Malignant rhabdoid tumor: Cyto-histologic correlation and immunohistochemical characterization of a rare pediatric malignancy and its differential diagnoses.

Malignant rhabdoid tumor of the kidney (MRTK) is a rare aggressive pediatric renal tumor which can be diagnosed via fine-needle aspiration (FNA) cytology and core biopsy. The diagnosis of MRTK is challenging, and requires morphologic, immunohistochemical and clinical correlation to distinguish it from other entities. The differential diagnosis includes Wilms tumor, desmoplastic small round cell tumor, rhabdomyosarcoma, synovial sarcoma, renal medullary carcinoma, and epithelioid sarcoma. Here we describe a case of MRTK diagnosed on renal cytology and core biopsy with immunohistochemistry and follow by nephrectomy with gross and morphologic findings.

TGF-ß Promotes Metabolic Reprogramming in Lung Fibroblasts via mTORC1-dependent ATF4 Activation.

Idiopathic pulmonary fibrosis is a fatal interstitial lung disease characterized by the TGF-ß (transforming growth factor-ß)-dependent differentiation of lung fibroblasts into myofibroblasts, which leads to excessive deposition of collagen proteins and progressive scarring. We have previously shown that synthesis of collagen by myofibroblasts requires de novo synthesis of glycine, the most abundant amino acid found in collagen protein. TGF-ß upregulates the expression of the enzymes of the de novo serine-glycine synthesis pathway in lung fibroblasts; however, the transcriptional and signaling regulators of this pathway remain incompletely understood. Here, we demonstrate that TGF-ß promotes accumulation of ATF4 (activating transcription factor 4), which is required for increased expression of the serine-glycine synthesis pathway enzymes in response to TGF-ß. We found that induction of the integrated stress response (ISR) contributes to TGF-ß-induced ATF4 activity; however, the primary driver of ATF4 downstream of TGF-ß is activation of mTORC1 (mTOR Complex 1). TGF-ß activates the PI3K-Akt-mTOR pathway, and inhibition of PI3K prevents activation of downstream signaling and induction of ATF4. Using a panel of mTOR inhibitors, we found that ATF4 activation is dependent on mTORC1, independent of mTORC2. Rapamycin, which incompletely and allosterically inhibits mTORC1, had no effect on TGF-ß-mediated induction of ATF4; however, Rapalink-1, which specifically targets the kinase domain of mTORC1, completely inhibited ATF4 induction and metabolic reprogramming downstream of TGF-ß. Our results provide insight into the mechanisms of metabolic reprogramming in myofibroblasts and clarify contradictory published findings on the role of mTOR inhibition in myofibroblast differentiation.

Current state of Grand Rounds in U.S. pathology training programs.

Grand Rounds are held with variable frequency in many academic pathology departments, but their exact goal is uncertain, and the type of subjects covered, and presenters have not been studied. We aimed to gather information about the current state of pathology grand rounds (PGR). We identified all US pathology residency programs accredited by the Accreditation Council for Graduate Medical Education (ACGME) and searched their websites for information regarding PGR, extracting data on their existence, frequency and timing. For a representative subgroup of institutions from all US regions and program sizes, we tabulated the 2017-2018 PGR titles and presenters (gender, degree(s), resident/fellow, faculty academic rank). We found that 71 of 142 (50%) ACGME-accredited programs had PGR, more often in programs with >12 residents (53/88, 60%). PGR were scheduled most commonly weekly, on Thursdays, and at noon. We analyzed 1019 PGR presentations from 41 institutions located in 26 US states. Among the 1105 presenters, 183 (16.56%) were trainees, 74 (6.7%) were non-academic, and 848 (76.7%) were faculty, 559 male and 289 female (M/F = 1.93). M/F ratio increased with academic rank, from 1.0 (117/115) for assistant, to 2.0 (135/68) for associate, and 2.9 (307/106) for full professors. Topics covered by PGR belonged to anatomic pathology (357), clinical pathology (209), research (184) or other medical or surgical specialties (149). Our study suggests that trainees are a major intended audience of pathology grand round. Unfortunately, there is a gender gap among pathology grand round presenters that widens with increasing academic rank of presenters.

Inhibition of Phosphoglycerate Dehydrogenase Attenuates Bleomycin-induced Pulmonary Fibrosis.

Organ fibrosis, including idiopathic pulmonary fibrosis, is associated with significant morbidity and mortality. Because currently available therapies have limited effect, there is a need to better understand the mechanisms by which organ fibrosis occurs. We have recently reported that transforming growth factor (TGF)-ß, a key cytokine that promotes fibrogenesis, induces the expression of the enzymes of the de novo serine and glycine synthesis pathway in human lung fibroblasts, and that phosphoglycerate dehydrogenase (PHGDH; the first and rate-limiting enzyme of the pathway) is required to promote collagen protein synthesis downstream of TGF-ß. In this study, we investigated whether inhibition of de novo serine and glycine synthesis attenuates lung fibrosis in vivo. We found that TGF-ß induces mRNA and protein expression of PHGDH in murine fibroblasts. Similarly, intratracheal administration of bleomycin resulted in increased expression of PHGDH in mouse lungs, localized to fibrotic regions. Using a newly developed small molecule inhibitor of PHGDH (NCT-503), we tested whether pharmacologic inhibition of PHGDH could inhibit fibrogenesis both in vitro and in vivo. Treatment of murine and human lung fibroblasts with NCT-503 decreased TGF-ß-induced collagen protein synthesis. Mice treated with the PHGDH inhibitor beginning 7 days after intratracheal instillation of bleomycin had attenuation of lung fibrosis. These results indicate that the de novo serine and glycine synthesis pathway is necessary for TGF-ß-induced collagen synthesis and bleomycin-induced pulmonary fibrosis. PHGDH and other enzymes in the de novo serine and glycine synthesis pathway may be a therapeutic target for treatment of fibrotic diseases, including idiopathic pulmonary fibrosis.

Experimental Lung Injury Reduces Krüppel-like Factor 2 to Increase Endothelial Permeability via Regulation of RAPGEF3-Rac1 Signaling.

RATIONALE: Acute respiratory distress syndrome (ARDS) is caused by widespread endothelial barrier disruption and uncontrolled cytokine storm. Genome-wide association studies (GWAS) have linked multiple genes to ARDS. Although mechanosensitive transcription factor Krüppel-like factor 2 (KLF2) is a major regulator of endothelial function, its role in regulating pulmonary vascular integrity in lung injury and ARDS-associated GWAS genes remains poorly understood. OBJECTIVES: To examine KLF2 expression in multiple animal models of acute lung injury and further elucidate the KLF2-mediated pathways involved in endothelial barrier disruption and cytokine storm in experimental lung injury. METHODS: Animal and in vitro models of acute lung injury were used to characterize KLF2 expression and its downstream effects responding to influenza A virus (A/WSN/33 [H1N1]), tumor necrosis factor-α, LPS, mechanical stretch/ventilation, or microvascular flow. KLF2 manipulation, permeability measurements, small GTPase activity, luciferase assays, chromatin immunoprecipitation assays, and network analyses were used to determine the mechanistic roles of KLF2 in regulating endothelial monolayer integrity, ARDS-associated GWAS genes, and lung pathophysiology. MEASUREMENTS AND MAIN RESULTS: KLF2 is significantly reduced in several animal models of acute lung injury. Microvascular endothelial KLF2 is significantly induced by capillary flow but reduced by pathologic cyclic stretch and inflammatory stimuli. KLF2 is a novel activator of small GTPase Ras-related C3 botulinum toxin substrate 1 by transcriptionally controlling Rap guanine nucleotide exchange factor 3/exchange factor directly activated by cyclic adenosine monophosphate, which maintains vascular integrity. KLF2 regulates multiple ARDS GWAS genes related to cytokine storm, oxidation, and coagulation in lung microvascular endothelium. KLF2 overexpression ameliorates LPS-induced lung injury in mice. CONCLUSIONS: Disruption of endothelial KLF2 results in dysregulation of lung microvascular homeostasis and contributes to lung pathology in ARDS.

Transforming Growth Factor (TGF)-ß Promotes de Novo Serine Synthesis for Collagen Production.

TGF-ß promotes excessive collagen deposition in fibrotic diseases such as idiopathic pulmonary fibrosis (IPF). The amino acid composition of collagen is unique due to its high (33%) glycine content. Here, we report that TGF-ß induces expression of glycolytic genes and increases glycolytic flux. TGF-ß also induces the expression of the enzymes of the de novo serine synthesis pathway (phosphoglycerate dehydrogenase (PHGDH), phosphoserine aminotransferase 1 (PSAT1), and phosphoserine phosphatase (PSPH)) and de novo glycine synthesis (serine hydroxymethyltransferase 2 (SHMT2)). Studies in fibroblasts with genetic attenuation of PHGDH or SHMT2 and pharmacologic inhibition of PHGDH showed that these enzymes are required for collagen synthesis. Furthermore, metabolic labeling experiments demonstrated carbon from glucose incorporated into collagen. Lungs from humans with IPF demonstrated increased expression of PHGDH and SHMT2. These results indicate that the de novo serine synthesis pathway is necessary for TGF-ß-induced collagen production and suggest that this pathway may be a therapeutic target for treatment of fibrotic diseases including IPF.

Lung-specific loss of α3 laminin worsens bleomycin-induced pulmonary fibrosis.

Laminins are heterotrimeric proteins that are secreted by the alveolar epithelium into the basement membrane, and their expression is altered in extracellular matrices from patients with pulmonary fibrosis. In a small number of patients with pulmonary fibrosis, we found that the normal basement membrane distribution of the α3 laminin subunit was lost in fibrotic regions of the lung. To determine if these changes play a causal role in the development of fibrosis, we generated mice lacking the α3 laminin subunit specifically in the lung epithelium by crossing mice expressing Cre recombinase driven by the surfactant protein C promoter (SPC-Cre) with mice expressing floxed alleles encoding the α3 laminin gene (Lama3(fl/fl)). These mice exhibited no developmental abnormalities in the lungs up to 6 months of age, but, compared with control mice, had worsened mortality, increased inflammation, and increased fibrosis after the intratracheal administration of bleomycin. Similarly, the severity of fibrosis induced by an adenovirus encoding an active form of transforming growth factor-ß was worse in mice deficient in α3 laminin in the lung. Taken together, our results suggest that the loss of α3 laminin in the lung epithelium does not affect lung development, but plays a causal role in the development of fibrosis in response to bleomycin or adenovirally delivered transforming growth factor-ß. Thus, we speculate that the loss of the normal basement membrane organization of α3 laminin that we observe in fibrotic regions from the lungs of patients with pulmonary fibrosis contributes to their disease progression.

Extraskeletal Ewing Sarcoma of the Gastrointestinal and Hepatobiliary Tract: Deceptive Immunophenotype Commonly Leads to Misdiagnosis.

Ewing sarcoma (ES) is an uncommon mesenchymal neoplasm that typically develops as a bone mass, although up to 30% arise in extraskeletal sites. ES of the gastrointestinal (GI) and hepatobiliary tract is rare and may be misdiagnosed as other, more common neoplasms that occur in these sites. However, the correct classification of extraskeletal ES is important for timely clinical management and prognostication. We reviewed our experience of ES in the GI and hepatobiliary tract in order to further highlight the clinicopathologic features of these neoplasms and document the potential for misdiagnosis in this setting. The archives and consultation files of 6 academic institutions were retrospectively queried for cases of ES occurring in the GI and hepatobiliary tract. The histologic slides and ancillary studies were reviewed and clinical data were retrieved for each case through the electronic medical records, when available. Twenty-three patients with ES in the GI and/or hepatobiliary tract were identified from 2000 to 2022. Of these, 11 were women and 12 were men with a median age of 38 years (range, 2 to 64). Tumor locations included the pancreas (n=5), liver (n=2), stomach (n=3), colorectum (n=3), and small intestine (n=5), as well as tumors involving multiple organs, pelvis and retroperitoneum (n=5). Tumor size varied between 2 cm and 18 cm. Twenty were primary and 3 were metastases. Of the 23 cases, only 17% were initially diagnosed as ES. The most common misdiagnoses involved various forms of neuroendocrine neoplasia due to expression of synaptophysin and other neuroendocrine markers (22%). A wide variety of diagnoses including GI stromal tumor was considered due to aberrant CD117 expression (4%). The diagnosis of ES was ultimately confirmed by detection of the EWSR1 rearrangement in 22 cases. The remaining case was diagnosed using traditional immunohistochemistry. Follow-up information was available in 20 cases, with follow-up time varying between 2 and 256 months. Six patients with follow-up died of disease between 6 and 60 months following initial presentation. Our data indicate ES in the GI and hepatobiliary tract is commonly misdiagnosed leading to a delay in therapy. In light of the attendant therapeutic and prognostic implications, ES should be considered in the differential diagnosis of any GI or hepatobiliary tumor with epithelioid and/or small round cell morphology.

Impacted gutkha presenting as an intrabronchial mass lesion leading to post-obstructive pneumonia.

A 66-year-old man presented with a chief complaint of difficulty breathing and productive cough. CT scan of the chest revealed an endobronchial mass with associated "tree-in-bud" opacities. A bronchoscopic biopsy of the mass was performed due to clinical suspicion of malignancy. Microscopic examination revealed inflamed endobronchial mucosa, granulation tissue and abundant fragments of uncharacterized organic material, compatible with aspiration. Detailed history revealed a history of chewing "gutkha", a form of smokeless tobacco comprising a mixture of betel nut and other condiments. Microscopic sections of a betel nut and the "gutkha mix" processed subsequently in the histology laboratory were found to be similar to the organic material found in the mass. Thus, a diagnosis of impacted betel nut mixture leading to post-obstructive pneumonia was rendered.

Particulate matter air pollution causes oxidant-mediated increase in gut permeability in mice.

BACKGROUND: Exposure to particulate matter (PM) air pollution may be an important environmental factor leading to exacerbations of inflammatory illnesses in the GI tract. PM can gain access to the gastrointestinal (GI) tract via swallowing of air or secretions from the upper airways or mucociliary clearance of inhaled particles. METHODS: We measured PM-induced cell death and mitochondrial ROS generation in Caco-2 cells stably expressing oxidant sensitive GFP localized to mitochondria in the absence or presence of an antioxidant. C57BL/6 mice were exposed to a very high dose of urban PM from Washington, DC (200 µg/mouse) or saline via gastric gavage and small bowel and colonic tissue were harvested for histologic evaluation, and RNA isolation up to 48 hours. Permeability to 4 kD dextran was measured at 48 hours. RESULTS: PM induced mitochondrial ROS generation and cell death in Caco-2 cells. PM also caused oxidant-dependent NF-κB activation, disruption of tight junctions and increased permeability of Caco-2 monolayers. Mice exposed to PM had increased intestinal permeability compared with PBS treated mice. In the small bowel, colocalization of the tight junction protein, ZO-1 was lower in the PM treated animals. In the small bowel and colon, PM exposed mice had higher levels of IL-6 mRNA and reduced levels of ZO-1 mRNA. Increased apoptosis was observed in the colon of PM exposed mice. CONCLUSIONS: Exposure to high doses of urban PM causes oxidant dependent GI epithelial cell death, disruption of tight junction proteins, inflammation and increased permeability in the gut in vitro and in vivo. These PM-induced changes may contribute to exacerbations of inflammatory disorders of the gut.

Inhalational exposure to particulate matter air pollution alters the composition of the gut microbiome.

Recent studies suggest an association between particulate matter (PM) air pollution and gastrointestinal (GI) disease. In addition to direct deposition, PM can be indirectly deposited in oropharynx via mucociliary clearance and upon swallowing of saliva and mucus. Within the GI tract, PM may alter the GI epithelium and gut microbiome. Our goal was to determine the effect of PM on gut microbiota in a murine model of PM exposure via inhalation. C57BL/6 mice were exposed via inhalation to either concentrated ambient particles or filtered air for 8-h per day, 5-days a week, for a total of 3-weeks. At exposure's end, GI tract tissues and feces were harvested, and gut microbiota was analyzed. Alpha-diversity was modestly altered with increased richness in PM-exposed mice compared to air-exposed mice in some parts of the GI tract. Most importantly, PM-induced alterations in the microbiota were very apparent in beta-diversity comparisons throughout the GI tract and appeared to increase from the proximal to distal parts. Changes in some genera suggest that distinct bacteria may have the capacity to bloom with PM exposure. Exposure to PM alters the microbiota throughout the GI tract which maybe a potential mechanism that explains PM induced inflammation in the GI tract.

Prolonged Exposures to Intermittent Hypoxia Promote Visceral White Adipose Tissue Inflammation in a Murine Model of Severe Sleep Apnea: Effect of Normoxic Recovery.

Study Objective: Increased visceral white adipose tissue (vWAT) mass results in infiltration of inflammatory macrophages that drive inflammation and insulin resistance. Patients with obstructive sleep apnea (OSA) suffer from increased prevalence of obesity, insulin resistance, and metabolic syndrome. Murine models of intermittent hypoxia (IH) mimicking moderate-severe OSA manifest insulin resistance following short-term IH. We examined in mice the effect of long-term IH on the inflammatory cellular changes within vWAT and the potential effect of normoxic recovery (IH-R). Methods: Male C57BL/6J mice were subjected to IH for 20 weeks, and a subset was allowed to recover in room air (RA) for 6 or 12 weeks (IH-R). Stromal vascular fraction was isolated from epididymal vWAT and mesenteric vWAT depots, and single-cell suspensions were prepared for flow cytometry analyses, reactive oxygen species (ROS), and metabolic assays. Results: IH reduced body weight and vWAT mass and IH-R resulted in catch-up weight and vWAT mass. IH-exposed vWAT exhibited increased macrophage counts (ATMs) that were only partially improved in IH-R. IH also caused a proinflammatory shift in ATMs (increased Ly6c(hi)(+) and CD36(+) ATMs). These changes were accompanied by increased vWAT insulin resistance with only partial improvements in IH-R. In addition, ATMs exhibited increased ROS production, altered metabolism, and changes in electron transport chain, which were only partially improved in IH-R. Conclusion: Prolonged exposures to IH during the sleep period induce pronounced vWAT inflammation and insulin resistance despite concomitant vWAT mass reductions. These changes are only partially reversible after 3 months of normoxic recovery. Thus, long-lasting OSA may preclude complete reversibility of metabolic changes.

HIF-1α is required for disturbed flow-induced metabolic reprogramming in human and porcine vascular endothelium.

Hemodynamic forces regulate vascular functions. Disturbed flow (DF) occurs in arterial bifurcations and curvatures, activates endothelial cells (ECs), and results in vascular inflammation and ultimately atherosclerosis. However, how DF alters EC metabolism, and whether resulting metabolic changes induce EC activation, is unknown. Using transcriptomics and bioenergetic analysis, we discovered that DF induces glycolysis and reduces mitochondrial respiratory capacity in human aortic ECs. DF-induced metabolic reprogramming required hypoxia inducible factor-1α (HIF-1α), downstream of NAD(P)H oxidase-4 (NOX4)-derived reactive oxygen species (ROS). HIF-1α increased glycolytic enzymes and pyruvate dehydrogenase kinase-1 (PDK-1), which reduces mitochondrial respiratory capacity. Swine aortic arch endothelia exhibited elevated ROS, NOX4, HIF-1α, and glycolytic enzyme and PDK1 expression, suggesting that DF leads to metabolic reprogramming in vivo. Inhibition of glycolysis reduced inflammation suggesting a causal relationship between flow-induced metabolic changes and EC activation. These findings highlight a previously uncharacterized role for flow-induced metabolic reprogramming and inflammation in ECs.

Intratracheal administration of influenza virus is superior to intranasal administration as a model of acute lung injury.

Infection of mice with human or murine adapted influenza A viruses results in a severe pneumonia. However, the results of studies from different laboratories show surprising variability, even in genetically similar strains. Differences in inoculum size related to the route of viral delivery (intranasal vs. intratracheal) might explain some of this variability. To test this hypothesis, mice were infected intranasally or intratracheally with different doses of influenza A virus (A/WSN/33 [H1N1]). Daily weights, a requirement for euthanasia, viral load in the lungs and brains, inflammatory cytokines, wet-to-dry ratio, total protein and histopathology of the infected mice were examined. With all doses of influenza tested, intranasal delivery resulted in less severe lung injury, as well as smaller and more variable viral loads in the lungs when compared with intratracheal delivery. Virus was not detected in the brain following either method of delivery. It is concluded that compared to intranasal infection, intratracheal infection with influenza A virus is a more reliable method to deliver virus to the lungs.

Impaired clearance of influenza A virus in obese, leptin receptor deficient mice is independent of leptin signaling in the lung epithelium and macrophages.

RATIONALE: During the recent H1N1 outbreak, obese patients had worsened lung injury and increased mortality. We used a murine model of influenza A pneumonia to test the hypothesis that leptin receptor deficiency might explain the enhanced mortality in obese patients. METHODS: We infected wild-type, obese mice globally deficient in the leptin receptor (db/db) and non-obese mice with tissue specific deletion of the leptin receptor in the lung epithelium (SPC-Cre/LepR fl/fl) or macrophages and alveolar type II cells (LysM-Cre/Lepr fl/fl) with influenza A virus (A/WSN/33 [H1N1]) (500 and 1500 pfu/mouse) and measured mortality, viral clearance and several markers of lung injury severity. RESULTS: The clearance of influenza A virus from the lungs of mice was impaired in obese mice globally deficient in the leptin receptor (db/db) compared to normal weight wild-type mice. In contrast, non-obese, SP-C-Cre+/+/LepR fl/fl and LysM-Cre+/+/LepR fl/fl had improved viral clearance after influenza A infection. In obese mice, mortality was increased compared with wild-type mice, while the SP-C-Cre+/+/LepR fl/fl and LysM-Cre+/+/LepR fl/fl mice exhibited improved survival. CONCLUSIONS: Global loss of the leptin receptor results in reduced viral clearance and worse outcomes following influenza A infection. These findings are not the result of the loss of leptin signaling in lung epithelial cells or macrophages. Our results suggest that factors associated with obesity or with leptin signaling in non-myeloid populations such as natural killer and T cells may be associated with worsened outcomes following influenza A infection.

ß2-Adrenergic agonists augment air pollution-induced IL-6 release and thrombosis.

Acute exposure to particulate matter (PM) air pollution causes thrombotic cardiovascular events, leading to increased mortality rates; however, the link between PM and cardiovascular dysfunction is not completely understood. We have previously shown that the release of IL-6 from alveolar macrophages is required for a prothrombotic state and acceleration of thrombosis following exposure to PM. Here, we determined that PM exposure results in the systemic release of catecholamines, which engage the ß2-adrenergic receptor (ß2AR) on murine alveolar macrophages and augment the release of IL-6. In mice, ß2AR signaling promoted the development of a prothrombotic state that was sufficient to accelerate arterial thrombosis. In primary human alveolar macrophages, administration of a ß2AR agonist augmented IL-6 release, while the addition of a beta blocker inhibited PM-induced IL-6 release. Genetic loss or pharmacologic inhibition of the ß2AR on murine alveolar macrophages attenuated PM-induced IL-6 release and prothrombotic state. Furthermore, exogenous ß2AR agonist therapy further augmented these responses in alveolar macrophages through generation of mitochondrial ROS and subsequent increase of adenylyl cyclase activity. Together, these results link the activation of the sympathetic nervous system by ß2AR signaling with metabolism, lung inflammation, and an enhanced susceptibility to thrombotic cardiovascular events.

Particulate matter-induced lung inflammation increases systemic levels of PAI-1 and activates coagulation through distinct mechanisms.

BACKGROUND: Exposure of human populations to ambient particulate matter (PM) air pollution significantly contributes to the mortality attributable to ischemic cardiovascular events. We reported that mice treated with intratracheally instilled PM develop a prothrombotic state that requires the release of IL-6 by alveolar macrophages. We sought to determine whether exposure of mice to PM increases the levels of PAI-1, a major regulator of thrombolysis, via a similar or distinct mechanism. METHODS AND PRINCIPAL FINDINGS: Adult, male C57BL/6 and IL-6 knock out (IL-6(-/-)) mice were exposed to either concentrated ambient PM less than 2.5 µm (CAPs) or filtered air 8 hours daily for 3 days or were exposed to either urban particulate matter or PBS via intratracheal instillation and examined 24 hours later. Exposure to CAPs or urban PM resulted in the IL-6 dependent activation of coagulation in the lung and systemically. PAI-1 mRNA and protein levels were higher in the lung and adipose tissue of mice treated with CAPs or PM compared with filtered air or PBS controls. The increase in PAI-1 was similar in wild-type and IL-6(-/-) mice but was absent in mice treated with etanercept, a TNF-α inhibitor. Treatment with etanercept did not prevent the PM-induced tendency toward thrombus formation. CONCLUSIONS: Mice exposed to inhaled PM exhibited a TNF-α-dependent increase in PAI-1 and an IL-6-dependent activation of coagulation. These results suggest that multiple mechanisms link PM-induced lung inflammation with the development of a prothrombotic state.