Methods to study lipid alterations in neutrophils and the subsequent formation of neutrophil extracellular traps

Research output: Contribution to journalArticle

Abstract

Lipid analysis performed by high performance thin layer chromatography (HPTLC) is a relatively simple, cost-effective method of analyzing a broad range of lipids. The function of lipids (e.g., in host-pathogen interactions or host entry) has been reported to play a crucial role in cellular processes. Here, we show a method to determine lipid composition, with a focus on the cholesterol level of primary blood-derived neutrophils, by HPTLC in comparison to high performance liquid chromatography (HPLC). The aim was to investigate the role of lipid/cholesterol alterations in the formation of neutrophil extracellular traps (NETs). NET release is known as a host defense mechanism to prevent pathogens from spreading within the host. Therefore, blood-derived human neutrophils were treated with methyl-β-cyclodextrin (MβCD) to induce lipid alterations in the cells. Using HPTLC and HPLC, we have shown that MβCD treatment of the cells leads to lipid alterations associated with a significant reduction in the cholesterol content of the cell. At the same time, MβCD treatment of the neutrophils led to the formation of NETs, as shown by immunofluorescence microscopy. In summary, here we present a detailed method to study lipid alterations in neutrophils and the formation of NETs.

Details

Authors
  • Graham Brogden
  • Ariane Neumann
  • Diab M. Husein
  • Friederike Reuner
  • Hassan Y. Naim
  • Maren Von Köckritz-Blickwede
Organisations
External organisations
  • University of Veterinary Medicine Hannover
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Cell and Molecular Biology

Keywords

  • Cholesterol, High Performance Liquid Chromatography (HPLC), High Performance Thin Layer Chromatography (HPTLC), Immunology, Issue 121, Methyl-β-cyclodextrin (MβCD), Neutrophil Extracellular Traps (NETs), Neutrophils
Original languageEnglish
Article numbere54667
JournalJournal of Visualized Experiments
Volume2017
Issue number121
Publication statusPublished - 2017 Mar 29
Publication categoryResearch
Peer-reviewedYes