The effect of the postnatal environment on altered fetal programming of adult vascular function in mice that lack endothelial nitric oxide synthase

Abstract

Objective: The purpose of this study was to investigate vascular reactivity in heterozygous and homozygous offspring with a genetic predisposition for hypertension after postnatal cross-fostering to mothers with the opposite genetic inheritance of the NOS3 knockout allele.

Study design: Homozygous NOS3 knockout (C57BL/6J-NOS3(-/-KO)) and wild-type mice (NOS3(+/+WT)) were bred to obtain heterozygous litters with a paternally derived (NOS3(+/-pat)) or maternally derived (NOS3(+/-mat)) knockout allele. After delivery, heterozygous and homozygous litters were cross-fostered to a mother with the opposite NOS3 gene status. Carotid arteries were placed in a wire myograph for isometric tension recording with the use of contractile and relaxant agents. Statistical analysis with 1-way analysis of variance and Neuman-Keuls post-hoc testing was performed.

Results: Increased sensitivity to phenylephrine and absent relaxation to acetylcholine in NOS3(+/-mat) was reversed with cross-fostering, and vasorelaxation to isoproterenol was increased. Contraction to calcium was increased in the cross-fostered paternally derived and wild-type litters.

Conclusion: Postnatal interventions may alter the adult vascular profile favorably that is the result of an abnormal intrauterine environment.

Figure 1

Schematic diagram of breeding of first generation heterozygous and homozygous mice from homozygous wild-type and knockout mice. The litters from the first generation were cross fostered to homozygous wild-type and knockout mothers to obtain the offspring used in the experiments.

Figure 2

Figure 2A. Average daily weight (grams) of cross fostered offspring from day 1 to 19 of life in NOS3^FKO/−pat (n=17), NOS3^FWT/−mat (n=19), NOS3^FWT/−KO (n=13), and NOS3^FKO/+WT (n=10) male and female pups. One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FKO/+WT vs. all other groups, †P < 0.05 NOS3^FKO/−pat vs. NOS3^FWT/−mat and NOS3^FWT/−KO, ‡P < 0.05 NOS3^FWT/−mat vs. NOS3^FWT/−KO. Figure 2B. Average weekly weight (grams) of female offspring from weeks 4 to 9 of adult life in NOS3^FKO/−pat (n=7), NOS3^FWT/−mat (n=12), NOS3^FWT/−KO (n=8), and NOS3^FKO/+WT (n=8). One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FKO/+WT vs. all other groups, †P < 0.05 NOS3^FKO/−pat vs. NOS3^FWT/−mat and NOS3^FWT/−KO.

Figure 3

Figure 3A. Phenylephrine (PE) concentration-response curves in the carotid arteries of NOS3^FKO/−pat (n=8), NOS3^FWT/−mat (n=7), NOS3^FWT/−KO (n=8), and NOS3^FKO/+WT (n=6) female mice. One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FWT/−KO vs. all other groups. †P < 0.05 NOS3^FKO/+WT vs. all other groups. Figure 3B. Phenylephrine (PE) concentration-response curves in the carotid arteries of NOS3^FKO/−pat (n=8), NOS3^FWT/−mat (n=7), NOS3^FWT/−KO (n=8), and NOS ^FKO/+WT (n=6) female mice in the presence of the non-specific NOS inhibitor L-NAME. One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FKO/+WT vs. all other groups.

Figure 4

Acetylcholine (Ach) concentration-response curves in the carotid arteries of NOS3^FKO/−pat (n=10), NOS3^FWT/−mat (n=10), NOS3^FWT/−KO (n=7), and NOS3^FKO/+WT (n=4) female mice. One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FKO/−pat vs. NOS3^FWT/−mat and NOS3^FWT/−KO, †P < 0.05 NOS3^FKO/+WT vs. NOS3^FWT/−mat and NOS3^FWT/−KO, ‡P < 0.05 NOS3^FWT/−mat vs. NOS3^FWT/−KO.

Figure 5

Isoproteronol (Isop) concentration-response curves in the carotid arteries of the NOS3^FKO/−pat (n=8), NOS3^FWT/−mat (n=7), NOS3^FWT/−KO (n=8), and NOS ^FKO/+WT (n=5) female mice. One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FKO/+WT vs. all other groups. †P < 0.05 NOS3^FKO/−pat vs. all other groups.

Figure 6

Calcium (Ca2+) concentration-response curves in the carotid arteries of NOS3^FKO/−pat (n=7), NOS3^FWT/−mat (n=8), NOS3^FWT/−KO (n=4), and NOS3^FKO/+WT (n=3) female mice. One-way ANOVA followed by Newman-Keuls tests were used. *P < 0.05 NOS3^FKO/−pat vs. NOS3^FWT/−mat.