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Inhibition of Calcium Influx Reduces Dysfunction and Apoptosis in Lipotoxic Pancreatic β-Cells via Regulation of Endoplasmic Reticulum Stress.

Zhou Y, Sun P, Wang T, Chen K, Zhu W, Wang H - PLoS ONE (2015)

Bottom Line: And whether the compounds could reduce palmitic acid-induced β-cell failure and the underlying mechanism were also investigated.It was found that both nifedipine and diazoxide protected MIN6 pancreatic β-cells and primary cultured murine islets from palmitic acid-induced apoptosis.Our results verified that nifedipine and diazoxide could reduce palmitic acid-induced endoplasmic reticulum stress to generate protective effects on pancreatic β-cells.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road, Shanghai, 201203, China.

ABSTRACT
Lipotoxicity plays an important role in pancreatic β-cell failure during the development of type 2 diabetes. Prolonged exposure of β-cells to elevated free fatty acids level could cause deterioration of β-cell function and induce cell apoptosis. Therefore, inhibition of fatty acids-induced β-cell dysfunction and apoptosis might provide benefit for the therapy of type 2 diabetes. The present study examined whether regulation of fatty acids-triggered calcium influx could protect pancreatic β-cells from lipotoxicity. Two small molecule compounds, L-type calcium channel blocker nifedipine and potassium channel activator diazoxide were used to inhibit palmitic acid-induced calcium influx. And whether the compounds could reduce palmitic acid-induced β-cell failure and the underlying mechanism were also investigated. It was found that both nifedipine and diazoxide protected MIN6 pancreatic β-cells and primary cultured murine islets from palmitic acid-induced apoptosis. Meanwhile, the impaired insulin secretion was also recovered to varying degrees by these two compounds. Our results verified that nifedipine and diazoxide could reduce palmitic acid-induced endoplasmic reticulum stress to generate protective effects on pancreatic β-cells. More importantly, it suggested that regulation of calcium influx by small molecule compounds might provide benefits for the prevention and therapy of type 2 diabetes.

No MeSH data available.


Related in: MedlinePlus

Nifedipine and diazoxide protected against PA-induced impairment of GSIS in pancreatic beta cells.(A) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the GSIS induced by 25 mM glucose in MIN6 cells were determined. *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (B) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the expression of PDX1 in nuclei and whole cell was determined by western blot. (C) The expression of PDX1 was calculated by optical density. ** p<0.01 compared to PA-treated alone group in nuclei PDX1 detection; ## p<0.01 compared to PA-treated alone group in whole cell PDX1 detection, n = 3. (D) After cultured islets were treated with PA in the presence/absence of different compounds for 48 h, the GSIS induced by 5 mM and 25 mM glucose were detected. * p<0.05; ** p<0.01; *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (E) Cultured islets were treated with PA in the presence/absence of different compounds for 48 h, then the PDX1 location was marked with red fluorescence, nuclei were dyed with DAPI to show blue fluorescence. Scale bar = 50 μm and referred to all panels. (F) The PDX1-positive cell rate was calculated as PDX1-postive nuclei number divided by total nuclei number in each islet. 10 islets were analyzed from six duplicated wells. * p<0.05 denote significant difference versus the PA-treated alone group, n = 6.
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pone.0132411.g006: Nifedipine and diazoxide protected against PA-induced impairment of GSIS in pancreatic beta cells.(A) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the GSIS induced by 25 mM glucose in MIN6 cells were determined. *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (B) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the expression of PDX1 in nuclei and whole cell was determined by western blot. (C) The expression of PDX1 was calculated by optical density. ** p<0.01 compared to PA-treated alone group in nuclei PDX1 detection; ## p<0.01 compared to PA-treated alone group in whole cell PDX1 detection, n = 3. (D) After cultured islets were treated with PA in the presence/absence of different compounds for 48 h, the GSIS induced by 5 mM and 25 mM glucose were detected. * p<0.05; ** p<0.01; *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (E) Cultured islets were treated with PA in the presence/absence of different compounds for 48 h, then the PDX1 location was marked with red fluorescence, nuclei were dyed with DAPI to show blue fluorescence. Scale bar = 50 μm and referred to all panels. (F) The PDX1-positive cell rate was calculated as PDX1-postive nuclei number divided by total nuclei number in each islet. 10 islets were analyzed from six duplicated wells. * p<0.05 denote significant difference versus the PA-treated alone group, n = 6.

Mentions: As nifedipine and diazoxide generated protective effect on PA-impaired β-cells, whether these two compounds could resist PA-induced impairment of insulin secretion in MIN6 β-cells and primary cultured islets was investigated. It was found that nifedipine at a concentration of 10 and 30 μM, diazoxide at 100 and 300 μM could partly restore PA-impaired GSIS in MIN6 cells (Fig 6A). We next detected the expression of PDX1 in nuclei and whole cell to investigate the translocation of PDX1. It was found that the nuclei PDX1 was significantly reduced after 48 h PA treatment in MIN6 cells (Fig 6B and 6C). However, both 10 μM nifedipine and 100 μM diazoxide increased the expression of nuclei PDX1 (Fig 6B and 6C). And PA-induced decreasing of PDX1 expression in whole cell was also significantly recovered by nifedipine and diazoxide (Fig 6B and 6C).


Inhibition of Calcium Influx Reduces Dysfunction and Apoptosis in Lipotoxic Pancreatic β-Cells via Regulation of Endoplasmic Reticulum Stress.

Zhou Y, Sun P, Wang T, Chen K, Zhu W, Wang H - PLoS ONE (2015)

Nifedipine and diazoxide protected against PA-induced impairment of GSIS in pancreatic beta cells.(A) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the GSIS induced by 25 mM glucose in MIN6 cells were determined. *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (B) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the expression of PDX1 in nuclei and whole cell was determined by western blot. (C) The expression of PDX1 was calculated by optical density. ** p<0.01 compared to PA-treated alone group in nuclei PDX1 detection; ## p<0.01 compared to PA-treated alone group in whole cell PDX1 detection, n = 3. (D) After cultured islets were treated with PA in the presence/absence of different compounds for 48 h, the GSIS induced by 5 mM and 25 mM glucose were detected. * p<0.05; ** p<0.01; *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (E) Cultured islets were treated with PA in the presence/absence of different compounds for 48 h, then the PDX1 location was marked with red fluorescence, nuclei were dyed with DAPI to show blue fluorescence. Scale bar = 50 μm and referred to all panels. (F) The PDX1-positive cell rate was calculated as PDX1-postive nuclei number divided by total nuclei number in each islet. 10 islets were analyzed from six duplicated wells. * p<0.05 denote significant difference versus the PA-treated alone group, n = 6.
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Related In: Results  -  Collection

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pone.0132411.g006: Nifedipine and diazoxide protected against PA-induced impairment of GSIS in pancreatic beta cells.(A) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the GSIS induced by 25 mM glucose in MIN6 cells were determined. *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (B) After MIN6 cells were treated with 0.5 mM PA in the presence/absence of nifedipine or diazoxide for 48 h, the expression of PDX1 in nuclei and whole cell was determined by western blot. (C) The expression of PDX1 was calculated by optical density. ** p<0.01 compared to PA-treated alone group in nuclei PDX1 detection; ## p<0.01 compared to PA-treated alone group in whole cell PDX1 detection, n = 3. (D) After cultured islets were treated with PA in the presence/absence of different compounds for 48 h, the GSIS induced by 5 mM and 25 mM glucose were detected. * p<0.05; ** p<0.01; *** p<0.001 denote significant difference versus the PA-treated alone group, n = 6. (E) Cultured islets were treated with PA in the presence/absence of different compounds for 48 h, then the PDX1 location was marked with red fluorescence, nuclei were dyed with DAPI to show blue fluorescence. Scale bar = 50 μm and referred to all panels. (F) The PDX1-positive cell rate was calculated as PDX1-postive nuclei number divided by total nuclei number in each islet. 10 islets were analyzed from six duplicated wells. * p<0.05 denote significant difference versus the PA-treated alone group, n = 6.
Mentions: As nifedipine and diazoxide generated protective effect on PA-impaired β-cells, whether these two compounds could resist PA-induced impairment of insulin secretion in MIN6 β-cells and primary cultured islets was investigated. It was found that nifedipine at a concentration of 10 and 30 μM, diazoxide at 100 and 300 μM could partly restore PA-impaired GSIS in MIN6 cells (Fig 6A). We next detected the expression of PDX1 in nuclei and whole cell to investigate the translocation of PDX1. It was found that the nuclei PDX1 was significantly reduced after 48 h PA treatment in MIN6 cells (Fig 6B and 6C). However, both 10 μM nifedipine and 100 μM diazoxide increased the expression of nuclei PDX1 (Fig 6B and 6C). And PA-induced decreasing of PDX1 expression in whole cell was also significantly recovered by nifedipine and diazoxide (Fig 6B and 6C).

Bottom Line: And whether the compounds could reduce palmitic acid-induced β-cell failure and the underlying mechanism were also investigated.It was found that both nifedipine and diazoxide protected MIN6 pancreatic β-cells and primary cultured murine islets from palmitic acid-induced apoptosis.Our results verified that nifedipine and diazoxide could reduce palmitic acid-induced endoplasmic reticulum stress to generate protective effects on pancreatic β-cells.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road, Shanghai, 201203, China.

ABSTRACT
Lipotoxicity plays an important role in pancreatic β-cell failure during the development of type 2 diabetes. Prolonged exposure of β-cells to elevated free fatty acids level could cause deterioration of β-cell function and induce cell apoptosis. Therefore, inhibition of fatty acids-induced β-cell dysfunction and apoptosis might provide benefit for the therapy of type 2 diabetes. The present study examined whether regulation of fatty acids-triggered calcium influx could protect pancreatic β-cells from lipotoxicity. Two small molecule compounds, L-type calcium channel blocker nifedipine and potassium channel activator diazoxide were used to inhibit palmitic acid-induced calcium influx. And whether the compounds could reduce palmitic acid-induced β-cell failure and the underlying mechanism were also investigated. It was found that both nifedipine and diazoxide protected MIN6 pancreatic β-cells and primary cultured murine islets from palmitic acid-induced apoptosis. Meanwhile, the impaired insulin secretion was also recovered to varying degrees by these two compounds. Our results verified that nifedipine and diazoxide could reduce palmitic acid-induced endoplasmic reticulum stress to generate protective effects on pancreatic β-cells. More importantly, it suggested that regulation of calcium influx by small molecule compounds might provide benefits for the prevention and therapy of type 2 diabetes.

No MeSH data available.


Related in: MedlinePlus