reaction. The tumor appeared with mixed MRI signals, and with involvement of the epiphysis and epiphyseal plates. Radiography and CT of the third case showed bone destruction in the right pubic ramus, with patchy punctate, cambered calcifications in the soft-tissue mass. MRI of the soft-tissue mass revealed isointensity on T1WI and heterogeneous hyperintensity on T2WI. Ossifications and the septum appeared as low T1WI and T2WI. Of the 34 patients, 1 patient presented with lymphoma involving the T12, L1 and L2 vertebrae. CT showed vertebral bone destruction, a soft-tissue mass and a

Hypermineralization and High Osteocyte Lacunar Density in Osteogenesis Imperfecta Type V Bone Indicate Exuberant Primary Bone Formation.

PubMed

Blouin, Stéphane; Fratzl-Zelman, Nadja; Glorieux, Francis H; Roschger, Paul; Klaushofer, Klaus; Marini, Joan C; Rauch, Frank

2017-09-01

In contrast to "classical" forms of osteogenesis imperfecta (OI) types I to IV, caused by a mutation in COL1A1/A2, OI type V is due to a gain-of-function mutation in the IFITM5 gene, encoding the interferon-induced transmembrane protein 5, or bone-restricted interferon-inducible transmembrane (IFITM)-like protein (BRIL). Its phenotype distinctly differs from OI types I to IV by absence of blue sclerae and dentinogenesis imperfecta, by the occurrence of ossification disorders such as hyperplastic callus and forearm interosseous membrane ossification. Little is known about the impact of the mutation on bone tissue/material level in untreated and bisphosphonate-treated patients. Therefore, investigations of transiliac bone biopsy samples from a cohort of OI type V children (n = 15, 8.7 ± 4 years old) untreated at baseline and a subset (n = 8) after pamidronate treatment (2.6 years in average) were performed. Quantitative backscattered electron imaging (qBEI) was used to determine bone mineralization density distribution (BMDD) as well as osteocyte lacunar density. The BMDD of type V OI bone was distinctly shifted toward a higher degree of mineralization. The most frequently occurring calcium concentration (CaPeak) in cortical (Ct) and cancellous (Cn) bone was markedly increased (+11.5%, +10.4%, respectively, p < 0.0001) compared to healthy reference values. Treatment with pamidronate resulted in only a slight enhancement of mineralization. The osteocyte lacunar density derived from sectioned bone area was elevated in OI type V Ct and Cn bone (+171%, p < 0.0001; +183.3%, p < 0.01; respectively) versus controls. The high osteocyte density was associated with an overall immature primary bone structure ("mesh-like") as visualized by polarized light microscopy. In summary, the bone material from OI type V patients is hypermineralized, similar to other forms of OI. The elevated osteocyte lacunar density in connection with lack of regular bone

Ewing's sarcoma of bone tumor cells produces MCSF that stimulates monocyte proliferation in a novel mouse model of Ewing's sarcoma of bone.

PubMed

Margulies, B S; DeBoyace, S D; Damron, T A; Allen, M J

2015-10-01

Ewing's sarcoma of bone is a primary childhood malignancy of bone that is treated with X-radiation therapy in combination with surgical excision and chemotherapy. To better study Ewing's sarcoma of bone we developed a novel model of primary Ewing's sarcoma of bone and then treated animals with X-radiation therapy. We identified that uncontrolled tumor resulted in lytic bone destruction while X-radiation therapy decreased lytic bone destruction and increased limb-length asymmetry, a common, crippling complication of X-radiation therapy. Osteoclasts were indentified adjacent to the tumor, however, we were unable to detect RANK-ligand in the Ewing's tumor cells in vitro, which lead us to investigate alternate mechanisms for osteoclast formation. Ewing's sarcoma tumor cells and archival Ewing's sarcoma of bone tumor biopsy samples were shown to express MCSF, which could promote osteoclast formation. Increased monocyte numbers were detected in peripheral blood and spleen in animals with untreated Ewing's sarcoma tumor while monocyte number in animals treated with x-radiation had normal numbers of monocytes. Our data suggest that our Ewing's sarcoma of bone model will be useful in the study Ewing's sarcoma tumor progression in parallel with the effects of chemotherapy and X-radiation therapy. Copyright © 2015 Elsevier Inc. All rights reserved.

Ewing's Sarcoma of Bone Tumor Cells Produce MCSF that Stimulates Monocyte Proliferation in a Novel Mouse Model of Ewing's Sarcoma of Bone

PubMed Central

Margulies, BS; DeBoyace, SD; Damron, TA; Allen, MJ

2015-01-01

Ewing's sarcoma of bone is a primary childhood malignancy of bone that is treated with X-radiation therapy in combination with surgical excision and chemotherapy. To better study Ewing's sarcoma of bone we developed a novel model of primary Ewing's sarcoma of bone and then treated animals with X-radiation therapy. We identified that uncontrolled tumor resulted in lytic bone destruction while X-radiation therapy decreased lytic bone destruction and increased limb-length asymmetry, a common, crippling complication of X-radiation therapy. Osteoclasts were indentified adjacent to the tumor, however, we were unable to detect RANK-ligand in the Ewing's tumor cells in vitro, which lead us to investigate alternate mechanisms for osteoclast formation. Ewing's sarcoma tumor cells and archival Ewing's sarcoma of bone tumor biopsy samples were shown to express MCSF, which could promote osteoclast formation. Increased monocyte numbers were detected in peripheral blood and spleen in animals with untreated Ewing's sarcoma tumor while monocyte number in animals treated with x-radiation had normal numbers of monocytes. Our data suggest that our Ewing's sarcoma of bone model will be useful in the study Ewing's sarcoma tumor progression in parallel with the effects of chemotherapy and X-radiation therapy. PMID:26051470

The Risk of Misdiagnosing the Primary Site Responsible for Bone Metastases in Patients With Chronic Lymphocytic Leukemia and a Second Primary Carcinoma

PubMed Central

Hatoum, Georges; Meshkin, Cyrus; Alkhunaizi, Sufana; Levene, Richard; Formoso-Onofrio, Julie

2015-01-01

Chronic lymphocytic leukemia (CLL) is a common malignancy which may coexist with other primary cancers. CLL is rarely the cause of solitary bone lesions; such lesions in the context of CLL are believed to result from either Richter’s transformation or metastasis from another primary malignancy. Renal cell carcinoma (RCC), on the other hand, is a malignancy which frequently metastasizes to bone and may cause an osteolytic solitary bone lesion. The origin of a solitary bone lesion in a patient with multiple potential primary malignancies has prognostic implications and affects treatment protocol, and as such must be diagnosed accurately. We describe a patient with CLL and a history of RCC who is found to have an incidental solitary bone lesion of the T11 vertebra. After two separate CT-guided biopsies revealed various lymphoid cell predominance and no evidence of RCC, treatment with low dose external beam radiation therapy (EBRT) was employed. Post-therapy MRI showed further propagation of the lesion. Surgical corpectomy was subsequently performed and postoperative pathology of the lesion was consistent with RCC. The patient was treated with bisphosphonates and a higher dose of EBRT. Our case illustrates the importance of surgical excisional biopsy for accurately diagnosing the primary source metastatic to the bone in a patient with CLL and another potential primary cancer. PMID:29147427

Bone pain caused by swelling of mouse ear capsule static xylene and effects on rat models of cervical spondylosis

NASA Astrophysics Data System (ADS)

Zhang, Xuhui; Xia, Lei; Hao, Shaojun; Chen, Weiliang; Guo, Junyi; Ma, Zhenzhen; Wang, Huamin; Kong, Xuejun; Wang, Hongyu; Zhang, Zhengchen

2018-04-01

To observe the effect of intravenous bone pain Capsule on the ear of mice induced by xylene, swelling of rat models of cervical spondylosis. Weighing 18 ˜ 21g 50 mice, male, were randomly divided into for five groups, which were fed with service for bone pain static capsule suspension, Jingfukang granule suspension 0.5%CMC liquid and the same volume of. Respectively to the mice ear drop of xylene 0.05 ml, 4h after cervical dislocation, the mice were sacrificed and the cut two ear, rapid analytical balance weighing, and calculate the ear swelling degree and the other to take the weight of 200 - 60 250g male SD rats, were randomly divided into for 6 groups, 10 rats in each group, of which 5 groups made cervical spondylosis model. Results: with the blank group than bone pain static capsule group and Jingfukang granule group can significantly reduce mouse auricular dimethylbenzene swelling, significantly reduce ear swelling degree (P < 0.01); the successful establishment of the rat model of cervical spondylosis. With the model group ratio, large, medium and small dose of bone pain static capsule group, Jingfukang granule group (P < 0.01) angle of swash plate of rats increased significantly, the high and middle dose of bone pain static capsule group, Jingfukang granule group can significantly reduce the rat X-ray scores (P < 0.05). Bone pain static capsule can significantly reduce mouse auricular dimethylbenzene swelling. The bone pain capsule has a good effect on the rat model of cervical spondylosis.

Predicting the bending properties of long bones: Insights from an experimental mouse model.

PubMed

Peacock, Sarah J; Coats, Brittney R; Kirkland, J Kyle; Tanner, Courtney A; Garland, Theodore; Middleton, Kevin M

2018-03-01

Analyses of bone cross-sectional geometry are frequently used by anthropologists and paleontologists to infer the loading histories of past populations. To address some underlying assumptions, we investigated the relative roles of genetics and exercise on bone cross-sectional geometry and bending mechanics in three mouse strains: high bone density (C3H/He), low bone density (C57BL/6), and a high-runner strain homozygous for the Myh4 Minimsc allele (MM). Weanlings of each strain were divided into exercise (wheel) or control (sedentary) treatment groups for a 7-week experimental period. Morphometrics of the femoral mid-diaphysis and mechanical testing were used to assess both theoretical and ex vivo bending mechanics. Across all measured morphological and bending traits, we found relatively small effects of exercise treatment compared to larger and more frequent interstrain differences. In the exercised group, total distance run over the experimental period was not a predictor of any morphological or bending traits. Cross-sectional geometry did not accurately predict bone response to loading. Results from this experimental model do not support hypothesized associations among extreme exercise, cross-sectional geometry, and bending mechanics. Our results suggest that analysis of cross-sectional geometry alone is insufficient to predict loading response, and questions the common assumption that cross-sectional geometry differences are indicative of differential loading history. © 2017 Wiley Periodicals, Inc.

Ionizing Radiation Affects Gene Expression in Mouse Skin and Bone

NASA Technical Reports Server (NTRS)

Terada, Masahiro; Tahimic, Candice; Sowa, Marianne B.; Schreurs, Ann-Sofie; Shirazi-Fard, Yasaman; Alwood, Joshua; Globus, Ruth K.

2017-01-01

Future long-duration space exploration beyond low earth orbit will increase human exposure to space radiation and microgravity conditions as well as associated risks to skeletal health. In animal studies, radiation exposure (greater than 1 Gy) is associated with pathological changes in bone structure, enhanced bone resorption, reduced bone formation and decreased bone mineral density, which can lead to skeletal fragility. Definitive measurements and detection of bone loss typically require large and specialized equipment which can make their application to long duration space missions logistically challenging. Towards the goal of developing non-invasive and less complicated monitoring methods to predict astronauts' health during spaceflight, we examined whether radiation induced gene expression changes in skin may be predictive of the responses of skeletal tissue to radiation exposure. We examined oxidative stress and growth arrest pathways in mouse skin and long bones by measuring gene expression levels via quantitative polymerase chain reaction (qPCR) after exposure to total body irradiation (IR). To investigate the effects of irradiation on gene expression, we used skin and femora (cortical shaft) from the following treatment groups: control (normally loaded, sham-irradiated), and IR (0.5 Gy 56Fe 600 MeV/n and 0.5 Gy 1H 150 MeV/n), euthanized at one and 11 days post-irradiation (IR). To determine the extent of bone loss, tibiae were harvested and cancellous microarchitecture in the proximal tibia quantified ex vivo using microcomputed tomography (microCT). Statistical analysis was performed using Student's t-test. At one day post-IR, expression of FGF18 in skin was significantly greater (3.8X) than sham-irradiated controls, but did not differ at 11 days post IR. Expression levels of other genes associated with antioxidant response (Nfe2l2, FoxO3 and Sod1) and the cell cycle (Trp53, Cdkn1a, Gadd45g) did not significantly differ between the control and IR groups

Development and characterization of a mouse floxed Bmp2 osteoblast cell line that retains osteoblast genotype and phenotype

PubMed Central

Wu, Li-an; Feng, Junsheng; Wang, Lynn; Mu, Yan-dong; Baker, Andrew; Donly, Kevin J.; Harris, Stephen E.; MacDougall, Mary; Chen, Shuo

2011-01-01

Bone morphogenetic protein 2 (Bmp2) is essential for osteoblast differentiation and osteogenesis. Generation of floxed Bmp2 osteoblast cell lines is a valuable tool for studying the effects of Bmp2 on osteoblast differentiation and its signaling pathways during skeletal metabolism. Due to relatively limited sources of primary osteoblasts, we have developed cell lines that serve as good surrogate models for the study of osteoblast cell differentiation and bone mineralization. In this study, we established and characterized immortalized mouse floxed Bmp2 osteoblast cell lines. Primary mouse floxed Bmp2 osteoblasts were transfected with pSV3-neo and clonally selected. These transfected cells were verified by PCR and immunohistochemistry. To determine the genotype and phenotype of the immortalized cells, cell morphology, proliferation, differentiation and mineralization were analyzed. Also, expression of osteoblast-related gene markers including Runx2, Osx, ATF4, Dlx3, bone sialoprotein, dentin matrix protein 1, osteonectin, osteocalcin and osteopontin were examined by quantitative RT-PCR and immunohistochemistry. These results showed that immortalized floxed Bmp2 osteoblasts had a higher proliferation rate but preserved their genotypic and phenotypic characteristics similar to the primary cells. Thus, we, for the first time, describe the development of immortalized mouse floxed Bmp2 osteoblast cell lines and present a useful model to study osteoblast biology mediated by BMP2 and its downstream signaling transduction pathways. PMID:21271257

Modularity in the Organization of Mouse Primary Visual Cortex

PubMed Central

Ji, Weiqing; Gămănuţ, Răzvan; Bista, Pawan; D’Souza, Rinaldo D.; Wang, Quanxin; Burkhalter, Andreas

2015-01-01

SUMMARY Layer 1 (L1) of primary visual cortex (V1) is the target of projections from many brain regions outside of V1. We found that inputs to the non-columnar mouse V1 from the dorsal lateral geniculate nucleus and feedback projections from multiple higher cortical areas to L1 are patchy. The patches are matched to a pattern of M2 muscarinic acetylcholine receptor expression at fixed locations of mouse, rat and monkey V1. Neurons in L2/3 aligned with M2-rich patches have high spatial acuity whereas cells in M2-poor zones exhibited high temporal acuity. Together M2+ and M2− zones form constant-size domains that are repeated across V1. Domains map subregions of the receptive field, such that multiple copies are contained within the point image. The results suggest that the modular network in mouse V1 selects spatiotemporally distinct clusters of neurons within the point image for top-down control and differential routing of inputs to cortical streams. PMID:26247867

Leukemia inhibitory factor: a novel bone-active cytokine.

PubMed

Reid, L R; Lowe, C; Cornish, J; Skinner, S J; Hilton, D J; Willson, T A; Gearing, D P; Martin, T J

1990-03-01

A number of cytokines have been found to be potent regulators of bone resorption and to share the properties originally attributed to osteoclast-activating factor. One such activity, differentiation-inducing factor (DIF, D-factor) from mouse spleen cells, shares a number of biological and biochemical properties with the recently characterized and cloned leukemia inhibitory factor (LIF). We have assessed the effects of recombinant LIF on bone resorption and other parameters in neonatal mouse calvaria. Both recombinant murine and human (h) LIFs stimulated 45Ca release from prelabeled calvaria in a dose-dependent manner. The increase in bone resorption was associated with an increase in the number of osteoclasts per mm2 bone. The osteolytic effect of hLIF were blocked by 10(-7) M indomethacin. hLIF also stimulated incorporation of [3H] thymidine into calvaria, but the dose-response relationship was distinct from that for bone resorption, and this effect was not blocked by indomethacin. Similarly, hLIF increased [3H]phenylalanine incorporation into calvaria, and this was also not inhibited by indomethacin. It is concluded that LIF stimulates bone resorption by a mechanism involving prostaglandin production, but that a distinct mechanism is responsible for its stimulation of DNA and protein synthesis. The primary structure of LIF differs from that of other fully characterized, bone-active cytokines, and it, thus, represents a novel factor which may be involved in the normal regulation of bone cell function.

AN INVESTIGATION OF THE MINERAL IN DUCTILE AND BRITTLE CORTICAL MOUSE BONE

PubMed Central

Rodriguez-Florez, Naiara; Garcia-Tunon, Esther; Mukadam, Quresh; Saiz, Eduardo; Oldknow, Karla J.; Farquharson, Colin; Millán, José Luis; Boyde, Alan; Shefelbine, Sandra J.

2015-01-01

Bone is a strong and tough material composed of apatite mineral, organic matter and water. Changes in composition and organization of these building blocks affect bone’s mechanical integrity. Skeletal disorders often affect bone’s mineral phase, either by variations in the collagen or directly altering mineralization. The aim of the current study was to explore the differences in the mineral of brittle and ductile cortical bone at the mineral (nm) and tissue (µm) levels using two mouse phenotypes. Osteogenesis imperfecta murine (oim−/−) mice were used to model brittle bone; PHOSPHO1 mutants (Phospho1−/−) had ductile bone. They were compared to their respective wild-type controls. Femora were defatted and ground to powder to measure average mineral crystal size using X-ray diffraction (XRD), and to monitor the bulk mineral to matrix ratio via thermogravimetric analysis (TGA). XRD scans were run after TGA for phase identification, to assess the fractions of hydroxyapatite and β-tricalcium phosphate. Tibiae were embedded to measure elastic properties with nanoindentation and the extent of mineralization with backscattered electron microscopy (qbSEM). Interestingly, the mineral of brittle oim−/− and ductile Phospho1−/− bones had many similar characteristics. Both pathology models had smaller apatite crystals, lower mineral to matrix ratio, and showed more thermal conversion to β-tricalcium phosphate than their wild-types, indicating deviations from stoichiometric hydroxyapatite in the original mineral. The degree of mineralization of the bone matrix was different for each strain: oim−/− were hypermineralized, while Phospho1−/− were hypomineralized. However, alterations in the mineral were associated with reduced tissue elastic moduli in both pathologies. Results revealed that despite having extremely different whole bone mechanics, the mineral of oim−/− and Phospho1−/− has several similar trends at smaller length scales. This

Gender-dependence of bone structure and properties in adult osteogenesis imperfecta murine model.

PubMed

Yao, Xiaomei; Carleton, Stephanie M; Kettle, Arin D; Melander, Jennifer; Phillips, Charlotte L; Wang, Yong

2013-06-01

Osteogenesis imperfecta (OI) is a dominant skeletal disorder characterized by bone fragility and deformities. Though the oim mouse model has been the most widely studied of the OI models, it has only recently been suggested to exhibit gender-dependent differences in bone mineralization. To characterize the impact of gender on the morphometry/ultra-structure, mechanical properties, and biochemical composition of oim bone on the congenic C57BL/J6 background, 4-month-old oim/oim, +/oim, and wild-type (wt) female and male tibiae were evaluated using micro-computed tomography, three-point bending, and Raman spectroscopy. Dramatic gender differences were evident in both cortical and trabecular bone morphological and geometric parameters. Male mice had inherently more bone and increased moment of inertia than genotype-matched female counterparts with corresponding increases in bone biomechanical strength. The primary influence of gender was structure/geometry in bone growth and mechanical properties, whereas the mineral/matrix composition and hydroxyproline content of bone were influenced primarily by the oim collagen mutation. This study provides evidence of the importance of gender in the evaluation and interpretation of potential therapeutic strategies when using mouse models of OI.

Gender-dependence of bone structure and properties in adult osteogenesis imperfecta murine model

PubMed Central

Yao, Xiaomei; Carleton, Stephanie M.; Kettle, Arin D; Melander, Jennifer; Phillips, Charlotte L.; Wang, Yong

2013-01-01

Osteogenesis imperfecta (OI) is a dominant skeletal disorder characterized by bone fragility and deformities. Though the oim mouse model has been the most widely studied of the OI models, it has only recently been suggested to exhibit gender-dependent differences in bone mineralization. To characterize the impact of gender on the morphometry/ultra-structure, mechanical properties, and biochemical composition of oim bone on the congenic C57BL/J6 background, 4-month-old oim/oim, +/oim, and wild-type (wt) female and male tibiae were evaluated using micro-computed tomography, three-point bending, and Raman spectroscopy. Dramatic gender differences were evident in both cortical and trabecular bone morphological and geometric parameters. Male mice had inherently more bone and increased moment of inertia than genotype-matched female counterparts with corresponding increases in bone biomechanical strength. The primary influence of gender was structure/geometry in bone growth and mechanical properties, whereas the mineral/matrix composition, hydroxyproline content of bone were influenced primarily by the oim collagen mutation. This study provides evidence of the importance of gender in the evaluation and interpretation of potential therapeutic strategies when using mouse models of OI. PMID:23536112

Primary Neoplasms of Bones in Mice: Retrospective Study and Review of Literature

PubMed Central

Kavirayani, A. M.; Sundberg, J. P.; Foreman, O.

2011-01-01

To compare and summarize the mechanisms, frequencies of occurrence, and classification schemes of spontaneous, experimental, and genetically engineered, mouse skeletal neoplasms, the literature was reviewed and archived case material at The Jackson Laboratory examined. The frequency of occurrence of spontaneous bone neoplasms was less than 1% for most strains, with the exceptions of osteomas in CF-1 (5.5% and 10% in two studies) and OF-1 outbred strains (35%), and osteosarcomas in NOD/ShiLtJ (11.5%) and NOD derived (7.1%) mice. The frequency was 100% for osteochondromas induced by conditional inactivation of exostoses (multiple) 1 (Ext1) in chondrocytes, osteosarcomas induced by tibial intramedullary inoculation of Moloney’s murine sarcoma virus, and osteosarcomas induced by conditional inactivation of Trp53-with or without inactivation of Rb1-in osteoblast precursors. Spontaneous osteogenic neoplasms were more frequent than spontaneous cartilaginous and vascular types. Malignant neoplasms were more frequent than benign ones. The age of occurrence for spontaneous neoplasms ranged from 37 to 720 (Mean 316.35) days for benign, and 35 to 990 (Mean 299.28) days for malignant neoplasms. In genetically engineered mice, the average age of occurrence ranged from 28 to 70 days for benign, and from 35 to 690 days for malignant neoplasms. Histologically, non-osteogenic neoplasms were similar across strains and mutant stocks; osteogenic neoplasms exhibited greater diversity. This comparison and summarization of mouse bone neoplasms provides valuable information for the selection of strains to create, compare, and validate models of bone neoplasms. PMID:21343597

Long-term primary culture of mouse mammary tumor cells: production of virus.

PubMed

Young, L J; Cardiff, R D; Ashley, R L

1975-05-01

Long-term primary cultures of mouse mammary tumor cells proved an excellent source of mouse mammary tumor virus (MMTV). Virus purified from these primary cultures had the same morphologic biochemical, immunologic, and biologic characteristics as MMTV. Quantitation of MMTV-protein equivalents released into the medium was measured by the radioimmunoassay for MMTV. Peak production levels were 20-40 mug MMTV protien equivalents/75-cm-2 flask/24 hours. These cultures produced MMTV for as long as 90 days. MMTV cultivation depended on the initial cell-plating density and hormones. Maximal MMTV release was obtained at a plating density of 1 times 10-6 cells/cm-2 in the presence of insulin and hydrocortisone. Insulin alone gave basal levels of MMTV, and hydrocortisone alone increased MMTV release only three-fold, but insulin and hydrocortisone together effected an eightfold increase in MMTV release. This suggested that hydrocortisone had a primary effect on MMTV release and insulin acted synergistically with hydrocortisone to maximize MMTV release.

Establishment of a patient-derived orthotopic osteosarcoma mouse model.

PubMed

Blattmann, Claudia; Thiemann, Markus; Stenzinger, Albrecht; Roth, Eva K; Dittmar, Anne; Witt, Hendrik; Lehner, Burkhard; Renker, Eva; Jugold, Manfred; Eichwald, Viktoria; Weichert, Wilko; Huber, Peter E; Kulozik, Andreas E

2015-04-30

Osteosarcoma (OS) is the most common pediatric primary malignant bone tumor. As the prognosis for patients following standard treatment did not improve for almost three decades, functional preclinical models that closely reflect important clinical cancer characteristics are urgently needed to develop and evaluate new treatment strategies. The objective of this study was to establish an orthotopic xenotransplanted mouse model using patient-derived tumor tissue. Fresh tumor tissue from an adolescent female patient with osteosarcoma after relapse was surgically xenografted into the right tibia of 6 immunodeficient BALB/c Nu/Nu mice as well as cultured into medium. Tumor growth was serially assessed by palpation and with magnetic resonance imaging (MRI). In parallel, a primary cell line of the same tumor was established. Histology and high-resolution array-based comparative genomic hybridization (aCGH) were used to investigate both phenotypic and genotypic characteristics of different passages of human xenografts and the cell line compared to the tissue of origin. A primary OS cell line and a primary patient-derived orthotopic xenotranplanted mouse model were established. MRI analyses and histopathology demonstrated an identical architecture in the primary tumor and in the xenografts. Array-CGH analyses of the cell line and all xenografts showed highly comparable patterns of genomic progression. So far, three further primary patient-derived orthotopic xenotranplanted mouse models could be established. We report the first orthotopic OS mouse model generated by transplantation of tumor fragments directly harvested from the patient. This model represents the morphologic and genomic identity of the primary tumor and provides a preclinical platform to evaluate new treatment strategies in OS.

Primary telangiectatic osteosarcoma of occipital bone: a case report and review of literature.

PubMed

Patibandla, Mohana Rao; Uppin, Shantveer G; Thotakura, Amit Kumar; Panigrahi, Manas K; Challa, Sundaram

2011-01-01

Telangiectatic osteosarcoma (TOS), an uncommon variant of osteosarcoma, involving skull bones is extremely rare. We present clinico-pathological, imageological and treatment outcome of a primary TOS of occipital bone in a 30-year-old woman and review the previously reported skull bone TOS. We suggest that TOS should be included in the differential diagnosis of destructive lytic lesions involving the skull bones. As radical surgical procedures are not applicable to skull bones, the outcome is poor even with adjuvant chemotherapy.

Columnar metaplasia in a surgical mouse model of gastro-esophageal reflux disease is not derived from bone marrow-derived cell.

PubMed

Aikou, Susumu; Aida, Junko; Takubo, Kaiyo; Yamagata, Yukinori; Seto, Yasuyuki; Kaminishi, Michio; Nomura, Sachiyo

2013-09-01

The incidence of esophageal adenocarcinoma has increased in the last 25 years. Columnar metaplasia in Barrett's mucosa is assumed to be a precancerous lesion for esophageal adenocarcinoma. However, the induction process of Barrett's mucosa is still unknown. To analyze the induction of esophageal columnar metaplasia, we established a mouse gastro-esophageal reflux disease (GERD) model with associated development of columnar metaplasia in the esophagus. C57BL/6 mice received side-to-side anastomosis of the esophagogastric junction with the jejunum, and mice were killed 10, 20, and 40 weeks after operation. To analyze the contribution of bone marrow-derived cells to columnar metaplasia in this surgical GERD model, some mice were transplanted with GFP-marked bone marrow after the operation. Seventy-three percent of the mice (16/22) showed thickened mucosa in esophagus and 41% of mice (9/22) developed columnar metaplasia 40 weeks after the operation with a mortality rate of 4%. Bone marrow-derived cells were not detected in columnar metaplastic epithelia. However, scattered epithelial cells in the thickened squamous epithelia in regions of esophagitis did show bone marrow derivation. The results demonstrate that reflux induced by esophago-jejunostomy in mice leads to the development of columnar metaplasia in the esophagus. However, bone marrow-derived cells do not contribute directly to columnar metaplasia in this mouse model. © 2013 Japanese Cancer Association.

Establishment and characterization of mouse bone marrow-derived mast cell hybridomas

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kawahara, Takeshi, E-mail: tkawafb@shinshu-u.ac.jp

2012-11-01

Interleukin (IL)-3-dependent mouse bone marrow-derived mast cells (BMMCs) are an important model for studying the function of mucosal-type mast cells. In the present study, BMMCs were successfully immortalized by cell fusion using a hypoxanthine-aminopterin-thymidine medium-sensitive variant of P815 mouse mastocytoma (P815-6TgR) as a partner cell line. The established mouse mast cell hybridomas (MMCHs) expressed {alpha}, {beta}, and {gamma} subunits of high-affinity immunoglobulin E (IgE) receptor (Fc{epsilon}RI) and possessed cytoplasmic granules devoid of or partially filled with electron-dense material. Four independent MMCH clones continuously proliferated without supplemental exogenous IL-3 and showed a degranulation response on stimulation with IgE+antigen. Furthermore, histamine synthesismore » and release by degranulation were confirmed in MMCH-D5, a MMCH clone that showed the strongest degranulation response. MMCH-D5 exhibited elevated levels of IL-3, IL-4, IL-13, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor (TNF)-{alpha}, and cyclooxygenase 2, and production of prostaglandin D{sub 2} and leukotriene C{sub 4} in response to IgE-induced stimulation. MMCH clones also expressed Toll-like receptors (TLRs) 1, 2, 4, and 6 and showed elevated levels of TNF-{alpha} expression in response to stimulation with TLR2 and TLR4 ligands. The MMCHs established using this method should be suitable for studies on Fc{epsilon}RI- and TLR-mediated effector functions of mast cells.« less

Sex steroids during bone growth: a comparative study between mouse models for hypogonadal and senile osteoporosis.

PubMed

Ophoff, J; Venken, K; Callewaert, F; Boonen, S; Bouillon, R; Vanderschueren, D

2009-10-01

In this study, the role of disturbed bone mineral acquisition during puberty in the pathogenesis of osteoporosis was studied. To this end, a mouse model for senile and hypogonadal osteoporosis was used. Longitudinal follow-up showed that bone fragility in both models results from deficient bone build-up during early puberty. Male osteoporosis may result from impaired bone growth. This study characterizes the mechanisms of deficient peak bone mass acquisition in models for senile (SAMP6) and hypogonadal (orchidectomized SAMR1) osteoporosis. Bone mineral acquisition was investigated longitudinally in SAMP6 and orchidectomized SAMR1 mice (eight to ten animals per group) using peripheral quantitative computed tomography and histomorphometry. Additionally, the effects of long-term 5alpha-dihydrotestosterone (DHT) and 17beta-estradiol (E2) replacement were studied. Statistical analysis was performed using ANOVA and Student's t test. SAMP6 mice showed an early (4 weeks) medullary expansion of the cortex due to impaired endocortical bone formation (-43%). Despite compensatory periosteal bone formation (+47%), cortical thickness was severely reduced in 20-week-old SAMP6 versus SAMR1. Orchidectomy reduced periosteal apposition between 4 and 8 weeks of age and resulted in high bone turnover and less trabecular bone gain in SAMP6 and SAMR1. DHT and E2 stimulated periosteal expansion and trabecular bone in orchidectomized SAMP6 and SAMR1. E2 stimulated endocortical apposition in SAMP6. Moreover, sex steroid action occurred between 4 and 8 weeks of age. Bone fragility in both models resulted from deficient bone build-up during early puberty. DHT and E2 improved bone mass acquisition in orchidectomized animals, suggesting a role for AR and ER in male skeletal development.

Effects of the activin A-myostatin-follistatin system on aging bone and muscle progenitor cells

PubMed Central

Bowser, Matthew; Herberg, Samuel; Arounleut, Phonepasong; Shi, Xingming; Fulzele, Sadanand; Hill, William D.; Isales, Carlos M.; Hamrick, Mark W.

2013-01-01

The activin A-myostatin-follistatin system is thought to play an important role in the regulation of muscle and bone mass throughout growth, development, and aging; however, the effects of these ligands on progenitor cell proliferation and differentiation in muscle and bone are not well understood. In addition, age-associated changes in the relative expression of these factors in musculoskeletal tissues have not been described. We therefore examined changes in protein levels of activin A, follistatin, and myostatin (GDF-8) in both muscle and bone with age in C57BL6 mice using ELISA. We then investigated the effects of activin A, myostatin and follistatin on the proliferation and differentiation of primary myoblasts and mouse bone marrow stromal cells (BMSCs) in vitro. Myostatin levels and the myostatin:follistatin ratio increased with age in the primarily slow-twitch mouse soleus muscle, whereas the pattern was reversed with age in the fast-twitch extensor digitorum longus muscle. Myostatin levels and the myostatin: follistatin ratio increased significantly (+75%) in mouse bone marrow with age, as did activin A levels (+17%). Follistatin increased the proliferation of primary myoblasts from both young and aged mice, whereas myostatin increased proliferation of younger myoblasts but decreased proliferation of older myoblasts. Myostatin reduced proliferation of both young and aged BMSCs in a dose-dependent fashion, and activin A increased mineralization in both young and aged BMSCs. Together these data suggest that aging in mice is accompanied by changes in the expression of activin A and myostatin, as well as changes in the response of bone and muscle progenitor cells to these factors. Myostatin appears to play a particularly important role in the impaired proliferative capacity of muscle and bone progenitor cells from aged mice. PMID:23178301

Tridax procumbens flavonoids promote osteoblast differentiation and bone formation.

PubMed

Al Mamun, Md Abdullah; Hosen, Mohammad Jakir; Islam, Kamrul; Khatun, Amina; Alam, M Masihul; Al-Bari, Md Abdul Alim

2015-11-18

Tridax procumbens flavonoids (TPFs) are well known for their medicinal properties among local natives. Besides traditionally used for dropsy, anemia, arthritis, gout, asthma, ulcer, piles, and urinary problems, it is also used in treating gastric problems, body pain, and rheumatic pains of joints. TPFs have been reported to increase osteogenic functioning in mesenchymal stem cells. Our previous study showed that TPFs were significantly suppressed the RANKL-induced differentiation of osteoclasts and bone resorption. However, the effects of TPFs to promote osteoblasts differentiation and bone formation remain unclear. TPFs were isolated from Tridax procumbens and investigated for their effects on osteoblasts differentiation and bone formation by using primary mouse calvarial osteoblasts. TPFs promoted osteoblast differentiation in a dose-dependent manner demonstrated by up-regulation of alkaline phosphatase and osteocalcin. TPFs also upregulated osteoblast differentiation related genes, including osteocalcin, osterix, and Runx2 in primary osteoblasts. TPFs treated primary osteoblast cells showed significant upregulation of bone morphogenetic proteins (BMPs) including Bmp-2, Bmp-4, and Bmp-7. Addition of noggin, a BMP specific-antagonist, inhibited TPFs induced upregulation of the osteocalcin, osterix, and Runx2. Our findings point towards the induction of osteoblast differentiation by TPFs and suggested that TPFs could be a potential anabolic agent to treat patients with bone loss-associated diseases such as osteoporosis.

CRISPR/Cas9-mediated reversibly immortalized mouse bone marrow stromal stem cells (BMSCs) retain multipotent features of mesenchymal stem cells (MSCs).

PubMed

Hu, Xue; Li, Li; Yu, Xinyi; Zhang, Ruyi; Yan, Shujuan; Zeng, Zongyue; Shu, Yi; Zhao, Chen; Wu, Xingye; Lei, Jiayan; Li, Yasha; Zhang, Wenwen; Yang, Chao; Wu, Ke; Wu, Ying; An, Liping; Huang, Shifeng; Ji, Xiaojuan; Gong, Cheng; Yuan, Chengfu; Zhang, Linghuan; Liu, Wei; Huang, Bo; Feng, Yixiao; Zhang, Bo; Haydon, Rex C; Luu, Hue H; Reid, Russell R; Lee, Michael J; Wolf, Jennifer Moriatis; Yu, Zebo; He, Tong-Chuan

2017-12-19

Mesenchymal stem cells (MSCs) are multipotent non-hematopoietic progenitor cells that can undergo self-renewal and differentiate into multi-lineages. Bone marrow stromal stem cells (BMSCs) represent one of the most commonly-used MSCs. In order to overcome the technical challenge of maintaining primary BMSCs in long-term culture, here we seek to establish reversibly immortalized mouse BMSCs (imBMSCs). By exploiting CRISPR/Cas9-based homology-directed-repair (HDR) mechanism, we target SV40T to mouse Rosa26 locus and efficiently immortalize mouse BMSCs (i.e., imBMSCs). We also immortalize BMSCs with retroviral vector SSR #41 and establish imBMSC41 as a control line. Both imBMSCs and imBMSC41 exhibit long-term proliferative capability although imBMSC41 cells have a higher proliferation rate. SV40T mRNA expression is 130% higher in imBMSC41 than that in imBMSCs. However, FLP expression leads to 86% reduction of SV40T expression in imBMSCs, compared with 63% in imBMSC41 cells. Quantitative genomic PCR analysis indicates that the average copy number of SV40T and hygromycin is 1.05 for imBMSCs and 2.07 for imBMSC41, respectively. Moreover, FLP expression removes 92% of SV40T in imBMSCs at the genome DNA level, compared with 58% of that in imBMSC41 cells, indicating CRISPR/Cas9 HDR-mediated immortalization of BMSCs can be more effectively reversed than that of retrovirus-mediated random integrations. Nonetheless, both imBMSCs and imBMSC41 lines express MSC markers and are highly responsive to BMP9-induced osteogenic, chondrogenic and adipogenic differentiation in vitro and in vivo . Thus, the engineered imBMSCs can be used as a promising alternative source of primary MSCs for basic and translational research in the fields of MSC biology and regenerative medicine.

CRISPR/Cas9-mediated reversibly immortalized mouse bone marrow stromal stem cells (BMSCs) retain multipotent features of mesenchymal stem cells (MSCs)

PubMed Central

Hu, Xue; Li, Li; Yu, Xinyi; Zhang, Ruyi; Yan, Shujuan; Zeng, Zongyue; Shu, Yi; Zhao, Chen; Wu, Xingye; Lei, Jiayan; Li, Yasha; Zhang, Wenwen; Yang, Chao; Wu, Ke; Wu, Ying; An, Liping; Huang, Shifeng; Ji, Xiaojuan; Gong, Cheng; Yuan, Chengfu; Zhang, Linghuan; Liu, Wei; Huang, Bo; Feng, Yixiao; Zhang, Bo; Haydon, Rex C.; Luu, Hue H.; Reid, Russell R.; Lee, Michael J.; Wolf, Jennifer Moriatis; Yu, Zebo; He, Tong-Chuan

2017-01-01

Mesenchymal stem cells (MSCs) are multipotent non-hematopoietic progenitor cells that can undergo self-renewal and differentiate into multi-lineages. Bone marrow stromal stem cells (BMSCs) represent one of the most commonly-used MSCs. In order to overcome the technical challenge of maintaining primary BMSCs in long-term culture, here we seek to establish reversibly immortalized mouse BMSCs (imBMSCs). By exploiting CRISPR/Cas9-based homology-directed-repair (HDR) mechanism, we target SV40T to mouse Rosa26 locus and efficiently immortalize mouse BMSCs (i.e., imBMSCs). We also immortalize BMSCs with retroviral vector SSR #41 and establish imBMSC41 as a control line. Both imBMSCs and imBMSC41 exhibit long-term proliferative capability although imBMSC41 cells have a higher proliferation rate. SV40T mRNA expression is 130% higher in imBMSC41 than that in imBMSCs. However, FLP expression leads to 86% reduction of SV40T expression in imBMSCs, compared with 63% in imBMSC41 cells. Quantitative genomic PCR analysis indicates that the average copy number of SV40T and hygromycin is 1.05 for imBMSCs and 2.07 for imBMSC41, respectively. Moreover, FLP expression removes 92% of SV40T in imBMSCs at the genome DNA level, compared with 58% of that in imBMSC41 cells, indicating CRISPR/Cas9 HDR-mediated immortalization of BMSCs can be more effectively reversed than that of retrovirus-mediated random integrations. Nonetheless, both imBMSCs and imBMSC41 lines express MSC markers and are highly responsive to BMP9-induced osteogenic, chondrogenic and adipogenic differentiation in vitro and in vivo. Thus, the engineered imBMSCs can be used as a promising alternative source of primary MSCs for basic and translational research in the fields of MSC biology and regenerative medicine. PMID:29340096

Nasal Bone Shape Is under Complex Epistatic Genetic Control in Mouse Interspecific Recombinant Congenic Strains

PubMed Central

Burgio, Gaétan; Baylac, Michel; Heyer, Evelyne; Montagutelli, Xavier

2012-01-01

Background Genetic determinism of cranial morphology in the mouse is still largely unknown, despite the localization of putative QTLs and the identification of genes associated with Mendelian skull malformations. To approach the dissection of this multigenic control, we have used a set of interspecific recombinant congenic strains (IRCS) produced between C57BL/6 and mice of the distant species Mus spretus (SEG/Pas). Each strain has inherited 1.3% of its genome from SEG/Pas under the form of few, small-sized, chromosomal segments. Results The shape of the nasal bone was studied using outline analysis combined with Fourier descriptors, and differential features were identified between IRCS BcG-66H and C57BL/6. An F2 cross between BcG-66H and C57BL/6 revealed that, out of the three SEG/Pas-derived chromosomal regions present in BcG-66H, two were involved. Segments on chromosomes 1 (∼32 Mb) and 18 (∼13 Mb) showed additive effect on nasal bone shape. The three chromosomal regions present in BcG-66H were isolated in congenic strains to study their individual effect. Epistatic interactions were assessed in bicongenic strains. Conclusions Our results show that, besides a strong individual effect, the QTL on chromosome 1 interacts with genes on chromosomes 13 and 18. This study demonstrates that nasal bone shape is under complex genetic control but can be efficiently dissected in the mouse using appropriate genetic tools and shape descriptors. PMID:22662199

Bone morphogenetic protein 9 (BMP9) induces effective bone formation from reversibly immortalized multipotent adipose-derived (iMAD) mesenchymal stem cells.

PubMed

Lu, Shun; Wang, Jing; Ye, Jixing; Zou, Yulong; Zhu, Yunxiao; Wei, Qiang; Wang, Xin; Tang, Shengli; Liu, Hao; Fan, Jiaming; Zhang, Fugui; Farina, Evan M; Mohammed, Maryam M; Song, Dongzhe; Liao, Junyi; Huang, Jiayi; Guo, Dan; Lu, Minpeng; Liu, Feng; Liu, Jianxiang; Li, Li; Ma, Chao; Hu, Xue; Lee, Michael J; Reid, Russell R; Ameer, Guillermo A; Zhou, Dongsheng; He, Tongchuan

2016-01-01

Regenerative medicine and bone tissue engineering using mesenchymal stem cells (MSCs) hold great promise as an effective approach to bone and skeletal reconstruction. While adipose tissue harbors MSC-like progenitors, or multipotent adipose-derived cells (MADs), it is important to identify and characterize potential biological factors that can effectively induce osteogenic differentiation of MADs. To overcome the time-consuming and technically challenging process of isolating and culturing primary MADs, here we establish and characterize the reversibly immortalized mouse multipotent adipose-derived cells (iMADs). The isolated mouse primary inguinal MAD cells are reversibly immortalized via the retrovirus-mediated expression of SV40 T antigen flanked with FRT sites. The iMADs are shown to express most common MSC markers. FLP-mediated removal of SV40 T antigen effectively reduces the proliferative activity and cell survival of iMADs, indicating the immortalization is reversible. Using the highly osteogenic BMP9, we find that the iMADs are highly responsive to BMP9 stimulation, express multiple lineage regulators, and undergo osteogenic differentiation in vitro upon BMP9 stimulation. Furthermore, we demonstrate that BMP9-stimulated iMADs form robust ectopic bone with a thermoresponsive biodegradable scaffold material. Collectively, our results demonstrate that the reversibly immortalized iMADs exhibit the characteristics of multipotent MSCs and are highly responsive to BMP9-induced osteogenic differentiation. Thus, the iMADs should provide a valuable resource for the study of MAD biology, which would ultimately enable us to develop novel and efficacious strategies for MAD-based bone tissue engineering.

Overexpression of bone sialoprotein leads to an uncoupling of bone formation and bone resorption in mice.

PubMed

Valverde, Paloma; Zhang, Jin; Fix, Amanda; Zhu, Ji; Ma, Wenli; Tu, Qisheng; Chen, Jake

2008-11-01

The purpose of this study was to determine the effects of bone sialoprotein (BSP) overexpression in bone metabolism in vivo by using a homozygous transgenic mouse line that constitutively overexpresses mouse BSP cDNA driven by the cytomegalovirus (CMV) promoter. CMV-BSP transgenic (TG) mice and wildtype mice were weighed, and their length, BMD, and trabecular bone volume were measured. Serum levels of RANKL, osteocalcin, osteoprotegerin (OPG), TRACP5b, and PTH were determined. Bone histomorphometry, von Kossa staining, RT-PCR analysis, Western blot, MTS assay, in vitro mineralization assay, and TRACP staining were also performed to delineate phenotypes of this transgenic mouse line. Compared with wildtype mice, adult TG mice exhibit mild dwarfism, lower values of BMD, and lower trabecular bone volume. TG mice serum contained increased calcium levels and decreased PTH levels, whereas the levels of phosphorus and magnesium were within normal limits. TG mice serum also exhibited lower levels of osteoblast differentiation markers and higher levels of markers, indicating osteoclastic activity and bone resorption. H&E staining, TRACP staining, and bone histomorphometry showed that adult TG bones were thinner and the number of giant osteoclasts in TG mice was higher, whereas there were no significant alterations in osteoblast numbers between TG mice and WT mice. Furthermore, the vertical length of the hypertrophic zone in TG mice was slightly enlarged. Moreover, ex vivo experiments indicated that overexpression of BSP decreased osteoblast population and increased osteoclastic activity. Partly because of its effects in enhancing osteoclastic activity and decreasing osteoblast population, BSP overexpression leads to an uncoupling of bone formation and resorption, which in turn results in osteopenia and mild dwarfism in mice. These findings are expected to help the development of therapies to metabolic bone diseases characterized by high serum level of BSP.

Overexpression of Bone Sialoprotein Leads to an Uncoupling of Bone Formation and Bone Resorption in Mice

PubMed Central

Valverde, Paloma; Zhang, Jin; Fix, Amanda; Zhu, Ji; Ma, Wenli; Tu, Qisheng; Chen, Jake

2008-01-01

The purpose of this study was to determine the effects of bone sialoprotein (BSP) overexpression in bone metabolism in vivo by using a homozygous transgenic mouse line that constitutively overexpresses mouse BSP cDNA driven by the cytomegalovirus (CMV) promoter. CMV-BSP transgenic (TG) mice and wildtype mice were weighed, and their length, BMD, and trabecular bone volume were measured. Serum levels of RANKL, osteocalcin, osteoprotegerin (OPG), TRACP5b, and PTH were determined. Bone histomorphometry, von Kossa staining, RT-PCR analysis, Western blot, MTS assay, in vitro mineralization assay, and TRACP staining were also performed to delineate phenotypes of this transgenic mouse line. Compared with wildtype mice, adult TG mice exhibit mild dwarfism, lower values of BMD, and lower trabecular bone volume. TG mice serum contained increased calcium levels and decreased PTH levels, whereas the levels of phosphorus and magnesium were within normal limits. TG mice serum also exhibited lower levels of osteoblast differentiation markers and higher levels of markers, indicating osteoclastic activity and bone resorption. H&E staining, TRACP staining, and bone histomorphometry showed that adult TG bones were thinner and the number of giant osteoclasts in TG mice was higher, whereas there were no significant alterations in osteoblast numbers between TG mice and WT mice. Furthermore, the vertical length of the hypertrophic zone in TG mice was slightly enlarged. Moreover, ex vivo experiments indicated that overexpression of BSP decreased osteoblast population and increased osteoclastic activity. Partly because of its effects in enhancing osteoclastic activity and decreasing osteoblast population, BSP overexpression leads to an uncoupling of bone formation and resorption, which in turn results in osteopenia and mild dwarfism in mice. These findings are expected to help the development of therapies to metabolic bone diseases characterized by high serum level of BSP. PMID:18597627

Osteoclast TGF-β Receptor Signaling Induces Wnt1 Secretion and Couples Bone Resorption to Bone Formation

PubMed Central

Weivoda, Megan M; Ruan, Ming; Pederson, Larry; Hachfeld, Christine; Davey, Rachel A; Zajac, Jeffrey D; Westendorf, Jennifer J; Khosla, Sundeep; Oursler, Merry Jo

2016-01-01

Osteoblast-mediated bone formation is coupled to osteoclast-mediated bone resorption. These processes become uncoupled with age, leading to increased risk for debilitating fractures. Therefore, understanding how osteoblasts are recruited to sites of resorption is vital to treating age-related bone loss. Osteoclasts release and activate TGF-β from the bone matrix. Here we show that osteoclastspecific inhibition of TGF-β receptor signaling in mice results in osteopenia due to reduced osteoblast numbers with no significant impact on osteoclast numbers or activity. TGF-β induced osteoclast expression of Wnt1, a protein crucial to normal bone formation, and this response was blocked by impaired TGF-β receptor signaling. Osteoclasts in aged murine bones had lower TGF-β signaling and Wnt1 expression in vivo. Ex vivo stimulation of osteoclasts derived from young or old mouse bone marrow macrophages showed no difference in TGF-β–induced Wnt1 expression. However, young osteoclasts expressed reduced Wnt1 when cultured on aged mouse bone chips compared to young mouse bone chips, consistent with decreased skeletal TGF-β availability with age. Therefore, osteoclast responses to TGF-β are essential for coupling bone resorption to bone formation, and modulating this pathway may provide opportunities to treat age-related bone loss. PMID:26108893

Protective effect of hawthorn extract against genotoxicity induced by cyclophosphamide in mouse bone marrow cells.

PubMed

Hosseinimehr, Seyed Jalal; Azadbakht, Mohammad; Abadi, Atefeh Jahan

2008-01-01

The preventive effect of hawthorn (Crataegus microphylla) fruit extract was investigated in mouse bone marrow cells against genotoxicity induced by cyclophosphamide. Mice were orally (gavages) pretreated with solutions of hawthorn extract which was prepared at five different doses (25, 50, 100, 200 and 400mg/kg b.w.) for seven consecutive days. Mice were injected intraperitoneally on the seventh day with cyclophosphamide (50mg/kg b.w.) and killed after 24h for the evaluation of micronucleated polychromatic erythrocytes (MnPCEs) and the ratio of PCE/(PCE+NCE) (polychromatic erythrocyte/polychromatic erythrocyte+normochromatic erythrocyte). All of five doses of extract significantly reduced MnPCEs induced by cyclophosphamide (P<0.0001). Hawthorn extract at dose 100mg/kg b.w. reduced MnPCEs 2.5 time and also completely normalized PCE/(PCE+NCE) ratio. Hawthorn extract exhibited concentration-dependent antioxidant activity on 1,1-diphenyl-2-picryl hydrazyl free radical. Hawthorn contains high amounts of phenolic compounds; the HPLC analysis showed that it contained chlorogenic acid, epicatechin and hyperoside. It is obvious that hawthorn, particularly flavonoids constituents with antioxidative activity, reduced the oxidative stress and genotoxicity induced by cyclophosphamide in mouse bone marrow cells. Copyright © 2007 Elsevier B.V. All rights reserved.

Survival of Free and Encapsulated Human and Rat Islet Xenografts Transplanted into the Mouse Bone Marrow

PubMed Central

Meier, Raphael P. H.; Seebach, Jörg D.; Morel, Philippe; Mahou, Redouan; Borot, Sophie; Giovannoni, Laurianne; Parnaud, Geraldine; Montanari, Elisa; Bosco, Domenico; Wandrey, Christine; Berney, Thierry; Bühler, Leo H.; Muller, Yannick D.

2014-01-01

Bone marrow was recently proposed as an alternative and potentially immune-privileged site for pancreatic islet transplantation. The aim of the present study was to assess the survival and rejection mechanisms of free and encapsulated xenogeneic islets transplanted into the medullary cavity of the femur, or under the kidney capsule of streptozotocin-induced diabetic C57BL/6 mice. The median survival of free rat islets transplanted into the bone marrow or under the kidney capsule was 9 and 14 days, respectively, whereas that of free human islets was shorter, 7 days (bone marrow) and 10 days (kidney capsule). Infiltrating CD8+ T cells and redistributed CD4+ T cells, and macrophages were detected around the transplanted islets in bone sections. Recipient mouse splenocytes proliferated in response to donor rat stimulator cells. One month after transplantation under both kidney capsule or into bone marrow, encapsulated rat islets had induced a similar degree of fibrotic reaction and still contained insulin positive cells. In conclusion, we successfully established a small animal model for xenogeneic islet transplantation into the bone marrow. The rejection of xenogeneic islets was associated with local and systemic T cell responses and macrophage recruitment. Although there was no evidence for immune-privilege, the bone marrow may represent a feasible site for encapsulated xenogeneic islet transplantation. PMID:24625569

Stromal derived factor-1 regulates bone morphogenetic protein 2-induced osteogenic differentiation of primary mesenchymal stem cells

PubMed Central

Hosogane, Naobumi; Huang, Zhiping; Rawlins, Bernard A.; Liu, Xia; Boachie-Adjei, Oheneba; Boskey, Adele L.; Zhu, Wei

2010-01-01

Stromal derived factor-1 (SDF-1) is a chemokine signaling molecule that binds to its transmembrane receptor CXC chemokine receptor-4 (CXCR4). While we previously detected that SDF-1 was co-required with bone morphogenetic protein 2 (BMP2) for differentiating mesenchymal C2C12 cells into osteoblastic cells, it is unknown whether SDF-1 is similarly involved in the osteogenic differentiation of mesenchymal stem cells (MSCs). Therefore, here we examined the role of SDF-1 signaling during BMP2-induced osteogenic differentiation of primary MSCs that were derived from human and mouse bone marrow. Our data showed that blocking of the SDF-1/CXCR4 signal axis or adding SDF-1 protein to MSCs significantly affected BMP2-induced alkaline phosphatase (ALP) activity and osteocalcin (OCN) synthesis, markers of preosteoblasts and mature osteoblasts, respectively. Moreover, disrupting the SDF-1 signaling impaired bone nodule mineralization during terminal differentiation of MSCs. Furthermore, we detected that blocking of the SDF-1 signaling inhibited the BMP2-induced early expression of Runt-related factor-2 (Runx2) and osterix (Osx), two “master” regulators of osteogenesis, and the SDF-1 effect was mediated via intracellular Smad and Erk activation. In conclusion, our results demonstrated a regulatory role of SDF-1 in BMP2-induced osteogenic differentiation of MSCs, as perturbing the SDF-1 signaling affected the differentiation of MSCs towards osteoblastic cells in response to BMP2 stimulation. These data provide novel insights into molecular mechanisms underlying MSC osteogenesis, and will contribute to the development of MSC therapies for enhancing bone formation and regeneration in broad orthopaedic situations. PMID:20362069

Dura Mater Stimulates Human Adipose-Derived Stromal Cells to Undergo Bone Formation in Mouse Calvarial Defects

PubMed Central

Levi, Benjamin; Nelson, Emily R.; Li, Shuli; James, Aaron W.; Hyun, Jeong S.; Montoro, Daniel T.; Lee, Min; Glotzbach, Jason P.; Commons, George W.; Longaker, Michael T.

2015-01-01

Human adipose-derived stromal cells (hASCs) have a proven capacity to aid in osseous repair of calvarial defects. However, the bone defect microenvironment necessary for osseous healing is not fully understood. In this study, we postulated that the cell-cell interaction between engrafted ASCs and host dura mater (DM) cells is critical for the healing of calvarial defects. hASCs were engrafted into critical sized calvarial mouse defects. The DM-hASC interaction was manipulated surgically by DM removal or by insertion of a semipermeable or nonpermeable membrane between DM and hASCs. Radiographic, histologic, and gene expression analyses were performed. Next, the hASC-DM interaction is assessed by conditioned media (CM) and coculture assays. Finally, bone morphogenetic protein (BMP) signaling from DM was investigated in vivo using novel BMP-2 and anti-BMP-2/4 slow releasing scaffolds. With intact DM, osseous healing occurs both from host DM and engrafted hASCs. Interference with the DM-hASC interaction dramatically reduced calvarial healing with abrogated BMP-2–Smad-1/5 signaling. Using CM and coculture assays, mouse DM cells stimulated hASC osteogenesis via BMP signaling. Through in vivo manipulation of the BMP-2 pathway, we found that BMP-2 plays an important role in DM stimulation of hASC osteogenesis in the context of calvarial bone healing. BMP-2 supplementation to a defect with disrupted DM allowed for bone formation in a nonhealing defect. DM is an osteogenic cell type that both participates in and stimulates osseous healing in a hASC-engrafted calvarial defect. Furthermore, DM-derived BMP-2 paracrine stimulation appears to play a key role for hASC mediated repair. PMID:21656608

Tumor-targeting Salmonella typhimurium A1-R inhibits human prostate cancer experimental bone metastasis in mouse models.

PubMed

Toneri, Makoto; Miwa, Shinji; Zhang, Yong; Hu, Cameron; Yano, Shuya; Matsumoto, Yasunori; Bouvet, Michael; Nakanishi, Hayao; Hoffman, Robert M; Zhao, Ming

2015-10-13

Bone metastasis is a frequent occurrence in prostate cancer patients and often is lethal. Zoledronic acid (ZOL) is often used for bone metastasis with limited efficacy. More effective models and treatment methods are required to improve the outcome of prostate cancer patients. In the present study, the effects of tumor-targeting Salmonella typhimurium A1-R were analyzed in vitro and in vivo on prostate cancer cells and experimental bone metastasis. Both ZOL and S. typhimurium A1-R inhibited the growth of PC-3 cells expressing red fluorescent protien in vitro. To investigate the efficacy of S. typhimurium A1-R on prostate cancer experimental bone metastasis, we established models of both early and advanced stage bone metastasis. The mice were treated with ZOL, S. typhimurium A1-R, and combination therapy of both ZOL and S. typhimurium A1-R. ZOL and S. typhimurium A1-R inhibited the growth of solitary bone metastases. S. typhimurium A1-R treatment significantly decreased bone metastasis and delayed the appearance of PC-3 bone metastases of multiple mouse models. Additionally, S. typhimurium A1-R treatment significantly improved the overall survival of the mice with multiple bone metastases. The results of the present study indicate that S. typhimurium A1-R is useful to prevent and inhibit prostate cancer bone metastasis and has potential for future clinical use in the adjuvant setting.

Host cell recruitment patterns by bone morphogenetic protein-2 releasing hyaluronic acid hydrogels in a mouse subcutaneous environment.

PubMed

Todeschi, Maria R; El Backly, Rania M; Varghese, Oommen P; Hilborn, Jöns; Cancedda, Ranieri; Mastrogiacomo, Maddalena

2017-07-01

This study aimed to identify host cell recruitment patterns in a mouse model in response to rhBMP-2 releasing hyaluronic acid hydrogels and influence of added nano-hydroxyapatite particles on rhBMP-2 release and pattern of bone formation. Implanted gels were retrieved after implantation and cells were enzymatically dissociated for flow cytometric analysis. Percentages of macrophages, progenitor endothelial cells and putative mesenchymal stem cells were measured. Implants were evaluated for BMP-2 release by ELISA and by histology to monitor tissue formation. Hyaluronic acid+BMP-2 gels influenced the inflammatory response in the bone healing microenvironment. Host-derived putative mesenchymal stem cells were major contributors. Addition of hydroxyapatite nanoparticles modified the release pattern of rhBMP-2, resulting in enhanced bone formation.

Mushroom Extracts Decrease Bone Resorption and Improve Bone Formation.

PubMed

Erjavec, Igor; Brkljacic, Jelena; Vukicevic, Slobodan; Jakopovic, Boris; Jakopovich, Ivan

2016-01-01

Mushroom extracts have shown promising effects in the treatment of cancer and various chronic diseases. Osteoporosis is considered one of the most widespread chronic diseases, for which currently available therapies show mixed results. In this research we investigated the in vitro effects of water extracts of the culinary-medicinal mushrooms Trametes versicolor, Grifola frondosa, Lentinus edodes, and Pleurotus ostreatus on a MC3T3-E1 mouse osteoblast-like cell line, primary rat osteoblasts, and primary rat osteoclasts. In an animal osteoporosis model, rats were ovariectomized and then fed 2 mushroom blends of G. frondosa and L. edodes for 42 days. Bone loss was monitored using densitometry (dual-energy X-ray absorptiometry) and micro computed tomography. In the concentration test, mushroom extracts showed no toxic effect on MC3T3-E1 cells; a dose of 24 µg/mL showed the most proliferative effect. Mushroom extracts of T. versicolor, G. frondosa, and L. edodes inhibited osteoclast activity, whereas the extract of L. edodes increased osteoblast mineralization and the production of osteocalcin, a specific osteoblastic marker. In animals, mushroom extracts did not prevent trabecular bone loss in the long bones. However, we show for the first time that the treatment with a combination of extracts from L. edodes and G. frondosa significantly reduced trabecular bone loss at the lumbar spine. Inhibitory properties of extracts from L. edodes on osteoclasts and the promotion of osteoblasts in vitro, together with the potential to decrease lumbar spine bone loss in an animal osteoporosis model, indicate that medicinal mushroom extracts can be considered as a preventive treatment and/or a supplement to pharmacotherapy to enhance its effectiveness and ameliorate its harmful side effects.

Primary homologies of the circumorbital bones of snakes.

PubMed

Palci, Alessandro; Caldwell, Michael W

2013-09-01

Some snakes have two circumorbital ossifications that in the current literature are usually referred to as the postorbital and supraorbital. We review the arguments that have been proposed to justify this interpretation and provide counter-arguments that reject those conjectures of primary homology based on the observation of 32 species of lizards and 81 species of snakes (both extant and fossil). We present similarity arguments, both topological and structural, for reinterpretation of the primary homologies of the dorsal and posterior orbital ossifications of snakes. Applying the test of similarity, we conclude that the posterior orbital ossification of snakes is topologically consistent as the homolog of the lacertilian jugal, and that the dorsal orbital ossification present in some snakes (e.g., pythons, Loxocemus, and Calabaria) is the homolog of the lacertilian postfrontal. We therefore propose that the terms postorbital and supraorbital should be abandoned as reference language for the circumorbital bones of snakes, and be replaced with the terms jugal and postfrontal, respectively. The primary homology claim for the snake "postorbital" fails the test of similarity, while the term "supraorbital" is an unnecessary and inaccurate application of the concept of a neomorphic ossification, for an element that passes the test of similarity as a postfrontal. This reinterpretation of the circumorbital bones of snakes is bound to have important repercussions for future phylogenetic analyses and consequently for our understanding of the origin and evolution of snakes. Copyright © 2013 Wiley Periodicals, Inc.

Effects of O-methylated (-)-epigallocatechin gallate (EGCG) on LPS-induced osteoclastogenesis, bone resorption, and alveolar bone loss in mice.

PubMed

Tominari, Tsukasa; Ichimaru, Ryota; Yoshinouchi, Shosei; Matsumoto, Chiho; Watanabe, Kenta; Hirata, Michiko; Grundler, Florian M W; Inada, Masaki; Miyaura, Chisato

2017-12-01

(-)-Epigallocatechin-3- O -gallate (EGCG), present in green tea, exhibits antioxidant and antiallergy effects. EGCG3″Me, a 3- O -methylated derivative of EGCG, has been reported to show similar biological functions; the inhibitory activity of EGCG3″Me in a mouse allergy model was more potent than that of EGCG, probably due to the efficiency of absorption from the intestine. However, the functional potency of these EGCGs is controversial in each disease model. We previously observed that EGCG suppressed inflammatory bone resorption and prevented alveolar bone loss in a mouse model of periodontosis. In this study, we examined the role of EGCG3″Me in bone resorption using a mouse model of periodontitis. Lipopolysaccharide (LPS)-induced osteoclast formation was suppressed by adding EGCG3″Me to cocultures of osteoblasts and bone marrow cells, and LPS-induced bone resorption was also inhibited by EGCG3″Me in calvarial organ cultures. EGCG3″Me acted on osteoblasts and suppressed prostaglandin E (PGE) production, which is critical for inflammatory bone resorption, by inhibiting the expression of COX-2 and mPGES-1, key enzymes for PGE synthesis. In osteoclast precursor macrophages, EGCG3″Me suppressed RANKL-dependent differentiation into mature osteoclasts. In a mouse model of periodontitis, LPS-induced bone resorption was suppressed by EGCG3″Me in organ culture of mouse alveolar bone, and the alveolar bone loss was further attenuated by the treatment of EGCG3″Me in the lower gingiva in vivo . EGCG3″Me may be a potential natural compound for the protection of inflammatory bone loss in periodontitis.

Mechanical Unloading of Mouse Bone in Microgravity Significantly Alters Cell Cycle Gene Set Expression

NASA Astrophysics Data System (ADS)

Blaber, Elizabeth; Dvorochkin, Natalya; Almeida, Eduardo; Kaplan, Warren; Burns, Brnedan

2012-07-01

unloading in spaceflight, we conducted genome wide microarray analysis of total RNA isolated from the mouse pelvis. Specifically, 16 week old mice were subjected to 15 days spaceflight onboard NASA's STS-131 space shuttle mission. The pelvis of the mice was dissected, the bone marrow was flushed and the bones were briefly stored in RNAlater. The pelvii were then homogenized, and RNA was isolated using TRIzol. RNA concentration and quality was measured using a Nanodrop spectrometer, and 0.8% agarose gel electrophoresis. Samples of cDNA were analyzed using an Affymetrix GeneChip\\S Gene 1.0 ST (Sense Target) Array System for Mouse and GenePattern Software. We normalized the ST gene arrays using Robust Multichip Average (RMA) normalization, which summarizes perfectly matched spots on the array through the median polish algorithm, rather than normalizing according to mismatched spots. We also used Limma for statistical analysis, using the BioConductor Limma Library by Gordon Smyth, and differential expression analysis to identify genes with significant changes in expression between the two experimental conditions. Finally we used GSEApreRanked for Gene Set Enrichment Analysis (GSEA), with Kolmogorov-Smirnov style statistics to identify groups of genes that are regulated together using the t-statistics derived from Limma. Preliminary results show that 6,603 genes expressed in pelvic bone had statistically significant alterations in spaceflight compared to ground controls. These prominently included cell cycle arrest molecules p21, and p18, cell survival molecule Crbp1, and cell cycle molecules cyclin D1, and Cdk1. Additionally, GSEA results indicated alterations in molecular targets of cyclin D1 and Cdk4, senescence pathways resulting from abnormal laminin maturation, cell-cell contacts via E-cadherin, and several pathways relating to protein translation and metabolism. In total 111 gene sets out of 2,488, about 4%, showed statistically significant set alterations. These

The inward rectifier potassium channel Kir2.1 is expressed in mouse neutrophils from bone marrow and liver.

PubMed

Masia, Ricard; Krause, Daniela S; Yellen, Gary

2015-02-01

Neutrophils are phagocytic cells that play a critical role in innate immunity by destroying bacterial pathogens. Channels belonging to the inward rectifier potassium channel subfamily 2 (Kir2 channels) have been described in other phagocytes (monocytes/macrophages and eosinophils) and in hematopoietic precursors of phagocytes. Their physiological function in these cells remains unclear, but some evidence suggests a role in growth factor-dependent proliferation and development. Expression of functional Kir2 channels has not been definitively demonstrated in mammalian neutrophils. Here, we show by RT-PCR that neutrophils from mouse bone marrow and liver express mRNA for the Kir2 subunit Kir2.1 but not for other subunits (Kir2.2, Kir2.3, and Kir2.4). In electrophysiological experiments, resting (unstimulated) neutrophils from mouse bone marrow and liver exhibit a constitutively active, external K(+)-dependent, strong inwardly rectifying current that constitutes the dominant current. The reversal potential is dependent on the external K(+) concentration in a Nernstian fashion, as expected for a K(+)-selective current. The current is not altered by changes in external or internal pH, and it is blocked by Ba(2+), Cs(+), and the Kir2-selective inhibitor ML133. The single-channel conductance is in agreement with previously reported values for Kir2.1 channels. These properties are characteristic of homomeric Kir2.1 channels. Current density in short-term cultures of bone marrow neutrophils is decreased in the absence of growth factors that are important for neutrophil proliferation [granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF)]. These results demonstrate that mouse neutrophils express functional Kir2.1 channels and suggest that these channels may be important for neutrophil function, possibly in a growth factor-dependent manner. Copyright © 2015 the American Physiological Society.

Ovariectomy-Induced Osteopenia Influences the Middle and Late Periods of Bone Healing in a Mouse Femoral Osteotomy Model.

PubMed

Pang, Jian; Ye, Meina; Gu, Xinfeng; Cao, Yuelong; Zheng, Yuxin; Guo, Hailing; Zhao, Yongfang; Zhan, Hongsheng; Shi, Yinyu

2015-08-01

It is known that bone healing is delayed in the presence of osteoporosis in humans. However, due to the complexities of the healing of osteoporotic fractures, animal models may be more appropriate for studying the effects of osteoporosis in more detail and for testing drugs on the fracture repair process. The purpose of this study was to investigate the influence of ovariectomy-induced osteopenia in bone healing in an open femoral osteotomy model, and to test the feasibility of this model for evaluating the healing process under osteopenic conditions. Ovariectomized (OVX) mouse models were employed to assess the effects of osteopenia on fracture healing, A mid-shaft femur osteotomy model was also established 3 weeks after ovariectomy as an osteopenic fracture group (OVX group). Femurs were then harvested at 2 weeks and 6 weeks after fracture for X-ray radiography, micro-computed tomography (micro-CT), histology, and biomechanical analysis. A sham-operated group (sham group) was used for comparison. The OVX mice had significantly lower bone volume density (BVF), volumetric bone mineral density (vBMD), and tissue mineral density (TMD) in the fracture calluses at 6 weeks (p<0.05), and similar trend was observed in 2 weeks. Additionally, larger calluses in OVX animals were observed via micro-CT and X-ray, but these did not result in better healing outcomes, as determined by biomechanical test at 6 weeks. Histological images of the healing fractures in the OVX mice found hastening of broken end resorption and delay of hard callus remodeling. The impaired biomechanical measurements in the OVX group (p<0.05) were consistent with micro-CT measurements and radiographic scoring, which also indicated delay in fracture healing of the OVX group. This study provided evidence that ovariectomy-induced osteopenia impair the middle and late bone healing process. These data also supported the validity of the mouse femoral osteotomy model in evaluating the process of bone healing

Automatic detection and quantitative analysis of cells in the mouse primary motor cortex

NASA Astrophysics Data System (ADS)

Meng, Yunlong; He, Yong; Wu, Jingpeng; Chen, Shangbin; Li, Anan; Gong, Hui

2014-09-01

Neuronal cells play very important role on metabolism regulation and mechanism control, so cell number is a fundamental determinant of brain function. Combined suitable cell-labeling approaches with recently proposed three-dimensional optical imaging techniques, whole mouse brain coronal sections can be acquired with 1-μm voxel resolution. We have developed a completely automatic pipeline to perform cell centroids detection, and provided three-dimensional quantitative information of cells in the primary motor cortex of C57BL/6 mouse. It involves four principal steps: i) preprocessing; ii) image binarization; iii) cell centroids extraction and contour segmentation; iv) laminar density estimation. Investigations on the presented method reveal promising detection accuracy in terms of recall and precision, with average recall rate 92.1% and average precision rate 86.2%. We also analyze laminar density distribution of cells from pial surface to corpus callosum from the output vectorizations of detected cell centroids in mouse primary motor cortex, and find significant cellular density distribution variations in different layers. This automatic cell centroids detection approach will be beneficial for fast cell-counting and accurate density estimation, as time-consuming and error-prone manual identification is avoided.

Bat-mouse bone marrow chimera: a novel animal model for dissecting the uniqueness of the bat immune system.

PubMed

Yong, Kylie Su Mei; Ng, Justin Han Jia; Her, Zhisheng; Hey, Ying Ying; Tan, Sue Yee; Tan, Wilson Wei Sheng; Irac, Sergio Erdal; Liu, Min; Chan, Xue Ying; Gunawan, Merry; Foo, Randy Jee Hiang; Low, Dolyce Hong Wen; Mendenhall, Ian Hewitt; Chionh, Yok Teng; Dutertre, Charles-Antoine; Chen, Qingfeng; Wang, Lin-Fa

2018-03-16

Bats are an important animal model with long lifespans, low incidences of tumorigenesis and an ability to asymptomatically harbour pathogens. Currently, in vivo studies of bats are hampered due to their low reproduction rates. To overcome this, we transplanted bat cells from bone marrow (BM) and spleen into an immunodeficient mouse strain NOD-scid IL-2R -/- (NSG), and have successfully established stable, long-term reconstitution of bat immune cells in mice (bat-mice). Immune functionality of our bat-mouse model was demonstrated through generation of antigen-specific antibody response by bat cells following immunization. Post-engraftment of total bat BM cells and splenocytes, bat immune cells survived, expanded and repopulated the mouse without any observable clinical abnormalities. Utilizing bat's remarkable immunological functions, this novel model has a potential to be transformed into a powerful platform for basic and translational research.

Early-onset type 2 diabetes impairs skeletal acquisition in the male TALLYHO/JngJ mouse.

PubMed

Devlin, M J; Van Vliet, M; Motyl, K; Karim, L; Brooks, D J; Louis, L; Conlon, C; Rosen, C J; Bouxsein, M L

2014-10-01

Type 2 diabetes (T2D) incidence in adolescents is rising and may interfere with peak bone mass acquisition. We tested the effects of early-onset T2D on bone mass, microarchitecture, and strength in the TALLYHO/JngJ mouse, which develops T2D by 8 weeks of age. We assessed metabolism and skeletal acquisition in male TALLYHO/JngJ and SWR/J controls (n = 8-10/group) from 4 weeks to 8 and 17 weeks of age. Tallyho mice were obese; had an approximately 2-fold higher leptin and percentage body fat; and had lower bone mineral density vs SWR at all time points (P < .03 for all). Tallyho had severe deficits in distal femur trabecular bone volume fraction (-54%), trabecular number (-27%), and connectivity density (-82%) (P < .01 for all). Bone formation was higher in Tallyho mice at 8 weeks but lower by 17 weeks of age vs SWR despite similar numbers of osteoblasts. Bone marrow adiposity was 7- to 50-fold higher in Tallyho vs SWR. In vitro, primary bone marrow stromal cell differentiation into osteoblast and adipocyte lineages was similar in SWR and Tallyho, suggesting skeletal deficits were not due to intrinsic defects in Tallyho bone-forming cells. These data suggest the Tallyho mouse might be a useful model to study the skeletal effects of adolescent T2D.

Isolation of Primary Mouse Trophoblast Cells and Trophoblast Invasion Assay

PubMed Central

Pennington, Kathleen A.; Schlitt, Jessica M.; Schulz, Laura C.

2012-01-01

The placenta is responsible for the transport of nutrients, gasses and growth factors to the fetus, as well as the elimination of wastes. Thus, defects in placental development have important consequences for the fetus and mother, and are a major cause of embryonic lethality. The major cell type of the fetal portion of the placenta is the trophoblast. Primary mouse placental trophoblast cells are a useful tool for studying normal and abnormal placental development, and unlike cell lines, may be isolated and used to study trophoblast at specific stages of pregnancy. In addition, primary cultures of trophoblast from transgenic mice may be used to study the role of particular genes in placental cells. The protocol presented here is based on the description by Thordarson et al.1, in which a percoll gradient is used to obtain a relatively pure trophoblast cell population from isolated mouse placentas. It is similar to the more widely used methods for human trophoblast cell isolation2-3. Purity may be assessed by immunocytochemical staining of the isolated cells for cytokeratin 74. Here, the isolated cells are then analyzed using a matrigel invasion assay to assess trophoblast invasiveness in vitro5-6. The invaded cells are analyzed by immunocytochemistry and stained for counting. PMID:22257865

Lithium treatment elongates primary cilia in the mouse brain and in cultured cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyoshi, Ko, E-mail: miyoshi@cc.okayama-u.ac.jp; Kasahara, Kyosuke; Miyazaki, Ikuko

2009-10-30

The molecular mechanisms underlying the therapeutic effects of lithium, a first-line antimanic mood stabilizer, have not yet been fully elucidated. Treatment of the algae Chlamydomonas reinhardtii with lithium has been shown to induce elongation of their flagella, which are analogous structures to vertebrate cilia. In the mouse brain, adenylyl cyclase 3 (AC3) and certain neuropeptide receptors colocalize to the primary cilium of neuronal cells, suggesting a chemosensory function for the primary cilium in the nervous system. Here we show that lithium treatment elongates primary cilia in the mouse brain and in cultured cells. Brain sections from mice chronically fed withmore » Li{sub 2}CO{sub 3} were subjected to immunofluorescence study. Primary cilia carrying both AC3 and the receptor for melanin-concentrating hormone (MCH) were elongated in the dorsal striatum and nucleus accumbens of lithium-fed mice, as compared to those of control animals. Moreover, lithium-treated NIH3T3 cells and cultured striatal neurons exhibited elongation of the primary cilia. The present results provide initial evidence that a psychotropic agent can affect ciliary length in the central nervous system, and furthermore suggest that lithium exerts its therapeutic effects via the upregulation of cilia-mediated MCH sensing. These findings thus contribute novel insights into the pathophysiology of bipolar mood disorder and other psychiatric diseases.« less

Resveratrol Inhibited Hydroquinone-Induced Cytotoxicity in Mouse Primary Hepatocytes

PubMed Central

Wang, Da-Hong; Ootsuki, Yoshie; Fujita, Hirofumi; Miyazaki, Masahiro; Yie, Qinxia; Tsutsui, Ken; Sano, Kuniaki; Masuoka, Noriyoshi; Ogino, Keiki

2012-01-01

Hydroquinone (1,4-benzenediol) has been widely used in clinical situations and the cosmetic industry because of its depigmenting effects. Most skin-lightening hydroquinone creams contain 4%–5% hydroquinone. We have investigated the role of resveratrol in prevention of hydroquinone induced cytotoxicity in mouse primary hepatocytes. We found that 400 µM hydroquinone exposure alone induced apoptosis of the cells and also resulted in a significant drop of cell viability compared with the control, and pretreatment of resveratrol to a final concentration of 0.5 mM 1 h before hydroquinone exposure did not show a significant improvement in the survival rate of the hepatocytes, however, relatively higher concentrations of resveratrol (≥1 mM) inhibited apoptosis of the mouse primary hepatocytes and increased cell viability in a dose-dependent manner, and in particular the survival rate of the hepatocytes was recovered from 28% to near 100% by 5 mM resveratrol. Interestingly, pretreatment with resveratrol for longer time (24 h), even in very low concentrations (50 µM, 100 µM), blocked the damage of hydroquinone to the cells. We also observed that resveratrol pretreatment suppressed hydroquinone-induced expression of cytochrome P450 2E1 mRNA dose-dependently. The present study suggests that resveratrol protected the cells against hydroquinone-induced toxicity through its antioxidant function and possibly suppressive effect on the expression of cytochrome P450 2E1. PMID:23202692

Resveratrol inhibited hydroquinone-induced cytotoxicity in mouse primary hepatocytes.

PubMed

Wang, Da-Hong; Ootsuki, Yoshie; Fujita, Hirofumi; Miyazaki, Masahiro; Yie, Qinxia; Tsutsui, Ken; Sano, Kuniaki; Masuoka, Noriyoshi; Ogino, Keiki

2012-09-19

Hydroquinone (1,4-benzenediol) has been widely used in clinical situations and the cosmetic industry because of its depigmenting effects. Most skin-lightening hydroquinone creams contain 4%-5% hydroquinone. We have investigated the role of resveratrol in prevention of hydroquinone induced cytotoxicity in mouse primary hepatocytes. We found that 400 µM hydroquinone exposure alone induced apoptosis of the cells and also resulted in a significant drop of cell viability compared with the control, and pretreatment of resveratrol to a final concentration of 0.5 mM 1 h before hydroquinone exposure did not show a significant improvement in the survival rate of the hepatocytes, however, relatively higher concentrations of resveratrol (≥1 mM) inhibited apoptosis of the mouse primary hepatocytes and increased cell viability in a dose-dependent manner, and in particular the survival rate of the hepatocytes was recovered from 28% to near 100% by 5 mM resveratrol. Interestingly, pretreatment with resveratrol for longer time (24 h), even in very low concentrations (50 µM, 100 µM), blocked the damage of hydroquinone to the cells. We also observed that resveratrol pretreatment suppressed hydroquinone-induced expression of cytochrome P450 2E1 mRNA dose-dependently. The present study suggests that resveratrol protected the cells against hydroquinone-induced toxicity through its antioxidant function and possibly suppressive effect on the expression of cytochrome P450 2E1.

Cytogenetic effects of sildenafil citrate (Viagra) on SWR/J mouse bone marrow cells.

PubMed

Abou-Tarboush, Faisal Mohamed; Abdel-Samad, Mohamed Fathy

2010-10-01

The present study was conducted to investigate the cytogenetic effects of sildenafil citrate in SWR/J mouse bone marrow cells. Thirty-six males and 36 females were used and divided into four groups. Each group contained 18 animals (9 males and 9 females), weighing 30-35 g. These animals were orally administered with a single dose of 13, 26 or 40 mg/kg sildenafil citrate solution. A control group received normal saline in an identical condition. The animals were sacrificed at 12, 24 or 48 h, after the treatment. Chromosome aberrations were investigated in 50 metaphases per animal. No significant differences in the percentages of mitotic indices or in the frequencies of chromosome aberrations were observed between treated male and female mice at any doses or at any time intervals used, therefore, data from the two sexes were pooled when analyzed statistically. No significant (p < 0.05) differences in the percentages of mitotic indices or in the frequencies of chromosome aberrations were observed between sildenafil citrate-treated groups and the control group at any doses or at any time intervals used. However, the percentages of centromeric adhesions increased significantly (p < 0.01) in treated groups as compared with the control group at all doses and at all time intervals used. In conclusion, the results of the present study suggest that sildenafil citrate does not have cytogenetic effects on mouse bone marrow cells, but the centromeric adhesions induced by this drug need further studies to confirm them and to investigate the possible mechanism(s) responsible for such effect.

Cytogenetic effects of sildenafil citrate (Viagra) on SWR/J mouse bone marrow cells

PubMed Central

Abou-Tarboush, Faisal Mohamed; Abdel-Samad, Mohamed Fathy

2010-01-01

The present study was conducted to investigate the cytogenetic effects of sildenafil citrate in SWR/J mouse bone marrow cells. Thirty-six males and 36 females were used and divided into four groups. Each group contained 18 animals (9 males and 9 females), weighing 30–35 g. These animals were orally administered with a single dose of 13, 26 or 40 mg/kg sildenafil citrate solution. A control group received normal saline in an identical condition. The animals were sacrificed at 12, 24 or 48 h, after the treatment. Chromosome aberrations were investigated in 50 metaphases per animal. No significant differences in the percentages of mitotic indices or in the frequencies of chromosome aberrations were observed between treated male and female mice at any doses or at any time intervals used, therefore, data from the two sexes were pooled when analyzed statistically. No significant (p < 0.05) differences in the percentages of mitotic indices or in the frequencies of chromosome aberrations were observed between sildenafil citrate-treated groups and the control group at any doses or at any time intervals used. However, the percentages of centromeric adhesions increased significantly (p < 0.01) in treated groups as compared with the control group at all doses and at all time intervals used. In conclusion, the results of the present study suggest that sildenafil citrate does not have cytogenetic effects on mouse bone marrow cells, but the centromeric adhesions induced by this drug need further studies to confirm them and to investigate the possible mechanism(s) responsible for such effect. PMID:23961094

Facial growth and development in unilateral cleft lip and palate: comparison between secondary alveolar bone grafting and primary periosteoplasty.

PubMed

Cagáňová, Veronika; Borský, Jiří; Smahel, Zbyněk; Velemínská, Jana

2014-01-01

To describe the effect of secondary alveolar bone grafting in patients with unilateral cleft lip and palate by comparison with a sample of patients who have undergone primary periosteoplasty. Cephalometric analysis of lateral x-ray films in a retrospective semilongitudinal study. Lateral x-ray films of 18 secondary alveolar bone grafting patients and 48 primary periosteoplasty patients at 10 years of age and again at 15 years of age. The treatment of secondary alveolar bone grafting patients included lip repair according to Tennison, palatoplasty including retropositioning, pharyngeal flap surgery, and secondary alveolar bone grafting. The lips of primary periosteoplasty patient were repaired using the methods of Tennison and Veau, followed by primary periosteoplasty, palatoplasty including retropositioning, and pharyngeal flap surgery. Lateral radiographs were assessed using classical morphometry. There were few significant differences at 10 years of age between the secondary alveolar bone grafting and primary periosteoplasty patients. At 15 years of age, there were several significant differences. Compared with primary periosteoplasty patients, subsequent development in patients who had undergone secondary alveolar bone grafting was characterized by a significantly better position of the upper and lower dentoalveolar components in relation to the facial plane, a higher increase in the global convexity of the soft profile, a significantly better maxillary inclination, and a more favorable development of vertical intermaxillary relationships. Craniofacial development in secondary alveolar bone grafting patients was better than that in primary periosteoplasty patients due to the more marked facial convexity, the increased prominence of the nose, and better vertical intermaxillary relationships.

Primary amines protect against retinal degeneration in mouse models of retinopathies

PubMed Central

Maeda, Akiko; Golczak, Marcin; Chen, Yu; Okano, Kiichiro; Kohno, Hideo; Shiose, Satomi; Ishikawa, Kaede; Harte, William; Palczewska, Grazyna; Maeda, Tadao; Palczewski, Krzysztof

2011-01-01

Vertebrate vision is initiated by photoisomerization of the visual pigment chromophore, 11-cis-retinal, and is maintained by continuous regeneration of this retinoid through a series of reactions termed the retinoid cycle. However, toxic side reaction products, especially those involving reactive aldehyde groups of the photoisomered product, all-trans-retinal, can cause severe retinal pathology. Here we lowered peak concentrations of free all-trans-retinal with primary amine-containing FDA-approved drugs that did not inhibit chromophore regeneration in mouse models of retinal degeneration. Schiff base adducts between all-trans-retinal and these amines were identified by mass spectrometry. Adducts were observed in mouse eyes only when an experimental drug protected the retina from degeneration in both short-term and long-term treatment experiments. This study demonstrates a molecular basis of all-trans-retinal-induced retinal pathology and identifies an assemblage of FDA-approved compounds with protective effects against this pathology in a mouse model that displays features of Stargardt’s and age-related retinal degeneration. PMID:22198730

Defective Bone Repair in C57Bl6 Mice With Acute Systemic Inflammation.

PubMed

Behrends, D A; Hui, D; Gao, C; Awlia, A; Al-Saran, Y; Li, A; Henderson, J E; Martineau, P A

2017-03-01

Bone repair is initiated with a local inflammatory response to injury. The presence of systemic inflammation impairs bone healing and often leads to malunion, although the underlying mechanisms remain poorly defined. Our research objective was to use a mouse model of cortical bone repair to determine the effect of systemic inflammation on cells in the bone healing microenvironment. QUESTION/PURPOSES: (1) Does systemic inflammation, induced by lipopolysaccharide (LPS) administration affect the quantity and quality of regenerating bone in primary bone healing? (2) Does systemic inflammation alter vascularization and the number or activity of inflammatory cells, osteoblasts, and osteoclasts in the bone healing microenvironment? Cortical defects were drilled in the femoral diaphysis of female and male C57BL/6 mice aged 5 to 9 months that were treated with daily systemic injections of LPS or physiologic saline as control for 7 days. Mice were euthanized at 1 week (Control, n = 7; LPS, n = 8), 2 weeks (Control, n = 7; LPS, n = 8), and 6 weeks (Control, n = 9; LPS, n = 8) after surgery. The quantity (bone volume per tissue volume [BV/TV]) and microarchitecture (trabecular separation and thickness, porosity) of bone in the defect were quantified with time using microCT. The presence or activity of vascular endothelial cells (CD34), macrophages (F4/80), osteoblasts (alkaline phosphatase [ALP]), and osteoclasts (tartrate-resistant acid phosphatase [TRAP]) were evaluated using histochemical analyses. Only one of eight defects was bridged completely 6 weeks after surgery in LPS-injected mouse bones compared with seven of nine defects in the control mouse bones (odds ratio [OR], 0.04; 95% CI, 0.003-0.560; p = 0.007). The decrease in cortical bone in LPS-treated mice was reflected in reduced BV/TV (21% ± 4% vs 39% ± 10%; p < 0.01), increased trabecular separation (240 ± 36 μm vs 171 ± 29 μm; p < 0.01), decreased trabecular thickness (81 ± 18 μm vs 110 ± 22 μm; p = 0

Increased bone formation in mice lacking apolipoprotein E.

PubMed

Schilling, Arndt F; Schinke, Thorsten; Münch, Christian; Gebauer, Matthias; Niemeier, Andreas; Priemel, Matthias; Streichert, Thomas; Rueger, Johannes M; Amling, Michael

2005-02-01

ApoE is a plasma protein that plays a major role in lipoprotein metabolism. Here we describe that ApoE expression is strongly induced on mineralization of primary osteoblast cultures. ApoE-deficient mice display an increased bone formation rate compared with wildtype controls, thereby showing that ApoE has a physiologic function in bone remodeling. Apolipoprotein E (ApoE) is a protein component of lipoproteins and facilitates their clearance from the circulation. This is confirmed by the phenotype of ApoE-deficient mice that have high plasma cholesterol levels and spontaneously develop atherosclerotic lesions. The bone phenotype of these mice has not been analyzed to date, although an association between certain ApoE alleles and BMD has been reported. Primary osteoblasts were isolated from newborn mouse calvariae and mineralized ex vivo. A genome-wide expression analysis was performed during the course of differentiation using the Affymetrix gene chip system. Bones from ApoE-deficient mice and wildtype controls were analyzed using radiography, micro CT imaging, and undecalcified histology. Cellular activities were assessed using dynamic histomorphometry and by measuring urinary collagen degradation products. Lipoprotein uptake assays were performed with (125)I-labeled triglyceride-rich lipoprotein-remnants (TRL-R) using primary osteoblasts from wildtype and ApoE-deficient mice. Serum concentrations of osteocalcin were determined by radioimmunoassay after hydroxyapatite chromatography. ApoE expression is strongly induced on mineralization of primary osteoblast cultures ex vivo. Mice lacking ApoE display a high bone mass phenotype that is caused by an increased bone formation rate, whereas bone resorption is not affected. This phenotype may be explained by a decreased uptake of triglyceride-rich lipoproteins by osteoblasts, resulting in elevated levels of undercarboxylated osteocalcin in the serum of ApoE-deficient mice. The specific induction of ApoE gene expression

Autologous serum improves bone formation in a primary stable silica-embedded nanohydroxyapatite bone substitute in combination with mesenchymal stem cells and rhBMP-2 in the sheep model

PubMed Central

Boos, Anja M; Weigand, Annika; Deschler, Gloria; Gerber, Thomas; Arkudas, Andreas; Kneser, Ulrich; Horch, Raymund E; Beier, Justus P

2014-01-01

New therapeutic strategies are required for critical size bone defects, because the gold standard of transplanting autologous bone from an unharmed area of the body often leads to several severe side effects and disadvantages for the patient. For years, tissue engineering approaches have been seeking a stable, axially vascularized transplantable bone replacement suitable for transplantation into the recipient bed with pre-existing insufficient conditions. For this reason, the arteriovenous loop model was developed and various bone substitutes have been vascularized. However, it has not been possible thus far to engineer a primary stable and axially vascularized transplantable bone substitute. For that purpose, a primary stable silica-embedded nanohydroxyapatite (HA) bone substitute in combination with blood, bone marrow, expanded, or directly retransplanted mesenchymal stem cells, recombinant human bone morphogenetic protein 2 (rhBMP-2), and different carrier materials (fibrin, cell culture medium, autologous serum) was tested subcutaneously for 4 or 12 weeks in the sheep model. Autologous serum lead to an early matrix change during degradation of the bone substitute and formation of new bone tissue. The best results were achieved in the group combining mesenchymal stem cells expanded with 60 μg/mL rhBMP-2 in autologous serum. Better ingrowth of fibrovascular tissue could be detected in the autologous serum group compared with the control (fibrin). Osteoclastic activity indicating an active bone remodeling process was observed after 4 weeks, particularly in the group with autologous serum and after 12 weeks in every experimental group. This study clearly demonstrates the positive effects of autologous serum in combination with mesenchymal stem cells and rhBMP-2 on bone formation in a primary stable silica-embedded nano-HA bone grafting material in the sheep model. In further experiments, the results will be transferred to the sheep arteriovenous loop model in

Tooth-bone morphogenesis during postnatal stages of mouse first molar development

PubMed Central

Lungová, Vlasta; Radlanski, Ralf J; Tucker, Abigail S; Renz, Herbert; Míšek, Ivan; Matalová, Eva

2011-01-01

The first mouse molar (M1) is the most common model for odontogenesis, with research particularly focused on prenatal development. However, the functional dentition forms postnatally, when the histogenesis and morphogenesis of the tooth is completed, the roots form and the tooth physically anchors into the jaw. In this work, M1 was studied from birth to eruption, assessing morphogenesis, proliferation and apoptosis, and correlating these with remodeling of the surrounding bony tissue. The M1 completed crown formation between postnatal (P) days 0–2, and the development of the tooth root was initiated at P4. From P2 until P12, cell proliferation in the dental epithelium reduced and shifted downward to the apical region of the forming root. In contrast, proliferation was maintained or increased in the mesenchymal cells of the dental follicle. At later stages, before tooth eruption (P20), cell proliferation suddenly ceased. This withdrawal from the cell cycle correlated with tooth mineralization and mesenchymal differentiation. Apoptosis was observed during all stages of M1 postnatal morphogenesis, playing a role in the removal of cells such as osteoblasts in the mandibular region and working together with osteoclasts to remodel the bone around the developing tooth. At more advanced developmental stages, apoptotic cells and bodies accumulated in the cell layers above the tooth cusps, in the path of eruption. Three-dimensional reconstruction of the developing postnatal tooth and bone indicates that the alveolar crypts form by resorption underneath the primordia, whereas the ridges form by active bone growth between the teeth and roots to form a functional complex. PMID:21418206

Sequencing analysis of mutations induced by N-ethyl-N-nitrosourea at different sampling times in mouse bone marrow.

PubMed

Wang, Jianyong; Chen, Tao

2010-03-01

In our previous study (Wang et al., 2004, Toxicol. Sci. 82: 124-128), we observed that the cII gene mutant frequency (MF) in the bone marrow of Big Blue mice showed significant increase as early as day 1, reached the maximum at day 3 and then decreased to a plateau by day 15 after a single dose of carcinogen N-ethyl-N-nitrosourea (ENU) treatment, which is different from the longer mutation manifestation time and the constancy of MFs after reaching their maximum in some other tissues. To determine the mechanism underlying the quick increase in MF and the peak formation in the mutant manifestation, we examined the mutation frequencies and spectra of the ENU-induced mutants collected from different sampling times in this study. The cII mutants from days 1, 3 and 120 after ENU treatment were randomly selected from different animals. The mutation frequencies were 33, 217, 305 and 144 x 10(-6) for control, days 1, 3, and 120, respectively. The mutation spectra at days 1 and 3 were significantly different from that at day 120. Considering that stem cells are responsible for the ultimate MF plateau (day 120) and transit cells are accountable for the earlier MF induction (days 1 or 3) in mouse bone marrow, we conclude that transit cells are much more sensitive to mutation induction than stem cells in mouse bone marrow, which resulted in the specific mutation manifestation induced by ENU.

Effects of gentamicin and monomer on bone. An in vitro study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pedersen, J.G.; Lund, B.

1988-01-01

Gentamicin-loaded bone cement is used with increasing frequency in primary and revision arthroplasty. Considering the high local concentration and the well-known toxic effect of gentamicin on the kidney, a similar inhibiting effect on bone tissue might be expected. In a series of in vitro studies using paired mouse calvaries cultured for 2 days, the authors found a dose-dependent decrease in the release of previously incorporated calcium-45 (UVCa) or tritiated proline and a decrease in alkaline phosphatase activity. In combination with methylmethacrylate, a small additional reduction in UVCa release and a marked decrease in alkaline phosphatase activity were recorded. These resultsmore » indicate that released gentamicin and monomer from antibiotic-supplemented bone cement depresses bone turnover and might thus play an important part in the pathogenesis of loosening.« less

Analysis of Mouse Growth Plate Development

PubMed Central

Mangiavini, Laura; Merceron, Christophe; Schipani, Ernestina

2016-01-01

To investigate skeletal development, pathophysiological mechanisms of cartilage and bone disease, and eventually assess innovative treatments, the mouse is a very important resource. During embryonic development, mesenchymal condensations are formed, and cells within these mesenchymal condensations either directly differentiate into osteoblasts and give origin to intramembranous bone, or differentiate into chondrocytes and form a cartilaginous anlage. The cartilaginous anlage or fetal growth plate is then replaced with bone. This process is also called endochondral bone development, and it is responsible for the generation of most of our skeleton. In this Review, we will discuss in detail the most common in vivo and in vitro techniques our laboratory is currently using for the analysis of the mouse fetal growth plate during development. PMID:26928664

Primary Ewing's Sarcoma of the Temporal Bone: A Rare Case Report and Literature Review.

PubMed

Gupta, Divya; Gulati, Achal; Purnima

2017-09-01

Ewing's sarcoma is a malignant, round cell tumor arising from the bones and primarily affecting children and adolescent, accounting for 3 % of all childhood malignancies. Although the long bones and the trunk are typically affected, rare cases of it involving isolated bones throughout the body have been reported. Involvement of the skull bones is rare, constituting 1-6 % of the total Ewing's sarcoma cases but those affecting the cranial bones are rarer still, constituting only 1 %. We describe an 8 months old infant having Ewing sarcoma, of the petrous and mastoid parts of temporal bone along with the occipital bone, whose clinical presentation mimicked mastoiditis with facial nerve palsy. We discuss the clinical and therapeutic course of an extensive primary Ewing sarcoma of the temporal bone, which was treated without performing surgery and review this entity's literature in detail.

Primary stability and self-tapping blades: biomechanical assessment of dental implants in medium-density bone.

PubMed

Kim, Yung-Soo; Lim, Young-Jun

2011-10-01

The aim of this biomechanical study was to assess the influence of self-tapping blades in terms of primary implant stability between implants with self-tapping blades and implants without self-tapping blades using five different analytic methods, especially in medium-density bone. Two different types of dental implants (4 × 10 mm) were tested: self-tapping and non-self-tapping. The fixture design including thread profiles was exactly the same between the two groups; the only difference was the presence of cutting blades on one half of the apical portion of the implant body. Solid rigid polyurethane blocks with corresponding densities were selected to simulate medium-density bone. Five mechanical assessments (insertion torque, resonance frequency analysis [RFA], reverse torque, pull-out and push in test) were performed for primary stability. Implants without self-tapping blades showed significantly higher values (P<0.001) in four biomechanical assessments, except RFA (P=0.684). However, a statistically significant correlation could not be detected between insertion torque values with the four different outcome variables (P>0.05). The outcomes of the present study indicate that the implant body design without self-tapping blades has a good primary stability compared with that with self-tapping blades in medium-density bone. Considering the RFA, a distinct layer of cortical bone on marginal bone will yield implant stability quotient values similar to those in medium-bone density when implants have the same diameter. © 2011 John Wiley & Sons A/S.

Lack of centrioles and primary cilia in STIL(-/-) mouse embryos.

PubMed

David, Ahuvit; Liu, Fengying; Tibelius, Alexandra; Vulprecht, Julia; Wald, Diana; Rothermel, Ulrike; Ohana, Reut; Seitel, Alexander; Metzger, Jasmin; Ashery-Padan, Ruth; Meinzer, Hans-Peter; Gröne, Hermann-Josef; Izraeli, Shai; Krämer, Alwin

2014-01-01

Although most animal cells contain centrosomes, consisting of a pair of centrioles, their precise contribution to cell division and embryonic development is unclear. Genetic ablation of STIL, an essential component of the centriole replication machinery in mammalian cells, causes embryonic lethality in mice around mid gestation associated with defective Hedgehog signaling. Here, we describe, by focused ion beam scanning electron microscopy, that STIL(-/-) mouse embryos do not contain centrioles or primary cilia, suggesting that these organelles are not essential for mammalian development until mid gestation. We further show that the lack of primary cilia explains the absence of Hedgehog signaling in STIL(-/-) cells. Exogenous re-expression of STIL or STIL microcephaly mutants compatible with human survival, induced non-templated, de novo generation of centrioles in STIL(-/-) cells. Thus, while the abscence of centrioles is compatible with mammalian gastrulation, lack of centrioles and primary cilia impairs Hedgehog signaling and further embryonic development.

Specific excitatory connectivity for feature integration in mouse primary visual cortex

PubMed Central

Molina-Luna, Patricia; Roth, Morgane M.

2017-01-01

Local excitatory connections in mouse primary visual cortex (V1) are stronger and more prevalent between neurons that share similar functional response features. However, the details of how functional rules for local connectivity shape neuronal responses in V1 remain unknown. We hypothesised that complex responses to visual stimuli may arise as a consequence of rules for selective excitatory connectivity within the local network in the superficial layers of mouse V1. In mouse V1 many neurons respond to overlapping grating stimuli (plaid stimuli) with highly selective and facilitatory responses, which are not simply predicted by responses to single gratings presented alone. This complexity is surprising, since excitatory neurons in V1 are considered to be mainly tuned to single preferred orientations. Here we examined the consequences for visual processing of two alternative connectivity schemes: in the first case, local connections are aligned with visual properties inherited from feedforward input (a ‘like-to-like’ scheme specifically connecting neurons that share similar preferred orientations); in the second case, local connections group neurons into excitatory subnetworks that combine and amplify multiple feedforward visual properties (a ‘feature binding’ scheme). By comparing predictions from large scale computational models with in vivo recordings of visual representations in mouse V1, we found that responses to plaid stimuli were best explained by assuming feature binding connectivity. Unlike under the like-to-like scheme, selective amplification within feature-binding excitatory subnetworks replicated experimentally observed facilitatory responses to plaid stimuli; explained selective plaid responses not predicted by grating selectivity; and was consistent with broad anatomical selectivity observed in mouse V1. Our results show that visual feature binding can occur through local recurrent mechanisms without requiring feedforward convergence, and

Chitosan-Graphene Oxide 3D scaffolds as Promising Tools for Bone Regeneration in Critical-Size Mouse Calvarial Defects.

PubMed

Hermenean, Anca; Codreanu, Ada; Herman, Hildegard; Balta, Cornel; Rosu, Marcel; Mihali, Ciprian Valentin; Ivan, Alexandra; Dinescu, Sorina; Ionita, Mariana; Costache, Marieta

2017-11-30

Limited self-regenerating capacity of human skeleton makes the reconstruction of critical size bone defect a significant challenge for clinical practice. Aimed for regenerating bone tissues, this study was designed to investigate osteogenic differentiation, along with bone repair capacity of 3D chitosan (CHT) scaffolds enriched with graphene oxide (GO) in critical-sized mouse calvarial defect. Histopathological/histomorphometry and scanning electron microscopy(SEM) analysis of the implants revealed larger amount of new bone in the CHT/GO-filled defects compared with CHT alone (p < 0.001). When combined with GO, CHT scaffolds synergistically promoted the increase of alkaline phosphatase activity both in vitro and in vivo experiments. This enhanced osteogenesis was corroborated with increased expression of bone morphogenetic protein (BMP) and Runx-2 up to week 4 post-implantation, which showed that GO facilitates the differentiation of osteoprogenitor cells. Meanwhile, osteogenesis was promoted by GO at the late stage as well, as indicated by the up-regulation of osteopontin and osteocalcin at week 8 and overexpressed at week 18, for both markers. Our data suggest that CHT/GO biomaterial could represent a promising tool for the reconstruction of large bone defects, without using exogenous living cells or growth factors.

In Vivo Axial Loading of the Mouse Tibia

PubMed Central

Melville, Katherine M.; Robling, Alexander G.

2015-01-01

Summary Non-invasive methods to apply controlled, cyclic loads to the living skeleton are used as an anabolic agent to stimulate new bone formation in adults and enhance bone mass accrual in growing animals. These methods are also invaluable for understanding bone signaling pathways. Our focus here is on a particular loading model: in vivo axial compression of the mouse tibia. An advantage of loading the tibia is that changes are present in both the cancellous envelope of the proximal tibia and the cortical bone of the tibial diaphysis. To load the tibia of the mouse axially in vivo, a cyclic compressive load is applied up to five times a week to a single tibia per mouse for a duration lasting from 1 day to 6 weeks. With the contralateral limb as an internal control, the anabolic response of the skeleton to mechanical stimuli can be studied in a pairwise experimental design. Here, we describe the key parameters that must be considered before beginning an in vivo mouse tibial loading experiment, including methods for in vivo strain gauging of the tibial midshaft, and then we describe general methods for loading the mouse tibia for an experiment lasting multiple days. PMID:25331046

A Surgical Procedure for Resecting the Mouse Rib: A Model for Large-Scale Long Bone Repair

PubMed Central

Funnell, John W.; Thein, Thu Zan Tun; Mariani, Francesca V.

2015-01-01

This protocol introduces researchers to a new model for large-scale bone repair utilizing the mouse rib. The procedure details the following: preparation of the animal for surgery, opening the thoracic body wall, exposing the desired rib from the surrounding intercostal muscles, excising the desired section of rib without inducing a pneumothorax, and closing the incisions. Compared to the bones of the appendicular skeleton, the ribs are highly accessible. In addition, no internal or external fixator is necessary since the adjacent ribs provide a natural fixation. The surgery uses commercially available supplies, is straightforward to learn, and well-tolerated by the animal. The procedure can be carried out with or without removing the surrounding periosteum, and therefore the contribution of the periosteum to repair can be assessed. Results indicate that if the periosteum is retained, robust repair occurs in 1 - 2 months. We expect that use of this protocol will stimulate research into rib repair and that the findings will facilitate the development of new ways to stimulate bone repair in other locations around the body. PMID:25651082

Live-Cell Imaging of Phagosome Motility in Primary Mouse RPE Cells.

PubMed

Hazim, Roni; Jiang, Mei; Esteve-Rudd, Julian; Diemer, Tanja; Lopes, Vanda S; Williams, David S

2016-01-01

The retinal pigment epithelium (RPE) is a post-mitotic epithelial monolayer situated between the light-sensitive photoreceptors and the choriocapillaris. Given its vital functions for healthy vision, the RPE is a primary target for insults that result in blinding diseases, including age-related macular degeneration (AMD). One such function is the phagocytosis and digestion of shed photoreceptor outer segments. In the present study, we examined the process of trafficking of outer segment disk membranes in live cultures of primary mouse RPE, using high speed spinning disk confocal microscopy. This approach has enabled us to track phagosomes, and determine parameters of their motility, which are important for their efficient degradation.

Scaffold-mediated BMP-2 minicircle DNA delivery accelerated bone repair in a mouse critical-size calvarial defect model.

PubMed

Keeney, Michael; Chung, Michael T; Zielins, Elizabeth R; Paik, Kevin J; McArdle, Adrian; Morrison, Shane D; Ransom, Ryan C; Barbhaiya, Namrata; Atashroo, David; Jacobson, Gunilla; Zare, Richard N; Longaker, Michael T; Wan, Derrick C; Yang, Fan

2016-08-01

Scaffold-mediated gene delivery holds great promise for tissue regeneration. However, previous attempts to induce bone regeneration using scaffold-mediated non-viral gene delivery rarely resulted in satisfactory healing. We report a novel platform with sustained release of minicircle DNA (MC) from PLGA scaffolds to accelerate bone repair. MC was encapsulated inside PLGA scaffolds using supercritical CO2 , which showed prolonged release of MC. Skull-derived osteoblasts transfected with BMP-2 MC in vitro result in higher osteocalcin gene expression and mineralized bone formation. When implanted in a critical-size mouse calvarial defect, scaffolds containing luciferase MC lead to robust in situ protein production up to at least 60 days. Scaffold-mediated BMP-2 MC delivery leads to substantially accelerated bone repair as early as two weeks, which continues to progress over 12 weeks. This platform represents an efficient, long-term nonviral gene delivery system, and may be applicable for enhancing repair of a broad range of tissues types. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2099-2107, 2016. © 2016 Wiley Periodicals, Inc.

Establishment of primary cultures for mouse ameloblasts as a model of their lifetime

DOE Office of Scientific and Technical Information (OSTI.GOV)

Suzawa, Tetsuo; Itoh, Nao; Department of Clinical Pharmacy, School of Pharmaceutical Science, Showa University

2006-07-07

To understand how the properties of ameloblasts are spatiotemporally regulated during amelogenesis, two primary cultures of ameloblasts in different stages of differentiation were established from mouse enamel epithelium. Mouse primary ameloblasts (MPAs) prepared from immature enamel epithelium (MPA-I) could proliferate, whereas those from mature enamel epithelium (MPA-M) could not. MPA-M but not MPA-I caused apoptosis during culture. The mRNA expression of amelogenin, a marker of immature ameloblasts, was down-regulated, and that of enamel matrix serine proteiase-1, a marker of mature ameloblasts, was induced in MPA-I during culture. Using green fluorescence protein as a reporter, a visualized reporter system was establishedmore » to analyze the promoter activity of the amelogenin gene. The region between -1102 bp and -261 bp was required for the reporter expression in MPA-I. These results suggest that MPAs are valuable in vitro models for investigation of ameloblast biology, and that the visualized system is useful for promoter analysis in MPAs.« less

Enhanced Wnt signaling improves bone mass and strength, but not brittleness, in the Col1a1(+/mov13) mouse model of type I Osteogenesis Imperfecta.

PubMed

Jacobsen, Christina M; Schwartz, Marissa A; Roberts, Heather J; Lim, Kyung-Eun; Spevak, Lyudmila; Boskey, Adele L; Zurakowski, David; Robling, Alexander G; Warman, Matthew L

2016-09-01

Osteogenesis Imperfecta (OI) comprises a group of genetic skeletal fragility disorders. The mildest form of OI, Osteogenesis Imperfecta type I, is frequently caused by haploinsufficiency mutations in COL1A1, the gene encoding the α1(I) chain of type 1 collagen. Children with OI type I have a 95-fold higher fracture rate compared to unaffected children. Therapies for OI type I in the pediatric population are limited to anti-catabolic agents. In adults with osteoporosis, anabolic therapies that enhance Wnt signaling in bone improve bone mass, and ongoing clinical trials are determining if these therapies also reduce fracture risk. We performed a proof-of-principle experiment in mice to determine whether enhancing Wnt signaling in bone could benefit children with OI type I. We crossed a mouse model of OI type I (Col1a1(+/Mov13)) with a high bone mass (HBM) mouse (Lrp5(+/p.A214V)) that has increased bone strength from enhanced Wnt signaling. Offspring that inherited the OI and HBM alleles had higher bone mass and strength than mice that inherited the OI allele alone. However, OI+HBM and OI mice still had bones with lower ductility compared to wild-type mice. We conclude that enhancing Wnt signaling does not make OI bone normal, but does improve bone properties that could reduce fracture risk. Therefore, agents that enhance Wnt signaling are likely to benefit children and adults with OI type 1. Copyright © 2016 Elsevier Inc. All rights reserved.

Trabecular Bone Score (TBS)—A Novel Method to Evaluate Bone Microarchitectural Texture in Patients With Primary Hyperparathyroidism

PubMed Central

Boutroy, Stephanie; Zhang, Chiyuan; McMahon, Donald Jay; Zhou, Bin; Wang, Ji; Udesky, Julia; Cremers, Serge; Sarquis, Marta; Guo, Xiang-Dong Edward; Hans, Didier

2013-01-01

Context: In the milder form of primary hyperparathyroidism (PHPT), cancellous bone, represented by areal bone mineral density at the lumbar spine by dual-energy x-ray absorptiometry (DXA), is preserved. This finding is in contrast to high-resolution peripheral quantitative computed tomography (HRpQCT) results of abnormal trabecular microstructure and epidemiological evidence for increased overall fracture risk in PHPT. Because DXA does not directly measure trabecular bone and HRpQCT is not widely available, we used trabecular bone score (TBS), a novel gray-level textural analysis applied to spine DXA images, to estimate indirectly trabecular microarchitecture. Objective: The purpose of this study was to assess TBS from spine DXA images in relation to HRpQCT indices and bone stiffness in radius and tibia in PHPT. Design and Setting: This was a cross-sectional study conducted in a referral center. Patients: Participants were 22 postmenopausal women with PHPT. Main Outcome Measures: Outcomes measured were areal bone mineral density by DXA, TBS indices derived from DXA images, HRpQCT standard measures, and bone stiffness assessed by finite element analysis at distal radius and tibia. Results: TBS in PHPT was low at 1.24, representing abnormal trabecular microstructure (normal ≥1.35). TBS was correlated with whole bone stiffness and all HRpQCT indices, except for trabecular thickness and trabecular stiffness at the radius. At the tibia, correlations were observed between TBS and volumetric densities, cortical thickness, trabecular bone volume, and whole bone stiffness. TBS correlated with all indices of trabecular microarchitecture, except trabecular thickness, after adjustment for body weight. Conclusion: TBS, a measurement technology readily available by DXA, shows promise in the clinical assessment of trabecular microstructure in PHPT. PMID:23526463

The effect of undersizing and tapping on bone to implant contact and implant primary stability: A histomorphometric study on bovine ribs.

PubMed

Di Stefano, Danilo Alessio; Perrotti, Vittoria; Greco, Gian Battista; Cappucci, Claudia; Arosio, Paolo; Piattelli, Adriano; Iezzi, Giovanna

2018-06-01

Implant site preparation may be adjusted to achieve the maximum possible primary stability. The aim of this investigation was to study the relation among bone-to-implant contact at insertion, bone density, and implant primary stability intra-operatively measured by a torque-measuring implant motor, when implant sites were undersized or tapped. Undersized (n=14), standard (n=13), and tapped (n=13) implant sites were prepared on 9 segments of bovine ribs. After measuring bone density using the implant motor, 40 implants were placed, and their primary stability assessed by measuring the integral of the torque-depth insertion curve. Bovine ribs were then processed histologically, the bone-to-implant contact measured and statistically correlated to bone density and the integral. Bone-to-implant contact and the integral of the torque-depth curve were significantly greater for undersized sites than tapped sites. Moreover, a correlation between bone to implant contact, the integral and bone density was found under all preparation conditions. The slope of the bone-to-implant/density and integral/density lines was significantly greater for undersized sites, while those corresponding to standard prepared and tapped sites did not differ significantly. The integral of the torque-depth curve provided reliable information about bone-to-implant contact and primary implant stability even in tapped or undersized sites. The linear relations found among the parameters suggests a connection between extent and modality of undersizing and the corresponding increase of the integral and, consequently, of primary stability. These results might help the physician determine the extent of undersizing needed to achieve the proper implant primary stability, according to the planned loading protocol.

Imaging Primary Mouse Sarcomas After Radiation Therapy Using Cathepsin-Activatable Fluorescent Imaging Agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cuneo, Kyle C.; Mito, Jeffrey K.; Javid, Melodi P.

2013-05-01

Purpose: Cathepsin-activated fluorescent probes can detect tumors in mice and in canine patients. We previously showed that these probes can detect microscopic residual sarcoma in the tumor bed of mice during gross total resection. Many patients with soft tissue sarcoma (STS) and other tumors undergo radiation therapy (RT) before surgery. This study assesses the effect of RT on the ability of cathepsin-activated probes to differentiate between normal and cancerous tissue. Methods and Materials: A genetically engineered mouse model of STS was used to generate primary hind limb sarcomas that were treated with hypofractionated RT. Mice were injected intravenously with cathepsin-activatedmore » fluorescent probes, and various tissues, including the tumor, were imaged using a hand-held imaging device. Resected tumor and normal muscle samples were harvested to assess cathepsin expression by Western blot. Uptake of activated probe was analyzed by flow cytometry and confocal microscopy. Parallel in vitro studies using mouse sarcoma cells were performed. Results: RT of primary STS in mice and mouse sarcoma cell lines caused no change in probe activation or cathepsin protease expression. Increasing radiation dose resulted in an upward trend in probe activation. Flow cytometry and immunofluorescence showed that a substantial proportion of probe-labeled cells were CD11b-positive tumor-associated immune cells. Conclusions: In this primary murine model of STS, RT did not affect the ability of cathepsin-activated probes to differentiate between tumor and normal muscle. Cathepsin-activated probes labeled tumor cells and tumor-associated macrophages. Our results suggest that it would be feasible to include patients who have received preoperative RT in clinical studies evaluating cathepsin-activated imaging probes.« less

Deproteinized bovine bone mineral particles and osseointegration of implants without primary bone contact: an experimental study in dogs.

PubMed

Sivolella, Stefano; Bressan, Eriberto; Salata, Luiz A; Quiñones, Maria E; Lang, Niklaus P; Botticelli, Daniele

2014-03-01

To evaluate the influence on osseointegration of Deproteinized bovine bone mineral (DBBM) particles used to fill defects of at least 1 mm around implants having no primary contact with bone. Premolars and first molars were extracted bilaterally from the mandible of six Labrador dogs. After 3 months of healing, mucoperiosteal full-thickness flaps were elevated, and one recipient site was prepared in the molar region of each hemi-mandible to place implants. These were installed with a deliberate circumferential and periapical space to the bone walls of 1.2 mm. All implants were stabilized with passive fixation plates to maintain the implants in situ and without any contact with the implant bed. The control sites were left to be filled with coagulum, while at the test sites, the residual gap was filled with DBBM. After 3 months of submerged healing, the animals were sacrificed. Ground sections were prepared and analyzed histomorphometrically. Mineralized bone-to-implant contact was 4.0% and 3.9% for control and test sites, respectively. The width of the residual defects was 0.48 mm and 0.88 mm at the control and test sites, respectively. The percentage of implant surface covered by a layer of dense connective tissue of 0.12 mm of width on average was 84.9% and 88.5% at the control and test sites, respectively. A minor and not predictable degree of contact or distance osteogenesis was obtained on the implant surface when primary contact of the implant surface with the implant bed had deliberately been avoided. DBBM grafting of the artificial gap did not favor osseointegration. Neither did it enhance the ability to bridge the gap with newly formed bone in an artificial defect wider than 1 mm. © 2013 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.

Zika virus-induced hyper excitation precedes death of mouse primary neuron.

PubMed

Gaburro, Julie; Bhatti, Asim; Sundaramoorthy, Vinod; Dearnley, Megan; Green, Diane; Nahavandi, Saeid; Paradkar, Prasad N; Duchemin, Jean-Bernard

2018-04-27

Zika virus infection in new born is linked to congenital syndromes, especially microcephaly. Studies have shown that these neuropathies are the result of significant death of neuronal progenitor cells in the central nervous system of the embryo, targeted by the virus. Although cell death via apoptosis is well acknowledged, little is known about possible pathogenic cellular mechanisms triggering cell death in neurons. We used in vitro embryonic mouse primary neuron cultures to study possible upstream cellular mechanisms of cell death. Neuronal networks were grown on microelectrode array and electrical activity was recorded at different times post Zika virus infection. In addition to this method, we used confocal microscopy and Q-PCR techniques to observe morphological and molecular changes after infection. Zika virus infection of mouse primary neurons triggers an early spiking excitation of neuron cultures, followed by dramatic loss of this activity. Using NMDA receptor antagonist, we show that this excitotoxicity mechanism, likely via glutamate, could also contribute to the observed nervous system defects in human embryos and could open new perspective regarding the causes of adult neuropathies. This model of excitotoxicity, in the context of neurotropic virus infection, highlights the significance of neuronal activity recording with microelectrode array and possibility of more than one lethal mechanism after Zika virus infection in the nervous system.

Monomethylarsonous acid (MMA+3) Inhibits IL-7 Signaling in Mouse Pre-B Cells

PubMed Central

Ezeh, Peace C.; Xu, Huan; Lauer, Fredine T.; Liu, Ke Jian; Hudson, Laurie G.; Burchiel, Scott W.

2016-01-01

Our previously published data show that As+3 in vivo and in vitro, at very low concentrations, inhibits lymphoid, but not myeloid stem cell development in mouse bone marrow. We also showed that the As+3 metabolite, monomethylarsonous acid (MMA+3), was responsible for the observed pre-B cell toxicity caused by As+3. Interleukin-7 (IL-7) is the primary growth factor responsible for pre-lymphoid development in mouse and human bone marrow, and Signal Transducer and Activator of Transcription 5 (STAT5) is a transcriptional factor in the IL-7 signaling pathway. We found that MMA+3 inhibited STAT5 phosphorylation at a concentration as low as 50 nM in mouse bone marrow pre-B cells. Inhibition of STAT5 phosphorylation by As+3 occurred only at a concentration of 500 nM. In the IL-7 dependent mouse pre-B 2E8 cell line, we also found selective inhibition of STAT5 phosphorylation by MMA+3, and this inhibition was dependent on effects on JAK3 phosphorylation. IL-7 receptor expression on 2E8 cell surface was also suppressed by 50 nM MMA+3 at 18 h. As further evidence for the inhibition of STAT5, we found that the induction of several genes required in B cell development, cyclin D1, E2A, EBF1, and PAX5, were selectively inhibited by MMA+3. Since 2E8 cells lack the enzymes responsible for the conversion of As+3 to MMA+3 in vitro, the results of these studies suggest that As+3 induced inhibition of pre-B cell formation in vivo is likely dependent on the formation of MMA+3 which in turn inhibits IL-7 signaling at several steps in mouse pre-B cells. PMID:26518055

Hypomorphic mutation in mouse Nppc gene causes retarded bone growth due to impaired endochondral ossification

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsuji, Takehito; Kondo, Eri; Yasoda, Akihiro

2008-11-07

Long bone abnormality (lbab/lbab) is a spontaneous mutant mouse characterized by dwarfism with shorter long bones. A missense mutation was reported in the Nppc gene, which encodes C-type natriuretic peptide (CNP), but it has not been confirmed whether this mutation is responsible for the dwarf phenotype. To verify that the mutation causes the dwarfism of lbab/lbab mice, we first investigated the effect of CNP in lbab/lbab mice. By transgenic rescue with chondrocyte-specific expression of CNP, the dwarf phenotype in lbab/lbab mice was completely compensated. Next, we revealed that CNP derived from the lbab allele retained only slight activity to inducemore » cGMP production through its receptor. Histological analysis showed that both proliferative and hypertrophic zones of chondrocytes in the growth plate of lbab/lbab mice were markedly reduced. Our results demonstrate that lbab/lbab mice have a hypomorphic mutation in the Nppc gene that is responsible for dwarfism caused by impaired endochondral ossification.« less

Quantitative Primary Tumor Indocyanine Green Measurements Predict Osteosarcoma Metastatic Lung Burden in a Mouse Model.

PubMed

Fourman, Mitchell S; Mahjoub, Adel; Mandell, Jon B; Yu, Shibing; Tebbets, Jessica C; Crasto, Jared A; Alexander, Peter E; Weiss, Kurt R

2018-03-01

Current preclinical osteosarcoma (OS) models largely focus on quantifying primary tumor burden. However, most fatalities from OS are caused by metastatic disease. The quantification of metastatic OS currently relies on CT, which is limited by motion artifact, requires intravenous contrast, and can be technically demanding in the preclinical setting. We describe the ability for indocyanine green (ICG) fluorescence angiography to quantify primary and metastatic OS in a previously validated orthotopic, immunocompetent mouse model. (1) Can near-infrared ICG fluorescence be used to attach a comparable, quantitative value to the primary OS tumor in our experimental mouse model? (2) Will primary tumor fluorescence differ in mice that go on to develop metastatic lung disease? (3) Does primary tumor fluorescence correlate with tumor volume measured with CT? Six groups of 4- to 6-week-old immunocompetent Balb/c mice (n = 6 per group) received paraphyseal injections into their left hindlimb proximal tibia consisting of variable numbers of K7M2 mouse OS cells. A hindlimb transfemoral amputation was performed 4 weeks after injection with euthanasia and lung extraction performed 10 weeks after injection. Histologic examination of lung and primary tumor specimens confirmed ICG localization only within the tumor bed. Mice with visible or palpable tumor growth had greater hindlimb fluorescence (3.5 ± 2.3 arbitrary perfusion units [APU], defined as the fluorescence pixel return normalized by the detector) compared with those with a negative examination (0.71 ± 0.38 APU, -2.7 ± 0.5 mean difference, 95% confidence interval -3.7 to -1.8, p < 0.001). A strong linear trend (r = 0.81, p < 0.01) was observed between primary tumor and lung fluorescence, suggesting that quantitative ICG tumor fluorescence is directly related to eventual metastatic burden. We did not find a correlation (r = 0.04, p = 0.45) between normalized primary tumor fluorescence and CT volumetric measurements. We

Anatomical Variation of the Tarsus in Common Inbred Mouse Strains.

PubMed

Richbourg, Heather A; Martin, Matthew J; Schachner, Emma R; McNulty, Margaret A

2017-03-01

Rodent models are used for a variety of orthopedic research applications; however, anatomy references include mostly artistic representations. Advanced imaging techniques, including micro-computed tomography (microCT), can provide more accurate representations of subtle anatomical characteristics. A recent microCT atlas of laboratory mouse (Mus musculus) anatomy depicts the central and tarsal bone III (T3) as a single bone, differing from previous references. Fusion of tarsal bones is generally characterized as pathological secondary to mutations associated with growth factors, and normal variation has not been documented in the mouse tarsus. Therefore, it is unclear if this fusion is a normal or a pathological characteristic. The aim of this study is to characterize the tarsus of the laboratory mouse and compare it to the rat and selected outgroup species (i.e., white-footed mouse) via microCT and histology to determine if the central and T3 are separate or fused into a single bone. Laboratory mice (C57/Bl6 [n = 17] and BalbC [n = 2]) and rats (n = 5) were scanned with microCT. A representative laboratory mouse from each strain was evaluated histologically via serial sagittal sections through the mid-tarsus. General pedal anatomy was similar between all species; however, the central and T3 bones were fused in all laboratory mice but not the rat or white-footed mouse. A band of hyaline cartilage was identified within the fused bone of the laboratory mice. We conclude that the fusion found is a normal characteristic in laboratory mice, but timing of the fusion remains ambiguous. Anat Rec, 300:450-459, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Primary mouse lung fibroblasts help macrophages to tackle Mycobacterium tuberculosis more efficiently and differentiate into myofibroblasts up on bacterial stimulation.

PubMed

Verma, Subash Chand; Agarwal, Pooja; Krishnan, Manju Y

2016-03-01

Keeping with their classical role in wound healing, fibroblasts of the lung take part in the resolution of tubercular granulomas. They are totally absent in nascent granulomas, but surround necrotizing granulomas, and are the majority of cells in healed granulomas. Lung fibroblasts may become infected with Mycobacterium tuberculosis (Mtb). Two previous studies suggested an immunomodulatory effect of fibroblasts on infected macrophages. In the present study, we looked at the role of primary mouse lung fibroblasts on naive or activated mouse bone marrow macrophages infected with Mtb and the effect of infection on fibroblast properties. We observed that with fibroblasts in the vicinity, infected naive macrophages restricted the bacterial growth, while activated macrophages turned more bactericidal with concomitant increase in nitrite production. Neutralizing IL-1α in fibroblast supernatant reduced the nitrite production by infected macrophages. Secretion of IL-6 and MCP-1 was down-regulated, while TNF-α was up-regulated in infected naive macrophages. In infected activated macrophages, the secretion of IL-6 was up-regulated, while that of MCP-1 and TNF-α was unaffected. The 'fibroblast effects' were enhanced when the fibroblasts too were infected. Mtb induced IL-1 secretion and pro-fibrotic responses by fibroblasts. Mtb-induced myofibroblast conversion was blocked by rapamycin suggesting cell signalling via mTOR. Copyright © 2015 Elsevier Ltd. All rights reserved.

Bone mineral density and bone size in men with primary osteoporosis and vertebral fractures.

PubMed

Vega, E; Ghiringhelli, G; Mautalen, C; Rey Valzacchi, G; Scaglia, H; Zylberstein, C

1998-05-01

The bone mineral density (BMD) at the lumbar spine, proximal femur, and total skeleton was evaluated in 38 men with primary osteoporosis and vertebral fractures. BMD of the patients was significantly reduced over all skeletal areas compared with controls. The Z-score of the lumbar spine (-2.8 +/- 0.9) was less than that of the other areas (P < 0.001) except the legs (-2.5 +/- 1.1) (p.n.s.) showing that bone loss had a tendency to be greater over the axial skeleton. Vertebral dimensions compared with age-matched controls were as follows: projected L2-L4 area (cm 2): 45.7 +/- 5.6 versus 53.7 +/- 3. 6 (P < 0.001); vertebral width (cm): 4.37 +/- 0.44 versus 4.90 +/- 0. 36 (P < 0.001). Serum biochemical parameters and testosterone levels were similar between osteoporotic and control men. We conclude that men with vertebral osteoporotic fractures have reduced vertebral BMD and vertebral dimensions compared with age-matched controls. Thus, these findings indicate that the achievement of a reduced bone size at the end of the growth period or a failure of periosteal increase during adult life is likely to contribute to the pathogenesis of the vertebral fractures observed in older men.

Radioprotective effects of hawthorn fruit extract against gamma irradiation in mouse bone marrow cells.

PubMed

Hosseinimehr, Seyed Jalal; Azadbakht, Mohammad; Mousavi, Seyedeh Maryam; Mahmoudzadeh, Aziz; Akhlaghpoor, Shahram

2007-01-01

The radioprotective effect of hawthorn (Crataegus microphylla) fruit extract against genotoxicity induced by gamma irradiation has been investigated in mouse bone marrow cells. A single intraperitoneal (ip) administration of hawthorn extract at doses of 25, 50, 100 and 200 mg/kg 1h prior to gamma irradiation (2 Gy) reduced the frequencies of micronucleated polychromatic erythrocytes (MnPCEs). All four doses of hawthorn extract significantly reduced the frequencies of MnPCEs and increased the PCE/PCE+NCE ratio (polychromatic erythrocyte/ polychromatic erythrocyte + normochromatic erythrocyte) in mice bone marrow compared with the non drug-treated irradiated control (p < 0.02-0.00001). The maximum reduction in MnPCEs was observed in mice treated with extract at a dose of 200 mg/kg. Administration of amifostine at dose 100 mg/kg and hawthorn at dose 200 mg/kg reduced the frequency of MnPCE almost 4.8 and 5.7 fold; respectively, after being exposed to 2 Gy of gamma rays, compare with the irradiated control group. Crataegus extract exhibited concentration-dependent activity on 1,1-diphenyl 2-picrylhydrazyl free radical showing that Crataegus contained high amounts of phenolic compounds and the HPLC analysis determined that it contained chlorogenic acid, epicatechin and hyperoside. It appeared that hawthorn extract with antioxidant activity reduced the genotoxicity induced by gamma irradiation in bone marrow cells.

Correlations Between Bone Mechanical Properties and Bone Composition Parameters in Mouse Models of Dominant and Recessive Osteogenesis Imperfecta and the Response to Anti-TGF-β Treatment.

PubMed

Bi, Xiaohong; Grafe, Ingo; Ding, Hao; Flores, Rene; Munivez, Elda; Jiang, Ming Ming; Dawson, Brian; Lee, Brendan; Ambrose, Catherine G

2017-02-01

Osteogenesis imperfecta (OI) is a group of genetic disorders characterized by brittle bones that are prone to fracture. Although previous studies in animal models investigated the mechanical properties and material composition of OI bone, little work has been conducted to statistically correlate these parameters to identify key compositional contributors to the impaired bone mechanical behaviors in OI. Further, although increased TGF-β signaling has been demonstrated as a contributing mechanism to the bone pathology in OI models, the relationship between mechanical properties and bone composition after anti-TGF-β treatment in OI has not been studied. Here, we performed follow-up analyses of femurs collected in an earlier study from OI mice with and without anti-TGF-β treatment from both recessive (Crtap -/- ) and dominant (Col1a2 +/P.G610C ) OI mouse models and WT mice. Mechanical properties were determined using three-point bending tests and evaluated for statistical correlation with molecular composition in bone tissue assessed by Raman spectroscopy. Statistical regression analysis was conducted to determine significant compositional determinants of mechanical integrity. Interestingly, we found differences in the relationships between bone composition and mechanical properties and in the response to anti-TGF-β treatment. Femurs of both OI models exhibited increased brittleness, which was associated with reduced collagen content and carbonate substitution. In the Col1a2 +/P.G610C femurs, reduced hydroxyapatite crystallinity was also found to be associated with increased brittleness, and increased mineral-to-collagen ratio was correlated with increased ultimate strength, elastic modulus, and bone brittleness. In both models of OI, regression analysis demonstrated that collagen content was an important predictor of the increased brittleness. In summary, this work provides new insights into the relationships between bone composition and material properties in

Decreasing maternal myostatin programs adult offspring bone strength in a mouse model of osteogenesis imperfecta

PubMed Central

Oestreich, Arin K.; Kamp, William M.; McCray, Marcus G.; Carleton, Stephanie M.; Karasseva, Natalia; Lenz, Kristin L.; Jeong, Youngjae; Daghlas, Salah A.; Yao, Xiaomei; Wang, Yong; Pfeiffer, Ferris M.; Ellersieck, Mark R.; Schulz, Laura C.; Phillips, Charlotte L.

2016-01-01

During fetal development, the uterine environment can have effects on offspring bone architecture and integrity that persist into adulthood; however, the biochemical and molecular mechanisms remain unknown. Myostatin is a negative regulator of muscle mass. Parental myostatin deficiency (Mstntm1Sjl/+) increases muscle mass in wild-type offspring, suggesting an intrauterine programming effect. Here, we hypothesized that Mstntm1Sjl/+ dams would also confer increased bone strength. In wild-type offspring, maternal myostatin deficiency altered fetal growth and calvarial collagen content of newborn mice and conferred a lasting impact on bone geometry and biomechanical integrity of offspring at 4 mo of age, the age of peak bone mass. Second, we sought to apply maternal myostatin deficiency to a mouse model with osteogenesis imperfecta (Col1a2oim), a heritable connective tissue disorder caused by abnormalities in the structure and/or synthesis of type I collagen. Femora of male Col1a2oim/+ offspring from natural mating of Mstntm1Sjl/+ dams to Col1a2oim/+sires had a 15% increase in torsional ultimate strength, a 29% increase in tensile strength, and a 24% increase in energy to failure compared with age, sex, and genotype-matched offspring from natural mating of Col1a2oim/+ dams to Col1a2oim/+ sires. Finally, increased bone biomechanical strength of Col1a2oim/+ offspring that had been transferred into Mstntm1Sjl/+ dams as blastocysts demonstrated that the effects of maternal myostatin deficiency were conferred by the postimplantation environment. Thus, targeting the gestational environment, and specifically prenatal myostatin pathways, provides a potential therapeutic window and an approach for treating osteogenesis imperfecta. PMID:27821779

Administration of soluble activin receptor 2B increases bone and muscle mass in a mouse model of osteogenesis imperfecta

PubMed Central

DiGirolamo, Douglas J.; Singhal, Vandana; Chang, Xiaoli; Lee, Se-Jin; Germain-Lee, Emily L.

2015-01-01

Osteogenesis imperfecta (OI) comprises a group of heritable connective tissue disorders generally defined by recurrent fractures, low bone mass, short stature and skeletal fragility. Beyond the skeletal complications of OI, many patients also report intolerance to physical activity, fatigue and muscle weakness. Indeed, recent studies have demonstrated that skeletal muscle is also negatively affected by OI, both directly and indirectly. Given the well-established interdependence of bone and skeletal muscle in both physiology and pathophysiology and the observations of skeletal muscle pathology in patients with OI, we investigated the therapeutic potential of simultaneous anabolic targeting of both bone and skeletal muscle using a soluble activin receptor 2B (ACVR2B) in a mouse model of type III OI (oim). Treatment of 12-week-old oim mice with ACVR2B for 4 weeks resulted in significant increases in both bone and muscle that were similar to those observed in healthy, wild-type littermates. This proof of concept study provides encouraging evidence for a holistic approach to treating the deleterious consequences of OI in the musculoskeletal system. PMID:26161291

Keratin 13 expression reprograms bone and brain metastases of human prostate cancer cells.

PubMed

Li, Qinlong; Yin, Lijuan; Jones, Lawrence W; Chu, Gina C-Y; Wu, Jason B-Y; Huang, Jen-Ming; Li, Quanlin; You, Sungyong; Kim, Jayoung; Lu, Yi-Tsung; Mrdenovic, Stefan; Wang, Ruoxiang; Freeman, Michael R; Garraway, Isla; Lewis, Michael S; Chung, Leland W K; Zhau, Haiyen E

2016-12-20

Lethal progression of prostate cancer metastasis can be improved by developing animal models that recapitulate the clinical conditions. We report here that cytokeratin 13 (KRT13), an intermediate filament protein, plays a directive role in prostate cancer bone, brain, and soft tissue metastases. KRT13 expression was elevated in bone, brain, and soft tissue metastatic prostate cancer cell lines and in primary and metastatic clinical prostate, lung, and breast cancer specimens. When KRT13 expression was determined at a single cell level in primary tumor tissues of 44 prostate cancer cases, KRT13 level predicted bone metastasis and the overall survival of prostate cancer patients. Genetically enforced KRT13 expression in human prostate cancer cell lines drove metastases toward mouse bone, brain and soft tissues through a RANKL-independent mechanism, as KRT13 altered the expression of genes associated with EMT, stemness, neuroendocrine/neuromimicry, osteomimicry, development, and extracellular matrices, but not receptor activator NF-κB ligand (RANKL) signaling networks in prostate cancer cells. Our results suggest new inhibitors targeting RANKL-independent pathways should be developed for the treatment of prostate cancer bone and soft tissue metastases.

Follow-up of bone lesions in an experimental multiple myeloma mouse model: description of an in vivo technique using radiography dedicated for mammography.

PubMed Central

Vanderkerken, K.; Goes, E.; De Raeve, H.; Radl, J.; Van Camp, B.

1996-01-01

The evolution of bone lesions in transplantable C57BL/KaLwRjj 5T mouse myeloma (MM) has been followed in vivo. Mice were anaesthetised and a radiograph of the pelvis and hind legs was performed by a radiograph dedicated for mammography. This is the first description of an in vivo technique under experimental conditions whereby the development of bone lesions owing to the MM growth was demonstrated. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 6 PMID:8664113

The effect of dipeptidyl peptidase-IV inhibition on bone in a mouse model of type 2 diabetes

PubMed Central

Gallagher, Emily Jane; Sun, Hui; Kornhauser, Caroline; Tobin-Hess, Aviva; Epstein, Sol; Yakar, Shoshana; LeRoith, Derek

2017-01-01

Background Individuals with type 2 diabetes (T2D) are at greater risk of bone fractures than those without diabetes. Certain oral diabetic medications may further increase the risk of fracture. Dipeptidyl peptidase-IV (DPP-IV) inhibitors are incretin-based therapies that are being increasingly used for the management of T2D. It has been hypothesized that these agents may reduce fracture risk in those with T2D. In this study, we used a mouse model of T2D to examine the effects of the DPP-IV inhibitor, MK-0626, on bone. Methods Male wild type (WT) and diabetic muscle-lysine-arginine (MKR) mice were treated with MK-0626, pioglitazone, alendronate or vehicle. The effects of treatment with MK-0626 on bone microarchitecture and turnover were compared with treatment with pioglitazone, alendronate and vehicle. Osteoblast differentiation was determined by alkaline phosphatase staining of bone marrow cells from WT and MKR mice after treatment with pioglitazone, MK-0626 or phosphate buffered saline. Results We found that MK-0626 had neutral effects on cortical and trabecular bone in diabetic mice. Pioglitazone had detrimental effects on the trabecular bone of WT but not of diabetic mice. Alendronate caused improvements in cortical and trabecular bone architecture in diabetic and WT mice. MK-0626 did not alter osteoblast differentiation, but pioglitazone impaired osteoblast differentiation in vitro. Conclusions Overall, the DPP-IV inhibitor, MK-0626, had no adverse effects on bone in an animal model of T2D or directly on osteoblasts in culture. These findings are reassuring as DPP-IV inhibitors are being widely used to treat patients with T2D who are already at an increased risk of fractures. PMID:24023014

Stimulation of host bone marrow stromal cells by sympathetic nerves promotes breast cancer bone metastasis in mice.

PubMed

Campbell, J Preston; Karolak, Matthew R; Ma, Yun; Perrien, Daniel S; Masood-Campbell, S Kathryn; Penner, Niki L; Munoz, Steve A; Zijlstra, Andries; Yang, Xiangli; Sterling, Julie A; Elefteriou, Florent

2012-07-01

Bone and lung metastases are responsible for the majority of deaths in patients with breast cancer. Following treatment of the primary cancer, emotional and psychosocial factors within this population precipitate time to recurrence and death, however the underlying mechanism(s) remain unclear. Using a mouse model of bone metastasis, we provide experimental evidence that activation of the sympathetic nervous system, which is one of many pathophysiological consequences of severe stress and depression, promotes MDA-231 breast cancer cell colonization of bone via a neurohormonal effect on the host bone marrow stroma. We demonstrate that induction of RANKL expression in bone marrow osteoblasts, following β2AR stimulation, increases the migration of metastatic MDA-231 cells in vitro, independently of SDF1-CXCR4 signaling. We also show that the stimulatory effect of endogenous (chronic stress) or pharmacologic sympathetic activation on breast cancer bone metastasis in vivo can be blocked with the β-blocker propranolol, and by knockdown of RANK expression in MDA-231 cells. These findings indicate that RANKL promotes breast cancer cell metastasis to bone via its pro-migratory effect on breast cancer cells, independently of its effect on bone turnover. The emerging clinical implication, supported by recent epidemiological studies, is that βAR-blockers and drugs interfering with RANKL signaling, such as Denosumab, could increase patient survival if used as adjuvant therapy to inhibit both the early colonization of bone by metastatic breast cancer cells and the initiation of the "vicious cycle" of bone destruction induced by these cells.

Contemporary adjuvant polymethyl methacrylate cementation optimally limits recurrence in primary giant cell tumor of bone patients compared to bone grafting: a systematic review and meta-analysis

PubMed Central

2013-01-01

Background Reports of recurrence following restructuring of primary giant cell tumor (GCT) defects using polymethyl methacrylate (PMMA) bone cementation or allogeneic bone graft with and without adjuvants for intralesional curettage vary widely. Systematic review and meta-analysis were conducted to investigate efficacy of PMMA bone cementation and allogeneic bone grafting following intralesional curettage for GCT. Methods Medline, EMBASE, Google Scholar, and Cochrane databases were searched for studies reporting GCT of bone treatment with PMMA cementation and/or bone grafting with or without adjuvant therapy following intralesional curettage of primary GCTs. Pooled risk ratios and 95% confidence intervals (CIs) for local recurrence risks were calculated by fixed-effects methods. Results Of 1,690 relevant titles, 6 eligible studies (1,293 patients) spanning March 2008 to December 2011 were identified in published data. Treatment outcomes of PMMA-only (n = 374), bone graft-only (n = 436), PMMA with or without adjuvant (PMMA + adjuvant; n = 594), and bone graft filling with or without adjuvant (bone graft + adjuvant; n = 699) were compared. Bone graft-only patients exhibited higher recurrence rates than PMMA-treated patients (RR 2.09, 95% CI (1.64, 2.66), Overall effect: Z = 6.00; P <0.001), and bone graft + adjuvant patients exhibited higher recurrence rates than PMMA + adjuvant patients (RR 1.66, 95% CI (1.21, 2.28), Overall effect: Z = 3.15, P = 0.002). Conclusions Local recurrence was minimal in PMMA cementation patients, suggesting that PMMA is preferable for routine clinical restructuring in eligible GCT patients. Relationships between tumor characteristics, other modern adjuvants, and recurrence require further exploration. PMID:23866921

Ovariectomy-induced osteopenia influences the middle and late periods of bone healing in a mouse femoral osteotomy model.

PubMed

Pang, Jian; Ye, Meina; Cao, Yuelong; Zheng, Yuxin; Guo, Hailing; Zhao, Yongfang; Zhan, Hongsheng; Shi, Yinyu

2014-10-09

Objective It is known that bone healing was delayed in the presence of osteoporosis in humans. However, due to the complexities of the healing of osteoporotic fractures, animal models may be more appropriate to study the effects of osteoporosis in more details and to test drugs on the fracture repair process. The purpose of this study was to investigate the influence of ovariectomy-induced osteopenia in bone healing in an open femoral osteotomy model, and to test the feasibility of this model for evaluating the healing process under osteopenic conditions. Methods In assessing the effects of osteopenia on fracture healing, ovariectomized mouse models were employed. A mid-shaft femur osteotomy model was also established 3 weeks after ovariectomy as an osteopenic fracture group (OVX group). Femurs were then harvested at 2 weeks and 6 weeks after fracture for X-ray radiography, micro-computed tomography (micro-CT), histology and biomechanical analysis. A sham-operated group (Sham group) was used for comparison. Results The OVX mice had significantly lower BVF, vBMD and TMD in the fracture calluses at 6 weeks (P < 0.05), and similar trend was observed in 2 weeks. Additionally, larger calluses in OVX animals were observed via micro-CT and X-ray, but these did not result in better healing outcomes as determined by biomechanical test at 6 weeks. Histological images of the healing fractures in the OVX mice found forward of broken end resorption and delay of hard callus remodeling. The impaired biomechanical measurements in the OVX group (P < 0.05) were consistent with micro-CT measurements and radiographic scoring, which also indicated delay in fracture healing of the OVX group. Conclusions This study provided evidences that ovariectomy-induced osteopenia impair the middle and late bone healing process once more. These data also supported the validity of the mouse femoral osteotomy model in evaluating the process of bone healing under osteopenic conditions.

Efficient CRISPR-mediated mutagenesis in primary immune cells using CrispRGold and a C57BL/6 Cas9 transgenic mouse line.

PubMed

Chu, Van Trung; Graf, Robin; Wirtz, Tristan; Weber, Timm; Favret, Jeremy; Li, Xun; Petsch, Kerstin; Tran, Ngoc Tung; Sieweke, Michael H; Berek, Claudia; Kühn, Ralf; Rajewsky, Klaus

2016-11-01

Applying clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9)-mediated mutagenesis to primary mouse immune cells, we used high-fidelity single guide RNAs (sgRNAs) designed with an sgRNA design tool (CrispRGold) to target genes in primary B cells, T cells, and macrophages isolated from a Cas9 transgenic mouse line. Using this system, we achieved an average knockout efficiency of 80% in B cells. On this basis, we established a robust small-scale CRISPR-mediated screen in these cells and identified genes essential for B-cell activation and plasma cell differentiation. This screening system does not require deep sequencing and may serve as a precedent for the application of CRISPR/Cas9 to primary mouse cells.

Intraperitoneal injection of thalidomide attenuates bone cancer pain and decreases spinal tumor necrosis factor-α expression in a mouse model.

PubMed

Gu, Xiaoping; Zheng, Yaguo; Ren, Bingxu; Zhang, Rui; Mei, Fengmei; Zhang, Juan; Ma, Zhengliang

2010-10-05

Tumor necrosis factor α (TNF-α) may have a pivotal role in the genesis of mechanical allodynia and thermal hyperalgesia during inflammatory and neuropathic pain. Thalidomide has been shown to selectively inhibit TNF-α production. Previous studies have suggested that thalidomide exerts anti-nociceptive effects in various pain models, but its effects on bone cancer pain have not previously been studied. Therefore, in the present study, we investigated the effect of thalidomide on bone cancer-induced hyperalgesia and up-regulated expression of spinal TNF-α in a mouse model. Osteosarcoma NCTC 2472 cells were implanted into the intramedullary space of the right femurs of C3H/HeJ mice to induce ongoing bone cancer related pain behaviors. At day 5, 7, 10 and 14 after operation, the expression of TNF-α in the spinal cord was higher in tumor-bearing mice compared to the sham mice. Intraperitoneal injection of thalidomide (50 mg/kg), started at day 1 after surgery and once daily thereafter until day 7, attenuated bone cancer-evoked mechanical allodynia and thermal hyperalgesia as well as the up-regulation of TNF-α in the spinal cord. These results suggest that thalidomide can efficiently alleviate bone cancer pain and it may be a useful alternative or adjunct therapy for bone cancer pain. Our data also suggest a role of spinal TNF-α in the development of bone cancer pain.

Lack of centrioles and primary cilia in STIL−/− mouse embryos

PubMed Central

David, Ahuvit; Liu, Fengying; Tibelius, Alexandra; Vulprecht, Julia; Wald, Diana; Rothermel, Ulrike; Ohana, Reut; Seitel, Alexander; Metzger, Jasmin; Ashery-Padan, Ruth; Meinzer, Hans-Peter; Gröne, Hermann-Josef; Izraeli, Shai; Krämer, Alwin

2014-01-01

Although most animal cells contain centrosomes, consisting of a pair of centrioles, their precise contribution to cell division and embryonic development is unclear. Genetic ablation of STIL, an essential component of the centriole replication machinery in mammalian cells, causes embryonic lethality in mice around mid gestation associated with defective Hedgehog signaling. Here, we describe, by focused ion beam scanning electron microscopy, that STIL−/− mouse embryos do not contain centrioles or primary cilia, suggesting that these organelles are not essential for mammalian development until mid gestation. We further show that the lack of primary cilia explains the absence of Hedgehog signaling in STIL−/− cells. Exogenous re-expression of STIL or STIL microcephaly mutants compatible with human survival, induced non-templated, de novo generation of centrioles in STIL−/− cells. Thus, while the abscence of centrioles is compatible with mammalian gastrulation, lack of centrioles and primary cilia impairs Hedgehog signaling and further embryonic development. PMID:25486474

The effect of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrow‑derived mesenchymal stem cells.

PubMed

Javanmard, F; Azadbakht, M; Pourmoradi, M

2016-01-01

In this study, the role of hydrostatic pressure on staurosporine-induced neural differentiation in mouse bone marrow mesenchymal stem cells were investigated. The cells were cultured in treatment medium containing 100 nM of staurosporine for 4 hours; then the cells were affected by hydrostatic pressure (0, 25,50, 100 mmHg). The percentage of cell viability by trypan blue staining and the percentage of cell death by Hoechst/PI differential staining were assessed. We obtained the total neurite length. Expression of β-tubulin III and GFAP (Glial fibrillary acidic protein) proteins were also analyzed by immunocytochemistry. The percentage of cell viability in treatments decreased relative to the increase in hydrostatic pressure and time (p Keywords: bone marrow mesenchymal stem cell, hydrostatic pressure, immunocytochemistry, neural differentiation, neurite length, cell differentiation.

Decreasing maternal myostatin programs adult offspring bone strength in a mouse model of osteogenesis imperfecta.

PubMed

Oestreich, Arin K; Kamp, William M; McCray, Marcus G; Carleton, Stephanie M; Karasseva, Natalia; Lenz, Kristin L; Jeong, Youngjae; Daghlas, Salah A; Yao, Xiaomei; Wang, Yong; Pfeiffer, Ferris M; Ellersieck, Mark R; Schulz, Laura C; Phillips, Charlotte L

2016-11-22

During fetal development, the uterine environment can have effects on offspring bone architecture and integrity that persist into adulthood; however, the biochemical and molecular mechanisms remain unknown. Myostatin is a negative regulator of muscle mass. Parental myostatin deficiency (Mstn tm1Sjl/+ ) increases muscle mass in wild-type offspring, suggesting an intrauterine programming effect. Here, we hypothesized that Mstn tm1Sjl/+ dams would also confer increased bone strength. In wild-type offspring, maternal myostatin deficiency altered fetal growth and calvarial collagen content of newborn mice and conferred a lasting impact on bone geometry and biomechanical integrity of offspring at 4 mo of age, the age of peak bone mass. Second, we sought to apply maternal myostatin deficiency to a mouse model with osteogenesis imperfecta (Col1a2 oim ), a heritable connective tissue disorder caused by abnormalities in the structure and/or synthesis of type I collagen. Femora of male Col1a2 oim/+ offspring from natural mating of Mstn tm1Sjl/+ dams to Col1a2 oim/+ sires had a 15% increase in torsional ultimate strength, a 29% increase in tensile strength, and a 24% increase in energy to failure compared with age, sex, and genotype-matched offspring from natural mating of Col1a2 oim/+ dams to Col1a2 oim/+ sires. Finally, increased bone biomechanical strength of Col1a2 oim/+ offspring that had been transferred into Mstn tm1Sjl/+ dams as blastocysts demonstrated that the effects of maternal myostatin deficiency were conferred by the postimplantation environment. Thus, targeting the gestational environment, and specifically prenatal myostatin pathways, provides a potential therapeutic window and an approach for treating osteogenesis imperfecta.

Suppression of the motor deficit in a mucolipidosis type IV mouse model by bone marrow transplantation

PubMed Central

Walker, Marquis T.; Montell, Craig

2016-01-01

Mucolipidosis IV (MLIV) is a severe lysosomal storage disorder, which results from loss of the TRPML1 channel. MLIV causes multiple impairments in young children, including severe motor deficits. Currently, there is no effective treatment. Using a Drosophila MLIV model, we showed previously that introduction of trpml+ in phagocytic glia rescued the locomotor deficit by removing early dying neurons, thereby preventing amplification of neuronal death from cytotoxicity. Because microglia, which are phagocytic cells in the mammalian brain, are bone marrow derived, and cross the blood–brain barrier, we used a mouse MLIV model to test the efficacy of bone marrow transplantation (BMT). We found that BMT suppressed the reduced myelination and the increased caspase-3 activity due to loss of TRPML1. Using a rotarod test, we demonstrated that early BMT greatly delayed the motor impairment in the mutant mice. These data offer the possibility that BMT might provide the first therapy for MLIV. PMID:27270598

Breed-specific incidence rates of canine primary bone tumors — A population based survey of dogs in Norway

PubMed Central

Anfinsen, Kristin P.; Grotmol, Tom; Bruland, Oyvind S.; Jonasdottir, Thora J.

2011-01-01

This is one of few published population-based studies describing breed specific rates of canine primary bone tumors. Incidence rates related to dog breeds could help clarify the impact of etiological factors such as birth weight, growth rate, and adult body weight/height on development of these tumors. The study population consisted of dogs within 4 large/giant breeds; Irish wolfhound (IW), Leonberger (LB), Newfoundland (NF), and Labrador retriever (LR), born between January 1st 1989 and December 31st 1998. Questionnaires distributed to owners of randomly selected dogs — fulfilling the criteria of breed, year of birth, and registration in the Norwegian Kennel Club — constituted the basis for this retrospective, population-based survey. Of the 3748 questionnaires received by owners, 1915 were completed, giving a response rate of 51%. Forty-three dogs had been diagnosed with primary bone tumors, based upon clinical examination and x-rays. The breeds IW and LB, with 126 and 72 cases per 10 000 dog years at risk (DYAR), respectively, had significantly higher incidence rates of primary bone tumors than NF and LR (P < 0.0001). Incidence rates for the latter were 11 and 2 cases per 10 000 DYAR, respectively. Pursuing a search for risk factors other than body size/weight is supported by the significantly different risks of developing primary bone tumors between similarly statured dogs, like NF and LB, observed in this study. Defining these breed-specific incidence rates enables subsequent case control studies, ultimately aiming to identify specific etiological factors for developing primary bone tumors. PMID:22210997

Oncostatin M promotes bone formation independently of resorption when signaling through leukemia inhibitory factor receptor in mice

PubMed Central

Walker, Emma C.; McGregor, Narelle E.; Poulton, Ingrid J.; Solano, Melissa; Pompolo, Sueli; Fernandes, Tania J.; Constable, Matthew J.; Nicholson, Geoff C.; Zhang, Jian-Guo; Nicola, Nicos A.; Gillespie, Matthew T.; Martin, T. John; Sims, Natalie A.

2010-01-01

Effective osteoporosis therapy requires agents that increase the amount and/or quality of bone. Any modification of osteoclast-mediated bone resorption by disease or drug treatment, however, elicits a parallel change in osteoblast-mediated bone formation because the processes are tightly coupled. Anabolic approaches now focus on uncoupling osteoblast action from osteoclast formation, for example, by inhibiting sclerostin, an inhibitor of bone formation that does not influence osteoclast differentiation. Here, we report that oncostatin M (OSM) is produced by osteoblasts and osteocytes in mouse bone and that it has distinct effects when acting through 2 different receptors, OSM receptor (OSMR) and leukemia inhibitory factor receptor (LIFR). Specifically, mouse OSM (mOSM) inhibited sclerostin production in a stromal cell line and in primary murine osteoblast cultures by acting through LIFR. In contrast, when acting through OSMR, mOSM stimulated RANKL production and osteoclast formation. A key role for OSMR in bone turnover was confirmed by the osteopetrotic phenotype of mice lacking OSMR. Furthermore, in contrast to the accepted model, in which mOSM acts only through OSMR, mOSM inhibited sclerostin expression in Osmr–/– osteoblasts and enhanced bone formation in vivo. These data reveal what we believe to be a novel pathway by which bone formation can be stimulated independently of bone resorption and provide new insights into OSMR and LIFR signaling that are relevant to other medical conditions, including cardiovascular and neurodegenerative diseases and cancer. PMID:20051625

Bone tumor

MedlinePlus

Tumor - bone; Bone cancer; Primary bone tumor; Secondary bone tumor; Bone tumor - benign ... The cause of bone tumors is unknown. They often occur in areas of the bone that grow rapidly. Possible causes include: Genetic defects ...

[Knockdown of dopamine receptor D2 upregulates the expression of adiogenic genes in mouse primary mesencephalic neurons].

PubMed

Ding, Jiaqi; Chen, Xiaoli; Lin, Jiaji; Zhu, Junling; Li, Zhuyi

2018-01-01

Objective To study the effects of dopamine receptor D2 (DRD2) on the adipogenesis genes in mouse primary mesencephalic neurons. Methods The lentiviral vectors which expressed specific shRNA targeting DRD2 were constructed to decrease DRD2 expression in mouse primary mesencephalic neurons. High throughput sequencing (HTS) analysis was used to investigate gene expression changes between the DRD2 knock-down group and the negative control group. Real-time quantitative PCR (qRT-PCR) and Western blot analysis were applied to verify the differently expressed genes. Fatty acids were measured by fatty acid detection kit. Results DRD2 expression was effectively down-regulated in mouse primary mesencephalic neurons by lentiviral vectors. HTS revealed adipogenesis genes were significantly up-regulated after DRD2 down-regulation, mainly including delta(14)-sterol reductase, acetyl-coenzyme A synthetase, insulin-induced gene 1 protein and especially stearoyl-coenzyme A desaturase 1 (SCD1, 4-fold upregulated). The qRT-PCR and Western blot analysis verified that SCD1 was upregulated 2.6 folds and 2 folds respectively by lentiviral DRD2-shRNA vectors. Moreover, the SCD1-related free fatty acids were significantly more increased than the negative control group. Conclusion DRD2 in primary mesencephalic neurons had a significant regulative effect on the adipogenesis genes. The up-regulation of SCD1 can accelerate the conversion of saturated fatty acids to monounsaturated fatty acids and prevent the damage of lipid toxicity to cells.

Structurally-diverse, PPARγ-activating environmental toxicants induce adipogenesis and suppress osteogenesis in bone marrow mesenchymal stromal cells

PubMed Central

Watt, James; Schlezinger, Jennifer J.

2015-01-01

Environmental obesogens are a newly recognized category of endocrine disrupting chemicals that have been implicated in contributing to the rising rates of obesity in the United States. While obesity is typically regarded as an increase in visceral fat, adipocyte accumulation in the bone has been linked to increased fracture risk, lower bone density, and osteoporosis. Exposure to environmental toxicants that activate peroxisome proliferator activated receptor γ (PPARγ), a critical regulator of the balance of differentiation between adipogenesis and osteogenesis, may contribute to the increasing prevalence of osteoporosis. However, induction of adipogenesis and suppression of osteogenesis are separable activities of PPARγ, and ligands may selectively alter these activities. It currently is unknown whether suppression of osteogenesis is a common toxic endpoint of environmental PPARγ ligands. Using a primary mouse bone marrow culture model, we tested the hypothesis that environmental toxicants acting as PPARγ agonists divert the differentiation pathway of bone marrow-derived multipotent mesenchymal stromal cells towards adipogenesis and away from osteogenesis. The toxicants tested included the organotins tributyltin and triphenyltin, a ubiquitous phthalate metabolite (mono-(2-ethylhexyl) phthalate, MEHP), and two brominated flame retardants (tetrabromobisphenol-a, TBBPA, and mono-(2-ethylhexyl) tetrabromophthalate, METBP). All of the compounds activated PPARγ1 and 2. All compounds increased adipogenesis (lipid accumulation, Fabp4 expression) and suppressed osteogenesis (alkaline phosphatase activity, Osx expression) in mouse primary bone marrow cultures, but with different potencies and efficacies. Despite structural dissimilarities, there was a strong negative correlation between efficacies to induce adipogenesis and suppress osteogenesis, with the organotins being distinct in their exceptional ability to suppress osteogenesis. As human exposure to a mixture of

Immunostimulatory effect of spinach aqueous extract on mouse macrophage-like J774.1 cells and mouse primary peritoneal macrophages.

PubMed

Ishida, Momoko; Ose, Saya; Nishi, Kosuke; Sugahara, Takuya

2016-07-01

We herein report the immunostimulatory effect of spinach aqueous extract (SAE) on mouse macrophage-like J774.1 cells and mouse primary peritoneal macrophages. SAE significantly enhanced the production of interleukin (IL)-6 and tumor necrosis factor-α by both J774.1 cells and peritoneal macrophages by enhancing the expression levels of these cytokine genes. In addition, the phagocytosis activity of J774.1 cells was facilitated by SAE. Immunoblot analysis revealed that SAE activates mitogen-activated protein kinase and nuclear factor-κB cascades. It was found that SAE activates macrophages through not only TLR4, but also other receptors. The production of IL-6 was significantly enhanced by peritoneal macrophages from SAE-administered BALB/c mice, suggesting that SAE has a potential to stimulate macrophage activity in vivo. Taken together, these data indicate that SAE would be a beneficial functional food with immunostimulatory effects on macrophages.

The effects of pilocarpine nitrate upon the mitotic index of mouse bone marrow cells.

PubMed

Prabhu, M P; Hegde, M J

1991-11-01

Aneuploidies are the most common chromosomal causes for spontaneous abortions and constitute a major part of genetic disorders among the neonates. Aneuploidy producing agents (aneugens) pose serious genetic hazards to the human population. Therefore, testing for aneuploidy induction should be part of the requirement in drug safety guidelines. The aneugenic potential of pilocarpine nitrate, an alkaloid drug used as an ophthalmic solution was screened by chromosome analysis studies in the bone marrow cells of mice. Using the technique developed by Miller and Adler (1989) we evaluated changes in the mitotic index (MI), induction of chromatid contraction and spreading (C-mitoses) and decrease of anaphase frequencies as indicators of the aneuploidy inducing potency of the drug. Concentrations of pilocarpine nitrate of 4, 8 and 12 mg/kg body weight were administered intraperitoneally to mice. Colchicine-treated and water-treated animals formed the positive and negative controls. The data obtained in the cytogenetic analysis of both dose-response and time-response studies showed a significant induction of C-mitotic effects in the mouse bone marrow. The positive results indicated that the drug is a potential aneugen and should be further evaluated.

Endochondral bone formation in embryonic mouse pre-metatarsals

NASA Technical Reports Server (NTRS)

Klement, B. J.; Spooner, B. S.

1992-01-01

Long term exposure to a reduced gravitational environment has a deleterious effect on bone. The developmental events which occur prior to initial bone deposition will provide insight into the regulation of mature bone physiology. We have characterized a system in which the events preceding bone formation take place in an isolated in vitro organ culture environment. We show that cultured pre-metatarsal tissue parallels development of pre-metatarsal tissue in the embryo. Both undergo mesenchyme differentiation and morphogenesis to form a cartilage rod, which resembles the future bone, followed by terminal chondrocyte differentiation in a definite morphogenetic pattern. These sequential steps occur prior to osteoblast maturation and bone matrix deposition in the developing organism. Alkaline phosphatase (ALP) activity is a distinctive enzymatic marker for mineralizing tissues. We have measured this activity throughout pre-metatarsal development and show (a) where in the tissue it is predominantly found, and (b) that this is indeed the mineralizing isoform of the enzyme.

Bone marrow cell migration to the heart in a chimeric mouse model of acute chagasic disease.

PubMed

Irion, Camila Iansen; Paredes, Bruno Diaz; Brasil, Guilherme Visconde; Cunha, Sandro Torrentes da; Paula, Luis Felipe; Carvalho, Alysson Roncally; Carvalho, Antonio Carlos Campos de; Carvalho, Adriana Bastos; Goldenberg, Regina Coeli Dos Santos

2017-08-01

Chagas disease is a public health problem caused by infection with the protozoan Trypanosoma cruzi. There is currently no effective therapy for Chagas disease. Although there is some evidence for the beneficial effect of bone marrow-derived cells in chagasic disease, the mechanisms underlying their effects in the heart are unknown. Reports have suggested that bone marrow cells are recruited to the chagasic heart; however, studies using chimeric mouse models of chagasic cardiomyopathy are rare. The aim of this study was to investigate the migration of bone marrow cells to the heart after T. cruzi infection in a model of chagasic disease in chimeric mice. To obtain chimerical mice, wild-type (WT) C57BL6 mice were exposed to full body irradiation (7 Gy), causing bone marrow ablation. Then, bone marrow cells from green fluorescent protein (GFP)-transgenic mice were infused into the mice. Graft effectiveness was confirmed by flow cytometry. Experimental mice were divided into four groups: (i) infected chimeric (iChim) mice; (ii) infected WT (iWT) mice, both of which received 3 × 104 trypomastigotes of the Brazil strain; (iii) non-infected chimeric (Chim) mice; and (iv) non-infected WT mice. At one-month post-infection, iChim and iWT mice showed first degree atrioventricular block with decreased heart rate and treadmill exercise parameters compared to those in the non-infected groups. iChim mice showed an increase in parasitaemia, myocarditis, and the presence of amastigote nests in the heart tissue compared to iWT mice. Flow cytometry analysis did not detect haematopoietic progenitor cells in the hearts of infected mice. Furthermore, GFP+ cardiomyocytes were not detected in the tissues of chimeric mice.

Promoting Endochondral Bone Repair Using Human Osteoarthritic Articular Chondrocytes.

PubMed

Bahney, Chelsea S; Jacobs, Linsey; Tamai, Robert; Hu, Diane; Luan, Tammy F; Wang, Miqi; Reddy, Sanjay; Park, Michelle; Limburg, Sonja; Kim, Hubert T; Marcucio, Ralph; Kuo, Alfred C

2016-03-01

Current tissue engineering strategies to heal critical-size bone defects through direct bone formation are limited by incomplete integration of grafts with host bone and incomplete graft vascularization. An alternative strategy for bone regeneration is the use of cartilage grafts that form bone through endochondral ossification. Endochondral cartilages stimulate angiogenesis and are remodeled into bone, but are found in very small quantities in growth plates and healing fractures. We sought to develop engineered endochondral cartilage grafts using osteoarthritic (OA) articular chondrocytes as a cell source. Such chondrocytes often undergo hypertrophy, which is a characteristic of endochondral cartilages. We compared the ability of unmodified human OA (hOA) cartilage and cartilage grafts formed in vitro from hOA chondrocytes to undergo endochondral ossification in mice. Scaffold-free engineered chondrocyte grafts were generated by pelleting chondrocytes, followed by culture with transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein 4. Samples derived from either primary or passaged chondrocytes were implanted subcutaneously into immunocompromised mice. Grafts derived from passaged chondrocytes from three patients were implanted into critical-size tibial defects in mice. Bone formation was assessed with histology after 4 weeks of implantation. The composition of tibial repair tissue was quantified with histomorphometry. Engineered cartilage grafts generated from passaged OA chondrocytes underwent endochondral ossification after implantation either subcutaneously or in bone. Cartilage grafts integrated with host bone at 15 out of 16 junctions. Grafts variably remodeled into woven bone, with the proportion of bony repair tissue in tibial defects ranging from 22% to 85% (average 48%). Bony repair tissue bridged the tibial defects in half of the animals. In contrast, unmodified OA cartilage and engineered grafts formed from primary chondrocytes did not

FDG-PET/CT Imaging Predicts Histopathologic Treatment Responses after Neoadjuvant Therapy in Adult Primary Bone Sarcomas

DOE PAGES

Benz, Matthias R.; Czernin, Johannes; Tap, William D.; ...

2010-01-01

Purpose . Tmore » he aim of this study was to prospectively evaluate whether FDG-PET allows an accurate assessment of histopathologic response to neoadjuvant treatment in adult patients with primary bone sarcomas. Methods . Twelve consecutive patients with resectable, primary high grade bone sarcomas were enrolled prospectively. FDG-PET/CT imaging was performed prior to the initiation and after completion of neoadjuvant treatment. Imaging findings were correlated with histopathologic response. Results . Histopathologic responders showed significantly more pronounced decreases in tumor FDG-SUVmax from baseline to late follow up than non-responders ( 64 ± 19 % versus 29 ± 30 %, resp.; P = .03 ). Using a 60% decrease in tumor FDG-uptake as a threshold for metabolic response correctly classified 3 of 4 histopathologic responders and 7 of 8 histopathologic non-responders as metabolic responders and non-responders, respectively (sensitivity, 75%; specificity, 88%). Conclusion . These results suggest that changes in FDG-SUVmax at the end of neoadjuvant treatment can identify histopathologic responders and non-responders in adult primary bone sarcoma patients.« less

Missense Mutations in LRP5 Associated with High Bone Mass Protect the Mouse Skeleton from Disuse- and Ovariectomy-Induced Osteopenia.

PubMed

Niziolek, Paul J; Bullock, Whitney; Warman, Matthew L; Robling, Alexander G

2015-01-01

The low density lipoprotein receptor-related protein-5 (LRP5), a co-receptor in the Wnt signaling pathway, modulates bone mass in humans and in mice. Lrp5 knock-out mice have severely impaired responsiveness to mechanical stimulation whereas Lrp5 gain-of-function knock-in and transgenic mice have enhanced responsiveness to mechanical stimulation. Those observations highlight the importance of Lrp5 protein in bone cell mechanotransduction. It is unclear if and how high bone mass-causing (HBM) point mutations in Lrp5 alter the bone-wasting effects of mechanical disuse. To address this issue we explored the skeletal effects of mechanical disuse using two models, tail suspension and Botulinum toxin-induced muscle paralysis, in two different Lrp5 HBM knock-in mouse models. A separate experiment employing estrogen withdrawal-induced bone loss by ovariectomy was also conducted as a control. Both disuse stimuli induced significant bone loss in WT mice, but Lrp5 A214V and G171V were partially or fully protected from the bone loss that normally results from disuse. Trabecular bone parameters among HBM mice were significantly affected by disuse in both models, but these data are consistent with DEXA data showing a failure to continue growing in HBM mice, rather than a loss of pre-existing bone. Ovariectomy in Lrp5 HBM mice resulted in similar protection from catabolism as was observed for the disuse experiments. In conclusion, the Lrp5 HBM alleles offer significant protection from the resorptive effects of disuse and from estrogen withdrawal, and consequently, present a potential mechanism to mimic with pharmaceutical intervention to protect against various bone-wasting stimuli.

Orientation selectivity of synaptic input to neurons in mouse and cat primary visual cortex.

PubMed

Tan, Andrew Y Y; Brown, Brandon D; Scholl, Benjamin; Mohanty, Deepankar; Priebe, Nicholas J

2011-08-24

Primary visual cortex (V1) is the site at which orientation selectivity emerges in mammals: visual thalamus afferents to V1 respond equally to all stimulus orientations, whereas their target V1 neurons respond selectively to stimulus orientation. The emergence of orientation selectivity in V1 has long served as a model for investigating cortical computation. Recent evidence for orientation selectivity in mouse V1 opens cortical computation to dissection by genetic and imaging tools, but also raises two essential questions: (1) How does orientation selectivity in mouse V1 neurons compare with that in previously described species? (2) What is the synaptic basis for orientation selectivity in mouse V1? A comparison of orientation selectivity in mouse and in cat, where such measures have traditionally been made, reveals that orientation selectivity in mouse V1 is weaker than in cat V1, but that spike threshold plays a similar role in narrowing selectivity between membrane potential and spike rate. To uncover the synaptic basis for orientation selectivity, we made whole-cell recordings in vivo from mouse V1 neurons, comparing neuronal input selectivity-based on membrane potential, synaptic excitation, and synaptic inhibition-to output selectivity based on spiking. We found that a neuron's excitatory and inhibitory inputs are selective for the same stimulus orientations as is its membrane potential response, and that inhibitory selectivity is not broader than excitatory selectivity. Inhibition has different dynamics than excitation, adapting more rapidly. In neurons with temporally modulated responses, the timing of excitation and inhibition was different in mice and cats.

Orientation Selectivity of Synaptic Input to Neurons in Mouse and Cat Primary Visual Cortex

PubMed Central

Tan (陈勇毅), Andrew Y. Y.; Brown, Brandon D.; Scholl, Benjamin; Mohanty, Deepankar; Priebe, Nicholas J.

2011-01-01

Primary visual cortex (V1) is the site at which orientation selectivity emerges in mammals: visual thalamus afferents to V1 respond equally to all stimulus orientations whereas their target V1 neurons respond selectively to stimulus orientation. The emergence of orientation selectivity in V1 has long served as a model for investigating cortical computation. Recent evidence for orientation selectivity in mouse V1 opens cortical computation to dissection by genetic and imaging tools, but also raises two essential questions: 1) how does orientation selectivity in mouse V1 neurons compare with that in previously described species? 2) what is the synaptic basis for orientation selectivity in mouse V1? A comparison of orientation selectivity in mouse and in cat, where such measures have traditionally been made, reveals that orientation selectivity in mouse V1 is weaker than in cat V1, but that spike threshold plays a similar role in narrowing selectivity between membrane potential and spike rate. To uncover the synaptic basis for orientation selectivity, we made whole-cell recordings in vivo from mouse V1 neurons, comparing neuronal input selectivity - based on membrane potential, synaptic excitation, and synaptic inhibition - to output selectivity based on spiking. We found that a neuron's excitatory and inhibitory inputs are selective for the same stimulus orientations as is its membrane potential response, and that inhibitory selectivity is not broader than excitatory selectivity. Inhibition has different dynamics than excitation, adapting more rapidly. In neurons with temporally modulated responses, the timing of excitation and inhibition was different in mice and cats. PMID:21865476

Bone morphogenetic protein signaling is impaired in an Hfe knockout mouse model of hemochromatosis

PubMed Central

Corradini, Elena; Garuti, Cinzia; Montosi, Giuliana; Ventura, Paolo; Andriopoulos, Billy; Lin, Herbert Y.; Pietrangelo, Antonello; Babitt, Jodie L.

2009-01-01

Background and Aims Mutations in HFE are the most common cause of the iron-overload disorder hereditary hemochromatosis (HH). Levels of the main iron regulatory hormone, hepcidin, are inappropriately low in HH mouse models and patients with HFE mutations, indicating that HFE regulates hepcidin. The bone morphogenetic protein 6 (BMP6)-SMAD signaling pathway is an important endogenous regulator of hepcidin expression. We investigated whether HFE is involved in BMP6-SMAD regulation of hepcidin expression. Methods The BMP6-SMAD pathway was examined in Hfe knockout (KO) mice and in wild-type (WT) mice as controls. Mice were placed on diets of varying iron content. Hepcidin induction by BMP6 was examined in primary hepatocytes from Hfe KO mice; data were compared with those of WT mice. Results Liver levels of Bmp6 mRNA were higher in Hfe KO mice; these were appropriate for the increased hepatic levels of iron in these mice, compared with WT mice. However, levels of hepatic phosphorylated Smad 1/5/8 protein (an intracellular mediator of Bmp6 signaling) and Id1 mRNA (a target gene of Bmp6) were inappropriately low for the body iron burden and Bmp6 mRNA levels in Hfe KO, compared with WT mice. BMP6 induction of hepcidin expression was reduced in Hfe KO hepatocytes compared with WT hepatocytes. Conclusions HFE is not involved in regulation of BMP6 by iron, but does regulate the downstream signals of BMP6 that are triggered by iron. PMID:19591830

Tributyltin engages multiple nuclear receptor pathways and suppresses osteogenesis in bone marrow multipotent stromal cells.

PubMed

Baker, Amelia H; Watt, James; Huang, Cassie K; Gerstenfeld, Louis C; Schlezinger, Jennifer J

2015-06-15

Organotins are members of the environmental obesogen class of contaminants because they activate peroxisome proliferator-activated receptor γ (PPARγ), the essential regulator of adipogenesis. Exposure to thiazolidinediones (PPARγ ligands used to treat type 2 diabetes) is associated with increased fractures. Diminished bone quality likely results from PPARγ's role in promoting adipogenesis while suppressing osteogenesis of bone marrow multipotent mesenchymal stromal cells (BM-MSC). We hypothesized that tributyltin (TBT) would be a potent modifier of BM-MSC differentiation and a negative regulator of bone formation. Organotins interact with both PPARγ and retinoid X receptors (RXR), suggesting that they activate multiple nuclear receptor pathways. To investigate the role of RXR in the actions of TBT, the effects of PPARγ (rosiglitazone) and RXR (bexarotene, LG100268) agonists were compared to the effects of TBT in BMS2 cells and primary mouse BM-MSC cultures. In BMS2 cells, TBT induced the expression of Fabp4, Abca1, and Tgm2 in an RXR-dependent manner. All agonists suppressed osteogenesis in primary mouse BM-MSC cultures, based on decreased alkaline phosphatase activity, mineralization, and expression of osteoblast-related genes. While rosiglitazone and TBT strongly activated adipogenesis, based on lipid accumulation and expression of adipocyte-related genes, the RXR agonists did not. Extending these analyses to other RXR heterodimers showed that TBT and the RXR agonists activated the liver X receptor pathway, whereas rosiglitazone did not. Application of either a PPARγ antagonist (T0070907) or an RXR antagonist (HX531) significantly reduced rosiglitazone-induced suppression of bone nodule formation. Only the RXR antagonist significantly reduced LG100268- and TBT-induced bone suppression. The RXR antagonist also inhibited LG100268- and TBT-induced expression of Abca1, an LXR target gene, in primary BM-MSC cultures. These results provide novel evidence that

Wnt inhibition enhances browning of mouse primary white adipocytes.

PubMed

Lo, Kinyui Alice; Ng, Pei Yi; Kabiri, Zahra; Virshup, David; Sun, Lei

2016-01-01

The global epidemic in obesity and metabolic syndrome requires novel approaches to tackle. White adipose tissue, traditionally seen as a passive energy-storage organ, can be induced to take on certain characteristics of brown fat in a process called browning. The "browned" white adipose tissue, or beige fat, is a potential anti-obesity target. Various signaling pathways can enhance browning. Wnt is a key regulator of adipocyte biology, but its role in browning has not been explored. In this study, we found that in primary mouse adipocytes derived from the inguinal depot, Wnt inhibition by both chemical and genetic methods significantly enhanced browning. The effect of Wnt inhibition on browning most likely targets the beige precursor cells in selected adipose depots.

The effects of hydroxyapatite coating and bone allograft on fixation of loaded experimental primary and revision implants.

PubMed

Søballe, Kjeld; Mouzin, Olivier R G; Kidder, Louis A; Overgaard, Søren; Bechtold, Joan E

2003-06-01

We used our established experimental model of revision joint replacement to examine the roles of hydroxyapatite coating and bone graft in improving the fixation of revision implants. The revision protocol uses the Søballe micromotion device in a preliminary 8-week period of implant instability for the presence of particulate polyethylene. During this procedure, a sclerotic endosteal bone rim forms, and a dense fibrous membrane is engendered, having macrophages with ingested polyethylene and high levels of inflammatory cytokines. At the time of revision after 8 weeks, the cavity is revised with either a titanium alloy (Ti) or a hydroxyapatite (HA) 6.0 mm plasma-sprayed implant, in the presence or absence of allograft packed into the initial 0.75 mm peri-implant gap. The contralateral limb is subjected to primary surgery with the same implant configuration, and serves as control. 8 implants were included in each of the 8 treatment groups (total 64 implants in 32 dogs). The observation period was 4 weeks after revision. Outcome measures are based on histomorphometry and mechanical pushout properties. The revision setting was always inferior to its primary counterpart. Bone graft improved the revision fixation in all treatment groups, as also did the HA coating. The sole exception was revision-grafted HA implants, which reached the same fixation as primary Ti and HA grafted implants. The revision, which was less active in general, seems to need the dual stimulation of bone graft and HA implant surface, to obtain the same level of fixation associated with primary implants. Our findings suggest that the combination of HA implant and bone graft may be of benefit in the clinical revision implant setting.

The effects of hydroxyapatite coating and bone allograft on fixation of loaded experimental primary and revision implants

PubMed Central

Søballe, Kjeld; Mouzin, Olivier R G; Kidder, Louis A; Overgaard, Søren; Bechtold, Joan E

2015-01-01

We used our established experimental model of revision joint replacement to examine the roles of hydroxyapatite coating and bone graft in improving the fixation of revision implants. The revision protocol uses the Søballe micromotion device in a preliminary 8-week period of implant instability for the presence of particulate polyethylene. During this procedure, a sclerotic endosteal bone rim forms, and a dense fibrous membrane is engendered, having macrophages with ingested polyethylene and high levels of inflammatory cytokines. At the time of revision after 8 weeks, the cavity is revised with either a titanium alloy (Ti) or a hydroxyapatite (HA) 6.0 mm plasma-sprayed implant, in the presence or absence of allograft packed into the initial 0.75 mm peri-implant gap. The contralateral limb is subjected to primary surgery with the same implant configuration, and serves as control. 8 implants were included in each of the 8 treatment groups (total 64 implants in 32 dogs). The observation period was 4 weeks after revision. Outcome measures are based on histomorphometry and mechanical pushout properties. The revision setting was always inferior to its primary counterpart. Bone graft improved the revision fixation in all treatment groups, as also did the HA coating. The sole exception was revision-grafted HA implants, which reached the same fixation as primary Ti and HA grafted implants. The revision, which was less active in general, seems to need the dual stimulation of bone graft and HA implant surface, to obtain the same level of fixation associated with primary implants. Our findings suggest that the combination of HA implant and bone graft may be of benefit in the clinical revision implant setting. PMID:12899541

Monomethylarsonous acid (MMA+3) Inhibits IL-7 Signaling in Mouse Pre-B Cells.

PubMed

Ezeh, Peace C; Xu, Huan; Lauer, Fredine T; Liu, Ke Jian; Hudson, Laurie G; Burchiel, Scott W

2016-02-01

Our previously published data show that As(+3) in vivo and in vitro, at very low concentrations, inhibits lymphoid, but not myeloid stem cell development in mouse bone marrow. We also showed that the As(+3) metabolite, monomethylarsonous acid (MMA(+3)), was responsible for the observed pre-B cell toxicity caused by As(+3). Interleukin-7 (IL-7) is the primary growth factor responsible for pre-lymphoid development in mouse and human bone marrow, and Signal Transducer and Activator of Transcription 5 (STAT5) is a transcriptional factor in the IL-7 signaling pathway. We found that MMA(+3) inhibited STAT5 phosphorylation at a concentration as low as 50 nM in mouse bone marrow pre-B cells. Inhibition of STAT5 phosphorylation by As(+3) occurred only at a concentration of 500 nM. In the IL-7 dependent mouse pre-B 2E8 cell line, we also found selective inhibition of STAT5 phosphorylation by MMA(+3), and this inhibition was dependent on effects on JAK3 phosphorylation. IL-7 receptor expression on 2E8 cell surface was also suppressed by 50 nM MMA(+3) at 18 h. As further evidence for the inhibition of STAT5, we found that the induction of several genes required in B cell development, cyclin D1, E2A, EBF1, and PAX5, were selectively inhibited by MMA(+3). Since 2E8 cells lack the enzymes responsible for the conversion of As(+3) to MMA(+3) in vitro, the results of these studies suggest that As(+3) induced inhibition of pre-B cell formation in vivo is likely dependent on the formation of MMA(+3) which in turn inhibits IL-7 signaling at several steps in mouse pre-B cells. © The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Intraperitoneal injection of thalidomide attenuates bone cancer pain and decreases spinal tumor necrosis factor-α expression in a mouse model

PubMed Central

2010-01-01

Background Tumor necrosis factor α (TNF-α) may have a pivotal role in the genesis of mechanical allodynia and thermal hyperalgesia during inflammatory and neuropathic pain. Thalidomide has been shown to selectively inhibit TNF-α production. Previous studies have suggested that thalidomide exerts anti-nociceptive effects in various pain models, but its effects on bone cancer pain have not previously been studied. Therefore, in the present study, we investigated the effect of thalidomide on bone cancer-induced hyperalgesia and up-regulated expression of spinal TNF-α in a mouse model. Results Osteosarcoma NCTC 2472 cells were implanted into the intramedullary space of the right femurs of C3H/HeJ mice to induce ongoing bone cancer related pain behaviors. At day 5, 7, 10 and 14 after operation, the expression of TNF-α in the spinal cord was higher in tumor-bearing mice compared to the sham mice. Intraperitoneal injection of thalidomide (50 mg/kg), started at day 1 after surgery and once daily thereafter until day 7, attenuated bone cancer-evoked mechanical allodynia and thermal hyperalgesia as well as the up-regulation of TNF-α in the spinal cord. Conclusions These results suggest that thalidomide can efficiently alleviate bone cancer pain and it may be a useful alternative or adjunct therapy for bone cancer pain. Our data also suggest a role of spinal TNF-α in the development of bone cancer pain. PMID:20923560

Characterization of neutrophils and macrophages from ex vivo cultured murine bone marrow for morphologic maturation and functional responses by imaging flow cytometry

PubMed Central

Pelletier, Margery G. H.; Szymczak, Klaudia; Barbeau, Anna M.; Prata, Gianna N.; O’Fallon, Kevin S.; Gaines, Peter

2016-01-01

Neutrophils and macrophages differentiate from common myeloid progenitors in the bone marrow, where they undergo nuclear morphologic changes during maturation. During this process, both cell types acquire critical innate immune functions that include phagocytosis of pathogens, and for neutrophils the release of nuclear material called nuclear extracellular traps (NETs). Primary cells used to study these functions are typically purified from mature mouse tissues, but bone marrow-derived ex vivo cultures provide more abundant numbers of progenitors and functionally mature cells. Routine analyses of these cells use conventional microscopy and flow cytometry, which present limitations; microscopy is laborious and subjective, whereas flow cytometry lacks spatial resolution. Here we describe methods to generate enriched populations of neutrophils or macrophages from cryopreserved mouse bone marrow cultured ex vivo, and to use imaging flow cytometry that combines the resolution of microscopy with flow cytometry to analyze cells for morphologic features, phagocytosis, and NETosis. PMID:27663441

Alcohol Devitalization and Replantation for Primary Malignant Bone Tumors of the Knee Joint

PubMed Central

ZHANG, Xihai; CHEN, Ge; WANG, Jun; TANG, Lian; YIN, Yiran

2017-01-01

Background: This paper is aimed at studying the therapeutic effects of in situ replantation of alcohol-devitalized bone segments to treat malignant bone tumors of the knee joint. Methods: We retrospectively analyzed clinical data for 45 patients from January 2013 to January 2016 who underwent replantation following alcohol-devitalization of bone segments and 40 who underwent prosthesis implantation. The two groups were comparable in basal clinical biometric data, including gender, age, tumor type and location, Enneking staging, and maximum tumor diameter. Radical tumor resection was combined with neoadjuvant chemotherapy following the two-implantation procedures. Results: The median follow-up time was 25 months, and the outcomes were compared. We found no differences in the length of bone lesions, surgery time, intraoperative blood loss, amount of postoperative drainage, and perioperative complications, which were just three for each method. We also found no significant differences in limb function scores, internal fixation imaging scores, tumor-free survival rate, and overall survival rate between the two groups. Replantation following alcohol-devitalization of tumor-bearing bone segment demonstrated similar clinical outcomes compared with prosthesis implantation in the treatment of primary malignant bone tumors of the knee joint. Conclusion: Both therapies enjoy good application safety and effectiveness. Because alcohol devitalization is inexpensive and easy to apply in the clinic, it should be considered a preferred method in the treatment of bone tumors. PMID:29308374

Critical-Size Calvarial Bone Defects Healing in a Mouse Model with Silk Scaffolds and SATB2- Modified iPSCs

PubMed Central

Ye, Jin-Hai; Xu, Yuan-Jin; Gao, Jun; Yan, Shi-Guo; Zhao, Jun; Tu, Qisheng; Zhang, Jin; Duan, Xue-Jing; Sommer, Cesar A.; Mostoslavsky, Gustavo; Kaplan, David; Wu, Yu-Nong; Zhang, Chen-Ping; Wang, Lin; Chen, Jake

2011-01-01

Induced pluripotent stem cells (iPSCs) can differentiate into mineralizing cells and thus have a great potential in application in engineered bone substitutes with bioactive scaffolds in regeneration medicine. In the current study we characterized and demonstrated the pluripotency and osteogenic differentiation of mouse iPSCs. To enhance the osteogenic differentiation of iPSCs, we then transduced the iPSCs with the potent transcription factor, nuclear matrix protein SATB2. We observed that in SATB2-overexpressing iPSCs there were increased mineral nodule formation and elevated mRNA levels of key osteogenic genes, osterix (OSX), Runx2, bone sialoprotein (BSP) and osteocalcin (OCN). Moreover, the mRNA levels of HoxA2 was reduced after SATB2 overexpression in iPSCs. The SATB2-overexpressing iPSCs were then combined with silk scaffolds and transplanted into critical-size calvarial bone defects created in nude mice. Five weeks post-surgery, radiological and micro-CT analysis revealed enhanced new bone formation in calvarial defects in SATB2 group. Histological analysis also showed increased new bone formation and mineralization in the SATB2 group. In conclusion, the results demonstrate that SATB2 facilitates the differentiation of iPSCs towards osteoblast-lineage cells by repressing HoxA2 and augmenting the functions of the osteoblast determinants Runx2, BSP and OCN. PMID:21492931

Fourier Transform Infrared Imaging Microspectroscopy and Tissue-Level Mechanical Testing Reveal Intraspecies Variation in Mouse Bone Mineral and Matrix Composition

PubMed Central

Courtland, Hayden-William; Nasser, Philip; Goldstone, Andrew B.; Spevak, Lyudmila; Boskey, Adele L.; Jepsen, Karl J.

2009-01-01

Fracture susceptibility is heritable and dependent upon bone morphology and quality. However, studies of bone quality are typically overshadowed by emphasis on bone geometry and bone mineral density. Given that differences in mineral and matrix composition exist in a variety of species, we hypothesized that genetic variation in bone quality and tissue-level mechanical properties would also exist within species. Sixteen-week-old female A/J, C57BL/6J (B6), and C3H/HeJ (C3H) inbred mouse femora were analyzed using Fourier transform infrared imaging and tissue-level mechanical testing for variation in mineral composition, mineral maturity, collagen cross-link ratio, and tissue-level mechanical properties. A/J femora had an increased mineral-to-matrix ratio compared to B6. The C3H mineral-to-matrix ratio was intermediate of A/J and B6. C3H femora had reduced acid phosphate and carbonate levels and an increased collagen cross-link ratio compared to A/J and B6. Modulus values paralleled mineral-to-matrix values, with A/J femora being the most stiff, B6 being the least stiff, and C3H having intermediate stiffness. In addition, work-to-failure varied among the strains, with the highly mineralized and brittle A/J femora performing the least amount of work-to-failure. Inbred mice are therefore able to differentially modulate the composition of their bone mineral and the maturity of their bone matrix in conjunction with tissue-level mechanical properties. These results suggest that specific combinations of bone quality and morphological traits are genetically regulated such that mechanically functional bones can be constructed in different ways. PMID:18855037

Fourier transform infrared imaging microspectroscopy and tissue-level mechanical testing reveal intraspecies variation in mouse bone mineral and matrix composition.

PubMed

Courtland, Hayden-William; Nasser, Philip; Goldstone, Andrew B; Spevak, Lyudmila; Boskey, Adele L; Jepsen, Karl J

2008-11-01

Fracture susceptibility is heritable and dependent upon bone morphology and quality. However, studies of bone quality are typically overshadowed by emphasis on bone geometry and bone mineral density. Given that differences in mineral and matrix composition exist in a variety of species, we hypothesized that genetic variation in bone quality and tissue-level mechanical properties would also exist within species. Sixteen-week-old female A/J, C57BL/6J (B6), and C3H/HeJ (C3H) inbred mouse femora were analyzed using Fourier transform infrared imaging and tissue-level mechanical testing for variation in mineral composition, mineral maturity, collagen cross-link ratio, and tissue-level mechanical properties. A/J femora had an increased mineral-to-matrix ratio compared to B6. The C3H mineral-to-matrix ratio was intermediate of A/J and B6. C3H femora had reduced acid phosphate and carbonate levels and an increased collagen cross-link ratio compared to A/J and B6. Modulus values paralleled mineral-to-matrix values, with A/J femora being the most stiff, B6 being the least stiff, and C3H having intermediate stiffness. In addition, work-to-failure varied among the strains, with the highly mineralized and brittle A/J femora performing the least amount of work-to-failure. Inbred mice are therefore able to differentially modulate the composition of their bone mineral and the maturity of their bone matrix in conjunction with tissue-level mechanical properties. These results suggest that specific combinations of bone quality and morphological traits are genetically regulated such that mechanically functional bones can be constructed in different ways.

[Action of two pyrazine-containing chemosignals on cells of bone marrow and testes in male house mouse Mus musculus L].

PubMed

Daev, E V; Vyborova, A M; Kazarova, V É; Dukel'skaia, A V

2012-01-01

Evolutionary conservative chemosignal 2,5-dimethylpyrazin that is pheromone in female mice has been shown to increase frequency of mitotic aberrations analyzed with aid of metaphasic and ana-telophasic analysis in bone marrow cells. Replacement of one of methyl radicals in the pheromone molecule by the carboxyl radical reveals specificity of action of the used derivative: the frequency of disturbances revealed only by the ana-telophasic analysis increases, whereas by the metaphasic analysis, no induction of disturbance is detected. In the sperm head abnormality test there is shown a rise of the anomalies by both compounds. Possible mechanisms of specific action of the tested substances on stability of genetic apparatus of the bone marrow dividing cells in the house mouse are discussed.

Bone marrow cell migration to the heart in a chimeric mouse model of acute chagasic disease

PubMed Central

Irion, Camila Iansen; Paredes, Bruno Diaz; Brasil, Guilherme Visconde; da Cunha, Sandro Torrentes; Paula, Luis Felipe; Carvalho, Alysson Roncally; de Carvalho, Antonio Carlos Campos; Carvalho, Adriana Bastos; Goldenberg, Regina Coeli dos Santos

2017-01-01

BACKGROUND Chagas disease is a public health problem caused by infection with the protozoan Trypanosoma cruzi. There is currently no effective therapy for Chagas disease. Although there is some evidence for the beneficial effect of bone marrow-derived cells in chagasic disease, the mechanisms underlying their effects in the heart are unknown. Reports have suggested that bone marrow cells are recruited to the chagasic heart; however, studies using chimeric mouse models of chagasic cardiomyopathy are rare. OBJECTIVES The aim of this study was to investigate the migration of bone marrow cells to the heart after T. cruzi infection in a model of chagasic disease in chimeric mice. METHODS To obtain chimerical mice, wild-type (WT) C57BL6 mice were exposed to full body irradiation (7 Gy), causing bone marrow ablation. Then, bone marrow cells from green fluorescent protein (GFP)-transgenic mice were infused into the mice. Graft effectiveness was confirmed by flow cytometry. Experimental mice were divided into four groups: (i) infected chimeric (iChim) mice; (ii) infected WT (iWT) mice, both of which received 3 × 104 trypomastigotes of the Brazil strain; (iii) non-infected chimeric (Chim) mice; and (iv) non-infected WT mice. FINDINGS At one-month post-infection, iChim and iWT mice showed first degree atrioventricular block with decreased heart rate and treadmill exercise parameters compared to those in the non-infected groups. MAIN CONCLUSIONS iChim mice showed an increase in parasitaemia, myocarditis, and the presence of amastigote nests in the heart tissue compared to iWT mice. Flow cytometry analysis did not detect haematopoietic progenitor cells in the hearts of infected mice. Furthermore, GFP+ cardiomyocytes were not detected in the tissues of chimeric mice. PMID:28767980

Treatment with soluble activin type IIB-receptor improves bone mass and strength in a mouse model of Duchenne muscular dystrophy.

PubMed

Puolakkainen, Tero; Ma, Hongqian; Kainulainen, Heikki; Pasternack, Arja; Rantalainen, Timo; Ritvos, Olli; Heikinheimo, Kristiina; Hulmi, Juha J; Kiviranta, Riku

2017-01-19

Inhibition of activin/myostatin pathway has emerged as a novel approach to increase muscle mass and bone strength. Duchenne muscular dystrophy (DMD) is a neuromuscular disorder that leads to progressive muscle degeneration and also high incidence of fractures. The aim of our study was to test whether inhibition of activin receptor IIB ligands with or without exercise could improve bone strength in the mdx mouse model for DMD. Thirty-two mdx mice were divided to running and non-running groups and to receive either PBS control or soluble activin type IIB-receptor (ActRIIB-Fc) once weekly for 7 weeks. Treatment of mdx mice with ActRIIB-Fc resulted in significantly increased body and muscle weights in both sedentary and exercising mice. Femoral μCT analysis showed increased bone volume and trabecular number (BV/TV +80%, Tb.N +70%, P < 0.05) in both ActRIIB-Fc treated groups. Running also resulted in increased bone volume and trabecular number in PBS-treated mice. However, there was no significant difference in trabecular bone structure or volumetric bone mineral density between the ActRIIB-Fc and ActRIIB-Fc-R indicating that running did not further improve bone structure in ActRIIB-Fc-treated mice. ActRIIB-Fc increased bone mass also in vertebrae (BV/TV +20%, Tb.N +30%, P < 0.05) but the effects were more modest. The number of osteoclasts was decreased in histological analysis and the expression of several osteoblast marker genes was increased in ActRIIB-Fc treated mice suggesting decreased bone resorption and increased bone formation in these mice. Increased bone mass in femurs translated into enhanced bone strength in biomechanical testing as the maximum force and stiffness were significantly elevated in ActRIIB-Fc-treated mice. Our results indicate that treatment of mdx mice with the soluble ActRIIB-Fc results in a robust increase in bone mass, without any additive effect by voluntary running. Thus ActRIIB-Fc could be an attractive option in the treatment

Raman spectroscopy of bone metastasis

NASA Astrophysics Data System (ADS)

Esmonde-White, Karen A.; Sottnik, Joseph; Morris, Michael; Keller, Evan

2012-02-01

Raman spectroscopy of bone has been used to characterize chemical changes occurring in diseases such as osteoporosis, osteoarthritis and osteomyelitis. Metastasis of cancer into bone causes changes to bone quality that are similar to those observed in osteoporosis, such as decreased bone strength, but with an accelerated timeframe. In particular, osteolytic (bone degrading) lesions in bone metastasis have a marked effect on patient quality of life because of increased risk of fractures, pain, and hypercalcemia. We use Raman spectroscopy to examine bone from two different mouse models of osteolytic bone metastasis. Raman spectroscopy measures physicochemical information which cannot be obtained through standard biochemical and histological measurements. This study was reviewed and approved by the University of Michigan University Committee on the Care and Use of Animals. Two mouse models of prostate cancer bone metastasis, RM1 (n=3) and PC3-luc (n=4) were examined. Tibiae were injected with RM1 or PC3-luc cancer cells, while the contralateral tibiae received a placebo injection for use as controls. After 2 weeks of incubation, the mice were sacrificed and the tibiae were examined by Raman microspectroscopy (λ=785 nm). Spectroscopic markers corresponding to mineral stoichiometry, bone mineralization, and mineral crystallinity were compared in spectra from the cancerous and control tibiae. X-ray imaging of the tibia confirmed extensive osteolysis in the RM1 mice, with tumor invasion into adjoining soft tissue and moderate osteolysis in the PC3-luc mice. Raman spectroscopic markers indicate that osteolytic lesions are less mineralized than normal bone tissue, with an altered mineral stoichiometry and crystallinity.

Polycythemia is associated with bone loss and reduced osteoblast activity in mice.

PubMed

Oikonomidou, P R; Casu, C; Yang, Z; Crielaard, B; Shim, J H; Rivella, S; Vogiatzi, M G

2016-04-01

Increased fragility has been described in humans with polycythemia vera (PV). Herein, we describe an osteoporotic phenotype associated with decreased osteoblast activity in a mouse model of PV and another mouse of polycythemia and elevated circulating erythropoietin (EPO). Our results are important for patients with PV or those treated with recombinant EPO (rEPO). PV and other myeloproliferative syndromes have been recently associated with an increased risk for fractures. However, the presence of osteoporosis in these patients has not been well documented. EPO, a hormone primarily known to stimulate erythropoiesis, has been shown recently to regulate bone homeostasis in mice. The aim of this study was to examine the bone phenotype of a mouse model of PV and compare it to that of animals with polycythemia caused by elevated circulating EPO. Bone mass and remodeling were evaluated by micro-computed tomography and histomorphometry. The JAK2(V617F) knock-in mouse, a model of human PV, manifests polycythemia and low circulating EPO levels. Results from this mouse were compared to wild type (wt) controls and the tg6 transgenic mouse that shows polycythemia caused by increased constitutive expression of EPO. Compared to wt, both JAK2(V617F) and tg6 mice had a decrease in trabecular bone mass. Tg6 mice showed an additional modest decrease in cortical thickness and cortical bone volume per tissue volume (P < 0.01) suggesting a more severe bone phenotype than JAK2(V617F). Decreased osteoblast numbers and bone formation along with normal osteoclast numbers and activity were found in both mice. This study indicates that PV is associated with low bone mass and decreased osteoblast activity in mice. Our results support future studies of osteoporosis in affected humans. Polycythemia caused by chronically elevated circulating EPO also results in bone loss, and implications on patients treated with rEPO should be evaluated.

Dietary coral calcium and zeolite protects bone in a mouse model for postmenopausal bone loss.

PubMed

Banu, Jameela; Varela, Erika; Guerra, Juan M; Halade, Ganesh; Williams, Paul J; Bahadur, Ali N; Hanaoka, Kokichi; Fernandes, Gabriel

2012-12-01

In patients diagnosed with osteoporosis, calcium is lost from bones making them weaker and easily susceptible to fractures. Supplementation of calcium is highly recommended for such conditions. However, the source of calcium plays an important role in the amount of calcium that is assimilated into bone. We hypothesize that naturally occurring coral calcium and zeolite may prevent ovariectomy-induced bone loss. We have measured bone loss in ovariectomized mice supplemented with coral calcium and Zeolite. Female C57BL/6 mice were either sham-operated or ovariectomized and fed diets containing coral calcium or zeolite for 6 months. Serum was analyzed for bone biochemical markers and cytokines. Bones were analyzed using dual x-ray absorbtiometry, peripheral quantitative computed tomography, and micro-computed tomography densitometry. In the distal femoral metaphysis, total bone and cortical bone mass was restored and the endocortical surface was significantly decreased in coral calcium and zeolite fed ovariectomized (OVX) mice. Trabecular number and the ratio of bone volume to total volume was higher in OVX mice after coral calcium and zeolite feeding, while trabecular separation decreased in the different treatment OVX groups. Coral calcium protected bone to a lesser extent in the proximal tibia and lumbar vertebrae. Overall, coral calcium and zeolite may protect postmenopausal bone loss. Copyright © 2012 Elsevier Inc. All rights reserved.

Neuroglobin overexpression inhibits oxygen-glucose deprivation-induced mitochondrial permeability transition pore opening in primary cultured mouse cortical neurons.

PubMed

Yu, Zhanyang; Liu, Ning; Li, Yadan; Xu, Jianfeng; Wang, Xiaoying

2013-08-01

Neuroglobin (Ngb) is an endogenous neuroprotective molecule against hypoxic/ischemic brain injury, but the underlying mechanisms remain largely undefined. Our recent study revealed that Ngb can bind to voltage-dependent anion channel (VDAC), a regulator of mitochondria permeability transition (MPT). In this study we examined the role of Ngb in MPT pore (mPTP) opening following oxygen-glucose deprivation (OGD) in primary cultured mouse cortical neurons. Co-immunoprecipitation (Co-IP) and immunocytochemistry showed that the binding between Ngb and VDAC was increased after OGD compared to normoxia, indicating the OGD-enhanced Ngb-VDAC interaction. Ngb overexpression protected primary mouse cortical neurons from OGD-induced neuronal death, to an extent comparable to mPTP opening inhibitor, cyclosporine A (CsA) pretreatment. We further measured the role of Ngb in OGD-induced mPTP opening using Ngb overexpression and knockdown approaches in primary cultured neurons, and recombinant Ngb exposure to isolated mitochondria. Same as CsA pretreatment, Ngb overexpression significantly reduced OGD-induced mPTP opening markers including mitochondria swelling, mitochondrial NAD(+) release, and cytochrome c (Cyt c) release in primary cultured neurons. Recombinant Ngb incubation significantly reduced OGD-induced NAD(+) release and Cyt c release from isolated mitochondria. In contrast, Ngb knockdown significantly increased OGD-induced neuron death, and increased OGD-induced mitochondrial NAD(+) release and Cyt c release as well, and these outcomes could be rescued by CsA pretreatment. In summary, our results demonstrated that Ngb overexpression can inhibit OGD-induced mPTP opening in primary cultured mouse cortical neurons, which may be one of the molecular mechanisms of Ngb's neuroprotection. Copyright © 2013 Elsevier Inc. All rights reserved.

Primary extraskeletal myxoid chondrosarcoma of bone: Report of three cases and review of the literature.

PubMed

Finos, L; Righi, A; Frisoni, T; Gambarotti, M; Ghinelli, C; Benini, S; Vanel, D; Picci, P

2017-05-01

Extraskeletal myxoid chondrosarcoma is a rare neoplasm of soft tissue. The usual location is in deep parts of the proximal extremities and limb girdles in middle-aged adults. The bone location as primary location is extremely rare and few cases are reported. We present three cases arising in bone with molecular confirmation using both RT-PCR and FISH analysis. Patients include two men and one woman with an age of 62, 69 and 73 years old. The mean size of the lesion was 13cm (range 8-18cm). Tumors arose in the iliac bone in two cases and in the proximal humerus in the other case. At time of diagnosis the three cases show bone cortex and soft tissue involvement. On imaging, lesions have a lobular pattern, are purely lytic, but take up contrast medium after injection. Two patients are alive with disease (local recurrence and lung metastasis) after five years and five years and six months, respectively and one patient died of disease two years after the diagnosis. The primary extraskeletal myxoid chondrosarcoma of bone seems to have a more aggressive behavior than the soft tissue counterpart. The molecular confirmation of diagnosis using RT-PCR is necessary to do the differential diagnosis with other entities, in particular with myoepithelioma that shows similar morphological features and EWSR1 and FUS genes rearrangement. Copyright © 2017 Elsevier GmbH. All rights reserved.

Benefit of Consolidative Radiation Therapy for Primary Bone Diffuse Large B-Cell Lymphoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tao, Randa; Allen, Pamela K.; Rodriguez, Alma

Purpose: Outcomes for patients with diffuse large B-cell lymphoma (DLBCL) differ according to the site of presentation. With effective chemotherapy, the need for consolidative radiation therapy (RT) is controversial. We investigated the influence of primary bone presentation and receipt of consolidative RT on progression-free survival (PFS) and overall survival (OS) in patients with DLBCL. Methods and Materials: We identified 102 patients with primary bone DLBCL treated consecutively from 1988 through 2013 and extracted clinical, pathologic, and treatment characteristics from the medical records. Survival outcomes were calculated by the Kaplan-Meier method, with factors affecting survival determined by log-rank tests. Univariate andmore » multivariate analyses were done with a Cox regression model. Results: The median age was 55 years (range, 16-87 years). The most common site of presentation was in the long bones. Sixty-five patients (63%) received R-CHOP–based chemotherapy, and 74 (72%) received rituximab. RT was given to 67 patients (66%), 47 with stage I to II and 20 with stage III to IV disease. The median RT dose was 44 Gy (range, 24.5-50 Gy). At a median follow-up time of 82 months, the 5-year PFS and OS rates were 80% and 82%, respectively. Receipt of RT was associated with improved 5-year PFS (88% RT vs 63% no RT, P=.0069) and OS (91% vs 68%, P=.0064). On multivariate analysis, the addition of RT significantly improved PFS (hazard ratio [HR] = 0.14, P=.014) with a trend toward an OS benefit (HR=0.30, P=.053). No significant difference in PFS or OS was found between patients treated with 30 to 35 Gy versus ≥36 Gy (P=.71 PFS and P=.31 OS). Conclusion: Patients with primary bone lymphoma treated with standard chemotherapy followed by RT can have excellent outcomes. The use of consolidative RT was associated with significant benefits in both PFS and OS.« less

Tridax procumbens flavonoids: a prospective bioactive compound increased osteoblast differentiation and trabecular bone formation.

PubMed

Al Mamun, Md Abdullah; Hosen, Mohammad Jakir; Khatun, Amina; Alam, M Masihul; Al-Bari, Md Abdul Alim

2017-09-08

The Tridax procumbens extracts (TPE) are known for their ethno-medicinal properties to increase osteogenic functioning in mesenchymal stem cells. Recently, we found that the T. procumbens flavonoids (TPF) significantly suppressed the RANKL-induced osteoclasts differentiation and bone resorption. The TPF also promoted osteoblasts differentiation and bone formation demonstrated by increasing bone formation markers in cultured mouse primary osteoblasts. However, the effects of the TPF on in vivo bone formation remain unclear. In this study, we investigated the effects of the TPF on in vivo bone formation, injected the TPF (20 mg/kg) twice a day in the low calcium diet mice and killed them after 21 day. Radiographic and histomorphometric analyses were performed on the dissected bones to determine the anabolic effects of the TPF. Bone mineral density and bone mineral content of the TPF-treated mice were significantly increased compared to the control mice. Bone formation-related indices like osteoblast number, osteoblast surface, bone volume, mineralizing surface, mineral apposition rate and bone formation rate were significantly increased in the TPF-treated mice compared to the control mice. Our findings point towards the stimulation of bone formation by TPF, suggested that the TPF could be a potential natural anabolic agent to treat patients with bone loss-associated diseases such as osteoporosis.

[Local injection of exogenous nerve growth factor improves early bone maturation of implants].

PubMed

Yao, Yang; Du, Yu; Gu, Xia; Guang, Meng-Kai; Huang, Bo; Gong, Ping

2018-04-01

To investigate the effects of nerve growth factor (NGF) in the osteogenic action of implants and the maturation and reconstruction changes in bone tissues in the early stage of osseointegration. The mouse implant model was established by placing titanium in the femoral head of the mouse and locally injecting NGF in the implant zone. On 1, 2 and 4 weeks after operation, stain samples were collected from animals using hematoxylin-eosin (HE) staining and Masson staining. The effect of NGF on the bone maturation was compared at different time points of early stage osseointegration. The results of HE and Masson staining indicated that the local injection of external NGF can up-regulate bone mass, amount of bone trabecula, and bone maturity in the mouse model. The mature bone rate in treatment group of 1 week and 4 weeks after operation were significantly higher than those in the control group (P<0.05). NGF can shorten the period of bone maturation.

A 4-year treatment with clodronate plus calcium and vitamin D supplements does not improve bone mass in primary biliary cirrhosis.

PubMed

Floreani, A; Carderi, I; Ferrara, F; Rizzotto, E R; Luisetto, G; Camozzi, V; Baldo, V

2007-06-01

International guidelines for managing osteoporosis in cirrhosis or severe cholestasis indicate a <-2.5 t-score as a cut-off for medical treatment, while no treatment is recommended in the case of osteopenia (t-scores ranging from -1.0 to -2.5). We conducted a prospective study in primary biliary cirrhosis with a view to optimizing the rationale for the medical treatment of bone loss. All naïve post-menopausal women with primary biliary cirrhosis were enrolled in the study. Bone metabolism was evaluated by measuring 25-hydroxy-vitamin D, parathyroid hormone, osteocalcin. Bone mineral density was assessed at the lumbar spine by dual-photon X-ray absorptiometry at the baseline and every 2 years for up to 4 years. Patients with either osteopenia or osteoporosis received the following treatment: oral calcium carbonate (1000 mg/day)+vitamin D3 (880 IU/day)+i.m. disodium clodronate 100mg every 10 days for 4 years. Ninety-six patients completed the study: 30 had a normal bone mineral density (group 1), 37 had osteopenia (group 2), 29 had osteoporosis (group 3). No significant differences in biochemical parameters of bone metabolism were observed between the three groups. A total of 288 bone mineral density measurements were taken. Linear regression analysis failed to reveal significant changes in t-score over the follow-up in all groups. A 4-year treatment with clodronate+calcium/vitamin D3 supplements does not significantly improve osteoporosis or osteopenia in primary biliary cirrhosis women in menopause, but prevents the natural bone loss in these patients. Extensive international trials are warranted to optimize the prevention and treatment of bone loss in primary biliary cirrhosis.

Radioprotective Effects of Sulfur-containing Mineral Water of Ramsar Hot Spring with High Natural Background Radiation on Mouse Bone Marrow Cells.

PubMed

Heidari, A H; Shabestani Monfared, A; Mozdarani, H; Mahmoudzadeh, A; Razzaghdoust, A

2017-12-01

We intend to study the inhibitory effect of sulfur compound in Ramsar hot spring mineral on tumor-genesis ability of high natural background radiation. The radioprotective effect of sulfur compounds was previously shown on radiation-induced chromosomal aberration, micronuclei in mouse bone marrow cells and human peripheral lymphocyte. Ramsar is known for having the highest level of natural background radiation on Earth. This study was performed to show the radioprotective effect of sulfur-containing Ramsar mineral water on mouse bone marrow cells. Mice were fed three types of water (drinking water, Ramsar radioactive water containing sulfur and Ramsar radioactive water whose sulfur was removed). Ten days after feeding, mice were irradiated by gamma rays (0, 2 and 4 Gy). 48 and 72 hours after irradiating, mice were killed and femurs were removed. Frequency of micronuclei was determined in bone marrow erythrocytes. A significant reduction was shown in the rate of micronuclei polychromatic erythrocyte in sulfur-containing hot spring water compared to sulfur-free water in hot spring mineral water. Gamma irradiation induced significant increases in micronuclei polychromatic erythrocyte (MNPCE) and decreases in polychromatic erythrocyte/polychromatic erythrocyte + normochromatic erythrocyte ratio (PCEs/PCEs+NCEs) (P < 0.001) in sulfur-containing hot spring water compared to sulfur-free hot spring mineral water. Also, apparently there was a significant difference between drinking water and sulfur-containing hot spring water in micronuclei polychromatic erythrocyte and polychromatic erythrocyte/polychromatic erythrocyte+ normochromatic erythrocyte ratio. The results indicate that sulfur-containing mineral water could result in a significant reduction in radiation-induced micronuclei representing the radioprotective effect of sulfur compounds.

Co-regulation of primary mouse hepatocyte viability and function by oxygen and matrix.

PubMed

Buck, Lorenna D; Inman, S Walker; Rusyn, Ivan; Griffith, Linda G

2014-05-01

Although oxygen and extracellular matrix cues both influence differentiation state and metabolic function of primary rat and human hepatocytes, relatively little is known about how these factors together regulate behaviors of primary mouse hepatocytes in culture. To determine the effects of pericellular oxygen tension on hepatocellular function, we employed two methods of altering oxygen concentration in the local cellular microenvironment of cells cultured in the presence or absence of an extracellular matrix (Matrigel) supplement. By systematically altering medium depth and gas phase oxygen tension, we created multiple oxygen regimes (hypoxic, normoxic, and hyperoxic) and measured the local oxygen concentrations in the pericellular environment using custom-designed oxygen microprobes. From these measurements of oxygen concentrations, we derived values of oxygen consumption rates under a spectrum of environmental contexts, thus providing the first reported estimates of these values for primary mouse hepatocytes. Oxygen tension and matrix microenvironment were found to synergistically regulate hepatocellular survival and function as assessed using quantitative image analysis for cells stained with vital dyes, and assessment of secretion of albumin. Hepatocellular viability was affected only at strongly hypoxic conditions. Surprisingly, albumin secretion rates were greatest at a moderately supra-physiological oxygen concentration, and this effect was mitigated at still greater supra-physiological concentrations. Matrigel enhanced the effects of oxygen on retention of function. This study underscores the importance of carefully controlling cell density, medium depth, and gas phase oxygen, as the effects of these parameters on local pericellular oxygen tension and subsequent hepatocellular function are profound. © 2014 Wiley Periodicals, Inc.

Co-regulation of Primary Mouse Hepatocyte Viability and Function by Oxygen and Matrix

PubMed Central

Buck, Lorenna D.; Inman, S. Walker; Rusyn, Ivan; Griffith, Linda G.

2014-01-01

Although oxygen and extracellular matrix cues both influence differentiation state and metabolic function of primary rat and human hepatocytes, relatively little is known about how these factors together regulate behaviors of primary mouse hepatocytes in culture. To determine the effects of pericellular oxygen tension on hepatocellular function, we employed 2 methods of altering oxygen concentration in the local cellular microenvironment of cells cultured in the presence or absence of an extracellular matrix (Matrigel) supplement. By systematically altering medium depth and gas phase oxygen tension, we created multiple oxygen regimes (hypoxic, normoxic, and hyperoxic) and measured the local oxygen concentrations in the pericellular environment using custom-designed oxygen microprobes. From these measurements of oxygen concentrations, we derived values of oxygen consumption rates under a spectrum of environmental contexts, thus providing the first reported estimates of these values for primary mouse hepatocytes. Oxygen tension and matrix microenvironment were found to synergistically regulate hepatocellular survival and function as assessed using quantitative image analysis for cells stained with vital dyes, and assessment of secretion of albumin. Hepatocellular viability was affected only at strongly hypoxic conditions. Surprisingly, albumin secretion rates were greatest at a moderately supra-physiological oxygen concentration, and this effect was mitigated at still greater supra-physiological concentrations. Matrigel enhanced the effects of oxygen on retention of function. This study underscores the importance of carefully controlling cell density, medium depth and gas phase oxygen, as the effects of these parameters on local pericellular oxygen tension and subsequent hepatocellular function are profound. PMID:24222008

Relationship between bone turnover and left ventricular function in primary hyperparathyroidism: The EPATH trial.

PubMed

Verheyen, Nicolas; Fahrleitner-Pammer, Astrid; Belyavskiy, Evgeny; Gruebler, Martin R; Dimai, Hans Peter; Amrein, Karin; Ablasser, Klemens; Martensen, Johann; Catena, Cristiana; Pieske-Kraigher, Elisabeth; Colantonio, Caterina; Voelkl, Jakob; Lang, Florian; Alesutan, Ioana; Meinitzer, Andreas; März, Winfried; Brussee, Helmut; Pieske, Burkert; Pilz, Stefan; Tomaschitz, Andreas

2017-01-01

Observational studies suggested a link between bone disease and left ventricular (LV) dysfunction that may be pronounced in hyperparathyroid conditions. We therefore aimed to test the hypothesis that circulating markers of bone turnover correlate with LV function in a cohort of patients with primary hyperparathyroidism (pHPT). Cross-sectional data of 155 subjects with pHPT were analyzed who participated in the "Eplerenone in Primary Hyperparathyroidism" (EPATH) Trial. Multivariate linear regression analyses with LV ejection fraction (LVEF, systolic function) or peak early transmitral filling velocity (e', diastolic function) as dependent variables and N-terminal propeptide of procollagen type 1 (P1NP), osteocalcin (OC), bone-specific alkaline phosphatase (BALP), or beta-crosslaps (CTX) as the respective independent variable were performed. Analyses were additionally adjusted for plasma parathyroid hormone, plasma calcium, age, sex, HbA1c, body mass index, mean 24-hours systolic blood pressure, smoking status, estimated glomerular filtration rate, antihypertensive treatment, osteoporosis treatment, 25-hydroxy vitamin D and N-terminal pro-brain B-type natriuretic peptide. Independent relationships were observed between P1NP and LVEF (adjusted β-coefficient = 0.201, P = 0.035) and e' (β = 0.188, P = 0.042), respectively. OC (β = 0.192, P = 0.039) and BALP (β = 0.198, P = 0.030) were each independently related with e'. CTX showed no correlations with LVEF or e'. In conclusion, high bone formation markers were independently and paradoxically related with better LV diastolic and, partly, better systolic function, in the setting of pHPT. Potentially cardio-protective properties of stimulated bone formation in the context of hyperparathyroidism should be explored in future studies.

Transgenic Expression of Osteoactivin/gpnmb Enhances Bone Formation In Vivo and Osteoprogenitor Differentiation Ex Vivo.

PubMed

Frara, Nagat; Abdelmagid, Samir M; Sondag, Gregory R; Moussa, Fouad M; Yingling, Vanessa R; Owen, Thomas A; Popoff, Steven N; Barbe, Mary F; Safadi, Fayez F

2016-01-01

Initial identification of osteoactivin (OA)/glycoprotein non-melanoma clone B (gpnmb) was demonstrated in an osteopetrotic rat model, where OA expression was increased threefold in mutant bones, compared to normal. OA mRNA and protein expression increase during active bone regeneration post-fracture, and primary rat osteoblasts show increased OA expression during differentiation in vitro. To further examine OA/gpnmb as an osteoinductive agent, we characterized the skeletal phenotype of transgenic mouse overexpressing OA/gpnmb under the CMV-promoter (OA-Tg). Western blot analysis showed increased OA/gpnmb in OA-Tg osteoblasts, compared to wild-type (WT). In OA-Tg mouse femurs versus WT littermates, micro-CT analysis showed increased trabecular bone volume and thickness, and cortical bone thickness; histomorphometry showed increased osteoblast numbers, bone formation and mineral apposition rates in OA-Tg mice; and biomechanical testing showed higher peak moment and stiffness. Given that OA/gpnmb is also over-expressed in osteoclasts in OA-Tg mice, we evaluated bone resorption by ELISA and histomorphometry, and observed decreased serum CTX-1 and RANK-L, and decreased osteoclast numbers in OA-Tg, compared to WT mice, indicating decreased bone remodeling in OA-Tg mice. The proliferation rate of OA-Tg osteoblasts in vitro was higher, compared to WT, as was alkaline phosphatase staining and activity, the latter indicating enhanced differentiation of OA-Tg osteoprogenitors. Quantitative RT-PCR analysis showed increased TGF-β1 and TGF-β receptors I and II expression in OA-Tg osteoblasts, compared to WT. Together, these data suggest that OA overexpression has an osteoinductive effect on bone mass in vivo and stimulates osteoprogenitor differentiation ex vivo. © 2015 Wiley Periodicals, Inc.

Hydroxyproline metabolism in mouse models of primary hyperoxaluria

PubMed Central

Holmes, Ross P.; Cramer, Scott D.; Takayama, Tatsuya; Salido, Eduardo

2012-01-01

Primary hyperoxaluria type 1 (PH1) and type 2 (PH2) are rare genetic diseases that result from deficiencies in glyoxylate metabolism. The increased oxalate synthesis that occurs can lead to kidney stone formation, deposition of calcium oxalate in the kidney and other tissues, and renal failure. Hydroxyproline (Hyp) catabolism, which occurs mainly in the liver and kidney, is a prominent source of glyoxylate and could account for a significant portion of the oxalate produced in PH. To determine the sensitivity of mouse models of PH1 and PH2 to Hyp-derived oxalate, animals were fed diets containing 1% Hyp. Urinary excretions of glycolate and oxalate were used to monitor Hyp catabolism and the kidneys were examined to assess pathological changes. Both strains of knockout (KO) mice excreted more oxalate than wild-type (WT) animals with Hyp feeding. After 4 wk of Hyp feeding, all mice deficient in glyoxylate reductase/hydroxypyruvate reductase (GRHPR KO) developed severe nephrocalcinosis in contrast to animals deficient in alanine-glyoxylate aminotransferase (AGXT KO) where nephrocalcinosis was milder and with a lower frequency. Plasma cystatin C measurements over 4-wk Hyp feeding indicated no significant loss of renal function in WT and AGXT KO animals, and significant and severe loss of renal function in GRHPR KO animals after 2 and 4 wk, respectively. These data suggest that GRHPR activity may be vital in the kidney for limiting the conversion of Hyp-derived glyoxylate to oxalate. As Hyp catabolism may make a major contribution to the oxalate produced in PH patients, Hyp feeding in these mouse models should be useful in understanding the mechanisms associated with calcium oxalate deposition in the kidney. PMID:22189945

Bone marrow-derived mesenchymal stem cells protect against lung injury in a mouse model of bronchopulmonary dysplasia.

PubMed

Luan, Yun; Ding, Wei; Ju, Zhi-Ye; Zhang, Zhao-Hua; Zhang, Xue; Kong, Feng

2015-03-01

The aim of the present study was to investigate the effect of bone marrow‑derived mesenchymal stem cells (BMSCs) in the treatment of lung injury in a mouse model of bronchopulmonary dysplasia (BPD) and examine the underlying mechanisms. A mouse model of BPD was created using continuous exposure to high oxygen levels for 14 days. BMSCs were isolated, cultured and then labeled with green fluorescent protein. Cells (1x106) were subsequently injected intravenously 1 h prior to high oxygen treatment. Animals were randomly divided into three groups (n=5 in each): Control group, BPD model group and BMSC injection group. At two weeks post‑treatment, the expression of transforming growth factor‑β1 (TGF‑β1), vascular endothelial growth factor (VEGF) and von Willebrand factor (vWF) was detected using immunohistochemical staining and immunofluorescence. Compared with the BPD model group, the body weight, airway structure and levels of TGF‑β1 and VEGF were significantly improved in the BMSC‑treated group. Immunofluorescence observations indicated that BMSCs were able to differentiate into cells expressing vWF and VEGF, which are markers of vascular tissues. The present study demonstrated that intravenous injection of BMSCs significantly improved lung damage in a neonatal mouse model of BPD at 14 days following hyperoxia‑induced injury. This provides novel information which may be used to guide further investigation into the use of stem cells in BPD.

Women Build Long Bones With Less Cortical Mass Relative to Body Size and Bone Size Compared With Men.

PubMed

Jepsen, Karl J; Bigelow, Erin M R; Schlecht, Stephen H

2015-08-01

The twofold greater lifetime risk of fracturing a bone for white women compared with white men and black women has been attributed in part to differences in how the skeletal system accumulates bone mass during growth. On average, women build more slender long bones with less cortical area compared with men. Although slender bones are known to have a naturally lower cortical area compared with wider bones, it remains unclear whether the relatively lower cortical area of women is consistent with their increased slenderness or is reduced beyond that expected for the sex-specific differences in bone size and body size. Whether this sexual dimorphism is consistent with ethnic background and is recapitulated in the widely used mouse model also remains unclear. We asked (1) do black women build bones with reduced cortical area compared with black men; (2) do white women build bones with reduced cortical area compared with white men; and (3) do female mice build bones with reduced cortical area compared with male mice? Bone strength and cross-sectional morphology of adult human and mouse bone were calculated from quantitative CT images of the femoral midshaft. The data were tested for normality and regression analyses were used to test for differences in cortical area between men and women after adjusting for body size and bone size by general linear model (GLM). Linear regression analysis showed that the femurs of black women had 11% lower cortical area compared with those of black men after adjusting for body size and bone size (women: mean=357.7 mm2; 95% confidence interval [CI], 347.9-367.5 mm2; men: mean=400.1 mm2; 95% CI, 391.5-408.7 mm2; effect size=1.2; p<0.001, GLM). Likewise, the femurs of white women had 12% less cortical area compared with those of white men after adjusting for body size and bone size (women: mean=350.1 mm2; 95% CI, 340.4-359.8 mm2; men: mean=394.3 mm2; 95% CI, 386.5-402.1 mm2; effect size=1.3; p<0.001, GLM). In contrast, female and male femora

The role of bone marrow-derived cells during the bone healing process in the GFP mouse bone marrow transplantation model.

PubMed

Tsujigiwa, Hidetsugu; Hirata, Yasuhisa; Katase, Naoki; Buery, Rosario Rivera; Tamamura, Ryo; Ito, Satoshi; Takagi, Shin; Iida, Seiji; Nagatsuka, Hitoshi

2013-03-01

Bone healing is a complex and multistep process in which the origin of the cells participating in bone repair is still unknown. The involvement of bone marrow-derived cells in tissue repair has been the subject of recent studies. In the present study, bone marrow-derived cells in bone healing were traced using the GFP bone marrow transplantation model. Bone marrow cells from C57BL/6-Tg (CAG-EGFP) were transplanted into C57BL/6 J wild mice. After transplantation, bone injury was created using a 1.0-mm drill. Bone healing was histologically assessed at 3, 7, 14, and 28 postoperative days. Immunohistochemistry for GFP; double-fluorescent immunohistochemistry for GFP-F4/80, GFP-CD34, and GFP-osteocalcin; and double-staining for GFP and tartrate-resistant acid phosphatase were performed. Bone marrow transplantation successfully replaced the hematopoietic cells into GFP-positive donor cells. Immunohistochemical analyses revealed that osteoblasts or osteocytes in the repair stage were GFP-negative, whereas osteoclasts in the repair and remodeling stages and hematopoietic cells were GFP-positive. The results indicated that bone marrow-derived cells might not differentiate into osteoblasts. The role of bone marrow-derived cells might be limited to adjustment of the microenvironment by differentiating into inflammatory cells, osteoclasts, or endothelial cells in immature blood vessels.

Enhanced cloning efficiency of mouse bone marrow macrophage progenitors correlates with increased content of CSF-1 receptor of their progeny at low oxygen tension.

PubMed

Flamant, Stéphane; Lebastard, Maï; Pescher, Pascale; Besmond, Claude; Milon, Geneviève; Marchal, Gilles

2003-10-01

Mononuclear phagocytes are located in every tissue of metazoan organisms. In this extravascular space, they are designated as macrophages and are known to sense and process many signals including the local oxygen tension (PO2), which ranges from 150 mmHg at the lung apices to around 40 mmHg in mixed venous blood and most organs, and to less than 10 mmHg in tissues where long-term and dynamic remodeling processes occur. Most tissue macrophages survive and maintain their differentiated status within an environment bathed by colony-stimulating factor (CSF)-1 through the CSF-1 receptor, encoded by the Csf1r gene. In order to investigate the mRNA expression profile of macrophages as a function of PO2, we developed an in vitro model in which monocyte-derived macrophages were generated from mouse bone marrow progenitor cells grown and maintained under low (36 mmHg) or atmospheric (142 mmHg) PO2, in the presence of L929-conditioned medium (L-CM) as a source of CSF-1. We show that CSF-1-reactive C57BL/6 bone marrow cells displayed an increased cloning efficiency under a PO2 of 36, compared with 142 mmHg. Furthermore, we provide evidence of the overexpression of both CSF-1 receptor protein and mRNA by mouse monocyte-derived macrophages generated from bone marrow under low PO2.

Age dependent regulation of bone-mass and renal function by the MEPE ASARM-motif

PubMed Central

Zelenchuk, Lesya V; Hedge, Anne-Marie; Rowe, Peter S N

2015-01-01

Context Mice with null mutations in Matrix Extracellular Phosphoglycoprotein (MEPE) have increased bone mass, increased trabecular density and abnormal cancellous bone (MN-mice). These defects worsen with age and MEPE over expression induces opposite effects. Also, Genome Wide Association studies show MEPE plays a major role in bone mass. We hypothesized the conserved C-terminal MEPE ASARM-motif is chiefly responsible for regulating bone mass and trabecular structure. Design To test our theory we over expressed C-terminal ASARM-peptide in MN-mice using the Col1α1 promoter (MNAt-mice). We then compared the bone and renal phenotypes of the MNAt-mouse with the MN-mouse and the X-linked hypophosphatemic rickets mouse (HYP). The HYP mouse over expresses ASARM-peptides and is defective for the PHEX gene. Results The MN-mouse developed increased bone mass, bone strength and trabecular abnormalities that worsened markedly with age. Defects in bone formation were chiefly responsible with suppressed sclerostin and increased active β-catenin. Increased uric acid levels also suggested abnormalities in purine-metabolism and a reduced fractional excretion of uric acid signaled additional renal transport changes. The MN mouse developed a worsening hyperphosphatemia and reduced FGF23 with age. An increase in the fractional excretion of phosphate (FEP) despite the hyperphosphatemia confirms an imbalance in kidney-intestinal phosphate regulation. Also, the MN mice showed an increased creatinine clearance suggesting hyperfiltration. A reversal of the MN bone-renal phenotype changes occurred with the MNAt mice including the apparent hyperfiltration. The MNAt mice also developed localized hypomineralization, hypophosphatemia and increased FGF23. Conclusions The C-terminal ASARM-motif plays a major role in regulating bone–mass and cancellous structure as mice age. In healthy mice, the processing and release of free ASARM-peptide is chiefly responsible for preserving normal bone and

Congenital Bone Fractures in Spinal Muscular Atrophy: Functional Role for SMN Protein in Bone Remodeling

PubMed Central

Shanmugarajan, Srinivasan; Swoboda, Kathryn J.; Iannaccone, Susan T.; Ries, William L.; Maria, Bernard L.; Reddy, Sakamuri V.

2009-01-01

Spinal muscular atrophy is the second most common fatal childhood disorder. Core clinical features include muscle weakness caused by degenerating lower motor neurons and a high incidence of bone fractures and hypercalcemia. Fractures further compromise quality of life by progression of joint contractures or additional loss of motor function. Recent observations suggest that bone disease in spinal muscular atrophy may not be attributed entirely to lower motor neuron degeneration. The presence of the spinal muscular atrophy disease-determining survival motor neuron gene (SMN), SMN expression, and differential splicing in bone-resorbing osteoclasts was recently discovered. Its ubiquitous expression and the differential expression of splice variants suggest that SMN has specific roles in bone cell function. SMN protein also interacts with osteoclast stimulatory factor. Mouse models of human spinal muscular atrophy disease suggest a potential role of SMN protein in skeletal development. Dual energy x-ray absorptiometry analysis demonstrated a substantial decrease in total bone area and poorly developed caudal vertebra in the mouse model. These mice also had pelvic bone fractures. Studies delineating SMN signaling mechanisms and gene transcription in a cell-specific manner will provide important molecular insights into the pathogenesis of bone disease in children with spinal muscular atrophy. Moreover, understanding bone remodeling in spinal muscular atrophy may lead to novel therapeutic approaches to enhance skeletal health and quality of life. This article reviews the skeletal complications associated with spinal muscular atrophy and describes a functional role for SMN protein in osteoclast development and bone resorption activity. PMID:17761651

Long-Term Follow-Up of Primary Medical Versus Surgical Treatment of Prolactinomas in Men: Effects on Hyperprolactinemia, Hypogonadism, and Bone Health.

PubMed

Andereggen, Lukas; Frey, Janine; Andres, Robert H; El-Koussy, Marwan; Beck, Jürgen; Seiler, Rolf W; Christ, Emanuel

2017-01-01

In men with prolactinomas, impaired bone density is the principle consequence of hyperprolactinemia-induced hypogonadism. Although dopamine agonists (DAs) are the first-line approach in prolactinomas, surgery can be considered in selected cases. In this study, we aimed to investigate the long-term control of hyperprolactinemia, hypogonadism, and bone health comparing primary medical and surgical therapy in men who had not had prior DA treatment. This is a retrospective case-note study of 44 consecutive men with prolactinomas and no prior DAs managed in a single tertiary referral center. Clinical, biochemical, and radiologic response to the first-line approach were analyzed in the 2 cohorts. Mean age at diagnosis was 47 years (range, 22-78 years). The prevalence of hypogonadism was 86%, and 27% of patients had pathologic bone density at baseline. The primary therapeutic strategy was surgery for 34% and DAs for 66% of patients. Median long-term follow-up was 63 months (range, 17-238 months). Long-term control of hyperprolactinemia required DAs in 53% of patients with primary surgical therapy, versus 90% of patients with primary medical therapy (P = 0.02). Hypogonadism was controlled in 73% of patients. The prevalence of patients with pathologic bone density was 37% at last follow-up, with no differences between the 2 therapeutic cohorts (P = 0.48). Despite control of hyperprolactinemia and hypogonadism in most patients independent of the primary treatment modality, the prevalence of impaired bone health status remains high, and osteodensitometry should be recommended. Copyright © 2016 Elsevier Inc. All rights reserved.

Transgenic Mouse Model for Reducing Oxidative Damage in Bone

NASA Technical Reports Server (NTRS)

Schreurs, A.-S.; Torres, S.; Truong, T.; Kumar, A.; Alwood, J. S.; Limoli, C. L.; Globus, R. K.

2014-01-01

Exposure to musculoskeletal disuse and radiation result in bone loss; we hypothesized that these catabolic treatments cause excess reactive oxygen species (ROS), and thereby alter the tight balance between bone resorption by osteoclasts and bone formation by osteoblasts, culminating in bone loss. To test this, we used transgenic mice which over-express the human gene for catalase, targeted to mitochondria (MCAT). Catalase is an anti-oxidant that converts the ROS hydrogen peroxide into water and oxygen. MCAT mice were shown previously to display reduced mitochondrial oxidative stress and radiosensitivity of the CNS compared to wild type controls (WT). As expected, MCAT mice expressed the transgene in skeletal tissue, and in marrow-derived osteoblasts and osteoclast precursors cultured ex vivo, and also showed greater catalase activity compared to wildtype (WT) mice (3-6 fold). Colony expansion in marrow cells cultured under osteoblastogenic conditions was 2-fold greater in the MCAT mice compared to WT mice, while the extent of mineralization was unaffected. MCAT mice had slightly longer tibiae than WT mice (2%, P less than 0.01), although cortical bone area was slightly lower in MCAT mice than WT mice (10%, p=0.09). To challenge the skeletal system, mice were treated by exposure to combined disuse (2 wk Hindlimb Unloading) and total body irradiation Cs(137) (2 Gy, 0.8 Gy/min), then bone parameters were analyzed by 2-factor ANOVA to detect possible interaction effects. Treatment caused a 2-fold increase (p=0.015) in malondialdehyde levels of bone tissue (ELISA) in WT mice, but had no effect in MCAT mice. These findings indicate that the transgene conferred protection from oxidative damage caused by treatment. Unexpected differences between WT and MCAT mice emerged in skeletal responses to treatment.. In WT mice, treatment did not alter osteoblastogenesis, cortical bone area, moment of inertia, or bone perimeter, whereas in MCAT mice, treatment increased these

A mouse model with postnatal endolymphatic hydrops and hearing loss

PubMed Central

Megerian, Cliff A.; Semaan, Maroun T.; Aftab, Saba; Kisley, Lauren B.; Zheng, Qing Yin; Pawlowski, Karen S.; Wright, Charles G.; Alagramam, Kumar N.

2010-01-01

Endolymphatic hydrops (ELH), hearing loss and neuronal degeneration occur together in a variety of clinically significant disorders, including Meniere’s disease (MD). However, the sequence of these pathological changes and their relationship to each other are not well understood. In this regard, an animal model that spontaneously develops these features postnatally would be useful for research purposes. A search for such a model led us to the PhexHyp-Duk mouse, a mutant allele of the Phex gene causing X-linked hypophosphatemic rickets. The hemizygous male (PhexHyp-Duk/Y) was previously reported to exhibit various abnormalities during adulthood, including thickening of bone, ELH and hearing loss. The reported inner-ear phenotype was suggestive of progressive pathology and spontaneous development of ELH postnatally, but not conclusive. The main focuses of this report are to further characterize the inner ear phenotype in PhexHyp-Duk/Y mice and to test the hypotheses that (a) the PhexHyp-Duk/Y mouse develops ELH and hearing loss postnatally, and (b) the development of ELH in the PhexHyp-Duk/Y mouse is associated with obstruction of the endolymphatic duct (ED) due to thickening of the surrounding bone. Auditory brainstem response (ABR) recordings at various times points and histological analysis of representative temporal bones reveal that PhexHyp-Duk/Y mice typically develop adult onset, asymmetric, progressive hearing loss closely followed by the onset of ELH. ABR and histological data show that functional degeneration precedes structural degeneration. The major degenerative correlate of hearing loss and ELH in the mutants is the primary loss of spiral ganglion cells. Further, PhexHyp-Duk/Y mice develop ELH without evidence of ED obstruction, supporting the idea that ELH can be induced by a mechanism other than the blockade of longitudinal flow of endolymphatic fluid, and occlusion of ED is not a prerequisite for the development of ELH in patients. PMID:18289812

Differential Bone Loss in Mouse Models of Colon Cancer Cachexia.

PubMed

Bonetto, Andrea; Kays, Joshua K; Parker, Valorie A; Matthews, Ryan R; Barreto, Rafael; Puppa, Melissa J; Kang, Kyung S; Carson, James A; Guise, Theresa A; Mohammad, Khalid S; Robling, Alexander G; Couch, Marion E; Koniaris, Leonidas G; Zimmers, Teresa A

2016-01-01

Cachexia is a distinctive feature of colorectal cancer associated with body weight loss and progressive muscle wasting. Several mechanisms responsible for muscle and fat wasting have been identified, however it is not known whether the physiologic and molecular crosstalk between muscle and bone tissue may also contribute to the cachectic phenotype in cancer patients. The purpose of this study was to clarify whether tumor growth associates with bone loss using several experimental models of colorectal cancer cachexia, namely C26, HT-29, and Apc Min/+ . The effects of cachexia on bone structure and strength were evaluated with dual energy X-ray absorptiometry (DXA), micro computed tomography (μCT), and three-point bending test. We found that all models showed tumor growth consistent with severe cachexia. While muscle wasting in C26 hosts was accompanied by moderate bone depletion, no loss of bone strength was observed. However, HT-29 tumor bearing mice showed bone abnormalities including significant reductions in whole-body bone mineral density (BMD), bone mineral content (BMC), femoral trabecular bone volume fraction (BV/TV), trabecular number (Tb.N), and trabecular thickness (Tb.Th), but no declines in strength. Similarly, cachexia in the Apc Min/+ mice was associated with significant decreases in BMD, BMC, BV/TV, Tb.N, and Tb.Th as well as decreased strength. Our data suggest that colorectal cancer is associated with muscle wasting and may be accompanied by bone loss dependent upon tumor type, burden, stage and duration of the disease. It is clear that preserving muscle mass promotes survival in cancer cachexia. Future studies will determine whether strategies aimed at preventing bone loss can also improve outcomes and survival in colorectal cancer cachexia.

Formononetin, a methoxy isoflavone, enhances bone regeneration in a mouse model of cortical bone defect.

PubMed

Singh, Krishna Bhan; Dixit, Manisha; Dev, Kapil; Maurya, Rakesh; Singh, Divya

2017-06-01

The bone regeneration and healing effect of formononetin was evaluated in a cortical bone defect model that predominantly heals by intramembranous ossification. For this study, female Balb/c mice were ovariectomised (OVx) and a drill-hole injury was generated in the midfemoral bones of all animals. Treatment with formononetin commenced the day after and continued for 21 d. Parathyroid hormone (PTH1-34) was used as a reference standard. Animals were killed at days 10 and 21. Femur bones were collected at the injury site for histomorphometry studies using microcomputed tomography (μCT) and confocal microscopy. RNA and protein were harvested from the region surrounding the drill-hole injury. For immunohistochemistry, 5 µm sections of decalcified femur bone adjoining the drill-hole site were cut. μCT analysis showed that formononetin promoted bone healing at days 10 and 21 and the healing effect observed was significantly better than in Ovx mice and equal to PTH treatment in many aspects. Formononetin also significantly enhanced bone regeneration as assessed by calcein-labelling studies. In addition, formononetin enhanced the expression of osteogenic markers at the injury site in a manner similar to PTH. Formononetin treatment also led to predominant runt-related transcription factor 2 and osteocalcin localisation at the injury site. These results support the potential of formononetin to be a bone-healing agent and are suggestive of its promising role in the fracture-repair process.

The role of the SIBLING, Bone Sialoprotein in skeletal biology - Contribution of mouse experimental genetics.

PubMed

Bouleftour, Wafa; Juignet, Laura; Bouet, Guenaelle; Granito, Renata Neves; Vanden-Bossche, Arnaud; Laroche, Norbert; Aubin, Jane E; Lafage-Proust, Marie-Hélène; Vico, Laurence; Malaval, Luc

2016-01-01

Bone Sialoprotein (BSP) is a member of the "Small Integrin-Binding Ligand N-linked Glycoproteins" (SIBLING) extracellular matrix protein family of mineralized tissues. BSP has been less studied than other SIBLING proteins such as Osteopontin (OPN), which is coexpressed with it in several skeletal cell types. Here we review the contribution of genetically engineered mice (BSP gene knockout and overexpression) to the understanding of the role of BSP in the bone organ. The studies made so far highlight the role of BSP in skeletal mineralization, as well as its importance for proper osteoblast and osteoclast differentiation and activity, most prominently in primary/repair bone. The absence of BSP also affects the local environment of the bone tissue, in particular hematopoiesis and vascularization. Interestingly, lack of BSP induces an overexpression of OPN, and the cognate protein could be responsible for some aspects of the BSP gene knockout skeletal phenotype, while replacing BSP for some of its functions. Such interplay between the partly overlapping functions of SIBLING proteins, as well as the network of cross-regulations in which they are involved should now be the focus of further work. Copyright © 2016 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

Miscellaneous indications in bone scintigraphy: metabolic bone diseases and malignant bone tumors.

PubMed

Cook, Gary J R; Gnanasegaran, Gopinath; Chua, Sue

2010-01-01

The diphosphonate bone scan is ideally suited to assess many global, focal or multifocal metabolic bone disorders and there remains a role for conventional bone scintigraphy in metabolic bone disorders at diagnosis, investigation of complications, and treatment response assessment. In contrast, the role of bone scintigraphy in the evaluation of primary malignant bone tumors has reduced with the improvement of morphologic imaging, such as computed tomography and magnetic resonance imaging. However, an increasing role for (18)F-fluorodeoxyglucose positron emission tomography and positron emission tomography/computed tomography is emerging as a functional assessment at diagnosis, staging, and neoadjuvant treatment response assessment.

Mineralization defects in cementum and craniofacial bone from loss of bone sialoprotein

PubMed Central

Foster, B.L.; Ao, M.; Willoughby, C.; Soenjaya, Y.; Holm, E.; Lukashova, L.; Tran, A. B.; Wimer, H.F.; Zerfas, P.M.; Nociti, F.H.; Kantovitz, K.R.; Quan, B.D.; Sone, E.D.; Goldberg, H.A.; Somerman, M.J.

2015-01-01

Bone sialoprotein (BSP) is a multifunctional extracellular matrix protein found in mineralized tissues, including bone, cartilage, tooth root cementum (both acellular and cellular types), and dentin. In order to define the role BSP plays in the process of biomineralization of these tissues, we analyzed cementogenesis, dentinogenesis, and osteogenesis (intramembranous and endochondral) in craniofacial bone in Bsp null mice and wild-type (WT) controls over a developmental period (1-60 days post natal; dpn) by histology, immunohistochemistry, undecalcified histochemistry, microcomputed tomography (microCT), scanning electron microscopy (SEM), transmission electron microscopy (TEM), and quantitative PCR (qPCR). Regions of intramembranous ossification in the alveolus, mandible, and calvaria presented delayed mineralization and osteoid accumulation, assessed by von Kossa and Goldner's trichrome stains at 1 and 14 dpn. Moreover, Bsp−/− mice featured increased cranial suture size at the early time point, 1 dpn. Immunostaining and PCR demonstrated that osteoblast markers, osterix, alkaline phosphatase, and osteopontin were unchanged in Bsp null mandibles compared to WT. Bsp−/− mouse molars featured a lack of functional acellular cementum formation by histology, SEM, and TEM, and subsequent loss of Sharpey's collagen fiber insertion into the tooth root structure. Bsp−/− mouse alveolar and mandibular bone featured equivalent or fewer osteoclasts at early ages (1 and 14 dpn), however, increased RANKL immunostaining and mRNA, and significantly increased number of osteoclast-like cells (2-5 fold) were found at later ages (26 and 60 dpn), corresponding to periodontal breakdown and severe alveolar bone resorption observed following molar teeth entering occlusion. Dentin formation was unperturbed in Bsp−/− mouse molars, with no delay in mineralization, no alteration in dentin dimensions, and no differences in odontoblast markers analyzed. No defects were identified

Differential Bone Loss in Mouse Models of Colon Cancer Cachexia

PubMed Central

Bonetto, Andrea; Kays, Joshua K.; Parker, Valorie A.; Matthews, Ryan R.; Barreto, Rafael; Puppa, Melissa J.; Kang, Kyung S.; Carson, James A.; Guise, Theresa A.; Mohammad, Khalid S.; Robling, Alexander G.; Couch, Marion E.; Koniaris, Leonidas G.; Zimmers, Teresa A.

2017-01-01

Cachexia is a distinctive feature of colorectal cancer associated with body weight loss and progressive muscle wasting. Several mechanisms responsible for muscle and fat wasting have been identified, however it is not known whether the physiologic and molecular crosstalk between muscle and bone tissue may also contribute to the cachectic phenotype in cancer patients. The purpose of this study was to clarify whether tumor growth associates with bone loss using several experimental models of colorectal cancer cachexia, namely C26, HT-29, and ApcMin/+. The effects of cachexia on bone structure and strength were evaluated with dual energy X-ray absorptiometry (DXA), micro computed tomography (μCT), and three-point bending test. We found that all models showed tumor growth consistent with severe cachexia. While muscle wasting in C26 hosts was accompanied by moderate bone depletion, no loss of bone strength was observed. However, HT-29 tumor bearing mice showed bone abnormalities including significant reductions in whole-body bone mineral density (BMD), bone mineral content (BMC), femoral trabecular bone volume fraction (BV/TV), trabecular number (Tb.N), and trabecular thickness (Tb.Th), but no declines in strength. Similarly, cachexia in the ApcMin/+ mice was associated with significant decreases in BMD, BMC, BV/TV, Tb.N, and Tb.Th as well as decreased strength. Our data suggest that colorectal cancer is associated with muscle wasting and may be accompanied by bone loss dependent upon tumor type, burden, stage and duration of the disease. It is clear that preserving muscle mass promotes survival in cancer cachexia. Future studies will determine whether strategies aimed at preventing bone loss can also improve outcomes and survival in colorectal cancer cachexia. PMID:28123369

Changes in Mouse Bone Turnover in Response to Microgravity

NASA Technical Reports Server (NTRS)

Cheng-Campbell, M.; Blaber, E.; Almeida, E.

2016-01-01

Mechanical unloading during spaceflight is known to adversely affect mammalian physiology. Our previous studies using the Animal Enclosure Module on short duration Shuttle missions enabled us to identify a deficit in stem cell based-tissue regeneration as being a significant concern for long-duration spaceflight. Specifically, we found that mechanical unloading in microgravity resulted in inhibition of differentiation of mesenchymal and hematopoietic stem cells in the bone marrow compartment. Also, we observed overexpression of a cell cycle arrest molecule, CDKN1a/p21, in osteoprecursor cells on the bone surface, chondroprogenitors in the articular cartilage, and in myofibers attached to bone tissue. Specifically in bone tissue during both short (15-day) and long (30-day) microgravity experiments, we observed significant loss of bone tissue and structure in both the pelvis and the femur. After 15-days of microgravity on STS-131, pelvic ischium displayed a 6.23% decrease in bone fraction (p=0.005) and 11.91% decrease in bone thickness (p=0.002). Furthermore, during long-duration spaceflight we observed onset of an accelerated aging-like phenotype and osteoarthritic disease state indicating that stem cells within the bone tissue fail to repair and regenerate tissues in a normal manner, leading to drastic tissue alterations in response to microgravity. The Rodent Research Hardware System provides the capability to investigate these effects during long-duration experiments on the International Space Station. During the Rodent Research-1 mission 10 16-week-old female C57Bl/6J mice were exposed to 37-days of microgravity. All flight animals were euthanized and frozen on orbit for future dissection. Ground (n=10) and vivarium controls (n=10) were housed and processed to match the flight animal timeline. During this study we collected pelvis, femur, and tibia from all animal groups to test the hypothesis that stem cell-based tissue regeneration is significantly altered

Age-dependent Changes in the Articular Cartilage and Subchondral Bone of C57BL/6 Mice after Surgical Destabilization of Medial Meniscus.

PubMed

Huang, Henry; Skelly, Jordan D; Ayers, David C; Song, Jie

2017-02-09

Age is the primary risk factor for osteoarthritis (OA), yet surgical OA mouse models such as destabilization of the medial meniscus (DMM) used for evaluating disease-modifying OA targets are frequently performed on young adult mice only. This study investigates how age affects cartilage and subchondral bone changes in mouse joints following DMM. DMM was performed on male C57BL/6 mice at 4 months (4 M), 12 months (12 M) and 19+ months (19 M+) and on females at 12 M and 18 M+. Two months after surgery, operated and unoperated contralateral knees were harvested and evaluated using cartilage histology scores and μCT quantification of subchondral bone plate thickness and osteophyte formation. The 12 M and 19 M+ male mice developed more cartilage erosions and thicker subchondral bone plates after DMM than 4 M males. The size of osteophytes trended up with age, while the bone volume fraction was significantly higher in the 19 M+ group. Furthermore, 12 M females developed milder OA than males as indicated by less cartilage degradation, less subchondral bone plate sclerosis and smaller osteophytes. Our results reveal distinct age/gender-dependent structural changes in joint cartilage and subchondral bone post-DMM, facilitating more thoughtful selection of murine age/gender when using this surgical technique for translational OA research.

Age-dependent Changes in the Articular Cartilage and Subchondral Bone of C57BL/6 Mice after Surgical Destabilization of Medial Meniscus

PubMed Central

Huang, Henry; Skelly, Jordan D.; Ayers, David C.; Song, Jie

2017-01-01

Age is the primary risk factor for osteoarthritis (OA), yet surgical OA mouse models such as destabilization of the medial meniscus (DMM) used for evaluating disease-modifying OA targets are frequently performed on young adult mice only. This study investigates how age affects cartilage and subchondral bone changes in mouse joints following DMM. DMM was performed on male C57BL/6 mice at 4 months (4 M), 12 months (12 M) and 19+ months (19 M+) and on females at 12 M and 18 M+. Two months after surgery, operated and unoperated contralateral knees were harvested and evaluated using cartilage histology scores and μCT quantification of subchondral bone plate thickness and osteophyte formation. The 12 M and 19 M+ male mice developed more cartilage erosions and thicker subchondral bone plates after DMM than 4 M males. The size of osteophytes trended up with age, while the bone volume fraction was significantly higher in the 19 M+ group. Furthermore, 12 M females developed milder OA than males as indicated by less cartilage degradation, less subchondral bone plate sclerosis and smaller osteophytes. Our results reveal distinct age/gender-dependent structural changes in joint cartilage and subchondral bone post-DMM, facilitating more thoughtful selection of murine age/gender when using this surgical technique for translational OA research. PMID:28181577

Absence of ERRα in Female Mice Confers Resistance to Bone Loss Induced by Age or Estrogen-Deficiency

PubMed Central

Rabier, Bénédicte; Monfoulet, Laurent; Dine, Julien; Macari, Claire; Espallergues, Julie; Horard, Béatrice; Giguère, Vincent; Cohen-Solal, Martine; Chassande, Olivier; Vanacker, Jean-Marc

2009-01-01

Background ERRα is an orphan member of the nuclear hormone receptor superfamily, which acts as a transcription factor and is involved in various metabolic processes. ERRα is also highly expressed in ossification zones during mouse development as well as in human bones and cell lines. Previous data have shown that this receptor up-modulates the expression of osteopontin, which acts as an inhibitor of bone mineralization and whose absence results in resistance to ovariectomy-induced bone loss. Altogether this suggests that ERRα may negatively regulate bone mass and could impact on bone fragility that occurs in the absence of estrogens. Methods/Principal Findings In this report, we have determined the in vivo effect of ERRα on bone, using knock-out mice. Relative to wild type animals, female ERRαKO bones do not age and are resistant to bone loss induced by estrogen-withdrawal. Strikingly male ERRαKO mice are indistinguishable from their wild type counterparts, both at the unchallenged or gonadectomized state. Using primary cell cultures originating from ERRαKO bone marrow, we also show that ERRα acts as an inhibitor of osteoblast differentiation. Conclusion/Significance Down-regulating ERRα could thus be beneficial against osteoporosis. PMID:19936213

The STR/ort mouse model of spontaneous osteoarthritis - an update.

PubMed

Staines, K A; Poulet, B; Wentworth, D N; Pitsillides, A A

2017-06-01

Osteoarthritis is a degenerative joint disease and a world-wide healthcare burden. Characterized by cartilage degradation, subchondral bone thickening and osteophyte formation, osteoarthritis inflicts much pain and suffering, for which there are currently no disease-modifying treatments available. Mouse models of osteoarthritis are proving critical in advancing our understanding of the underpinning molecular mechanisms. The STR/ort mouse is a well-recognized model which develops a natural form of osteoarthritis very similar to the human disease. In this Review we discuss the use of the STR/ort mouse in understanding this multifactorial disease with an emphasis on recent advances in its genetics and its bone, endochondral and immune phenotypes. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Sex Steroid Actions in Male Bone

PubMed Central

Laurent, Michaël R.; Claessens, Frank; Gielen, Evelien; Lagerquist, Marie K.; Vandenput, Liesbeth; Börjesson, Anna E.; Ohlsson, Claes

2014-01-01

Sex steroids are chief regulators of gender differences in the skeleton, and male gender is one of the strongest protective factors against osteoporotic fractures. This advantage in bone strength relies mainly on greater cortical bone expansion during pubertal peak bone mass acquisition and superior skeletal maintenance during aging. During both these phases, estrogens acting via estrogen receptor-α in osteoblast lineage cells are crucial for male cortical and trabecular bone, as evident from conditional genetic mouse models, epidemiological studies, rare genetic conditions, genome-wide meta-analyses, and recent interventional trials. Genetic mouse models have also demonstrated a direct role for androgens independent of aromatization on trabecular bone via the androgen receptor in osteoblasts and osteocytes, although the target cell for their key effects on periosteal bone formation remains elusive. Low serum estradiol predicts incident fractures, but the highest risk occurs in men with additionally low T and high SHBG. Still, the possible clinical utility of serum sex steroids for fracture prediction is unknown. It is likely that sex steroid actions on male bone metabolism rely also on extraskeletal mechanisms and cross talk with other signaling pathways. We propose that estrogens influence fracture risk in aging men via direct effects on bone, whereas androgens exert an additional antifracture effect mainly via extraskeletal parameters such as muscle mass and propensity to fall. Given the demographic trends of increased longevity and consequent rise of osteoporosis, an increased understanding of how sex steroids influence male bone health remains a high research priority. PMID:25202834

Bone Marrow Transplantation Results in Human Donor Blood Cells Acquiring and Displaying Mouse Recipient Class I MHC and CD45 Antigens on Their Surface

PubMed Central

Yamanaka, Nobuko; Wong, Christine J.; Gertsenstein, Marina; Casper, Robert F.; Nagy, Andras; Rogers, Ian M.

2009-01-01

Background Mouse models of human disease are invaluable for determining the differentiation ability and functional capacity of stem cells. The best example is bone marrow transplants for studies of hematopoietic stem cells. For organ studies, the interpretation of the data can be difficult as transdifferentiation, cell fusion or surface antigen transfer (trogocytosis) can be misinterpreted as differentiation. These events have not been investigated in hematopoietic stem cell transplant models. Methodology/Principal Findings In this study we investigated fusion and trogocytosis involving blood cells during bone marrow transplantation using a xenograft model. We report that using a standard SCID repopulating assay almost 100% of the human donor cells appear as hybrid blood cells containing both mouse and human surface antigens. Conclusion/Significance Hybrid cells are not the result of cell-cell fusion events but appear to be due to efficient surface antigen transfer, a process referred to as trogocytosis. Antigen transfer appears to be non-random and includes all donor cells regardless of sub-type. We also demonstrate that irradiation preconditioning enhances the frequency of hybrid cells and that trogocytosis is evident in non-blood cells in chimera mice. PMID:20046883

Dissecting modes of action of non-genotoxic carcinogens in primary mouse hepatocytes.

PubMed

Schaap, Mirjam M; Zwart, Edwin P; Wackers, Paul F K; Huijskens, Ilse; van de Water, Bob; Breit, Timo M; van Steeg, Harry; Jonker, Martijs J; Luijten, Mirjam

2012-11-01

Under REACH, the European Community Regulation on chemicals, the testing strategy for carcinogenicity is based on in vitro and in vivo genotoxicity assays. Given that non-genotoxic carcinogens are negative for genotoxicity and chronic bioassays are no longer regularly performed, this class of carcinogens will go undetected. Therefore, test systems detecting non-genotoxic carcinogens, or even better their modes of action, are required. Here, we investigated whether gene expression profiling in primary hepatocytes can be used to distinguish different modes of action of non-genotoxic carcinogens. For this, primary mouse hepatocytes were exposed to 16 non-genotoxic carcinogens with diverse modes of action. Upon profiling, pathway analysis was performed to obtain insight into the biological relevance of the observed changes in gene expression. Subsequently, both a supervised and an unsupervised comparison approach were applied to recognize the modes of action at the transcriptomic level. These analyses resulted in the detection of three of eight compound classes, that is, peroxisome proliferators, metalloids and skin tumor promotors. In conclusion, gene expression profiles in primary hepatocytes, at least in rodent hepatocytes, appear to be useful to detect some, certainly not all, modes of action of non-genotoxic carcinogens.

Bone Marrow Failure Secondary to Cytokinesis Failure

DTIC Science & Technology

2015-12-01

SUPPLEMENTARY NOTES 14. ABSTRACT Fanconi anemia (FA) is a human genetic disease characterized by a progressive bone marrow failure and heightened...Fanconi anemia (FA) is the most commonly inherited bone marrow failure syndrome. FA patients develop bone marrow failure during the first decade of...experiments proposed in specific aims 1- 3 (Tasks 1-3). Task 1: To determine whether HSCs from Fanconi anemia mouse models have increased cytokinesis

Cancer-associated bone disease.

PubMed

Rizzoli, R; Body, J-J; Brandi, M-L; Cannata-Andia, J; Chappard, D; El Maghraoui, A; Glüer, C C; Kendler, D; Napoli, N; Papaioannou, A; Pierroz, D D; Rahme, M; Van Poznak, C H; de Villiers, T J; El Hajj Fuleihan, G

2013-12-01

Bone is commonly affected in cancer. Cancer-induced bone disease results from the primary disease, or from therapies against the primary condition, causing bone fragility. Bone-modifying agents, such as bisphosphonates and denosumab, are efficacious in preventing and delaying cancer-related bone disease. With evidence-based care pathways, guidelines assist physicians in clinical decision-making. Of the 57 million deaths in 2008 worldwide, almost two thirds were due to non-communicable diseases, led by cardiovascular diseases and cancers. Bone is a commonly affected organ in cancer, and although the incidence of metastatic bone disease is not well defined, it is estimated that around half of patients who die from cancer in the USA each year have bone involvement. Furthermore, cancer-induced bone disease can result from the primary disease itself, either due to circulating bone resorbing substances or metastatic bone disease, such as commonly occurs with breast, lung and prostate cancer, or from therapies administered to treat the primary condition thus causing bone loss and fractures. Treatment-induced osteoporosis may occur in the setting of glucocorticoid therapy or oestrogen deprivation therapy, chemotherapy-induced ovarian failure and androgen deprivation therapy. Tumour skeletal-related events include pathologic fractures, spinal cord compression, surgery and radiotherapy to bone and may or may not include hypercalcaemia of malignancy while skeletal complication refers to pain and other symptoms. Some evidence demonstrates the efficacy of various interventions including bone-modifying agents, such as bisphosphonates and denosumab, in preventing or delaying cancer-related bone disease. The latter includes treatment of patients with metastatic skeletal lesions in general, adjuvant treatment of breast and prostate cancer in particular, and the prevention of cancer-associated bone disease. This has led to the development of guidelines by several societies and

Cancer-associated bone disease

PubMed Central

Body, J.-J.; Brandi, M.-L.; Cannata-Andia, J.; Chappard, D.; El Maghraoui, A.; Glüer, C.C.; Kendler, D.; Napoli, N.; Papaioannou, A.; Pierroz, D.D.; Rahme, M.; Van Poznak, C.H.; de Villiers, T.J.; El Hajj Fuleihan, G.

2016-01-01

Bone is commonly affected in cancer. Cancer-induced bone disease results from the primary disease, or from therapies against the primary condition, causing bone fragility. Bone-modifying agents, such as bisphosphonates and denosumab, are efficacious in preventing and delaying cancer-related bone disease. With evidence-based care pathways, guidelines assist physicians in clinical decision-making. Of the 57 million deaths in 2008 worldwide, almost two thirds were due to non-communicable diseases, led by cardiovascular diseases and cancers. Bone is a commonly affected organ in cancer, and although the incidence of metastatic bone disease is not well defined, it is estimated that around half of patients who die from cancer in the USA each year have bone involvement. Furthermore, cancer-induced bone disease can result from the primary disease itself, either due to circulating bone resorbing substances or metastatic bone disease, such as commonly occurs with breast, lung and prostate cancer, or from therapies administered to treat the primary condition thus causing bone loss and fractures. Treatment-induced osteoporosis may occur in the setting of glucocorticoid therapy or oestrogen deprivation therapy, chemotherapy-induced ovarian failure and androgen deprivation therapy. Tumour skeletal-related events include pathologic fractures, spinal cord compression, surgery and radiotherapy to bone and may or may not include hypercalcaemia of malignancy while skeletal complication refers to pain and other symptoms. Some evidence demonstrates the efficacy of various interventions including bone-modifying agents, such as bisphosphonates and denosumab, in preventing or delaying cancer-related bone disease. The latter includes treatment of patients with metastatic skeletal lesions in general, adjuvant treatment of breast and prostate cancer in particular, and the prevention of cancer-associated bone disease. This has led to the development of guidelines by several societies and

Epigallocatechin gallate (EGCG) suppresses lipopolysaccharide-induced inflammatory bone resorption, and protects against alveolar bone loss in mice.

PubMed

Tominari, Tsukasa; Matsumoto, Chiho; Watanabe, Kenta; Hirata, Michiko; Grundler, Florian M W; Miyaura, Chisato; Inada, Masaki

2015-01-01

Epigallocatechin gallate (EGCG), a major polyphenol in green tea, possesses antioxidant properties and regulates various cell functions. Here, we examined the function of EGCG in inflammatory bone resorption. In calvarial organ cultures, lipopolysaccharide (LPS)-induced bone resorption was clearly suppressed by EGCG. In osteoblasts, EGCG suppressed the LPS-induced expression of COX-2 and mPGES-1 mRNAs, as well as prostaglandin E2 production, and also suppressed RANKL expression, which is essential for osteoclast differentiation. LPS-induced bone resorption of mandibular alveolar bones was attenuated by EGCG in vitro, and the loss of mouse alveolar bone mass was inhibited by the catechin in vivo.

Mouse Bone Marrow VSELs Exhibit Differentiation into Three Embryonic Germ Lineages and Germ & Hematopoietic Cells in Culture.

PubMed

Shaikh, Ambreen; Anand, Sandhya; Kapoor, Sona; Ganguly, Ranita; Bhartiya, Deepa

2017-04-01

Very small embryonic-like stem cells (VSELs) have been reported in various adult tissues, express pluripotent and primordial germ cells (PGCs) specific markers, are mobilized under stress/disease conditions, give rise to tissue committed progenitors and thus help regenerate and maintain homeostasis. The aim of the present study was to evaluate in vitro differentiation potential of VSELs using a quantitative approach. VSELs were collected from mouse bone marrow after 4 days of 5-fluorouracil (5-FU, 150 mg/Kg) treatment, further enriched by size based filtration and cultured on a feeder support in the presence of specific differentiation media. Cultured VSELs were found to differentiate into all three embryonic germ cell lineages, germ and hematopoietic cells after 14 days in culture. This was confirmed by studying Nestin, PDX-1, NKX2.5, DAZL, CD45 and other markers expression by various approaches. Very small, CD45 negative cells collected and enriched from GFP positive 5-FU treated mice bone marrow transitioned into CD45 positive cells in vitro thus demonstrating that VSELs can give rise to hematopoietic stem cells (HSCs). We envision that VSELs may be responsible for plasticity and ability of bone marrow cells to give rise to non-hematopoietic tissue progenitors of all 3 germ layers. Moreover the ability of VSELs to differentiate into germ cells as well as all the three lineages provides further evidence to support their pluripotent state and confirms developmental link between bone marrow VSELs and PGCs. The property of quiescence, no risk of teratoma formation and autologus source, make pluripotent VSELs a potential candidate to facilitate endogenous regeneration compared to cell replacement strategy envisioned using embryonic and induced pluripotent stem cells.

Noncanonical Wnt5a-Ca(2+) -NFAT signaling axis in pesticide induced bone marrow aplasia mouse model: A study to explore the novel mechanism of pesticide toxicity.

PubMed

Chattopadhyay, Sukalpa; Chatterjee, Ritam; Law, Sujata

2016-10-01

According to case-control studies, long-term pesticide exposure can cause bone marrow aplasia like hematopoietic degenerative disease leading to impaired hematopoiesis and increased risk of aplastic anemia in human subjects. However, the exact mechanism of pesticide mediated hematotoxicity still remains elusive. In this study, we investigated the role of noncanonical Wnt signaling pathway, a crucial regulator of adult hematopoiesis, in pesticide induced bone marrow aplasia mouse model. Aplasia mouse model was developed following inhalation and dermal exposure of 5% aqueous mixture of common agriculturally used pesticides for 6 h/day for 5 days a week up to 90 days. After that, blood hemogram, marrow smear, cellularity, scanning electron microscopy, extramedullary hematopoiesis and flowcytometric expression analysis of noncanonical Wnt signaling components, such as Wnt 5a, fzd5, NFAT, IFN-γ, intracellular Ca(2+) level were evaluated in the bone marrow hematopoietic stem/progenitor compartment of the control and pesticide induced aplasia groups of animals. Results showed that pesticide exposed mice were anemic with peripheral blood pancytopenia, hypocellular degenerative marrow, and extramedullary hematopoiesis in the spleen. Upon pesticide exposure, Wnt 5a expression was severely downregulated with a decline in intracellular Ca(2+) level. Moreover, downstream of Wnt5a, we observed sharp downregulation of NFATc2 transcription factor expression, the major target of pesticide toxicity and its target molecule IFN-γ. Taken together, our result suggests that deregulation of Wnt5a-Ca(2+) -NFAT signaling axis in the hematopoietic stem/progenitor compartment plays a crucial role behind the pathogenesis of pesticide mediated bone marrow aplasia by limiting primitive hematopoietic stem cells' ability to maintain hematopoietic homeostasis and reconstitution mechanism in vivo during xenobiotic stress leading to ineffective hematopoiesis and evolution of bone marrow aplasia. �

Transgenic Mouse Model for Reducing Oxidative Damage in Bone

NASA Technical Reports Server (NTRS)

Schreurs, Ann-Sofie; Torres, S.; Truong, T.; Moyer, E. L.; Kumar, A.; Tahimic, Candice C. G.; Alwood, J. S.; Limoli, C. L.; Globus, R. K.

2016-01-01

Bone loss can occur due to many challenges such age, radiation, microgravity, and Reactive Oxygen Species (ROS) play a critical role in bone resorption by osteoclasts (Bartell et al. 2014). We hypothesize that suppression of excess ROS in skeletal cells, both osteoblasts and osteoclasts, regulates skeletal growth and remodeling. To test our hypothesis, we used transgenic mCAT mice which overexpress the human anti-oxidant catalase gene targeted to the mitochondria, the main site for endogenous ROS production. mCAT mice have a longer life-span than wildtype controls and have been used to study various age-related disorders. To stimulate remodeling, 16 week old mCAT mice or wildtype mice were exposed to treatment (hindlimb-unloading and total body-irradiation) or sham treatment conditions (control). Tissues were harvested 2 weeks later for skeletal analysis (microcomputed tomography), biochemical analysis (gene expression and oxidative damage measurements), and ex vivo bone marrow derived cell culture (osteoblastogenesis and osteoclastogenesis). mCAT mice expressed the transgene and displayed elevated catalase activity in skeletal tissue and marrow-derived osteoblasts and osteoclasts grown ex vivo. In addition, when challenged with treatment, bone tissues from wildtype mice showed elevated levels of malondialdehyde (MDA), indicating oxidative damage) whereas mCAT mice did not. Correlation analysis revealed that increased catalase activity significantly correlated with decreased MDA levels and that increased oxidative damage correlated with decreased percent bone volume (BVTV). In addition, ex-vivo cultured osteoblast colony growth correlated with catalase activity in the osteoblasts. Thus, we showed that these transgenic mice can be used as a model to study the relationship between markers of oxidative damage and skeletal properties. mCAT mice displayed reduced BVTV and trabecular number relative to wildtype mice, as well as increased structural model index in the

The effects of imidacloprid combined with endosulfan on IgE-mediated mouse bone marrow-derived mast cell degranulation and anaphylaxis.

PubMed

Shi, Lin-Bo; Xu, Hua-Ping; Wu, Yu-Jie; Li, Xin; Gao, Jin-Yan; Chen, Hong-Bing

2018-06-01

Low levels of endosulfan are known to stimulate mast cells to release allergic mediators, while imidacloprid can inhibit IgE-mediated mast cell degranulation. However, little information about the effects of both pesticides together on mast cell degranulation is available. To measure the effects, IgE-activated mouse bone marrow-derived mast cells (BMMCs) were treated with imidacloprid and endosulfan, individually, and simultaneously at equi-molar concentrations in tenfold steps ranging from 10 -4 to 10 -11  M, followed by measuring several allergy-related parameters expressed in BMMCs: the mediator production and influx of Ca 2+ , the phosphorylation content of NF-κB in the FcεRI signaling pathway. Then, the effects of the mixtures on IgE-induced passive systemic anaphylaxis (PSA) of BALB/c was detectded. This study clearly showed that the application of equi-molar mixtures of both pesticides with 10 -4 -10 -5  M significantly inhibited the IgE-mediated mouse bone marrow-derived mast cells degranulation in vitro and 10 -4  M of them decreased IgE-mediated PSA in vivo, as the application of imidacloprid at the same concentration alone did. Morever endosulfan alone had no remarkable stimulatory effects on any of the factors measured. In conclusion, simultaneous application of equi-molar concentrations of both pesticides generally showed highly similar responses compared to the responses to imidacloprid alone, suggesting that the effects of the mixture could be solely attributed to the effects of imidacloprid. Copyright © 2018 Elsevier Inc. All rights reserved.

The potential of materials analysis by electron rutherford backscattering as illustrated by a case study of mouse bones and related compounds.

PubMed

Vos, Maarten; Tökési, Károly; Benkö, Ilona

2013-06-01

Electron Rutherford backscattering (ERBS) is a new technique that could be developed into a tool for materials analysis. Here we try to establish a methodology for the use of ERBS for materials analysis of more complex samples using bone minerals as a test case. For this purpose, we also studied several reference samples containing Ca: calcium carbonate (CaCO(3)) and hydroxyapatite and mouse bone powder. A very good understanding of the spectra of CaCO(3) and hydroxyapatite was obtained. Quantitative interpretation of the bone spectrum is more challenging. A good fit of these spectra is only obtained with the same peak widths as used for the hydroxyapatite sample, if one allows for the presence of impurity atoms with a mass close to that of Na and Mg. Our conclusion is that a meaningful interpretation of spectra of more complex samples in terms of composition is indeed possible, but only if widths of the peaks contributing to the spectra are known. Knowledge of the peak widths can either be developed by the study of reference samples (as was done here) or potentially be derived from theory.

Anemia in patient with primary hyperoxaluria and bone marrow involvement by oxalate crystals.

PubMed

Mykytiv, Vitaliy; Campoy Garcia, Fiz

2018-06-01

We present a rare case of anaemia secondary to bone marrow infiltration by oxalate crystals and renal failure in a patient diagnosed with primary hyperoxaluria. In our case, the anaemia was recovered after the double liver and kidney transplantation, the latter was performed on two occasions after the failure of the first graft. Copyright © 2017 King Faisal Specialist Hospital & Research Centre. Published by Elsevier Ltd. All rights reserved.

Postradiation atrophy of mature bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ergun, H.; Howland, W.J.

1980-01-01

The primary event of radiation damage to bone is atrophy and true necrosis of bone is uncommon. The postradiation atrophic changes of bone are the result of combined cellular and vascular damage, the former being more important. The damage to the osteoblast resulting in decreased matrix production is apparently the primary histopathologic event. Radiation damaged bone is susceptible to superimposed complications of fracture, infection, necrosis, and sarcoma. The primary radiographic evidence of atrophy, localized osteopenia, is late in appearing. Contrary to former views, the mature bone is quite radiosensitive and reacts quickly to even small doses of radiation. The differentiationmore » of postirradiation atrophy and metastasis may be difficult. Biopsy should be the last resort because of the possibility of causing true necrosis in atrophic bone by trauma and infection.« less

A Progressive Translational Mouse Model of Human VCP Disease: The VCP R155H/+ Mouse

PubMed Central

Nalbandian, Angèle; Llewellyn, Katrina J.; Badadani, Mallikarjun; Yin, Hong Z.; Nguyen, Christopher; Katheria, Veeral; Watts, Giles; Mukherjee, Jogeshwar; Vesa, Jouni; Caiozzo, Vincent; Mozaffar, Tahseen; Weiss, John H.; Kimonis, Virginia E.

2012-01-01

Introduction Mutations in the valosin containing protein (VCP) gene cause hereditary Inclusion Body Myopathy (hIBM) associated with Paget disease of bone (PDB), and frontotemporal dementia (FTD). More recently they have been linked to 2% of familial ALS cases. A knock-in mouse model offers the opportunity to study VCP-associated pathogenesis. Methods The VCPR155H/+ knock-in mouse model was assessed for muscle strength, immunohistochemical, Western, apoptosis, autophagy and MicroPET/CT imaging analyses. Results VCPR155H/+ mice developed significant progressive muscle weakness, and the quadriceps and brain developed progressive cytoplasmic accumulation of TDP-43, ubiquitin-positive inclusion bodies and increased LC3-II staining. MicroCT analyses revealed Paget-like lesions at the ends of long bones. Spinal cord demonstrated neurodegenerative changes, ubiquitin, and TDP-43 pathology of motor neurons. Discussion VCPR155H/+ knock-in mice represent an excellent pre-clinical model for understanding VCP-associated disease mechanisms and future treatments. PMID:23169451

Comparative analysis of TCDD-induced AhR-mediated gene expression in human, mouse and rat primary B cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kovalova, Natalia, E-mail: kovalova@msu.edu

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental pollutant that activates the aryl hydrocarbon receptor (AhR) resulting in altered gene expression. In vivo, in vitro, and ex vivo studies have demonstrated that B cells are directly impaired by TCDD, and are a sensitive target as evidenced by suppression of antibody responses. The window of sensitivity to TCDD-induced suppression of IgM secretion among mouse, rat and human B cells is similar. Specifically, TCDD must be present within the initial 12 h post B cell stimulation, indicating that TCDD disrupts early signaling network(s) necessary for B lymphocyte activation and differentiation. Therefore, we hypothesized thatmore » TCDD treatment across three different species (mouse, rat and human) triggers a conserved, B cell-specific mechanism that is involved in TCDD-induced immunosuppression. RNA sequencing (RNA-Seq) was used to identify B cell-specific orthologous genes that are differentially expressed in response to TCDD in primary mouse, rat and human B cells. Time course studies identified TCDD-elicited differential expression of 515 human, 2371 mouse and 712 rat orthologous genes over the 24-h period. 28 orthologs were differentially expressed in response to TCDD in all three species. Overrepresented pathways enriched in all three species included cytokine-cytokine receptor interaction, ECM-receptor interaction, focal adhesion, regulation of actin cytoskeleton and pathways in cancer. Differentially expressed genes functionally associated with cell-cell signaling in humans, immune response in mice, and oxidation reduction in rats. Overall, these results suggest that despite the conservation of the AhR and its signaling mechanism, TCDD elicits species-specific gene expression changes. - Highlights: • Kovalova TAAP Highlights Nov. 2016 • RNA-Seq identified TCDD-induced gene expression in PWM-activated primary B cells. • TCDD elicited differential expression of 515 human, 2371 mouse

ANTIBODY FORMATION BY TRANSPLANTED BONE MARROW, SPLEEN, LYMPH NODE AND THYMUS CELLS IN IRRADIATED RECIPIENTS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoner, R.D.; Bond, V.P.

1963-01-14

Immunological competence of immunized mouse bone marrow, spleen, lymph node, and thymus cells was demonstrated when specific recall tetanus antitoxin responses were elicited after transfer of these cells to isologous irradiated mice or rats. Lesser amounts of antibody were obtained as the genetic strain distance was increased between the relation of donor and host in the parental to F/sub 1/ and in the homologous combination within the same species. It was not possible in the heterologous situation to elicit significant amounts of antibody from rat bone marrow and other lymphoid cells following their transplantation into irradiated mice. Minimal but notmore » significant antibody responses were elicited from cells obtained from immunized rat spleen and thymus tissue. In a few experiments, it was possible to elicit antibody formation from a buffy coat suspension of circulating white cells following their transfer to irradiated recipients. Isologous nonimmunized bone marrow did not stimulate or hasten recovery of the ability to eiicit secondary antibody responses in previously immunized irradiated mice. The capacity to elicit primary antibody responses to tetanus toxoid was depressed in parental-bone-marrow-protected F/sub 1/ mice when these chimeras exhibited varying degrees of secondary disease. The depression of primary antibody responses in irradiated F/sub 1/ mice given parental bone marrow provides evidence for a donor mediated immunological depression of antibody synthesis by host-lymphoid tissues. (auth)« less

Bayesian pretest probability estimation for primary malignant bone tumors based on the Surveillance, Epidemiology and End Results Program (SEER) database.

PubMed

Benndorf, Matthias; Neubauer, Jakob; Langer, Mathias; Kotter, Elmar

2017-03-01

In the diagnostic process of primary bone tumors, patient age, tumor localization and to a lesser extent sex affect the differential diagnosis. We therefore aim to develop a pretest probability calculator for primary malignant bone tumors based on population data taking these variables into account. We access the SEER (Surveillance, Epidemiology and End Results Program of the National Cancer Institute, 2015 release) database and analyze data of all primary malignant bone tumors diagnosed between 1973 and 2012. We record age at diagnosis, tumor localization according to the International Classification of Diseases (ICD-O-3) and sex. We take relative probability of the single tumor entity as a surrogate parameter for unadjusted pretest probability. We build a probabilistic (naïve Bayes) classifier to calculate pretest probabilities adjusted for age, tumor localization and sex. We analyze data from 12,931 patients (647 chondroblastic osteosarcomas, 3659 chondrosarcomas, 1080 chordomas, 185 dedifferentiated chondrosarcomas, 2006 Ewing's sarcomas, 281 fibroblastic osteosarcomas, 129 fibrosarcomas, 291 fibrous malignant histiocytomas, 289 malignant giant cell tumors, 238 myxoid chondrosarcomas, 3730 osteosarcomas, 252 parosteal osteosarcomas, 144 telangiectatic osteosarcomas). We make our probability calculator accessible at http://ebm-radiology.com/bayesbone/index.html . We provide exhaustive tables for age and localization data. Results from tenfold cross-validation show that in 79.8 % of cases the pretest probability is correctly raised. Our approach employs population data to calculate relative pretest probabilities for primary malignant bone tumors. The calculator is not diagnostic in nature. However, resulting probabilities might serve as an initial evaluation of probabilities of tumors on the differential diagnosis list.

A primary phosphorus-deficient skeletal phenotype in juvenile Atlantic salmon Salmo salar: the uncoupling of bone formation and mineralization.

PubMed

Witten, P E; Owen, M A G; Fontanillas, R; Soenens, M; McGurk, C; Obach, A

2016-02-01

To understand the effect of low dietary phosphorus (P) intake on the vertebral column of Atlantic salmon Salmo salar, a primary P deficiency was induced in post-smolts. The dietary P provision was reduced by 50% for a period of 10 weeks under controlled conditions. The animal's skeleton was subsequently analysed by radiology, histological examination, histochemical detection of minerals in bones and scales and chemical mineral analysis. This is the first account of how a primary P deficiency affects the skeleton in S. salar at the cellular and at the micro-anatomical level. Animals that received the P-deficient diet displayed known signs of P deficiency including reduced growth and soft, pliable opercula. Bone and scale mineral content decreased by c. 50%. On radiographs, vertebral bodies appear small, undersized and with enlarged intervertebral spaces. Contrary to the X-ray-based diagnosis, the histological examination revealed that vertebral bodies had a regular size and regular internal bone structures; intervertebral spaces were not enlarged. Bone matrix formation was continuous and uninterrupted, albeit without traces of mineralization. Likewise, scale growth continues with regular annuli formation, but new scale matrix remains without minerals. The 10 week long experiment generated a homogeneous osteomalacia of vertebral bodies without apparent induction of skeletal malformations. The experiment shows that bone formation and bone mineralization are, to a large degree, independent processes in the fish examined. Therefore, a deficit in mineralization must not be the only cause of the alterations of the vertebral bone structure observed in farmed S. salar. It is discussed how the observed uncoupling of bone formation and mineralization helps to better diagnose, understand and prevent P deficiency-related malformations in farmed S. salar. © 2015 The Authors.Journal of Fish Biology published by John Wiley & Sons Ltd on behalf of The Fisheries Society of the

Ex-vivo transduced autologous skin fibroblasts expressing human Lim Mineralization Protein-3 efficiently form new bone in animal models

PubMed Central

Lattanzi, Wanda; Parrilla, Claudio; Fetoni, Annarita; Logroscino, Giandomenico; Straface, Giuseppe; Pecorini, Giovanni; Stigliano, Egidio; Tampieri, Anna; Bedini, Rossella; Pecci, Raffaella; Michetti, Fabrizio; Gambotto, Andrea; Robbins, Paul D.; Pola, Enrico

2012-01-01

Local gene transfer of the human LIM Mineralization Protein (LMP), a novel intracellular positive regulator of the osteoblast differentiation program, can induce efficient bone formation in rodents. In order to develop a clinically relevant gene therapy approach to facilitate bone healing, we have used primary dermal fibroblasts transduced ex vivo with Ad.LMP3 and seeded on an hydroxyapatite/collagen matrix prior to autologous implantation. Here we demonstrate that genetically modified autologous dermal fibroblasts expressing Ad.LMP-3 are able to induce ectopic bone formation following implantation of the matrix into the mouse triceps and paravertebral muscles. Moreover, implantation of the Ad.LMP-3-modified dermal fibroblasts into a rat mandibular bone critical size defect model results in efficient healing as determined by X-ray, histology and three dimensional micro computed tomography (3DμCT). These results demonstrate the effectiveness of the non-secreted intracellular osteogenic factor LMP-3, in inducing bone formation in vivo. Moreover, the utilization of autologous dermal fibroblasts implanted on a biomaterial represents a promising approach for possible future clinical applications aimed at inducing new bone formation. PMID:18633445

The development of inter-strain variation in cortical and trabecular traits during growth of the mouse lumbar vertebral body.

PubMed

Ramcharan, M A; Faillace, M E; Guengerich, Z; Williams, V A; Jepsen, K J

2017-03-01

How cortical and trabecular bone co-develop to establish a mechanically functional structure is not well understood. Comparing early postnatal differences in morphology of lumbar vertebral bodies for three inbred mouse strains identified coordinated changes within and between cortical and trabecular traits. These early coordinate changes defined the phenotypic differences among the inbred mouse strains. Age-related changes in cortical and trabecular traits have been well studied; however, very little is known about how these bone tissues co-develop from day 1 of postnatal growth to establish functional structures by adulthood. In this study, we aimed to establish how cortical and trabecular tissues within the lumbar vertebral body change during growth for three inbred mouse strains that express wide variation in adult bone structure and function. Bone traits were quantified for lumbar vertebral bodies of female A/J, C57BL/6J (B6), and C3H/HeJ (C3H) inbred mouse strains from 1 to 105 days of age (n = 6-10 mice/age/strain). Inter-strain differences in external bone size were observed as early as 1 day of age. Reciprocal and rapid changes in the trabecular bone volume fraction and alignment in the direction of axial compression were observed by 7 days of age. Importantly, the inter-strain difference in adult trabecular bone volume fraction was established by 7 days of age. Early variation in external bone size and trabecular architecture was followed by progressive increases in cortical area between 28 and 105 days of age, with the greatest increases in cortical area seen in the mouse strain with the lowest trabecular mass. Establishing the temporal changes in bone morphology for three inbred mouse strains revealed that genetic variation in adult trabecular traits were established early in postnatal development. Early variation in trabecular architecture preceded strain-specific increases in cortical area and changes in cortical thickness. This study

Prostate Cancer Metastases Alter Bone Mineral and Matrix Composition Independent of Effects on Bone Architecture in Mice A Quantitative Study Using microCT and Raman Spectroscopy

PubMed Central

Bi, Xiaohong; Sterling, Julie A.; Merkel, Alyssa R.; Perrien, Daniel S.; Nyman, Jeffry; Mahadevan-Jansen, Anita

2013-01-01

Prostate cancer is the most common primary tumor and the second leading cause of cancer-related deaths in men in the United States. Prostate cancer bone metastases are characterized by abnormal bone remodeling processes and result in a variety of skeletal morbidities. Prevention of skeletal complications is a crucial element in prostate cancer management. This study investigated prostate cancer-induced alterations in the molecular composition and morphological structure of metastasis-bearing bones in a mouse model of prostate cancer using Raman spectroscopy and micro-computed tomography (microCT). LNCaP C4-2B prostate cancer cells were injected into the right tibiae of 5-week old male SCID mice. Upon sacrifice at 8 weeks post tumor inoculation, two out of the ten tumor-bearing tibiae showed only osteoblastic lesions in the radiographs, 4 osteolytic lesions only and 4 mixed with osteoblastic and osteolytic lesions.. Carbonate substitution was significantly increased while there was a marked reduction in the level of collagen mineralization, mineral crystallinity, and carbonate:matrix ratio in the cortex of the intact tumor-bearing tibiae compared to contralateral controls. MicroCT analysis revealed a significant reduction in bone volume/total volume, trabecular number and trabecular thickness, as well as significant increase in bone surface/volume ratio in tibiae with osteolytic lesions, suggesting active bone remodeling and bone loss. None of the changes in bone compositional properties were correlated with lesion area from radiographs or the changes in bone architecture from microCT. This study indicates that LNCaP C4-2B prostate cancer metastases alter bone tissue composition independent of changes in architecture, and altered bone quality may be an important contributor to fracture risk in these patients. Raman spectroscopy may provide a new avenue of investigation into interactions between tumor and bone microenvironment. PMID:23867219

ANGPTL2 increases bone metastasis of breast cancer cells through enhancing CXCR4 signaling

PubMed Central

Masuda, Tetsuro; Endo, Motoyoshi; Yamamoto, Yutaka; Odagiri, Haruki; Kadomatsu, Tsuyoshi; Nakamura, Takayuki; Tanoue, Hironori; Ito, Hitoshi; Yugami, Masaki; Miyata, Keishi; Morinaga, Jun; Horiguchi, Haruki; Motokawa, Ikuyo; Terada, Kazutoyo; Morioka, Masaki Suimye; Manabe, Ichiro; Iwase, Hirotaka; Mizuta, Hiroshi; Oike, Yuichi

2015-01-01

Bone metastasis of breast cancer cells is a major concern, as it causes increased morbidity and mortality in patients. Bone tissue-derived CXCL12 preferentially recruits breast cancer cells expressing CXCR4 to bone metastatic sites. Thus, understanding how CXCR4 expression is regulated in breast cancer cells could suggest approaches to decrease bone metastasis of breast tumor cells. Here, we show that tumor cell-derived angiopoietin-like protein 2 (ANGPTL2) increases responsiveness of breast cancer cells to CXCL12 by promoting up-regulation of CXCR4 in those cells. In addition, we used a xenograft mouse model established by intracardiac injection of tumor cells to show that ANGPTL2 knockdown in breast cancer cells attenuates tumor cell responsiveness to CXCL12 by decreasing CXCR4 expression in those cells, thereby decreasing bone metastasis. Finally, we found that ANGPTL2 and CXCR4 expression levels within primary tumor tissues from breast cancer patients are positively correlated. We conclude that tumor cell-derived ANGPTL2 may increase bone metastasis by enhancing breast tumor cell responsiveness to CXCL12 signaling through up-regulation of tumor cell CXCR4 expression. These findings may suggest novel therapeutic approaches to treat metastatic breast cancer. PMID:25773070

Effect on bone density of postoperative calcium and vitamin-D supplementation in patients with primary hyperparathyroidism: A retrospective study.

PubMed

Nordenström, Erik; Westerdahl, Johan; Bergenfelz, Anders

2009-05-01

Primary hyperparathyroidism (pHPT) is associated with decreased bone density and increased fracture risk. A significant number of pHPT patients have low calcium intake and suffer from vitamin deficiency. Thus, we adopted a policy of postoperative supplements with calcium and vitamin D after parathyroid surgery. In this study, we investigated if this policy enhanced the postoperative increase in bone density. Forty-two consecutive patients (83% female) were studied. The first 21 patients received no supplements, whereas the following 21 patients received 1,000 g calcium and 800 IU hydroxy D: -vitamin daily (Ca-D group) for 1 year postoperatively. The patients were monitored with bone density and biochemistry pre- and at 1 year postoperatively. Preoperatively, the patients without vitamin D supplementation (non-Ca-D group) did neither differ in biochemistry, clinical features, nor in bone density from patients in Ca-D group. Postoperatively, there was a tendency that patients in Ca-D group increased their bone density, at all sites measured, in a greater extent than patients that did not receive calcium and vitamin D supplementation. In conclusion, based on our results, it is difficult to give a recommendation of vitamin D supplementation in routine use following surgery for primary hyperparathyroidism. Based on the present data, a calculation of sample size for a future randomized controlled trial is presented.

Kinetochore identification in micronuclei in mouse bone-marrow erythrocytes: an assay for the detection of aneuploidy-inducing agents.

PubMed

Gudi, R; Sandhu, S S; Athwal, R S

1990-10-01

An in vivo micronucleus assay using mouse bone marrow for identifying the ability of chemicals to induce aneuploidy and/or chromosome breaks is described. Micronucleus formation in bone-marrow erythrocytes of mice is commonly used as an index for evaluating the clastogenicity of environmental agents. However, micronuclei may also originate from intact lagging chromosomes resulting from the effect of aneuploidy-inducing agents. We have used immunofluorescent staining using anti-kinetochore antibodies to classify micronuclei for the presence or absence of kinetochores. Micronuclei positive for kinetochores are assumed to contain intact chromosomes and result from induced aneuploidy; while those negative for kinetochores contain acentric chromosomal fragments and originate from clastogenic events. The assay was evaluated using X-irradiation (a known clastogen) and vincristine sulfate (an aneuploidy-inducing agent). A dose-related response for the induction of micronuclei was observed for both agents. Micronuclei induced by X-irradiation were negative for kinetochores while the majority of the micronuclei resulting from vincristine treatment contained kinetochores. Thus, the micronucleus assay in combination with immunofluorescent staining for kinetochores may provide a useful method to simultaneously assess the ability of chemicals to induce aneuploidy and/or chromosome breaks.

Localization of sarcomeric proteins during myofibril assembly in cultured mouse primary skeletal myotubes

PubMed Central

White, Jennifer; Barro, Marietta V.; Makarenkova, Helen P.; Sanger, Joseph W.; Sanger, Jean M.

2014-01-01

It is important to understand how muscle forms normally in order to understand muscle diseases that result in abnormal muscle formation. Although the structure of myofibrils is well understood, the process through which the myofibril components form organized contractile units is not clear. Based on the staining of muscle proteins in avian embryonic cardiomyocytes, we previously proposed that myofibrils formation occurred in steps that began with premyofibrils followed by nascent myofibrils and ending with mature myofibrils. The purpose of this study was to determine whether the premyofibril model of myofibrillogenesis developed from studies developed from studies in avian cardiomyocytes was supported by our current studies of myofibril assembly in mouse skeletal muscle. Emphasis was on establishing how the key sarcomeric proteins, F-actin, non-muscle myosin II, muscle myosin II, and α-actinin were organized in the three stages of myofibril assembly. The results also test previous reports that non-muscle myosins II A and B are components of the Z-Bands of mature myofibrils, data that are inconsistent with the premyofibril model. We have also determined that in mouse muscle cells, telethonin is a late assembling protein that is present only in the Z-Bands of mature myofibrils. This result of using specific telethonin antibodies supports the approach of using YFP-tagged proteins to determine where and when these YFP-sarcomeric fusion proteins are localized. The data presented in this study on cultures of primary mouse skeletal myocytes are consistent with the premyofibril model of myofibrillogenesis previously proposed for both avian cardiac and skeletal muscle cells. PMID:25125171

Loss of bone sialoprotein leads to impaired endochondral bone development and mineralization.

PubMed

Holm, Erik; Aubin, Jane E; Hunter, Graeme K; Beier, Frank; Goldberg, Harvey A

2015-02-01

Bone sialoprotein (BSP) is an anionic phosphoprotein in the extracellular matrix of mineralized tissues, and a promoter of biomineralization and osteoblast development. Previous studies on the Bsp-deficient mouse (Bsp(-/-)) have demonstrated a significant bone and periodontal tissue phenotype in adulthood. However, the role of BSP during early long bone development is not known. To address this, early endochondral ossification in the Bsp(-/-) mouse was studied. Embryonic day 15.5 (E15.5) wild-type (WT) tibiae showed early stages of ossification that were absent in Bsp(-/-) mice. At E16.5, mineralization had commenced in the Bsp(-/-) mice, but staining for mineral was less intense and more dispersed compared with that in WT controls. Tibiae from Bsp(-/-) mice also demonstrated decreased mineralization and shortened length at postnatal day 0.5 (P0.5) compared to WT bones. There was no detectable difference in the number of tartrate-resistant acid phosphatase-positive foci at P0.5, although the P0.5 Bsp(-/-) tibiae had decreased Vegfα expression compared with WT tissue. Due to the shortened tibiae the growth plates were examined and determined to be of normal overall length. However, the length of the resting zone was increased in P0.5 Bsp(-/-) tibiae whereas that of the proliferative zone was decreased, with no change in the hypertrophic zone length of Bsp(-/-) mice. A reduction in cells positive for Ki-67, an S-phase cell-cycle marker, was noted in the proliferative zone. Decreased numbers of TUNEL-positive hypertrophic chondrocytes were also apparent in the Bsp(-/-) tibial growth plates, suggesting decreased apoptosis. Expression of the osteogenic markers Alp1, Col1a1, Sp7, Runx2, and Bglap was reduced in the endochondral bone of the neonatal Bsp(-/-) compared to WT tibiae. These results suggest that BSP is an important and multifaceted protein that regulates both chondrocyte proliferation and apoptosis as well as transition from cartilage to bone during

Novel Wnt Regulator NEL-Like Molecule-1 Antagonizes Adipogenesis and Augments Osteogenesis Induced by Bone Morphogenetic Protein 2

PubMed Central

Shen, Jia; James, Aaron W.; Zhang, Xinli; Pang, Shen; Zara, Janette N.; Asatrian, Greg; Chiang, Michael; Lee, Min; Khadarian, Kevork; Nguyen, Alan; Lee, Kevin S.; Siu, Ronald K.; Tetradis, Sotirios; Ting, Kang; Soo, Chia

2017-01-01

The differentiation factor NEL-like molecule-1 (NELL-1) has been reported as osteoinductive in multiple in vivo preclinical models. Bone morphogenetic protein (BMP)-2 is used clinically for skeletal repair, but in vivo administration can induce abnormal, adipose-filled, poor-quality bone. We demonstrate that NELL-1 combined with BMP2 significantly optimizes osteogenesis in a rodent femoral segmental defect model by minimizing the formation of BMP2-induced adipose-filled cystlike bone. In vitro studies using the mouse bone marrow stromal cell line M2-10B4 and human primary bone marrow stromal cells have confirmed that NELL-1 enhances BMP2-induced osteogenesis and inhibits BMP2-induced adipogenesis. Importantly, the ability of NELL-1 to direct BMP2-treated cells toward osteogenesis and away from adipogenesis requires intact canonical Wnt signaling. Overall, these studies establish the feasibility of combining NELL-1 with BMP2 to improve clinical bone regeneration and provide mechanistic insight into canonical Wnt pathway activity during NELL-1 and BMP2 osteogenesis. The novel abilities of NELL-1 to stimulate Wnt signaling and to repress adipogenesis may highlight new treatment approaches for bone loss in osteoporosis. PMID:26772960

Basement membrane of mouse bone marrow sinusoids shows distinctive structure and proteoglycan composition: a high resolution ultrastructural study.

PubMed

Inoue, S; Osmond, D G

2001-11-01

Venous sinusoids in bone marrow are the site of a large-scale traffic of cells between the extravascular hemopoietic compartment and the blood stream. The wall of the sinusoids consists solely of a basement membrane interposed between a layer of endothelial cells and an incomplete covering of adventitial cells. To examine its possible structural specialization, the basement membrane of bone marrow sinusoids has now been examined by high resolution electron microscopy of perfusion-fixed mouse bone marrow. The basement membrane layer was discontinuous, consisting of irregular masses of amorphous material within a uniform 60-nm-wide space between apposing endothelial cells and adventitial cell processes. At maximal magnifications, the material was resolved as a random arrangement of components lacking the "cord network" formation seen in basement membranes elsewhere. Individual components exhibited distinctive ultrastructural features whose molecular identity has previously been established. By these morphological criteria, the basement membrane contained unusually abundant chondroitin sulfate proteoglycan (CSPG) revealed by 3-nm-wide "double tracks," and moderate amounts of both laminin as dense irregular coils and type IV collagen as 1-1.5-nm-wide filaments, together with less conspicuous amounts of amyloid P forming pentagonal frames. In contrast, 4.5-5-nm-wide "double tracks" characteristic of heparan sulfate proteoglycan (HSPG) were absent. The findings demonstrate that, in comparison with "typical" basement membranes in other tissues, the bone marrow sinusoidal basement membrane is uniquely specialized in several respects. Its discontinuous nature, lack of network organization, and absence of HSPG, a molecule that normally helps to maintain membrane integrity, may facilitate disassembly and reassembly of basement membrane material in concert with movements of adventitial cell processes as maturing hemopoietic cells pass through the sinusoidal wall: the

Postradiation atrophy of mature bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Erguen, H.; Howland, W.J.

1980-01-01

The growing number of oncological patients subjected to radiotherapy require the diagnostic radiologist to be aware of expected bone changes following irradiation and the differentiation of this entity from metastasis. The primary event of radiation damage to bone is atrophy and true necrosis of bone is uncommon. The postradiation atrophic changes of bone are the result of combined cellular and vascular damage, the former being more important. The damage to the osteoblast resulting in decreased matrix production is apparently the primary histopathologic event. Radiation damaged bone is susceptible to superimposed complications of fracture, infection, necrosis, and sarcoma. The primary radiographicmore » evidence of atrophy, localized osteopenia, is late in appearing, mainly because of the relative insensitivity of radiographs in detecting demineralization. Contrary to former views, the mature bone is quite radiosensitive and reacts quickly to even small doses of radiation. In vivo midrodensitometric analysis and radionuclide bone and bone marrow scans can reveal early changes following irradiation. The differentiation of postirradiation atrophy and metastasis may be difficult. Biopsy should be the last resort because of the possibility of causing true necrosis in atrophic bone by trauma and infection.« less

Postradiation atrophy of mature bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ergun, H.; Howland, W.J.

1980-01-01

The growing number of oncological patients subjected to radiotherapy require the diagnostic radiologist to be aware of expected bone changes following irradiation and the differentiation of this entity from metastasis. The primary event of radiation damage to bone is atrophy and true necrosis of bone is uncommon. The postradiation atrophic changes of bone are the result of combined cellular and vascular damage, the former being more important. The damage to the osteoblast resulting in decreased matrix production is apparently the primary histopathologic event. Radiation damaged bone is susceptible to superimposed complications of fracture, infection, necrosis, and sarcoma. The primary radiographicmore » evidence of atrophy, localized osteopenia, is late in appearing, mainly because of the relative insensitivity of radiographs in detecing demineralization. Contrary to former views, the mature bone is quite radiosensitive and reacts quickly to even small doses of radiation. In vivo midrodensitometric analysis and radionuclide bone and bone marrow scans can reveal early changes following irradiation. The differentiation of postirradiation atrophy and metastasis may be difficult. Biopsy should be the last resort because of the possibility of causing true necrosis in atrophic bone by trauma and infection.« less

Embryonic stem cell therapy improves bone quality in a model of impaired fracture healing in the mouse; tracked temporally using in vivo micro-CT.

PubMed

Taiani, J T; Buie, H R; Campbell, G M; Manske, S L; Krawetz, R J; Rancourt, D E; Boyd, S K; Matyas, J R

2014-07-01

In the current study, we used an estrogen-deficient mouse model of osteoporosis to test the efficacy of a cell-generated bone tissue construct for bone augmentation of an impaired healing fracture. A reduction in new bone formation at the defect site was observed in ovariectomized fractures compared to the control group using repeated measures in vivo micro-computed tomography (μCT) imaging over 4 weeks. A significant increase in the bone mineral density (BMD), trabecular bone volume ratio, and trabecular number, thickness and connectivity were associated with fracture repair in the control group, whereas the fractured bones of the ovariectomized mice exhibited a loss in all of these parameters (p<0.001). In a separate group, ovariectomized fractures were treated with murine embryonic stem (ES) cell-derived osteoblasts loaded in a three-dimensional collagen I gel and recovery of the bone at the defect site was observed. A significant increase in the trabecular bone volume ratio (p<0.001) and trabecular number (p<0.01) was observed by 4 weeks in the fractures treated with cell-loaded collagen matrix compared to those treated with collagen I alone. The stem cell-derived osteoblasts were identified at the fracture site at 4 weeks post-implantation through in situ hybridization histochemistry. Although this cell tracking method was effective, the formation of an ectopic cellular nodule adjacent to the knee joints of two mice suggested that alternative in vivo cell tracking methods should be employed in order to definitively assess migration of the implanted cells. To our knowledge, this study is the first of its kind to examine the efficacy of stem cell therapy for fracture repair in an osteoporosis-related fracture model in vivo. The findings presented provide novel insight into the use of stem cell therapies for bone injuries. Copyright © 2014 Elsevier Inc. All rights reserved.

Gallium modulates osteoclastic bone resorption in vitro without affecting osteoblasts

PubMed Central

Verron, Elise; Masson, Martial; Khoshniat, Solmaz; Duplomb, Laurence; Wittrant, Yohann; Baud'huin, Marc; Badran, Zahi; Bujoli, Bruno; Janvier, Pascal; Scimeca, Jean-Claude; Bouler, Jean-Michel; Guicheux, Jérôme

2010-01-01

Background and purpose: Gallium (Ga) has been shown to be effective in the treatment of disorders associated with accelerated bone loss, including cancer-related hypercalcemia and Paget's disease. These clinical applications suggest that Ga could reduce bone resorption. However, few studies have studied the effects of Ga on osteoclastic resorption. Here, we have explored the effects of Ga on bone cells in vitro. Experimental approach: In different osteoclastic models [osteoclasts isolated from long bones of neonatal rabbits (RBC), murine RAW 264.7 cells and human CD14-positive cells], we have performed resorption activity tests, staining for tartrate resistant acid phosphatase (TRAP), real-time polymerase chain reaction analysis, viability and apoptotic assays. We also evaluated the effect of Ga on osteoblasts in terms of proliferation, viability and activity by using an osteoblastic cell line (MC3T3-E1) and primary mouse osteoblasts. Key results: Gallium dose-dependently (0–100 µM) inhibited the in vitro resorption activity of RBC and induced a significant decrease in the expression level of transcripts coding for osteoclastic markers in RAW 264.7 cells. Ga also dramatically reduced the formation of TRAP-positive multinucleated cells. Ga down-regulated in a dose-dependant manner the expression of the transcription factor NFATc1. However, Ga did not affect the viability or activity of primary and MC3T3-E1 osteoblasts. Conclusions and implications: Gallium exhibits a dose-dependent anti-osteoclastic effect by reducing in vitro osteoclastic resorption, differentiation and formation without negatively affecting osteoblasts. We provide evidence that this inhibitory mechanism involves down-regulation of NFATc1 expression, a master regulator of RANK-induced osteoclastic differentiation. PMID:20397300

The Role of Peripheral Nerve Function in Age-Related Bone Loss and Changes in Bone Adaptation

DTIC Science & Technology

2013-10-01

mechanical loading (months 6-18): 2a. Strain gage analysis of bone strain during tibial compression (months 6-7) 2b. Capsaicin or vehicle treatment...of neonatal mice (months 6-8) 2c. Tibial compression of capsaicin- and vehicle-injected mice (months 8-10) 2d. Micro-computed tomography of mouse...the endosteal and periosteal surfaces. Capsaicin treatment altered bone formation rate parameters in the tibias of treated mice (Table 2). There

Targeting G-Protein Signaling for the Therapeutics of Prostate Tumor Bone Metastases and the Associated Chronic Bone Pain

DTIC Science & Technology

2015-09-01

results in increased activity/expression of key pain-sensing receptor channels, such as TRPV1 , such that the channels are constitutively activated...Keywords: Prostate Cancer Bone Metastasis, Bone Cancer Pain, Heterotrimeric G protein betagamma subunits, G protein coupled receptors (GPCRs), TRPV1 ...vitro, as well as mediating GPCR-regulated TRPV1 channel function in cultured mouse sensory neurons (Aim 1) Major Goal/Objective 1: Determine the

Targeting G-Protein Signaling for the Therapeutics of Prostate Tumor Bone Metastases and the Associated Chronic Bone Pain

DTIC Science & Technology

2013-07-01

results in increased activity/expression of key pain-sensing receptor channels, such as TRPV1 , such that the channels are constitutively activated...Keywords: Prostate Cancer Bone Metastasis, Bone Cancer Pain, Heterotrimeric G protein betagamma subunits, G protein coupled receptors (GPCRs), TRPV1 ...cell growth, migration and invasion in vitro, as well as mediating GPCR-regulated TRPV1 channel function in cultured mouse sensory neurons (Aim 1

Examining tissue composition, whole-bone morphology and mechanical behavior of GorabPrx1 mice tibiae: A mouse model of premature aging.

PubMed

Yang, Haisheng; Albiol, Laia; Chan, Wing-Lee; Wulsten, Dag; Seliger, Anne; Thelen, Michael; Thiele, Tobias; Spevak, Lyudmila; Boskey, Adele; Kornak, Uwe; Checa, Sara; Willie, Bettina M

2017-12-08

Gerodermia osteodysplastica (GO) is a segmental progeroid disorder caused by loss-of-function mutations in the GORAB gene, associated with early onset osteoporosis and bone fragility. A conditional mouse model of GO (Gorab Prx1 ) was generated in which the Gorab gene was deleted in long bones. We examined the biomechanical/functional relevance of the Gorab Prx1 mutants as a premature aging model by characterizing bone composition, tissue-level strains, and whole-bone morphology and mechanical properties of the tibia. MicroCT imaging showed that Gorab Prx1 tibiae had an increased anterior convex curvature and decreased cortical cross-sectional area, cortical thickness and moments of inertia, compared to littermate control (LC) tibiae. Fourier transform infrared (FTIR) imaging indicated a 34% decrease in mineral/matrix ratio and a 27% increase in acid phosphate content in the posterior metaphyseal cortex of the Gorab Prx1 tibiae (p < .05), suggesting delayed mineralization. In vivo strain gauge measurement and finite element analysis showed ∼two times higher tissue-level strains within the Gorab Prx1 tibiae relative to LC tibiae when subjected to axial compressive loads of the same magnitude. Three-point bending tests suggested that Gorab Prx1 tibiae were weaker and more brittle, as indicated by decreasing whole-bone strength (46%), stiffness (55%), work-to-fracture (61%) and post-yield displacement (47%). Many of these morphological and biomechanical characteristics of the Gorab Prx1 tibia recapitulated changes in other animal models of skeletal aging. Future studies are necessary to confirm how our observations might guide the way to a better understanding and treatment of GO. Copyright © 2017 Elsevier Ltd. All rights reserved.

Skeletal development of mice lacking bone sialoprotein (BSP)--impairment of long bone growth and progressive establishment of high trabecular bone mass.

PubMed

Bouleftour, Wafa; Boudiffa, Maya; Wade-Gueye, Ndeye Marième; Bouët, Guénaëlle; Cardelli, Marco; Laroche, Norbert; Vanden-Bossche, Arnaud; Thomas, Mireille; Bonnelye, Edith; Aubin, Jane E; Vico, Laurence; Lafage-Proust, Marie Hélène; Malaval, Luc

2014-01-01

Adult Ibsp-knockout mice (BSP-/-) display shorter stature, lower bone turnover and higher trabecular bone mass than wild type, the latter resulting from impaired bone resorption. Unexpectedly, BSP knockout also affects reproductive behavior, as female mice do not construct a proper "nest" for their offsprings. Multiple crossing experiments nonetheless indicated that the shorter stature and lower weight of BSP-/- mice, since birth and throughout life, as well as their shorter femur and tibia bones are independent of the genotype of the mothers, and thus reflect genetic inheritance. In BSP-/- newborns, µCT analysis revealed a delay in membranous primary ossification, with wider cranial sutures, as well as thinner femoral cortical bone and lower tissue mineral density, reflected in lower expression of bone formation markers. However, trabecular bone volume and osteoclast parameters of long bones do not differ between genotypes. Three weeks after birth, osteoclast number and surface drop in the mutants, concomitant with trabecular bone accumulation. The growth plates present a thinner hypertrophic zone in newborns with lower whole bone expression of IGF-1 and higher IHH in 6 days old BSP-/- mice. At 3 weeks the proliferating zone is thinner and the hypertrophic zone thicker in BSP-/- than in BSP+/+ mice of either sex, maybe reflecting a combination of lower chondrocyte proliferation and impaired cartilage resorption. Six days old BSP-/- mice display lower osteoblast marker expression but higher MEPE and higher osteopontin(Opn)/Runx2 ratio. Serum Opn is higher in mutants at day 6 and in adults. Thus, lack of BSP alters long bone growth and membranous/cortical primary bone formation and mineralization. Endochondral development is however normal in mutant mice and the accumulation of trabecular bone observed in adults develops progressively in the weeks following birth. Compensatory high Opn may allow normal endochondral development in BSP-/- mice, while impairing

Analysis of differences in bone removal during femoral box osteotomy for primary total knee arthroplasty.

PubMed

Graceffa, Angelo; Indelli, Pier Francesco; Basnett, Kaitlyn; Marcucci, Massimiliano

2014-01-01

this study was conducted to compare the quantity of intercondylar bone removed during femoral box osteotomy for implantation of three contemporary posterior stabilized (PS) total knee arthroplasty designs: Sigma PS (DePuy), Vanguard (Biomet) and Persona (Zimmer). we compared the maximum volumetric bone resection required for the housing of the PS mechanism of these three designs. Bone removal by each PS box cutting jig was three-dimensionally measured. The differences between the three designs were analyzed by the Kruskal-Wallis test. The Mann-Whitney U-test was used for pairwise comparisons. The level of significance was set at p<0.05. for small-size implants, the average box osteotomy volume of Persona was significantly smaller than the Vanguard and Sigma PS volumes (p=0.003). The mean difference between Vanguard and Sigma PS (p=0.01) was also significant. For medium size implants, the mean difference between Persona and Sigma PS (p=0.008) and the mean difference between Vanguard and Sigma PS (p=0.01) were statistically significant. For large size implants, the mean difference between Vanguard and Sigma PS (p=0.01) and the mean difference between Sigma PS and Persona (p=0.008) were statistically significant. irrespective of implant size, the Persona cutting jig always resected significantly less bone than did Vanguard and Sigma PS. although this study does not establish any clinical relevance of removing more or less bone at primary TKA, its results suggest that if a PS design is indicated, it is preferable to select a model which resects less distal femoral bone.

BMP-non-responsive Sca1+ CD73+ CD44+ mouse bone marrow derived osteoprogenitor cells respond to combination of VEGF and BMP-6 to display enhanced osteoblastic differentiation and ectopic bone formation.

PubMed

Madhu, Vedavathi; Li, Ching-Ju; Dighe, Abhijit S; Balian, Gary; Cui, Quanjun

2014-01-01

Clinical trials on fracture repair have challenged the effectiveness of bone morphogenetic proteins (BMPs) but suggest that delivery of mesenchymal stem cells (MSCs) might be beneficial. It has also been reported that BMPs could not increase mineralization in several MSCs populations, which adds ambiguity to the use of BMPs. However, an exogenous supply of MSCs combined with vascular endothelial growth factor (VEGF) and BMPs is reported to synergistically enhance fracture repair in animal models. To elucidate the mechanism of this synergy, we investigated the osteoblastic differentiation of cloned mouse bone marrow derived MSCs (D1 cells) in vitro in response to human recombinant proteins of VEGF, BMPs (-2, -4, -6, -9) and the combination of VEGF with BMP-6 (most potent BMP). We further investigated ectopic bone formation induced by MSCs pre-conditioned with VEGF, BMP-6 or both. No significant increase in mineralization, phosphorylation of Smads 1/5/8 and expression of the ALP, COL1A1 and osterix genes was observed upon addition of VEGF or BMPs alone to the cells in culture. The lack of CD105, Alk1 and Alk6 expression in D1 cells correlated with poor response to BMPs indicating that a greater care in the selection of MSCs is necessary. Interestingly, the combination of VEGF and BMP-6 significantly increased the expression of ALP, COL1A1 and osterix genes and D1 cells pre-conditioned with VEGF and BMP-6 induced greater bone formation in vivo than the non-conditioned control cells or the cells pre-conditioned with either VEGF or BMP-6 alone. This enhanced bone formation by MSCs correlated with higher CADM1 expression and OPG/RANKL ratio in the implants. Thus, combined action of VEGF and BMP on MSCs enhances osteoblastic differentiation of MSCs and increases their bone forming ability, which cannot be achieved through use of BMPs alone. This strategy can be effectively used for bone repair.

Stromal Gene Expression and Function in Primary Breast Tumors that Metastasize to Bone Cancer

DTIC Science & Technology

2006-07-01

surrounding bone microenvironment were investigated by purifying endothelial cells from tumor-burdened and non-tumor burdened spines . 4T1...of Balb/c mice. Fresh resected tissue (normal fat pad, primary tumor tissue or the metastatic sites spine , femur and lung) was obtained and cell... Hedgehog signalling pathway: Lasp1, CREBBP/EP300 inhibitory protein 1 and FoxP1. Of interest as well are a number of differentially regulated ESTs

Protective effects of total saponins from stem and leaf of Panax ginseng against cyclophosphamide-induced genotoxicity and apoptosis in mouse bone marrow cells and peripheral lymphocyte cells.

PubMed

Zhang, Qiu Hua; Wu, Chun Fu; Duan, Lian; Yang, Jing Yu

2008-01-01

Cyclophosphamide (CP), commonly used anti-cancer, induces oxidative stress and is cytotoxic to normal cells. It is very important to choice the protective agent combined CP to reduce the side effects in cancer treatment. Ginsenosides are biological active constituents of Panax ginseng C.A. Meyer that acts as the tonic agent for the cancer patients to reduce the side effects in the clinic application. Because CP is a pro-oxidant agent and induces oxidative stress by the generation of free radicals to decrease the activities of anti-oxidant enzymes, the protective effects of the total saponins from stem and leaf of P. ginseng C.A. Meyer (TSPG) act as an anti-oxidant agent against the decreased anti-oxidant enzymes, the genotoxicity and apoptosis induced by CP was carried out. The alkaline single cell gel electrophoresis was employed to detect DNA damage; flow cytometry assay and AO/EB staining assay were employed to measure cell apoptosis; the enzymatic anti-oxidants (T-SOD, CAT and GPx) and non-enzymatic anti-oxidant (GSH) were measured by the various colorimetric methods. CP induced the significant DNA damage in mouse peripheral lymphocytes in time- and dose-dependent manners, inhibited the activities of T-SOD, GPx and CAT, and decreased the contents of GSH in mouse blood, triggered bone marrow cell apoptosis at 6 and 12h. TSPG significantly reduced CP-induced DNA damages in bone marrow cells and peripheral lymphocyte cells, antagonized CP-induced reduction of T-SOD, GPx, CAT activities and the GSH contents, decreased the bone marrow cell apoptosis induced by CP. TSPG, significantly reduced the genotoxicity of CP in bone marrow cells and peripheral lymphocyte cells, and decreased the apoptotic cell number induced by CP in bone marrow cells. The effects of TSPG on T-SOD, GPx, CAT activities and GSH contents might partially contribute to its protective effects on CP-induced cell toxicities.

Oleoyl serine, an endogenous N-acyl amide, modulates bone remodeling and mass.

PubMed

Smoum, Reem; Bar, Arik; Tan, Bo; Milman, Garry; Attar-Namdar, Malka; Ofek, Orr; Stuart, Jordyn M; Bajayo, Alon; Tam, Joseph; Kram, Vardit; O'Dell, David; Walker, Michael J; Bradshaw, Heather B; Bab, Itai; Mechoulam, Raphael

2010-10-12

Bone mass is determined by a continuous remodeling process, whereby the mineralized matrix is being removed by osteoclasts and subsequently replaced with newly formed bone tissue produced by osteoblasts. Here we report the presence of endogenous amides of long-chain fatty acids with amino acids or with ethanolamine (N-acyl amides) in mouse bone. Of these compounds, N-oleoyl-l-serine (OS) had the highest activity in an osteoblast proliferation assay. In these cells, OS triggers a Gi-protein-coupled receptor and Erk1/2. It also mitigates osteoclast number by promoting osteoclast apoptosis through the inhibition of Erk1/2 phosphorylation and receptor activator of nuclear-κB ligand (RANKL) expression in bone marrow stromal cells and osteoblasts. In intact mice, OS moderately increases bone volume density mainly by inhibiting bone resorption. However, in a mouse ovariectomy (OVX) model for osteoporosis, OS effectively rescues bone loss by increasing bone formation and markedly restraining bone resorption. The differential effect of exogenous OS in the OVX vs. intact animals is apparently a result of an OVX-induced decrease in skeletal OS levels. These data show that OS is a previously unexplored lipid regulator of bone remodeling. It represents a lead to antiosteoporotic drug discovery, advantageous to currently available therapies, which are essentially either proformative or antiresorptive.

Implantable Self-Powered Low-Level Laser Cure System for Mouse Embryonic Osteoblasts' Proliferation and Differentiation.

PubMed

Tang, Wei; Tian, Jingjing; Zheng, Qiang; Yan, Lin; Wang, Jiangxue; Li, Zhou; Wang, Zhong Lin

2015-08-25

Bone remodeling or orthodontic treatment is usually a long-term process. It is highly desirable to speed up the process for effective medical treatment. In this work, a self-powered low-level laser cure system for osteogenesis is developed using the power generated by the triboelectric nanogenerator. It is found that the system significantly accelerated the mouse embryonic osteoblasts' proliferation and differentiation, which is essential for bone and tooth healing. The system is further demonstrated to be driven by a living creature's motions, such as human walking or a mouse's breathing, suggesting its practical use as a portable or implantable clinical cure for bone remodeling or orthodontic treatment.

Antitumor activity of the investigational proteasome inhibitor MLN9708 in mouse models of B-cell and plasma cell malignancies.

PubMed

Lee, Edmund C; Fitzgerald, Michael; Bannerman, Bret; Donelan, Jill; Bano, Kristen; Terkelsen, Jennifer; Bradley, Daniel P; Subakan, Ozlem; Silva, Matthew D; Liu, Ray; Pickard, Michael; Li, Zhi; Tayber, Olga; Li, Ping; Hales, Paul; Carsillo, Mary; Neppalli, Vishala T; Berger, Allison J; Kupperman, Erik; Manfredi, Mark; Bolen, Joseph B; Van Ness, Brian; Janz, Siegfried

2011-12-01

The clinical success of the first-in-class proteasome inhibitor bortezomib (VELCADE) has validated the proteasome as a therapeutic target for treating human cancers. MLN9708 is an investigational proteasome inhibitor that, compared with bortezomib, has improved pharmacokinetics, pharmacodynamics, and antitumor activity in preclinical studies. Here, we focused on evaluating the in vivo activity of MLN2238 (the biologically active form of MLN9708) in a variety of mouse models of hematologic malignancies, including tumor xenograft models derived from a human lymphoma cell line and primary human lymphoma tissue, and genetically engineered mouse (GEM) models of plasma cell malignancies (PCM). Both cell line-derived OCI-Ly10 and primary human lymphoma-derived PHTX22L xenograft models of diffuse large B-cell lymphoma were used to evaluate the pharmacodynamics and antitumor effects of MLN2238 and bortezomib. The iMyc(Cα)/Bcl-X(L) GEM model was used to assess their effects on de novo PCM and overall survival. The newly developed DP54-Luc-disseminated model of iMyc(Cα)/Bcl-X(L) was used to determine antitumor activity and effects on osteolytic bone disease. MLN2238 has an improved pharmacodynamic profile and antitumor activity compared with bortezomib in both OCI-Ly10 and PHTX22L models. Although both MLN2238 and bortezomib prolonged overall survival, reduced splenomegaly, and attenuated IgG2a levels in the iMyc(Cα)/Bcl-X(L) GEM model, only MLN2238 alleviated osteolytic bone disease in the DP54-Luc model. Our results clearly showed the antitumor activity of MLN2238 in a variety of mouse models of B-cell lymphoma and PCM, supporting its clinical development. MLN9708 is being evaluated in multiple phase I and I/II trials. ©2011 AACR.

Antitumor Activity of the Investigational Proteasome Inhibitor MLN9708 in Mouse Models of B-cell and Plasma Cell Malignancies

PubMed Central

Lee, Edmund C.; Fitzgerald, Michael; Bannerman, Bret; Donelan, Jill; Bano, Kristen; Terkelsen, Jennifer; Bradley, Daniel P.; Subakan, Ozlem; Silva, Matthew D.; Liu, Ray; Pickard, Michael; Li, Zhi; Tayber, Olga; Li, Ping; Hales, Paul; Carsillo, Mary; Neppalli, Vishala T.; Berger, Allison J.; Kupperman, Erik; Manfredi, Mark; Bolen, Joseph B.; Van Ness, Brian; Janz, Siegfried

2012-01-01

Purpose The clinical success of the first-in-class proteasome inhibitor bortezomib (VELCADE) has validated the proteasome as a therapeutic target for treating human cancers. MLN9708 is an investigational proteasome inhibitor that, compared with bortezomib, has improved pharmacokinetics, pharmacodynamics, and antitumor activity in preclinical studies. Here, we focused on evaluating the in vivo activity of MLN2238 (the biologically active form of MLN9708) in a variety of mouse models of hematologic malignancies, including tumor xenograft models derived from a human lymphoma cell line and primary human lymphoma tissue, and genetically engineered mouse (GEM) models of plasma cell malignancies (PCM). Experimental Design Both cell line–derived OCI-Ly10 and primary human lymphoma–derived PHTX22L xenograft models of diffuse large B-cell lymphoma were used to evaluate the pharmacodynamics and antitumor effects of MLN2238 and bortezomib. The iMycCα/Bcl-XL GEM model was used to assess their effects on de novo PCM and overall survival. The newly developed DP54-Luc–disseminated model of iMycCα/ Bcl-XL was used to determine antitumor activity and effects on osteolytic bone disease. Results MLN2238 has an improved pharmacodynamic profile and antitumor activity compared with bortezomib in both OCI-Ly10 and PHTX22L models. Although both MLN2238 and bortezomib prolonged overall survival, reduced splenomegaly, and attenuated IgG2a levels in the iMycCα/Bcl-XL GEM model, only MLN2238 alleviated osteolytic bone disease in the DP54-Luc model. Conclusions Our results clearly showed the antitumor activity of MLN2238 in a variety of mouse models of B-cell lymphoma and PCM, supporting its clinical development. MLN9708 is being evaluated in multiple phase I and I/II trials. PMID:21903769

Tumorigenic target cell regions in bone marrow studied by localized dosimetry of 239Pu, 241Am and 233U in the mouse femur.

PubMed

Lord, B I; Austin, A L; Ellender, M; Haines, J W; Harrison, J D

2001-06-01

To study the temporal change in microdistribution of plutonium-239, americium-241 and uranium-233 in the mouse distal femur and to compare and combine calculated radiation doses with those obtained previously for the femoral shaft. Also, to relate doses to relative risks of osteosarcoma and acute myeloid leukaemia. Computer-based image analysis of neutron-induced and alpha-track autoradiographs of sections of mouse femora was used to quantify the microdistribution of (239)Pu, (241)Am and (233)U from 1 to 448 days after intraperitoneal injection. Localized dose-rates and cumulative doses over this period were calculated for different regions of the marrow spaces in trabecular bone. The results were then combined with previous data for doses to the cortical marrow of the femoral shaft. A morphometric analysis of the distal femur was carried out. Initial deposition on endosteal surfaces and dose-rates near to the trabecular surfaces at 1 day were two to four times greater than corresponding results for cortical bone. Burial was most rapid for (233)U, about twice the rate in cortical bone. As in cortical bone, subsequent uptake into the marrow was seen for (239)Pu and (241)Am but not (233)U. Cumulative doses to 448 days for different regions of trabecular marrow were greater than corresponding values for cortical marrow for each radionuclide. Combined doses reflected the greater overall volume of cortical marrow. Cumulative radiation doses to the 10 microm thick band of marrow adjacent to all endosteal surfaces were in the ratio of approximately 7:3:1 for (239)Pu:(241)Am:(233)U. This ratio is not inconsistent with observed incidences of osteosarcoma induction by the three nuclides. Analysis of doses to different depths of marrow, however, showed that although ratios were probably not significantly different to that for a 10 microm depth, better correlations with osteosarcomagenic risk were obtained with 20-40 microm depths. For acute myeloid leukaemia, the closest

CDK2 phosphorylation of Smad2 disrupts TGF-beta transcriptional regulation in resistant primary bone marrow myeloma cells.

PubMed

Baughn, Linda B; Di Liberto, Maurizio; Niesvizky, Ruben; Cho, Hearn J; Jayabalan, David; Lane, Joseph; Liu, Fang; Chen-Kiang, Selina

2009-02-15

Resistance to growth suppression by TGF-beta1 is common in cancer; however, mutations in this pathway are rare in hematopoietic malignancies. In multiple myeloma, a fatal cancer of plasma cells, malignant cells accumulate in the TGF-beta-rich bone marrow due to loss of both cell cycle and apoptotic controls. Herein we show that TGF-beta activates Smad2 but fails to induce cell cycle arrest or apoptosis in primary bone marrow myeloma and human myeloma cell lines due to its inability to activate G(1) cyclin-dependent kinase (CDK) inhibitors (p15(INK4b), p21(CIP1/WAF1), p27(KIP1), p57(KIP2)) or to repress c-myc and Bcl-2 transcription. Correlating with aberrant activation of CDKs, CDK-dependent phosphorylation of Smad2 on Thr(8) (pT8), a modification linked to impaired Smad activity, is elevated in primary bone marrow myeloma cells, even in asymptomatic monoclonal gammopathy of undetermined significance. Moreover, CDK2 is the predominant CDK that phosphorylates Smad2 on T8 in myeloma cells, leading to inhibition of Smad2-Smad4 association that precludes transcriptional regulation by Smad2. Our findings provide the first direct evidence that pT8 Smad2 couples dysregulation of CDK2 to TGF-beta resistance in primary cancer cells, and they suggest that disruption of Smad2 function by CDK2 phosphorylation acts as a mechanism for TGF-beta resistance in multiple myeloma.

In vitro and in vivo evidence for orphan nuclear receptor RORα function in bone metabolism

PubMed Central

Meyer, Thomas; Kneissel, Michaela; Mariani, Jean; Fournier, Brigitte

2000-01-01

Bone is a major target site for steroid hormone action. Steroid hormones like cortisol, vitamin D, and estradiol are responsible for principal events associated with bone formation and resorption. Over the past decade, new members of the nuclear hormone gene family have been identified that lack known ligands. These orphan receptors can be used to uncover signaling molecules that regulate yet unidentified physiological networks. In the present study the function of retinoic acid receptor-related orphan receptor (ROR) α in bone metabolism has been examined. We showed that RORα and RORγ, but not RORβ, are expressed in mesenchymal stem cells derived from bone marrow. Interestingly, for RORα we observed an increased messenger signal expression between control cells and cells undergoing osteogenic differentiation. Furthermore, the direct activation of mouse bone sialoprotein by RORα, typically 7-fold, has been shown. In contrast, transient overexpression of RORα overrides the activation of the osteocalcin promoter by 1α,25-dihydroxyvitamin D3. In addition, we have investigated bone mass parameters and bone geometry in the mouse mutant staggerer (sg/sg), a mouse strain that carries a deletion within the RORα gene. Homozygote mutants have thin long bones compared with the heterozygote animals and wild-type littermates. More interestingly, the bones of the sg/sg animals are osteopenic as indicated by the comparison of bone mineral contents of sg/sg animals to the heterozygote and wild-type animals. We conclude that these in vitro and in vivo results suggest a function for RORα in bone biology. RORα most likely acts by direct modulation of a bone matrix component. PMID:10900268

Magnetic resonance guided focused ultrasound surgery (MRgFUS) of bone metastases: From primary pain palliation to local tumor control

NASA Astrophysics Data System (ADS)

Napoli, A.; Leonardi, A.; Andrani, F.; Boni, F.; Anzidei, M.; Catalano, C.

2017-03-01

Purpose: To evaluate the clinical performance of MRgFUS in primary pain palliation of painful bone metastases and in local tumor control. Materials and Methods: We enrolled 26 consecutive patients (female/male 12/14; age: 64.7±7.5yrs) with painful bone metastases. Before and 3 months after MRgFUS treatment pain severity and pain interference scores were assessed according to Brief Pain Inventory-Quality of Life (BPI-QoL) criteria and patients underwent both CT and MRI. Local tumor control was evaluated according to lesion size, density and perfusion at CT, dynamic contrast enhancement at MRI (Discovery 750HD, GE; Gd-Bopta, Bracco) and metabolic activity at PET or scintigraphy. Patients were classified as responders or non-responders. Results: No treatment-related adverse events were recorded during the study. As statistically significant difference between baseline and follow-up values for both pain severity and pain interference scores was observed (p<0.05). Increased bone density was observed in 9/26 (34.6%) patients. Non-Perfused Volume values ranged between 20% and 92%. There was no difference in NPV values between responders and non-responders (46.7±24.2% [25 - 90 %] vs. 45±24.9% [20 - 93 %]; p=0.7). In 6 patients (5 prostate and 1 breast primary cancer) there was nearly absence of metabolic activity after treatment (mean SUV=1.2). Conclusion: MRgFUS can be safely and effectively used as the primary treatment for pain palliation in patients with painful bone metastases; moreover our experience demonstrated also a potential role for the MRgFUS in local tumor control.

Inhibition of experimental bone resorption and osteoclast formation and survival by 2-aminoethanesulphonic acid.

PubMed

Koide, M; Okahashi, N; Tanaka, R; Kazuno, K; Shibasaki, K; Yamazaki, Y; Kaneko, K; Ueda, N; Ohguchi, M; Ishihara, Y; Noguchi, T; Nishihara, T

1999-09-01

It is known that bone resorption is mediated by osteoclasts, and lipopolysaccharide (LPS) and inflammatory mediators such as interleukin-1 (IL-1) and prostaglandin E2 (PGE2) induce osteoclast differentiation from haemopoietic cells, 2-aminoethanesulphonic acid, which is known as taurine, is an important nutrient and is added to most synthetic human infant milk formulas. In this study, it was found that 2-aminoethanesulphonic acid inhibits the stimulation of bone resorption mediated by LPS of the periodontopathic microorganism Actinobacillus actinomycetemcomitans Y4 in organ cultures of newborn mouse calvaria. The effect of 2-aminoethanesulphonic acid on the development and survival of osteoclast-like multinucleated cells produced in a mouse bone-marrow culture system was also examined. 2-aminoethanesulphonic acid (100 microg/ml) suppressed the formation of these osteoclast-like cells in the presence of LPS of A. actinomycetemcomitans Y4, IL-1alpha or PGE2 in mouse marrow cultures. On the other hand, 2-aminoethanesulphonic acid did not inhibit 1alpha, 25-dihydroxyvitamin D3-mediated osteoclast differentiation. Although IL-1alpha elongated the survival of the osteoclast-like cells, 2-aminoethanesulphonic acid blocked the supportive effect of IL-1alpha on osteoclast survival. 2-aminoethanesulphonic acid showed no effect on the growth of mouse osteoblasts. Finally, it was found that 2-aminoethanesulphonic acid inhibited alveolar bone resorption in experimental periodontitis in hamsters. These results suggest that 2-aminoethanesulphonic acid is an effective agent in preventing inflammatory bone resorption in periodontal diseases.

Leptin regulation of bone resorption by the sympathetic nervous system and CART.

PubMed

Elefteriou, Florent; Ahn, Jong Deok; Takeda, Shu; Starbuck, Michael; Yang, Xiangli; Liu, Xiuyun; Kondo, Hisataka; Richards, William G; Bannon, Tony W; Noda, Masaki; Clement, Karine; Vaisse, Christian; Karsenty, Gerard

2005-03-24

Bone remodelling, the mechanism by which vertebrates regulate bone mass, comprises two phases, namely resorption by osteoclasts and formation by osteoblasts; osteoblasts are multifunctional cells also controlling osteoclast differentiation. Sympathetic signalling via beta2-adrenergic receptors (Adrb2) present on osteoblasts controls bone formation downstream of leptin. Here we show, by analysing Adrb2-deficient mice, that the sympathetic nervous system favours bone resorption by increasing expression in osteoblast progenitor cells of the osteoclast differentiation factor Rankl. This sympathetic function requires phosphorylation (by protein kinase A) of ATF4, a cell-specific CREB-related transcription factor essential for osteoblast differentiation and function. That bone resorption cannot increase in gonadectomized Adrb2-deficient mice highlights the biological importance of this regulation, but also contrasts sharply with the increase in bone resorption characterizing another hypogonadic mouse with low sympathetic tone, the ob/ob mouse. This discrepancy is explained, in part, by the fact that CART ('cocaine amphetamine regulated transcript'), a neuropeptide whose expression is controlled by leptin and nearly abolished in ob/ob mice, inhibits bone resorption by modulating Rankl expression. Our study establishes that leptin-regulated neural pathways control both aspects of bone remodelling, and demonstrates that integrity of sympathetic signalling is necessary for the increase in bone resorption caused by gonadal failure.

Stromal Gene Expression and Function in Primary Breast Tumors that Metastasize to Bone Cancer

DTIC Science & Technology

2004-07-01

by quantitative RT-QPCR. Further analysis of epithelium from matched spine metastases revealed some genes that were up-regulated further at the...9-fold in matched bone metastases. The expression in spine metastases was verified by in situ hybridisation whilst the expression of stefin Al in...growth in the mammary gland of Balb/c mice. Fresh resected tissue (normal fat pad, primary tumor tissue or the metastatic sites spine , femur and lung) was

Effects of Polymethoxyflavonoids on Bone Loss Induced by Estrogen Deficiency and by LPS-Dependent Inflammation in Mice.

PubMed

Matsumoto, Shigeru; Tominari, Tsukasa; Matsumoto, Chiho; Yoshinouchi, Shosei; Ichimaru, Ryota; Watanabe, Kenta; Hirata, Michiko; Grundler, Florian M W; Miyaura, Chisato; Inada, Masaki

2018-01-20

Polymethoxyflavonoids (PMFs) are a family of the natural compounds that mainly compise nobiletin, tangeretin, heptamethoxyflavone (HMF), and tetramethoxyflavone (TMF) in citrus fruits. PMFs have shown various biological functions, including anti-oxidative effects. We previously showed that nobiletin, tangeretin, and HMF all inhibited interleukin (IL)-1-mediated osteoclast differentiation via the inhibition of prostaglandin E2 synthesis. In this study, we created an original mixture of PMFs (nobiletin, tangeretin, HMF, and TMF) and examined whether or not PMFs exhibit co-operative inhibitory effects on osteoclastogenesis and bone resorption. In a coculture of bone marrow cells and osteoblasts, PMFs dose-dependently inhibited IL-1-induced osteoclast differentiation and bone resorption. The optimum concentration of PMFs was lower than that of nobiletin alone in the suppression of osteoclast differentiation, suggesting that the potency of PMFs was stronger than that of nobiletin in vitro. The oral administration of PMFs recovered the femoral bone loss induced by estrogen deficiency in ovariectomized mice. We further tested the effects of PMFs on lipopolysaccharide-induced bone resorption in mouse alveolar bone. In an ex vivo experimental model for periodontitis, PMFs significantly suppressed the bone-resorbing activity in organ cultures of mouse alveolar bone. These results indicate that a mixture of purified nobiletin, tangeretin, HMF, and TMF exhibits a co-operative inhibitory effect for the protection against bone loss in a mouse model of bone disease, suggesting that PMFs may be potential candidates for the prevention of bone resorption diseases, such as osteoporosis and periodontitis.

Spontaneous osteosarcoma of the femur in a non-obese diabetic mouse

PubMed Central

Hong, Sunhwa; Lee, Hyun-A; Choe, Ohmok; Chung, Youngho

2011-01-01

An abnormal swelling was identified in the distal portion of the right femur in a 1-year-old non-obese diabetic (NOD) mouse. Grossly, a large mass of the distal femur was observed in the right femur. Lesions were poorly marginated, associated with destruction of the cancellous and cortical elements of the bone, and showed ossification within the soft tissue component. Histologically, the tumor was identified as a poorly differentiated sarcoma. Histopathologic examination of the bone masses revealed invasive proliferation of poorly differentiated neoplastic mesenchymal cells forming streams, bundles, and nests, which resulted in destruction of normal bone. Neoplastic cells exhibited random variation in cellular appearance and arrangement, as well as matrix composition and abundance. Haphazard and often intermingling patterns of osteogenic, chondroblastic, lipoblastic, and angiogenic tissues were present. Larger areas of neoplastic bone and hyaline cartilage contained multiple large areas of hemorrhage and necrosis bordered by neoplastic cells. The mass was diagnosed as an osteosarcoma. To our knowledge, this is the first spontaneous osteosarcoma in an NOD mouse. PMID:21998615

Bone x-ray

MedlinePlus

... different views of the bone may be uncomfortable. Why the Test is Performed A bone x-ray ... neoplasia (MEN) II Multiple myeloma Osgood-Schlatter disease Osteogenesis imperfecta Osteomalacia Paget's disease Primary hyperparathyroidism Rickets Risks There ...

Heterogenic transplantation of bone marrow-derived rhesus macaque mesenchymal stem cells ameliorates liver fibrosis induced by carbon tetrachloride in mouse

PubMed Central

Yan, Yaping; Wang, Junfeng; Duan, Yanchao; Li, Shanshan; Yan, Li; Wang, Hong; Chen, Bingbing; Sang, Xiongbo; Ji, Weizhi

2018-01-01

Liver fibrosis is a disease that causes high morbidity and has become a major health problem. Liver fibrosis can lead to the end stage of liver diseases (livercirrhosisand hepatocellularcarcinoma). Currently, liver transplantation is the only effective treatment for end-stage liver disease. However, the shortage of organ donors, high cost of medical surgery, immunological rejection and transplantation complications severely hamper liver transplantation therapy. Mesenchymal stem cells (MSCs) have been regarded as promising cells for clinical applications in stem cell therapy in the treatment of liver diseases due to their unique multipotent differentiation capacity, immunoregulation and paracrine effects. Although liver fibrosis improvements by MSC transplantation in preclinical experiments as well as clinical trials have been reported, the in vivo fate of MSCs after transportation and their therapeutic mechanisms remain unclear. In this present study, we isolated MSCs from the bone marrow of rhesus macaques. The cells exhibited typical MSC markers and could differentiate into chondrocytes, osteocytes, and adipocytes, which were not affected by labeling with enhanced green fluorescent protein (EGFP). The harvested MSCs respond to interferon-γ stimulation and have the ability to inhibit lymphocyte proliferation in vitro. EGFP-labeled MSCs (1 × 106 cells) were transplanted into mice with carbon tetrachloride-induced liver fibrosis via tail vein injection. The ability of the heterogenic MSC infusion to ameliorate liver fibrosis in mice was evaluated by a blood plasma chemistry index, pathological examination and liver fibrosis-associated gene expression. Additionally, a small number of MSCs that homed and engrafted in the mouse liver tissues were evaluated by immunofluorescence analysis. Our results showed that the transplantation of heterogenic MSCs derived from monkey bone marrow can be used to treat liver fibrosis in the mouse model and that the paracrine

Primary aneurysmal bone cyst of coronoid process

PubMed Central

Goyal, Amit; Tyagi, Isha; Syal, Rajan; Agrawal, Tanu; Jain, Manoj

2006-01-01

Background Aneurysmal bone cysts are relatively uncommon in the facial skeleton. These usually affect the mandible but origin from the coronoid process is even rarer. To the best of our knowledge, this is the first reported case of a coronoid process aneurysmal bone cyst presenting as temporal fossa swelling. Case presentation A 17 year old boy presented with a progressively increasing swelling in the left temporal region developed over the previous 8 months. An expansile lytic cystic lesion originating from the coronoid process of the left mandible and extending into the infratemporal and temporal fossa regions was found on CT scan. It was removed by a superior approach to the infratemporal fossa. Conclusion Aneurysmal bone cyst of the coronoid process can attain enormous dimensions until the temporal region is also involved. A superior approach to the infratemporal fossa is a reasonable approach for such cases, providing wide exposure and access to all parts of the lesion and ensuring better control and complete excision. PMID:16533409

Effects of alkylphenols on bone metabolism in vivo and in vitro.

PubMed

Hagiwara, Hiromi; Sugizaki, Toshinori; Tsukamoto, Yu; Senoh, Emi; Goto, Tadashi; Ishihara, Yoko

2008-09-01

Alkylphenols are endocrine disruptors that show estrogen-like effects in various wildlife species. However, little information is available about the action of these chemicals on bone metabolism. We investigated the effects of alkylphenols, such as nonylphenol (NP) and octylphenol (OP), on the formation of bone using several culture systems for osteoclasts and osteoblasts, as well as in vivo experiments. NP and OP dose-dependently inhibited the formation of tartrate-resistant acid phosphatase-positive multinucleated cells (osteoclasts) in cocultures of mouse spleen cells or mouse bone marrow cells with ST2 cells. However, beta-estradiol at 10(-9)M to 10(-6)M did not affect this process. In contrast, neither compound affected the proliferation and differentiation of rat calvarial osteoblast-like cells (ROB cells). When NP or OP (0.1mg/kg body weight) was administered subcutaneously to pregnant mice at 10 days, 12 days and 14 days post-coitus, fetuses at 17.5 days post-coitus showed stimulation of sternebrae bone calcification. Our findings suggest that alkylphenols have critical effects on the formation of bone by non-estrogenic effects.

Muscle-Bone Interactions in Pediatric Bone Diseases.

PubMed

Veilleux, Louis-Nicolas; Rauch, Frank

2017-10-01

Here, we review the skeletal effects of pediatric muscle disorders as well as muscle impairment in pediatric bone disorders. When starting in utero, muscle disorders can lead to congenital multiple contractures. Pediatric-onset muscle weakness such as cerebral palsy, Duchenne muscular dystrophy, spinal muscular atrophy, or spina bifida typically are associated with small diameter of long-bone shafts, low density of metaphyseal bone, and increased fracture incidence in the lower extremities, in particular, the distal femur. Primary bone diseases can affect muscles through generic mechanisms, such as decreased physical activity or in disease-specific ways. For example, the collagen defect underlying the bone fragility of osteogenesis imperfecta may also affect muscle force generation or transmission. Transforming growth factor beta released from bone in Camurati Engelman disease may decrease muscle function. Considering muscle-bone interactions does not only contribute to the understanding of musculoskeletal disorders but also can identify new targets for therapeutic interventions.

Increased formation of autophagosomes in ectromelia virus-infected primary culture of murine bone marrow-derived macrophages.

PubMed

Martyniszyn, L; Szulc-Dąbrowska, L; Boratyńska-Jasińska, A; Niemiałtowski, M

2013-01-01

Induction of autophagy by ectromelia virus (ECTV) in primary cultures of bone marrow-derived macrophages (BMDMs) was investigated. The results showed that ECTV infection of BMDMs resulted in increased formation of autophagosomes, increased level of LC3-II protein present in aggregates and extensive cytoplasmic vacuolization. These data indicate an increased autophagic activity in BMDMs during ECTV infection.

The Terminator mouse is a diphtheria toxin-receptor knock-in mouse strain for rapid and efficient enrichment of desired cell lineages.

PubMed

Guo, Jian-Kan; Shi, Hongmei; Koraishy, Farrukh; Marlier, Arnaud; Ding, Zhaowei; Shan, Alan; Cantley, Lloyd G

2013-11-01

Biomedical research often requires primary cultures of specific cell types, which are challenging to obtain at high purity in a reproducible manner. Here we engineered the murine Rosa26 locus by introducing the diphtheria toxin receptor flanked by loxP sites. The resultant strain was nicknamed the Terminator mouse. This approach results in diphtheria toxin-receptor expression in all non-Cre expressing cell types, making these cells susceptible to diphtheria toxin exposure. In primary cultures of kidney cells derived from the Terminator mouse, over 99.99% of cells were dead within 72 h of diphtheria toxin treatment. After crossing the Terminator with the podocin-Cre (podocyte specific) mouse or the Ggt-Cre (proximal tubule specific) mouse, diphtheria toxin treatment killed non-Cre expressing cells but spared podocytes and proximal tubule cells, respectively, enriching the primary cultures to over 99% purity, based on both western blotting and immunostaining of marker proteins. Thus, the Terminator mouse can be a useful tool to selectively and reproducibly obtain even low-abundant cell types at high quantity and purity.

Differential expression of liver and kidney proteins in a mouse model for primary hyperoxaluria type I.

PubMed

Hernández-Fernaud, Juan R; Salido, Eduardo

2010-11-01

Mutations in the alanine-glyoxylate aminotransferase gene (AGXT) are responsible for primary hyperoxaluria type I, a rare disease characterized by excessive hepatic oxalate production that leads to renal failure. A deeper understanding of the changes in the metabolic pathways secondary to the lack of AGXT expression is needed in order to explore substrate depletion as a therapeutic strategy to limit oxalate production in primary hyperoxaluria type I. We have developed an Agxt knockout (AgxtKO) mouse that reproduces some key features of primary hyperoxaluria type I. To improve our understanding of the metabolic adjustments subsequent to AGXT deficiency, we performed a proteomic analysis of the changes in expression levels of various subcellular fractions of liver and kidney metabolism linked to the lack of AGXT. In this article, we report specific changes in the liver and kidney proteome of AgxtKO mice that point to significant variations in gluconeogenesis, glycolysis and fatty acid pathways. Journal compilation © 2010 FEBS. No claim to original German government works.

Micro-CT evaluation of bone defects: applications to osteolytic bone metastases, bone cysts, and fracture.

PubMed

Buie, Helen R; Bosma, Nick A; Downey, Charlene M; Jirik, Frank R; Boyd, Steven K

2013-11-01

Bone defects can occur in various forms and present challenges to performing a standard micro-CT evaluation of bone quality because most measures are suited to homogeneous structures rather than ones with spatially focal abnormalities. Such defects are commonly associated with pain and fragility. Research involving bone defects requires quantitative approaches to be developed if micro-CT is to be employed. In this study, we demonstrate that measures of inter-microarchitectural bone spacing are sensitive to the presence of focal defects in the proximal tibia of two distinctly different mouse models: a burr-hole model for fracture healing research, and a model of osteolytic bone metastases. In these models, the cortical and trabecular bone compartments were both affected by the defect and were, therefore, evaluated as a single unit to avoid splitting the defects into multiple analysis regions. The burr-hole defect increased mean spacing (Sp) by 27.6%, spacing standard deviation (SpSD) by 113%, and maximum spacing (Spmax) by 72.8%. Regression modeling revealed SpSD (β=0.974, p<0.0001) to be a significant predictor of the defect volume (R(2)=0.949) and Spmax (β=0.712, p<0.0001) and SpSD (β=0.271, p=0.022) to be significant predictors of the defect diameter (R(2)=0.954). In the mice with osteolytic bone metastases, spacing parameters followed similar patterns of change as reflected by other imaging technologies, specifically bioluminescence data which is indicative of tumor burden. These data highlight the sensitivity of spacing measurements to bone architectural abnormalities from 3D micro-CT data and provide a tool for quantitative evaluation of defects within a bone. Copyright © 2013 IPEM. Published by Elsevier Ltd. All rights reserved.

Oleoyl serine, an endogenous N-acyl amide, modulates bone remodeling and mass

PubMed Central

Smoum, Reem; Bar, Arik; Tan, Bo; Milman, Garry; Attar-Namdar, Malka; Ofek, Orr; Stuart, Jordyn M.; Bajayo, Alon; Tam, Joseph; Kram, Vardit; O'Dell, David; Walker, Michael J.; Bradshaw, Heather B.; Bab, Itai; Mechoulam, Raphael

2010-01-01

Bone mass is determined by a continuous remodeling process, whereby the mineralized matrix is being removed by osteoclasts and subsequently replaced with newly formed bone tissue produced by osteoblasts. Here we report the presence of endogenous amides of long-chain fatty acids with amino acids or with ethanolamine (N-acyl amides) in mouse bone. Of these compounds, N-oleoyl-l-serine (OS) had the highest activity in an osteoblast proliferation assay. In these cells, OS triggers a Gi-protein-coupled receptor and Erk1/2. It also mitigates osteoclast number by promoting osteoclast apoptosis through the inhibition of Erk1/2 phosphorylation and receptor activator of nuclear-κB ligand (RANKL) expression in bone marrow stromal cells and osteoblasts. In intact mice, OS moderately increases bone volume density mainly by inhibiting bone resorption. However, in a mouse ovariectomy (OVX) model for osteoporosis, OS effectively rescues bone loss by increasing bone formation and markedly restraining bone resorption. The differential effect of exogenous OS in the OVX vs. intact animals is apparently a result of an OVX-induced decrease in skeletal OS levels. These data show that OS is a previously unexplored lipid regulator of bone remodeling. It represents a lead to antiosteoporotic drug discovery, advantageous to currently available therapies, which are essentially either proformative or antiresorptive. PMID:20876113

Hepatocyte growth factor improves bone regeneration via the bone morphogenetic protein‑2‑mediated NF‑κB signaling pathway.

PubMed

Zhen, Ruixin; Yang, Jianing; Wang, Yu; Li, Yubo; Chen, Bin; Song, Youxin; Ma, Guiyun; Yang, Bo

2018-04-01

Bone regeneration is an important process associated with the treatment of osteonecrosis, which is caused by various factors. Hepatocyte growth factor (HGF) is an active biological factor that has multifunctional roles in cell biology, life sciences and clinical medicine. It has previously been suggested that bone morphogenetic protein (BMP)‑2 exerts beneficial roles in bone formation, repair and angiogenesis in the femoral head. The present study aimed to investigate the benefits and molecular mechanisms of HGF in bone regeneration. The viability of osteoblasts and osteoclasts were studied in vitro. In addition, the expression levels of tumor necrosis factor (TNF)‑α, monocyte chemotactic protein (MCP)‑1, interleukin (IL)‑1 and IL‑6 were detected in a mouse fracture model following treatment with HGF. The expression and activity of nuclear factor (NF)‑κB were also analyzed in osteocytes post‑treatment with HGF. Histological analysis was used to determine the therapeutic effects of HGF on mice with fractures. The migration and differentiation of osteoblasts and osteoclasts were investigated in HGF‑incubated cells. Furthermore, angiogenesis and BMP‑2 expression were analyzed in the mouse fracture model post‑treatment with HGF. The results indicated that HGF regulates the cell viability of osteoblasts and osteoclasts, and also balanced the ratio between osteoblasts and osteoclasts. In addition, HGF decreased the serum expression levels of TNF‑α, MCP‑1, IL‑1 and IL‑6 in experimental mice. The results of a mechanistic analysis demonstrated that HGF upregulated p65, IκB kinase‑β and IκBα expression in osteoblasts from experimental mice. In addition, the expression levels of vascular endothelial growth factor, BMP‑2 receptor, receptor activator of NF‑κB ligand and macrophage colony‑stimulating factor were upregulated by HGF, which may effectively promote blood vessel regeneration, and contribute to the formation and

Hydrocortisone effect of arylsulfatase A in primary mouse brain cell cultures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marcelo, A.; Pieringer, R.A.

The primary goal of this study was to study the mechanism of action of hydrocortisone (HC) on arylsulfatase A (ASA) in primary cultures of cells that were dissociated from the brains of embryonic mice. Cells were cultured in a defined medium in the absence or in the presence of 3 ..mu..M HC. The specific activity of ASA in nontreated cells was 1.297 U/mg (U = ..mu..mol/hr) while the value for the HC-treated cells was 0.783 U/mg. The authors data shows that HC inhibits ASA activity in these cultures cells (p < 0.001). The determination of the ASA enzyme activity wasmore » assayed primarily with the artificial substrate p-nitrocatechol sulfate. However, the natural substrate (cerebroside /sup 35/S-sulfate) also as active and correlated linearly with the activity of p-nitrocatechol sulfate. Purified ASA was isolated from calf brains and used to generate an antibody (Ab) against ASA. The specificity of the Ab for the ASA protein of cell cultures was tested in Ouchterlony double immunodiffusion studies. The Ab was used in a competitive enzyme-linked immunosorbent assay to quantify the number of ASA molecules in the cell extracts from the embryonic mouse cell cultures. Preliminary data suggest that HC decreases the number of ASA molecules.« less

Relationship between markers of body fat and calcaneal bone stiffness differs between preschool and primary school children: results from the IDEFICS baseline survey.

PubMed

Sioen, Isabelle; Mouratidou, Theodora; Herrmann, Diana; De Henauw, Stefaan; Kaufman, Jean-Marc; Molnár, Dénes; Moreno, Luis A; Marild, Staffan; Barba, Gianvincenzo; Siani, Alfonso; Gianfagna, Francesco; Tornaritis, Michael; Veidebaum, Toomas; Ahrens, Wolfgang

2012-10-01

The aim of this study was to investigate the relationship between markers of body fat and bone status assessed as calcaneal bone stiffness in a large sample of European healthy pre- and primary school children. Participants were 7,447 children from the IDEFICS study (spread over eight different European countries), age 6.1 ± 1.8 years (range 2.1-9.9), 50.5 % boys. Anthropometric measurements (weight, height, bioelectrical impedance, waist and hip circumference, and tricipital and subscapular skinfold thickness) as well as quantitative ultrasonographic measurements to determine calcaneal stiffness index (SI) were performed. Partial correlation analysis, linear regression analysis, and ANCOVA were stratified by sex and age group: preschool boys (n = 1,699) and girls (n = 1,599) and primary school boys (n = 2,062) and girls (n = 2,087). In the overall study population, the average calcaneal SI was equal to 80.2 ± 14.0, ranging 42.4-153. The results showed that preschool children with higher body fat had lower calcaneal SI (significant correlation coefficients between -0.05 and -0.20), while primary school children with higher body fat had higher calcaneal SI (significant correlation coefficients between 0.05 and 0.13). After adjusting for fat-free mass, both preschool and primary school children showed an inverse relationship between body fat and calcaneal stiffness. To conclude, body fat is negatively associated with calcaneal bone stiffness in children after adjustment for fat-free mass. Fat-free mass may confound the association in primary school children but not in preschool children. Muscle mass may therefore be an important determinant of bone stiffness.

Modulation of bone resorption by phosphorylation state of bone sialoprotein.

PubMed

Curtin, Paul; McHugh, Kevin P; Zhou, Hai-Yan; Flückiger, Rudolf; Goldhaber, Paul; Oppenheim, Frank G; Salih, Erdjan

2009-07-28

We have determined transmembrane protein tyrosine phosphorylation (outside-in signaling) in cultured osteoclasts and macrophages in response to added native purified bone sialoprotein (nBSP) and its dephosphorylated form (dBSP). There were selective/differential and potent inhibitory effects by dBSP and minimal effect by nBSP on intracellular tyrosine phosphorylation in macrophages and osteoclasts. Further studies on the downstream gene expression effects led to identification of a large number of differentially expressed genes in response to nBSP relative to dBSP in both macrophages and osteoclasts. These studies were extended to a bone resorption model using live mouse neonatal calvarial bone organ cultures stimulated by parathyroid hormone (PTH) to undergo bone resorption. Inclusion of nBSP in such cultures showed no effect on type I collagen telopeptide fragment release, hence overall bone resorption, whereas addition of dBSP abolished the PTH-induced bone resorption. The inhibition of bone resorption by dBSP was shown to be unique since in complementary experiments use of integrin receptor binding ligand, GRGDS peptide, offered only partial reduction on overall bone resorption. Quantitative RANKL analysis indicated that mechanistically the PTH-induced bone resorption was inhibited by dBSP via down-regulation of the osteoblastic RANKL production. This conclusion was supported by the RANKL analysis in cultured MC3T3-E1 osteoblast cells. Overall, these studies provided direct evidence for the involvement of covalently bound phosphates on BSP in receptor mediated "outside-in" signaling via transmembrane tyrosine phosphorylation with concurrent effects on downstream gene expressions. The use of a live bone organ culture system augmented these results with further evidence that links the observed in vivo variable state of phosphorylation with bone remodeling.

In utero transplantation of adult bone marrow decreases perinatal lethality and rescues the bone phenotype in the knockin murine model for classical, dominant osteogenesis imperfecta

PubMed Central

Panaroni, Cristina; Gioia, Roberta; Lupi, Anna; Besio, Roberta; Goldstein, Steven A.; Kreider, Jaclynn; Leikin, Sergey; Vera, Juan Carlos; Mertz, Edward L.; Perilli, Egon; Baruffaldi, Fabio; Villa, Isabella; Farina, Aurora; Casasco, Marco; Cetta, Giuseppe; Rossi, Antonio; Frattini, Annalisa; Marini, Joan C.; Vezzoni, Paolo

2009-01-01

Autosomal dominant osteogenesis imperfecta (OI) caused by glycine substitutions in type I collagen is a paradigmatic disorder for stem cell therapy. Bone marrow transplantation in OI children has produced a low engraftment rate, but surprisingly encouraging symptomatic improvements. In utero transplantation (IUT) may hold even more promise. However, systematic studies of both methods have so far been limited to a recessive mouse model. In this study, we evaluated intrauterine transplantation of adult bone marrow into heterozygous BrtlIV mice. Brtl is a knockin mouse with a classical glycine substitution in type I collagen [α1(I)-Gly349Cys], dominant trait transmission, and a phenotype resembling moderately severe and lethal OI. Adult bone marrow donor cells from enhanced green fluorescent protein (eGFP) transgenic mice engrafted in hematopoietic and nonhematopoietic tissues differentiated to trabecular and cortical bone cells and synthesized up to 20% of all type I collagen in the host bone. The transplantation eliminated the perinatal lethality of heterozygous BrtlIV mice. At 2 months of age, femora of treated Brtl mice had significant improvement in geometric parameters (P < .05) versus untreated Brtl mice, and their mechanical properties attained wild-type values. Our results suggest that the engrafted cells form bone with higher efficiency than the endogenous cells, supporting IUT as a promising approach for the treatment of genetic bone diseases. PMID:19414862

The First Scube3 Mutant Mouse Line with Pleiotropic Phenotypic Alterations

PubMed Central

Fuchs, Helmut; Sabrautzki, Sibylle; Przemeck, Gerhard K. H.; Leuchtenberger, Stefanie; Lorenz-Depiereux, Bettina; Becker, Lore; Rathkolb, Birgit; Horsch, Marion; Garrett, Lillian; Östereicher, Manuela A.; Hans, Wolfgang; Abe, Koichiro; Sagawa, Nobuho; Rozman, Jan; Vargas-Panesso, Ingrid L.; Sandholzer, Michael; Lisse, Thomas S.; Adler, Thure; Aguilar-Pimentel, Juan Antonio; Calzada-Wack, Julia; Ehrhard, Nicole; Elvert, Ralf; Gau, Christine; Hölter, Sabine M.; Micklich, Katja; Moreth, Kristin; Prehn, Cornelia; Puk, Oliver; Racz, Ildiko; Stoeger, Claudia; Vernaleken, Alexandra; Michel, Dian; Diener, Susanne; Wieland, Thomas; Adamski, Jerzy; Bekeredjian, Raffi; Busch, Dirk H.; Favor, John; Graw, Jochen; Klingenspor, Martin; Lengger, Christoph; Maier, Holger; Neff, Frauke; Ollert, Markus; Stoeger, Tobias; Yildirim, Ali Önder; Strom, Tim M.; Zimmer, Andreas; Wolf, Eckhard; Wurst, Wolfgang; Klopstock, Thomas; Beckers, Johannes; Gailus-Durner, Valerie; Hrabé de Angelis, Martin

2016-01-01

The vertebrate Scube (Signal peptide, CUB, and EGF-like domain-containing protein) family consists of three independent members, Scube1–3, which encode secreted cell surface-associated membrane glycoproteins. Limited information about the general function of this gene family is available, and their roles during adulthood. Here, we present the first Scube3 mutant mouse line (Scube3N294K/N294K), which clearly shows phenotypic alterations by carrying a missense mutation in exon 8, and thus contributes to our understanding of SCUBE3 functions. We performed a detailed phenotypic characterization in the German Mouse Clinic (GMC). Scube3N294K/N294K mutants showed morphological abnormalities of the skeleton, alterations of parameters relevant for bone metabolism, changes in renal function, and hearing impairments. These findings correlate with characteristics of the rare metabolic bone disorder Paget disease of bone (PDB), associated with the chromosomal region of human SCUBE3. In addition, alterations in energy metabolism, behavior, and neurological functions were detected in Scube3N294K/N294K mice. The Scube3N294K/N294K mutant mouse line may serve as a new model for further studying the effect of impaired SCUBE3 gene function. PMID:27815347

Three-dimensional (3D) printing of mouse primary hepatocytes to generate 3D hepatic structure.

PubMed

Kim, Yohan; Kang, Kyojin; Jeong, Jaemin; Paik, Seung Sam; Kim, Ji Sook; Park, Su A; Kim, Wan Doo; Park, Jisun; Choi, Dongho

2017-02-01

The major problem in producing artificial livers is that primary hepatocytes cannot be cultured for many days. Recently, 3-dimensional (3D) printing technology draws attention and this technology regarded as a useful tool for current cell biology. By using the 3D bio-printing, these problems can be resolved. To generate 3D bio-printed structures (25 mm × 25 mm), cells-alginate constructs were fabricated by 3D bio-printing system. Mouse primary hepatocytes were isolated from the livers of 6-8 weeks old mice by a 2-step collagenase method. Samples of 4 × 10 7 hepatocytes with 80%-90% viability were printed with 3% alginate solution, and cultured with well-defined culture medium for primary hepatocytes. To confirm functional ability of hepatocytes cultured on 3D alginate scaffold, we conducted quantitative real-time polymerase chain reaction and immunofluorescence with hepatic marker genes. Isolated primary hepatocytes were printed with alginate. The 3D printed hepatocytes remained alive for 14 days. Gene expression levels of Albumin , HNF-4α and Foxa3 were gradually increased in the 3D structures. Immunofluorescence analysis showed that the primary hepatocytes produced hepatic-specific proteins over the same period of time. Our research indicates that 3D bio-printing technique can be used for long-term culture of primary hepatocytes. It can therefore be used for drug screening and as a potential method of producing artificial livers.

Optimizing tamoxifen-inducible Cre/loxp system to reduce tamoxifen effect on bone turnover in long bones of young mice.

PubMed

Zhong, Zhendong A; Sun, Weihua; Chen, Haiyan; Zhang, Hongliang; Lay, Yu-An E; Lane, Nancy E; Yao, Wei

2015-12-01

For tamoxifen-dependent Cre recombinase, also known as CreER recombinase, tamoxifen (TAM) is used to activate the Cre to generate time- and tissue-specific mouse mutants. TAM is a potent CreER system inducer; however, TAM is also an active selective estrogen receptor modulator (SERM) that can influence bone homeostasis. The purpose of this study was to optimize the TAM dose for Cre recombinase activation while minimizing the effects of TAM on bone turnover in young growing mice. To evaluate the effects of TAM on bone turnover and bone mass, 1-month-old wild-type male and female mice were intraperitoneally injected with TAM at 0, 1, 10 or 100mg/kg/day for four consecutive days, or 100, 300 mg/kg/day for one day. The distal femurs were analyzed one month after the last TAM injection by microCT, mechanical test, and surface-based bone histomorphometry. Similar doses of TAM were used in Col1 (2.3 kb)-CreERT2; mT/mG reporter male mice to evaluate the dose-dependent efficacy of Cre-ER activation in bone tissue. A TAM dose of 100 mg/kg × 4 days significantly increased trabecular bone volume/total volume (BV/TV) of the distal femur, femur length, bone strength, and serum bone turnover markers compared to the 0mg control group. In contrast, TAM doses ≤ 10 mg/kg did not significantly change any of these parameters compared to the 0mg group, although a higher bone strength was observed in the 10mg group. Surface-based histomorphometry revealed that the 100mg/kg dose of TAM dose significantly increased trabecular bone formation and decreased periosteal bone formation at 1-week post-TAM treatment. Using the reporter mouse model Col1-CreERT2; mT/mG, we found that 10mg/kg TAM induced Col1-CreERT2 activity in bone at a comparable level to the 100mg/kg dose. TAM treatment at 100mg/kg/day × 4 days significantly affects bone homeostasis, resulting in an anabolic bone effect on trabecular bone in 1-month-old male mice. However, a lower dose of TAM at 10 mg/kg/day × 4 days can

Mechanical Loading Attenuates Radiation-Induced Bone Loss in Bone Marrow Transplanted Mice.

PubMed

Govey, Peter M; Zhang, Yue; Donahue, Henry J

2016-01-01

Exposure of bone to ionizing radiation, as occurs during radiotherapy for some localized malignancies and blood or bone marrow cancers, as well as during space travel, incites dose-dependent bone morbidity and increased fracture risk. Rapid trabecular and endosteal bone loss reflects acutely increased osteoclastic resorption as well as decreased bone formation due to depletion of osteoprogenitors. Because of this dysregulation of bone turnover, bone's capacity to respond to a mechanical loading stimulus in the aftermath of irradiation is unknown. We employed a mouse model of total body irradiation and bone marrow transplantation simulating treatment of hematologic cancers, hypothesizing that compression loading would attenuate bone loss. Furthermore, we hypothesized that loading would upregulate donor cell presence in loaded tibias due to increased engraftment and proliferation. We lethally irradiated 16 female C57Bl/6J mice at age 16 wks with 10.75 Gy, then IV-injected 20 million GFP(+) total bone marrow cells. That same day, we initiated 3 wks compression loading (1200 cycles 5x/wk, 10 N) in the right tibia of 10 of these mice while 6 mice were irradiated, non-mechanically-loaded controls. As anticipated, before-and-after microCT scans demonstrated loss of trabecular bone (-48.2% Tb.BV/TV) and cortical thickness (-8.3%) at 3 wks following irradiation. However, loaded bones lost 31% less Tb.BV/TV and 8% less cortical thickness (both p<0.001). Loaded bones also had significant increases in trabecular thickness and tissue mineral densities from baseline. Mechanical loading did not affect donor cell engraftment. Importantly, these results demonstrate that both cortical and trabecular bone exposed to high-dose therapeutic radiation remain capable of an anabolic response to mechanical loading. These findings inform our management of bone health in cases of radiation exposure.

Intrasocket reactive soft tissue for primary closure after augmentation of extraction sites with severe bone loss before implant placement.

PubMed

Mardinger, Ofer; Chaushu, Gavriel; Ghelfan, Oded; Nissan, Joseph

2009-06-01

The normal bone resorption after tooth extraction can be significantly aggravated in the case of pre-existing severe bone loss and chronic infection. Bone augmentation procedures have been proposed, but they require adequate closure of soft tissues. We propose the use of intrasocket reactive tissue to cover extraction sites augmented by bovine bone mineral graft to promote the success of the graft procedure. The study included 24 patients with severe bone loss and chronic pathology in 27 sites. The intrasocket reactive soft tissue was elevated from the bony walls in a subperiosteal plane. Porous bovine or allograft bone mineral was placed in the extraction site without membranes, and the intrasocket reactive soft tissue was sutured over the grafting material to seal the coronal portion of the socket. Twenty-seven implants were placed 6 months after bone augmentation. Healing progressed uneventfully. Postoperative morbidity was minimal. There was no leakage or infection of the grafting material. The mean time to implant placement was 7.8 months. Supplemental augmentation was not needed. There were no implant failures. Follow-up ranged from 6 to 36 months (mean, 15 months). All implants were rehabilitated with fixed prostheses. Intrasocket reactive soft tissue can be used predictably to obtain primary closure of augmented extraction sites with severe bone loss with minimal postoperative morbidity.

Global deletion of tetraspanin CD82 attenuates bone growth and enhances bone marrow adipogenesis.

PubMed

Bergsma, Alexis; Ganguly, Sourik S; Dick, Daniel; Williams, Bart O; Miranti, Cindy K

2018-05-18

CD82 is a widely expressed member of the tetraspanin family of transmembrane proteins known to control cell signaling, adhesion, and migration. Tetraspanin CD82 is induced over 9-fold during osteoclast differentiation in vitro; however, its role in bone homeostasis is unknown. A globally deleted CD82 mouse model was used to assess the bone phenotype. Based on microCT and 4-point bending tests, CD82-deficient bones are smaller in diameter and weaker, but display no changes in bone density. Histomorphometry shows a decrease in size, erosion perimeter, and number of osteoclasts in situ, with a corresponding increase in trabecular surface area, specifically in male mice. Male-specific alterations are observed in trabecular structure by microCT and in vitro differentiated osteoclasts are morphologically abnormal. Histomorphometry did not reveal a significant reduction in osteoblast number; however, dynamic labeling reveals a significant decrease in bone growth. Consistent with defects in OB function, OB differentiation and mineralization are defective in vitro, whereas adipogenesis is enhanced. There is a corresponding increase in bone marrow adipocytes in situ. Thus, combined defects in both osteoclasts and osteoblasts can account for the observed bone phenotypes, and suggests a role for CD82 in both bone mesenchyme and myeloid cells. Copyright © 2018 Elsevier Inc. All rights reserved.

Dietary phlorizin enhances osteoblastogenic bone formation through enhancing β-catenin activity via GSK-3β inhibition in a model of senile osteoporosis.

PubMed

Antika, Lucia Dwi; Lee, Eun-Jung; Kim, Yun-Ho; Kang, Min-Kyung; Park, Sin-Hye; Kim, Dong Yeon; Oh, Hyeongjoo; Choi, Yean-Jung; Kang, Young-Hee

2017-11-01

Osteoporosis is one of the most prevalent forms of age-related bone diseases. Increased bone loss with advancing age has become a grave public health concern. This study examined whether phlorizin and phloretin, dihydrochalcones in apple peels, inhibited senile osteoporosis through enhancing osteoblastogenic bone formation in cell-based and aged mouse models. Submicromolar phloretin and phlorizin markedly stimulated osteoblast differentiation of MC3T3-E1 cells with increased transcription of Runx2 and osteocalcin. Senescence-accelerated resistant mouse strain prone-6 (SAMP6) mice were orally supplemented with 10 mg/kg phlorizin and phloretin daily for 12 weeks. Male senescence-accelerated resistant mouse strain R1 mice were employed as a nonosteoporotic age-matched control. Oral administration of ploretin and phorizin boosted bone mineralization in all the bones of femur, tibia and vertebra of SAMP6. In particular, phlorizin reduced serum RANKL/OPG ratio and diminished TRAP-positive osteoclasts in trabecular bones of SAMP6. Additionally, treating phlorizin to SAMP6 inhibited the osteoporotic resorption in distal femoral bones through up-regulating expression of BMP-2 and collagen-1 and decreasing production of matrix-degrading cathepsin K and MMP-9. Finally, phlorizin and phloretin antagonized GSK-3β induction and β-catenin phosphorylation in osteoblasts and aged mouse bones. Therefore, phlorizin and phloretin were potential therapeutic agents encumbering senile osteoporosis through promoting bone-forming osteoblastogenesis via modulation of GSK-3β/β-catenin-dependent signaling. Copyright © 2017 Elsevier Inc. All rights reserved.

Ridge preservation using a composite bone graft and a bioabsorbable membrane with and without primary wound closure: a comparative clinical trial.

PubMed

Engler-Hamm, Daniel; Cheung, Wai S; Yen, Alec; Stark, Paul C; Griffin, Terrence

2011-03-01

The aim of this single-masked, randomized controlled clinical trial is to compare hard and soft tissue changes after ridge preservation performed with (control, RPc) and without (test, RPe) primary soft tissue closure in a split-mouth design. Eleven patients completed this 6-month trial. Extraction and ridge preservation were performed using a composite bone graft of inorganic bovine-derived hydroxyapatite matrix and cell binding peptide P-15 (ABM/P-15), demineralized freeze-dried bone allograft, and a copolymer bioabsorbable membrane. Primary wound closure was achieved on the control sites (RPc), whereas test sites (RPe) left the membrane exposed. Pocket probing depth on adjacent teeth, repositioning of the mucogingival junction, bone width, bone fill, and postoperative discomfort were assessed. Bone cores were obtained for histological examination. Intragroup analyses for both groups demonstrated statistically significant mean reductions in probing depth (RPc: 0.42 mm, P = 0.012; RPe: 0.25 mm, P = 0.012) and bone width (RPc: 3 mm, P = 0.002; RPe: 3.42 mm, P <0.001). However, intergroup analysis did not find these parameters to be statistically different at 6 months. The test group showed statistically significant mean change in bone fill (7.21 mm; P <0.001). Compared to the control group, the test group showed statistically significant lower mean postoperative discomfort (RPc 4 versus RPe 2; P = 0.002). Histomorphometric analysis showed presence of 0% to 40% of ABM/P-15 and 5% to 20% of new bone formation in both groups. Comparison of clinical variables between the two groups at 6 months revealed that the mucogingival junction was statistically significantly more coronally displaced in the control group than in the test group, with a mean of 3.83 mm versus 1.21 mm (P = 0.002). Ridge preservation without flap advancement preserves more keratinized tissue and has less postoperative discomfort and swelling. Although ridge preservation is performed with either method

Atmospheric Oxygen Inhibits Growth and Differentiation of Marrow-Derived Mouse Mesenchymal Stem Cells via a p53 Dependent Mechanism: Implications for Long-Term Culture Expansion

PubMed Central

Boregowda, Siddaraju; Krishnappa, Veena; Chambers, Jeremy; LoGrasso, Phillip V.; Lai, Wen-Tzu; Ortiz, Luis A.; Phinney, Donald G.

2013-01-01

Large scale expansion of human mesenchymal stem cells (MSCs) is routinely performed for clinical therapy. In contrast, developing protocols for large scale expansion of primary mouse MSCs has been more difficult due to unique aspects of rodent biology. Currently, established methods to isolate mouse MSCs select for rapidly dividing subpopulations that emerge from bone marrow cultures following long-term (months) expansion in atmospheric oxygen. Herein, we demonstrate that exposure to atmospheric oxygen rapidly induced p53, TOP2A and BAX expression and mitochondrial ROS generation in primary mouse MSCs resulting in oxidative stress, reduced cell viability and inhibition of cell proliferation. Alternatively, procurement and culture in 5% oxygen supported more prolific expansion of the CD45−ve/CD44+ve cell fraction in marrow, produced increased MSC yields following immuno-depletion, and supported sustained MSC growth resulting in a 2300-fold increase in cumulative cell yield by 4th passage. MSCs cultured in 5% oxygen also exhibited enhanced tri-lineage differentiation. The oxygen-induced stress response was dependent upon p53 since siRNA mediated knockdown of p53 in wild type cells or exposure of p53−/− MSCs to atmospheric oxygen failed to induce ROS generation, reduce viability, or arrest cell growth. These data indicate that long-term culture expansion of mouse MSCs in atmospheric oxygen selects for clones with absent or impaired p53 function, which allows cells to escape oxygen-induced growth inhibition. In contrast, expansion in 5% oxygen generates large numbers of primary mouse MSCs that retain sensitivity to atmospheric oxygen, and therefore a functional p53 protein, even after long-term expansion in vitro. PMID:22367737

Mechanical Loading Attenuates Radiation-Induced Bone Loss in Bone Marrow Transplanted Mice

PubMed Central

Govey, Peter M.; Zhang, Yue; Donahue, Henry J.

2016-01-01

Exposure of bone to ionizing radiation, as occurs during radiotherapy for some localized malignancies and blood or bone marrow cancers, as well as during space travel, incites dose-dependent bone morbidity and increased fracture risk. Rapid trabecular and endosteal bone loss reflects acutely increased osteoclastic resorption as well as decreased bone formation due to depletion of osteoprogenitors. Because of this dysregulation of bone turnover, bone’s capacity to respond to a mechanical loading stimulus in the aftermath of irradiation is unknown. We employed a mouse model of total body irradiation and bone marrow transplantation simulating treatment of hematologic cancers, hypothesizing that compression loading would attenuate bone loss. Furthermore, we hypothesized that loading would upregulate donor cell presence in loaded tibias due to increased engraftment and proliferation. We lethally irradiated 16 female C57Bl/6J mice at age 16 wks with 10.75 Gy, then IV-injected 20 million GFP(+) total bone marrow cells. That same day, we initiated 3 wks compression loading (1200 cycles 5x/wk, 10 N) in the right tibia of 10 of these mice while 6 mice were irradiated, non-mechanically-loaded controls. As anticipated, before-and-after microCT scans demonstrated loss of trabecular bone (-48.2% Tb.BV/TV) and cortical thickness (-8.3%) at 3 wks following irradiation. However, loaded bones lost 31% less Tb.BV/TV and 8% less cortical thickness (both p<0.001). Loaded bones also had significant increases in trabecular thickness and tissue mineral densities from baseline. Mechanical loading did not affect donor cell engraftment. Importantly, these results demonstrate that both cortical and trabecular bone exposed to high-dose therapeutic radiation remain capable of an anabolic response to mechanical loading. These findings inform our management of bone health in cases of radiation exposure. PMID:27936104

Systemic Delivery of Oncolytic Adenoviruses Targeting Transforming Growth Factor-β Inhibits Established Bone Metastasis in a Prostate Cancer Mouse Model

PubMed Central

Hu, Zebin; Gupta, Janhavi; Zhang, Zhenwei; Gerseny, Helen; Berg, Arthur; Chen, Yun Ju; Zhang, Zhiling; Du, Hongyan; Brendler, Charles B.; Xiao, Xianghui; Pienta, Kenneth J.; Guise, Theresa; Lee, Chung; Stern, Paula H.; Stock, Stuart

2012-01-01

Abstract We have examined whether Ad.sTβRFc and TAd.sTβRFc, two oncolytic viruses expressing soluble transforming growth factor-β receptor II fused with human Fc (sTGFβRIIFc), can be developed to treat bone metastasis of prostate cancer. Incubation of PC-3 and DU-145 prostate tumor cells with Ad.sTβRFc and TAd.sTβRFc produced sTGFβRIIFc and viral replication; sTGFβRIIFc caused inhibition of TGF-β-mediated SMAD2 and SMAD3 phosphorylation. Ad(E1-).sTβRFc, an E1– adenovirus, produced sTGFβRIIFc but failed to replicate in tumor cells. To examine the antitumor response of adenoviral vectors, PC-3-luc cells were injected into the left heart ventricle of nude mice. On day 9, mice were subjected to whole-body bioluminescence imaging (BLI). Mice bearing hind-limb tumors were administered viral vectors via the tail vein on days 10, 13, and 17 (2.5×1010 viral particles per injection per mouse, each injection in a 0.1-ml volume), and subjected to BLI and X-ray radiography weekly until day 53. Ad.sTβRFc, TAd.sTβRFc, and Ad(E1-).sTβRFc caused significant inhibition of tumor growth; however, Ad.sTβRFc was the most effective among all the vectors. Only Ad.sTβRFc and TAd.sTβRFc inhibited tumor-induced hypercalcemia. Histomorphometric and synchrotron micro-computed tomographic analysis of isolated bones indicated that Ad.sTβRFc induced significant reduction in tumor burden, osteoclast number, and trabecular and cortical bone destruction. These studies suggest that Ad.sTβRFc and TAd.sTβRFc can be developed as potential new therapies for prostate cancer bone metastasis. PMID:22551458

Applications of transgenics in studies of bone sialoprotein.

PubMed

Zhang, Jin; Tu, Qisheng; Chen, Jake

2009-07-01

Bone sialoprotein (BSP) is a major non-collagenous protein in mineralizing connective tissues such as dentin, cementum and calcified cartilage tissues. As a member of the Small Integrin-Binding Ligand, N-linked Glycoprotein (SIBLING) gene family of glycoproteins, BSP is involved in regulating hydroxyapatite crystal formation in bones and teeth, and has long been used as a marker gene for osteogenic differentiation. In the most recent decade, new discoveries in BSP gene expression and regulation, bone remodeling, bone metastasis, and bone tissue engineering have been achieved with the help of transgenic mice. In this review, we discuss these new discoveries obtained from the literatures and from our own laboratory, which were derived from the use of transgenic mouse mutants related to BSP gene or its promoter activity.

Bone marrow endothelial progenitors augment atherosclerotic plaque regression in a mouse model of plasma lipid lowering

PubMed Central

Yao, Longbiao; Heuser-Baker, Janet; Herlea-Pana, Oana; Iida, Ryuji; Wang, Qilong; Zou, Ming-Hui; Barlic-Dicen, Jana

2012-01-01

The major event initiating atherosclerosis is hypercholesterolemia-induced disruption of vascular endothelium integrity. In settings of endothelial damage, endothelial progenitor cells (EPCs) are mobilized from bone marrow into circulation and home to sites of vascular injury where they aid endothelial regeneration. Given the beneficial effects of EPCs in vascular repair, we hypothesized that these cells play a pivotal role in atherosclerosis regression. We tested our hypothesis in the atherosclerosis-prone mouse model in which hypercholesterolemia, one of the main factors affecting EPC homeostasis, is reversible (Reversa mice). In these mice normalization of plasma lipids decreased atherosclerotic burden; however, plaque regression was incomplete. To explore whether endothelial progenitors contribute to atherosclerosis regression, bone marrow EPCs from a transgenic strain expressing green fluorescent protein under the control of endothelial cell-specific Tie2 promoter (Tie2-GFP+) were isolated. These cells were then adoptively transferred into atheroregressing Reversa recipients where they augmented plaque regression induced by reversal of hypercholesterolemia. Advanced plaque regression correlated with engraftment of Tie2-GFP+ EPCs into endothelium and resulted in an increase in atheroprotective nitric oxide and improved vascular relaxation. Similarly augmented plaque regression was also detected in regressing Reversa mice treated with the stem cell mobilizer AMD3100 which also mobilizes EPCs to peripheral blood. We conclude that correction of hypercholesterolemia in Reversa mice leads to partial plaque regression that can be augmented by AMD3100 treatment or by adoptive transfer of EPCs. This suggests that direct cell therapy or indirect progenitor cell mobilization therapy may be used in combination with statins to treat atherosclerosis. PMID:23081735

Tenascin-W inhibits proliferation and differentiation of preosteoblasts during endochondral bone formation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kimura, Hiroaki; Akiyama, Haruhiko; Nakamura, Takashi

We identified a cDNA encoding mouse Tenascin-W (TN-W) upregulated by bone morphogenetic protein (Bmp)2 in ATDC5 osteo-chondroprogenitors. In adult mice, TN-W was markedly expressed in bone. In mouse embryos, during endochondral bone formation TN-W was localized in perichondrium/periosteum, but not in trabecular and cortical bones. During bone fracture repair, cells in the newly formed perichondrium/periosteum surrounding the cartilaginous callus expressed TN-W. Furthermore, TN-W was detectable in perichondrium/periosteum of Runx2-null and Osterix-null embryos, indicating that TN-W is expressed in preosteoblasts. In CFU-F and -O cells, TN-W had no effect on initiation of osteogenesis of bone marrow cells, and in MC3T3-E1 osteoblasticmore » cells TN-W inhibited cell proliferation and Col1a1 expression. In addition, TN-W suppressed canonical Wnt signaling which stimulates osteoblastic differentiation. Our results indicate that TN-W is a novel marker of preosteoblasts in early stage of osteogenesis, and that TN-W inhibits cell proliferation and differentiation of preosteoblasts mediated by canonical Wnt signaling.« less

CD146 expression on primary nonhematopoietic bone marrow stem cells is correlated with in situ localization

PubMed Central

Tormin, Ariane; Li, Ou; Brune, Jan Claas; Walsh, Stuart; Schütz, Birgit; Ehinger, Mats; Ditzel, Nicholas; Kassem, Moustapha

2011-01-01

Nonhematopoietic bone marrow mesenchymal stem cells (BM-MSCs) are of central importance for bone marrow stroma and the hematopoietic environment. However, the exact phenotype and anatomical distribution of specified MSC populations in the marrow are unknown. We characterized the phenotype of primary human BM-MSCs and found that all assayable colony-forming units-fibroblast (CFU-Fs) were highly and exclusively enriched not only in the lin−/CD271+/CD45−/CD146+ stem-cell fraction, but also in lin−/CD271+/CD45−/CD146−/low cells. Both populations, regardless of CD146 expression, shared a similar phenotype and genotype, gave rise to typical cultured stromal cells, and formed bone and hematopoietic stroma in vivo. Interestingly, CD146 was up-regulated in normoxia and down-regulated in hypoxia. This was correlated with in situ localization differences, with CD146 coexpressing reticular cells located in perivascular regions, whereas bone-lining MSCs expressed CD271 alone. In both regions, CD34+ hematopoietic stem/progenitor cells were located in close proximity to MSCs. These novel findings show that the expression of CD146 differentiates between perivascular versus endosteal localization of non-hematopoietic BM-MSC populations, which may be useful for the study of the hematopoietic environment. PMID:21415267

Histomorphometric Parameters of the Growth Plate and Trabecular Bone in Wild-Type and Trefoil Factor Family 3 (Tff3)-Deficient Mice Analyzed by Free and Open-Source Image Processing Software.

PubMed

Bijelić, Nikola; Belovari, Tatjana; Stolnik, Dunja; Lovrić, Ivana; Baus Lončar, Mirela

2017-08-01

Trefoil factor family 3 (Tff3) peptide is present during intrauterine endochondral ossification in mice, and its deficiency affects cancellous bone quality in secondary ossification centers of mouse tibiae. The aim of this study was to quantitatively analyze parameters describing the growth plate and primary ossification centers in tibiae of 1-month-old wild-type and Tff3 knock-out mice (n=5 per genotype) by using free and open-source software. Digital photographs of the growth plates and trabecular bone were processed by open-source computer programs GIMP and FIJI. Histomorphometric parameters were calculated using measurements made with FIJI. Tff3 knock-out mice had significantly smaller trabecular number and significantly larger trabecular separation. Trabecular bone volume, trabecular bone surface, and trabecular thickness showed no significant difference between the two groups. Although such histomorphological differences were found in the cancellous bone structure, no significant differences were found in the epiphyseal plate histomorphology. Tff3 peptide probably has an effect on the formation and quality of the cancellous bone in the primary ossification centers, but not through disrupting the epiphyseal plate morphology. This work emphasizes the benefits of using free and open-source programs for morphological studies in life sciences.

Cytology of Bone.

PubMed

Barger, Anne M

2017-01-01

Cytology of bone is a useful diagnostic tool. Aspiration of lytic or proliferative lesions can assist with the diagnosis of inflammatory or neoplastic processes. Bacterial, fungal, and protozoal organisms can result in significant osteomyelitis, and these organisms can be identified on cytology. Neoplasms of bone including primary bone tumors such as osteosarcoma, chondrosarcoma, fibrosarcoma, synovial cell sarcoma, and histiocytic sarcoma and tumors of bone marrow including plasma cell neoplasia and lymphoma and metastatic neoplasia can result in significant bone lysis or proliferation and can be diagnosed effectively with cytology. Copyright © 2016 Elsevier Inc. All rights reserved.

Role of FGF/FGFR signaling in skeletal development and homeostasis: learning from mouse models

PubMed Central

Su, Nan; Jin, Min; Chen, Lin

2014-01-01

Fibroblast growth factor (FGF)/fibroblast growth factor receptor (FGFR) signaling plays essential roles in bone development and diseases. Missense mutations in FGFs and FGFRs in humans can cause various congenital bone diseases, including chondrodysplasia syndromes, craniosynostosis syndromes and syndromes with dysregulated phosphate metabolism. FGF/FGFR signaling is also an important pathway involved in the maintenance of adult bone homeostasis. Multiple kinds of mouse models, mimicking human skeleton diseases caused by missense mutations in FGFs and FGFRs, have been established by knock-in/out and transgenic technologies. These genetically modified mice provide good models for studying the role of FGF/FGFR signaling in skeleton development and homeostasis. In this review, we summarize the mouse models of FGF signaling-related skeleton diseases and recent progresses regarding the molecular mechanisms, underlying the role of FGFs/FGFRs in the regulation of bone development and homeostasis. This review also provides a perspective view on future works to explore the roles of FGF signaling in skeletal development and homeostasis. PMID:26273516

Self-perception and quality of life in adolescents during treatment for a primary malignant bone tumour.

PubMed

van Riel, Christel A H P; Meijer-van den Bergh, Esther E M; Kemps, Hennie L M; Feuth, Ton; Schreuder, Hendrik W B; Hoogerbrugge, Peter M; De Groot, Imelda J M; Mavinkurve-Groothuis, Annelies M C

2014-06-01

Adolescents experience physical and psychosocial changes as part of their normal development. It can be hypothesized that they have lower scores on Quality of Life (QoL) and self-perception when additional changes occur due to cancer treatment. The purpose of our study was to assess self-perception and QoL of adolescents during or up to three months after adjuvant treatment for a primary malignant bone tumour. Ten adolescent patients (median age of 15 years) were included. Every patient was matched with two healthy peers. Participants completed the dutch version of the Self Perception Profile of Adolescents (SPPA) to measure self-perception and the KIDSCREEN-52 questionnaire for QoL. For both instruments, normative data were available. Adolescents with a bone tumour had consistently lower scores on QoL as compared to healthy peers. Significantly on domains: physical well-being (P < 0.002), autonomy (P = 0.02), social support (P = 0.04) and school environment (P = 0.02). Scores on self-perception in this group were similar in both the study and control group. Adolescents with a primary malignant bone tumour during or up to three months after adjuvant treatment had lower scores on QoL (KIDSCREEN-52), significantly on domains of physical well-being and social functioning. Unlike most other quality of life instruments, the KIDSCREEN-52 contains different areas of social functioning and has shown to be a useful instrument in our patient group. Scores on self-perception in this group were similar in both study and control group. Copyright © 2014 Elsevier Ltd. All rights reserved.

Rethinking the nature of fibrolamellar bone: an integrative biological revision of sauropod plexiform bone formation.

PubMed

Stein, Koen; Prondvai, Edina

2014-02-01

We present novel findings on sauropod bone histology that cast doubt on general palaeohistological concepts concerning the true nature of woven bone in primary cortical bone and its role in the rapid growth and giant body sizes of sauropod dinosaurs. By preparing and investigating longitudinal thin sections of sauropod long bones, of which transverse thin sections were published previously, we found that the amount of woven bone in the primary complex has been largely overestimated. Using comparative cellular and light-extinction characteristics in the two section planes, we revealed that the majority of the bony lamina consists of longitudinally organized primary bone, whereas woven bone is usually represented only by a layer a few cells thin in the laminae. Previous arguments on sauropod biology, which have been based on the overestimated amount, misinterpreted formation process and misjudged role of woven bone in the plexiform bone formation of sauropod dinosaurs, are thereby rejected. To explain the observed pattern in fossil bones, we review the most recent advances in bone biology concerning bone formation processes at the cellular and tissue levels. Differentiation between static and dynamic osteogenesis (SO and DO) and the revealed characteristics of SO- versus DO-derived bone tissues shed light on several questions raised by our palaeohistological results and permit identification of these bone tissues in fossils with high confidence. By presenting the methods generally used for investigating fossil bones, we show that the major cause of overestimation of the amount of woven bone in previous palaeohistological studies is the almost exclusive usage of transverse sections. In these sections, cells and crystallites of the longitudinally organized primary bone are cut transversely, thus cells appear rounded and crystallites remain dark under crossed plane polarizers, thereby giving the false impression of woven bone. In order to avoid further confusion in

In vivo analysis of biocompatibility and vascularization of the synthetic bone grafting substitute NanoBone.

PubMed

Abshagen, K; Schrodi, I; Gerber, T; Vollmar, B

2009-11-01

One of the major challenges in the application of bone substitutes is adequate vascularization and biocompatibility of the implant. Thus, the temporal course of neovascularization and the microvascular inflammatory response of implants of NanoBone (fully synthetic nanocrystalline bone grafting material) were studied in vivo by using the mouse dorsal skinfold chamber model. Angiogenesis, microhemodynamics, and leukocyte-endothelial cell interaction were analyzed repetitively after implantation in the center and in the border zone of the implant up to 15 days. Both NanoBone granules and plates exhibited high biocompatibility comparable to that of cancellous bone, as indicated by a lack of venular leukocyte activation after implantation. In both synthetic NanoBone groups, signs of angiogenesis could be observed even at day 5 after implantation, whereas granules showed higher functional vessel density compared with NanoBone plates. The angiogenic response of the cancellous bone was markedly accelerated in the center of the implant tissue. Histologically, implant tissue showed an ingrowth of vascularized fibrous tissue into the material combined with an increased number of foreign-body giant cells. In conclusion, NanoBone, particularly in granular form, showed high biocompatibility and high angiogenic response, thus improving the healing of bone defects. Our results underline that, beside the composition and nanostructure, the macrostructure is also of importance for the incorporation of the biomaterial by the host tissue. (c) 2008 Wiley Periodicals, Inc.

Bone tissue engineering: a review in bone biomimetics and drug delivery strategies.

PubMed

Porter, Joshua R; Ruckh, Timothy T; Popat, Ketul C

2009-01-01

Critical-sized defects in bone, whether induced by primary tumor resection, trauma, or selective surgery have in many cases presented insurmountable challenges to the current gold standard treatment for bone repair. The primary purpose of a tissue-engineered scaffold is to use engineering principles to incite and promote the natural healing process of bone which does not occur in critical-sized defects. A synthetic bone scaffold must be biocompatible, biodegradable to allow native tissue integration, and mimic the multidimensional hierarchical structure of native bone. In addition to being physically and chemically biomimetic, an ideal scaffold is capable of eluting bioactive molecules (e.g., BMPs, TGF-betas, etc., to accelerate extracellular matrix production and tissue integration) or drugs (e.g., antibiotics, cisplatin, etc., to prevent undesired biological response such as sepsis or cancer recurrence) in a temporally and spatially controlled manner. Various biomaterials including ceramics, metals, polymers, and composites have been investigated for their potential as bone scaffold materials. However, due to their tunable physiochemical properties, biocompatibility, and controllable biodegradability, polymers have emerged as the principal material in bone tissue engineering. This article briefly reviews the physiological and anatomical characteristics of native bone, describes key technologies in mimicking the physical and chemical environment of bone using synthetic materials, and provides an overview of local drug delivery as it pertains to bone tissue engineering is included. (c) 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009.

Osteoprotective effects of osthole in a mouse model of 5/6 nephrectomy through inhibiting osteoclast formation.

PubMed

Li, Xiaofeng; Xue, Chunchun; Wang, Libo; Tang, Dezhi; Huang, Jian; Zhao, Yongjian; Chen, Yan; Zhao, Dongfeng; Shi, Qi; Wang, Yongjun; Shu, Bing

2016-10-01

The present study aimed to investigate the effects of osthole on osteoclast formation and bone loss in a mouse model of 5/6 nephrectomy. The mice in control and osthole groups were treated 1 month following 5/6 nephrectomy with either a placebo or osthole, respectively. At 2 months post‑nephrectomy, the L4 vertebrae were harvested. The bone mineral density (BMD) of cancellous bone was measured using micro‑CT and tartrate‑resistant acid phosphatase (TRAP) staining was performed to evaluate osteoclast formation. Immunohistochemistry staining and reverse transcription‑quantitative polymerase chain reaction were performed to detect the expression of nuclear factor of activated T‑cells, cytoplasmic‑1 (NFATc‑1), c‑Fos, cathepsin K, Trap, matrix metalloproteinase 9 (Mmp9), osteoprotegerin (Opg) and receptor activator for nuclear factor‑κB ligand (Rankl). Bone marrow cells were cultured with osthole, and osteoclast formation was shown by TRAP staining. Primary calvaria osteoblasts were cultured with osthole, and expression levels of Opg and Rankl were detected. Compared with the sham group, the BMD of mice in model group was significantly reduced. The numbers of osteoclasts and the expression levels of NFATc‑1, c‑Fos, cathepsin K and Mmp9 were significantly increased. Compared with the control group, the mice in the osthole group exhibited increased BMD of the L4 vertebrae, a reduction in osteoclast numbers and decreased expression levels of NFATc‑1, c‑Fos, cathepsin K and Mmp9. In vitro experiments also showed that osteoclast formation was decreased following treatment with osthole. Osteoprotegerin (Opg)/receptor activator for nuclear factor‑κB ligand (Rankl) was upregulated by osthole treatment in the L4 vertebrae and in primary cultures of calvarial osteoblasts. Osthole inhibited osteoclast formation and partially reversed the bone loss induced by 5/6 nephrectomy in mice through the upregulation of OPG/RANKL.

Metastases to bones.

PubMed

Wick, Mark R

2014-01-01

Metastatic tumors involving the bones may derive from a number of visceral primary sites, and they can assume several histological appearances. In selected instances, diagnostic confusion with some primary bone tumors may eventuate, necessitating the use of adjunctive pathologic studies to reach a final interpretation. This review considers metastatic osseous neoplasms in the small-cell, large-polygonal-cell, and spindle-cell-pleomorphic microscopic categories. The use of immunohistology and molecular analysis to study such tumors is discussed. Copyright © 2014 Elsevier Inc. All rights reserved.

Primary versus revision arthroscopic reconstruction with remplissage for shoulder instability with moderate bone loss.

PubMed

McCabe, Michael P; Weinberg, Douglas; Field, Larry D; O'Brien, Michael J; Hobgood, E Rhett; Savoie, Felix H

2014-04-01

This study aims to evaluate our outcomes of arthroscopic remplissage in this setting. A retrospective review was performed to identify patients who underwent arthroscopic remplissage of an engaging Hill-Sachs lesion along with anterior capsulolabral reconstruction for anterior glenohumeral instability with moderate glenohumeral bone loss at our institution. Thirty-five patients, with a minimum of 2 years' follow-up, were identified. We assessed the American Shoulder and Elbow Surgeons score, incidence of recurrent instability, and postoperative Rowe instability score. Follow-up was available for 30 patients (31 shoulders). The mean age was 24.6 years, with a mean follow-up period of 41 months. Prior instability surgery had failed in 11 patients, and they underwent capsulolabral reconstruction and remplissage ("revision surgery"). The failure rate in revision cases (36%) was significantly higher than the failure rate in primary surgery cases (0%) (P = .01). Failure resulted from trauma in all 4 patients, and none required further surgery. The mean American Shoulder and Elbow Surgeons score for all patients improved from 50 preoperatively to 91 postoperatively (P < .001), with no significant postoperative difference between primary and revision patients (P = .13). The patients with clinical failure showed nonsignificant improvement from 41 preoperatively to 72 postoperatively (P = .08). The mean postoperative Rowe score for the entire cohort was 90. The Rowe score was significantly lower in the 4 cases of failure than in the 27 non-failure cases (51 v 96, P < .001). In our experience, aggressive capsulolabral reconstruction with remplissage in traumatic instability patients with moderate bone loss and engaging humeral Hill-Sachs lesions yields acceptable outcomes for primary instability surgery. However, a significantly higher failure rate occurred when arthroscopic reconstruction with remplissage was performed in the revision setting. Level IV, therapeutic

Targeting skeletal endothelium to ameliorate bone loss.

PubMed

Xu, Ren; Yallowitz, Alisha; Qin, An; Wu, Zhuhao; Shin, Dong Yeon; Kim, Jung-Min; Debnath, Shawon; Ji, Gang; Bostrom, Mathias P; Yang, Xu; Zhang, Chao; Dong, Han; Kermani, Pouneh; Lalani, Sarfaraz; Li, Na; Liu, Yifang; Poulos, Michael G; Wach, Amanda; Zhang, Yi; Inoue, Kazuki; Di Lorenzo, Annarita; Zhao, Baohong; Butler, Jason M; Shim, Jae-Hyuck; Glimcher, Laurie H; Greenblatt, Matthew B

2018-06-01

Recent studies have identified a specialized subset of CD31 hi endomucin hi (CD31 hi EMCN hi ) vascular endothelium that positively regulates bone formation. However, it remains unclear how CD31 hi EMCN hi endothelium levels are coupled to anabolic bone formation. Mice with an osteoblast-specific deletion of Shn3, which have markedly elevated bone formation, demonstrated an increase in CD31 hi EMCN hi endothelium. Transcriptomic analysis identified SLIT3 as an osteoblast-derived, SHN3-regulated proangiogenic factor. Genetic deletion of Slit3 reduced skeletal CD31 hi EMCN hi endothelium, resulted in low bone mass because of impaired bone formation and partially reversed the high bone mass phenotype of Shn3 -/- mice. This coupling between osteoblasts and CD31 hi EMCN hi endothelium is essential for bone healing, as shown by defective fracture repair in SLIT3-mutant mice and enhanced fracture repair in SHN3-mutant mice. Finally, administration of recombinant SLIT3 both enhanced bone fracture healing and counteracted bone loss in a mouse model of postmenopausal osteoporosis. Thus, drugs that target the SLIT3 pathway may represent a new approach for vascular-targeted osteoanabolic therapy to treat bone loss.

Effect of sclerostin antibody treatment in a mouse model of severe osteogenesis imperfecta.

PubMed

Roschger, Andreas; Roschger, Paul; Keplingter, Petra; Klaushofer, Klaus; Abdullah, Sami; Kneissel, Michaela; Rauch, Frank

2014-09-01

Osteogenesis imperfecta (OI) is a heritable bone fragility disorder that is usually caused by mutations affecting collagen type I production in osteoblasts. Stimulation of bone formation through sclerostin antibody treatment (Sost-ab) has shown promising results in mouse models of relatively mild OI. We assessed the effect of once-weekly intravenous Sost-ab injections for 4weeks in male Col1a1(Jrt)/+mice, a model of severe dominant OI, starting either at 4weeks (growing mice) or at 20weeks (adult mice) of age. Sost-ab had no effect on weight or femur length. In OI mice, no significant treatment-associated differences in serum markers of bone formation (alkaline phosphatase activity, procollagen type I N-propeptide) or resorption (C-telopeptide of collagen type I) were found. Micro-CT analyses at the femur showed that Sost-ab treatment was associated with higher trabecular bone volume and higher cortical thickness in wild type mice at both ages and in growing OI mice, but not in adult OI mice. Three-point bending tests of the femur showed that in wild type but not in OI mice, Sost-ab was associated with higher ultimate load and work to failure. Quantitative backscattered electron imaging of the femur did not show any effect of Sost-ab on CaPeak (the most frequently occurring calcium concentration in the bone mineral density distribution), regardless of genotype, age or measurement location. Thus, Sost-ab had a larger effect in wild type than in Col1a1(Jrt)/+mice. Previous studies had found marked improvements of Sost-ab on bone mass and strength in an OI mouse model with a milder phenotype. Our data therefore suggest that Sost-ab is less effective in a more severely affected OI mouse model. Copyright © 2014 Elsevier Inc. All rights reserved.

INF-γ encoding plasmid administration triggers bone loss and disrupts bone marrow microenvironment.

PubMed

Agas, Dimitrios; Gusmão Silva, Guilherme; Laus, Fulvio; Marchegiani, Andrea; Capitani, Melania; Vullo, Cecilia; Catone, Giuseppe; Lacava, Giovanna; Concetti, Antonio; Marchetti, Luigi; Sabbieti, Maria Giovanna

2017-02-01

IFN-γ is a pleotropic cytokine produced in the bone microenvironment. Although IFN-γ is known to play a critical role on bone remodeling, its function is not fully elucidated. Consistently, outcomes on the effects of IFN-γ recombinant protein on bone loss are contradictory among reports. In our work we explored, for the first time, the role of IFN-γ encoding plasmid (pIFN-γ) in a mouse model of osteopenia induced by ovariectomy and in the sham-operated counterpart to estimate its effects in skeletal homeostasis. Ovariectomy produced a dramatic decrease of bone mineral density (BMD). pINF-γ injected mice showed a pathologic bone and bone marrow phenotype; the disrupted cortical and trabecular bone microarchitecture was accompanied by an increased release of pro-inflammatory cytokine by bone marrow cells. Moreover, mesenchymal stem cells' (MSCs) commitment to osteoblast was found impaired, as evidenced by the decline of osterix-positive (Osx + ) cells within the mid-diaphyseal area of femurs. For instance, a reduction and redistribution of CXCL12 cells have been found, in accordance with bone marrow morphological alterations. As similar effects were observed both in sham-operated and in ovariectomized mice, our studies proved that an increased IFN-γ synthesis in bone marrow might be sufficient to induce inflammatory and catabolic responses even in the absence of pathologic predisposing substrates. In addition, the obtained data might raise questions about pIFN-γ's safety when it is used as vaccine adjuvant. © 2017 Society for Endocrinology.

Evolutionary Patterns of Bone Histology and Bone Compactness in Xenarthran Mammal Long Bones

PubMed Central

Straehl, Fiona R.; Scheyer, Torsten M.; Forasiepi, Analía M.; MacPhee, Ross D.; Sánchez-Villagra, Marcelo R.

2013-01-01

Bone microstructure reflects physiological characteristics and has been shown to contain phylogenetic and ecological signals. Although mammalian long bone histology is receiving increasing attention, systematic examination of the main clades has not yet been performed. Here we describe the long bone microstructure of Xenarthra based on thin sections representing twenty-two species. Additionally, patterns in bone compactness of humeri and femora are investigated. The primary bone tissue of xenarthran long bones is composed of a mixture of woven, parallel-fibered and lamellar bone. The vascular canals have a longitudinal, reticular or radial orientation and are mostly arranged in an irregular manner. Concentric rows of vascular canals and laminar organization of the tissue are only found in anteater bones. The long bones of adult specimens are marked by dense Haversian bone, a feature that has been noted for most groups of mammals. In the long bones of armadillos, secondary osteons have an oblique orientation within the three-dimensional bone tissue, thus resulting in their irregular shape when the bones are sectioned transversely. Secondary remodeling is generally more extensive in large taxa than in small taxa, and this could be caused by increased loading. Lines of arrested growth are assumed to be present in all specimens, but they are restricted to the outermost layer in bones of armadillos and are often masked by secondary remodeling in large taxa. Parameters of bone compactness show a pattern in the femur that separates Cingulata and Pilosa (Folivora and Vermilingua), with cingulates having a lower compactness than pilosans. In addition, cingulates show an allometric relationship between humeral and femoral bone compactness. PMID:23874932

Evolutionary patterns of bone histology and bone compactness in xenarthran mammal long bones.

PubMed

Straehl, Fiona R; Scheyer, Torsten M; Forasiepi, Analía M; MacPhee, Ross D; Sánchez-Villagra, Marcelo R

2013-01-01

Bone microstructure reflects physiological characteristics and has been shown to contain phylogenetic and ecological signals. Although mammalian long bone histology is receiving increasing attention, systematic examination of the main clades has not yet been performed. Here we describe the long bone microstructure of Xenarthra based on thin sections representing twenty-two species. Additionally, patterns in bone compactness of humeri and femora are investigated. The primary bone tissue of xenarthran long bones is composed of a mixture of woven, parallel-fibered and lamellar bone. The vascular canals have a longitudinal, reticular or radial orientation and are mostly arranged in an irregular manner. Concentric rows of vascular canals and laminar organization of the tissue are only found in anteater bones. The long bones of adult specimens are marked by dense Haversian bone, a feature that has been noted for most groups of mammals. In the long bones of armadillos, secondary osteons have an oblique orientation within the three-dimensional bone tissue, thus resulting in their irregular shape when the bones are sectioned transversely. Secondary remodeling is generally more extensive in large taxa than in small taxa, and this could be caused by increased loading. Lines of arrested growth are assumed to be present in all specimens, but they are restricted to the outermost layer in bones of armadillos and are often masked by secondary remodeling in large taxa. Parameters of bone compactness show a pattern in the femur that separates Cingulata and Pilosa (Folivora and Vermilingua), with cingulates having a lower compactness than pilosans. In addition, cingulates show an allometric relationship between humeral and femoral bone compactness.

Excess TGF-β mediates muscle weakness associated with bone metastases in mice

PubMed Central

Reiken, Steven; Xie, Wenjun; Andersson, Daniel C.; John, Sutha; Chiechi, Antonella; Wright, Laura E.; Umanskaya, Alisa; Niewolna, Maria; Trivedi, Trupti; Charkhzarrin, Sahba; Khatiwada, Pooja; Wronska, Anetta; Haynes, Ashley; Benassi, Maria Serena; Witzmann, Frank A.; Zhen, Gehua; Wang, Xiao; Cao, Xu; Roodman, G. David; Marks, Andrew R.; Guise, Theresa A.

2015-01-01

Cancer-associated muscle weakness is poorly understood and there is no effective treatment. Here, we find that seven different mouse models of human osteolytic bone metastases, representing breast, lung and prostate cancers, as well as multiple myeloma exhibited impaired muscle function, implicating a role for the tumor-bone microenvironment in cancer-associated muscle weakness. We found that TGF-β, released from the bone surface as a result of metastasis-induced bone destruction upregulated NADPH oxidase 4 (Nox4), resulting in elevated oxidization of skeletal muscle proteins, including the ryanodine receptor/calcium (Ca2+) release channel (RyR1). The oxidized RyR1 channels leaked Ca2+, resulting in lower intracellular signaling required for proper muscle contraction. We found that inhibiting RyR1 leak, TGF-β signaling, TGF-β release from bone or Nox4 all improved muscle function in mice with MDA-MB-231 bone metastases. Humans with breast cancer- or lung cancer-associated bone metastases also had oxidized skeletal muscle RyR1 that is not seen in normal muscle. Similarly, skeletal muscle weakness, higher levels of Nox4 protein and Nox4 binding to RyR1, and oxidation of RyR1 were present in a mouse model of Camurati-Engelmann disease, a non-malignant metabolic bone disorder associated with increased TGF-β activity. Thus, metastasis-induced TGF-β release from bone contributes to muscle weakness by decreasing Ca2+-induced muscle force production. PMID:26457758

Applications of Transgenics in Studies of Bone Sialoprotein

PubMed Central

Zhang, Jin; Tu, Qisheng; Chen, Jake

2010-01-01

Bone sialoprotein (BSP) is a major non-collagenous protein in mineralizing connective tissues such as dentin, cementum and calcified cartilage tissues. As a member of the SIBLING (Small Integrin-Binding Ligand, N-linked Glycoprotein) gene family of glycoproteins, BSP is involved in regulating hydroxyapatite crystal formation in bones and teeth, and has long been used as a marker gene for osteogenic differentiation. In the most recent decade, new discoveries in BSP gene expression and regulation, bone remodeling, bone metastasis, and bone tissue engineering have been achieved with the help of transgenic mice. In this review, we discuss these new discoveries obtained from the literatures and from our own laboratory, which were derived from the use of transgenic mouse mutants related to BSP gene or its promoter activity. PMID:19326395

Rat astrocytes are more supportive for mouse OPC self-renewal than mouse astrocytes in culture.

PubMed

Cheng, Xuejun; Xie, Binghua; Qi, Jiajun; Zhao, Xiaofeng; Zhang, Zunyi; Qiu, Mengsheng; Yang, Junlin

2017-09-01

Mouse primary oligodendrocyte precursor cells (OPCs) are increasingly used to study the molecular mechanisms underlying the phenotype changes in oligodendrocyte differentiation and axonal myelination observed in transgenic or mutant mouse models. However, mouse OPCs are much more difficult to be isolated by the simple dissociation culture of brain tissues than their rat counterparts. To date, the mechanisms underlying the species difference in OPC preparation remain obscure. In this study, we showed that astrocytes from rats have a stronger effect than those from mouse in promoting OPC proliferation and survival in vitro. Mouse astrocytes displayed significantly weaker viability in culture and reduced potential in maintaining OPC self-renewal, as confirmed by culturing OPCs with conditioned media from rat or mouse astrocytes. These results explained the reason for why stratified cultures of OPCs and astrocytes are difficult to be achieved in mouse CNS tissues. Based on these findings, we adopted inactivated rat astrocytes as feeder cells to support the self-renewal of mouse cortical OPCs and preparation of high-purity mouse OPCs. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 907-916, 2017. © 2016 Wiley Periodicals, Inc.

Chimeric Mouse model to track the migration of bone marrow derived cells in glioblastoma following anti-angiogenic treatments.

PubMed

Achyut, B R; Shankar, Adarsh; Iskander, A S M; Ara, Roxan; Knight, Robert A; Scicli, Alfonso G; Arbab, Ali S

2016-01-01

Bone marrow derived cells (BMDCs) have been shown to contribute in the tumor development. In vivo animal models to investigate the role of BMDCs in tumor development are poorly explored. We established a novel chimeric mouse model using as low as 5 × 10(6) GFP+ BM cells in athymic nude mice, which resulted in >70% engraftment within 14 d. In addition, chimera was established in NOD-SCID mice, which displayed >70% with in 28 d. Since anti-angiogenic therapies (AAT) were used as an adjuvant against VEGF-VEGFR pathway to normalize blood vessels in glioblastoma (GBM), which resulted into marked hypoxia and recruited BMDCs to the tumor microenvironment (TME). We exploited chimeric mice in athymic nude background to develop orthotopic U251 tumor and tested receptor tyrosine kinase inhibitors and CXCR4 antagonist against GBM. We were able to track GFP+ BMDCs in the tumor brain using highly sensitive multispectral optical imaging instrument. Increased tumor growth associated with the infiltration of GFP+ BMDCs acquiring suppressive myeloid and endothelial phenotypes was seen in TME following treatments. Immunofluorescence study showed GFP+ cells accumulated at the site of VEGF, SDF1 and PDGF expression, and at the periphery of the tumors following treatments. In conclusion, we developed a preclinical chimeric model of GBM and phenotypes of tumor infiltrated BMDCs were investigated in context of AATs. Chimeric mouse model could be used to study detailed cellular and molecular mechanisms of interaction of BMDCs and TME in cancer.

Transcriptional profile of fibroblasts obtained from the primary site, lymph node and bone marrow of breast cancer patients.

PubMed

Del Valle, Paulo Roberto; Milani, Cintia; Brentani, Maria Mitzi; Katayama, Maria Lucia Hirata; de Lyra, Eduardo Carneiro; Carraro, Dirce Maria; Brentani, Helena; Puga, Renato; Lima, Leandro A; Rozenchan, Patricia Bortman; Nunes, Bárbara Dos Santos; Góes, João Carlos Guedes Sampaio; Azevedo Koike Folgueira, Maria Aparecida

2014-09-01

Cancer-associated fibroblasts (CAF) influence tumor development at primary as well as in metastatic sites, but there have been no direct comparisons of the transcriptional profiles of stromal cells from different tumor sites. In this study, we used customized cDNA microarrays to compare the gene expression profile of stromal cells from primary tumor (CAF, n = 4), lymph node metastasis (N+, n = 3) and bone marrow (BM, n = 4) obtained from breast cancer patients. Biological validation was done in another 16 samples by RT-qPCR. Differences between CAF vs N+, CAF vs BM and N+ vs BM were represented by 20, 235 and 245 genes, respectively (SAM test, FDR < 0.01). Functional analysis revealed that genes related to development and morphogenesis were overrepresented. In a biological validation set, NOTCH2 was confirmed to be more expressed in N+ (vs CAF) and ADCY2, HECTD1, HNMT, LOX, MACF1, SLC1A3 and USP16 more expressed in BM (vs CAF). Only small differences were observed in the transcriptional profiles of fibroblasts from the primary tumor and lymph node of breast cancer patients, whereas greater differences were observed between bone marrow stromal cells and the other two sites. These differences may reflect the activities of distinct differentiation programs.

Transcriptional profile of fibroblasts obtained from the primary site, lymph node and bone marrow of breast cancer patients

PubMed Central

Del Valle, Paulo Roberto; Milani, Cintia; Brentani, Maria Mitzi; Katayama, Maria Lucia Hirata; de Lyra, Eduardo Carneiro; Carraro, Dirce Maria; Brentani, Helena; Puga, Renato; Lima, Leandro A.; Rozenchan, Patricia Bortman; Nunes, Bárbara dos Santos; Góes, João Carlos Guedes Sampaio; Azevedo Koike Folgueira, Maria Aparecida

2014-01-01

Cancer-associated fibroblasts (CAF) influence tumor development at primary as well as in metastatic sites, but there have been no direct comparisons of the transcriptional profiles of stromal cells from different tumor sites. In this study, we used customized cDNA microarrays to compare the gene expression profile of stromal cells from primary tumor (CAF, n = 4), lymph node metastasis (N+, n = 3) and bone marrow (BM, n = 4) obtained from breast cancer patients. Biological validation was done in another 16 samples by RT-qPCR. Differences between CAF vs N+, CAF vs BM and N+ vs BM were represented by 20, 235 and 245 genes, respectively (SAM test, FDR < 0.01). Functional analysis revealed that genes related to development and morphogenesis were overrepresented. In a biological validation set, NOTCH2 was confirmed to be more expressed in N+ (vs CAF) and ADCY2, HECTD1, HNMT, LOX, MACF1, SLC1A3 and USP16 more expressed in BM (vs CAF). Only small differences were observed in the transcriptional profiles of fibroblasts from the primary tumor and lymph node of breast cancer patients, whereas greater differences were observed between bone marrow stromal cells and the other two sites. These differences may reflect the activities of distinct differentiation programs. PMID:25249769

Interaction of primary mast cells with Borrelia burgdorferi (sensu stricto): role in transmission and dissemination in C57BL/6 mice.

PubMed

Bernard, Quentin; Wang, Zhenping; Di Nardo, Anna; Boulanger, Nathalie

2017-06-27

Borrelia burgdorferi (sensu lato), the causative agent of Lyme borreliosis is a bacterium transmitted by hard ticks, Ixodes spp. Bacteria are injected into the host skin during the tick blood meal with tick saliva. There, Borrelia and saliva interact together with skin cells such as keratinocytes, fibroblasts, mast cells and other specific immune cells before disseminating to target organs. To study the role of mast cells in the transmission of Lyme borreliosis, we isolated mouse primary mast cells from bone marrow and incubated them in the presence of Borrelia burgdorferi (sensu stricto) and tick salivary gland extract. We further analyzed their potential role in vivo, in a mouse model of deficient in mast cells (Kit wsh-/- mice). To our knowledge, we report here for the first time the bacteria ability to induce the inflammatory response of mouse primary mast cells. We show that OspC, a major surface lipoprotein involved in the early transmission of Borrelia, induces the degranulation of primary mast cells but has a limited effect on the overall inflammatory response of these cells. In contrast, whole bacteria have an opposite effect. We also show that mast cell activation is significantly inhibited by tick salivary gland extract. Finally, we demonstrate that mast cells are likely not the only host cells involved in the early transmission and dissemination of Borrelia since the use of mast cell deficient Kit wsh-/- mice shows a limited impact on these two processes in the context of this mouse genetic background. The absence of mast cells did not change the replication rate of Borrelia in the skin. However, in the absence of mast cells, Borrelia dissemination to the joints was faster. Mast cells do not control skin bacterial proliferation during primary infection and the establishment of the primary infection, as shown in the C57BL/6 mouse model studied. Nevertheless, the Borrelia induced cytotokine modulation on mast cells might be involved in long term and

Effects of major histocompatibility complex class II knockout on mouse bone mechanical properties during development

NASA Technical Reports Server (NTRS)

Simske, Steven J.; Bateman, Ted A.; Smith, Erin E.; Ferguson, Virginia L.; Chapes, Stephen K.

2002-01-01

We investigated the effect of major histocompatibility complex class II (MHC II) knockout on the development of the mouse peripheral skeleton. These C2D mice had less skeletal development at 8, 12 and 16 weeks of age compared to wild-type C57BL/6J (B6) male mice. The C2D mice had decreased femur mechanical, geometric and compositional measurements compared to wild type mice at each of these ages. C2D femur stiffness (S), peak force in 3-pt bending (Pm), and mineral mass (Min-M) were 74%, 64% and 66%, respectively, of corresponding B6 values at 8 weeks of age. Similar differences were measured at 12 weeks (for which C2D femoral S, Pm and Min-M were 71%, 72% and 73%, respectively, of corresponding B6 values) and at 16 weeks (for which C2D femoral S, Pm and Min-M were 80%, 66% and 61%, respectively, of corresponding B6 values). MHC II knockout delays the development of adult bone properties and is accompanied by lower body mass compared to wild-type controls.

Depleting dietary valine permits nonmyeloablative mouse hematopoietic stem cell transplantation.

PubMed

Taya, Yuki; Ota, Yasunori; Wilkinson, Adam C; Kanazawa, Ayano; Watarai, Hiroshi; Kasai, Masataka; Nakauchi, Hiromitsu; Yamazaki, Satoshi

2016-12-02

A specialized bone marrow microenvironment (niche) regulates hematopoietic stem cell (HSC) self-renewal and commitment. For successful donor-HSC engraftment, the niche must be emptied via myeloablative irradiation or chemotherapy. However, myeloablation can cause severe complications and even mortality. Here we report that the essential amino acid valine is indispensable for the proliferation and maintenance of HSCs. Both mouse and human HSCs failed to proliferate when cultured in valine-depleted conditions. In mice fed a valine-restricted diet, HSC frequency fell dramatically within 1 week. Furthermore, dietary valine restriction emptied the mouse bone marrow niche and afforded donor-HSC engraftment without chemoirradiative myeloablation. These findings indicate a critical role for valine in HSC maintenance and suggest that dietary valine restriction may reduce iatrogenic complications in HSC transplantation. Copyright © 2016, American Association for the Advancement of Science.

Biorhythms, deciduous enamel thickness, and primary bone growth: a test of the Havers-Halberg Oscillation hypothesis.

PubMed

Mahoney, Patrick; Miszkiewicz, Justyna J; Pitfield, Rosie; Schlecht, Stephen H; Deter, Chris; Guatelli-Steinberg, Debbie

2016-06-01

Across mammalian species, the periodicity with which enamel layers form (Retzius periodicity) in permanent teeth corresponds with average body mass and the pace of life history. According to the Havers-Halberg Oscillation hypothesis (HHO), Retzius periodicity (RP) is a manifestation of a biorhythm that is also expressed in lamellar bone. Potentially, these links provide a basis for investigating aspects of a species' biology from fossilized teeth. Here, we tested intra-specific predictions of this hypothesis on skeletal samples of human juveniles. We measured daily enamel growth increments to calculate RP in deciduous molars (n = 25). Correlations were sought between RP, molar average and relative enamel thickness (AET, RET), and the average amount of primary bone growth (n = 7) in humeri of age-matched juveniles. Results show a previously undescribed relationship between RP and enamel thickness. Reduced major axis regression reveals RP is significantly and positively correlated with AET and RET, and scales isometrically. The direction of the correlation was opposite to HHO predictions as currently understood for human adults. Juveniles with higher RPs and thicker enamel had increased primary bone formation, which suggests a coordinating biorhythm. However, the direction of the correspondence was, again, opposite to predictions. Next, we compared RP from deciduous molars with new data for permanent molars, and with previously published values. The lowermost RP of 4 and 5 days in deciduous enamel extends below the lowermost RP of 6 days in permanent enamel. A lowered range of RP values in deciduous enamel implies that the underlying biorhythm might change with age. Our results develop the intra-specific HHO hypothesis. © 2016 Anatomical Society.

Is it safe to preserve the deltoid when resecting the proximal humerus for a primary malignant bone tumour? A comparative study.

PubMed

Cladière-Nassif, V; Bourdet, C; Audard, V; Babinet, A; Anract, P; Biau, D

2017-09-01

Resection of the proximal humerus for the primary malignant bone tumour sometimes requires en bloc resection of the deltoid. However, there is no information in the literature which helps a surgeon decide whether to preserve the deltoid or not. The aim of this study was to determine whether retaining the deltoid at the time of resection would increase the rate of local recurrence. We also sought to identify the variables that persuade expert surgeons to choose a deltoid sparing rather than deltoid resecting procedure. We reviewed 45 patients who had undergone resection of a primary malignant tumour of the proximal humerus. There were 29 in the deltoid sparing group and 16 in the deltoid resecting group. Imaging studies were reviewed to assess tumour extension and soft-tissue involvement. The presence of a fat rim separating the tumour from the deltoid on MRI was particularly noted. The cumulative probability of local recurrence was calculated in a competing risk scenario. There was no significant difference (adjusted p = 0.89) in the cumulative probability of local recurrence between the deltoid sparing (7%, 95% confidence interval (CI) 1 to 20) and the deltoid resecting group (26%, 95% CI 8 to 50). Patients were more likely to be selected for a deltoid sparing procedure if they presented with a small tumour (p = 0.0064) with less bone involvement (p = 0.032) and a continuous fat rim on MRI (p = 0.002) and if the axillary nerve could be identified (p = 0.037). A deltoid sparing procedure can provide good local control after resection of the proximal humerus for a primary malignant bone tumour. A smaller tumour, the presence of a continuous fat rim and the identification of the axillary nerve on pre-operative MRI will persuade surgeons to opt for a deltoid resecting procedure. Cite this article: Bone Joint J 2017;99-B:1244-9. ©2017 The British Editorial Society of Bone & Joint Surgery.

Gain-of-function mutation in FGFR3 in mice leads to decreased bone mass by affecting both osteoblastogenesis and osteoclastogenesis

PubMed Central

Su, Nan; Sun, Qidi; Li, Can; Lu, Xiumin; Qi, Huabing; Chen, Siyu; Yang, Jing; Du, Xiaolan; Zhao, Ling; He, Qifen; Jin, Min; Shen, Yue; Chen, Di; Chen, Lin

2010-01-01

Achondroplasia (ACH) is a short-limbed dwarfism resulting from gain-of-function mutations in fibroblast growth factor receptor 3 (FGFR3). Previous studies have shown that ACH patients have impaired chondrogenesis, but the effects of FGFR3 on bone formation and bone remodeling at adult stages of ACH have not been fully investigated. Using micro-computed tomography and histomorphometric analyses, we found that 2-month-old Fgfr3G369C/+ mice (mouse model mimicking human ACH) showed decreased bone mass due to reduced trabecular bone volume and bone mineral density, defect in bone mineralization and increased osteoclast numbers and activity. Compared with primary cultures of bone marrow stromal cells (BMSCs) from wild-type mice, Fgfr3G369C/+ cultures showed decreased cell proliferation, increased osteogenic differentiation including up-regulation of alkaline phosphatase activity and expressions of osteoblast marker genes, and reduced bone matrix mineralization. Furthermore, our studies also suggest that decreased cell proliferation and enhanced osteogenic differentiation observed in Fgfr3G369C/+ BMSCs are caused by up-regulation of p38 phosphorylation and that enhanced Erk1/2 activity is responsible for the impaired bone matrix mineralization. In addition, in vitro osteoclast formation and bone resorption assays demonstrated that osteoclast numbers and bone resorption area were increased in cultured bone marrow cells derived from Fgfr3G369C/+ mice. These findings demonstrate that gain-of-function mutation in FGFR3 leads to decreased bone mass by regulating both osteoblast and osteoclast activities. Our studies provide new insight into the mechanism underlying the development of ACH. PMID:20053668

Change in Mouse Bone Turnover in Response to Microgravity on RR-1

NASA Technical Reports Server (NTRS)

Cheng-Campbell, Margareth A.; Blaber, Elizabeth A.; Almeida, Eduardo A. C.

2016-01-01

Mechanical unloading during spaceflight is known to adversely affect mammalian physiology. Our previous studies using the Animal Enclosure Module on short duration Shuttle missions enabled us to identify a deficit in stem cell based-tissue regeneration as being a significant concern for long-duration spaceflight. Specifically, we found that mechanical unloading in microgravity resulted in inhibition of differentiation of mesenchymal and hematopoietic stem cells in the bone marrow compartment. Also, we observed overexpression of a cell cycle arrest molecule, CDKN1ap21, in osteoprecursor cells on the bone surface, chondroprogenitors in the articular cartilage, and in myofibers attached to bone tissue. Specifically in bone tissue during both short (15-day) and long (30-day) microgravity experiments, we observed significant loss of bone tissue and structure in both the pelvis and the femur. After 15-days of microgravity on STS-131, pelvic ischium displayed a 6.23 decrease in bone fraction (p0.005) and 11.91 decrease in bone thickness (p0.002). Furthermore, during long-duration spaceflight we observed onset of an accelerated aging-like phenotype and osteoarthritic disease state indicating that stem cells within the bone tissue fail to repair and regenerate tissues in a normal manner, leading to drastic tissue alterations in response to microgravity. The Rodent Research Hardware System provides the capability to investigate these effects during long-duration experiments on the International Space Station. During the Rodent Research-1 mission 10 16-week-old female C57Bl6J mice were exposed to 37-days of microgravity. All flight animals were euthanized and frozen on orbit for future dissection. Ground (n10) and vivarium controls (n10) were housed and processed to match the flight animal timeline. During this study we collected pelvis, femur, and tibia from all animal groups to test the hypothesis that stem cell-based tissue regeneration is significantly altered after 37

Suppression of Hepatic Cyp1a2 by Total Ginsenosides in Lipopolysaccharide-Treated Mice and Primary Mouse Hepatocytes.

PubMed

Sun, Haiyan; Yan, Yijing; Xu, Chenshu; Wan, Hongxia; Liu, Dong

2016-03-23

The roots of Panax ginseng (ginseng) have been extensively used in traditional Chinese medicine. However, herb-drug interactions between ginseng and other co-administered drugs are not fully understood concerning the effect of ginseng on drug metabolism and clearance. The current study aimed to elucidate the effect of total ginsenosides, a typical ginseng extract, on the regulation of Cyp1a2, a key enzyme to regulate drug metabolism under the normal and inflammatory conditions in mice. Female C57BL/6J mice treated with vehicle and lipopolysaccharide (LPS) were intragastrically administered ginseng extract for 7 days before hepatic P450 expression was analyzed. Primary mouse hepatocytes were also employed to further explore the effects of total ginsenosides on Cyp1a2 expression. The results showed that total ginsenosides in P. ginseng extract exhibited a concentration-dependent suppression on Cyp1a2 mRNA and protein level in both mice and primary mouse hepatocytes. Notably, the inhibitory effects of total ginsenosides on Cyp1a2 mRNA and protein expression were further enhanced following LPS treatment. Therefore, future research is warranted to investigate the role of ginsenosides in the regulation of hepatic CYP450s. Moreover, consumption of ginseng as food or supplement should be monitored for patients on combinational therapy, especially those with inflammatory diseases.

Anabolic activity of ursolic acid in bone: Stimulating osteoblast differentiation in vitro and inducing new bone formation in vivo.

PubMed

Lee, Su-Ui; Park, Sang-Joon; Kwak, Han Bok; Oh, Jaemin; Min, Yong Ki; Kim, Seong Hwan

2008-01-01

In the field of osteoporosis, there has been growing interest in anabolic agents that enhance bone mass and improve bone architecture. In this study, we demonstrated that the ubiquitous plant triterpenoid, ursolic acid, enhances differentiation and mineralization of osteoblasts in vitro. We found that ursolic acid induced the expression of osteoblast-specific genes with the activation of mitogen-activated protein kinases, nuclear factor-kappaB, and activator protein-1. Additionally, noggin, an antagonist of bone morphogenetic proteins (BMPs), inhibited ursolic acid-induced osteoblast differentiation. Noggin also inhibited the activation of Smad and the induction of BMP-2 mRNA expression by ursolic acid in the late stage of osteoblast differentiation. Importantly, ursolic acid was shown to have bone-forming activity in vivo in a mouse calvarial bone formation model. A high proportion of positive immunostaining of BMP-2 was found in the nuclear region of woven bone formed by ursolic acid. These results suggested that ursolic acid has the anabolic potential to stimulate osteoblast differentiation and enhance new bone formation.

Novel osteoinductive photo-cross-linkable chitosan-lactide-fibrinogen hydrogels enhance bone regeneration in critical size segmental bone defects

PubMed Central

Kim, Sungwoo; Bedigrew, Katherine; Guda, Teja; Maloney, William J.; Park, Sangwon; Wenke, Joseph C.; Yang, Yunzhi Peter

2014-01-01

The purpose of this study was to develop and characterize a novel photo-cross-linkable chitosan-lactide-fibrinogen (CLF) hydrogel and evaluate the efficacy of bone morphogenetic protein-2 (BMP-2) containing CLF hydrogel for osteogenesis in vitro and in vivo. We synthesized the CLF hydrogels and characterized their chemical structure, degradation rate, compressive modulus, and in vitro BMP-2 release kinetics. We evaluated bioactivities of the BMP-2 containing CLF hydrogels (0, 50, 100, and 500 ng/ml) in vitro using W-20-17 preosteoblast mouse bone marrow stromal cells and C2C12 mouse myoblast cells. The effect of BMP-2 containing CLF gels (0, 0.5, 1, 2, and 5μg) on bone formation was evaluated using rat critical size segmental bone defects for 4 weeks. FTIR spectra and SEM images showed chemical and structural changes by addition of fibrinogen into chitosan-lactide copolymer. Incorporation of fibrinogen molecules significantly increased compressive modulus of the hydrogels. In vitro BMP-2 release study showed initial burst releases from the CLF hydrogels followed by sustained releases, regardless of the concentration of the BMP-2 over 4 weeks. Cells in all groups were viable in the presence of the hydrogels regardless of BMP-2 doses, indicating non-cytotoxicity of hydrogels. Alkaline phosphate activity and mineralization of cells exhibited dose dependence on BMP-2 containing CLF hydrogels. Radiographs, microcomputed tomography, and histology confirmed that the BMP-2 containing CLF hydrogels prompted neo-osteogenesis and accelerated healing of the defects in a dose-dependent manner. Thus the CLF hydrogel is a promising delivery system of growth factors for bone regeneration. PMID:25174669

EARLY ONSET OF CRANIOSYNOSTOSIS IN AN APERT MOUSE MODEL REVEALS CRITICAL FEATURES OF THIS PATHOLOGY

PubMed Central

Holmes, Greg; Rothschild, Gerson; Roy, Upal Basu; Deng, Chu-Xia; Mansukhani, Alka; Basilico, Claudio

2009-01-01

Activating mutations of FGFRs1–3 cause craniosynostosis (CS), the premature fusion of cranial bones, in man and mouse. The mechanisms by which such mutations lead to CS have been variously ascribed to increased osteoblast proliferation, differentiation, and apoptosis, but it is not always clear how these disturbances relate to the process of suture fusion. We have reassessed coronal suture fusion in an Apert Fgfr2 (S252W) mouse model. We find that the critical event of CS is the early loss of basal sutural mesenchyme as the osteogenic fronts, expressing activated Fgfr2, unite to form a contiguous skeletogenic membrane. A mild increase in osteoprogenitor proliferation precedes but does not accompany this event, and apoptosis is insignificant. On the other hand, the more apical coronal suture initially forms appropriately but then undergoes fusion, albeit at a slower rate, accompanied by a significant decrease in osteoprogenitor proliferation, and increased osteoblast maturation. Apoptosis now accompanies fusion, but is restricted to bone fronts in contact with one another. We correlated these in vivo observations with the intrinsic effects of the activated Fgfr2 S252W mutation in primary osteoblasts in culture, which show an increased capacity for both proliferation and differentiation. Our studies suggest that the major determinant of Fgfr2-induced craniosynostosis is the failure to respond to signals that would halt the recruitment or the advancement of osteoprogenitor cells at the sites where sutures should normally form. PMID:19389359

Effects of the 1, 4-dihydropyridine L-type calcium channel blocker benidipine on bone marrow stromal cells.

PubMed

Ma, Zhong-ping; Liao, Jia-cheng; Zhao, Chang; Cai, Dao-zhang

2015-08-01

Osteoporosis (OP) often increases the risk of bone fracture and other complications and is a major clinical problem. Previous studies have found that high blood pressure is associated with bone formation abnormalities, resulting in increased calcium loss. We have investigated the effect of the antihypertensive drug benidipine on bone marrow stromal cell (BMSC) differentiation into osteoblasts and bone formation under osteoporotic conditions. We used a combination of in vitro and in vivo approaches to test the hypothesis that benidipine promotes murine BMSC differentiation into osteoblasts. Alkaline phosphatase (ALP), osteocalcin (OCN), runt-related transcription factor 2 (RUNX2), β-catenin, and low-density lipoprotein receptor-related protein 5 (LRP5) protein expression was evaluated in primary femoral BMSCs from C57/BL6 mice cultured under osteogenic conditions for 2 weeks to examine the effects of benidipine. An ovariectomized (OVX) mouse model was used to investigate the effect of benidipine treatment for 3 months in vivo. We found that ALP, OCN, and RUNX2 expression was up-regulated and WNT/β-catenin signaling was enhanced in vitro and in vivo. In OVX mice that were intragastrically administered benidipine, bone parameters (trabecular thickness, bone mineral density, and trabecular number) in the distal femoral metaphysis were significantly increased compared with control OVX mice. Consistently, benidipine promoted BMSC differentiation into osteoblasts and protected against bone loss in OVX mice. Therefore, benidipine might be a suitable candidate for the treatment of patients with postmenopausal osteoporosis and hypertension.

Recombinant mouse periostin ameliorates coronal sutures fusion in Twist1+/- mice.

PubMed

Bai, Shanshan; Li, Dong; Xu, Liang; Duan, Huichuan; Yuan, Jie; Wei, Min

2018-04-17

Saethre-Chotzen syndrome is an autosomal dominantly inherited disorder caused by mutations in the twist family basic helix-loop-helix transcription factor 1 (TWIST1) gene. Surgical procedures are frequently required to reduce morphological and functional defects in patients with Saethre-Chotzen syndrome. Therefore, the development of noninvasive procedures to treat Saethre-Chotzen syndrome is critical. We identified that periostin, which is an extracellular matrix protein that plays an important role in both bone and connective tissues, is downregulated in craniosynostosis patients. We aimed to verify the effects of different concentrations (0, 50, 100, and 200 μg/l) of recombinant mouse periostin in Twist1 +/- mice (a mouse model of Saethre-Chotzen syndrome) coronal suture cells in vitro and in vivo. Cell proliferation, migration, and osteogenic differentiation were observed and detected. Twist1 +/- mice were also injected with recombinant mouse periostin to verify the treatment effects. Cell Counting Kit-8 results showed that recombinant mouse periostin inhibited the proliferation of suture-derived cells in a time- and concentration-dependent manner. Cell migration was also suppressed when treated with recombinant mouse periostin. Real-time quantitative PCR and Western blotting results suggested that messenger ribonucleic acid and protein expression of alkaline phosphatase, bone sialoprotein, collagen type I, and osteocalcin were all downregulated after treatment with recombinant mouse periostin. However, the expression of Wnt-3a, Wnt-1, and β-catenin were upregulated. The in vivo results demonstrated that periostin-treated Twist1 +/- mice showed patent coronal sutures in comparison with non-treated Twist1 +/- mice which have coronal craniosynostosis. Our results suggest that recombinant mouse periostin can inhibit coronal suture cell proliferation and migration and suppress osteogenic differentiation of suture-derived cells via Wnt canonical signaling, as

Immortalization and characterization of osteoblast cell lines generated from wild-type and Nmp4-null mouse bone marrow stromal cells using murine telomerase reverse transcriptase (mTERT).

PubMed

Alvarez, Marta B; Childress, Paul; Philip, Binu K; Gerard-O'Riley, Rita; Hanlon, Michael; Herbert, Brittney-Shea; Robling, Alexander G; Pavalko, Fredrick M; Bidwell, Joseph P

2012-05-01

Intermittent parathyroid hormone (PTH) adds new bone to the osteoporotic skeleton; the transcription factor Nmp4/CIZ represses PTH-induced bone formation in mice and as a consequence is a potential drug target for improving hormone clinical efficacy. To explore the impact of Nmp4/CIZ on osteoblast phenotype, we immortalized bone marrow stromal cells from wildtype (WT) and Nmp4-knockout (KO) mice using murine telomerase reverse transcriptase. Clonal lines were initially chosen based on their positive staining for alkaline phosphatase and capacity for mineralization. Disabling Nmp4/CIZ had no gross impact on osteoblast phenotype development. WT and KO clones exhibited identical sustained growth, reduced population doubling times, extended maintenance of the mature osteoblast phenotype, and competency for differentiating toward the osteoblast and adipocyte lineages. Additional screening of the immortalized cells for PTH-responsiveness permitted further studies with single WT and KO clones. We recently demonstrated that PTH-induced c-fos femoral mRNA expression is enhanced in Nmp4-KO mice and in the present study we observed that hormone stimulated either an equivalent or modestly enhanced increase in c-fos mRNA expression in both primary null and KO clone cells depending on PTH concentration. The null primary osteoblasts and KO clone cells exhibited a transiently enhanced response to bone morphogenetic protein 2 (BMP2). The clones exhibited lower and higher expressions of the PTH receptor (Pthr1) and the BMP2 receptor (Bmpr1a, Alk3), respectively, as compared to primary cells. These immortalized cell lines will provide a valuable tool for disentangling the complex functional roles underlying Nmp4/CIZ regulation of bone anabolism. Copyright © 2011 Wiley Periodicals, Inc.

The role of bone sialoprotein in the tendon-bone insertion.

PubMed

Marinovich, Ryan; Soenjaya, Yohannes; Wallace, Gregory Q; Zuskov, Andre; Dunkman, Andrew; Foster, Brian L; Ao, Min; Bartman, Kevin; Lam, Vida; Rizkalla, Amin; Beier, Frank; Somerman, Martha J; Holdsworth, David W; Soslowsky, Louis J; Lagugné-Labarthet, François; Goldberg, Harvey A

2016-01-01

Tendons/ligaments insert into bone via a transitional structure, the enthesis, which is susceptible to injury and difficult to repair. Fibrocartilaginous entheses contain fibrocartilage in their transitional zone, part of which is mineralized. Mineral-associated proteins within this zone have not been adequately characterized. Members of the Small Integrin Binding Ligand N-linked Glycoprotein (SIBLING) family are acidic phosphoproteins expressed in mineralized tissues. Here we show that two SIBLING proteins, bone sialoprotein (BSP) and osteopontin (OPN), are present in the mouse enthesis. Histological analyses indicate that the calcified zone of the quadriceps tendon enthesis is longer in Bsp(-/-) mice, however no difference is apparent in the supraspinatus tendon enthesis. In an analysis of mineral content within the calcified zone, micro-CT and Raman spectroscopy reveal that the mineral content in the calcified fibrocartilage of the quadriceps tendon enthesis are similar between wild type and Bsp(-/-) mice. Mechanical testing of the patellar tendon shows that while the tendons fail under similar loads, the Bsp(-/-) patellar tendon is 7.5% larger in cross sectional area than wild type tendons, resulting in a 16.5% reduction in failure stress. However, Picrosirius Red staining shows no difference in collagen organization. Data collected here indicate that BSP is present in the calcified fibrocartilage of murine entheses and suggest that BSP plays a regulatory role in this structure, influencing the growth of the calcified fibrocartilage in addition to the weakening of the tendon mechanical properties. Based on the phenotype of the Bsp(-/-) mouse enthesis, and the known in vitro functional properties of the protein, BSP may be a useful therapeutic molecule in the reattachment of tendons and ligaments to bone. Copyright © 2016 International Society of Matrix Biology. All rights reserved.

The Role of Bone Sialoprotein in the Tendon-Bone Insertion

PubMed Central

Marinovich, Ryan; Soenjaya, Yohannes; Wallace, Gregory Q.; Zuskov, Andre; Dunkman, Andrew; Foster, Brian L.; Ao, Min; Bartman, Kevin; Lam, Vida; Rizkalla, Amin; Beier, Frank; Somerman, Martha J.; Holdsworth, David W.; Soslowsky, Louis J.; Lagugné-Labarthet, François; Goldberg, Harvey A.

2016-01-01

Tendons/ligaments insert into bone via a transitional structure, the enthesis, which is susceptible to injury and difficult to repair. Fibrocartilaginous entheses contain fibrocartilage in their transitional zone, part of which is mineralized. Mineral-associated proteins within this zone have not been adequately characterized. Members of the Small Integrin Binding Ligand N-Linked Glycoprotein (SIBLING) family are acidic phosphoproteins expressed in mineralized tissues. Here we show that two SIBLING proteins, bone sialoprotein (BSP) and osteopontin (OPN), are present in the mouse enthesis. Histological analyses indicate that the calcified zone of the quadriceps tendon enthesis is longer in Bsp−/− mice, however no difference is apparent in the supraspinatus tendon enthesis. In an analysis of mineral content within the calcified zone, micro-CT and Raman spectroscopy reveal that the mineral content in the calcified fibrocartilage of the quadriceps tendon enthesis are similar between wild type and Bsp−/− mice. Mechanical testing of the patellar tendon shows that while the tendons fail under similar loads, the Bsp−/− patellar tendon is 7.5% larger in cross sectional area than wild type tendons, resulting in a 16.5% reduction in failure stress. However, picrosirius red staining shows no difference in collagen organization. Data collected here indicate that BSP is present in the calcified fibrocartilage of murine entheses and suggest that BSP plays a regulatory role in this structure, influencing the growth of the calcified fibrocartilage in addition to the weakening of the tendon mechanical properties. Based on the phenotype of the Bsp−/− mouse enthesis, and the known in vitro functional properties of the protein, BSP may be a useful therapeutic molecule in the reattachment of tendons and ligaments to bone. PMID:26826499

Rapid ex vivo imaging of PAIII prostate to bone tumor with SWIFT-MRI.

PubMed

Luhach, Ihor; Idiyatullin, Djaudat; Lynch, Conor C; Corum, Curt; Martinez, Gary V; Garwood, Michael; Gillies, Robert J

2014-09-01

The limiting factor for MRI of skeletal/mineralized tissue is fast transverse relaxation. A recent advancement in MRI technology, SWIFT (Sweep Imaging with Fourier Transform), is emerging as a new approach to overcome this difficulty. Among other techniques like UTE, ZTE, and WASPI, the application of SWIFT technology has the strong potential to impact preclinical and clinical imaging, particularly in the context of primary or metastatic bone cancers because it has the added advantage of imaging water in mineralized tissues of bone allowing MRI images to be obtained of tissues previously visible only with modalities such as computed tomography (CT). The goal of the current study is to examine the feasibility of SWIFT for the assessment of the prostate cancer induced changes in bone formation (osteogenesis) and destruction (osteolysis) in ex vivo specimens. A luciferase expressing prostate cancer cell line (PAIII) or saline control was inoculated directly into the tibia of 6-week-old immunocompromised male mice. Tumor growth was assessed weekly for 3 weeks before euthanasia and dissection of the tumor bearing and sham tibias. The ex vivo mouse tibia specimens were imaged with a 9.4 Tesla (T) and 7T MRI systems. SWIFT images are compared with traditional gradient-echo and spin-echo MRI images as well as CT and histological sections. SWIFT images with nominal resolution of 78 μm are obtained with the tumor and different bone structures identified. Prostate cancer induced changes in the bone microstructure are visible in SWIFT images, which is supported by spin-echo, high resolution CT and histological analysis. SWIFT MRI is capable of high-quality high-resolution ex vivo imaging of bone tumor and surrounding bone and soft tissues. Furthermore, SWIFT MRI shows promise for in vivo bone tumor imaging, with the added benefits of nonexposure to ionizing radiation, quietness, and speed. Copyright © 2013 Wiley Periodicals, Inc.

Disorders of bone and bone mineral metabolism.

PubMed

Komoroski, Monica; Azad, Nasrin; Camacho, Pauline

2014-01-01

Metabolic bone disorders are very common in the general population and untreated, they can cause a variety of neurologic symptoms. These diseases include osteoporosis, vitamin D deficiency, Paget's disease, and alterations in calcium, phosphorus, and magnesium metabolism. Diagnosis is made through analysis of metabolic bone blood chemistries as well as radiologic studies such as dual energy X-ray absorptiometry (DXA) scans, bone scans, and X-rays. Treatment options have advanced significantly in the past decade for osteoporosis and Paget's disease and mainly include antiresorptive therapy. New recommendations for treatment of primary hyperparathyroidism are discussed as well as therapy for calcium, phosphorus, and mineral disorders. © 2014 Elsevier B.V. All rights reserved.

MMP-8, A Breast Cancer Bone Metastasis Suppressor Gene

DTIC Science & Technology

2006-08-01

new protein synthesis. This event is particularly important in situations such as tissue repair following injury . PTH and TGF-b1 stimulated LTBP-1...osteoclastogenesis inhi- bitory factor in the stimulation of osteoclast formation by parathyroid hormone in mouse bone cells. Eur J Endo - crinol 142:661–664...done to determine cross-sectional area, bone volume, and perios - teal perimeter (Ps.Pm). The endocortical sur- face was outlined, and the analysis

Dietary Strontium Increases Bone Mineral Density in Intact Zebrafish (Danio rerio): A Potential Model System for Bone Research

PubMed Central

Padgett-Vasquez, Steve; Garris, Heath W.; Nagy, Tim R.; D'Abramo, Louis R.; Watts, Stephen A.

2010-01-01

Abstract Zebrafish (Danio rerio) skeletal bone possesses properties similar to human bone, which suggests that they may be used as a model to study mineralization characteristics of the human Haversian system, as well as human bone diseases. One prerequisite for the use of zebrafish as an alternative osteoporotic bone model is to determine whether their bone displays functional plasticity similar to that observed in other bone models. Strontium citrate was supplemented into a laboratory-prepared diet (45% crude protein) to produce dietary strontium levels of 0%, 0.63%, 1.26%, 1.89%, and 2.43% and fed ad libitum twice daily for 12 weeks to 28-day-old intact zebrafish. Length was determined at 4-week intervals, and both weight and length were recorded at 12 weeks. At 12 weeks, seven zebrafish from each dietary level were analyzed for total bone mineral density by microcomputed tomography. Dietary strontium citrate supplementation significantly (p < 0.05) increased zebrafish whole-body and spinal column bone mineral density. In addition, trace amounts of strontium were incorporated into the scale matrix in those zebrafish that consumed strontium-supplemented diets. These findings suggest that zebrafish bone displays plasticity similar to that reported for other bone models (i.e., rat, mouse, and monkey) that received supplements of strontium compounds and zebrafish should be viewed as an increasingly valuable bone model. PMID:20874492

Establishment and proteomic characterization of NCC-LMS1-C1, a novel cell line of primary leiomyosarcoma of the bone.

PubMed

Sakumoto, Marimu; Takahashi, Mami; Oyama, Rieko; Takai, Yoko; Kito, Fusako; Shiozawa, Kumiko; Qiao, Zhiwei; Yoshida, Akihiko; Endo, Makoto; Kawai, Akira; Kondo, Tadashi

2017-10-01

Leiomyosarcoma (LMS) is one of most aggressive mesenchymal malignancies that differentiate towards smooth muscle. The clinical outcome of LMS patients is poor; as such, there is an urgent need for novel therapeutic approaches. Experimental models such as patient-derived cell lines are invaluable tools for pre-clinical studies. In the present study, we established a stable cell line from the tumor tissue of a patient with a primary LMS of the bone. Despite the urgent need for novel therapeutic strategies in LMS, there are only a few LMS cell lines available in public cell banks, none of which are primary to the bone. Bone primary LMS tumor tissues were sampled to establish cell lines. Morphological and proteomic analyses were performed and sensitivity to pazopanib was evaluated. NCC-LMS1-C1 cells were maintained for over 100 passages. The cells exhibited a spindle shape and aggressive growth; they also expressed smooth muscle actin, reflecting the original LMS tissue (i.e. smooth muscle cells). The cells also showed tumor characteristics such as colony formation on soft agar and sensitivity to pazopanib, doxorubicin and cisplatin, with half-maximal inhibitory concentrations of 4.5, 0.11 and 20 μM, respectively. Proteomic analyses by mass spectrometry and antibody array revealed some differences in the protein expression profiles of these cells as compared to the original tumor tissue. Our results indicate that the NCC-LMS1-C1 cell lines will be useful for LMS research. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

[Isolation, purification and primary culture of adult mouse cardiac fibroblasts].

PubMed

Li, Rujun; Gong, Kaizheng; Zhang, Zhengang

2017-01-01

Objective To establish a method for primary culture of adult mouse cardiac fibroblasts. Methods Myocardial tissues from adult mice were digested with 1 g/L trypsin and 0.8 g/L collagenase IV by oscillating water bath for a short time repeatedly. Cardiac fibroblasts and myocardial cells were isolated with differential adhesion method. Immunofluorescence staining was used to assess the purity of cardiac fibroblasts. The cell morphology was observed under an inverted phase contrast microscope. The proliferation of cardiac fibroblasts was analyzed by growth curve and CCK-8 assay. The Smad2/3 phosphorylation induced by TGF-β1 was detected by Western blotting. Results After 90 minutes of differential adhesion, adherent fibroblasts formed spherical cell mass and after 3 days, cells were spindle-shaped and proliferated rapidly. Cells were confluent after 5 days and the growth curve presented nearly "S" shape. The positive expression rate of vimentin was 95%. CCK-8 assay showed that the optimal cell proliferating activity was found from day 3 to day 5. The level of phosphorylated Smad2/3 obviously increased at the second passage induced by TGF-β1. Conclusion This method is economical and stable to isolate cardiac fibroblasts with high activity and high purity from adult mice.

A novel osteogenic oxysterol compound for therapeutic development to promote bone growth: activation of hedgehog signaling and osteogenesis through smoothened binding.

PubMed

Montgomery, Scott R; Nargizyan, Taya; Meliton, Vicente; Nachtergaele, Sigrid; Rohatgi, Rajat; Stappenbeck, Frank; Jung, Michael E; Johnson, Jared S; Aghdasi, Bayan; Tian, Haijun; Weintraub, Gil; Inoue, Hirokazu; Atti, Elisa; Tetradis, Sotirios; Pereira, Renata C; Hokugo, Akishige; Alobaidaan, Raed; Tan, Yanlin; Hahn, Theodor J; Wang, Jeffrey C; Parhami, Farhad

2014-08-01

Osteogenic factors are often used in orthopedics to promote bone growth, improve fracture healing, and induce spine fusion. Osteogenic oxysterols are naturally occurring molecules that were shown to induce osteogenic differentiation in vitro and promote spine fusion in vivo. The purpose of this study was to identify an osteogenic oxysterol more suitable for clinical development than those previously reported, and evaluate its ability to promote osteogenesis in vitro and spine fusion in rats in vivo. Among more than 100 oxysterol analogues synthesized, Oxy133 induced significant expression of osteogenic markers Runx2, osterix (OSX), alkaline phosphatase (ALP), bone sialoprotein (BSP), and osteocalcin (OCN) in C3H10T1/2 mouse embryonic fibroblasts and in M2-10B4 mouse marrow stromal cells. Oxy133-induced activation of an 8X-Gli luciferase reporter, its direct binding to Smoothened, and the inhibition of Oxy133-induced osteogenic effects by the Hedgehog (Hh) pathway inhibitor, cyclopamine, demonstrated the role of Hh pathway in mediating osteogenic responses to Oxy133. Oxy133 did not stimulate osteogenesis via BMP or Wnt signaling. Oxy133 induced the expression of OSX, BSP, and OCN, and stimulated robust mineralization in primary human mesenchymal stem cells. In vivo, bilateral spine fusion occurred through endochondral ossification and was observed in animals treated with Oxy133 at the fusion site on X-ray after 4 weeks and confirmed with manual assessment, micro-CT (µCT), and histology after 8 weeks, with equal efficiency to recombinant human bone morphogenetic protein-2 (rhBMP-2). Unlike rhBMP-2, Oxy133 did not induce adipogenesis in the fusion mass and resulted in denser bone evidenced by greater bone volume/tissue volume (BV/TV) ratio and smaller trabecular separation. Findings here suggest that Oxy133 has significant potential as an osteogenic molecule with greater ease of synthesis and improved time to fusion compared to previously studied oxysterols. Small

Mousepox detected in a research facility: case report and failure of mouse antibody production testing to identify Ectromelia virus in contaminated mouse serum.

PubMed

Labelle, Philippe; Hahn, Nina E; Fraser, Jenelle K; Kendall, Lonnie V; Ziman, Melanie; James, Edward; Shastri, Nilabh; Griffey, Stephen M

2009-04-01

An outbreak of mousepox in a research institution was caused by Ectromelia-contaminated mouse serum that had been used for bone marrow cell culture and the cells subsequently injected into the footpads of mice. The disease initially was diagnosed by identification of gross and microscopic lesions typical for Ectromelia infection, including foci of necrosis in the liver and spleen and eosinophilic intracytoplasmic inclusion bodies in the skin. The source of infection was determined by PCR analysis to be serum obtained from a commercial vendor. To determine whether viral growth in tissue culture was required to induce viral infection, 36 mice (BALB/cJ, C57BL/6J) were experimentally exposed intraperitoneally, intradermally (footpad), or intranasally to contaminated serum or bone marrow cell cultures using the contaminated serum in the culture medium. Mice were euthanized when clinical signs developed or after 12 wk. Necropsy, PCR of spleen, and serum ELISA were performed on all mice. Mice injected with cell cultures and their cage contacts developed mousepox, antibodies to Ectromelia, and lesions, whereas mice injected with serum without cells did not. Mouse antibody production, a tool commonly used to screen biologic materials for viral contamination, failed to detect active Ectromelia contamination in mouse serum.

Lipid Osteoclastokines Regulate Breast Cancer Bone Metastasis

PubMed Central

Krzeszinski, Jing Y.; Schwaid, Adam G.; Cheng, Wing Yin; Jin, Zixue; Gallegos, Zachary R.; Saghatelian, Alan

2017-01-01

Bone metastasis is a deadly consequence of cancers, in which osteoclast forms a vicious cycle with tumor cells. Bone metastasis attenuation by clinical usage of osteoclast inhibitors and in our osteopetrotic mouse genetic models with β-catenin constitutive activation or peroxisome proliferator-activated receptor γ deficiency fully support the important role of osteoclast in driving the bone metastatic niche. However, the mechanisms for this “partnership in crime” are underexplored. Here we show that osteoclasts reprogram their lipid secretion to support cancer cells. Metabolomic profiling reveals elevated prometastatic arachidonic acid (AA) but reduced antimetastatic lysophosphatidylcholines (LPCs). This shift in lipid osteoclastokines synergistically stimulates tumor cell proliferation, migration, survival, and expression of prometastatic genes. Pharmacologically, combined treatment with LPCs and BW-755C, an inhibitor of AA signaling via blocking lipoxygenase and cyclooxygenase, impedes breast cancer bone metastasis. Our findings elucidate key paracrine mechanisms for the osteoclast-cancer vicious cycle and uncover important therapeutic targets for bone metastasis. PMID:27967239

Turnover of bone marrow-derived cells in the irradiated mouse cornea

PubMed Central

Chinnery, Holly R; Humphries, Timothy; Clare, Adam; Dixon, Ariane E; Howes, Kristen; Moran, Caitlin B; Scott, Danielle; Zakrzewski, Marianna; Pearlman, Eric; McMenamin, Paul G

2008-01-01

In light of an increasing awareness of the presence of bone marrow (BM)-derived macrophages in the normal cornea and their uncertain role in corneal diseases, it is important that the turnover rate of these resident immune cells be established. The baseline density and distribution of macrophages in the corneal stroma was investigated in Cx3cr1gfp transgenic mice in which all monocyte-derived cells express enhanced green fluorescent protein (eGFP). To quantify turnover, BM-derived cells from transgenic eGFP mice were transplanted into whole-body irradiated wild-type recipients. Additionally, wild-type BM-derived cells were injected into irradiated Cx3cr1+/gfp recipients, creating reverse chimeras. At 2, 4 and 8 weeks post-reconstitution, the number of eGFP+ cells in each corneal whole mount was calculated using epifluorescence microscopy, immunofluorescence staining and confocal microscopy. The total density of myeloid-derived cells in the normal Cx3cr1+/gfp cornea was 366 cells/mm2. In BM chimeras 2 weeks post-reconstitution, 24% of the myeloid-derived cells had been replenished and were predominantly located in the anterior stroma. By 8 weeks post-reconstitution 75% of the myeloid-derived cells had been replaced and these cells were distributed uniformly throughout the stroma. All donor eGFP+ cells expressed low to moderate levels of CD45 and CD11b, with approximately 25% coexpressing major histocompatibility complex class II, a phenotype characteristic of previous descriptions of corneal stromal macrophages. In conclusion, 75% of the myeloid-derived cells in the mouse corneal stroma are replenished after 8 weeks. These data provide a strong basis for functional investigations of the role of resident stromal macrophages versus non-haematopoietic cells using BM chimeric mice in models of corneal inflammation. PMID:18540963

Neonatal bone marrow transplantation prevents bone pathology in a mouse model of mucopolysaccharidosis type I.

PubMed

Pievani, Alice; Azario, Isabella; Antolini, Laura; Shimada, Tsutomu; Patel, Pravin; Remoli, Cristina; Rambaldi, Benedetta; Valsecchi, Maria Grazia; Riminucci, Mara; Biondi, Andrea; Tomatsu, Shunji; Serafini, Marta

2015-03-05

Neonatal bone marrow transplantation (BMT) could offer a novel therapeutic opportunity for genetic disorders by providing sustainable levels of the missing protein at birth, thus preventing tissue damage. We tested this concept in mucopolysaccharidosis type I (MPS IH; Hurler syndrome), a lysosomal storage disorder caused by deficiency of α-l-iduronidase. MPS IH is characterized by a broad spectrum of clinical manifestations, including severe progressive skeletal abnormalities. Although BMT increases the life span of patients with MPS IH, musculoskeletal manifestations are only minimally responsive if the timing of BMT delays, suggesting already irreversible bone damage. In this study, we tested the hypothesis that transplanting normal BM into newborn MPS I mice soon after birth can prevent skeletal dysplasia. We observed that neonatal BMT was effective at restoring α-l-iduronidase activity and clearing elevated glycosaminoglycans in blood and multiple organs. At 37 weeks of age, we observed an almost complete normalization of all bone tissue parameters, using radiographic, microcomputed tomography, biochemical, and histological analyses. Overall, the magnitude of improvements correlated with the extent of hematopoietic engraftment. We conclude that BMT at a very early stage in life markedly reduces signs and symptoms of MPS I before they appear. © 2015 by The American Society of Hematology.

Lymphatic Endothelial Cells Produce M-CSF, Causing Massive Bone Loss in Mice.

PubMed

Wang, Wensheng; Wang, Hua; Zhou, Xichao; Li, Xing; Sun, Wen; Dellinger, Michael; Boyce, Brendan F; Xing, Lianping

2017-05-01

Gorham-Stout disease (GSD) is a rare bone disorder characterized by aggressive osteolysis associated with lymphatic vessel invasion within bone marrow cavities. The etiology of GSD is not known, and there is no effective therapy or animal model for the disease. Here, we investigated if lymphatic endothelial cells (LECs) affect osteoclasts (OCs) to cause a GSD osteolytic phenotype in mice. We examined the effect of a mouse LEC line on osteoclastogenesis in co-cultures. LECs significantly increased receptor activator of NF-κB ligand (RANKL)-mediated OC formation and bone resorption. LECs expressed high levels of macrophage colony-stimulating factor (M-CSF), but not RANKL, interleukin-6 (IL-6), and tumor necrosis factor (TNF). LEC-mediated OC formation and bone resorption were blocked by an M-CSF neutralizing antibody or Ki20227, an inhibitor of the M-CSF receptor, c-Fms. We injected LECs into the tibias of wild-type (WT) mice and observed massive osteolysis on X-ray and micro-CT scans. Histology showed that LEC-injected tibias had significant trabecular and cortical bone loss and increased OC numbers. M-CSF protein levels were significantly higher in serum and bone marrow plasma of mice given intra-tibial LEC injections. Immunofluorescence staining showed extensive replacement of bone and marrow by podoplanin+ LECs. Treatment of LEC-injected mice with Ki20227 significantly decreased tibial bone destruction. In addition, lymphatic vessels in a GSD bone sample were stained positively for M-CSF. Thus, LECs cause bone destruction in vivo in mice by secreting M-CSF, which promotes OC formation and activation. Blocking M-CSF signaling may represent a new therapeutic approach for treatment of patients with GSD. Furthermore, tibial injection of LECs is a useful mouse model to study GSD. © 2017 American Society for Bone and Mineral Research. © 2017 American Society for Bone and Mineral Research.

The Small Molecule Inhibitor G6 Significantly Reduces Bone Marrow Fibrosis and the Mutant Burden in a Mouse Model of Jak2-Mediated Myelofibrosis

PubMed Central

Kirabo, Annet; Park, Sung O.; Wamsley, Heather L.; Gali, Meghanath; Baskin, Rebekah; Reinhard, Mary K.; Zhao, Zhizhuang J.; Bisht, Kirpal S.; Keserű, György M.; Cogle, Christopher R.; Sayeski, Peter P.

2013-01-01

Philadelphia chromosome–negative myeloproliferative neoplasms, including polycythemia vera, essential thrombocytosis, and myelofibrosis, are disorders characterized by abnormal hematopoiesis. Among these myeloproliferative neoplasms, myelofibrosis has the most unfavorable prognosis. Furthermore, currently available therapies for myelofibrosis have little to no efficacy in the bone marrow and hence, are palliative. We recently developed a Janus kinase 2 (Jak2) small molecule inhibitor called G6 and found that it exhibits marked efficacy in a xenograft model of Jak2-V617F–mediated hyperplasia and a transgenic mouse model of Jak2-V617F–mediated polycythemia vera/essential thrombocytosis. However, its efficacy in Jak2-mediated myelofibrosis has not previously been examined. Here, we hypothesized that G6 would be efficacious in Jak2-V617F–mediated myelofibrosis. To test this, mice expressing the human Jak2-V617F cDNA under the control of the vav promoter were administered G6 or vehicle control solution, and efficacy was determined by measuring parameters within the peripheral blood, liver, spleen, and bone marrow. We found that G6 significantly reduced extramedullary hematopoiesis in the liver and splenomegaly. In the bone marrow, G6 significantly reduced pathogenic Jak/STAT signaling by 53%, megakaryocytic hyperplasia by 70%, and the Jak2 mutant burden by 68%. Furthermore, G6 significantly improved the myeloid to erythroid ratio and significantly reversed the myelofibrosis. Collectively, these results indicate that G6 is efficacious in Jak2-V617F–mediated myelofibrosis, and given its bone marrow efficacy, it may alter the natural history of this disease. PMID:22796437

Palmitoylethanolamide Blunts Amyloid-β42-Induced Astrocyte Activation and Improves Neuronal Survival in Primary Mouse Cortical Astrocyte-Neuron Co-Cultures.

PubMed

Beggiato, Sarah; Borelli, Andrea Celeste; Ferraro, Luca; Tanganelli, Sergio; Antonelli, Tiziana; Tomasini, Maria Cristina

2018-01-01

Based on the pivotal role of astrocytes in brain homeostasis and the strong metabolic cooperation existing between neurons and astrocytes, it has been suggested that astrocytic dysfunctions might cause and/or contribute to neuroinflammation and neurodegenerative processes. Therapeutic approaches aimed at both neuroprotection and neuroinflammation reduction may prove particularly effective in slowing the progression of these diseases. The endogenous lipid mediator palmitoylethanolamide (PEA) displayed neuroprotective and anti(neuro)inflammatory properties, and demonstrated interesting potential as a novel treatment for Alzheimer's disease. We firstly evaluated whether astrocytes could participate in regulating the Aβ42-induced neuronal damage, by using primary mouse astrocytes cell cultures and mixed astrocytes-neurons cultures. Furthermore, the possible protective effects of PEA against Aβ42-induced neuronal toxicity have also been investigated by evaluating neuronal viability, apoptosis, and morphometric parameters. The presence of astrocytes pre-exposed to Aβ42 (0.5μM; 24 h) induced a reduction of neuronal viability in primary mouse astrocytes-neurons co-cultures. Furthermore, under these experimental conditions, an increase in the number of neuronal apoptotic nuclei and a decrease in the number of MAP-2 positive neurons were observed. Finally, astrocytic Aβ42 pre-exposure induced an increase in the number of neurite aggregations/100μm as compared to control (i.e., untreated) astrocytes-neurons co-cultures. These effects were not observed in neurons cultured in the presence of astrocytes pre-exposed to PEA (0.1μM), applied 1 h before and maintained during Aβ42 treatment. Astrocytes contribute to Aβ42-induced neurotoxicity and PEA, by blunting Aβ42-induced astrocyte activation, improved neuronal survival in mouse astrocyte-neuron co-cultures.

ZIP4 silencing improves bone loss in pancreatic cancer

PubMed Central

Yang, Jingxuan; Ding, Hao; LeBrun, Drake; Ding, Kai; Houchen, Courtney W.; Postier, Russell G.; Ambrose, Catherine G.; Li, Zhaoshen; Bi, Xiaohong; Li, Min

2015-01-01

Metabolic bone disorders are associated with several types of human cancers. Pancreatic cancer patients usually suffer from severe nutrition deficiency, muscle wasting, and loss of bone mass. We have previously found that silencing of a zinc transporter ZIP4 prolongs the survival and reduces the severity of the cachexia in vivo. However, the role of ZIP4 in the pancreatic cancer related bone loss remains unknown. In this study we investigated the effect of ZIP4 knockdown on the bone structure, composition and mechanical properties of femurs in an orthotopic xenograft mouse model. Our data showed that silencing of ZIP4 resulted in increased bone tissue mineral density, decreased bone crystallinity and restoration of bone strength through the RANK/RANKL pathway. The results further support the impact of ZIP4 on the progression of pancreatic cancer, and suggest its potential significance as a therapeutic target for treating patients with such devastating disease and cancer related disorders. PMID:26305676

Bone morphogenetic protein and bone metastasis, implication and therapeutic potential.