The expression and antilipolytic role of phosphodiesterase 4 in rat adipocytes in vitro

Elevated concentrations of plasma free fatty acids (FFA) may cause insulin resistance. Inhibition of lipolysis reduces FFA availability and improves insulin sensitivity. Lipolysis is stimulated by increased concentration of cAMP. Phosphodiesterases (PDEs) hydrolyze cAMP and limit stimulation of lipolysis. Phosphodiesterase 313 (PDE3B) is the major isoform of PDE in rat adipocytes and mediates the antilipolytic effect of insulin. Phosphodiesterases 4 (PDE4) activity has also been detected in rat adipocytes, but its expression and physiological role are not clear.; In the first component of this dissertation research, the antilipolytic effect of Korean ginseng extract (Panax ginseng; KGE) in rat adipocytes and the signaling pathway for KGE antilipolysis were investigated. Adipocytes were isolated from rat adipose tissue by collagenase digestion. The ability of KGE to inhibit lipolysis was assessed by measuring glycerol release into the incubation medium. PDE inhibitors were applied to investigate the signaling pathway for KGE antilipolysis. The present study showed that insulin inhibited lipolysis by 42.4% compared to the basal lipolysis (p < 0.002). The specific PDE3 inhibitor cilostamide completely reversed insulin antilipolysis, whereas the specific PDE4 inhibitor rolipram did not reduce insulin antilipolysis. KGE inhibited lipolysis by 49% compared to the basal lipolysis (p < 0.002). Cilostamide and rolipram reduced KGE antilipolysis to 43% and 23.4% compared to basal conditions with the inhibitors, respectively. Moreover, combination of the PDE3 and PDE4 inhibitors completely reversed KGE antilipolysis. In contrast with insulin, KGE did not affect phosphorylation of protein kinase B (PKB). These data suggest that ginseng mimicked the antilipolytic effect of insulin through an alternative signaling pathway. KGE also partially affected PDE4-mediated antilipolysis in rat adipocytes.; In the second component of this project, the expression of PDE4 in rat adipocytes was investigated at the gene and protein levels. Reverse transcription-polymerase chain reaction (RT PCR) using published primers allowed amplification of fragments encoding PDE3B (530bp), PDE4A (233bp), PDE4B (786bp), PDE4C (539bp), and PDE4D (262bp) sequences. (Abstract shortened by UMI.)...