Possible relationship among resveratrol, PPARα and PDE.

<p>These diagrams present our hypothesis about short- and long-term effects of resveratrol, as shown in the text.</p

Docking models and analysis of PPARα residues required for binding to resveratrol.

<p>(A) The four docking modes of resveratrol predicted using the GOLD 3.0 docking program [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0120865#pone.0120865.ref024" target="_blank">24</a>] with protein co-ordination data from the PPARα-GW409544 complex structure (PDB ID: 1K7L) and a standard docking protocol. (B) Superimposition of docking mode II of resveratrol (orange) on the structure of PPARα bound to GW409544, a potent PPARα agonist (green). Only the amino acids located near to GW409544 are displayed. The hydrogen bonds of Tyr314 and Tyr464 are shown as dashed green lines. (C) Binding free energies (∆∆Gbind (kcal/mol)) of the indicated PPARα amino acid residues, calculated by alanine scanning using data for the four predicted docking modes. (D) Activation of wild-type (WT) PPARα and its mutants by 5, 50 μM resveratrol or Wy-14643. BAECs were transiently transfected with PPRE-luc, wild-type or mutant GS-hPPARα, and pSV-β-gal. The data are presented as relative luciferase activities normalized to those of the β-galactosidase standard and as 1 for cells treated with DMSO (control), and represent the mean ± SD of three independent wells of cells. Similar results were obtained by two additional experiments. The data were calculated the relative luciferase activity in cells transfected with wild-type PPARα.</p

Inhibition of PDE enhances the activation of PPARα by resveratrol, especially at higher doses.

<p>(A) The dose-dependent activation of PPARα by resveratrol, T4HS and 4-PAP in BAECs transiently transfected with PPRE-luc, GS-hPPARα, and pSV-β-gal. Following transfection, the cells were incubated for 24 h with resveratrol, T4HS or 4-PAP at the indicated concentrations. Data were normalized to the β-galactosidase standard and represent the mean ± SD of three independent wells of cells. The right graph corresponds to the lower area marked by a dashed rectangle in left graph. (B) cAMP-dependent enhancement of PPARα activation by resveratrol, T4HS or 4-PAP. BAECs transiently transfected with PPRE-luc, GS-hPPARα, and pSV-β-gal were incubated for 24 h with 5 μM compounds in the presence or absence of 25 μM rolipram, a PDE4 inhibitor, or 25 μM forskolin, an adenylate cyclase activator. Luciferase data were normalized to the β-galactosidase standard and represent the mean ± SD of three independent wells. *<i>p</i> < 0.05, ***<i>p</i> < 0.001 (unpaired <i>t</i>-test) compared with control cells treated with the same compound. (C) The inhibition of PDE by resveratrol, T4HS, and rolipram. Data represent the mean ± SD of three independent wells of cells. Similar results were obtained by two additional experiments. The IC₅₀ values are shown in the Table. ***<i>p</i> < 0.001 (unpaired <i>t</i>-test) compared with rolipram. ^###<i>p</i> < 0.001 (unpaired <i>t</i>-test) compared with resveratrol. Similar results were obtained by two additional experiments in (A-C).</p

Diastereoselective Synthesis of 6″‑(<i>Z</i>)- and 6″‑(<i>E</i>)‑Fluoro Analogues of Anti-hepatitis B Virus Agent Entecavir and Its Evaluation of the Activity and Toxicity Profile of the Diastereomers

A method for the diastereoselective synthesis of 6″-(<i>Z</i>)- and 6″‑(<i>E</i>)-fluorinated analogues of the anti-HBV agent entecavir has been developed. Construction of the methylenecyclopentane skeleton of the target molecules has been accomplished by radical-mediated 5-<i>exo</i>-<i>dig</i> cyclization of the selenides <b>6</b> and <b>15</b> having the phenylsulfanylethynyl structure as a radical accepting moiety. In the radical reaction of the TBS-protected precursor <b>6</b>, (<i>Z</i>)-<i>anti</i>-<b>12</b> was formed as a major product. On the other hand, TIPS-protected <b>15</b> gave (<i>E</i>)-<i>anti</i>-<b>12</b>. The sulfur-extrusive stannylation of <i>anti</i>-<b>12</b> furnished a mixture of geometric isomers of the respective vinylstannane, whereas benzoyl-protected <b>17</b> underwent the stannylation in the manner of retention of configuration. Following XeF₂-mediated fluorination, introduction of the purine base and deoxygenation of the resulting carbocyclic guanosine gave the target (<i>E</i>)- and (<i>Z</i>)-<b>3</b> after deprotection. Evaluation of the anti-HBV activity of <b>3</b> revealed that fluorine-substitution at the 6″-position of entecavir gave rise to a reduction in the cytotoxicity in HepG2 cells with retention of the antiviral activity