Inhibits prostaglandin synthesis, ..
sites of prostaglandin-E2 synthesis in ..
Prostaglandins (PGs) are autocrine/paracrinemodulators in the bone metabolism (). Among them, PGE has beenrecognized as an important mediator of bone remodeling (). It has been previously reported thatPGE stimulates OPG synthesis via the activation of p38mitogen-activated protein (MAP) kinase, p44/p42 MAP kinase andstress activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK)in osteoblast-like MC3T3-E1 cells (). However, the detailed mechanismunderlying PGE-stimulated OPG synthesis in osteoblastsremains to be fully elucidated.
Blockade of prostaglandin synthesis by ..
SMC migration from vascular media into the endothelial layer is an important vascular remodeling process during ICF (, ). Since the DA is chronically exposed to a high dose of PGE2 during gestation, we hypothesized that chronic exposure to EP4 stimuli may alter DA SMC migration. We found that chronic treatment with an EP4 agonist, ONO-AE1-329 (10–6 M) for long exposures (>2 days) increased SMC migration in a time-dependent manner (Figure A, black bars). We also observed similar increases in DA SMC migration after exposure to PGE2 (10–6 M) (data not shown). Since HA-rich matrices are important for cell migration and proliferation (, ), we measured HA production in culture media. We found that an increase in HA production in culture media was accompanied by an increase in SMC migration (Figure A, filled circles). To clarify whether an increase in HA production was responsible for the effect of ONO-AE1-329 on SMC migration, HA was removed from the culture media every day by replacing condition medium with fresh medium or by adding hyaluronidase (0.02 and 0.05 mg/ml) to the media to digest the secreted HA. The removal of HA from the culture medium significantly decreased the stimulatory effects of ONO-AE1-329 (Figure B). In addition, we investigated whether ONO-AE1-329’s effect was inhibited when HA synthase (HAS) was inhibited. We found that HAS2 was largely responsible for ONO-AE1-329–mediated HA production in DA, although HA is synthesized by 3 isoforms of HAS, namely HAS1, HAS2, and HAS3 (). We tested 2 different siRNAs for HAS2 for their capacity to decrease the expression of HAS2 mRNA and HA production in DA SMCs. We found that transfection of one siRNA for HAS2 significantly inhibited ONO-AE1-329–mediated HAS2 expression (~90% decrease) and thus the DA SMC migration, whereas that of a nontargeting negative control siRNA did not (Figure B). Moreover, HA supplementation (50 ng/ml) in the absence of ONO-AE1-329 increased DA SMC migration to the same level as treatment with ONO-AE1-329 alone (10–6 M) (Figure B). These findings indicate that EP4-induced HA production plays a critical role in SMC migration in DA.