Inhaled diesel exhaust particles result in microbiome-related systemic inflammation and altered cardiovascular disease biomarkers in C57Bl/6 male mice

Abstract

Background The gut microbiota plays a vital role in host homeostasis and is associated with inflammation and cardiovascular disease (CVD) risk. Exposure to particulate matter (PM) is a known mediator of inflammation and CVD and is reported to promote dysbiosis and decreased intestinal integrity. However, the role of inhaled traffic-generated PM on the gut microbiome and its corresponding systemic effects are not well-characterized. Thus, we investigated the hypothesis that exposure to inhaled diesel exhaust particles (DEP) alters the gut microbiome and promotes microbial-related inflammation and CVD biomarkers. 4–6-week-old male C57Bl/6 mice on either a low-fat (LF, 10% fat) or high-fat (HF, 45% fat) diet were exposed via oropharyngeal aspiration to 35 μg DEP suspended in 35 μl saline or saline only (CON) 2x/week for 30 days. To determine whether probiotics could prevent diet or DEP exposure mediated alterations in the gut microbiome or systemic outcomes, a subset of animals on the HF diet were treated orally with 0.3 g/day (~ 7.5 × 108 CFU/day) of Winclove Ecologic® Barrier probiotics throughout the study. Results Our results show that inhaled DEP exposure alters gut microbial profiles, including reducing Actinobacteria and expanding Verrucomicrobia and Proteobacteria. We observed increased circulating LPS, altered circulating cytokines (IL-1α, IL-3, IL-13, IL-15, G-CSF, LIF, MIP-2, and TNF-α), and CVD biomarkers (siCAM, PAI-1, sP-Selectin, thrombomodulin, and PECAM) in DEP-exposed and/or HF diet mice. Furthermore, probiotics attenuated the observed reduction of Actinobacteria and expansion of Proteobacteria in DEP-exposed and HF-diet mice. Probiotics mitigated circulating cytokines (IL-3, IL-13, G-CSF, RANTES, and TNF- α) and CVD biomarkers (siCAM, PAI-1, sP-Selectin, thrombomodulin, and PECAM) in respect to DEP-exposure and/or HF diet. Conclusion Key findings of this study are that inhaled DEP exposure alters small intestinal microbial profiles that play a role in systemic inflammation and early CVD biomarkers. Probiotic treatment in this study was fundamental in understanding the role of inhaled DEP on the microbiome and related systemic inflammatory and CVD biomarkers.

Probiotics Function as Immunomodulators in the Intestine in C57Bl/6 Male Mice Exposed to Inhaled Diesel Exhaust Particles on a High-Fat Diet

“…Thus, we propose that the HF diet is likely contributing to the increased baseline expression of TNF-α in the intestines of the CON group in this study. Moreover, we suggest that the reduction in TNF-α with DEP exposure is correlated to the observed expansion in Escherichia, which we previously reported [37]. When comparing PRO + CON to CON, we find a significant reduction in TNF-α expression, which we attribute to the known anti-inflammatory effects of probiotics, namely Lactococcus and Lactobacillus strains [65,66].…”

“…Furthermore, we observe a significant increase in TNF-α with DEP + PRO compared to its respective CON + PRO control, suggesting that the immunomodulatory effects of probiotics normalized the immune response in the intestines which resulted in an augmented response with DEP exposure. We have previously reported that probiotic treatment mitigates the expansion of Escherichia observed with DEP exposure in the HF diet mice, confirming the possible role of Escherichia in TNF-α signaling in these mice [37].…”

“…Another study found that the fecal microbiome of MMP-9 −/− mice showed an expansion of Proteobacteria compared to wild-type mice, providing evidence that MMP-9 expression is correlative to the presence of certain microbial profiles. Interestingly, we previously reported expansion of Proteobacteria with DEP exposure in the intestines from these same study mice, yet we observe an increase in MMP-9 protein expression throughout the duodenum and jejunum [37]. We see the highest expression of MMP-9 in the duodenum of DEP-exposed animals, which could result from the ingestion of particles encountering the duodenum at the highest concentration before dispersing as luminal contents move throughout the small intestine.…”

“…We observed a global and regional increase in MUC2 protein expression with DEP exposure ± probiotics. From these same study animals, we previously reported an increase in Akkermansia with DEP exposure ± probiotics [37]. Akkermansia is a novel bacteria known for its mucin degrading capabilities and importance in mucosal integrity homeostasis [43].…”

“…Some 4-6-w-old C57Bl/6 wild-type male mice (Taconic, New York, NY, USA, C57BL/NTac) on a high-fat (HF) diet consisting of 45% fat (HF, Research Diets #D12451, n = 48) were randomly assigned to be exposed via oropharyngeal aspiration (OA) to 35 µg diesel exhaust particles purchased from National Institute of Standards and Technology (NIST, Standard reference material #2975), suspended in 35 µL 0.9% sterile saline (n = 24), or sterile saline only (CON, n = 24) twice a week for 4 w. The exposure methodology for this study has been previously published by our lab [36,37]. Briefly, animals were housed four to a cage, had access to food and water ad libitum, and were maintained on a 12-h light/dark cycle in humidity and temperature-controlled rooms.…”