Macrophages reprogram their lipid metabolism in response to activation signals. However, a systems-level understanding of how different pro-inflammatory stimuli reshape the macrophage lipidome is ...lacking. Here, we use complementary “shotgun” and isotope tracer mass spectrometry approaches to define the changes in lipid biosynthesis, import, and composition of macrophages induced by various Toll-like receptors (TLRs) and inflammatory cytokines. “Shotgun” lipidomics data revealed that different TLRs and cytokines induce macrophages to acquire distinct lipidomes, indicating their specificity in reshaping lipid composition. Mechanistic studies showed that differential reprogramming of lipid composition is mediated by the opposing effects of MyD88- and TRIF-interferon-signaling pathways. Finally, we applied these insights to show that perturbing reprogramming of lipid composition can enhance inflammation and promote host defense to bacterial challenge. These studies provide a framework for understanding how inflammatory stimuli reprogram lipid composition of macrophages while providing a knowledge platform to exploit differential lipidomics to influence immunity.
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•A quantitative profiling of the mouse macrophage lipidome activated by immune stimuli•Macrophages alter lipid composition in a TLR-specific manner•MyD88-dependent TLRs alter lipid composition by increasing de novo MUFA synthesis•Inhibiting MUFA synthesis increases inflammation generated by MyD88-dependent TLRs
Using a combination of shotgun lipidomics and stable-isotope tracing, Hsieh et al. show that distinct pro-inflammatory stimuli reshape the macrophage lipid composition in a signal-specific manner and that targeting this change can increase immunity. Thus, the study provides an in-depth resource and framework for understanding this lipidomic response while suggesting approaches for future therapy.
Immune cells, such as macrophages, reprogram their lipid metabolism in response to the activation of pattern recognition receptors (e.g., TLRs, NLRs) and cytokine receptors (e.g., interferons, ...interleukins). Profiling these changes can be achieved with shotgun mass spectrometry. This protocol provides step-by-step instructions on the generation and stimulation of bone marrow-derived macrophages (BMDMs), sample collection, and lipid extraction for profiling the macrophage lipidome.
For complete details on the use and execution of this protocol, please refer to Hsieh et al. (2020).
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•Protocol for profiling mouse macrophage lipidome with direct infusion mass spectrometry•Provides quantitative measurements of immune cell lipid composition•Includes cell culture, cell imaging, sample preparation, and data output analysis•Can be adapted for different lipidomics-mass spectrometry platforms
Immune cells, such as macrophages, reprogram their lipid metabolism in response to the activation of pattern recognition receptors (e.g., TLRs, NLRs) and cytokine receptors (e.g., interferons, interleukins). Profiling these changes can be achieved with shotgun mass spectrometry. This protocol provides step-by-step instructions on the generation and stimulation of bone marrow-derived macrophages (BMDMs), sample collection, and lipid extraction for profiling the macrophage lipidome.
Electronic cigarette use is on the rise despite a number of reports linking electronic cigarettes with adverse health outcomes. Recent studies have suggested that alterations in lipid signaling may ...be one mechanism by which electronic cigarettes contribute to lung pulmonary function. Vitamin E acetate, for example, is synthetic form of Vitamin E transported via lipids, found to be associated with electronic cigarette associated lung injury. Lipids are absolutely critical for normal lung physiology and perturbations in a number of lipid pathways have been associated with respiratory illness. Is it conceivable that electronic cigarette use even in seemingly healthy cohorts are associated with alterations in lipid pathways?
To investigate quantitative alterations in the plasma lipidome associated with electronic cigarette use in healthy we obtained plasma samples from 119 male and female participants with who were either: (1) chronic tobacco cigarette (TC) smokers (> 12 months of self-reported TC use), (2) chronic Electronic cigarette (EC) users (> 12 months of self-reported EC use), or (3) non-users. We measured quantitative lipid species across different lipid sub-classes from plasma samples using the Sciex Lipidyzer.
We found that male and female tobacco and electronic cigarette users had distinct lipidome signatures across a number of lipid species although the vast majority of lipids were unchanged when compared to non-users. Intriguingly, we found that female but not male electronic cigarette users had lower levels of plasmalogens, critical glycerophospholipids secreted by alveoli and required for normal surfactant function.
In summary, our study does not reveal striking changes associated with electronic cigarette use but we observed sex-specific changes in lipids known to be critical for lung function.
Nonalcoholic fatty liver disease (NAFLD) is a leading cause of chronic liver disease and cirrhosis. NAFLD is mediated by changes in lipid metabolism and known risk factors include obesity, metabolic ...syndrome, and diabetes. The aim of this study was to better understand differences in the lipid composition of individuals with NAFLD compared to controls, by performing direct infusion lipidomics on serum biospecimens from a cohort study of adults in Mexico.
A nested case-control study was conducted with a sample of 98 NAFLD cases and 100 healthy controls who are participating in an on-going, longitudinal study in Mexico. NAFLD cases were clinically confirmed using elevated liver enzyme tests and liver ultrasound or liver ultrasound elastography, after excluding alcohol abuse, and 100 controls were identified as having at least two consecutive normal alanine aminotransferase (ALT) and aspartate aminotransferase (AST) (< 40 U/L) results in a 6-month period, and a normal liver ultrasound elastography result in January 2018. Samples were analyzed on the Sciex Lipidyzer Platform and quantified with normalization to serum volume. As many as 1100 lipid species can be identified using the Lipidyzer targeted multiple-reaction monitoring list. The association between serum lipids and NAFLD was investigated using analysis of covariance, random forest analysis, and by generating receiver operator characteristic (ROC) curves.
NAFLD cases had differences in total amounts of serum cholesterol esters, lysophosphatidylcholines, sphingomyelins, and triacylglycerols (TAGs), however, other lipid subclasses were similar to controls. Analysis of individual TAG species revealed increased incorporation of saturated fatty acyl tails in serum of NAFLD cases. After adjusting for age, sex, body mass index, and PNPLA3 genotype, a combined panel of ten lipids predicted case or control status better than an area under the ROC curve of 0.83.
These preliminary results indicate that the serum lipidome differs in patients with NAFLD, compared to healthy controls, and suggest that assessing the desaturation state of TAGs or a specific lipid panel may be useful clinical tools for the diagnosis of NAFLD.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Glaucoma is the second most common cause of blindness in the world. It is a multifactorial disease with several risk factors, of which intraocular pressure (IOP) is a primary contributing factor. IOP ...measurements are used for glaucoma diagnosis and patient monitoring. IOP has wide diurnal fluctuation and is dependent on body posture, so the occasional measurements done by the eye care expert in the clinic can be misleading. Here we show that microfluidic principles can be used to develop an implantable sensor that has a limit of detection of 1 mm Hg, high sensitivity and excellent reproducibility. This device has a simple optical interface that enables IOP to be read with a smartphone camera. This sensor, with its ease of fabrication and simple design, as well as its allowance for IOP home monitoring, offers a promising approach for better care of patients with glaucoma.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Abstract
Necrotizing fasciitis (NF) is an aggressive skin and soft tissue infection characterized by massive tissue destruction. Current treatment strategies are limited to aggressive and repeated ...surgical debridement, coupled with systemic antibiotics. Thus, new therapeutic approaches that preserve tissue integrity would be beneficial. Cholesterol-dependent cytolysins (CDCs) are toxins produced by select gram-positive microbes implicated in NF. CDCs recognize cholesterol in membranes of cells for pore formation and subsequent cellular and tissue damage. Recently, we have shown that interferon signaling is able to protect cells from CDCs via reprogramming cholesterol homeostasis. This observation led to us to ask if pharmacological targeting of cholesterol metabolism provide similar protection. Liver X Receptor (LXRs) are key transcriptional regulators of cholesterol metabolism. We find that pharmacological LXR activation protects mice from Streptolysin O (SLO)-induced tissue damage. LXR stimulation in various cell types, such as macrophages, neutrophils, and endothelial cells induced resistance to CDCs by abrogating the ability of CDCs to directly bind to cholesterol in the plasma membrane. Ongoing mechanistic studies are investigating which metabolic pathways induced by LXRs (e.g., cholesterol efflux, biosynthesis, import, or trafficking) are responsible for the protective effects. Translationally, LXR stimulation may serve as an adjunctive therapeutics for necrotizing fasciitis and other CDC-mediated pathologies (e.g., pneumonia, pharyngitis, and septic cardiomyopathy).
P01 HL146358
Malignant tumors exhibit heterogeneous metabolic reprogramming, hindering the identification of translatable vulnerabilities for metabolism-targeted therapy. How molecular alterations in tumors ...promote metabolic diversity and distinct targetable dependencies remains poorly defined. Here we create a resource consisting of lipidomic, transcriptomic, and genomic data from 156 molecularly diverse glioblastoma (GBM) tumors and derivative models. Through integrated analysis of the GBM lipidome with molecular datasets, we identify CDKN2A deletion remodels the GBM lipidome, notably redistributing oxidizable polyunsaturated fatty acids into distinct lipid compartments. Consequently, CDKN2A-deleted GBMs display higher lipid peroxidation, selectively priming tumors for ferroptosis. Together, this study presents a molecular and lipidomic resource of clinical and preclinical GBM specimens, which we leverage to detect a therapeutically exploitable link between a recurring molecular lesion and altered lipid metabolism in GBM.
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•Molecular and lipidomic resource of over 150 GBM tumors and derivative models•Unbiased multi-omic analysis identifies CDKN2A as a regulator of lipid metabolism•CDKN2A deletion reduces oxidizable PUFA sequestration into lipid droplets•GBMs with CDKN2A deletion are susceptible to lipid peroxidation and ferroptosis
Minami et al. integrate lipidomic, transcriptomic, and genomic profiling data to identify altered lipid metabolism in CDKN2A-deleted glioblastoma (GBM). CDKN2A deletion remodels the distribution of polyunsaturated fatty acids into different lipid compartments, sensitizing GBMs with CDKN2A loss to lipid peroxidation and ferroptosis in vitro and in vivo.
Abstract
Glioblastoma (GBM) is the most common malignant brain cancer in adults, enriched in a small subpopulation of glioma stem cells (GSC), which can drive tumor recurrence and therapeutic ...resistance. Considerable evidence suggests that the endogenous levels of unsaturated fatty acids (FA) are crucial regulators of GSCs survival and self-renewal. Stearoyl-CoA desaturase-1 (SCD-1) is the most abundant desaturase in humans. We have previously shown that SCD1 activity is required for GSCs self-renewal and brain tumor initiation. However, SCD1 orthologous isoform, SCD5, has been poorly characterized and its potential role in GBM has not been previously reported. We have observed that SCD5 is highly enriched in GSC both at the mRNA and protein levels. Genetic downregulation of SCD5 in GSCs led to a remarkable decrease in stem cell markers, impaired cell viability and the ability to form neurospheres. Further, the downregulation of SCD5 in GSCs orthotopically implanted in mice resulted in delayed tumor growth and extended overall survival. Shotgun lipidomics in GSCs after either SCD1 or SCD5 knock-down revealed a largely distinctive lipidome profile, highlighting the divergent role of these two isoforms in GBM lipid metabolism. Surprisingly, lipidomics analysis showed that both SCD1 and SCD5 are required to synthesize a variety of lipid species involved in receptor tyrosine kinase (RTKs) and GPCRs signal transduction, directly linking FA synthesis with the oncogenic signaling. We confirmed these results by immunoblot analysis. Using specific tagging and immunofluorescence analysis, we observed that, despite a spatial overlap in SCD1 and SCD5 expression, SCD5 is uniquely present in some subcellular locations. This suggests that different functions of these isoforms could be related to different subcellular localization. Altogether, our results underscore a novel function of SCD isoforms in GSCs metabolism and highlight SCD5 as a potential therapeutic target for GBM.
Modern biomarker and translational research as well as personalized health care studies rely heavily on powerful omics’ technologies, including metabolomics and lipidomics. However, to translate ...metabolomics and lipidomics discoveries into a high-throughput clinical setting, standardization is of utmost importance. Here, we compared and benchmarked a quantitative lipidomics platform. The employed Lipidyzer platform is based on lipid class separation by means of differential mobility spectrometry with subsequent multiple reaction monitoring. Quantitation is achieved by the use of 54 deuterated internal standards and an automated informatics approach. We investigated the platform performance across nine laboratories using NIST SRM 1950–Metabolites in Frozen Human Plasma, and three NIST Candidate Reference Materials 8231–Frozen Human Plasma Suite for Metabolomics (high triglyceride, diabetic, and African-American plasma). In addition, we comparatively analyzed 59 plasma samples from individuals with familial hypercholesterolemia from a clinical cohort study. We provide evidence that the more practical methyl-tert-butyl ether extraction outperforms the classic Bligh and Dyer approach and compare our results with two previously published ring trials. In summary, we present standardized lipidomics protocols, allowing for the highly reproducible analysis of several hundred human plasma lipids, and present detailed molecular information for potentially disease relevant and ethnicity-related materials.
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