Chicken intestinal organoids: a novel in vitro system to study mucosal vaccine targeting

Lead Research Organisation: University of Edinburgh
Department Name: The Roslin Institute

Abstract

The project seeks to fully characterise an avian minigut model and to exploit the model's multifunctional potential. The ultimate goal is to use these chicken miniguts to investigate new strategies to target mucosal vaccines to M cells and enhance their uptake across the gut epithelium. Prior to this application, the expertise to prepare and maintain these intestinal organoids from the chicken intestine has been established in the applicants' labs at The Roslin Institute.

Objective 1: Characterise the optimal growth conditions and cellular composition of avian gut organoids
Preliminary data show chicken organoids, similar to mouse organoids, can be prepared and maintained from chicken small intestine using mammalian growth factors. Recently, we cloned chicken epithelial growth factor, R-spondin 1 and Noggin and the proteins were produced. The optimal concentrations required for growth and differentiation into crypt and villus domains will be established. Detailed cellular and molecular characterisation of the organoids is of great importance to show that chicken organoids are excellent in vitro models for the chicken gut. The composition of organoids prepared from different parts of the intestine will be analysed using state-of-the art technology present at the Roslin Institute. Bioimaging (FACS, live imaging, confocal and multiphoton microscopy), transcriptomics (RNAsequencing) and bioinformatics will be used to compare the cellular composition and gene expression profiles of these organoids to that in the gut in vivo.

Objective 2: Study antigen uptake by M cells in the gut epithelium
Little is known of the immunobiology of chicken M cells. Therefore the cellular and molecular mechanisms which influence antigen uptake in the gut epithelium will be studied. We have cloned and produced chicken RANKL, and in homology to the mammalian protein, its capacity to induce chicken M cells will be tested. Antigen uptake in the presence or absence of M cells will be studied using fluorescent pathogens, beads coated with PAMPs and surface modified nanoparticles such as PLGA. The expertise of MSD on adjuvants and vaccines will ensure that the student investigates the state-of-the-art mucosal delivery systems and how they influence the uptake of different vaccine preparations across the gut epithelium. The aim is to identify biomarkers that correlate with induction of beneficial innate responses and maintenance of gut integrity.

Objective 3: to study host-pathogen interactions at the gut epithelium
The organoids will also be used to study the cellular and molecular responses of the gut epithelium following exposure to important avian pathogens such as Eimeria and Salmonella. Organoids will be exposed to Eimeria sporozoites and attachment, invasion and proliferation within the epithelial cells monitored by bioimaging and transcriptomics as above. Our data show Salmonella in mammalian gut takes advantage of the M cells' capacity to transcytose antigen to infect the host. In parallel the student with also study how S. Typhimurium interacts with the avian gut epithelium.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M014819/1 01/10/2015 30/11/2020
1658198 Studentship BB/M014819/1 01/10/2015 30/09/2019 Sebastian Cotton
 
Description We have developed an in vitro system to grow chicken intestinal organoids, which have the potential to be implemented in both research and industry intestinal-based activities. This intestinal model will reduce the use of animals in research.
Exploitation Route This in vitro model system can be utilised in research and industry for use in chicken intestinal applications.
Sectors Agriculture, Food and Drink,Environment,Pharmaceuticals and Medical Biotechnology

 
Description A patent application regarding this work has been filed.
First Year Of Impact 2020
 
Description General Scheme - Lonneke Vervelde - Chicken Intestinal Organoids: A Novel In Vitro System To Study Mucosal Vaccine Targeting
Amount £24,000 (GBP)
Funding ID WT 5906748 
Organisation Intervet International B.V. 
Sector Private
Country Netherlands
Start 09/2017 
End 08/2021
 
Description Research Grant - Lonneke Vervelde - Development of in vitro chicken enterocyte and organoid cultures
Amount £136,852 (GBP)
Funding ID WT 6571120 
Organisation Novozymes 
Sector Public
Country Denmark
Start 12/2018 
End 11/2019
 
Description Short term support for Dr Sam Sives
Amount £8,692 (GBP)
Funding ID BB-R506564-1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 01/2020 
End 03/2021
 
Description MSD Animal Health 
Organisation MSD Animal Health
Country United Kingdom 
Sector Private 
PI Contribution Our research team has provided adequate facilities, materials, equipment, supervision and training for the my work on the Project.
Collaborator Contribution MSD provided me with a placement at its premises for 10 days in 2019. During the placement MSD provided adequate facilities, equipment, supervision and training for my work on the Project, as well as paying the travel and accommodation costs incurred during the placement.
Impact Development of a technique to isolate sporozoites from Eimeria.
Start Year 2017
 
Description Invited lecture at MSD-Animal Health 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact Invited lecture at MSD-Animal Health in Boxmeer the Netherlands, with WebEx connection to all other location o MSD -AH around the world.
Year(s) Of Engagement Activity 2019
 
Description Talk given at RDSVS 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Talk on project given to qualified veterinary surgeons as part of their 10 year reunion, demonstrating the different career paths available outside of general practice. This sparked questions and discussions on the project field as well as research careers in general.
Year(s) Of Engagement Activity 2018
 
Description Talk given to MSD DIscovery & Technology Dept 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Industry/Business
Results and Impact Talk summarising the findings of the project so far to ~15 members of MSD's Discovery and Technology department. MSD used the content of the talk to suggest ideas for further research in this field, and invited further presentations on this topic to other poultry-based industries/businesses.
Year(s) Of Engagement Activity 2019