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Chondrocytes-Specific Expression of Osteoprotegerin Modulates Osteoclast Formation in Metaphyseal Bone.

Wang B, Jin H, Shu B, Mira RR, Chen D - Sci Rep (2015)

Bottom Line: TRAP staining showed that osteoclast numbers were reduced in both secondary ossification center and proximal metaphysis.In contrast, the mineral apposition rates were not changed in Col2-Opg transgenic mice.Our results demonstrated that OPG expression in chondrocyte increases bone mass in the proximal metaphysis of tibiae through negative regulation of osteoclast formation.

View Article: PubMed Central - PubMed

Affiliation: Key Lab of Hormone and Development (Ministry of Health), Metabolic Diseases Hospital and Tianjin Institute of Endocrinology, Tianjin Medical University, Tianjin 300070, China.

ABSTRACT
Bone marrow stromal cells/osteoblasts were originally thought to be the major player in regulating osteoclast differentiation through expressing RANKL/OPG cytokines. Recent studies have established that chondrocytes also express RANKL/OPG and support osteoclast formation. Till now, the in vivo function of chondrocyte-produced OPG in osteoclast formation and postnatal bone growth has not been directly investigated. In this study, chondrocyte-specific Opg transgenic mice were generated by using type II collagen promoter. The Col2-Opg transgenic mice showed delayed formation of secondary ossification center and localized increase of bone mass in proximal metaphysis of tibiae. TRAP staining showed that osteoclast numbers were reduced in both secondary ossification center and proximal metaphysis. This finding was further confirmed by in vitro chondrocyte/spleen cell co-culture assay. In contrast, the mineral apposition rates were not changed in Col2-Opg transgenic mice. TUNEL staining revealed more apoptotic hypertrophic chondrocytes in the growth plate of Col2-Opg mice. Flow cytometry analysis showed fewer RANK-expressing cells in the marrow of Col2a1-Opg mice, suggesting the role of OPG in blocking the differentiation of early mesenchymal progenitors into RANK-expressing pre-osteoclasts. Our results demonstrated that OPG expression in chondrocyte increases bone mass in the proximal metaphysis of tibiae through negative regulation of osteoclast formation.

No MeSH data available.


Related in: MedlinePlus

Opg-Flag transgene was expressed in chondrocytes in Col2-Opg mice.(A) The diagram shows the Col2-Opg transgene construct. (B) Expression of OPG-Flag protein was detected in chondrocytes derived from Col2-Opg transgenic mice by Western blot analysis using the anti-Flag antibody. (C) Expression of Opg-Flag mRNA was examined in multiple tissues by RT-PCR using the transgene specific primers. (D–G) Expression levels of Opg mRNA and protein in primary sternal chondrocytes, calvarial pre-osteoblasts and bone marrow stromal cells were examined. Data are presented as means ± SD (n = 3–4 per group).
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f1: Opg-Flag transgene was expressed in chondrocytes in Col2-Opg mice.(A) The diagram shows the Col2-Opg transgene construct. (B) Expression of OPG-Flag protein was detected in chondrocytes derived from Col2-Opg transgenic mice by Western blot analysis using the anti-Flag antibody. (C) Expression of Opg-Flag mRNA was examined in multiple tissues by RT-PCR using the transgene specific primers. (D–G) Expression levels of Opg mRNA and protein in primary sternal chondrocytes, calvarial pre-osteoblasts and bone marrow stromal cells were examined. Data are presented as means ± SD (n = 3–4 per group).

Mentions: In this study, we generated Col2-Opg transgenic mice in which expression of the Opg-Flag transgene was targeted to chondrocytes using the 1.0 kb type II collagen promoter (Col2a1) (Fig. 1A)192021. Two independent lines of Col2-Opg transgenic mice were established and both of them displayed similar phenotypes. Col2-Opg transgenic mice are viable, fertile with normal body size and have no any gross, physical, or behavioral abnormalities.


Chondrocytes-Specific Expression of Osteoprotegerin Modulates Osteoclast Formation in Metaphyseal Bone.

Wang B, Jin H, Shu B, Mira RR, Chen D - Sci Rep (2015)

Opg-Flag transgene was expressed in chondrocytes in Col2-Opg mice.(A) The diagram shows the Col2-Opg transgene construct. (B) Expression of OPG-Flag protein was detected in chondrocytes derived from Col2-Opg transgenic mice by Western blot analysis using the anti-Flag antibody. (C) Expression of Opg-Flag mRNA was examined in multiple tissues by RT-PCR using the transgene specific primers. (D–G) Expression levels of Opg mRNA and protein in primary sternal chondrocytes, calvarial pre-osteoblasts and bone marrow stromal cells were examined. Data are presented as means ± SD (n = 3–4 per group).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556963&req=5

f1: Opg-Flag transgene was expressed in chondrocytes in Col2-Opg mice.(A) The diagram shows the Col2-Opg transgene construct. (B) Expression of OPG-Flag protein was detected in chondrocytes derived from Col2-Opg transgenic mice by Western blot analysis using the anti-Flag antibody. (C) Expression of Opg-Flag mRNA was examined in multiple tissues by RT-PCR using the transgene specific primers. (D–G) Expression levels of Opg mRNA and protein in primary sternal chondrocytes, calvarial pre-osteoblasts and bone marrow stromal cells were examined. Data are presented as means ± SD (n = 3–4 per group).
Mentions: In this study, we generated Col2-Opg transgenic mice in which expression of the Opg-Flag transgene was targeted to chondrocytes using the 1.0 kb type II collagen promoter (Col2a1) (Fig. 1A)192021. Two independent lines of Col2-Opg transgenic mice were established and both of them displayed similar phenotypes. Col2-Opg transgenic mice are viable, fertile with normal body size and have no any gross, physical, or behavioral abnormalities.

Bottom Line: TRAP staining showed that osteoclast numbers were reduced in both secondary ossification center and proximal metaphysis.In contrast, the mineral apposition rates were not changed in Col2-Opg transgenic mice.Our results demonstrated that OPG expression in chondrocyte increases bone mass in the proximal metaphysis of tibiae through negative regulation of osteoclast formation.

View Article: PubMed Central - PubMed

Affiliation: Key Lab of Hormone and Development (Ministry of Health), Metabolic Diseases Hospital and Tianjin Institute of Endocrinology, Tianjin Medical University, Tianjin 300070, China.

ABSTRACT
Bone marrow stromal cells/osteoblasts were originally thought to be the major player in regulating osteoclast differentiation through expressing RANKL/OPG cytokines. Recent studies have established that chondrocytes also express RANKL/OPG and support osteoclast formation. Till now, the in vivo function of chondrocyte-produced OPG in osteoclast formation and postnatal bone growth has not been directly investigated. In this study, chondrocyte-specific Opg transgenic mice were generated by using type II collagen promoter. The Col2-Opg transgenic mice showed delayed formation of secondary ossification center and localized increase of bone mass in proximal metaphysis of tibiae. TRAP staining showed that osteoclast numbers were reduced in both secondary ossification center and proximal metaphysis. This finding was further confirmed by in vitro chondrocyte/spleen cell co-culture assay. In contrast, the mineral apposition rates were not changed in Col2-Opg transgenic mice. TUNEL staining revealed more apoptotic hypertrophic chondrocytes in the growth plate of Col2-Opg mice. Flow cytometry analysis showed fewer RANK-expressing cells in the marrow of Col2a1-Opg mice, suggesting the role of OPG in blocking the differentiation of early mesenchymal progenitors into RANK-expressing pre-osteoclasts. Our results demonstrated that OPG expression in chondrocyte increases bone mass in the proximal metaphysis of tibiae through negative regulation of osteoclast formation.

No MeSH data available.


Related in: MedlinePlus