Journal: Stem cell research & therapy

Article Title: High glucose suppresses embryonic stem cell differentiation into cardiomyocytes : High glucose inhibits ES cell cardiogenesis.

doi: 10.1186/s13287-016-0446-5

Figure Lengend Snippet: Fig. 4 High glucose suppresses cardiomyocyte-specific marker expression. a mRNA levels of cardiomyocyte structure proteins, MEF2C, and TNNT2. b Representative images of TNNT2 immunofluorescent staining. c Quantitative fluorescent density data of TNNT2 immunofluorescent staining. d MEF2C and TNNT2 protein expression at differentiation day 9. Gene expression was assessed at EB formation day 5 and differentiation day 1, 3, 5, and 7. Experiments were repeated three times (n = 3). EB embryoid body derived from E14 or GR-E14 cells, HG high glucose (25 mM), LG low glucose (5 mM). Values were dedicated as mean ± SD. *Indicates significant difference compared with the other group

Article Snippet: Samples were blocked for 1 hour with 10% heat-inactivated donkey serum in PBS and incubated with the following antibodies: TNNT2 (1:200, Origene, Rockville, MD, USA), NKX2.5 (1:200, Sigma-Aldrich, St. Louis, MO, USA), and HCN1 (1:200, Sigma-Aldrich, St. Louis, MO, USA), overnight at 4 °C.

Techniques: Marker, Expressing, Staining, Gene Expression, Derivative Assay

Journal: Circulation Research

Article Title: Induction of Cardiomyocyte-Like Cells in Infarct Hearts by Gene Transfer of Gata4, Mef2c, and Tbx5

doi: 10.1161/circresaha.112.271148

Figure Lengend Snippet: Figure 5. Induction of cardiomyocyte-like cells by a polycistronic vector. A, Schematic representation of the 3F2A retrovirus containing GMT for polycistronic gene expression. B, Western blot analysis of 293 cells transfected with 3F2A for 2 days. C, Immunocytochemistry for Gata4 and vimentin in mouse fibroblasts transduced with 3F2A. D, Gata4 and Mef2c were expressed in the same cells after transduction of 3F2A. E, 3F2A retrovirus induced α-actinin expression and cross striations. The cell, indicated by arrows in the first and second panels, is enlarged in the third to fifth panels. White boxes are enlarged in the insets. F, Z-stack image of the GFP+/ α-actinin+ cell in E. G and H, Quantitative analyses of the α-actinin+ to GFP+ cell ratios (G) and the proportions of α-actinin+/GFP+ cells showing cross striations (H) (120 sections from 5 mice in control, 140 section from 5 mice in GMT, and 110 sections from 5 mice in 3F2A). I, The 3F2A-expressing cells also showed cTnT expression. White boxes in the first panels are enlarged in the second to fourth panels. J, Schematic representation to analyze mRNA expression in GFP- or 3F2A/GFP-infected cells. GFP+ cells were sorted by FACS after 1 week. K, 3F2A/GFP-transduced cells expressed significantly higher cardiac genes compared with controls determined by qPCR (n=3). Representative data are shown in each panel. All data are presented as mean±SEM. *P<0.05;**P<0.01 versus control. Scale bars, 50 μm.

Article Snippet: Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, blocked, and then incubated with primary antibodies against α-actinin, GFP, cTnT, vimentin, CD31, SMA (Sigma), collagen 1, Mef2c (Aviva Systems Biology), Gata4 (Santa Cruz Biotechnology), ANP (Chemicon), Nkx2.5 (Santa Cruz), or SM- MHC (Biomedical Technologies), with secondary antibodies conjugated to Alexa 488 or 546.

Techniques: Plasmid Preparation, Gene Expression, Western Blot, Transfection, Immunocytochemistry, Transduction, Expressing, Control, Infection