Probing changes in brain esterified oxylipin concentrations during the early stages of pathogenesis in Alzheimer's Disease transgenic rats.

Despite known pathological hallmarks of Alzheimer's Disease (AD) including neuronal loss, gliosis (inflammation), beta-amyloid plaque deposition and neurofibrillary tangle accumulation in the brain, little is known about inflammation resolution in early AD pathogenesis. In the brain, inflammation and resolution pathways are mediated by free oxylipins which are mostly bound (i.e. esterified), and therefore must be released (i.e. become free) to exert bioactivity. Recently, we showed reductions in brain esterified pro-resolving oxylipins in a transgenic rat model of AD (TgF344-AD rat) at 15 months of age, suggesting deficits in the source and availability of free pro-resolving oxylipins. In the present study, we tested whether these changes are discernable earlier in the disease process, i.e., at age of 10 months. We observed significant reductions in esterified pro-resolving 8(9)-epoxyeicosatrienoic acid (8(9)-EpETrE), 13-hydroxyoctadecatrienoic acid (13-HOTrE) and 15-hydroxyeicosapentaenoic acid (15-HEPE) oxylipins, and in pro-inflammatory 13-hydroxy-octadecadienoic acid (13-HODE), 20-hydroxy-eicosatetraenoic acid (20-HETE), 15-deoxy-prostaglandin J2 (15-deoxy-PGJ2) and prostaglandin E2 (PGE2) oxylipins in male and/or female transgenic AD rats compared to wildtype controls. These findings point to a deficit in esterified pro-resolving lipid mediators in the early stages of AD, concident with. changes in esterified lipid mediators involved in promoting inflammation.

Hippocampal but Not Serum Cytokine Levels Are Altered by Traffic-Related Air Pollution in TgF344-AD and Wildtype Fischer 344 Rats in a Sex- and Age-Dependent Manner.

Epidemiological studies have demonstrated that air pollution is a significant risk factor for age-related dementia, including Alzheimer's disease (AD). It has been posited that traffic-related air pollution (TRAP) promotes AD neuropathology by exacerbating neuroinflammation. To test this hypothesis, serum and hippocampal cytokines were quantified in male and female TgF344-AD rats and wildtype (WT) Fischer 344 littermates exposed to TRAP or filtered air (FA) from 1 to 15 months of age. Luminex™ rat 23-cytokine panel assays were used to measure the levels of hippocampal and serum cytokines in 3-, 6-, 10-, and 15-month-old rats (corresponding to 2, 5, 9, and 14 months of exposure, respectively). Age had a pronounced effect on both serum and hippocampal cytokines; however, age-related changes in hippocampus were not mirrored in the serum and vice versa. Age-related changes in serum cytokine levels were not influenced by sex, genotype, or TRAP exposure. However, in the hippocampus, in 3-month-old TgF344-AD and WT animals, TRAP increased IL-1ß in females while increasing TNF ɑin males. In 6-month-old animals, TRAP increased hippocampal levels of M-CSF in TgF344-AD and WT females but had no significant effect in males. At 10 and 15 months of age, there were minimal effects of TRAP, genotype or sex on hippocampal cytokines. These observations demonstrate that TRAP triggers an early inflammatory response in the hippocampus that differs with sex and age and is not reflected in the serum cytokine profile. The relationship of TRAP effects on cytokines to disease progression remains to be determined.

Chronic exposure to traffic-related air pollution reduces lipid mediators of linoleic acid and soluble epoxide hydrolase in serum of female rats.

Chronic exposure to traffic-related air pollution (TRAP) is known to promote systemic inflammation, which is thought to underlie respiratory, cardiovascular, metabolic and neurological disorders. It is not known whether chronic TRAP exposure dampens inflammation resolution, the homeostatic process for stopping inflammation and repairing damaged cells. In vivo, inflammation resolution is facilitated by bioactive lipid mediators known as oxylipins, which are derived from the oxidation of polyunsaturated fatty acids. To understand the effects of chronic TRAP exposure on lipid-mediated inflammation resolution pathways, we measured total (i.e. free+bound) pro-inflammatory and pro-resolving lipid mediators in serum of female rats exposed to TRAP or filtered air (FA) for 14 months. Compared to rats exposed to FA, TRAP-exposed rats showed a significant 36-48% reduction in fatty acid alcohols, specifically, 9-hydroxyoctadecadienoic acid (9-HODE), 11,12-dihydroxyeicosatetraenoic acid (11,12-DiHETE) and 16,17-dihydroxydocosapentaenoic acid (16, 17-DiHDPA). The decrease in fatty acid diols (11,12-DiHETE and 16, 17-DiHDPA) corresponded to a significant 34-39% reduction in the diol to epoxide ratio, a marker of soluble epoxide hydrolase activity; this enzyme is typically upregulated during inflammation. The findings demonstrate that 14 months exposure to TRAP reduced pro-inflammatory 9-HODE concentration and dampened soluble epoxide hydrolase activation, suggesting adaptive immune changes in lipid mediator pathways involved in inflammation resolution.

Chronic exposure to ambient traffic-related air pollution (TRAP) alters gut microbial abundance and bile acid metabolism in a transgenic rat model of Alzheimer's disease.

Background: Traffic-related air pollution (TRAP) is linked to increased risk for age-related dementia, including Alzheimer's disease (AD). The gut microbiome is posited to influence AD risk, and an increase in microbial-derived secondary bile acids (BAs) is observed in AD patients. We recently reported that chronic exposure to ambient TRAP modified AD risk in a sex-dependent manner in the TgF344 AD (TG) rat. Objectives: In this study, we used samples from the same cohort to test our hypothesis that TRAP sex-dependently produces gut dysbiosis and increases secondary BAs to a larger extent in the TG rat relative to wildtype (WT) controls. Methods: Male and female TG and age-matched WT rats were exposed to either filtered air (FA) or TRAP from 28 days up to 15 months of age (n = 5-6). Tissue samples were collected after 9 or 14months of exposure. Results: At 10 months of age, TRAP tended to decrease the alpha diversity as well as the beneficial taxa Lactobacillus and Ruminococcus flavefaciens uniquely in male TG rats as determined by 16 S rDNA sequencing. A basal decrease in Firmicutes/Bacteroidetes (F/B) ratio was also noted in TG rats at 10 months. At 15 months of age, TRAP altered inflammation-related bacteria in the gut of female rats from both genotypes. BAs were more affected by chronic TRAP exposure in females, with a general trend of increase in host-produced unconjugated primary and microbiota-produced secondary BAs. Most of the mRNAs of the hepatic BA-processing genes were not altered by TRAP, except for a down-regulation of the BA-uptake transporter Ntcp in males. Conclusion: In conclusion, chronic TRAP exposure produced distinct gut dysbiosis and altered BA homeostasis in a sex and host genotype-specific manner.

The Effects of Chronic Exposure to Ambient Traffic-Related Air Pollution on Alzheimer's Disease Phenotypes in Wildtype and Genetically Predisposed Male and Female Rats.

BACKGROUND: Epidemiological data link traffic-related air pollution (TRAP) to increased risk of Alzheimer's disease (AD). Preclinical data corroborating this association are largely from studies of male animals exposed acutely or subchronically to high levels of isolated fractions of TRAP. What remains unclear is whether chronic exposure to ambient TRAP modifies AD risk and the influence of sex on this interaction. OBJECTIVES: This study sought to assess effects of chronic exposure to ambient TRAP on the time to onset and severity of AD phenotypes in a preclinical model and to determine whether sex or genetic susceptibility influences outcomes. METHODS: Male and female TgF344-AD rats that express human AD risk genes and wildtype littermates were housed in a vivarium adjacent to a heavily trafficked tunnel in Northern California and exposed for up to 14 months to filtered air (FA) or TRAP drawn from the tunnel and delivered to animals unchanged in real time. Refractive particles in the brain and AD phenotypes were quantified in 3-, 6-, 10-, and 15-month-old animals using hyperspectral imaging, behavioral testing, and neuropathologic measures. RESULTS: Particulate matter (PM) concentrations in TRAP exposure chambers fluctuated with traffic flow but remained below 24-h PM with aerodynamic diameter less than or equal to 2.5 micrometers (PM2.5) U.S. National Ambient Air Quality Standards limits. Ultrafine PM was a predominant component of TRAP. Nano-sized refractive particles were detected in the hippocampus of TRAP animals. TRAP-exposed animals had more amyloid plaque deposition, higher hyperphosphorylated tau levels, more neuronal cell loss, and greater cognitive deficits in an age-, genotype-, and sex-dependent manner. TRAP-exposed animals also had more microglial cell activation, but not astrogliosis. DISCUSSION: These data demonstrate that chronic exposure to ambient TRAP promoted AD phenotypes in wildtype and genetically susceptible rats. TRAP effects varied according to age, sex, and genotype, suggesting that AD progression depends on complex interactions between environment and genetics. These findings suggest current PM2.5 regulations are insufficient to protect the aging brain. https://doi.org/10.1289/EHP8905.

Pathological Cardiopulmonary Evaluation of Rats Chronically Exposed to Traffic-Related Air Pollution.

BACKGROUND: Traffic-related air pollution (TRAP) is made up of complex mixtures of particulate matter, gases and volatile compounds. However, the effects of TRAP on the cardiopulmonary system in most animal studies have been tested using acute exposure to singular pollutants. The cardiopulmonary effects and molecular mechanisms in animals that are chronically exposed to unmodified air pollution as a whole have yet to be studied. Additionally, sex-dependent toxicity of TRAP exposure has rarely been evaluated. OBJECTIVES: This study sought to assess the cardiopulmonary effect of chronic exposure to unmodified, real-world TRAP in both female and male rats. METHODS: Four-week-old male and female rats were exposed to TRAP or filtered air for 14 months in a novel facility drawing air from a major freeway tunnel system in Northern California. Inflammation and oxidative stress markers were examined in the lung, heart, spleen, and plasma, and TRAP deposits were quantified in the lungs of both male and female rats. RESULTS: Elemental analysis showed higher levels of eight elements in the female lungs and one element in the male lungs. Expression of genes related to fibrosis, aging, oxidative stress, and inflammation were higher in the rat hearts exposed to TRAP, with female rats being more susceptible than males. Enhanced collagen accumulation was found only in the TRAP-exposed female hearts. Plasma cytokine secretion was higher in both female and male rats, but inflammatory macrophages were higher only in TRAP-exposed male spleens. DISCUSSION: Our results in rats suggest pathological consequences from chronic TRAP exposure, including sex differences indicating females may be more susceptible to TRAP-induced cardiac fibrosis. https://doi.org/10.1289/EHP7045.

Developmental exposure to near roadway pollution produces behavioral phenotypes relevant to neurodevelopmental disorders in juvenile rats.

Epidemiological studies consistently implicate traffic-related air pollution (TRAP) and/or proximity to heavily trafficked roads as risk factors for developmental delays and neurodevelopmental disorders (NDDs); however, there are limited preclinical data demonstrating a causal relationship. To test the effects of TRAP, pregnant rat dams were transported to a vivarium adjacent to a major freeway tunnel system in northern California where they were exposed to TRAP drawn directly from the face of the tunnel or filtered air (FA). Offspring remained housed under the exposure condition into which they were born and were tested in a variety of behavioral assays between postnatal day 4 and 50. To assess the effects of near roadway exposure, offspring of dams housed in a standard research vivarium were tested at the laboratory. An additional group of dams was transported halfway to the facility and then back to the laboratory to control for the effect of potential transport stress. Near roadway exposure delayed growth and development of psychomotor reflexes and elicited abnormal activity in open field locomotion. Near roadway exposure also reduced isolation-induced 40-kHz pup ultrasonic vocalizations, with the TRAP group having the lowest number of call emissions. TRAP affected some components of social communication, evidenced by reduced neonatal pup ultrasonic calling and altered juvenile reciprocal social interactions. These findings confirm that living in close proximity to highly trafficked roadways during early life alters neurodevelopment.

Effects of early life exposure to traffic-related air pollution on brain development in juvenile Sprague-Dawley rats.

Epidemiological studies link traffic-related air pollution (TRAP) to increased risk for various neurodevelopmental disorders (NDDs); however, there are limited preclinical data demonstrating a causal relationship between TRAP and adverse neurodevelopmental outcomes. Moreover, much of the preclinical literature reports effects of concentrated ambient particles or diesel exhaust that do not recapitulate the complexity of real-world TRAP exposures. To assess the developmental neurotoxicity of more realistic TRAP exposures, we exposed male and female rats during gestation and early postnatal development to TRAP drawn directly from a traffic tunnel in Northern California and delivered to animals in real-time. We compared NDD-relevant neuropathological outcomes at postnatal days 51-55 in TRAP-exposed animals versus control subjects exposed to filtered air. As indicated by immunohistochemical analyses, TRAP significantly increased microglial infiltration in the CA1 hippocampus, but decreased astrogliosis in the dentate gyrus. TRAP exposure had no persistent effect on pro-inflammatory cytokine levels in the male or female brain, but did significantly elevate the anti-inflammatory cytokine IL-10 in females. In male rats, TRAP significantly increased hippocampal neurogenesis, while in females, TRAP increased granule cell layer width. TRAP had no effect on apoptosis in either sex. Magnetic resonance imaging revealed that TRAP-exposed females, but not males, also exhibited decreased lateral ventricular volume, which was correlated with increased granule cell layer width in the hippocampus in females. Collectively, these data indicate that exposure to real-world levels of TRAP during gestation and early postnatal development modulate neurodevelopment, corroborating epidemiological evidence of an association between TRAP exposure and increased risk of NDDs.

Cell-extrinsic consequences of epithelial stress: activation of protumorigenic tissue phenotypes.

INTRODUCTION: Tumors are characterized by alterations in the epithelial and stromal compartments, which both contribute to tumor promotion. However, where, when, and how the tumor stroma develops is still poorly understood. We previously demonstrated that DNA damage or telomere malfunction induces an activin A-dependent epithelial stress response that activates cell-intrinsic and cell-extrinsic consequences in mortal, nontumorigenic human mammary epithelial cells (HMECs and vHMECs). Here we show that this epithelial stress response also induces protumorigenic phenotypes in neighboring primary fibroblasts, recapitulating many of the characteristics associated with formation of the tumor stroma (for example, desmoplasia). METHODS: The contribution of extrinsic and intrinsic DNA damage to acquisition of desmoplastic phenotypes was investigated in primary human mammary fibroblasts (HMFs) co-cultured with vHMECs with telomere malfunction (TRF2-vHMEC) or in HMFs directly treated with DNA-damaging agents, respectively. Fibroblast reprogramming was assessed by monitoring increases in levels of selected protumorigenic molecules with quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, and immunocytochemistry. Dependence of the induced phenotypes on activin A was evaluated by addition of exogenous activin A or activin A silencing. In vitro findings were validated in vivo, in preinvasive ductal carcinoma in situ (DCIS) lesions by using immunohistochemistry and telomere-specific fluorescent in situ hybridization. RESULTS: HMFs either cocultured with TRF2-vHMEC or directly exposed to exogenous activin A or PGE2 show increased expression of cytokines and growth factors, deposition of extracellular matrix (ECM) proteins, and a shift toward aerobic glycolysis. In turn, these "activated" fibroblasts secrete factors that promote epithelial cell motility. Interestingly, cell-intrinsic DNA damage in HMFs induces some, but not all, of the molecules induced as a consequence of cell-extrinsic DNA damage. The response to cell-extrinsic DNA damage characterized in vitro is recapitulated in vivo in DCIS lesions, which exhibit telomere loss, heightened DNA damage response, and increased activin A and cyclooxygenase-2 expression. These lesions are surrounded by a stroma characterized by increased expression of α smooth muscle actin and endothelial and immune cell infiltration. CONCLUSIONS: Thus, synergy between stromal and epithelial interactions, even at the initiating stages of carcinogenesis, appears necessary for the acquisition of malignancy and provides novel insights into where, when, and how the tumor stroma develops, allowing new therapeutic strategies.