Publications by authors named "Amina S Woods"

104 Publications

Streamlined Analysis of Cardiolipins in Prokaryotic and Eukaryotic Samples Using a Norharmane Matrix by MALDI-MSI.

J Am Soc Mass Spectrom 2020 Dec 5;31(12):2495-2502. Epub 2020 Oct 5.

Department of Microbial Pathogenesis, School of Dentistry, University of Maryland, Baltimore 21201, Maryland, United States.

Cardiolipins (CLs) are an important, regulated lipid class both in prokaryotic and eukaryotic cells, yet they remain largely unexplored by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) in tissues. To date, no in-depth optimization studies of label-free visualization of CLs in complex biological samples have been reported. Here we report a streamlined modification to our previously reported MALDI-MSI method for detection of endogenous CLs in prokaryotic and eukaryotic cells based on preparation with norharmane (NRM) matrix. Notably, the use of NRM matrix permitted sensitive detection (4.7 pg/mm) of spotted CL synthetic standards. By contrast, four other MALDI matrices commonly used for lipid analysis failed to generate CL ions. Using this NRM-based method, endogenous CLs were detected from two types of complex biological samples: dried bacterial arrays and mouse tissue sections. In both cases, using NRM resulted in a better signal/noise for CL ions than the other matrices. Furthermore, inclusion of a washing step improved CL detection from tissue and this combined tissue preparation method (washing and NRM matrix) was used to profile normal mouse lung. Mouse lung yielded 26 unique CLs that were mapped and identified. Consistent with previous findings, CLs containing polyunsaturated fatty acids (PUFAs) were found in abundance in the airway and vascular features of the lung. This work represents a comprehensive investigation of detection conditions for CL using MALDI-MSI in complex biological samples that resulted in a streamlined method that enables future studies of the biological role(s) of CL in tissue.
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http://dx.doi.org/10.1021/jasms.0c00201DOI Listing
December 2020

On-Tissue Derivatization of Lipopolysaccharide for Detection of Lipid A Using MALDI-MSI.

Anal Chem 2020 10 28;92(20):13667-13671. Epub 2020 Sep 28.

Department of Microbial Pathogenesis, School of Dentistry, University of Maryland, Baltimore, Maryland 21201, United States.

We developed a method to directly detect and map the Gram-negative bacterial virulence factor lipid A derived from lipopolysaccharide (LPS) by coupling acid hydrolysis with matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). As the structure of lipid A (endotoxin) determines the innate immune outcome during infection, the ability to map its location within an infected organ or animal is needed to understand localized inflammatory responses that results during host-pathogen interactions. We previously demonstrated detection of free lipid A from infected tissue; however detection of lipid A derived from intact (smooth) LPS from host-pathogen MSI studies, proved elusive. Here, we detected LPS-derived lipid A from the Gram-negative pathogens, (, / 1797) and (, / 1446) using on-tissue acid hydrolysis to cleave the glycosidic linkage between the polysaccharide (core and O-antigen) and lipid A moieties of LPS. Using accurate mass methods, the ion corresponding to the major and lipid A species (/ 1797 and 1446, respectively) were unambiguously discriminated from complex tissue substrates. Further, we evaluated potential delocalization and signal loss of other tissue lipids and found no evidence for either, making this LPS-to-Lipid A-MSI (LLA-MSI) method, compatible with simultaneous host-pathogen lipid imaging following acid hydrolysis. This spatially sensitive technique is the first step in mapping host-influenced de novo lipid A modifications, such as those associated with antimicrobial resistance phenotypes, during Gram-negative bacterial infection and will advance our understanding of the host-pathogen interface.
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http://dx.doi.org/10.1021/acs.analchem.0c02566DOI Listing
October 2020

The Role of Peripheral Opioid Receptors in Triggering Heroin-induced Brain Hypoxia.

Sci Rep 2020 01 21;10(1):833. Epub 2020 Jan 21.

Behavioral Neuroscience Branch, National Institute on Drug Abuse - Intramural Research Program, National Institutes of Health, DHHS, 333 Cassell Drive, Baltimore, MD, 21224, USA.

While it is known that opioid receptors (ORs) are densely expressed in both the brain and periphery, it is widely accepted that hypoxic effects of opioids result solely from their direct action in the CNS. To examine the role of peripheral ORs in triggering brain hypoxia, we used oxygen sensors in freely moving rats to examine how naloxone-HCl and naloxone-methiodide, the latter which is commonly believed to be peripherally restricted, affect brain oxygen responses induced by intravenous heroin at low, human-relevant doses. Similar to naloxone-HCl, naloxone-methiodide at a relatively low dose (2 mg/kg) fully blocked heroin-induced decreases in brain oxygen levels. As measured by mass spectrometry, naloxone-methiodide was found to be ~40-fold less permeable than naloxone-HCl across the blood-brain barrier, thus acting as a selective blocker of peripheral ORs. Despite this selectivity, a low but detectable amount of naloxone was found in brain tissue after naloxone-methiodide administration, potentially influencing our results. Therefore, we examined the effects of naloxone-methiodide at a very low dose (0.2 mg/kg; at which naloxone was undetectable in brain tissue) and found that this drug still powerfully attenuates heroin-induced brain oxygen responses. These data demonstrate the role of peripheral ORs in triggering heroin-induced respiratory depression and subsequent brain hypoxia.
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http://dx.doi.org/10.1038/s41598-020-57768-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6972941PMC
January 2020

Cocaine-induced endocannabinoid signaling mediated by sigma-1 receptors and extracellular vesicle secretion.

Elife 2019 10 9;8. Epub 2019 Oct 9.

Electrophysiology Research Section, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Baltimore, United States.

Cocaine is an addictive drug that acts in brain reward areas. Recent evidence suggests that cocaine stimulates synthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG) in midbrain, increasing dopamine neuron activity via disinhibition. Although a mechanism for cocaine-stimulated 2-AG synthesis is known, our understanding of 2-AG release is limited. In NG108 cells and mouse midbrain tissue, we find that 2-AG is localized in non-synaptic extracellular vesicles (EVs) that are secreted in the presence of cocaine via interaction with the chaperone protein sigma-1 receptor (Sig-1R). The release of EVs occurs when cocaine causes dissociation of the Sig-1R from ADP-ribosylation factor (ARF6), a G-protein regulating EV trafficking, leading to activation of myosin light chain kinase (MLCK). Blockade of Sig-1R function, or inhibition of ARF6 or MLCK also prevented cocaine-induced EV release and cocaine-stimulated 2-AG-modulation of inhibitory synapses in DA neurons. Our results implicate the Sig-1R-ARF6 complex in control of EV release and demonstrate that cocaine-mediated 2-AG release can occur via EVs.
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http://dx.doi.org/10.7554/eLife.47209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6850780PMC
October 2019

A New Integrative Theory of Brain-Body-Ecosystem Medicine: From the Hippocratic Holistic View of Medicine to Our Modern Society.

Int J Environ Res Public Health 2019 08 28;16(17). Epub 2019 Aug 28.

Department of Biomedical Sciences, University of Modena and Reggio Emilia, 41124 Modena, Italy.

Humans are increasingly aware that their fate will depend on the wisdom they apply in interacting with the ecosystem. Its health is defined as the condition in which the ecosystem can deliver and continuously renew its fundamental services. A healthy ecosystem allows optimal interactions between humans and the other biotic/abiotic components, and only in a healthy ecosystem can humans survive and efficiently reproduce. Thus, both the human and ecosystem health should be considered together in view of their interdependence. The present article suggests that this relationship could be considered starting from the Hippocrates (460 BC-370 BC) work "On Airs, Waters, and Places" to derive useful medical and philosophical implications for medicine which is indeed a topic that involves scientific as well as philosophical concepts that implicate a background broader than the human body. The brain-body-ecosystem medicine is proposed as a new more complete approach to safeguarding human health. Epidemiological data demonstrate that exploitation of the environment resulting in ecosystem damage affects human health and in several instances these diseases can be detected by modifications in the heart-brain interactions that can be diagnosed through the analysis of changes in heart rate variability.
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http://dx.doi.org/10.3390/ijerph16173136DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6747255PMC
August 2019

Histidine, the less interactive cousin of arginine.

Eur J Mass Spectrom (Chichester) 2019 Apr;25(2):212-218

Structural Biology Unit, NIDA IRP, NIH, Baltimore, MD, USA.

Electrostatic interactions are one of the main factors influencing biomolecular conformation. The formation of noncovalent complexes by electrostatic interactions is governed by certain amino acid residues and post-translational modifications. It has been demonstrated that adjacent arginine forms noncovalent complex with phosphate; however, histidine noncovalent complexes have rarely been investigated. In the present work, we compare the interaction between basic epitopes (NLRRITRVN, SHHGLHSTPD) and diverse acidic and aromatic-rich peptides using both MALDI and ESI Mass spectrometry. We show that adjacent histidines can also form stable noncovalent bonds and that those bonds are probably formed by a salt bridge between the phosphate or the acid residues and the histidines. However, noncovalent complexes with the arginine epitopes form more readily and are stronger than those with histidine-containing epitopes.
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http://dx.doi.org/10.1177/1469066718791793DOI Listing
April 2019

An In Vitro Study of Aromatic Stacking of Drug Molecules.

J Am Soc Mass Spectrom 2019 Jul 4;30(7):1199-1203. Epub 2019 Apr 4.

Structural Biology Unit, Integrative Neuroscience Branch, NIDA IRP, NIH, Baltimore, MD, USA.

In this paper, drug-drug chemical interactions between two different aromatic compounds were studied by mass spectrometry. Specifically, we examined non-covalent complexes (NCX) between paclitaxel, a chemotherapeutic compound, and medications widely used in palliative care for depression, psychosis, and anxiety. It is unknown whether psychotropic medications directly interact with paclitaxel. Here, we use a simple and rapid electrospray ionization mass spectrometry in vitro assay, which has been predictive in the case of neuropeptides, to measure the relative strength of non-covalent interactions. This chemical interaction is most likely due to the overlap of aromatic rings of π-orbitals between paclitaxel and five commonly used medications: diazepam, clonozepam, sertraline, fluoxetine, and haloperidol. Molecular modeling illustrates that differences in the stability of the NCXs are likely due to the distance between the aromatic rings present in both the paclitaxel and antidepressant medications. Graphical Abstract.
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http://dx.doi.org/10.1007/s13361-019-02166-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7520095PMC
July 2019

Differential composition of DHA and very-long-chain PUFAs in rod and cone photoreceptors.

J Lipid Res 2018 09 9;59(9):1586-1596. Epub 2018 Jul 9.

Departments of Ophthalmology University of Oklahoma Health Sciences Center, Oklahoma City, OK.

Long-chain PUFAs (LC-PUFAs; C20-C22; e.g., DHA and arachidonic acid) are highly enriched in vertebrate retina, where they are elongated to very-long-chain PUFAs (VLC-PUFAs; C 28) by the elongation of very-long-chain fatty acids-4 (ELOVL4) enzyme. These fatty acids play essential roles in modulating neuronal function and health. The relevance of different lipid requirements in rods and cones to disease processes, such as age-related macular degeneration, however, remains unclear. To better understand the role of LC-PUFAs and VLC-PUFAs in the retina, we investigated the lipid compositions of whole retinas or photoreceptor outer segment (OS) membranes in rodents with rod- or cone-dominant retinas. We analyzed fatty acid methyl esters and the molecular species of glycerophospholipids (phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine) by GC-MS/GC-flame ionization detection and ESI-MS/MS, respectively. We found that whole retinas and OS membranes in rod-dominant animals compared with cone-dominant animals had higher amounts of LC-PUFAs and VLC-PUFAs. Compared with those of rod-dominant animals, retinas and OS membranes from cone-dominant animals also had about 2-fold lower levels of di-DHA (22:6/22:6) molecular species of glycerophospholipids. Because PUFAs are necessary for optimal G protein-coupled receptor signaling in rods, these findings suggest that cones may not have the same lipid requirements as rods.
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http://dx.doi.org/10.1194/jlr.M082495DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6121944PMC
September 2018

A Mouse Model of Schnyder Corneal Dystrophy with the N100S Point Mutation.

Sci Rep 2018 07 5;8(1):10219. Epub 2018 Jul 5.

Department of Ophthalmology, University of Cincinnati, Cincinnati, OH, USA.

Schnyder corneal dystrophy (SCD) is a rare autosomal dominant disease in humans, characterized by abnormal deposition of cholesterol and phospholipids in cornea caused by mutations in the UbiA prenyltransferase domain containing 1 (UBIAD1) gene. In this study, we generated a mouse line carrying Ubiad1 N100S point mutation using the CRISPR/Cas9 technique to investigate the pathogenesis of SCD. In vivo confocal microscopy revealed hyper-reflective dot-like deposits in the anterior cornea in heterozygotes and homozygotes. No significant change was found in corneal epithelial barrier function or wound healing. Electron microscopy revealed abnormal mitochondrial morphology in corneal epithelial, stromal, and endothelial cells. Mitochondrial DNA copy number assay showed 1.27 ± 0.07 fold change in homozygotes versus 0.98 ± 0.05 variation in wild type mice (P < 0.05). Lipidomic analysis indicated abnormal metabolism of glycerophosphoglycerols, a lipid class found in mitochondria. Four (34:1, 34:2, 36:2, and 44:8) of the 11 glycerophosphoglycerols species identified by mass spectrometry showed a significant increase in homozygous corneas compared with heterozygous and wild-type mouse corneas. Unexpectedly, we did not find a difference in the corneal cholesterol level between different genotypes by filipin staining or lipidomic analysis. The Ubiad1 mouse provides a promising animal model of SCD revealing that mitochondrial dysfunction is a prominent component of the disease. The different phenotype in human and mouse may due to difference in cholesterol metabolism between species.
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http://dx.doi.org/10.1038/s41598-018-28545-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6033878PMC
July 2018

Macrophages Shed Excess Cholesterol in Unique Extracellular Structures Containing Cholesterol Microdomains.

Arterioscler Thromb Vasc Biol 2018 07 31;38(7):1504-1518. Epub 2018 May 31.

From the Experimental Atherosclerosis Section, National Heart, Lung, and Blood Institute (X.J., Y.L., J.C., R.F., H.S.K.)

Objective: Cells use various mechanisms to maintain cellular cholesterol homeostasis including efflux of cholesterol from the cellular plasma membrane to cholesterol acceptors such as HDLs (high-density lipoproteins). Little is known about the transfer of cholesterol from cells into the extracellular matrix. Using a unique monoclonal antibody that detects ordered cholesterol arrays (ie, cholesterol micro[or nano]-domains), we previously identified that particles containing these cholesterol domains accumulate in the extracellular matrix during cholesterol enrichment of human monocyte-derived macrophages and are found in atherosclerotic lesions. In this study, we further investigate these deposited particles containing cholesterol microdomains and discover their unexpected morphology.

Approach And Results: Although appearing spherical at the resolution of the conventional fluorescence microscope, super-resolution immunofluorescence and atomic force microscopy of in situ cholesterol microdomains, and immunoelectron microscopy of isolated cholesterol microdomains revealed that the microdomains are not vesicles or 3-dimensional crystals but rather appear as branching irregularly shaped deposits of varying size. These cholesterol microdomain-containing deposits are shed from the plasma membrane into the extracellular matrix.

Conclusions: To date, research on cellular excretion of excess cholesterol has demonstrated cellular cholesterol efflux in the form of membranous vesicles and discoidal HDL particles released into the fluid-phase medium. Shedding of plasma membrane cholesterol microdomains provides an additional mechanism for cells such as macrophages to maintain plasma membrane cholesterol homeostasis. Furthermore, recognition that macrophages shed cholesterol microdomains into the extracellular matrix is important to our understanding of extracellular buildup of cholesterol in atherosclerosis.
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http://dx.doi.org/10.1161/ATVBAHA.118.311269DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6023747PMC
July 2018

AP-MALDI Mass Spectrometry Imaging of Gangliosides Using 2,6-Dihydroxyacetophenone.

J Am Soc Mass Spectrom 2018 07 16;29(7):1463-1472. Epub 2018 Mar 16.

Integrative Neuroscience, NIDA IRP, NIH, 333 Cassell Drive, Room 1119, Baltimore, MD, 21224, USA.

Matrix-assisted laser/desorption ionization (MALDI) mass spectrometry imaging (MSI) is widely used as a unique tool to record the distribution of a large range of biomolecules in tissues. 2,6-Dihydroxyacetophenone (DHA) matrix has been shown to provide efficient ionization of lipids, especially gangliosides. The major drawback for DHA as it applies to MS imaging is that it sublimes under vacuum (low pressure) at the extended time necessary to complete both high spatial and mass resolution MSI studies of whole organs. To overcome the problem of sublimation, we used an atmospheric pressure (AP)-MALDI source to obtain high spatial resolution images of lipids in the brain using a high mass resolution mass spectrometer. Additionally, the advantages of atmospheric pressure and DHA for imaging gangliosides are highlighted. The imaging of [M-H] and [M-HO-H] mass peaks for GD gangliosides showed different distribution, most likely reflecting the different spatial distribution of GD and GD species in the brain. Graphical Abstract ᅟ.
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http://dx.doi.org/10.1007/s13361-018-1928-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7549319PMC
July 2018

Mass Spectrometric Imaging of Ceramide Biomarkers Tracks Therapeutic Response in Traumatic Brain Injury.

ACS Chem Neurosci 2017 10 14;8(10):2266-2274. Epub 2017 Aug 14.

Structural Biology Unit, Integrative Neuroscience Branch, NIH/NIDA-IRP , Baltimore, Maryland 21224, United States.

Traumatic brain injury (TBI) is a serious public health problem and the leading cause of death in children and young adults. It also contributes to a substantial number of cases of permanent disability. As lipids make up over 50% of the brain mass and play a key role in both membrane structure and cell signaling, their profile is of particular interest. In this study, we show that advanced mass spectrometry imaging (MSI) has sufficient technical accuracy and reproducibility to demonstrate the anatomical distribution of 50 μm diameter microdomains that show changes in brain ceramide levels in a rat model of controlled cortical impact (CCI) 3 days post injury with and without treatment. Adult male Sprague-Dawley rats received one strike and were euthanized 3 days post trauma. Brain MS images showed increase in ceramides in CCI animals compared to control as well as significant reduction in ceramides in CCI treated animals, demonstrating therapeutic effect of a peptide agonist. The data also suggests the presence of diffuse changes outside of the injured area. These results shed light on the extent of biochemical and structural changes in the brain after traumatic brain injury and could help to evaluate the efficacy of treatments.
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http://dx.doi.org/10.1021/acschemneuro.7b00189DOI Listing
October 2017

Laser Desorption/Ionization Mass Spectrometric Imaging of Endogenous Lipids from Rat Brain Tissue Implanted with Silver Nanoparticles.

J Am Soc Mass Spectrom 2017 08 21;28(8):1716-1728. Epub 2017 Apr 21.

Structural Biology Unit, NIDA IRP, NIH, Baltimore, MD, USA.

Mass spectrometry imaging (MSI) of tissue implanted with silver nanoparticulate (AgNP) matrix generates reproducible imaging of lipids in rodent models of disease and injury. Gas-phase production and acceleration of size-selected 8 nm AgNP is followed by controlled ion beam rastering and soft landing implantation of 500 eV AgNP into tissue. Focused 337 nm laser desorption produces high quality images for most lipid classes in rat brain tissue (in positive mode: galactoceramides, diacylglycerols, ceramides, phosphatidylcholines, cholesteryl ester, and cholesterol, and in negative ion mode: phosphatidylethanolamides, sulfatides, phosphatidylinositol, and sphingomyelins). Image reproducibility in serial sections of brain tissue is achieved within <10% tolerance by selecting argentated instead of alkali cationized ions. The imaging of brain tissues spotted with pure standards was used to demonstrate that Ag cationized ceramide and diacylglycerol ions are from intact, endogenous species. In contrast, almost all Ag cationized fatty acid ions are a result of fragmentations of numerous lipid types having the fatty acid as a subunit. Almost no argentated intact fatty acid ions come from the pure fatty acid standard on tissue. Graphical Abstract ᅟ.
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http://dx.doi.org/10.1007/s13361-017-1665-4DOI Listing
August 2017

Novel Bivalent Ligands Based on the Sumanirole Pharmacophore Reveal Dopamine D Receptor (DR) Biased Agonism.

J Med Chem 2017 04 16;60(7):2890-2907. Epub 2017 Mar 16.

Molecular Targets and Medications Discovery Branch, National Institute on Drug Abuse-Intramural Research Program, National Institutes of Health , 333 Cassell Drive, Baltimore, Maryland 21224, United States.

The development of bivalent ligands has attracted interest as a way to potentially improve the selectivity and/or affinity for a specific receptor subtype. The ability to bind two distinct receptor binding sites simultaneously can allow the selective activation of specific G-protein dependent or β-arrestin-mediated cascade pathways. Herein, we developed an extended SAR study using sumanirole (1) as the primary pharmacophore. We found that substitutions in the N-1- and/or N-5-positions, physiochemical properties of those substituents, and secondary aromatic pharmacophores can enhance agonist efficacy for the cAMP inhibition mediated by G-proteins, while reducing or suppressing potency and efficacy toward β-arrestin recruitment. Compound 19 was identified as a new lead for its selective D G-protein biased agonism with an EC in the subnanomolar range. Structure-activity correlations were observed between substitutions in positions N-1 and/or N-5 of 1 and the capacity of the new bivalent compounds to selectively activate G-proteins versus β-arrestin recruitment in DR-BRET functional assays.
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http://dx.doi.org/10.1021/acs.jmedchem.6b01875DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7594663PMC
April 2017

The Authors Reply.

Kidney Int 2016 11;90(5):1130-1131

Structural Biology Unit, NIDA IRP, NIH, Baltimore, Maryland, USA. Electronic address:

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http://dx.doi.org/10.1016/j.kint.2016.08.002DOI Listing
November 2016

Ethanol Induced Brain Lipid Changes in Mice Assessed by Mass Spectrometry.

ACS Chem Neurosci 2016 08 20;7(8):1148-56. Epub 2016 Jun 20.

Structural Biology Unit, NIDA IRP, NIH , Baltimore, Maryland 21224, United States.

Alcohol abuse is a chronic disease characterized by the consumption of alcohol at a level that interferes with physical and mental health and causes serious and persistent changes in the brain. Lipid metabolism is of particular interest due to its high concentration in the brain. Lipids are the main component of cell membranes, are involved in cell signaling, signal transduction, and energy storage. In this study, we analyzed lipid composition of chronically ethanol exposed mouse brains. Juvenile (JUV) and adult (ADU) mice were placed on a daily limited-access ethanol intake model for 52 days. After euthanasia, brains were harvested, and total lipids were extracted from brain homogenates. Samples were analyzed using high resolution mass spectrometry and processed by multivariate and univariate statistical analysis. Significant lipid changes were observed in different classes including sphingolipids, fatty acids, lysophosphatidylcholines, and other glycerophospholipids.
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http://dx.doi.org/10.1021/acschemneuro.6b00120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577215PMC
August 2016

Mass spectrometry imaging of rat brain lipid profile changes over time following traumatic brain injury.

J Neurosci Methods 2016 10 10;272:19-32. Epub 2016 Feb 10.

Structural Biology Unit, Integrative Neuroscience Branch, NIH/NIDA-IRP, Baltimore, MD 21224, United States. Electronic address:

Background: Mild traumatic brain injury (TBI) is a common public health issue that may contribute to chronic degenerative disorders. Membrane lipids play a key role in tissue responses to injury, both as cell signals and as components of membrane structure and cell signaling. This study demonstrates the ability of high resolution mass spectrometry imaging (MSI) to assess sequences of responses of lipid species in a rat controlled cortical impact model for concussion.

New Method: A matrix of implanted silver nanoparticles was implanted superficially in brain sections for matrix-assisted laser desorption (MALDI) imaging of 50μm diameter microdomains across unfixed cryostat sections of rat brain. Ion-mobility time-of-flight MS was used to analyze and map changes over time in brain lipid composition in a rats after Controlled Cortical Impact (CCI) TBI.

Results: Brain MS images showed changes in sphingolipids near the CCI site, including increased ceramides and decreased sphingomyelins, accompanied by changes in glycerophospholipids and cholesterol derivatives. The kinetics differed for each lipid class; for example ceramides increased as early as 1 day after the injury whereas other lipids changes occurred between 3 and 7 days post injury.

Comparison With Existing Method(s): Silver nanoparticles MALDI matrix is a sensitive new tool for revealing previously undetectable cellular injury response and remodeling in neural, glial and vascular structure of the brain.

Conclusions: Lipid biochemical and structural changes after TBI could help highlighting molecules that can be used to determine the severity of such injuries as well as to evaluate the efficacy of potential treatments.
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http://dx.doi.org/10.1016/j.jneumeth.2016.02.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577232PMC
October 2016

Lipid imaging within the normal rat kidney using silver nanoparticles by matrix-assisted laser desorption/ionization mass spectrometry.

Kidney Int 2015 Jul 11;88(1):186-92. Epub 2015 Feb 11.

Structural Biology Unit, NIDA IRP, NIH, Baltimore, Maryland, USA.

The well-characterized cellular and structural components of the kidney show distinct regional compositions and distribution of lipids. In order to more fully analyze the renal lipidome we developed a matrix-assisted laser desorption/ionization mass spectrometry approach for imaging that may be used to pinpoint sites of changes from normal in pathological conditions. This was accomplished by implanting sagittal cryostat rat kidney sections with a stable, quantifiable and reproducible uniform layer of silver using a magnetron sputtering source to form silver nanoparticles. Thirty-eight lipid species including seven ceramides, eight diacylglycerols, 22 triacylglycerols, and cholesterol were detected and imaged in positive ion mode. Thirty-six lipid species consisting of seven sphingomyelins, 10 phosphatidylethanolamines, one phosphatidylglycerol, seven phosphatidylinositols, and 11 sulfatides were imaged in negative ion mode for a total of seventy-four high-resolution lipidome maps of the normal kidney. Thus, our approach is a powerful tool not only for studying structural changes in animal models of disease, but also for diagnosing and tracking stages of disease in human kidney tissue biopsies.
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http://dx.doi.org/10.1038/ki.2015.3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4527327PMC
July 2015

Chronic ethanol consumption profoundly alters regional brain ceramide and sphingomyelin content in rodents.

ACS Chem Neurosci 2015 Feb 1;6(2):247-59. Epub 2014 Dec 1.

Structural Biology Unit, NIDA IRP, NIH , Baltimore, Maryland 21224, United States.

Ceramides (CER) are involved in alcohol-induced neuroinflammation. In a mouse model of chronic alcohol exposure, 16 CER and 18 sphingomyelin (SM) concentrations from whole brain lipid extracts were measured using electrospray mass spectrometry. All 18 CER concentrations in alcohol exposed adults increased significantly (range: 25-607%); in juveniles, 6 CER decreased (range: -9 to -37%). In contrast, only three SM decreased in adult and one increased significantly in juvenile. Next, regional identification at 50 μm spatial resolution from coronal sections was obtained with matrix implanted laser desorption/ionization mass spectrometry imaging (MILDI-MSI) by implanting silver nanoparticulate matrices followed by focused laser desorption. Most of the CER and SM quantified in whole brain extracts were detected in MILDI images. Coronal sections from three brain levels show qualitative regional changes in CER-SM ion intensities, as a function of group and brain region, in cortex, striatum, accumbens, habenula, and hippocampus. Highly correlated changes in certain white matter CER-SM pairs occur in regions across all groups, including the hippocampus and the lateral (but not medial) cerebellar cortex of adult mice. Our data provide the first microscale MS evidence of regional lipid intensity variations induced by alcohol.
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http://dx.doi.org/10.1021/cn500174cDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4372063PMC
February 2015

MALDI-Ion Mobility Mass Spectrometry of Lipids in Negative Ion Mode.

Anal Methods 2014 Jul;6(14):5001-5007

Integrative Neuroscience, NIDA IRP, NIH, Baltimore, MD 21224, USA.

Profiling and imaging MALDI mass spectrometry (MS) allows detection and localization of biomolecules in tissue, of which lipids are a major component. However, due to the nature of this technique, complexity of tissue and need for a chemical matrix, the recorded signal is complex and can be difficult to assign. Ion mobility adds a dimension that provides coarse shape information, separating isobaric lipids, peptides, and oligonucleotides along distinct familial trend lines before mass analysis. Previous work using MALDI-ion mobility mass spectrometry to analyze and image lipids has been conducted mainly in positive ion mode, although several lipid classes ionize preferentially in negative ion mode. This work highlights recent data acquired in negative ion mode to detect glycerophosphoethanolamines (PEs), glycerophosphoserines (PSs), glycerophosphoglycerols (PGs), glycerolphosphoinositols (PIs), glycerophosphates (PAs), sulfatides (STs), and gangliosides from standard tissue extracts and directly from mouse brain tissue. In particular, this study focused on changes in ion mobility based upon lipid head groups, composition of radyl chain (# of carbons and double bonds), diacyl versus plasmalogen species, and hydroxylation of species. Finally, a MALDI-ion mobility imaging run was conducted in negative ion mode, resulting in the successful ion mapping of several lipid species.
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http://dx.doi.org/10.1039/C4AY00320ADOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4078893PMC
July 2014

Imaging of lipids in rat heart by MALDI-MS with silver nanoparticles.

Anal Bioanal Chem 2014 Feb 6;406(5):1377-86. Epub 2013 Dec 6.

Structural Biology Unit, NIDA IRP, NIH, 333 Cassell Drive, Room 1120, Baltimore, MD, 21224, USA.

Lipids are a major component of heart tissue and perform several important functions such as energy storage, signaling, and as building blocks of biological membranes. The heart lipidome is quite diverse consisting of glycerophospholipids such as phosphatidylcholines (PCs), phosphatidylethanolamines (PEs), phosphatidylinositols (PIs), phosphatidylglycerols (PGs), cardiolipins (CLs), and glycerolipids, mainly triacylglycerols (TAGs). In this study, mass spectrometry imaging (MSI) enabled by matrix implantation of ionized silver nanoparticles (AgNP) was used to map several classes of lipids in heart tissue. The use of AgNP matrix implantation was motivated by our previous work showing that implantation doses of only 10(14)/cm(2) of 2 nm gold nanoparticulates into the first 10 nm of the near surface of the tissue enabled detection of most brain lipids (including neutral lipid species such as cerebrosides) more efficiently than traditional organic MALDI matrices. Herein, a similar implantation of 500 eV AgNP(-) across the entire heart tissue section results in a quick, reproducible, solvent-free, uniform matrix concentration of 6 nm AgNP residing near the tissue surface. MALDI-MSI analysis of either positive or negative ions produce high-quality images of several heart lipid species. In negative ion mode, 24 lipid species [16 PEs, 4 PIs, 1 PG, 1 CL, 2 sphingomyelins (SMs)] were imaged. Positive ion images were also obtained from 29 lipid species (10 PCs, 5 PEs, 5 SMs, 9 TAGs) with the TAG species being heavily concentrated in vascular regions of the heart.
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http://dx.doi.org/10.1007/s00216-013-7525-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5523126PMC
February 2014

Imaging of noncovalent complexes by MALDI-MS.

J Am Soc Mass Spectrom 2013 Dec 2;24(12):1950-6. Epub 2013 Oct 2.

National Institute on Drug Abuse, Intramural Research Program, National Institutes of Health, Baltimore, MD, 21224, USA.

Noncovalent interactions govern how molecules communicate. Mass spectrometry is an important and versatile tool for the analysis of noncovalent complexes (NCX). Electrospray mass spectrometry (ESI-MS) is the most widely used MS technique for the study of NCXs because of its softer ionization and easy compatibility with the solution phase of NCX mixtures. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has also been used to study NCXs. However, successful analysis depends upon several experimental factors, such as matrix selection, solution pH, and instrumental parameters. In this study, we employ MALDI imaging mass spectrometry to investigate the location and formation of NCXs, involving both peptides and proteins, in a MALDI sample spot.
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http://dx.doi.org/10.1007/s13361-013-0745-3DOI Listing
December 2013

Rapid sensitization of physiological, neuronal, and locomotor effects of nicotine: critical role of peripheral drug actions.

J Neurosci 2013 Jun;33(24):9937-49

In-Vivo Electrophysiology Unit, Behavioral Neuroscience Branch, National Institute on Drug Abuse-Intramural Research Program, National Institutes of Health, Department of Health and Human Services, Baltimore, Maryland 21224, USA.

Repeated exposure to nicotine and other psychostimulant drugs produces persistent increases in their psychomotor and physiological effects (sensitization), a phenomenon related to the drugs' reinforcing properties and abuse potential. Here we examined the role of peripheral actions of nicotine in nicotine-induced sensitization of centrally mediated physiological parameters (brain, muscle, and skin temperatures), cortical and VTA EEG, neck EMG activity, and locomotion in freely moving rats. Repeated injections of intravenous nicotine (30 μg/kg) induced sensitization of the drug's effects on all these measures. In contrast, repeated injections of the peripherally acting analog of nicotine, nicotine pyrrolidine methiodide (nicotine(PM), 30 μg/kg, i.v.) resulted in habituation (tolerance) of the same physiological, neuronal, and behavioral measures. However, after repeated nicotine exposure, acute nicotine(PM) injections induced nicotine-like physiological responses: powerful cortical and VTA EEG desynchronization, EMG activation, a large brain temperature increase, but weaker hyperlocomotion. Additionally, both the acute locomotor response to nicotine and nicotine-induced locomotor sensitization were attenuated by blockade of peripheral nicotinic receptors by hexamethonium (3 mg/kg, i.v.). These data suggest that the peripheral actions of nicotine, which precede its direct central actions, serve as a conditioned interoceptive cue capable of eliciting nicotine-like physiological and neural responses after repeated nicotine exposure. Thus, by providing a neural signal to the CNS that is repeatedly paired with the direct central effects of nicotine, the drug's peripheral actions play a critical role in the development of nicotine-induced physiological, neural, and behavioral sensitization.
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http://dx.doi.org/10.1523/JNEUROSCI.4940-12.2013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3682376PMC
June 2013

Gangliosides and ceramides change in a mouse model of blast induced traumatic brain injury.

ACS Chem Neurosci 2013 Apr 17;4(4):594-600. Epub 2013 Jan 17.

Structural Biology Unit, NIDA IRP, National Institutes of Health, Baltimore, MD 21224, USA.

Explosive detonations generate atmospheric pressure changes that produce nonpenetrating blast induced "mild" traumatic brain injury (bTBI). The structural basis for mild bTBI has been extremely controversial. The present study applies matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging to track the distribution of gangliosides in mouse brain tissue that were exposed to very low level of explosive detonations (2.5-5.5 psi peak overpressure). We observed major increases of the ganglioside GM2 in the hippocampus, thalamus, and hypothalamus after a single blast exposure. Moreover, these changes were accompanied by depletion of ceramides. No neurological or brain structural signs of injury could be inferred using standard light microscopic techniques. The first source of variability is generated by the Latency between blast and tissue sampling (peak intensity of the blast wave). These findings suggest that subtle molecular changes in intracellular membranes and plasmalemma compartments may be biomarkers for biological responses to mild bTBI. This is also the first report of a GM2 increase in the brains of mature mice from a nongenetic etiology.
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http://dx.doi.org/10.1021/cn300216hDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3629744PMC
April 2013

MALDI/post ionization-ion mobility mass spectrometry of noncovalent complexes of dopamine receptors' epitopes.

J Proteome Res 2013 Apr 25;12(4):1668-77. Epub 2013 Mar 25.

NIDA IRP, NIH Structural Biology Unit Cellular Neurobiology Branch , 333 Cassell Drive, Baltimore, Maryland 21224, United States.

Protein domains involved in receptor heteromer formation are disordered and rich in the amino acids necessary for the formation of noncovalent complexes (NCX). We present mass spectral NCX data from proteins and protein receptors' epitopes obtained by combining ion mobility (IM) and MALDI. We focus on NCX involved in heteromer formation occurring between epitopes of the Dopamine D2 (D2R) and Adenosine A2A receptors (A2AR) as well as D2R and the α2 nicotinic (NR) receptor's subunit. The IM data yield information on the gas phase conformation of the singly charged NCX which are observed either directly from MALDI or as codesorbed neutrals that are subsequently postionized by a time-delayed excimer laser pulse directed onto a portion of the neutral plume created by the MALDI desorption laser. Imaging mass spectrometry of the matrix/epitope dried droplet surface shows that the acidic and basic epitopes and their NCX are found to be spatially collocated within regions as small as 25 × 50 μm(2). Subtle differences in the relative abundance of protonated and cationized NCX and epitopes are measured in spatial regions near the sodium-rich outer border of the droplet.
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http://dx.doi.org/10.1021/pr301004wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4144030PMC
April 2013

A new interpretative paradigm for Conformational Protein Diseases.

Curr Protein Pept Sci 2013 Mar;14(2):141-60

IRCCS San Camillo, Lido Venezia, Italy.

Conformational Protein Diseases (CPDs) comprise over forty clinically and pathologically diverse disorders in which specific altered proteins accumulate in cells or tissues of the body. The most studied are Alzheimerβ's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, prion diseases, inclusion body myopathy, and the systemic amyloidoses. They are characterised by three dimensional conformational alterations, which are often rich in β- structure. Proteins in this non-native conformation are highly stable, resistant to degradation, and have an enhanced tendency to aggregate with like protein molecules. The misfolded proteins can impart their anomalous properties to soluble, monomeric proteins with the same amino acid sequence by a process that has been likened to seeded crystallization. However, these potentially pathogenic proteins also have important physiological actions, which have not completely characterized. This opens up the question of what process transforms physiological actions into pathological actions and most intriguing, is why potentially dangerous proteins have been maintained during evolution and are present from yeasts to humans. In the present paper, we introduce the concept of mis-exaptation and of mis-tinkering since they may help in clarifying some of the double edged sword aspects of these proteins. Against this background an original interpretative paradigm for CPDs will be given in the frame of the previously proposed Red Queen Theory of Aging.
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http://dx.doi.org/10.2174/1389203711314020006DOI Listing
March 2013

Cellular membrane phospholipids act as a depository for quaternary amine containing drugs thus competing with the acetylcholine/nicotinic receptor.

J Proteome Res 2012 Jun 30;11(6):3382-9. Epub 2012 Apr 30.

Integrative Neuroscience, NIDA IRP, NIH , Baltimore, Maryland 21224, United States.

We previously demonstrated that ammonium- or guanidinium-phosphate interactions are key to forming noncovalent complexes (NCXs) through salt bridge formation with G-protein coupled receptors (GPCR), which are immersed in the cell membrane's lipids. The present work highlights MALDI ion mobility coupled to orthogonal time-of-flight mass spectrometry (MALDI IM oTOF MS) as a method to determine qualitative and relative quantitative affinity of drugs to form NCXs with targeted GPCRs' epitopes in a model system using, bis-quaternary amine based drugs, α- and β- subunit epitopes of the nicotinic acetylcholine receptor' (nAChR) and phospholipids. Bis-quaternary amines proved to have a strong affinity for all nAChR epitopes and negatively charged phospholipids, even in the presence of the physiological neurotransmitter acetylcholine. Ion mobility baseline separated isobaric phosphatidyl ethanolamine and a matrix cluster, providing an accurate estimate for phospholipid counts. Overall this technique is a powerful method for screening drugs' interactions with targeted lipids and protein respectively containing quaternary amines and guanidinium moieties.
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http://dx.doi.org/10.1021/pr300184gDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4144022PMC
June 2012

Cell growth on different types of ultrananocrystalline diamond thin films.

J Funct Biomater 2012 Aug 16;3(3):588-600. Epub 2012 Aug 16.

Materials Science Division, Argonne National Laboratory, 9700 S. Cass Ave, Argonne, IL 60439, USA.

Unique functional materials provide a platform as scaffolds for cell/tissue regeneration. Investigation of cell-materials' chemical and biological interactions will enable the application of more functional materials in the area of bioengineering, which provides a pathway to the novel treatment for patients who suffer from tissue/organ damage and face the limitation of donation sources. Many studies have been made into tissue/organ regeneration. Development of new substrate materials as platforms for cell/tissue regeneration is a key research area. Studies discussed in this paper focus on the investigation of novel ultrananocrystalline diamond (UNCD) films as substrate/scaffold materials for developmental biology. Specially designed quartz dishes have been coated with different types of UNCD films and cells were subsequently seeded on those films. Results showed the cells' growth on UNCD-coated culture dishes are similar to cell culture dishes with little retardation, indicating that UNCD films have no or little inhibition on cell proliferation and are potentially appealing as substrate/scaffold materials. The mechanisms of cell adhesion on UNCD surfaces are proposed based on the experimental results. The comparisons of cell cultures on diamond-powder-seeded culture dishes and on UNCD-coated dishes with matrix-assisted laser desorption/ionization-time-of-flight mass spectroscopy (MALDI-TOF MS) and X-ray photoelectron spectroscopy (XPS) analyses provided valuable data to support the mechanisms proposed to explain the adhesion and proliferation of cells on the surface of the UNCD platform.
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http://dx.doi.org/10.3390/jfb3030588DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4031007PMC
August 2012

Optimization of automated matrix deposition for biomolecular mapping using a spotter.

J Mass Spectrom 2011 Oct;46(10):1046-50

Cellular Neurobiology, NIDA IRP, National Institutes of Health, Baltimore, MD, USA.

Imaging mass spectrometry using matrix-assisted laser desorption/ionization allows the detailed mapping of biomolecules directly from tissue. Matrix deposition is the key step for successful imaging. The appropriate concentration and deposition of matrix is critical for extraction, desorption, and ionization of molecules from tissue without losing molecular localization. The main challenge to meet these criteria is to deposit matrix droplets homogeneously on the tissue section. This work shows how a chemical inkjet printer was used for this purpose resulting in the imaging of phosphatidylcholines and sulfatides. The intricacies involved in effective matrix deposition are discussed.
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http://dx.doi.org/10.1002/jms.1986DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3198816PMC
October 2011

Analysis of native biological surfaces using a 100 kV massive gold cluster source.

Anal Chem 2011 Nov 18;83(22):8448-53. Epub 2011 Oct 18.

Department of Chemistry, Texas A&M University, College Station, Texas 77843-3255, United States.

In the present work, the advantages of a new, 100 kV platform equipped with a massive gold cluster source for the analysis of native biological surfaces are shown. Inspection of the molecular ion emission as a function of projectile size demonstrates a secondary ion yield increase of ~100× for 520 keV Au(400)(4+) as compared to 130 keV Au(3)(1+) and 43 keV C(60). In particular, yields of tens of percent of molecular ions per projectile impact for the most abundant components can be observed with the 520 keV Au(400)(4+) probe. A comparison between 520 keV Au(400)(4+) time-of-flight-secondary ion mass spectrometry (TOF-SIMS) and matrix assisted laser desorption ionization-mass spectrometry (MALDI-MS) data showed a similar pattern and similar relative intensities of lipid components across a rat brain sagittal section. The abundant secondary ion yield of analyte-specific ions makes 520 keV Au(400)(4+) projectiles an attractive probe for submicrometer molecular mapping of native surfaces.
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http://dx.doi.org/10.1021/ac201481rDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3236644PMC
November 2011