Prostaglandin D2 Added during the Differentiation of 3T3-L1 Cells Suppresses Adipogenesis via Dysfunction of D-Prostanoid Receptor P1 and P2

Abstract

We previously reported that the addition of prostaglandin, (PG)D2, and its chemically stable analog, 11-deoxy-11-methylene-PGD2 (11d-11m-PGD2), during the maturation phase of 3T3-L1 cells promotes adipogenesis. In the present study, we aimed to elucidate the effects of the addition of PGD2 or 11d-11m-PGD2 to 3T3-L1 cells during the differentiation phase on adipogenesis. We found that both PGD2 and 11d-11m-PGD2 suppressed adipogenesis through the downregulation of peroxisome proliferator-activated receptor gamma (PPARγ) expression. However, the latter suppressed adipogenesis more potently than PGD2, most likely because of its higher resistance to spontaneous transformation into PGJ2 derivatives. In addition, this anti-adipogenic effect was attenuated by the coexistence of an IP receptor agonist, suggesting that the effect depends on the intensity of the signaling from the IP receptor. The D-prostanoid receptors 1 (DP1) and 2 (DP2, also known as a chemoattractant receptor-homologous molecule expressed on Th2 cells) are receptors for PGD2. The inhibitory effects of PGD2 and 11d-11m-PGD2 on adipogenesis were slightly attenuated by a DP2 agonist. Furthermore, the addition of PGD2 and 11d-11m-PGD2 during the differentiation phase reduced the DP1 and DP2 expression during the maturation phase. Overall, these results indicated that the addition of PGD2 or 11d-11m-PGD2 during the differentiation phase suppresses adipogenesis via the dysfunction of DP1 and DP2. Therefore, unidentified receptor(s) for both molecules may be involved in the suppression of adipogenesis.

Eicosapentaenoic Acid Induces the Inhibition of Adipogenesis by Reducing the Effect of PPARγ Activator and Mediating PKA Activation and Increased COX-2 Expression in 3T3-L1 Cells at the Differentiation Stage

“…Pre-adipogenic mouse 3T3-L1 cells (JCRB9014; JCRB Cell Bank, Osaka, Japan) were cultured following previously established protocols [ 24 , 25 ]. In summary, 3T3-L1 cells were seeded in 35- or 60-mm dishes containing growth medium (GM) at a density of 1 × 10 5 or 2 × 10 5 , respectively, in the growth stage.…”

“…To quantify intracellular triacylglycerol (TAG) and proteins, the same methods as previously described [ 24 , 25 ] were employed. Cultured mature adipocytes were collected and suspended in phosphate-buffered saline (PBS) without Ca 2+ and Mg 2+ (PBS [-]) containing 0.05% trypsin and 0.53 mM EDTA.…”

“…The gene expression levels were quantitatively analyzed following previously established methods [ 24 , 25 ]. In summary, reverse transcription (RT) was performed using M-MLV reverse transcriptase (ribonuclease H activity caused by point mutation), using total RNA (1 g) that was extracted from the cells using acid guanidium thiocyanate/phenol/chloroform after 24 and 48 h after the differentiation stage and on days 6 of the maturation stage.…”