Journal: Pancreas

Article Title: Short-Term Modulation of Extracellular Signal-Regulated Kinase 1/2 and Stress-Activated Protein Kinase/c-Jun NH2-Terminal Kinase in Pancreatic Islets by Glucose and Palmitate

doi: 10.1097/mpa.0b013e31819fef03

Figure Lengend Snippet: FIGURE 1. Time-course response of ERK1/2 (A), SAPK/JNK (B), and AKT (Ser473) phosphorylations (C). Groups of 200 to 300 freshly isolated islets were preincubated for 30 minutes at 37-C in Krebs-Henseleit buffer with 5.6-mmol/L glucose. This solution was then replaced, and islets were maintained for 1.5, 5, 15, 30, 45, or 60 minutes in Krebs-Henseleit buffer containing 10-mmol/L glucose. Protein (100 Kg) extracts from homogenized islets were resolved in SDS-PAGE, transferred to nitrocellulose membranes, and blotted (IB) with specific antibodies. Scanning densitometry was performed on autoradiograms from independent experiments. The results are expressed as mean (SEM). A: pERK1/2: n = 6; ERK1/2: n = 3; B: pSAPK/JNK: n = 4; SAPK/JNK: n = 3; C: pAKT (Ser473): n = 4; AKT1: n = 3; AKT2: n = 3; *P G 0.05 versus control islets (CTL). CTL indicates control; IB, immunoblotting; pAKT, phospho-AKT; pERK1/2, phospho-ERK1/2; pSAPK/JNK, phospho-SAPK/JNK. The same abbreviations are used in Figures 2Y5.

Article Snippet: The reagents for protein assay (BioRad Protein Assay-Dye Reagent Concentrate), the apparatus for sodium dodecyl sulfateYpolyacrylamide gel electrophoresis (SDS-PAGE), and the immunoblotting and nitrocellulose paper (0.45 Km) were obtained from Bio-Rad (Melville, NY).

Techniques: Isolation, SDS Page, Control, Western Blot

Journal: Pancreas

Article Title: Short-Term Modulation of Extracellular Signal-Regulated Kinase 1/2 and Stress-Activated Protein Kinase/c-Jun NH2-Terminal Kinase in Pancreatic Islets by Glucose and Palmitate

doi: 10.1097/mpa.0b013e31819fef03

Figure Lengend Snippet: FIGURE 2. Effect of palmitate on glucose-induced phosphorylation of ERK1/2 (A), SAPK/JNK (B), and AKT (Ser473; C) in rat pancreatic islets. Groups of 200 to 300 freshly isolated islets were preincubated for 30 minutes at 37-C in Krebs-Henseleit 5.6-mmol/L glucose. This solution was then replaced, and islets were maintained for 5 minutes in Krebs-Henseleit containing 5.6-mmol/L and 10-mmol/L glucose or 10-mmol/L glucose plus 0.1-mmol/L palmitate. Protein (100 Kg) extracts from homogenized islets were resolved in SDS-PAGE, transferred to nitrocellulose membranes and blotted (IB) with specific antibody. Scanning densitometry was performed on autoradiograms from independent experiments. The results are expressed as mean (SEM). A: pERK1/2: n = 6; ERK1/2: n = 3; B: pSAPK/JNK: n = 4; SAPK/JNK: n = 3; C: pAKT (Ser473): n = 7; AKT1: n = 3; AKT2: n = 3; *P G 0.05 versus islets exposed to G5.6 or **P G 0.05 versus islets exposed to G10.

Article Snippet: The reagents for protein assay (BioRad Protein Assay-Dye Reagent Concentrate), the apparatus for sodium dodecyl sulfateYpolyacrylamide gel electrophoresis (SDS-PAGE), and the immunoblotting and nitrocellulose paper (0.45 Km) were obtained from Bio-Rad (Melville, NY).

Techniques: Phospho-proteomics, Isolation, SDS Page

Journal: Pancreas

Article Title: Short-Term Modulation of Extracellular Signal-Regulated Kinase 1/2 and Stress-Activated Protein Kinase/c-Jun NH2-Terminal Kinase in Pancreatic Islets by Glucose and Palmitate

doi: 10.1097/mpa.0b013e31819fef03

Figure Lengend Snippet: FIGURE 3. Effect of PI3K inhibitors (LY294002) on glucose-induced changes of ERK1/2, SAPK/JNK and AKT (Ser473) phosphorylation. Groups of 200 to 300 freshly islets were preincubated in the absence or presence of LY294002. The islets were then incubated for 5 minutes in Krebs-Henseleit buffer containing 5.6-mmol/L and 10-mmol/L glucose and 10-mmol/L glucose plus LY294002. Total protein extracts from homogenized islets were separated by SDS-PAGE, transferred to nitrocellulose membranes, and blotted (IB) with pAKT (Ser473), pERK1/2, and pSAPK/JNK antibodies. Scanning densitometry was performed on autoradiograms from independent experiments. The results are expressed as mean (SEM). A: pERK1/2: n = 3; ERK1/2: n = 3; pSAPK/JNK: n = 3; SAPK/JNK: n = 3; pAKT (Ser473): n = 3; AKT1: n = 3, AKT2: n = 3; *P G 0.05 versus islets exposed to G5.6 or **P G 0.05 versus islets exposed to G10.

Article Snippet: The reagents for protein assay (BioRad Protein Assay-Dye Reagent Concentrate), the apparatus for sodium dodecyl sulfateYpolyacrylamide gel electrophoresis (SDS-PAGE), and the immunoblotting and nitrocellulose paper (0.45 Km) were obtained from Bio-Rad (Melville, NY).

Techniques: Phospho-proteomics, Incubation, SDS Page

Journal: Pancreas

Article Title: Short-Term Modulation of Extracellular Signal-Regulated Kinase 1/2 and Stress-Activated Protein Kinase/c-Jun NH2-Terminal Kinase in Pancreatic Islets by Glucose and Palmitate

doi: 10.1097/mpa.0b013e31819fef03

Figure Lengend Snippet: FIGURE 4. Effect of ceramide synthesis blockage (by fumonisin B1) on changes of AKT (Ser473), ERK1/2, and SAPK/JNK phosphorylations in islets of rats. Groups of 200 to 300 fresh islets were preincubated in the absence or presence of fumonisin B1. The islets were then incubated for 5 minutes in Krebs-Henseleit buffer containing 5.6-mmol/L and 10-mmol/L glucose and 10-mmol/L glucose plus fumonisin B1. Total protein extracts from homogenized islets were separated by SDS-PAGE, transferred to nitrocellulose membranes, and blotted (IB) with pAKT (Ser473), pERK1/2, and pSAPK/JNK antibodies. Scanning densitometry was performed on autoradiograms from independent experiments. The results are expressed as mean (SEM) [pERK1/2: n = 4, ERK1/2: n = 3; pSAPK/JNK: n = 3, SAPK/JNK: n = 3; pAKT (Ser473): n = 4; AKT1: n = 3, AKT2: n = 3; *P G 0.05 versus islets exposed to G5.6, **P G 0.05 versus islets exposed to G10, or #P G 0.05 versus islets exposed to G10P].

Article Snippet: The reagents for protein assay (BioRad Protein Assay-Dye Reagent Concentrate), the apparatus for sodium dodecyl sulfateYpolyacrylamide gel electrophoresis (SDS-PAGE), and the immunoblotting and nitrocellulose paper (0.45 Km) were obtained from Bio-Rad (Melville, NY).

Techniques: Incubation, SDS Page