In utero and early life exposure to diesel exhaust air pollution increases adult susceptibility to heart failure in mice

Abstract

Background: Fine particulate air pollution (PM2.5) is a global health concern, as exposure to PM2.5 has consistently been found to be associated with increased cardiovascular morbidity and mortality. Although adult exposure to traffic related PM2.5, which is largely derived from diesel exhaust (DE), has been associated with increased cardiac hypertrophy, there are limited investigations into the potential effect of in utero and early life exposure on adult susceptibility to heart disease. In this study, we investigate the effect of in utero and early life exposure to DE on adult susceptibility to heart failure.

Methods: Female C57BL/6 J mice were exposed to either filtered air (FA) or DE for 3 weeks (≈ 300 μg/m3 PM2.5 for 6 hours/day, 5 days/week) and then introduced to male breeders for timed matings. Female mice were exposed to either FA or DE throughout pregnancy and until offspring were 3 weeks of age. Offspring were then transferred to either FA or DE for an additional 8 weeks of exposure. At 12 weeks of age, male offspring underwent a baseline echocardiographic assessment, followed by a sham or transverse aortic constriction (TAC) surgery to induce pressure overload. Following sacrifice three weeks post surgery, ventricles were processed for histology to assess myocardial fibrosis and individual cardiomyocyte hypertrophy. mRNA from lung tissue was isolated to measure expression of inflammatory cytokines IL6 and TNFα.

Results: We observed that mice exposed to DE during in utero and early life development have significantly increased susceptibility to cardiac hypertrophy, systolic failure, myocardial fibrosis, and pulmonary congestion following TAC surgery compared to FA control, or adult DE exposed mice. In utero and early life DE exposure also strongly modified the inflammatory cytokine response in the adult lung.

Conclusions: We conclude that exposure to diesel exhaust air pollution during in utero and early life development in mice increases adult susceptibility to heart failure. The results of this study may imply that the effects of air pollution on cardiovascular disease in human populations may be strongly mediated through a 'fetal origins' of adult disease pathway. Further investigations on this potential pathway of disease are warranted.

Figure 1

Schematic diagram indicating diesel exhaust (DE) exposure paradigm. Four exposure groups tested the effect of developmental or adult exposure to DE or filtered air (FA) exposures on adult cardiac function. Brown color in horizontal arrows represents DE exposure (300 μg/m³, 6 hours per day, 5 days per week), while white color represents FA exposure.

Figure 2

Echocardiographic assessment of cardiac hypertrophy and function at 12 week of age, prior to TAC/Sham surgeries. (A) Calculated LV Mass normalized to body weight and (B) percentage ejection fraction (%EF) in FA/FA (n = 21), FA/DE (n = 7), DE/FA (n = 14), and DE/DE (n = 12) groups.

Figure 3

Echocardiographic assessment of calculated LV Mass normalized to body weight over time following transverse aortic constriction (TAC) or sham surgeries. (A) FA/FA (n = 7 sham, n = 8 TAC) vs. FA/DE (n = 3 sham, n = 3 TAC), (B) FA/FA vs. DE/FA (n = 5 sham, n = 5 TAC), (C) FA/FA vs. DE/DE (n = 5 sham, n = 4 TAC) groups, and (D) DE/FA vs. DE/DE.

Figure 4

Percentage ejection fraction (%EF) determined by echocardiography over time following transverse aortic constriction (TAC) or sham surgeries. (A) FA/FA (n = 7 sham, n = 8 TAC) vs. FA/DE (n = 3 sham, n = 3 TAC), (B) FA/FA vs. DE/FA (n = 5 sham, n = 5 TAC), (C) FA/FA vs. DE/DE (n = 5 sham, n = 4 TAC) groups, and (D) DE/FA vs. DE/DE.

Figure 5

Gravimetric analysis of ventricle weight at necropsy. Representative images of sagittal sections of hearts from mice three weeks post sham or transverse aortic constriction (TAC) surgeries (A-H), scale bar = 1 mm. Gravimetric analysis of ventricles normalized to tibia length (VW/tibia length) in FA/FA (n = 7 sham, n = 8 TAC), FA/DE (n = 3 sham, n = 3 TAC), DE/FA (n = 5 sham, n = 5 TAC) and DE/DE (n = 5 sham, n = 4 TAC) groups (I). Effect of TAC by exposure group was also represented as a fold effect of TAC on VW/tibia length normalized to sham mice (J).

Figure 6

Assessment of cardiac fibrosis by Masson’s Trichrome staining. Representative images (4X objective) of sagittal sections of left ventricle free wall from mice three weeks post sham or transverse aortic constriction (TAC) surgeries (A-H), scale bar = 0.5 mm. Fibrotic regions of cardiac tissues are determined by blue staining in FA/FA (n = 5 sham, n = 5 TAC), FA/DE (n = 3 sham, n = 3 TAC), DE/FA (n = 5 sham, n = 5 TAC) and DE/DE (n = 5 sham, n = 4 TAC) groups. Assessment of percentage highly fibrotic regions was quantified (I).

Figure 7

Measurement of individual cardiomyocyte hypertrophy. Representative images (40X objective) of sagittal sections of left ventricle free wall from mice three weeks post sham or transverse aortic constriction (TAC) surgeries (A-H), scale bar = 50 μm. Individual cardiomyocyte areas were traced, quantified, and analyzed in FA/FA (n = 5 sham, n = 5 TAC), FA/DE (n = 3 sham, n = 3 TAC), DE/FA (n = 5 sham, n = 5 TAC) and DE/DE (n = 5 sham, n = 4 TAC) groups (I).

Figure 8

Effects of DE and TAC on lung weight and inflammatory cytokine expression. (A) Lung weights as measured at necropsy were collected and analyzed in FA/FA (n = 7 sham, n = 8 TAC), FA/DE (n = 3 sham, n = 3 TAC), DE/FA (n = 5 sham, n = 5 TAC) and DE/DE (n = 5 sham, n = 4 TAC) groups. Relative real-time PCR analysis of IL6 (B) and TNFα (C) mRNA content in the lung as expressed as negative delta delta CT from FA/FA sham normalized to GAPDH.