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7-Chloroarctinone-b as a new selective PPARgamma antagonist potently blocks adipocyte differentiation.

Li YT, Li L, Chen J, Hu TC, Huang J, Guo YW, Jiang HL, Shen X - Acta Pharmacol. Sin. (2009)

Bottom Line: A new thiophene-acetylene type of natural product, 7-chloroarctinone-b (CAB), isolated from the roots of Rhaponticum uniflorum, was discovered as a novel PPARgamma antagonist capable of inhibiting rosiglitazone-induced PPARgamma transcriptional activity.SPR analysis suggested that CAB bound tightly to PPARgamma and considerably antagonized the potent PPARgamma agonist rosiglitazone-stimulated PPARgamma-LBD/RXRalpha-LBD binding.CAB could efficiently antagonize both hormone and rosiglitazone-induced adipocyte differentiation in cell culture.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China.

ABSTRACT

Aim: Peroxisome proliferator-activated receptor gamma (PPARgamma) is a therapeutic target for obesity, cancer and diabetes mellitus. In order to develop potent lead compounds for obesity treatment, we screened a natural product library for novel PPARgamma antagonists with inhibitory effects on adipocyte differentiation.

Methods: Surface plasmon resonance (SPR) technology and cell-based transactivation assay were used to screen for PPARgamma antagonists. To investigate the antagonistic mechanism of the active compound, we measured its effect on PPARgamma/RXRalpha heterodimerization and PPARgamma co-activator recruitment using yeast two-hybrid assay, Gal4/UAS cell-based assay and SPR based assay. The 3T3-L1 cell differentiation assay was used to evaluate the effect of the active compound on adipocyte differentiation.

Results: A new thiophene-acetylene type of natural product, 7-chloroarctinone-b (CAB), isolated from the roots of Rhaponticum uniflorum, was discovered as a novel PPARgamma antagonist capable of inhibiting rosiglitazone-induced PPARgamma transcriptional activity. SPR analysis suggested that CAB bound tightly to PPARgamma and considerably antagonized the potent PPARgamma agonist rosiglitazone-stimulated PPARgamma-LBD/RXRalpha-LBD binding. Gal4/UAS and yeast two-hybrid assays were used to evaluate the antagonistic activity of CAB on rosiglitazone-induced recruitment of the coactivator for PPARgamma. CAB could efficiently antagonize both hormone and rosiglitazone-induced adipocyte differentiation in cell culture.

Conclusion: CAB shows antagonistic activity to PPARgamma and can block the adipocyte differentiation, indicating it may be of potential use as a lead therapeutic compound for obesity.

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Related in: MedlinePlus

CAB inhibited PPARγ transcriptional activity as evaluated by transactivation assay. HEK293T cells were co-transfected with PPARγ, RXRα and reporter gene, while the control plasmids were treated (A) with or (B) without rosiglitazone (2.5 μmol/L) and increasing concentrations of CAB for 18 h. The cells were harvested for luciferase and β-galactosidase assays. The luciferase activity was normalized with β-galactosidase activity. The concentration of GW9662 (PPARγ antagonist) was 1 μmol/L.
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fig3: CAB inhibited PPARγ transcriptional activity as evaluated by transactivation assay. HEK293T cells were co-transfected with PPARγ, RXRα and reporter gene, while the control plasmids were treated (A) with or (B) without rosiglitazone (2.5 μmol/L) and increasing concentrations of CAB for 18 h. The cells were harvested for luciferase and β-galactosidase assays. The luciferase activity was normalized with β-galactosidase activity. The concentration of GW9662 (PPARγ antagonist) was 1 μmol/L.

Mentions: To further investigate the ability of CAB in modulation of PPARγ transcription, transactivation assay was performed. In the assay, HEK-293T cells were cotransfected with full-length PPARγ, RXRα, and a reporter plasmid containing the PPAR response element. Transfected cells were incubated with different concentrations of CAB with or without rosiglitazone. As shown in Figure 3A, co-treatment with 2.5 μmol/L rosiglitazone and increasing concentrations of CAB resulted in a dose-dependent decrease in rosiglitazone-stimulated PPARγ transcriptional activity. As shown in Figure 3B, CAB itself could also inhibit PPARγ basal transcriptional activity induced by endogenous PPARγ ligands. These results indicate that CAB is a dose-dependent inhibitor of PPARγ transcriptional activity whether stimulated by its agonist, rosiglitazone, or by other ligands.


7-Chloroarctinone-b as a new selective PPARgamma antagonist potently blocks adipocyte differentiation.

Li YT, Li L, Chen J, Hu TC, Huang J, Guo YW, Jiang HL, Shen X - Acta Pharmacol. Sin. (2009)

CAB inhibited PPARγ transcriptional activity as evaluated by transactivation assay. HEK293T cells were co-transfected with PPARγ, RXRα and reporter gene, while the control plasmids were treated (A) with or (B) without rosiglitazone (2.5 μmol/L) and increasing concentrations of CAB for 18 h. The cells were harvested for luciferase and β-galactosidase assays. The luciferase activity was normalized with β-galactosidase activity. The concentration of GW9662 (PPARγ antagonist) was 1 μmol/L.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4007185&req=5

fig3: CAB inhibited PPARγ transcriptional activity as evaluated by transactivation assay. HEK293T cells were co-transfected with PPARγ, RXRα and reporter gene, while the control plasmids were treated (A) with or (B) without rosiglitazone (2.5 μmol/L) and increasing concentrations of CAB for 18 h. The cells were harvested for luciferase and β-galactosidase assays. The luciferase activity was normalized with β-galactosidase activity. The concentration of GW9662 (PPARγ antagonist) was 1 μmol/L.
Mentions: To further investigate the ability of CAB in modulation of PPARγ transcription, transactivation assay was performed. In the assay, HEK-293T cells were cotransfected with full-length PPARγ, RXRα, and a reporter plasmid containing the PPAR response element. Transfected cells were incubated with different concentrations of CAB with or without rosiglitazone. As shown in Figure 3A, co-treatment with 2.5 μmol/L rosiglitazone and increasing concentrations of CAB resulted in a dose-dependent decrease in rosiglitazone-stimulated PPARγ transcriptional activity. As shown in Figure 3B, CAB itself could also inhibit PPARγ basal transcriptional activity induced by endogenous PPARγ ligands. These results indicate that CAB is a dose-dependent inhibitor of PPARγ transcriptional activity whether stimulated by its agonist, rosiglitazone, or by other ligands.

Bottom Line: A new thiophene-acetylene type of natural product, 7-chloroarctinone-b (CAB), isolated from the roots of Rhaponticum uniflorum, was discovered as a novel PPARgamma antagonist capable of inhibiting rosiglitazone-induced PPARgamma transcriptional activity.SPR analysis suggested that CAB bound tightly to PPARgamma and considerably antagonized the potent PPARgamma agonist rosiglitazone-stimulated PPARgamma-LBD/RXRalpha-LBD binding.CAB could efficiently antagonize both hormone and rosiglitazone-induced adipocyte differentiation in cell culture.

View Article: PubMed Central - PubMed

Affiliation: School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China.

ABSTRACT

Aim: Peroxisome proliferator-activated receptor gamma (PPARgamma) is a therapeutic target for obesity, cancer and diabetes mellitus. In order to develop potent lead compounds for obesity treatment, we screened a natural product library for novel PPARgamma antagonists with inhibitory effects on adipocyte differentiation.

Methods: Surface plasmon resonance (SPR) technology and cell-based transactivation assay were used to screen for PPARgamma antagonists. To investigate the antagonistic mechanism of the active compound, we measured its effect on PPARgamma/RXRalpha heterodimerization and PPARgamma co-activator recruitment using yeast two-hybrid assay, Gal4/UAS cell-based assay and SPR based assay. The 3T3-L1 cell differentiation assay was used to evaluate the effect of the active compound on adipocyte differentiation.

Results: A new thiophene-acetylene type of natural product, 7-chloroarctinone-b (CAB), isolated from the roots of Rhaponticum uniflorum, was discovered as a novel PPARgamma antagonist capable of inhibiting rosiglitazone-induced PPARgamma transcriptional activity. SPR analysis suggested that CAB bound tightly to PPARgamma and considerably antagonized the potent PPARgamma agonist rosiglitazone-stimulated PPARgamma-LBD/RXRalpha-LBD binding. Gal4/UAS and yeast two-hybrid assays were used to evaluate the antagonistic activity of CAB on rosiglitazone-induced recruitment of the coactivator for PPARgamma. CAB could efficiently antagonize both hormone and rosiglitazone-induced adipocyte differentiation in cell culture.

Conclusion: CAB shows antagonistic activity to PPARgamma and can block the adipocyte differentiation, indicating it may be of potential use as a lead therapeutic compound for obesity.

Show MeSH
Related in: MedlinePlus