The hypoxic environment of cystic fibrosis airways allows the persistence of facultative anaerobic bacteria, which can produce short-chain fatty acids (SCFAs) through fermentation. However, the relevance of SCFAs in cystic fibrosis lung disease is unknown. We show that SCFAs are present in sputum samples from cystic fibrosis patients in millimolar concentrations (mean±SEM 1.99±0.36 mM).SCFAs positively correlated with sputum neutrophil count and higher SCFAs were predictive for impaired nitric oxide production. We studied the effects of the SCFAs acetate, propionate and butyrate on airway inflammatory responses using epithelial cell lines and primary cell cultures. SCFAs in concentrations present in cystic fibrosis airways (0.5-2.5 mM) affected the release of granulocytemacrophage colony-stimulating factor, granulocyte colony-stimulating factor and interleukin (IL)-6. SCFAs also resulted in higher IL-8 release from stimulated cystic fibrosis transmembrane conductance regulator (CFTR) F508del-mutant compared to wild-type CFTR-corrected bronchial epithelial cells. At 25 mM propionate reduced IL-8 release in control but not primary cystic fibrosis epithelial cells. Low (0.5-2.5 mM) SCFA concentrations increased, while high (25-50 mM) concentrations decreased inducible nitric oxide synthase expression. In addition, SCFAs affected the growth of Pseudomonas aeruginosa in a concentration-and pH-dependent manner.Thus, our data suggest that SCFAs contribute to cystic fibrosis-specific alterations of responses to airway infection and inflammation. @ERSpublications Short-chain fatty acids contribute to CF-specific alterations of responses to airway infection and inflammation
Dihydrorhodamine (DHR) 123 is a fluorophore commonly used for measuring reactive oxygen species (ROS), often after exposing cells to ultraviolet (UV) irradiation or oxidative burst inducers such as Phorbol 12‐myristate 13‐acetate (PMA). However, the negative effects of UV irradiation on oxidation of DHR123 itself to green fluorescence rhodamine (R) 123 under different experimental conditions (e.g., different buffers, media, cells, ROS detection techniques) have not been fully appreciated. We determined the effect of UV on DHR123 fluorescence, using a cell‐free system, and A549 epithelial cells, NIH/3T3 fibroblast cells, Jurkat T cells, primary human T cells, HL‐60 neutrophils and primary human neutrophils. We found that UV irradiation rapidly increases green fluorescence of DHR123 in cell‐free solutions. The intensity of green fluorescence increases with increasing amounts of DHR123 and UV exposure. The fluorescence increase was greater in Roswell Park Memorial Institute medium (RPMI) than DMEM media. The presence of DMSO (0–1.25%, v/v) in RPMI further increases the fluorescence signal. Phosphate buffered solution (PBS) and Hanks' Balanced Salt Solution (HBSS) generate considerable background signal with DHR123, and increasing DMSO concentration greatly increases the fluorescence signal in these buffers. However, after UV irradiation the amount of DHR123 that remains unoxidized generates sufficient fluorescence signal to measure the ROS produced by H2O2 and peroxidase in vitro or Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase‐mediated ROS production within HL‐60 neutrophils or primary human neutrophils. We conclude that UV irradiation oxidizes DHR123 to generate Rhodamine 123 (R123) green fluorescence signal, and that the R123 present in the culture supernatant could give erroneous results in plate reader assays. However, flow cytometry and fluorescence microscopy reliably detect ROS in cells such as neutrophils. Overall, avoiding false‐positive results when detecting ROS using DHR123 requires selection of, agonists, the correct buffers, media, cell types, and measurement techniques.
Cigarette smoke (CS) is the main cause of chronic obstructive pulmonary disease. Surfactant protein D (SP-D) is an important anti-inflammatory protein that regulates host immune defense in the lungs. Here, we investigated the role of SP-D in a murine model of CS-induced inflammation. Pulmonary SP-D localization and abundance was compared between smoker and non-smoker individuals. For in vivo studies, wildtype, and SP-D-deficient mice were exposed to CS for either 12 weeks or 3 days. Moreover, the effect of therapeutic administration of recombinant fragment of human SP-D on the acute CS-induced changes was evaluated. Pulmonary SP-D appeared with heterogenous expression in human smokers, while mouse lung SP-D was uniformly upregulated after CS exposure. We found that SP-D-deficient mice were more susceptible to CS-induced macrophage-rich airway inflammation. SP-D deficiency influenced local pro-inflammatory cytokine levels, with increased CCL3 and interleukin-6 but decreased CXCL1. Furthermore, CS exposure caused significant upregulation of pro-inflammatory ceramides and related ceramide synthase gene transcripts in SP-D-deficient mice compared to wildtype littermates. Administration of recombinant fragment of human SP-D (rfhSP-D) alleviated CS-induced macrophage infiltration and prevented induction of ceramide synthase gene expression. Finally, rfhSP-D treatment attenuated CS-induced human epithelial cell apoptosis in vitro. Our results indicate that SP-D deficiency aggravates CS-induced lung inflammation partly through regulation of ceramide synthesis and that local SP-D enrichment rescues CS-induced inflammation.
The challenges of identifying and eliminating racial disparities regarding the exposure, transmission, prevention, and treatment of communicable diseases within the healthcare system have been a mounting concern since the COVID-19 pandemic began. The African, Caribbean, and Black (ACB) populations in Canada represent a fast-expanding and underprivileged community, which have been previously found to have higher susceptibility to communicable diseases and lower sensitivity to intervention measures. Currently, there is insufficient evidence to adequately identify racial patterns in the prevalence and healthcare utilization among the ACB population within the context of the ongoing pandemic. Our proposed study will explore the association between the social determinants of health (SDH) and COVID-19 health outcomes in ACB populations in high-income countries (UK, US, Australia). We will explore the literary evidence through a systematic review (SR) of COVID-19 literature covering the period between December 2019 and October 2020. The objectives include investigating the effect of SDH on the ACB populations’ risk to COVID-19 health outcomes, including COVID-19 infection incidence, severity of disease, hospitalization, mortality and barriers to the treatment and management of COVID-19 for Black people in Canada. In addition, this project aims to investigate the effect of COVID-19 on ACB communities in Ontario by examining the challenges that front-line healthcare workers and administrators have during this pandemic as it pertains to service provisions to ACB communities. A systematic review of original and review studies will be conducted based on the publications on eleven databases (MEDLINE, Web of Science, Cochrane Library, CINAHL, NHS EDD, Global Health, PsychInfo, PubMed, Scopus, Proquest, and Taylor and Francis Online Journals) published between December 2019 to October 2020. Primary outcomes will include the rate of COVID-19 infection. The systematic review will include a meta-analysis of available quantitative data, as well as a narrative synthesis of qualitative studies. This systematic review will be among the first to report racial disparities in COVID-19 infection among the ACB population in Canada. Through synthesizing population data regarding the risk factors on various levels, the findings from this systematic review will provide recommendations for future research and evidence for clinical practitioners and social workers. Overall, a better understanding of the nature and consequences of racial disparities during the pandemic will provide policy directions for effective interventions and resilience-building in the post-pandemic era.
Only a few extracellular soluble proteins are known to modulate apoptosis. We considered that surfactant-associated protein D (SP-D), an innate immune collectin present on many mucosal surfaces, could regulate apoptosis. Although SP-D is known to be important for immune cell homeostasis, whether SP-D affects apoptosis is unknown. In this study we aimed to determine the effects of SP-D on Jurkat T cells and human T cells dying by apoptosis. Here we show that SP-D binds to Jurkat T cells and delays the progression of Fas (CD95)-Fas ligand and TRAIL-TRAIL receptor induced, but not TNF-TNF receptor-mediated apoptosis. SP-D exerts its effects by reducing the activation of initiator caspase-8 and executioner caspase-3. SP-D also delays the surface exposure of phosphatidylserine. The effect of SP-D was ablated by the presence of caspase-8 inhibitor, but not by intrinsic pathway inhibitors. The binding ability of SP-D to dying cells decreases during the early stages of apoptosis, suggesting the release of apoptotic cell surface targets during apoptosis. SP-D also delays FasL-induced death of primary human T cells. SP-D delaying the progression of the extrinsic pathway of apoptosis could have important implications in regulating immune cell homeostasis at mucosal surfaces.
IntroductionIn 2001, 50%–55% of French-speaking minority communities did not have access to health services in French in Canada. Although Canada is officially a bilingual country, reports indicate that many healthcare services offered in French in Anglophone provinces are insufficient or substandard, leading to healthcare discrepancies among Canada’s minority Francophone communities.ObjectivesThe primary aim of this scoping systematic review was to identify existing gaps in HIV-care delivery to Francophone minorities living with HIV in Canada.Study designScoping systematic review.Data sourcesSearch for studies published between 1990 and November 2019 reporting on health and healthcare in Francophone populations in Canada. Nine databases were searched, including Medline, Cumulative Index to Nursing and Allied Health Literature, the Cochrane Library, the National Health Service Economic Development Database, Global Health, PsychInfo, PubMed, Scopus and Web of Science.Study selectionEnglish or French language studies that include data on French-speaking people with HIV in an Anglophone majority Canadian province.ResultsThe literature search resulted in 294 studies. A total of 230 studies were excluded after duplicates were removed. The full texts of 43 potentially relevant papers were retrieved for evaluation and data extraction. Forty-one studies were further excluded based on failure to meet the inclusion criteria leaving two qualitative studies that met our inclusion criteria. These two studies reported on barriers on access to specialised care by Francophone and highlighted difficulties experienced by healthcare professionals in providing quality healthcare to Francophone patients in Ontario and Manitoba.ConclusionThe findings of this scoping systematic review highlight the need for more HIV research on linguistic minority communities and should inform health policymaking and HIV/AIDS community organisations in providing HIV care to Francophone immigrants and Canadians.
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