Yonsei Med J.  2005 Feb;46(1):149-154. 10.3349/ymj.2005.46.1.149.

Rolipram, a Phosphodiesterase 4 Inhibitor, Stimulates Inducible cAMP Early Repressor Expression in Osteoblasts

• Cho ES
• Yu JH
• Kim MS
• Yim M

• KMID: 2158125
• DOI: http://doi.org/10.3349/ymj.2005.46.1.149

Abstract

Phosphodiesterase (PDE) 4 inhibitors have been shown to induce the cAMP-mediated signaling pathway by inhibiting cAMP hydrolysis. This study investigated the effect of a PDE4 inhibitor on the expression of the inducible cAMP early repressor (ICER), which is an endogenous inhibitor of CRE- mediated transcription, in osteoblastic cells. RT-PCR analysis revealed that rolipram, a PDE4 inhibitor, stimulates the ICER mRNA in a dose dependent manner. The induction of ICER mRNA expression by rolipram was suppressed by the inhibitors of protein kinase A (PKA) and p38 MAPK, suggesting the involvement of PKA and p38 MAPK activation in ICER expression by rolipram. It was previously shown that rolipram induced the expression of TNF-related activation-induced cytokine (TRANCE, also known as RANKL, ODF, or OPGL) in osteoblasts. This paper provides evidences that a transcriptional repressor like ICER might modulate TRANCE mRNA expression by rolipram in osteoblasts.

Keyword

Osteoblast; cyclic AMP; PDE4 inhibitor; ICER

MeSH Terms

3', 5'-Cyclic-Nucleotide Phosphodiesterase/*antagonists & inhibitors
Animals
Animals, Outbred Strains
Cyclic AMP-Dependent Protein Kinases/metabolism
DNA-Binding Proteins/genetics/*metabolism
Gene Expression/drug effects
Mice
Osteoblasts/*drug effects/metabolism
Phosphodiesterase Inhibitors/*pharmacology
Research Support, Non-U.S. Gov't
Rolipram/*pharmacology
Transcription Factors/genetics/*metabolism
p38 Mitogen-Activated Protein Kinases/metabolism

Figure

• 

  Fig. 1 Effect of rolipram, a PDE4 inhibitor, on ICER mRNA expression in UAMS-32 cells. A. The cells were treated with 5 µM rolipram for the indicated times, or B. with the indicated concentrations of rolipram for 3 hrs. The total RNA was prepared and reverse transcribed in the presence (+RT) or absence (-RT) of reverse transcriptase, and the cDNA was amplified using specific primers designed for the genes of ICER-I (700 bp) and -II (262 bp). M=0.1 kb DNA marker.

• 

  Fig. 2 The involvement of signal transduction pathways in ICER mRNA expression by rolipram. A. The UAMS-32 cells were preincubated in the presence or absence of 30µ M of H89, 50 µM of PD98059, or 50 µM of SB203580 (inhibitors of PKA, ERK, and p38 MAPK, respectively) for 30 min, and then treated with 5µM rolipram for 3 hrs. Total RNA was extracted from the cells and the expression of the ICER was examined by RT-PCR analysis. M=0.1kb DNA marker. B. The UAMS-32 cells were incubated with 5µM rolipram for the indicated times. The total cellular protein was analyzed by immunoblotting with the antibodies for phospho-p38 after which the membranes was stripped and incubated with the anti-p38 antibodies.

• 

  Fig. 3 The attenuation of rolipram-induced TRANCE mRNA expression requires the transcriptional repressor in osteoblasts. A. UAMS-32 cells were treated with 5 µM rolipram, 1 µg/ml cycloheximide (CHX) or rolipram and CHX (the CHX was added 1hr before the rolipram). The total RNA was extracted from the cells at the indicated times. Total RNA was analyzed by Northern blot using probes for TRANCE and GAPDH. B. primary mouse calvarial cells were treated in a similar manner shown in A.

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