Figure 2.

Characterization of the Dual Human NKX2.5^EGFP/+-COUP-TFII^mCherry/+ Stem Cell Line (COUP-red) as Atrial Reporter

(A) Atrial-directed pin-embryoid body (spin-EB) protocol and treatment with retinoic acid (RA).

(B) mCherry expression together with GFP and bright-field (BF) images at D7 and D14 of differentiation in RA and control (CT) differentiations. Scale bar, 100 μm.

(C) Representative flow cytometry plots depicting the percentage of GFP-positive (G⁺) or mCherry (M⁺) cells at D14 of differentiation in RA or CT differentiations.

(D) Averaged G/M percentage calculated from four independent differentiations (n = 4): RA condition (G⁺/M⁺: 41% ± 2%, G⁻/M⁺: 52% ± 3%, G⁺/M⁻: 4% ± 1%, G⁻/M⁻: 4% ± 1%) versus CT (G⁺/M⁺: 12% ± 1%, G⁻/M⁺: 14% ± 2%, G⁺/M⁻: 61% ± 4%, G⁻/M⁻: 13% ± 2%). Data are displayed as means ± SEM

(E) Relative mRNA expression of cardiac troponin (TNNT2) in sorted G/M populations at day 20 of differentiation. Data are displayed as means ± SEM; ^∗p < 0.05.

(F) Immunostaining of cardiac troponin (cTnI) together with endogenous GFP expression and DAPI as nuclear staining in unsorted cultures after dissociation and re-plating. Scale bars, 25 μm (left), 2.5 μm (right).

See also Figure S3.