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Insulin elevates leptin secretion and mRNA levels via cyclic AMP in 3T3-L1 adipocytes deprived of glucose

View Article: PubMed Central - PubMed

ABSTRACT

Aims: Leptin plays an important role in the pathogenesis of obesity and diabetes, yet the regulatory mechanisms of this hormone have not been fully elucidated. In this study, we aimed to clarify the roles of insulin and glucose in leptin secretion and mRNA production using inhibitors of insulin signal transduction in adipocytes cultured under glucose-free or normal conditions.

Methods: Differentiated 3T3-L1 adipocytes were stimulated with insulin in combination with inhibitors for phosphoinositide 3-kinase (PI3K), Akt, and phosphodiesterase 3B (PDE3B), as well as epinephrine and a cyclic AMP (cAMP) analog under glucose-free or normal conditions. After 8 h of stimulation, leptin protein levels in the media and leptin mRNA expression levels in the adipocytes were measured.

Results: Insulin significantly increased the secretion and mRNA levels of leptin under the depletion of glucose. Glucose augmented basal leptin secretion without insulin, while glucose ified insulin-induced leptin mRNA upregulation. The PI3K inhibitor BEZ-235, the Akt inhibitor MK-2206, and the PDE3B inhibitor cilostazol attenuated the insulin stimulation of leptin secretion, but did not suppress the insulin-induced leptin mRNA upregulation with glucose depletion. In contrast to the glucose-free condition, insulin failed to upregulate leptin mRNA in the presence of glucose. The cAMP analog dibutyryl cAMP and epinephrine decreased both leptin secretion and mRNA regardless of glucose supplementation.

Conclusion: Insulin alone stimulates leptin secretion and elevates leptin mRNA levels via cAMP under the lack of glucose metabolism, while glucose is a significant and ambivalent effector on the insulin effects of leptin.

No MeSH data available.


Regulation of leptin secretion and mRNA levels by the insulin signaling pathway in 3T3-L1 adipocytes under glucose depletion. 3T3-L1 adipocytes were incubated for 8 h in glucose-free DMEM containing insulin (0.5 μM) with or without the PI3K inhibitor BEZ-235 (0.1, 1, 10 μM) and the Akt inhibitor MK-2206 (0.1, 1, 10 μM). (A and B) Effects of BEZ-235, (C) wortmannin, and (D and E) MK-2206 on leptin secretion and mRNA expression. (F) MTS assay after 24 h-incubation. *p < 0.05, **p < 0.01, and ***p < 0.001. n.s., not significant. Data are expressed as means ± SE.
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fig0010: Regulation of leptin secretion and mRNA levels by the insulin signaling pathway in 3T3-L1 adipocytes under glucose depletion. 3T3-L1 adipocytes were incubated for 8 h in glucose-free DMEM containing insulin (0.5 μM) with or without the PI3K inhibitor BEZ-235 (0.1, 1, 10 μM) and the Akt inhibitor MK-2206 (0.1, 1, 10 μM). (A and B) Effects of BEZ-235, (C) wortmannin, and (D and E) MK-2206 on leptin secretion and mRNA expression. (F) MTS assay after 24 h-incubation. *p < 0.05, **p < 0.01, and ***p < 0.001. n.s., not significant. Data are expressed as means ± SE.

Mentions: We then examined the effects of insulin signal inhibitors on leptin secretion and mRNA levels in 3T3-L1 adipocytes deprived of glucose. The PI3K inhibitor BEZ-235 attenuated the insulin stimulation of leptin secretion in a dose-dependent manner (Fig. 2A). In contrast, BEZ-235 did not suppress the upregulation of leptin mRNA by insulin (Fig. 2B). Another PI3K inhibitor, wortomannin, also inhibited the insulin-induced leptin secretion (Fig. 2C). The effects of the Akt inhibitor MK-2206 were similar. MK-2206 dose-dependently inhibited insulin-induced leptin secretion but not its mRNA expression with glucose depletion (Fig. 2D and E). Neither BEZ-235 nor MK-2206 affected cell viability assessed by MTS assay (Fig. 2F).


Insulin elevates leptin secretion and mRNA levels via cyclic AMP in 3T3-L1 adipocytes deprived of glucose
Regulation of leptin secretion and mRNA levels by the insulin signaling pathway in 3T3-L1 adipocytes under glucose depletion. 3T3-L1 adipocytes were incubated for 8 h in glucose-free DMEM containing insulin (0.5 μM) with or without the PI3K inhibitor BEZ-235 (0.1, 1, 10 μM) and the Akt inhibitor MK-2206 (0.1, 1, 10 μM). (A and B) Effects of BEZ-235, (C) wortmannin, and (D and E) MK-2206 on leptin secretion and mRNA expression. (F) MTS assay after 24 h-incubation. *p < 0.05, **p < 0.01, and ***p < 0.001. n.s., not significant. Data are expressed as means ± SE.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5121139&req=5

fig0010: Regulation of leptin secretion and mRNA levels by the insulin signaling pathway in 3T3-L1 adipocytes under glucose depletion. 3T3-L1 adipocytes were incubated for 8 h in glucose-free DMEM containing insulin (0.5 μM) with or without the PI3K inhibitor BEZ-235 (0.1, 1, 10 μM) and the Akt inhibitor MK-2206 (0.1, 1, 10 μM). (A and B) Effects of BEZ-235, (C) wortmannin, and (D and E) MK-2206 on leptin secretion and mRNA expression. (F) MTS assay after 24 h-incubation. *p < 0.05, **p < 0.01, and ***p < 0.001. n.s., not significant. Data are expressed as means ± SE.
Mentions: We then examined the effects of insulin signal inhibitors on leptin secretion and mRNA levels in 3T3-L1 adipocytes deprived of glucose. The PI3K inhibitor BEZ-235 attenuated the insulin stimulation of leptin secretion in a dose-dependent manner (Fig. 2A). In contrast, BEZ-235 did not suppress the upregulation of leptin mRNA by insulin (Fig. 2B). Another PI3K inhibitor, wortomannin, also inhibited the insulin-induced leptin secretion (Fig. 2C). The effects of the Akt inhibitor MK-2206 were similar. MK-2206 dose-dependently inhibited insulin-induced leptin secretion but not its mRNA expression with glucose depletion (Fig. 2D and E). Neither BEZ-235 nor MK-2206 affected cell viability assessed by MTS assay (Fig. 2F).

View Article: PubMed Central - PubMed

ABSTRACT

Aims: Leptin plays an important role in the pathogenesis of obesity and diabetes, yet the regulatory mechanisms of this hormone have not been fully elucidated. In this study, we aimed to clarify the roles of insulin and glucose in leptin secretion and mRNA production using inhibitors of insulin signal transduction in adipocytes cultured under glucose-free or normal conditions.

Methods: Differentiated 3T3-L1 adipocytes were stimulated with insulin in combination with inhibitors for phosphoinositide 3-kinase (PI3K), Akt, and phosphodiesterase 3B (PDE3B), as well as epinephrine and a cyclic AMP (cAMP) analog under glucose-free or normal conditions. After 8 h of stimulation, leptin protein levels in the media and leptin mRNA expression levels in the adipocytes were measured.

Results: Insulin significantly increased the secretion and mRNA levels of leptin under the depletion of glucose. Glucose augmented basal leptin secretion without insulin, while glucose ified insulin-induced leptin mRNA upregulation. The PI3K inhibitor BEZ-235, the Akt inhibitor MK-2206, and the PDE3B inhibitor cilostazol attenuated the insulin stimulation of leptin secretion, but did not suppress the insulin-induced leptin mRNA upregulation with glucose depletion. In contrast to the glucose-free condition, insulin failed to upregulate leptin mRNA in the presence of glucose. The cAMP analog dibutyryl cAMP and epinephrine decreased both leptin secretion and mRNA regardless of glucose supplementation.

Conclusion: Insulin alone stimulates leptin secretion and elevates leptin mRNA levels via cAMP under the lack of glucose metabolism, while glucose is a significant and ambivalent effector on the insulin effects of leptin.

No MeSH data available.