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Observing heterogeneity in neutrophil phagosomes

A postgraduate research opportunity at the University of Otago.

Details

Close date
Sunday, 27 February 2022
Academic background
Sciences, Health Sciences
Host campus
Christchurch
Qualification
Honours
Department
Pathology and Biomedical Science (Christchurch)
Supervisor
Professor Tony Kettle

Overview

Neutrophils – the most abundant white blood cell – fight infections by engulfing bacteria into phagosomes, and then killing them with oxidants, proteases and bactericidal peptides. Oxidants are produced when an NADPH-oxidase assembles on phagosomal membranes and generates superoxide, which is used by myeloperoxidase to oxidize chloride to the toxic oxidant hypochlorous acid (HOCl). Recently, our group demonstrated that there is considerable heterogeneity between individual neutrophil phagosomes in their ability to produce HOCl. Formation of HOCl occurred immediately upon phagocytosis in most phagosomes but there was a distinct group of phagosomes in which HOCl was not produced until well after bacteria should have been killed. These results suggest that the killing mechanisms neutrophils deploy into individual phagosomes may vary, with some providing more lethal conditions than others. By using a variety of different mechanisms to kill ingested bacteria, neutrophils may restrict bacteria from evolving virulence factors that maintain viability within phagosomes.

The aims of this project are to establish why oxidant production varies between individual phagosomes in neutrophils, and whether phagosomal heterogeneity also extends to the deployment of cytotoxic peptides and proteins.

The student will learn to isolate neutrophils from human blood, and culture and count bacteria. They will use live cell microscopy to investigate the time course and extent of oxidant production in individual neutrophil phagosomes using specific probes for HOCl and superoxide. Oxidant production inside neutrophils will be quantified by flow cytometry. The student will also use immunochemistry to stain neutrophils for specific components of the NADPH-oxidase and myeloperoxidase to assess whether these oxidative enzymes are associated with all or a limited number of phagosomes. In addition, the student will use immunochemistry to stain neutrophils for other major cytotoxic peptides and proteins, such as defensins and elastase, to establish whether their content varies between phagosomes.

Preferred student expertise

The student should have a background in immunology and microbiology and an interest in biochemistry.

Further information

This is one of a number of projects on offer for the 2022 intake of BBiomedSc(Hons) at the University of Otago, Christchurch campus.

UOC BBiomedSc(Hons)

Professor Tony Kettle

Centre for Free Radical Research

Department of Pathology and Biomedical Science

Contact

Professor Tony Kettle
Email   tony.kettle@otago.ac.nz