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Involvement of Prolyl Hydroxylase Domain Protein in the Rosiglitazone-Induced Suppression of Osteoblast Differentiation.

Kang JH, Kwak HJ, Choi HE, Kim J, Hong S, Kim OH, Oh BC, Cheon HG - PLoS ONE (2015)

Bottom Line: PHD inhibitors and knockdown of each isoform prevented the inhibitory effects of rosiglitazone on osteoblast differentiation and increased the expression of Runx2, a transcription factor essential for osteoblastogenesis.Furthermore, both increased PHD isoform expressions and reduced osteoblast differentiation by rosiglitazone were prevented by PPARγ antagonists, indicating these effects were mediated via PPARγ activation.In vivo oral administration of rosiglitazone to female ICR mice for 8 weeks reduced bone mineral densities and plasma alkaline phosphatase (ALP) activity, and increased PHD expression in femoral primary bone marrow cells and the ubiquitination of Runx2.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Medicine, Gachon University, Incheon, Republic of Korea.

ABSTRACT
Rosiglitazone is a well-known anti-diabetic drug that increases insulin sensitivity via peroxisome proliferator-activated receptor γ (PPARγ) activation, but unfortunately it causes bone loss in animals and humans. A previous study showed that prolyl hydroxylase domain protein (PHD) plays a role in rosiglitazone-induced adipocyte differentiation. Based on the inverse relationship between adipocyte and osteoblast differentiation, we investigated whether PHD is involved in the effects of rosiglitazone on osteoblast differentiation. Rosiglitazone inhibited osteoblast differentiation in a concentration-dependent manner, and in parallel induced three PHD isoforms (PHD1, 2, and 3). PHD inhibitors and knockdown of each isoform prevented the inhibitory effects of rosiglitazone on osteoblast differentiation and increased the expression of Runx2, a transcription factor essential for osteoblastogenesis. MG-132, a proteasomal inhibitor also prevented the rosiglitazone-induced degradation of Runx2. Furthermore, both increased PHD isoform expressions and reduced osteoblast differentiation by rosiglitazone were prevented by PPARγ antagonists, indicating these effects were mediated via PPARγ activation. In vivo oral administration of rosiglitazone to female ICR mice for 8 weeks reduced bone mineral densities and plasma alkaline phosphatase (ALP) activity, and increased PHD expression in femoral primary bone marrow cells and the ubiquitination of Runx2. Together, this suggests that the rosiglitazone-induced suppression of osteoblast differentiation is at least partly induced via PPARγ-mediated PHD induction and subsequent promotion of the ubiquitination and degradation of Runx2.

No MeSH data available.


Related in: MedlinePlus

Effects of PPARγ antagonists on rosiglitazone-induced adipocyte differentiation.Primary bone marrow cells were treated with BADGE (20 μM) or GW9662 (1 μM) in the presence of rosiglitazone (10 μM) for 12 days. Osteoblast differentiation was determined by Alizarin red staining (A), and the mRNA levels of PHD1, 2, and 3, and osteogenic marker genes were analyzed by RT-PCR (B). Densitometric analysis was conducted using UN-SCAN-IT gel ver. 5.1 software (Silk Scientific) and is expressed as means ± SEMs. *P<0.05 vs control (C), #P<0.05 vs differentiated (D), ✝P<0.05 vs DR. Experiments were conducted three times and representative results are shown. C, control (no differentiation); D, differentiated in DAG medium; DR, DAG in the presence of rosiglitazone; BD, BADGE; GW, GW9662.
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pone.0139093.g005: Effects of PPARγ antagonists on rosiglitazone-induced adipocyte differentiation.Primary bone marrow cells were treated with BADGE (20 μM) or GW9662 (1 μM) in the presence of rosiglitazone (10 μM) for 12 days. Osteoblast differentiation was determined by Alizarin red staining (A), and the mRNA levels of PHD1, 2, and 3, and osteogenic marker genes were analyzed by RT-PCR (B). Densitometric analysis was conducted using UN-SCAN-IT gel ver. 5.1 software (Silk Scientific) and is expressed as means ± SEMs. *P<0.05 vs control (C), #P<0.05 vs differentiated (D), ✝P<0.05 vs DR. Experiments were conducted three times and representative results are shown. C, control (no differentiation); D, differentiated in DAG medium; DR, DAG in the presence of rosiglitazone; BD, BADGE; GW, GW9662.

Mentions: To determine whether the effect of rosiglitazone on osteoblast differentiation is mediated by PPARγ, we examined the effects of two well known PPARγ antagonists, that is, BADGE and GW9662. Both antagonists partially inhibited the rosiglitazone-induced suppression of osteoblast differentiation (Fig 5A), and inhibited rosiglitazone-induced PHD isotype expressions (Fig 5B). As shown in Fig 5B, the mRNA expression of Runx2, ALP and osterix decreased when primary bone marrow cells were treated with rosiglitazone, and this decrease prevented by the presence of BADGE or GW9662. Together, these results suggest that rosiglitazone-induced PHD upregulation results from the activation of PPARγ by rosiglitazone.


Involvement of Prolyl Hydroxylase Domain Protein in the Rosiglitazone-Induced Suppression of Osteoblast Differentiation.

Kang JH, Kwak HJ, Choi HE, Kim J, Hong S, Kim OH, Oh BC, Cheon HG - PLoS ONE (2015)

Effects of PPARγ antagonists on rosiglitazone-induced adipocyte differentiation.Primary bone marrow cells were treated with BADGE (20 μM) or GW9662 (1 μM) in the presence of rosiglitazone (10 μM) for 12 days. Osteoblast differentiation was determined by Alizarin red staining (A), and the mRNA levels of PHD1, 2, and 3, and osteogenic marker genes were analyzed by RT-PCR (B). Densitometric analysis was conducted using UN-SCAN-IT gel ver. 5.1 software (Silk Scientific) and is expressed as means ± SEMs. *P<0.05 vs control (C), #P<0.05 vs differentiated (D), ✝P<0.05 vs DR. Experiments were conducted three times and representative results are shown. C, control (no differentiation); D, differentiated in DAG medium; DR, DAG in the presence of rosiglitazone; BD, BADGE; GW, GW9662.
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Related In: Results  -  Collection

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pone.0139093.g005: Effects of PPARγ antagonists on rosiglitazone-induced adipocyte differentiation.Primary bone marrow cells were treated with BADGE (20 μM) or GW9662 (1 μM) in the presence of rosiglitazone (10 μM) for 12 days. Osteoblast differentiation was determined by Alizarin red staining (A), and the mRNA levels of PHD1, 2, and 3, and osteogenic marker genes were analyzed by RT-PCR (B). Densitometric analysis was conducted using UN-SCAN-IT gel ver. 5.1 software (Silk Scientific) and is expressed as means ± SEMs. *P<0.05 vs control (C), #P<0.05 vs differentiated (D), ✝P<0.05 vs DR. Experiments were conducted three times and representative results are shown. C, control (no differentiation); D, differentiated in DAG medium; DR, DAG in the presence of rosiglitazone; BD, BADGE; GW, GW9662.
Mentions: To determine whether the effect of rosiglitazone on osteoblast differentiation is mediated by PPARγ, we examined the effects of two well known PPARγ antagonists, that is, BADGE and GW9662. Both antagonists partially inhibited the rosiglitazone-induced suppression of osteoblast differentiation (Fig 5A), and inhibited rosiglitazone-induced PHD isotype expressions (Fig 5B). As shown in Fig 5B, the mRNA expression of Runx2, ALP and osterix decreased when primary bone marrow cells were treated with rosiglitazone, and this decrease prevented by the presence of BADGE or GW9662. Together, these results suggest that rosiglitazone-induced PHD upregulation results from the activation of PPARγ by rosiglitazone.

Bottom Line: PHD inhibitors and knockdown of each isoform prevented the inhibitory effects of rosiglitazone on osteoblast differentiation and increased the expression of Runx2, a transcription factor essential for osteoblastogenesis.Furthermore, both increased PHD isoform expressions and reduced osteoblast differentiation by rosiglitazone were prevented by PPARγ antagonists, indicating these effects were mediated via PPARγ activation.In vivo oral administration of rosiglitazone to female ICR mice for 8 weeks reduced bone mineral densities and plasma alkaline phosphatase (ALP) activity, and increased PHD expression in femoral primary bone marrow cells and the ubiquitination of Runx2.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Medicine, Gachon University, Incheon, Republic of Korea.

ABSTRACT
Rosiglitazone is a well-known anti-diabetic drug that increases insulin sensitivity via peroxisome proliferator-activated receptor γ (PPARγ) activation, but unfortunately it causes bone loss in animals and humans. A previous study showed that prolyl hydroxylase domain protein (PHD) plays a role in rosiglitazone-induced adipocyte differentiation. Based on the inverse relationship between adipocyte and osteoblast differentiation, we investigated whether PHD is involved in the effects of rosiglitazone on osteoblast differentiation. Rosiglitazone inhibited osteoblast differentiation in a concentration-dependent manner, and in parallel induced three PHD isoforms (PHD1, 2, and 3). PHD inhibitors and knockdown of each isoform prevented the inhibitory effects of rosiglitazone on osteoblast differentiation and increased the expression of Runx2, a transcription factor essential for osteoblastogenesis. MG-132, a proteasomal inhibitor also prevented the rosiglitazone-induced degradation of Runx2. Furthermore, both increased PHD isoform expressions and reduced osteoblast differentiation by rosiglitazone were prevented by PPARγ antagonists, indicating these effects were mediated via PPARγ activation. In vivo oral administration of rosiglitazone to female ICR mice for 8 weeks reduced bone mineral densities and plasma alkaline phosphatase (ALP) activity, and increased PHD expression in femoral primary bone marrow cells and the ubiquitination of Runx2. Together, this suggests that the rosiglitazone-induced suppression of osteoblast differentiation is at least partly induced via PPARγ-mediated PHD induction and subsequent promotion of the ubiquitination and degradation of Runx2.

No MeSH data available.


Related in: MedlinePlus