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Targeting FoxO1 with AS1842856 suppresses adipogenesis.

Zou P, Liu L, Zheng L, Liu L, Stoneman RE, Cho A, Emery A, Gilbert ER, Cheng Z - Cell Cycle (2014)

Bottom Line: As FoxO1 has been implicated in adipogenic regulation, we investigated the kinetics of FoxO1 activation during adipocyte differentiation, and tested the effects of FoxO1 antagonist (AS1842856) on adipogenesis.We found for the first time that the kinetics of FoxO1 activation follows a series of sigmoid curves, and reveals the phases relevant to clonal expansion, cell cycle arrest, and the regulation of PPARγ, adiponectin, and mitochondrial proteins (complexes I and III).Importantly, persistent inhibition of FoxO1 with AS1842856 almost completely suppressed adipocyte differentiation, while selective inhibition in specific stages had differential effects on adipogenesis.

View Article: PubMed Central - PubMed

Affiliation: a Department of Human Nutrition, Foods and Exercise; Fralin Life Science Institute; College of Agriculture and Life Science; Virginia Tech , Blacksburg , VA USA.

ABSTRACT
Hyperplasia (i.e., increased adipogenesis) contributes to excess adiposity, the hallmark of obesity that can trigger metabolic complications. As FoxO1 has been implicated in adipogenic regulation, we investigated the kinetics of FoxO1 activation during adipocyte differentiation, and tested the effects of FoxO1 antagonist (AS1842856) on adipogenesis. We found for the first time that the kinetics of FoxO1 activation follows a series of sigmoid curves, and reveals the phases relevant to clonal expansion, cell cycle arrest, and the regulation of PPARγ, adiponectin, and mitochondrial proteins (complexes I and III). In addition, multiple activation-inactivation transitions exist in the stage of terminal differentiation. Importantly, persistent inhibition of FoxO1 with AS1842856 almost completely suppressed adipocyte differentiation, while selective inhibition in specific stages had differential effects on adipogenesis. Our data present a new view of FoxO1 in adipogenic regulation, and suggest AS1842856 can be an anti-obesity agent that warrants further investigation.

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(See previous page). Effects of phase-specific FoxO1 inhibition on adipogenesis. (A and B) Images of cells that were maintained in basal medium (A), and that underwent differentiation induced with the protocol as described in Materials and Methods (B). (C) Images of cells that were treated with AS1842856 during stage BMII (days 7–12) and underwent differentiation induction as in (B). (D) Images of cells that were treated with AS1842856 during stage DMI (days 3–4) and underwent differentiation induction as in (B). (E) Images of cells that were treated with AS1842856 during stage BMI (days 1–2) and underwent differentiation induction as in (B). (F) Images of cells that were treated with AS1842856 during stage DMII (days 5–6) and underwent differentiation induction as in (B). The final concentration of AS1842856 was 0.1 μM. All the images were captured on day 12, and the microscope was set at 100X. (G) Measurement of extracted Oil red O retained in cells by the absorbance (O.D.) at 510 nm (n = 6). Treatments of ‘A’–‘F’ refer to the individual treatments as described in panels A-F. * P < 0 .05; **, P < 0 .01; and ***, P < 0 .0001.
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f0005: (See previous page). Effects of phase-specific FoxO1 inhibition on adipogenesis. (A and B) Images of cells that were maintained in basal medium (A), and that underwent differentiation induced with the protocol as described in Materials and Methods (B). (C) Images of cells that were treated with AS1842856 during stage BMII (days 7–12) and underwent differentiation induction as in (B). (D) Images of cells that were treated with AS1842856 during stage DMI (days 3–4) and underwent differentiation induction as in (B). (E) Images of cells that were treated with AS1842856 during stage BMI (days 1–2) and underwent differentiation induction as in (B). (F) Images of cells that were treated with AS1842856 during stage DMII (days 5–6) and underwent differentiation induction as in (B). The final concentration of AS1842856 was 0.1 μM. All the images were captured on day 12, and the microscope was set at 100X. (G) Measurement of extracted Oil red O retained in cells by the absorbance (O.D.) at 510 nm (n = 6). Treatments of ‘A’–‘F’ refer to the individual treatments as described in panels A-F. * P < 0 .05; **, P < 0 .01; and ***, P < 0 .0001.

Mentions: To test the importance of the additional transitions of IP2→AP3 for adipogenesis, we added AS1842856 (0.1 μM) to BMII at the end of day 6, locking FoxO1 in an inhibited state during days 7-12 (Fig. S2A). As controls, mature adipocytes showed significant accumulation of lipid droplets and strong Oil red O staining, which were not observed in the pre-adipocytes (Fig. 5A and B; Fig. S3A and B). By contrast, treatment of the cells with AS1842856 from day 7 to day 12 potently prevented adipogenesis regardless of differentiation induction (Fig. 5C; Fig. S3C). Thus, FoxO1 re-activation and the progression to IP2→AP3 are critical for adipogenesis. We then moved the treatments to earlier stages (Fig. S2B-D). Addition of AS1842856 (0.1 μM) to DMI led to a similar phenotype than to BMII (Fig. 5D; Fig. S3D). However, when it was supplemented in BMI and DMII, AS1842856 suppressed adipogenesis to a lesser extent than in DMI, suggestive of a milder disturbance of FoxO1 action in adipogenesis (Fig. 5E and F; Fig. S3E and F).Figure 5


Targeting FoxO1 with AS1842856 suppresses adipogenesis.

Zou P, Liu L, Zheng L, Liu L, Stoneman RE, Cho A, Emery A, Gilbert ER, Cheng Z - Cell Cycle (2014)

(See previous page). Effects of phase-specific FoxO1 inhibition on adipogenesis. (A and B) Images of cells that were maintained in basal medium (A), and that underwent differentiation induced with the protocol as described in Materials and Methods (B). (C) Images of cells that were treated with AS1842856 during stage BMII (days 7–12) and underwent differentiation induction as in (B). (D) Images of cells that were treated with AS1842856 during stage DMI (days 3–4) and underwent differentiation induction as in (B). (E) Images of cells that were treated with AS1842856 during stage BMI (days 1–2) and underwent differentiation induction as in (B). (F) Images of cells that were treated with AS1842856 during stage DMII (days 5–6) and underwent differentiation induction as in (B). The final concentration of AS1842856 was 0.1 μM. All the images were captured on day 12, and the microscope was set at 100X. (G) Measurement of extracted Oil red O retained in cells by the absorbance (O.D.) at 510 nm (n = 6). Treatments of ‘A’–‘F’ refer to the individual treatments as described in panels A-F. * P < 0 .05; **, P < 0 .01; and ***, P < 0 .0001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4613185&req=5

f0005: (See previous page). Effects of phase-specific FoxO1 inhibition on adipogenesis. (A and B) Images of cells that were maintained in basal medium (A), and that underwent differentiation induced with the protocol as described in Materials and Methods (B). (C) Images of cells that were treated with AS1842856 during stage BMII (days 7–12) and underwent differentiation induction as in (B). (D) Images of cells that were treated with AS1842856 during stage DMI (days 3–4) and underwent differentiation induction as in (B). (E) Images of cells that were treated with AS1842856 during stage BMI (days 1–2) and underwent differentiation induction as in (B). (F) Images of cells that were treated with AS1842856 during stage DMII (days 5–6) and underwent differentiation induction as in (B). The final concentration of AS1842856 was 0.1 μM. All the images were captured on day 12, and the microscope was set at 100X. (G) Measurement of extracted Oil red O retained in cells by the absorbance (O.D.) at 510 nm (n = 6). Treatments of ‘A’–‘F’ refer to the individual treatments as described in panels A-F. * P < 0 .05; **, P < 0 .01; and ***, P < 0 .0001.
Mentions: To test the importance of the additional transitions of IP2→AP3 for adipogenesis, we added AS1842856 (0.1 μM) to BMII at the end of day 6, locking FoxO1 in an inhibited state during days 7-12 (Fig. S2A). As controls, mature adipocytes showed significant accumulation of lipid droplets and strong Oil red O staining, which were not observed in the pre-adipocytes (Fig. 5A and B; Fig. S3A and B). By contrast, treatment of the cells with AS1842856 from day 7 to day 12 potently prevented adipogenesis regardless of differentiation induction (Fig. 5C; Fig. S3C). Thus, FoxO1 re-activation and the progression to IP2→AP3 are critical for adipogenesis. We then moved the treatments to earlier stages (Fig. S2B-D). Addition of AS1842856 (0.1 μM) to DMI led to a similar phenotype than to BMII (Fig. 5D; Fig. S3D). However, when it was supplemented in BMI and DMII, AS1842856 suppressed adipogenesis to a lesser extent than in DMI, suggestive of a milder disturbance of FoxO1 action in adipogenesis (Fig. 5E and F; Fig. S3E and F).Figure 5

Bottom Line: As FoxO1 has been implicated in adipogenic regulation, we investigated the kinetics of FoxO1 activation during adipocyte differentiation, and tested the effects of FoxO1 antagonist (AS1842856) on adipogenesis.We found for the first time that the kinetics of FoxO1 activation follows a series of sigmoid curves, and reveals the phases relevant to clonal expansion, cell cycle arrest, and the regulation of PPARγ, adiponectin, and mitochondrial proteins (complexes I and III).Importantly, persistent inhibition of FoxO1 with AS1842856 almost completely suppressed adipocyte differentiation, while selective inhibition in specific stages had differential effects on adipogenesis.

View Article: PubMed Central - PubMed

Affiliation: a Department of Human Nutrition, Foods and Exercise; Fralin Life Science Institute; College of Agriculture and Life Science; Virginia Tech , Blacksburg , VA USA.

ABSTRACT
Hyperplasia (i.e., increased adipogenesis) contributes to excess adiposity, the hallmark of obesity that can trigger metabolic complications. As FoxO1 has been implicated in adipogenic regulation, we investigated the kinetics of FoxO1 activation during adipocyte differentiation, and tested the effects of FoxO1 antagonist (AS1842856) on adipogenesis. We found for the first time that the kinetics of FoxO1 activation follows a series of sigmoid curves, and reveals the phases relevant to clonal expansion, cell cycle arrest, and the regulation of PPARγ, adiponectin, and mitochondrial proteins (complexes I and III). In addition, multiple activation-inactivation transitions exist in the stage of terminal differentiation. Importantly, persistent inhibition of FoxO1 with AS1842856 almost completely suppressed adipocyte differentiation, while selective inhibition in specific stages had differential effects on adipogenesis. Our data present a new view of FoxO1 in adipogenic regulation, and suggest AS1842856 can be an anti-obesity agent that warrants further investigation.

Show MeSH
Related in: MedlinePlus