Journal: Cell Cycle

Article Title: Isoproterenol directs hair follicle-associated pluripotent (HAP) stem cells to differentiate in vitro to cardiac muscle cells which can be induced to form beating heart-muscle tissue sheets

doi: 10.1080/15384101.2016.1146837

Figure Lengend Snippet: The combination of isoproterenol, activin A, BMP4, and bFGF induced HAP stem cells in vibrissa hair follicles to form beating cardiac-muscle tissue sheets. (A) Schematic diagram of upper parts of five hair follicles forming cardiac-muscle tissue sheets (upper panel). Bright-field image of upper part of hair follicle differentiatimg to cardiac muscle cells and immunostaining (middle panel). Red = cTnT, Blue = DAPI, Bars = 100 µm. Cardiac-muscle tissue sheet formation process (lower panel). Bar = 500 µm. (B) Immunostaining of beating cardiac-muscle tissue sheets. cTnT-positive cardiac muscle cells were extensively distributed within the sheets. Red = cTnT, Blue = DAPI. Bars = 100 µm.

Article Snippet: Beating cardiac muscle cells and cardiac-muscle tissue sheets were visualized and recorded with a BioStation IM-Q (Nikon, Tokyo, Japan) and video microscope camera (ScopPad-500, GelleX, Tokyo, Japan).

Techniques: Immunostaining

Journal: Cell Cycle

Article Title: Isoproterenol directs hair follicle-associated pluripotent (HAP) stem cells to differentiate in vitro to cardiac muscle cells which can be induced to form beating heart-muscle tissue sheets

doi: 10.1080/15384101.2016.1146837

Figure Lengend Snippet: Flow cytometry analysis of beating cardiac-muscle tissue sheets. (A) Flow cytometry analysis of harvested cardiac-muscle tissue sheets showed that the sheets contain 38.88 ± 6.64% cTnT-positive cells (left panel); 22.19 ± 6.36% CD31-positive cells (middle panel); and 29.58 ± 6.11% SMA-positive cells (right panel). (B) Intercellular Ca2+ imaging of cardiac-muscle tissue sheets. Phase contrast image of cardiac muscle cells loaded with Flou-3 (white dashed area). Images were obtained every 150 msec (left panel). Bar =100 µm. Fluo-3 image at 4 points (I, II, III, IV) showing time course of Fluo-3 intensity change (middle and left panel). (C) High rate of formation of the beating cardiac-muscle tissue sheets from HAP stem cells. Hair follicles were cultured in medium supplemented with isoproterenol, activin A, BMP4 and bFGF, where cardiac-muscle tissue sheets formed at 65.0 ± 5.77%. No cardiac tissue sheets formed in non-supplemented medium or medium supplemented with isoproterenol alone.

Article Snippet: Beating cardiac muscle cells and cardiac-muscle tissue sheets were visualized and recorded with a BioStation IM-Q (Nikon, Tokyo, Japan) and video microscope camera (ScopPad-500, GelleX, Tokyo, Japan).

Techniques: Flow Cytometry, Imaging, Cell Culture

Journal: Angewandte Chemie (International ed. in English)

Article Title: Ultrafluorogenic Coumarin-Tetrazine Probes for Real-Time Biological Imaging **

doi: 10.1002/anie.201403890

Figure Lengend Snippet: a) Mitochondrial imaging: OVCA-429 cells with RFP-tagged mitochondria were incubated with an anti-mitochondria-TCO antibody, rinsed briefly, and then imaged after addition of 100nM HELIOS-388H in PBS. Colocalization analysis in ImageJ (Costes auto-threshold) was used to generate the merged image in the right-hand panel. b) Actin imaging: COS-1 cells were incubated with phalloidin-TCO (1µg/mL) and DRAQ5 nuclear counterstain (1µM, BioStatus), rinsed briefly, and then imaged upon addition of the indicated HELIOS probe at 100nM. Control images were collected at matched dye concentrations in the absence of phalloidin-TCO.

Article Snippet: Colocalization analysis in ImageJ (Costes auto-threshold) was used to generate the merged image in the right-hand panel. b) Actin imaging: COS-1 cells were incubated with phalloidin-TCO (1µg/mL) and DRAQ5 nuclear counterstain (1µM, BioStatus), rinsed briefly, and then imaged upon addition of the indicated HELIOS probe at 100nM.

Techniques: Imaging, Incubation

Journal: Toxicology and applied pharmacology

Article Title: Video-based kinetic analysis of calcification in live osteogenic human embryonic stem cell cultures reveals the developmentally toxic effect of Snus tobacco extract

doi: 10.1016/j.taap.2018.11.006

Figure Lengend Snippet: (A) No detrimental effects of nicotine on culture morphology could be detected in photomicrographs of cultures in the tested range. (B) Time-course calcification of nicotine treated osteogenically differentiating hESCs as shown every 12 hours. (C) Segmented areas in images of nicotine treated samples normalized to the solvent control. (D) Calcification rates of representative concentrations determined from images taken at 12h intervals. (B-D) show selected concentrations only for ease of readability. (E) Concentration-response calcification curve across all tested concentrations generated using image-quantified calcium measurement. (F) Arsenazo III reagent-based endpoint calcification curve. B-E n=3 biological replicates 10 technical replicates ea; F n=3, 5 technical replicates ea ± SD. *P<0.05, One Way ANOVA versus untreated. Statistics shown only for d10 and d20 for ease of readability.

Article Snippet: To do so, live cultures of osteogenically induced and non-osteogenically induced hESCs were imaged as time-lapse videos using the Nikon Biostation CT to follow their commitment into the osteogenic lineage.

Techniques: Concentration Assay, Generated