Repeated Intratracheal Instillation of PM10 Induces Lipid Reshaping in Lung Parenchyma and in Extra-Pulmonary Tissues

<div><p>Adverse health effects of air pollution attributed mainly to airborne particulate matter have been well documented in the last couple of decades. Short term exposure, referring to a few hours exposure, to high ambient PM10 concentration is linked to increased hospitalization rates for cardiovascular events, typically 24 h after air pollution peaks. Particulate matter exposure is related to pulmonary and cardiovascular diseases, with increased oxidative stress and inflammatory status. Previously, we have demonstrated that repeated intratracheal instillation of PM10sum in BALB/c mice leads to respiratory tract inflammation, creating in lung a condition which could potentially evolve in a systemic toxic reaction. Additionally, plasma membrane and tissue lipids are easily affected by oxidative stress and directly correlated with inflammatory products. With this aim, in the present investigation using the same model, we analyzed the toxic potential of PM10sum exposure on lipid plasma membrane composition, lipid peroxidation and the mechanisms of cells protection in multiple organs such as lung, heart, liver and brain. Obtained results indicated that PM10 exposure led to lung lipid reshaping, in particular phospholipid and cholesterol content increases; concomitantly, the generation of oxidative stress caused lipid peroxidation. In liver we found significant changes in lipid content, mainly due to an increase of phosphatidylcholine, and in total fatty acid composition with a more pronounced level of docosahexaenoic acid; these changes were statistically correlated to lung molecular markers. Heart and brain were similarly affected; heart was significantly enriched in triglycerides in half of the PM10sum treated mice. These results demonstrated a direct involvement of PM10sum in affecting lipid metabolism and oxidative stress in peripheral tissues that might be related to the serious systemic air-pollution effects on human health.</p></div

Phospholipid fatty acid composition of Hearts from Sham and PM10sum- treated mice.

<p>n.d. not detected;</p><p>*p<0.05;</p><p>**p<0.01 PM10 vs. Sham.</p><p>Phospholipid fatty acid composition of Hearts from Sham and PM10sum- treated mice.</p

Immunoblotting analysis in lung, liver, brain and heart parenchyma from sham and PM10sum-treated mice, 24 h after the third intratracheal instillation.

<p>(A) Western blot analysis of HO-1, Cyp1B1 as optical density (OD) quantified by Kodak Image Station. The proteins have been normalized to β-actin and each protein in PM10 treated group has been normalized onto respective sham group. (B) Representative Western blotting showing HO-1, Cyp1B1 and β-actin in lung, liver, brain and heart parenchyma from sham and PM10sum-treated mice, 24 h after the third intratracheal instillation. All the data are expressed as mean ± S.E. Sham vs. PM10sum-treated: * = p<0.05.</p

Histology of lung and heart tissue of sham (A, C) and PM10sum-treated (B, D) mice, 24 h after the third intratracheal instillation.

<p>Each figure represents the status evidenced examining 6 sham and 6 PM10sum-treated mice. A and B bars  = 50 µm. C and D bars  = 25 µm. Lung results modified from Farina et al., 2013 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0106855#pone.0106855-Farina1" target="_blank">[24]</a>.</p

Fatty acid composition changes in lung, liver, heart and brain of PM10sum-treated mice.

<p>Data were calculated as percentage variation relative to mean sham value. U.I. =  Unsaturation Index (sum of the % unsaturated fatty acids multiplied by their number of double bonds). Statistical analysis was performed on percentage fatty acid distribution by two way ANOVA coupled to Bonferroni's test. n = 6 for PM10sum and n = 5 for Sham mice. * = p<0.05. Mean data and statistical results reported in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0106855#pone.0106855.s002" target="_blank">Table S1</a>.</p

Correlation Heat Map Matrix between Blood and lung parameter couple with liver heart and brain data.

<p>Pearson's correlations were estimated using SPSS platform and transformed in a color code matrix. Only correlation with an R greater than 0.6 were coded; positive correlation are indicated with a red scale (red indicates statistical significant ones with p<0.05 or 0.01), while negative correlation are indicated with a blue scale (dark blue indicates statistical significant ones with p<0.05 or 0.01).</p

Protein, DNA, and lipid subclass changes in lung, liver, heart and brain of PM10sum-treated mice.

<p>Data were calculated as percentage variation relative to mean sham value. Statistical analysis was performed on actual amounts by a two way ANOVA coupled to Bonferroni's test. n = 6 for PM10sum and n = 5 for Sham mice. * = p<0.05. Mean data and statistical results reported in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0106855#pone.0106855.s002" target="_blank">Table S1</a>.</p

Phospholipid fatty acid composition of Lung from Sham and PM10sum- treated mice.

<p>n.d. not detected;</p><p>*p<0.05;</p><p>**p<0.01 PM10 vs. Sham.</p><p>Phospholipid fatty acid composition of Lung from Sham and PM10sum- treated mice.</p

Triglyceride (TAG) content and total fatty acid composition (%) of Heart from Sham and PM10sum treated mice. PM10 mice were divided into 2 groups: A) with low content of Triglycerides and B) with high content of Triglycerides.

<p>U.I. =  Unsaturation Index (sum of the % unsaturated fatty acids multiplied by their number of double bonds).</p><p>Mean ± S.D.</p><p>*p<0.05;</p><p>**p<0.01 vs. Sham.</p><p>Triglyceride (TAG) content and total fatty acid composition (%) of Heart from Sham and PM10sum treated mice. PM10 mice were divided into 2 groups: A) with low content of Triglycerides and B) with high content of Triglycerides.</p

Catalase (Panel A, U/mg Hb+SD) and Superoxide dismutase (Panel B U/g HB+SD) activities in PTN and wild type mice erythrocytes after space flight.

<p>MDS-ISS (black bars), Ground MDS controls (line bars) and vivarium (white bars). * p<0.05 MDS-ISS vs Ground MDS control. Statistical analysis performed only on PTN mice.</p