Insulin-Like Growth Factor I Does Not Drive New Bone Formation in Experimental Arthritis.

Introduction: Insulin like growth factor (IGF)-I can act on a variety of cells involved in cartilage and bone repair, yet IGF-I has not been studied extensively in the context of inflammatory arthritis. The objective of this study was to investigate whether IGF-I overexpression in the osteoblast lineage could lead to increased reparative or pathological bone formation in rheumatoid arthritis and/or spondyloarthritis respectively.

Methods: Mice overexpressing IGF-I in the osteoblast lineage (Ob-IGF-I+/-) line 324-7 were studied during collagen induced arthritis and in the DBA/1 aging model for ankylosing enthesitis.

Combined experience of six independent laboratories attempting to create an Ewing sarcoma mouse model.

Ewing sarcoma (ES) involves a tumor-specific chromosomal translocation that produces the EWS-FLI1 protein, which is required for the growth of ES cells both in vitro and in vivo. However, an EWS-FLI1-driven transgenic mouse model is not currently available. Here, we present data from six independent laboratories seeking an alternative approach to express EWS-FLI1 in different murine tissues.

Impact of targeted PPARγ disruption on bone remodeling.

Peroxisome proliferator-activated receptor gamma (PPARγ), known as the master regulator of adipogenesis, has been regarded as a promising target for new anti-osteoporosis therapy due to its role in regulating bone marrow mesenchymal stem/progenitor cell (BMSC) lineage commitment. However, the precise mechanism underlying PPARγ regulation of bone is not clear as a bone-specific PPARγ conditional knockout (cKO) study has not been conducted and evidence showed that deletion of PPARγ in other tissues also have profound effect on bone. In this study, we show that mice deficiency of PPARγ in cells expressing a 3.

Regulatory T-cells and cAMP suppress effector T-cells independently of PKA-CREM/ICER: a potential role for Epac.

cAMP signalling is both a major pathway as well as a key therapeutic target for inducing immune tolerance and is involved in Treg cell (regulatory T-cell) function. To achieve potent immunoregulation, cAMP can act through several downstream effectors. One proposed mechanism is that cAMP-mediated suppression, including immunosuppression by Treg cells, results from activation of PKA (protein kinase A) leading to the induction of the transcription factor ICER (inducible cAMP early repressor).

Bmp2 in osteoblasts of periosteum and trabecular bone links bone formation to vascularization and mesenchymal stem cells.

We generated a new Bmp2 conditional-knockout allele without a neo cassette that removes the Bmp2 gene from osteoblasts (Bmp2-cKO(ob)) using the 3.6Col1a1-Cre transgenic model. Bones of Bmp2-cKO(ob) mice are thinner, with increased brittleness.

Forum on aging and skeletal health: summary of the proceedings of an ASBMR workshop.

With the aging of the population, the scope of the problem of age-related bone loss and osteoporosis will continue to increase. As such, it is critical to obtain a better understanding of the factors determining the acquisition and loss of bone mass from childhood to senescence. While there have been significant advances in recent years in our understanding of both the basic biology of aging and a clinical definition of age-related frailty, few of these concepts in aging research have been evaluated adequately for their relevance and application to skeletal aging or fracture prevention.

Selective osteoblast overexpression of IGF-I in mice prevents low protein-induced deterioration of bone strength and material level properties.

Protein deficiency is frequently observed in elderly osteoporotic patients. Undernutrition leads to decreased levels of IGF-I, an important factor in regulating bone homeostasis throughout life. IGF-I is produced in the liver and locally in the skeleton.

Epidermal growth factor receptor plays an anabolic role in bone metabolism in vivo.

While the epidermal growth factor receptor (EGFR)-mediated signaling pathway has been shown to have vital roles in many developmental and pathologic processes, its functions in the development and homeostasis of the skeletal system has been poorly defined. To address its in vivo role, we constructed transgenic and pharmacologic mouse models and used peripheral quantitative computed tomography (pQCT), micro-computed tomography (µCT) and histomorphometry to analyze their trabecular and cortical bone phenotypes. We initially deleted the EGFR in preosteoblasts/osteoblasts using a Cre/loxP system (Col-Cre Egfr(f/f)), but no bone phenotype was observed because of incomplete deletion of the Egfr genomic locus.

PDE8 regulates rapid Teff cell adhesion and proliferation independent of ICER.

Background: Abolishing the inhibitory signal of intracellular cAMP by phosphodiesterases (PDEs) is a prerequisite for effector T (Teff) cell function. While PDE4 plays a prominent role, its control of cAMP levels in Teff cells is not exclusive. T cell activation has been shown to induce PDE8, a PDE isoform with 40- to 100-fold greater affinity for cAMP than PDE4.

Col3.6-HSD2 transgenic mice: a glucocorticoid loss-of-function model spanning early and late osteoblast differentiation.

The goal of this study was to characterize the bone phenotype and molecular alterations in Col3.6-HSD2 mice in which a 3.6-kb Col1a1 promoter fragment drives 11beta-HSD2 expression broadly in the osteoblast lineage to reduce glucocorticoid signaling.

Dicer inactivation in osteoprogenitor cells compromises fetal survival and bone formation, while excision in differentiated osteoblasts increases bone mass in the adult mouse.

MicroRNA attenuation of protein translation has emerged as an important regulator of mesenchymal cell differentiation into the osteoblast lineage. A compelling question is the extent to which miR biogenesis is obligatory for bone formation. Here we show conditional deletion of the Dicer enzyme in osteoprogenitors by Col1a1-Cre compromised fetal survival after E14.

Parathyroid hormone induction of cyclooxygenase-2 in murine osteoblasts: role of the calcium-calcineurin-NFAT pathway.

Murine MC3T3-E1 and MC-4 cells were stably transfected with -371/+70 bp of the murine cyclooxygenase-2 (COX-2) promoter fused to a luciferase reporter (Pluc371) or with Pluc371 carrying site-directed mutations. Mutations were made in (1) the cAMP response element (CRE) at -57/-52 bp, (2) the activating protein-1 (AP-1)-binding site at -69/-63 bp, (3) the nuclear factor of activated T-cells (NFAT)-binding site at -77/-73 bp, and (4) both the AP-1 and NFAT sites, which comprise a composite consensus sequence for NFAT/AP-1. Single mutation of CRE, AP-1, or NFAT sites decreased parathyroid hormone (PTH)-stimulated COX-2 promoter activity 40% to 60%, whereas joint mutation of NFAT and AP-1 abrogated the induction.

Effects of global or targeted deletion of the EP4 receptor on the response of osteoblasts to prostaglandin in vitro and on bone histomorphometry in aged mice.

Because global deletion of the prostaglandin EP4 receptor results in neonatal lethality, we generated a mouse with targeted EP4 receptor deletion using Cre-LoxP methodology and a 2.3 kb collagen I a1 promoter driving Cre recombinase that is selective for osteoblastic cells. We compared wild type (WT), global heterozygote (G-HET), targeted heterozygote (T-HET) and knockout (KO) mice.

Calcitonin induces expression of the inducible cAMP early repressor in osteoclasts.

The cAMP response element modulator gene (Crem) encodes a variety of transcriptional regulators including the inducible cAMP early repressor, ICER. We previously showed that Crem knockout mice, which are deficient in CREM and ICER factors, display slightly increased long bone mass and decreased osteoclast number. These data are consistent with the notion that Crem regulates bone mass in part through an effect on osteoclast formation and/or function.

Osteopenia in transgenic mice with osteoblast-targeted expression of the inducible cAMP early repressor.

ICER is a member of the CREM family of basic leucine zipper transcription factors that acts as a dominant negative regulator of gene transcription. Four different isoforms of ICER (I, Igamma, II and IIgamma) are transcribed from the P2 promoter of the Crem gene. We previously found that each of the ICER isoforms is induced by parathyroid hormone in osteoblasts.

CREM deficiency in mice alters the response of bone to intermittent parathyroid hormone treatment.

CREM belongs to the ATF/CREB family of basic leucine zipper transcription factors. We previously showed that PTH induces ICER (inducible cAMP early repressor) in osteoblasts. ICER proteins, which are transcribed from the P2 promoter of the Crem gene, act as transcriptional attenuators.

Rearrangement of a conditional allele regardless of inheritance of a Cre recombinase transgene.

To generate conditional gene knockouts in osteoblasts, we previously developed transgenic mice in which Cre recombinase cDNA was cloned downstream of a 3.6 or 2.3 kb fragment of the rat Col1a1 promoter (Col3.

Osteoblast deletion of exon 3 of the androgen receptor gene results in trabecular bone loss in adult male mice.

Unlabelled: The mechanism of androgen action on bone was studied in male mice with the AR deleted in mature osteoblasts. These mice had decreased trabecular bone volume associated with a decrease in trabecular number, suggesting that androgens may act directly on osteoblasts to maintain trabecular bone.

Introduction: Androgens modulate bone cell activity and are important for the maintenance of bone mass.

Analysis of internal deletions of a rat Col1a1 promoter fragment in transfected ROS17/2.8 cells.

The aim of this paper is identification of regulatory sequences downstream of -1683 base pairs (bp) in the rat Col1a1 promoter important for expression in osteoblasts. Previous findings suggest that a rat Col1a1 gene fragment extending from -1719 to + 115 bp linked to the chloramphenicol acetyl transferase (CAT) reporter gene (ColCAT1719) is highly and selectively expressed in osteoblasts. Three internal deletions within the ColCAT1719 construct were generated and stably transfected into ROS 17/2.

Transgenic mice with osteoblast-targeted insulin-like growth factor-I show increased bone remodeling.

To determine the effects of locally-expressed insulin-like growth factor (IGF-I) on bone remodeling, a transgene was produced in which murine IGF-I cDNA was cloned downstream of a gene fragment comprising 3.6 kb of 5' upstream regulatory sequence and most of the first intron of the rat Col1a1 gene. The construct was expressed at the mRNA and protein level in transfected osteoblasts.

Osteoblast differentiation and skeletal development are regulated by Mdm2-p53 signaling.

Mdm2 is required to negatively regulate p53 activity at the peri-implantation stage of early mouse development. However, the absolute requirement for Mdm2 throughout embryogenesis and in organogenesis is unknown. To explore Mdm2-p53 signaling in osteogenesis, Mdm2-conditional mice were bred with Col3.

Postnatal growth and bone mass in mice with IGF-I haploinsufficiency.

We examined the influence of IGF-I haploinsufficiency on growth, bone mass and osteoblast differentiation in Igf1 heterozygous knockout (HET) mice. Cohorts of male and female wild type (WT) and HET mice in the outbred CD-1 background were analyzed at 1, 2, 4, 8, 12, 15 and 18 months of age for body weight, serum IGF-I and bone morphometry. Compared to WT mice, HET mice had 20-30% lower serum IGF-I levels in both genders and in all age groups.

Expression and activity of osteoblast-targeted Cre recombinase transgenes in murine skeletal tissues.

The Cre/loxP recombination system can be used to circumvent many of the limitations of generalized gene ablation in mice. Here we present the development and characterization of transgenic mice in which Cre recombinase has been targeted to cells of the osteoblast lineage with 2.3 kb (Col 2.

Transgenic expression of 11beta-hydroxysteroid dehydrogenase type 2 in osteoblasts reveals an anabolic role for endogenous glucocorticoids in bone.

Glucocorticoid excess leads to bone loss, primarily by decreasing bone formation. However, a variety of in vitro models show that glucocorticoids can promote osteogenesis. To elucidate the role of endogenous glucocorticoids in bone metabolism, we developed transgenic (TG) mice in which a 2.