Journal: International Journal of Molecular Sciences

Article Title: MiR-30b Is Involved in the Homocysteine-Induced Apoptosis in Human Coronary Artery Endothelial Cells by Regulating the Expression of Caspase 3

doi: 10.3390/ijms160817682

Figure Lengend Snippet: ( A ) The impact on expression of caspase 3 mRNA by different concentrations Hcy on HCAECs. The time of treatment was 24 h. Compared with the control group, * p < 0.05, ** p < 0.01, n = 3 in each group; and ( B ) The impact on expression of caspase 3 protein by different concentrations Hcy on HCAECs. The time of treatment was 24 h. From left to right is the control group, Hcy 0.1 mmol/L group, Hcy 0.3 mmol/L group, Hcy 0.6 mmol/L group, Hcy 1.0 mmol/L group, Hcy 1.3 mmol/L group, Hcy 1.6 mmol/L group, Hcy 2.0 mmol/L group.

Article Snippet: Then, the PVDF membrane was incubated overnight at 4 °C with anti-β-actin (Sigma, Saint Louis, MO, USA), anti-caspase-3 antibody (Cell Signaling Technology, Shanghai, China) diluted by Tris-Buffered Saline and Tween (TBST).

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: MiR-30b Is Involved in the Homocysteine-Induced Apoptosis in Human Coronary Artery Endothelial Cells by Regulating the Expression of Caspase 3

doi: 10.3390/ijms160817682

Figure Lengend Snippet: The expression of miR-30b and caspase-3 mRNA after overexpression of miR-30b in Hcy-induced HCAECs. The concentration of Hcy was 1 mmol/L, both the time of transfection and treatment of Hcy were 24 h. ( A ) shows the transfection of miR-30b mimic significantly up-regulated the expression of miR-30b which Hcy reduced; and ( B ) shows the overexpression of miR-30b down-regulated the expression of caspase-3 mRNA which Hcy induced. Compared with the control group, compared with the Hcy + miR-30b negative group, p < 0.05, n = 6 in each group.

Article Snippet: Then, the PVDF membrane was incubated overnight at 4 °C with anti-β-actin (Sigma, Saint Louis, MO, USA), anti-caspase-3 antibody (Cell Signaling Technology, Shanghai, China) diluted by Tris-Buffered Saline and Tween (TBST).

Techniques: Expressing, Over Expression, Concentration Assay, Transfection

Journal: International Journal of Molecular Sciences

Article Title: MiR-30b Is Involved in the Homocysteine-Induced Apoptosis in Human Coronary Artery Endothelial Cells by Regulating the Expression of Caspase 3

doi: 10.3390/ijms160817682

Figure Lengend Snippet: The expression of procaspase-3 and cleaved caspase-3 protein after overexpression of miR-30b in Hcy-induced HCAECs, the concentration of Hcy was 1 mmol/L, both the time of transfection and treatment of Hcy were 24 h.

Article Snippet: Then, the PVDF membrane was incubated overnight at 4 °C with anti-β-actin (Sigma, Saint Louis, MO, USA), anti-caspase-3 antibody (Cell Signaling Technology, Shanghai, China) diluted by Tris-Buffered Saline and Tween (TBST).

Techniques: Expressing, Over Expression, Concentration Assay, Transfection

Journal: International Journal of Molecular Sciences

Article Title: MiR-30b Is Involved in the Homocysteine-Induced Apoptosis in Human Coronary Artery Endothelial Cells by Regulating the Expression of Caspase 3

doi: 10.3390/ijms160817682

Figure Lengend Snippet: Primer for real-time qPCR.

Article Snippet: Then, the PVDF membrane was incubated overnight at 4 °C with anti-β-actin (Sigma, Saint Louis, MO, USA), anti-caspase-3 antibody (Cell Signaling Technology, Shanghai, China) diluted by Tris-Buffered Saline and Tween (TBST).

Techniques: Sequencing

Journal: The Journal of Experimental Medicine

Article Title: Cerebral Ischemia Enhances Polyamine Oxidation: Identification of Enzymatically Formed 3-Aminopropanal as an Endogenous Mediator of Neuronal and Glial Cell Death

doi:

Figure Lengend Snippet: Inhibition of caspase 1 but not of caspase 3 blocks 3-aminopropanal–induced glial apoptosis. Cells were pretreated with ( a ) the caspase 1 inhibitor (Ac-YVAD-CMK) or ( b ) the caspase 3 inhibitor (Ac-DEVD-CHO) at concentrations 0.4 ( triangles ) or 40 μM ( circles ) for 3 h, followed by treatment with 3-aminopropanal for an additional 5 h, and then were analyzed for cell viability by the 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide assay. Controls consisted of DMSO-treated cells ( squares ) to assess for nonspecific solvent effects. Data are mean ± SE, n = 3 wells/experiment.

Article Snippet: For all experiments using a short duration of 3-aminopropanal exposure (5 min to 2 h in 96-well plates), the cells were washed at the times indicated, and then incubated in Opti-MEM I for up to 20 h. Where indicated, cells were pretreated with the caspase 1 inhibitor II Ac-YVAD-CMK (BACHEM, Torrance, CA) or the caspase-3 inhibitor Ac-DEVD-CHO (Peptides International, Louisville, KY) in DMSO for 3 h followed by addition of 3-aminopropanal for an additional 5 h. DMSO vehicle controls were performed concurrently.

Techniques: Inhibition

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Berberine Protects Against Palmitate-Induced Apoptosis in Tubular Epithelial Cells by Promoting Fatty Acid Oxidation

doi: 10.12659/MSM.908927

Figure Lengend Snippet: Berberine ameliorated palmitate-induced apoptosis in HK-2 cells. ( A ) Representative Western blot analyses of cleaved-caspase3 expression in HK-2 cells treated with different concentrations of palmitate for 24 h. ( B ) Densitometric analysis of cleaved-caspase3 expression in Figure A . ( C ) Representative Western blot analyses of cleaved-caspase3 protein expression in HK-2 cells treated with or without 450 μmol/L palmitate in the presence or absence of 12.5 μmol/L berberine. ( D ) Densitometric analysis of cleaved-caspase3 expression in Figure C . ( E ) Representative cytograms of apoptosis. ( F ) Quantification of cell apoptosis. All the statistical data are expressed as the mean ±SEM; n=3. * p<0.05 vs. 0 μmol/L or control group, & p<0.05 vs. 300 μmol/L, # p<0.05 vs. PA group.

Article Snippet: Then, the protein samples were mixed with loading buffer, heated at 100°C for 5 min, separated by electrophoresis in 10% or 12% tris-glycine polyacrylamide gradient gels, transferred onto PVDF membrane (Millipore, USA), blocked with 5% non-fat milk for 1 h at room temperature, and then incubated overnight at 4°C with different primary antibodies: mouse anti-carnitine palmitoyltransferase 1A (CPT1A, 1: 1000, Abcam, UK), rabbit anti-cleaved-caspase3 (1: 1000, CST, USA), rabbit anti-peroxisome proliferator-activated receptor α (PPARα, 1: 500, Santa Cruz, USA), rabbit anti-PPARγ co-activator-1α (PGC1α, 1: 1000, Novus, USA), or mouse anti-β-actin (1: 5000, Sungene Biotech, China).

Techniques: Western Blot, Expressing

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Berberine Protects Against Palmitate-Induced Apoptosis in Tubular Epithelial Cells by Promoting Fatty Acid Oxidation

doi: 10.12659/MSM.908927

Figure Lengend Snippet: CPT1A inhibitor counteracted the protective effect of BBR against PA-induced lipid accumulation and apoptosis in HK-2 cells. HK-2 cells were pretreated with or without 40 μmol/L Etomoxir for 24 h, and then treated with 12.5 μmol/L BBR in the presence or absence of 450 μmol/L PA for 24 h. Intracellular lipid content was detected by Oil Red O staining (400×) ( A ) and Nile Red staining (400×) ( B ). The lower panels showed enlarged views of the boxed areas in the upper panels in Figure A . The rightmost panels showed that enlarged views of the boxed areas in the left panels in Figure B . ( C ) Fluorescent intensity from 5 randomly selected microscopic fields per group in Figure B was captured and analyzed. ( D ) Representative Western blot analyses of cleaved-caspase3 protein expression. ( E ) Densitometric analysis of cleaved-caspase3 protein expression in Figure D . ( F ) Representative cytograms of apoptosis. ( G ) Quantification of cell apoptosis. All the statistical data are shown as the mean ±SEM, n=3. * p<0.05 vs. control group. # p<0.05 vs. PA group. & p<0.05 vs. PA+BBR group.

Article Snippet: Then, the protein samples were mixed with loading buffer, heated at 100°C for 5 min, separated by electrophoresis in 10% or 12% tris-glycine polyacrylamide gradient gels, transferred onto PVDF membrane (Millipore, USA), blocked with 5% non-fat milk for 1 h at room temperature, and then incubated overnight at 4°C with different primary antibodies: mouse anti-carnitine palmitoyltransferase 1A (CPT1A, 1: 1000, Abcam, UK), rabbit anti-cleaved-caspase3 (1: 1000, CST, USA), rabbit anti-peroxisome proliferator-activated receptor α (PPARα, 1: 500, Santa Cruz, USA), rabbit anti-PPARγ co-activator-1α (PGC1α, 1: 1000, Novus, USA), or mouse anti-β-actin (1: 5000, Sungene Biotech, China).

Techniques: Staining, Western Blot, Expressing