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Lipidomic characterization and localization of phospholipids in the human lung.

ABSTRACT: Lipids play a central role in lung physiology and pathology; however, a comprehensive lipidomic characterization of human pulmonary cells relevant to disease has not been performed. The cells involved in lung host defense, including alveolar macrophages (AMs), bronchial epithelial cells (BECs), and alveolar type II cells (ATIIs), were isolated from human subjects and lipidomic analysis by LC-MS and LC-MS/MS was performed. Additionally, pieces of lung tissue from the same donors were analyzed by MALDI imaging MS in order to determine lipid localization in the tissue. The unique distribution of phospholipids in ATIIs, BECs, and AMs from human subjects was accomplished by subjecting the large number of identified phospholipid molecular species to univariant statistical analysis. Specific MALDI images were generated based on the univariant statistical analysis data to reveal the location of specific cell types within the human lung slice. While the complex composition and function of the lipidome in various disease states is currently poorly understood, this method could be useful for the characterization of lipid alterations in pulmonary disease and may aid in a better understanding of disease pathogenesis.

SUBMITTER: Zemski Berry KA

PROVIDER: S-EPMC5408611 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Lipidomic characterization and localization of phospholipids in the human lung.

Zemski Berry Karin A KA Murphy Robert C RC Kosmider Beata B Mason Robert J RJ

Journal of lipid research 20170309 5

Lipids play a central role in lung physiology and pathology; however, a comprehensive lipidomic characterization of human pulmonary cells relevant to disease has not been performed. The cells involved in lung host defense, including alveolar macrophages (AMs), bronchial epithelial cells (BECs), and alveolar type II cells (ATIIs), were isolated from human subjects and lipidomic analysis by LC-MS and LC-MS/MS was performed. Additionally, pieces of lung tissue from the same donors were analyzed by ...[more]

PMID: 28280112

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Project description:There is a wide variety of extracellular vesicles (EVs) that differ in size and cargo composition. EVs isolated from human plasma or serum carry lipid, protein, and RNA cargo that provides insights to the regulation of normal physiological processes, and to pathological states. Specific populations of EVs have been proposed to contain protein and RNA cargo that are biomarkers for neurologic and systemic diseases. Although there is a considerable amount of evidence that circulating lipids are biomarkers for multiple disease states, it not clear if these lipid biomarkers are enriched in EVs, or if specific populations of EVs are enriched for particular classes of lipid. A highly reproducible workflow for the analysis of lipid content in EVs isolated from human plasma or serum would facilitate this area of research. Here we optimized an MS/MSALL workflow for the untargeted analysis of the lipid content in EVs isolated from human serum. A simple sequential ultracentrifugation protocol isolated three distinct types of serum EVs that were identified based on size, targeted protein, and untargeted lipidomic analyses. EVs in the upper and middle fractions were approximately 140 nm in diameter, while EVs in the pellet were approximately 110 nm in diameter. EVs in the upper most buoyant fractions contained the highest concentration of lipids, were enriched with phospholipids, and immunopositive for the cytoskeletal markers actin, α-actinin, and the mitochondrial protein mitofillin, but negative for the typical EV markers CD63, TSG101, and flotillin. A central fraction of EVs was devoid of cytoskeletal and mitochondrial markers, and positive for CD63, and TSG101, but negative for flotillin. The EV pellet contained no cytoskeletal or mitochondrial markers, but was positive for CD63, TSG101, and flotillin. The EV pellet contained the lowest concentration of most lipids, but was enriched with ceramide. These results provided new insights into the lipid composition of EVs isolated from serum using a simple ultracentrifugation isolation method suitable for lipidomic analysis by mass spectrometry.

Dataset Information

Lipidomic characterization and localization of phospholipids in the human lung.

Publications

Lipidomic characterization and localization of phospholipids in the human lung.

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