Biotechnology and Biological Sciences Doctoral Training Programme
   
   
  

Extracellular Vesicle Loading Mechanisms - The Lipid Prospective

Lab rotation project description

This rotation project is to build the skills required for lipid specimen imaging and spectroscopy.

Cell types will be taken and imaged using fluorescent markers for lipids and lipid related proteins. These will be quantified using live cell optical techniques and combined with both cholesterol inhibitors and hypoxia.

The primary goal is to learn quantitative optical imaging of live cells, including image analysis, statistics and related dissemination of knowledge.

However the outcomes will be used to help target the transfections required for the photoxidation and electron microscopy aspect of the linked project.

Fact file

Research theme

MCO

Location

QMC

Rotation

LR1,2,3

Contact

Dr Kenton Arkill, School of Medicine

2nd supervisor


BBSRC Doctoral Training Partnerships
 

Linked PhD Project Outline

Linked PhD Project Outline - snippet

Extracellular vesicles have recently attracted significant attention from the biomedical community, due to their potential use as biomarkers. Vesicles carry both an RNA and protein cargo, the detection of which researchers believe could form the basis to develop a ‘liquid biopsy’, to non-invasively diagnose and monitor disease. Due to the ease of detecting RNA molecules, compared to the stringent antibody requirements to detect proteins, the RNA cargo of vesicles has become the key focus of these studies. Despite developing an understanding of different RNA molecules found within vesicles resulting from specific disease states, there is no clear understanding of the selection process for vesicle cargos. Therefore, identifying the vesicle loading mechanism will be critically important to not only the development of the use of vesicles as a diagnostic, but also to unlock their therapeutic potential as biological delivery agents.

We will test the hypothesis that the lipid curvature and biophysics of vesicles determines their cargo. To test this the following should be achieved:

  • Aim 1: Build fluorescent tags for mRNA with additional free radical activation (APEX II or miniSOG) to allow preferential staining in electron microscopy.
  • Aim 2: Use a depletion technique (e.g. Rapamycin analogue) for specific lipids within cells to determine defining lipid content and formation pathway.
  • Aim 3: Using reformed exosomes to determine the biophysical properties of the lipids in normal and depleted systems.
  • Aim 4: Determine if this holds true for systems over producing exosomes such as hypoxia to elude to mechanistic pathways.

Triangle project with

Victoria James:

Do the biophysical properties of exosomes relate to their role as in vivo extracellular messengers?

Amanda Wright:

Optical techniques for characterising membrane structure and dynamics

 

 

 

 

 

Biotechnology and Biological Sciences Doctoral Training Programme

The University of Nottingham
University Park
Nottingham, NG7 2RD

Tel: +44 (0) 115 8466946
Email: bbdtp@nottingham.ac.uk