Does early mismatched nutrition predispose to hypertension and atherosclerosis, in male mice?

Abstract

Background: A link between early mismatched nutritional environment and development of components of the metabolic syndrome later in life has been shown in epidemiological and animal data. The aim of this study was to investigate whether an early mismatched nutrition produced by catch-up growth after fetal protein restriction could induce the appearance of hypertension and/or atherosclerosis in adult male mice.

Methodology/principal findings: Wild-type C57BL6/J or LDLr-/- dams were fed a low protein (LP) or a control (C) diet during gestation. Catch-up growth was induced in LP offspring by feeding dams with a control diet and by culling the litter to 4 pups against 8 in controls. At weaning, male mice were fed either standard chow or an obesogenic diet (OB), leading to 4 experimental groups. Blood pressure (BP) and heart rate (HR) were assessed in conscious unrestrained wild-type mice by telemetry. Atherosclerosis plaque area was measured in aortic root sections of LDLr-/- mice. We found that: (1) postnatal OB diet increased significantly BP (P<0.0001) and HR (P<0.008) in 3-month old OB-C and OB-LP offspring, respectively; (2) that maternal LP diet induced a significant higher BP (P<0.009) and HR (P<0.004) and (3) an altered circadian rhythm in addition to higher plasma corticosterone concentration in 9 months-old LP offspring; (4) that, although LP offspring showed higher plasma total cholesterol than control offspring, atherosclerosis assessed in aortic roots of 6-mo old mice featured increased plaque area due to OB feeding but not due to early mismatched nutrition.

Conclusions/significance: These results indicate a long-term effect of early mismatched nutrition on the appearance of hypertension independently of obesity, while no effect on atherosclerosis was noticed at this age.

Figure 1. Blood glucose concentration and area under the curve (AUC) during intraperitoneal glucose tolerance test.

The test was performed in 3-mo (A and B), 9-mo (C and D) C57BL6 male mice and 6-mo LDLr−/− (E and F). Values are presented as mean ± SEM with n = 5 to 8 individuals per group. Statistical analysis by Two-way ANOVA indicates a significant influence of post-weaning OB diet for AUCs at 3-mo (B; P<0.0001) and 9-mo (D; P<0.0001) in C57BL6 mice (exp 1) and at 6-mo (F; P<0.0001) in LDLr−/− mice (exp 2).

Figure 2. Long-term recording blood pressures and heart rate in 9-mo male C57BL6 mice (exp1).

Recording of SBP (A), DBP (B) pressure and HR (C) were assessed during 24 h in n = 5 animals per group. Values are presented as mean values of SBP, DBP and HR calculated for each 60-minutes sequence of recording. Shaded zone on the X-axis indicates night period (activity period for mice).

Figure 3. Short-term recording of SBP and HR after an acute treatment with L-NAME (A–D) and phenylephrine (E–H).

Recording of cardiovascular parameters was assessed in 9-mo male C57BL6 mice (exp1) during 1 h in n = 5 animals per group. Values are presented as mean values of SBP or HR ± SEM calculated for each 5 minutes sequence of recording. Arrows indicate the time of injection. Δ values of SBP and HR were calculated by subtracting the mean values of the 5 time-point after to the mean values of the 5 time-point before injection. Statistical analysis by Two-way ANOVA indicates a significant influence of post-weaning OB diet (P = 0.0127) as well as a significant influence of maternal LP diet (P = 0.0429) for ΔHR after phenylephrine treatment.

Figure 4. Atherosclerosis plaque area assessment in aortic root of 6-mo LDLr−/− male mice.

(A) Representative images of oil red-O staining of the aortic root sections in the 4 experimental groups (scale bar = 200 µm). (B) The plaque area values are presented as mean values ± SEM for n = 6 animals per group. Two-way ANOVA indicates a significant influence of post-weaning OB diet (P = 0.0002) on plaque surface.

Figure 5. Relative abundance of gene mRNA assessed in livers of 6-mo LDLr−/− male mice.

Values are presented as mean relative abundance ± SEM for n = 6 animals per group. Two-way ANOVA indicated a significant influence of post weaning OB diet for SREBP-1c (P = 0.0002), HMG-CoA synthase (P = 0.004) and PPARγ (P = 0.0002) as well as a significant influence of maternal LP diet for SREBP-2 (P = 0.034) and HMG-CoA synthase (P = 0.0406). HMG-CoA reductase showed a significant interaction effect (P = 0.0243). Bonferroni post-test has been performed to assess statistical differences in mRNA relative abundance within chow fed vs. OB fed animals within C and LP groups. * P<0.05 and ** P<0.01.