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From 1967 until 2020, [Community] has had 85 million litres of pulp and paper mill effluent dumped every day into an estuary that borders the community. Despite long-term concerns about cancer in the community, a federal government appointed Joint Environmental Health Monitoring Committee, mandated to oversee the health of the community, has never addressed [Community] concerns. In this study we accessed the 2013 Canadian Cancer Registry microfile data, and using the standard geographical classification code, accessed the cancer data for [Community], and provided comparable data for all Nova Scotia First Nations, as well as the county, provincial, and national population level data. We determined that digestive organ cancers, respiratory organ cancers, male genital organ cancers, and urinary tract cancers are higher in [Community] than at all comparable levels. Female breast and genital organ cancers are lowest in [Community] than at all other comparable levels. We note the limitation of this study as not being able to capture cancer data for off-reserve members at the time of diagnosis and the lapse in availability of up-to-date CCR data. This study demonstrates that cancer data can be compiled for First Nation communities using the standard geographic code, and although not a comprehensive count of all diagnoses for the registered members of [Community], it is the first study to provide data for those who lived in [Community] at the time of diagnosis. Moreover, it highlights the lack of capacity (or will) by Joint Environmental Health Monitoring Committee to uphold their fiduciary duty.

Atherosclerosis is characterised by disturbed blood flow due to abnormal lipid and cell accumulation within arterial layers. Modified cholesterol forms such as oxidised low-density lipoprotein (oxLDL) enter cells altering their phenotype, triggering over exuberant repair and arterial occlusion, myocardial infarction or stroke. We hypothesised that oxLDL enters vascular wall cells and induces IL-1β secretion, potentially via a caspase-1/NLRP3 mechanism. Human coronary artery endothelial cells (HCAEC) and smooth muscle cells (VSMC), isolated from different donors, were cultured and stimulated (primed) with pro-inflammatory cytokines TNFα and IL-1α (10 ng/ml each, for 48 hours), followed by incubation with human oxLDL (10-50ug/ml) for - up to 6 hours. Inhibitors of Caspase-1 (YVAD), NLRP3 (MCC950) and Gasdermin D (Disulfiram) were added 1h before oxLDL. Cell lysates and culture supernatants were collected and analysed for IL-1β using ELISA. Microscopy imaging showed oxLDL entered stimulated cells and formed particles. OxLDL at 20 and 50 ug/ml induced the maximum release of IL-1β from stimulated HCASMCs and HCAECs respectively compared to control. Inhibition of either NLRP3, Caspase-1 or Gasdermin D significantly reduced the release of IL-1β (4-fold, p0.0001; 14-fold, p0.0001, 1.5-fold, p0.0003 respectively) in HCAEC. In contrast, in HCASMCs, only Caspase-1 inhibition reduced release of IL-1β (2.1-fold, P0.0001). HCAECs and HCASMCs elicited release of IL-1β in response to the same stimulus via different mechanisms. In HCAECs, released IL-1β potentially exits via a GSDMD-induced membrane pore. These data suggest that caspase-1 or gasdermin D inhibition are likely to be effective vessel wall cell specific strategies for the reduction of atherosclerosis.

IntroductionDespite epidemiological associations between community acquired pneumonia (CAP) and myocardial infarction, mechanisms that modify cardiovascular disease during CAP are not well defined. In particular, largely due to a lack of relevant experimental models, the effect of pneumonia on atherosclerotic plaques is unclear. We describe the development of a murine model of the commonest cause of CAP, Streptococcus pneumoniae pneumonia, on a background of established atherosclerosis. We go on to use our model to investigate the effects of pneumococcal pneumonia on atherosclerosis.MethodsC57BL/6J and ApoE-/- mice were fed a high fat diet to promote atherosclerotic plaque formation. Mice were then infected with a range of S. pneumoniae serotypes (1, 4 or 14) with the aim of establishing a model to study atherosclerotic plaque evolution after pneumonia and bacteremia. Laser capture microdissection of plaque macrophages enabled transcriptomic analysis.ResultsIntratracheal instillation of S. pneumoniae in mice fed a cholate containing diet resulted in low survival rates following infection, suggestive of increased susceptibility to severe infection. Optimization steps resulted in a final model of male ApoE-/- mice fed a Western diet then infected by intranasal instillation of serotype 4 (TIGR4) S. pneumoniae followed by antibiotic administration. This protocol resulted in high rates of bacteremia (88.9%) and survival (88.5%). Pneumonia resulted in increased aortic sinus plaque macrophage content 2 weeks post pneumonia but not at 8 weeks, and no difference in plaque burden or other plaque vulnerability markers were found at either time point. Microarray and qPCR analysis of plaque macrophages identified downregulation of two E3 ubiquitin ligases, Huwe1 and Itch, following pneumonia. Treatment with atorvastatin failed to alter plaque macrophage content or other plaque features.DiscussionWithout antibiotics, ApoE-/- mice fed a high fat diet were highly susceptible to mortality following S. pneumoniae infection. The major infection associated change in plaque morphology was an early increase in plaque macrophages. Our results also hint at a role for the ubiquitin proteasome system in the response to pneumococcal infection in the plaque microenvironment.

For over fifty years, Pictou Landing First Nation (PLFN), a small Mi'kmaw community on the northern shore of mainland Nova Scotia, Canada, has been told by a Joint Environmental Health Monitoring Committee (JEHMC) mandated to oversee the health of the community that their health has not been impacted by exposure to 85 million litres of pulp mill effluent dumped every day into what was once a culturally significant body of water bordering their community. Yet, based on lived experience, the community knows otherwise, and despite countless dollars spent on government and industry-sponsored research, their concerns have not gone away. Using biopolitical theory, we explore why JEHMC never fully implemented its mandate. We will use a Mi'kmaw environmental 'theoretical' framework to demonstrate that indicators of a relational epistemology and ontology that have been consistently and persistently overlooked in Indigenous environmental health research demands that Indigenous connections to the air, land and water must be taken into consideration to get a full understanding of environmental health impacts. Guided by the principle of Etuaptmumk (Two-Eyed Seeing), which brings together the strengths of both western and Indigenous knowledge, and employing a community-based participatory research approach, we use data that could have been accessed by the JEHMC that might have signaled that human health studies were warranted. Further, we developed an environmental health survey that more appropriately assesses the impacts on the community. Finally, we will discuss how an Indigenous-developed framework can adequately assess the impacts of land displacement and environmental dispossession on the health of Indigenous communities and illustrate how our framework can serve as a guide to others when exploring Indigenous environmental health more broadly.

Highlights • Brain endothelial cells mediate detrimental actions of IL-1 in cerebral ischemia. • Neuronal cholinergic IL-1R1 also mediate detrimental actions of IL-1 in brain injury. • Brain endothelial IL-1 actions reduce cortical perfusion after cerebral ischemia. • Ubiquitous IL-1R1 deletion does not affect injury, suggesting compensatory mechanisms., The cytokine interleukin-1 (IL-1) is a key contributor to neuroinflammation and brain injury, yet mechanisms by which IL-1 triggers neuronal injury remain unknown. Here we induced conditional deletion of IL-1R1 in brain endothelial cells, neurons and blood cells to assess site-specific IL-1 actions in a model of cerebral ischaemia in mice. Tamoxifen treatment of IL-1R1 floxed (fl/fl) mice crossed with mice expressing tamoxifen-inducible Cre-recombinase under the Slco1c1 promoter resulted in brain endothelium-specific deletion of IL-1R1 and a significant decrease in infarct size (29%), blood-brain barrier (BBB) breakdown (53%) and neurological deficit (40%) compared to vehicle-treated or control (IL-1R1fl/fl) mice. Absence of brain endothelial IL-1 signalling improved cerebral blood flow, followed by reduced neutrophil infiltration and vascular activation 24 h after brain injury. Conditional IL-1R1 deletion in neurons using tamoxifen inducible nestin-Cre mice resulted in reduced neuronal injury (25%) and altered microglia-neuron interactions, without affecting cerebral perfusion or vascular activation. Deletion of IL-1R1 specifically in cholinergic neurons reduced infarct size, brain oedema and improved functional outcome. Ubiquitous deletion of IL-1R1 had no effect on brain injury, suggesting beneficial compensatory mechanisms on other cells against the detrimental effects of IL-1 on endothelial cells and neurons. We also show that IL-1R1 signalling deletion in platelets or myeloid cells does not contribute to brain injury after experimental stroke. Thus, brain endothelial and neuronal (cholinergic) IL-1R1 mediate detrimental actions of IL-1 in the brain in ischaemic stroke. Cell-specific targeting of IL-1R1 in the brain could therefore have therapeutic benefits in stroke and other cerebrovascular diseases.

Vascular tissue-engineered in vitro models are increasing in popularity but often lack the complexity required to examine the underlying inflammatory response of diseases such as atherosclerosis. Most models also focus on the interactions between the endothelial cells and smooth muscle cells. However, fibroblasts found in the tunica adventitia are known to express pro-inflammatory cytokines such as Tumour Necrosis Factor Alpha (TNF-α) and Interleukin-1 (IL-1) which significantly affect the expression of adhesion proteins, such as Intercellular Adhesion Molecule 1 (ICAM-1) directing leukocyte infiltration and the recurrence of acute coronary syndromes. Consequently, the aim was to develop an in vitro co-culture model to examine whether the TNF-α induced expression of ICAM-1 could be up-regulated in endothelial and smooth muscle cells when co-cultured with fibroblasts. Human Umbilical Vein Endothelial Cells (HUVEC), Human Umbilical Artery Smooth Muscle Cells (HUASMC) and Normal Human Dermal Fibroblasts (NHDF) were cultured either alone or in co-culture and were stimulated with 25 U/mL TNF-α for up to 12 hours to assess the surface expression of ICAM-1. NHDF increased HUVEC TNF-α-induced ICAM-1 expression after 12 hours, in a contact-dependent manner, when compared to monoculture (p

Background Clinical studies consistently find an increase in the risk of acute coronary syndrome (ACS) in the weeks following pneumonia, although the mechanisms underlying this finding are unknown. ACS most commonly occurs as a result of thrombosis at the site of ruptured atherosclerotic plaques. We hypothesized that the systemic inflammatory response to pneumococcal pneumonia leads to acute localized inflammatory changes within established atherosclerotic plaques, favoring plaque instability and rupture, thereby resulting in ACS. Methods Male ApoE-/- mice, a well-established model of atherosclerosis, were fed an atherogenic diet for 7–8 weeks before intranasal infection with Streptococcus pneumoniae or mock infection. Mice were sacrificed 2 or 8 weeks post-infection. Formalin-fixed, paraffin-embedded aortic sinus plaque sections were analyzed to assess markers of plaque vulnerability to rupture. To characterise post-pneumonic plaque macrophage phenotype, aortic sinus plaque cryosections 2 weeks post pneumonia/mock infection were immunostained for MAC-3 to identify macrophage-rich areas. These plaque regions were collected using laser capture microdissection and RNA extracted for microarray analysis. Results S. pneumoniae infection was associated with increased aortic sinus atherosclerotic plaque macrophage content (18.1 vs. 8.0%; P < 0.05) at 2 weeks post infection, but no significant difference in aortic sinus plaque burden, plaque smooth muscle or collagen content. There was no significant difference in any of these plaque vulnerability markers at 8 weeks post infection. Microarray analysis of laser capture micro-dissected plaque macrophages identified downregulation of the expression of three genes coding for specific E3 ubiquitin ligases following pneumonia. Pathway analysis identified a significant perturbation in the ubiquitin proteasome system pathway as a result. Conclusion In this murine model, pneumococcal pneumonia resulted in increased atherosclerotic plaque macrophage content, a marker of plaque instability, at 2 weeks post infection. Pneumonia may therefore lead to an increased propensity for atherosclerotic plaques to rupture soon after pneumonia, due to infiltration of macrophages into the plaque. Disclosures All authors: No reported disclosures.

Background : Designing an effective survey for gathering primary health data using a community-based participatory research (CBPR) approach in Indigenous communities in Canada has its challenges. Yet, the Pictou Landing First Nation (PFLN) Native Women’s Group (NWG) and academic research partners achieved a 59% response rate. Objectives : To share lessons learned with both campus and community-based research teams engaged in CBPR involving Indigenous communities on the process of team development, and particularly survey development and execution, as well as the factors that led to a reliable and valid household level environmental health survey that achieved a 59% response rate. Methods : Multiple debriefings conducted over the course of the 12-month data collection period allowed us to modify our protocol to fit with community oscillations. Results/Lessons Learned : Unique aspects of CBPR allowed for the development of a culturally appropriate survey protocol and culturally relevant variables that reflected the concerns of the NWG, and presenting preliminary data to the community also encouraged community buy-in to participate. Conclusions : Sharing lessons learned in this project are intended to have positive implications for future CBPR projects wanting to collect primary health survey data involving Indigenous communities.

Bacterial infection contributes to diverse noninfectious diseases and worsens outcome after stroke. Streptococcus pneumoniae, the most common infection in patients at risk of stroke, is a major cause of prolonged hospitalization and death of stroke patients, but how infection impacts clinical outcome is not known.We induced sustained pulmonary infection by a human S. pneumoniae isolate in naive and comorbid rodents to investigate the effect of infection on vascular and inflammatory responses prior to and after cerebral ischemia.S. pneumoniae infection triggered atherogenesis, led to systemic induction of interleukin (IL) 1, and profoundly exacerbated (50-90%) ischemic brain injury in rats and mice, a response that was more severe in combination with old age and atherosclerosis. Systemic blockade of IL-1 with IL-1 receptor antagonist (IL-1Ra) fully reversed infection-induced exacerbation of brain injury and functional impairment caused by cerebral ischemia. We show that infection-induced systemic inflammation mediates its effects via increasing platelet activation and microvascular coagulation in the brain after cerebral ischemia, as confirmed by reduced brain injury in response to blockade of platelet glycoprotein (GP) Ibα. IL-1 and platelet-mediated signals converge on microglia, as both IL-1Ra and GPIbα blockade reversed the production of IL-1α by microglia in response to cerebral ischemia in infected animals.S. pneumoniae infection augments atherosclerosis and exacerbates ischemic brain injury via IL-1 and platelet-mediated systemic inflammation. These mechanisms may contribute to diverse cardio- and cerebrovascular pathologies in humans.