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Fig. 2 | Stem Cell Research & Therapy

Fig. 2

From: High glucose suppresses embryonic stem cell differentiation into cardiomyocytes

Fig. 2

High glucose suppresses GR-E14 cell cardiogenesis. a Schematic diagram of ES cell cardiomyocytes. Hanging-drop culture was performed with one drop of 30 μl medium per 1000 cells for 3 days for EB formation, and then suspension culture was done in a 10-cm petri dish for another 2 days for EB growth followed by differentiation. b The numbers of EB attached to the culture plate on day 1 after seeding the formed EBs (differentiation). c The numbers of EB attached to the culture plate on day 5 of differentiation. GR-E14 cells adapted to low glucose were used for hanging-drop culture in low glucose (5 mM, LG) or high glucose (25 mM) medium (LG2HG), parent E14 cells maintained in high glucose medium were used for hanging-drop culture in HG or low glucose medium (HG2LG). d The percentage of contracting EBs in GR-E14 cells under LG/HG condition. Fifty EBs per group (n = 3) were used for cardiogenesis and the contracting EB percentage was calculated by the formula: (number of contracting EBs/number of total EBs) × 100%. e Numbers of contracting colonies from GR-E14 cells under LG/HG conditions. At day 5 of EB formation, an aliquot of 10 μl EB suspension, containing 20 EBs (equivalent to 5 × 105 cells), was seeded into a well of a 6-well culture plate for cardiogenesis. f Percentage of contracting colonies from GR-E14 cells under LG/HG conditions. The contracting colony percentage was calculated by the formula: (number of contracting colonies/number of total colonies) × 100%. g Frequencies of contracting colonies from GR-E14 under LG/HG conditions. h The frequencies of TNNT2+ cardiomyocytes at day 6 (left), day 8 (middle) and day 10 (right) after differentiation. HG high glucose (25 mM, red), LG low glucose (5 mM, gray),. i Percentages of TNNT2+ cardiomyocytes differentiated from GR-E14 cells. TNNT2+ cell percentages (%) = (TNNT2+ cell number/total cell number) × 100%. Total cell number is 10,000 per group. j The percentage of contracting EBs in D3 cells under LG/HG conditions. The protocol was the same as that of GR-E14. k Percentage of contracting colonies from D3 cells under LG/HG conditions. The protocol was the same as that of GR-E14. E14 E14 cells maintained under 25 mM glucose condition, GR-E14 glucose-responsive E14 cell line, which was gradually adapted to 5 mM glucose medium from 25 mM glucose medium. All experiments were repeated three times (n = 3). Data were expressed as mean ± SD. *Indicate significant difference compared with the other group(s)

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