Figure 1.

Lz expansion is associated with increasing levels of circulating and endothelial IGF2

(A) Weights of micro-dissected Lz.

(B) Linear correlation analyses between fetal and Lz weights: p = 0.002 (E14), p < 0.0001 (E16), and p < 0.0001 (E19) (n = 46–189 placentae from n > 10 L per group in [A] and [B]).

(C) Levels of IGF2 (ng/mL) in plasma of wild-type fetuses.

(D) Linear correlation analyses between fetal weights and circulating IGF2: p < 0.0001 (E16 and E19) (n = 70–79 per group in [C] and [D]).

(E) Igf2 mRNA in situ hybridization (blue) in E14 wild-type Lz (red arrows—FPEC, feto-placental endothelial cells; AS, antisense probe; inset with S, sense probe; scale bars, 20 μm).

(F) Relative Igf2 mRNA expression levels measured by qRT-PCR in FPEC from wild-type Lz (n = 6–7 per group).

(G) Imprinted genes that rank within top 100 expressed genes in E16 wild-type FPEC (FPKM, fragments per kilobase million; n = 4).

(H) Double immunostaining for IGF2 and CD31 in E19 wild-type placenta. Endothelial cells are very thin and hard to detect except where the cytoplasm is more voluminous around the nucleus, with intense IGF2 stain (white arrows). Transmembrane glycoprotein CD31 immunostaining is in the membrane and largely marks endothelial intercellular junctions (scale bars, 20 μm).

(I) Semi-quantitative measurement of IGF2 protein in FPEC versus trophoblast cells (E19 wild-type Lz, n = 60 cells per group from two placentae). White arrows—endothelial cells; scale bars, 50 μm. For (E), (H), and (I): FC, fetal capillaries; MBS, maternal blood spaces; LT, labyrinthine trophoblast cells; S-TGC, sinusoidal trophoblast giant cells. Data in (A), (C), (F), (G), and (I) are presented as averages ± standard deviation (SD); ^∗∗∗p < 0.001 calculated by one-way ANOVA plus Tukey’s multiple comparisons test in (A) and (F) or by unpaired t test with Welch’s correction in (C) and (I). See also Table S1.