Temporal expression of PTHrP during endochondral bone formation in mouse and intramembranous bone formation in an in vivo rabbit model

Objective To support the preclinical evaluation of therapeutics that target chondrogenesis, our goal was to generate a rat strain that can noninvasively report endogenous chondrogenic activity. Design A transgene was constructed in which the dual expression of bioluminescent (firefly luciferase) and fluorescent (mCherry) reporters is controlled by regulatory sequences from rat Col2a1. Candidate lines were established on a Lewis background and characterized by serial bioluminescence imaging as well as ex vivo measurement of molecular reporter levels in several tissues. The sensitivity and specificity of the reporter strain were assessed in models of orthotopic and ectopic chondrogenesis. Results Substantial bioluminescence signal was detected from cartilaginous regions, including the appendicular synovial joints, spine, sternum, nose, and pinnae. Bioluminescent radiance was intense at 1 month of age, rapidly declined with continued development, yet remained detectable in 2-year-old animals. Explant imaging and immunohistochemistry confirmed that both molecular reporters were localized to cartilage. Implantation of wild-type bone marrow stromal cells into osteochondral defects made in both young adult and aged reporter rats led to a time-dependent elevation of intra-articular reporter activity concurrent with cartilaginous tissue repair. To stimulate ectopic, endochondral bone formation, bone morphogenetic protein 2 was overexpressed in the gastrocnemius muscle, which led to bioluminescent signal that closely preceded heterotopic ossification. Conclusions This strain can help develop strategies to stimulate cartilage repair and endochondral bone formation or to inhibit chondrogenesis associated with heterotopic ossification.

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Recombinant Vgr-1/BMP-6-expressing tumors induce fibrosis and endochondral bone formation in vivo.

The Journal of Cell Biology ◽

10.1083/jcb.126.6.1595 ◽

1994 ◽

Vol 126 (6) ◽

pp. 1595-1609 ◽

Cited By ~ 119

Author(s):

S E Gitelman ◽

M S Kobrin ◽

J Q Ye ◽

A R Lopez ◽

A Lee ◽

...

Keyword(s):

Bone Formation ◽

Tumor Cells ◽

Cho Cells ◽

Subcutaneous Tissue ◽

Endochondral Bone Formation ◽

Related Factor ◽

Tgf Beta ◽

Endochondral Bone ◽

Hypertrophic Cartilage

Members of the TGF-beta superfamily appear to modulate mesenchymal differentiation, including the processes of cartilage and bone formation. Nothing is yet known about the function of the TGF-beta-related factor vgr-1, also called bone morphogenetic protein-6 (BMP-6), and only limited studies have been conducted on the most closely related factors BMP-5, osteogenic protein-1 (OP-1) or BMP-7, and OP-2. Because vgr-1 mRNA has been localized in hypertrophic cartilage, this factor may play a vital role in endochondral bone formation. We developed antibodies to vgr-1, and documented that vgr-1 protein was expressed in hypertrophic cartilage of mice. To further characterize the role of this protein in bone differentiation, we generated CHO cells that overexpressed recombinant murine vgr-1 protein. Western blot analysis documented that recombinant vgr-1 protein was secreted into the media and was proteolytically processed to yield the mature vgr-1 molecule. To assess the biological activity of recombinant vgr-1 in vivo, we introduced the vgr-1-expressing CHO cells directly into the subcutaneous tissue of athymic nude mice. CHO-vgr-1 cells produced localized tumors, and the continuous secretion of vgr-1 resulted in tumors with a strikingly different gross and histological appearance as compared to the parental CHO cells. The tumors of control CHO cells were hemorrhagic, necrotic, and friable, whereas the CHO-vgr-1 tumors were dense, firm, and fibrotic. In contrast with control CHO tumors, the nests of CHO-vgr-1 tumor cells were surrounded by extensive connective tissue, which contained large regions of cartilage and bone. Further analysis indicated that secretion of vgr-1 from the transfected CHO tumor cells induced the surrounding host mesenchymal cells to develop along the endochondral bone pathway. These findings suggest that endochondral bone formation.

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Bone Marrow Mesenchymal Stem Cells Form Ectopic Woven Bone In Vivo Through Endochondral Bone Formation

Artificial Organs ◽

10.1111/j.1525-1594.2009.00728.x ◽

2009 ◽

Vol 33 (4) ◽

pp. 301-308 ◽

Cited By ~ 16

Author(s):

Sophia Chia-Ning Chang ◽

Ching-Lung Tai ◽

Hui-Ying Chung ◽

Tsung-Min Lin ◽

Long-Bin Jeng

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Bone Formation ◽

Endochondral Bone Formation ◽

Endochondral Bone ◽

Woven Bone ◽

Marrow Mesenchymal Stem Cells

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Selective Deficiency of the “Bone-related”Runx2-II Unexpectedly Preserves Osteoblast-mediated Skeletogenesis

Journal of Biological Chemistry ◽

10.1074/jbc.m401109200 ◽

2004 ◽

Vol 279 (19) ◽

pp. 20307-20313 ◽

Cited By ~ 66

Author(s):

Zhou-Sheng Xiao ◽

Anita B. Hjelmeland ◽

L. D. Quarles

Keyword(s):

Bone Formation ◽

Ex Vivo ◽

Mesenchymal Cells ◽

Endochondral Bone Formation ◽

Endochondral Bone ◽

Chondrocyte Maturation ◽

Intramembranous Bone ◽

Exon 1 ◽

Metaphyseal Bone ◽

Deficient Mice

Runx2 (runt-related transcription factor 2) is a master regulator of skeletogenesis. Distinct promoters in the Runx2 gene transcribe the “bone-related”Runx2-II and non-osseousRunx2-I isoforms that differ only in their respective N termini. Existing mutant mouse models with both isoforms deleted exhibit an arrest of osteoblast and chondrocyte maturation and the complete absence of mineralized bone, but they do not distinguish the separate functions of the two N-terminal isoforms. To elucidate the function of the bone-related isoform, we generated selectiveRunx2-II-deficient mice by the targeted deletion of the distal promoter and exon 1. HomozygousRunx2-II-deficient (Runx2-II-/-) mice unexpectedly formed axial, appendicular, and craniofacial bones derived from either intramembranous ossification or mesenchymal cells of the bone collar, but they failed to form the posterior cranium and other bones derived from endochondral ossification. HeterozygousRunx2-II-deficient mice had grossly normal skeletons, but were osteopenic. The commitment of mesenchymal cellsex vivoto the osteoblast lineage occurred inRunx2-II-/-mice, but osteoblastic gene expression was impaired. Chondrocyte maturation appeared normal, but the zone of hypertrophic chondrocytes was not transformed into metaphyseal bone, leading to widened growth plates inRunx2-II-/-mice. Compensatory increments inRunx2-I expression occurred inRunx2-II-/-mice but were not sufficient to normalize osteoblastic maturation or transcriptional activity. Our findings support distinct functions ofRunx2-II and -I in the control of skeletogenesis.Runx2-I is sufficient for early osteoblastogenesis and intramembranous bone formation, whereasRunx2-II is necessary for complete osteoblastic maturation and endochondral bone formation.

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The distribution of stanniocalcin 1 protein in fetal mouse tissues suggests a role in bone and muscle development

Journal of Endocrinology ◽

10.1677/joe.0.1650457 ◽

2000 ◽

Vol 165 (2) ◽

pp. 457-466 ◽

Cited By ~ 36

Author(s):

WQ Jiang ◽

AC Chang ◽

M Satoh ◽

Y Furuichi ◽

PP Tam ◽

...

Keyword(s):

Bone Formation ◽

Muscle Development ◽

Myogenic Differentiation ◽

Endochondral Bone Formation ◽

Endochondral Bone ◽

Mouse Fetus ◽

Developing Heart ◽

Intramembranous Bone ◽

Mouse Tissues ◽

Specific Localization

We previously isolated a mammalian gene STC1 that encodes a glycoprotein related to stanniocalcin (STC), a fish hormone that plays a major role in calcium homeostasis. However, the mammalian STC1 gene is expressed in a variety of adult tissues in contrast to fish where STC is expressed only in one unique gland, the corpuscles of Stannius. This suggested that STC1 may have wider autocrine/paracrine functions in mammals. In the present study, using immunocytochemistry, we showed that STC1 protein is localized in the developing bone and muscle of the mouse fetus. During endochondral bone formation, STC1 is found principally in prechondrocytes and prehypertrophic chondrocytes. During intramembranous bone formation STC1 is present in the mesenchyme that is about to undergo ossification. STC1 is also found in the myocardiocytes of the developing heart and at all stages of differentiation from myoblasts to myotube formation in developing skeletal muscle. The specific localization of STC1 to chondrocytes and muscle cells suggests a role for this protein in chondrogenic and myogenic differentiation.

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Ablation of Ephrin B2 in Col2 Expressing Cells Delays Fracture Repair

Endocrinology ◽

10.1210/endocr/bqaa179 ◽

2020 ◽

Vol 161 (12) ◽

Author(s):

Yongmei Wang ◽

Lin Ling ◽

Faming Tian ◽

Sun Hee Won Kim ◽

Sunita Ho ◽

...

Keyword(s):

Bone Formation ◽

Cell Activation ◽

Bone Development ◽

Type Ii Collagen ◽

Fracture Site ◽

Fracture Repair ◽

Endochondral Bone Formation ◽

Endochondral Bone ◽

Intramembranous Bone ◽

Endothelial Growth Factor

Abstract Ephrin B2 is critical for endochondral bone development. In this study, we investigated its role in fracture repair by deleting ephrin B2 in type II collagen (Col.2) expressing cells. We used a nonstable tibia fracture model to evaluate fracture repair at 3 sites: intramembranous bone formation, endochondral bone formation, and intramedullary bone formation. We observed that during fracture repair, deletion of ephrin B2 impaired periosteal stem cell activation, inhibited their proliferation, decreased their survival, and blocked their differentiation into osteoblasts and chondrocytes. In addition, deletion of ephrin B2 decreased vascular endothelial growth factor production as well as vascular invasion into the fracture site. These changes led to reduced cartilage to bone conversion in the callus with decreased new bone formation, resulting in impaired fracture repair. Our data indicate that ephrin B2 in Col2-expressing cells is a critical regulator of fracture repair, pointing to a new and potentially targetable mechanism to enhance fracture repair.

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