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. 2021 Aug 24;36(8):109610. doi: 10.1016/j.celrep.2021.109610

Figure 2.

Figure 2

Dynamic expression of CXCR4 and CXCL12 in the OFT and developing SLVs

(A) Schematic representation of E10.5–E11.5 heart (sagittal view) showing endocardium (green), endocardial cushions (yellow), and distal-proximal axis of OFT. Sectors (a), (b), and (c) identify distal, medial, and proximal OFT cushion, respectively; transverse section from region (b) (medial OFT) is also depicted. Double-headed arrow indicates valve-forming region.

(B–D’’) E11.5 wild-type OFT immunostained with anti-CXCR4 and PECAM-1 antibodies: distal region (a) in (B)–(B”); medial region (b) in (C)–(C”); and proximal region (c) in (D)–(D”). Boxed areas in (B)–(D) are shown enlarged in neighboring panels; double-headed arrows in (B) and (C) indicate valve primordia. Arrows in (B’), (B”), (C’), (C”), (D’), and (D”) show CXCR4+ ECs; yellow dashed lines indicate sub-endocardial CXCR4+ MCs, and white dashed lines indicate endocardium.

(E) Bar graphs show relative distribution of CXCR4+ MCs (PECAM-1−) and ECs (PECAM-1+) within the OFT (valve-forming region and proximal OFT cushion) at E11.5.

(F–G’) E12.5 (F–F’’) and E14.5 (G and G’) AoV, immunostained as above. Faint puncta in (F’) are shown enlarged in inset box; arrows in (F)–(F’’) indicate CXCR4+ ECs in SLV commissures.

(H) Sagittal section of E11.5 wild-type OFT (immunostained as above). Boxed areas showing distal (i) and proximal (ii) OFT are enlarged in neighboring panels. Arrowheads indicate CXCR4+ MCs (PECAM-1−) in proximal cushion.

(I) Graph shows proportion of CXCR4+ MCs with punctate CXCR4 in valve-forming and proximal cushion regions of E11.5 OFT (n = 4) and E12.5 SLVs (n = 4) as a percentage of total CXCR4+ MCs. Error bars represent ± SD; ∗∗∗p ≤ 0.001 (unpaired Student’s t test).

(J–N) Cxcl12GFP/+ E11.5 OFT cushion (J–L) and E12.5 SLV (M and N) sections immunostained with anti-GFP, anti-CXCR4, and anti-PECAM-1 antibodies. Boxed region in (K) is shown enlarged in neighboring panels; arrowheads indicate GFP+/CXCR4+ cells.

(O) Schematic showing relative levels of punctate or membranous CXCR4 (red) in MCs and ECs, and CXCL12 in the OFT wall and cushions (green) in distal, medial, and proximal OFT.

Ant, anterior; EC, endocardial cell; MC, mesenchymal cell; NC, non-coronary; OFT, outflow tract; RA, right atrium; RV, right ventricle. Scale bars represent 100 μm. See also Figure S2.