antibodies against total-extracellular signal-regulated kinase (t-erk)-1,2  (Cell Signaling Technology Inc)

Journal: Biomedicines

Article Title: Differential Roles of CD36 in Regulating Muscle Insulin Response Depend on Palmitic Acid Load

doi: 10.3390/biomedicines11030729

Figure Lengend Snippet: Loss of CD36 impairs insulin signaling in C2C12 myotubes without PA loading. ( A ) KEGG pathway analysis of differentiated C2C12 cells after transfection with scrambled siCont or siCD36. ( B ) Gene expression of Slc2a4, Irs1, Irs2, and Pi3kr1 in an insulin-signaling pathway was analyzed by qRT-PCR in differentiated C2C12 myotubes after transfection with siCont or siCD36. ( C ) Western blot analysis and quantification of insulin- induced CD36, P-AKT (S473), total AKT (T-AKT), P-ERK (T202/Y204), and total ERK (T-ERK) expression levels in differentiated C2C12 cells after transfection with siCont or siCD36. Values are presented as the mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001 versus the siCont group.

Article Snippet: After washing with TBST, the membranes were incubated with primary antibodies, including rabbit antibody phospho-AKT (P-AKT, S473) (#4060), total AKT (T-AKT) (#ab187783), phospho-p44/42 MAPK (P-ERK, T202/Y204) (#4370), total ERK (T-ERK) (#4695), cytochrome c (Cyto-C, #4280), cytochrome c oxidase subunit 4 (COX-4, #4850T), glucose-regulated protein 78 (GRP-78, #3177), C/EBP homologous protein (CHOP, #2895T) purchased from Cell Signaling Technology (Danvers, MA, USA), goat antibody anti CD36 (CD36, #AF2519) purchased from R&D system (Minneapolis, MN, USA), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, #AB0037) from Abways (Shanghai, China).

Techniques: Transfection, Expressing, Quantitative RT-PCR, Western Blot

Journal: Biomedicines

Article Title: Differential Roles of CD36 in Regulating Muscle Insulin Response Depend on Palmitic Acid Load

doi: 10.3390/biomedicines11030729

Figure Lengend Snippet: Loss of CD36 protects against insulin resistance in response to PA overload. Differentiated C2C12 myotubes transfected with siCont or siCD36 were treated with BSA or PA (200 μM) for 24 h. ( A ) The expression levels of CD36 were measured to assess the efficiency of knockdown. Lipid accumulations in differentiated C2C12 cells after CD36 siRNA or siCont transfection were examined by Oil Red O staining ( B ) and TG contents ( C ) under the conditions of BSA and PA stimulation. The area of lipid droplets (μm 2 ) was calculated using the Image J system (scale bar, 10 μm). ( D ) Differentiated C2C12 myotubes transfected with CD36 siRNA or siCont were treated with PA (300 μM) for 15 min, and then treated with insulin (10 nM) for 30 min. Whole-cell lysates were subjected to WB analysis with antibodies against CD36, P-AKT (S473), total AKT (T-AKT), P-ERK (T202/Y204), total ERK (T-ERK), and GAPDH. ( E ) Vectoror CD36 overexpressing (CD36 OE) C2C12 myotubes were treated with PA (300 μM) for 15 min, and then treated with insulin (10 nM) for the indicated times. Whole-cell lysates were subjected to WB analysis with antibodies against CD36, P-AKT (S473), total AKT (T-AKT), P-ERK (T202/Y204), total ERK (T-ERK), and GAPDH. Values are presented as the mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001 versus the control groups.

Article Snippet: After washing with TBST, the membranes were incubated with primary antibodies, including rabbit antibody phospho-AKT (P-AKT, S473) (#4060), total AKT (T-AKT) (#ab187783), phospho-p44/42 MAPK (P-ERK, T202/Y204) (#4370), total ERK (T-ERK) (#4695), cytochrome c (Cyto-C, #4280), cytochrome c oxidase subunit 4 (COX-4, #4850T), glucose-regulated protein 78 (GRP-78, #3177), C/EBP homologous protein (CHOP, #2895T) purchased from Cell Signaling Technology (Danvers, MA, USA), goat antibody anti CD36 (CD36, #AF2519) purchased from R&D system (Minneapolis, MN, USA), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, #AB0037) from Abways (Shanghai, China).

Techniques: Transfection, Expressing, Staining