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FTO Inhibits Insulin Secretion and Promotes NF-κB Activation through Positively Regulating ROS Production in Pancreatic β cells.

Fan HQ, He W, Xu KF, Wang ZX, Xu XY, Chen H - PLoS ONE (2015)

Bottom Line: FTO overexpression in MIN6 cells achieved by lentivirus delivery significantly inhibits insulin secretion in the presence of glucose stimulus as well as KCl.FTO silence has no effect on insulin secretion of MIN6 cells.Furthermore, reactive oxygen species (ROS) production and NF-κB activation are significantly promoted by FTO overexpression.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, China.

ABSTRACT
FTO (Fat mass and obesity-associated) is associated with increased risk of obesity and type 2 diabetes incurrence. Pancreas islet β cells dysfunction and insulin resistance are major causes of type 2 diabetes. However, whether FTO plays an important functional role in pancreatic β cells as well as the related molecular mechanism is still unclear. In the present study, the tissue expression profile of FTO was firstly determined using quantitative PCR and western blot. FTO is widely expressed in various tissues and presented with relative high expression in pancreas tissue, especially in endocrine pancreas. FTO overexpression in MIN6 cells achieved by lentivirus delivery significantly inhibits insulin secretion in the presence of glucose stimulus as well as KCl. FTO silence has no effect on insulin secretion of MIN6 cells. However, FTO overexpression doesn't affect the transcription of insulin gene. Furthermore, reactive oxygen species (ROS) production and NF-κB activation are significantly promoted by FTO overexpression. Inhibition of intracellular ROS production by N-acetyl-L-cysteine (NAC) can alleviate NF-κB activation and restore the insulin secretion mediated by FTO overexpression. A whole transcript-microarray is employed to analyze the differential gene expression mediated by FTO overexpression. The genes which are modulated by FTO are involved in many important biological pathways such as G-protein coupled receptor signaling and NF-κB signaling. Therefore, our study indicates that FTO may contribute to pancreas islet β cells dysfunction and the inhibition of FTO activity is a potential target for the treatment of diabetes.

No MeSH data available.


Related in: MedlinePlus

The effect of FTO expression on insulin secretion.(A) FTO overexpression in MIN6 cells via lentivirus delivery using Western blot. β-actin was loaded as an internal control. (B) Detection of FTO in MIN6 cells transfected with FTO shRNA and control shRNA. β-actin was loaded as an internal control. (C) Insulin secretion of MIN6 cells with FTO overexpression at 60 min after 2 mM, 20 mM glucose or 50 mM KCl stimulation. MIN6 cells were stably transfected with pLVX-IRES-ZsGreen (CON) or pLVX-IRES-ZsGreen-FTO (pLVX-FTO). (D) Effect of FTO silence on the insulin secretion in MIN6 cells. MIN6 cells transfected with FTO shRNA 2 or shRNA 3 were analyzed with insulin secretion at 60 min after the glucose or KCl stimulation. Data were presented as mean ± SD. The symbol ** denotes significant statistical difference (p < 0.01).
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pone.0127705.g002: The effect of FTO expression on insulin secretion.(A) FTO overexpression in MIN6 cells via lentivirus delivery using Western blot. β-actin was loaded as an internal control. (B) Detection of FTO in MIN6 cells transfected with FTO shRNA and control shRNA. β-actin was loaded as an internal control. (C) Insulin secretion of MIN6 cells with FTO overexpression at 60 min after 2 mM, 20 mM glucose or 50 mM KCl stimulation. MIN6 cells were stably transfected with pLVX-IRES-ZsGreen (CON) or pLVX-IRES-ZsGreen-FTO (pLVX-FTO). (D) Effect of FTO silence on the insulin secretion in MIN6 cells. MIN6 cells transfected with FTO shRNA 2 or shRNA 3 were analyzed with insulin secretion at 60 min after the glucose or KCl stimulation. Data were presented as mean ± SD. The symbol ** denotes significant statistical difference (p < 0.01).

Mentions: FTO polymorphisms are associated with type 2 diabetes which is a metabolic disorder in the context of insulin resistance and relative lack of insulin due to dysfunction of pancreas islet cells. Our profile data shows FTO gene may play an important role in pancreas tissues. We further explore the biological role of FTO in pancreas islet cells. FTO overexpression in MIN6 cells was archived by lentivirus delivery and was detected by Western blot analysis (Fig 2A). Furthermore, we knock down FTO in MIN6 cells via RNAi lentivirus. FTO expression was effectively silenced by shFTO-2 and 3 (Fig 2B). Insulin secretion assay was employed to determine whether FTO affects insulin secretion. MIN6 cells were incubated with glucose at different concentration and insulin secretion was determined at 10 min and 60 min when incubation. Our results show that FTO overexpression significantly suppresses the insulin secretion at 10 min (S1A Fig) and the secretion at 60 min (Fig 2C) in the presence of low and high concentration of glucose. Glucose-free secretagogues were also performed and the same effect was observed in potassium chloride (KCl) treatment (S1A Fig and Fig 2C). Insulin secretion experiment shows that FTO knockdown only slightly increase the insulin secretion compared with the control group (Fig 2D and S1B Fig). Our results indicated that FTO affect the insulin secretion in pancreas islet cells and this may contribute type 2 diabetes occurrences.


FTO Inhibits Insulin Secretion and Promotes NF-κB Activation through Positively Regulating ROS Production in Pancreatic β cells.

Fan HQ, He W, Xu KF, Wang ZX, Xu XY, Chen H - PLoS ONE (2015)

The effect of FTO expression on insulin secretion.(A) FTO overexpression in MIN6 cells via lentivirus delivery using Western blot. β-actin was loaded as an internal control. (B) Detection of FTO in MIN6 cells transfected with FTO shRNA and control shRNA. β-actin was loaded as an internal control. (C) Insulin secretion of MIN6 cells with FTO overexpression at 60 min after 2 mM, 20 mM glucose or 50 mM KCl stimulation. MIN6 cells were stably transfected with pLVX-IRES-ZsGreen (CON) or pLVX-IRES-ZsGreen-FTO (pLVX-FTO). (D) Effect of FTO silence on the insulin secretion in MIN6 cells. MIN6 cells transfected with FTO shRNA 2 or shRNA 3 were analyzed with insulin secretion at 60 min after the glucose or KCl stimulation. Data were presented as mean ± SD. The symbol ** denotes significant statistical difference (p < 0.01).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4446323&req=5

pone.0127705.g002: The effect of FTO expression on insulin secretion.(A) FTO overexpression in MIN6 cells via lentivirus delivery using Western blot. β-actin was loaded as an internal control. (B) Detection of FTO in MIN6 cells transfected with FTO shRNA and control shRNA. β-actin was loaded as an internal control. (C) Insulin secretion of MIN6 cells with FTO overexpression at 60 min after 2 mM, 20 mM glucose or 50 mM KCl stimulation. MIN6 cells were stably transfected with pLVX-IRES-ZsGreen (CON) or pLVX-IRES-ZsGreen-FTO (pLVX-FTO). (D) Effect of FTO silence on the insulin secretion in MIN6 cells. MIN6 cells transfected with FTO shRNA 2 or shRNA 3 were analyzed with insulin secretion at 60 min after the glucose or KCl stimulation. Data were presented as mean ± SD. The symbol ** denotes significant statistical difference (p < 0.01).
Mentions: FTO polymorphisms are associated with type 2 diabetes which is a metabolic disorder in the context of insulin resistance and relative lack of insulin due to dysfunction of pancreas islet cells. Our profile data shows FTO gene may play an important role in pancreas tissues. We further explore the biological role of FTO in pancreas islet cells. FTO overexpression in MIN6 cells was archived by lentivirus delivery and was detected by Western blot analysis (Fig 2A). Furthermore, we knock down FTO in MIN6 cells via RNAi lentivirus. FTO expression was effectively silenced by shFTO-2 and 3 (Fig 2B). Insulin secretion assay was employed to determine whether FTO affects insulin secretion. MIN6 cells were incubated with glucose at different concentration and insulin secretion was determined at 10 min and 60 min when incubation. Our results show that FTO overexpression significantly suppresses the insulin secretion at 10 min (S1A Fig) and the secretion at 60 min (Fig 2C) in the presence of low and high concentration of glucose. Glucose-free secretagogues were also performed and the same effect was observed in potassium chloride (KCl) treatment (S1A Fig and Fig 2C). Insulin secretion experiment shows that FTO knockdown only slightly increase the insulin secretion compared with the control group (Fig 2D and S1B Fig). Our results indicated that FTO affect the insulin secretion in pancreas islet cells and this may contribute type 2 diabetes occurrences.

Bottom Line: FTO overexpression in MIN6 cells achieved by lentivirus delivery significantly inhibits insulin secretion in the presence of glucose stimulus as well as KCl.FTO silence has no effect on insulin secretion of MIN6 cells.Furthermore, reactive oxygen species (ROS) production and NF-κB activation are significantly promoted by FTO overexpression.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology, the First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, China.

ABSTRACT
FTO (Fat mass and obesity-associated) is associated with increased risk of obesity and type 2 diabetes incurrence. Pancreas islet β cells dysfunction and insulin resistance are major causes of type 2 diabetes. However, whether FTO plays an important functional role in pancreatic β cells as well as the related molecular mechanism is still unclear. In the present study, the tissue expression profile of FTO was firstly determined using quantitative PCR and western blot. FTO is widely expressed in various tissues and presented with relative high expression in pancreas tissue, especially in endocrine pancreas. FTO overexpression in MIN6 cells achieved by lentivirus delivery significantly inhibits insulin secretion in the presence of glucose stimulus as well as KCl. FTO silence has no effect on insulin secretion of MIN6 cells. However, FTO overexpression doesn't affect the transcription of insulin gene. Furthermore, reactive oxygen species (ROS) production and NF-κB activation are significantly promoted by FTO overexpression. Inhibition of intracellular ROS production by N-acetyl-L-cysteine (NAC) can alleviate NF-κB activation and restore the insulin secretion mediated by FTO overexpression. A whole transcript-microarray is employed to analyze the differential gene expression mediated by FTO overexpression. The genes which are modulated by FTO are involved in many important biological pathways such as G-protein coupled receptor signaling and NF-κB signaling. Therefore, our study indicates that FTO may contribute to pancreas islet β cells dysfunction and the inhibition of FTO activity is a potential target for the treatment of diabetes.

No MeSH data available.


Related in: MedlinePlus