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. 2022 Feb 22;11(5):e024008. doi: 10.1161/JAHA.121.024008

Figure 2. GSNOR−⁄− mice exhibited placental insufficiency during pregnancy and showed alteration of VEGF pathway.

Figure 2

Nonpregnant and pregnant (17.5 d) (N=4–12 mothers per group) C57Bl/6J (B6) and GSNOR−⁄− (knockout [KO]) mice were examined. A, Fetal number and (B) weight were significantly lower in KO mice. C, NADH‐dependent GSNOR enzymatic activity was determined in placental tissue at 17.5 days of gestation. GSNOR activity is enriched in B6 placentas, whereas it is completely absent in the KO placentas. D, E, Placental weight was not significantly different between the 2 strains, whereas placental efficiency was significantly lower in KO mice as compared with control. F, G, Placental vascularization determined using isolectin immunostaining, was significantly lower in KO as compared with B6 at late gestation. SNO‐VEGF was measured using Biotin‐switch assay. H, Representative blots shown for S‐nitrosylated and total VEGF in the placenta. Omission of ascorbate was used as the negative control. I, Nitrosylation of VEGF was significantly lower in KO placentas as compared with B6 at late gestation. J, K, VEGFR2 and eNOS protein levels were determined in the placentas at 17.d of gestation. Both VEGFR2 and eNOS protein levels were significantly lower in in GSNOR−⁄− placentas as compared with B6 placentas at 17.5 days of gestation. For statistical significance, 2‐way ANOVA with Newman‐Keuls for post hoc analysis or Student’s t test were performed. Results are shown as mean±SEM. ***P<0.001, **P<0.01, *P<0.05. B6 indicates C57Bl/6J mice; GSNOR, S‐nitrosoglutathione reductase; KO, GSNOR−⁄− mice; and NP, nonpregnant.