Developing stem cell-derived macrophages to study host pathogen interactions and advance strategies to combat globally important disease in the pig

Why did we fund this project?

This award aims to optimise the derivation of macrophages from porcine stem cells for the study of African Swine Fever Virus (ASFV) and Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), replacing the need to isolate macrophages from pigs.

ASFV and PRRSV are highly transmissible diseases with severe economic and welfare implications for the pig farming industry. ASFV has a high mortality rate, and with no vaccine available, containment and culling are the only strategies available for managing an outbreak. PRRSV impacts piglets and pregnant sows, with high morbidity and mortality in piglets and abortions in sows. Both ASFV and PRRSV preferentially replicate in macrophages and are therefore used for research into new treatments or mechanisms of infection. Primary macrophages are the current gold standard but as they do not proliferate in culture, must be continually sourced from sampled or sacrificed animals. Dr Thomas Burdon has developed a system to derive macrophages from pig stem cells. The stem cells can be grown indefinitely, genetically modified in vitro and readily differentiate into macrophages, reducing the reliance on pigs for viral research.

Thomas will optimise the protocol maximising the efficiency of macrophage generation to enable easier uptake of the cells by the virology research community. He will use fluorescent reporter cells to screen culture conditions and monitor the differentiation process. Thomas will also immortalise macrophage progenitor cells, differentiated from the pig stem cells, as novel cell lines enabling faster differentiation into macrophages. All the macrophages developed will be tested for their ability to propagate ASFV and PRRSV and the new protocols for the cells disseminated via a web resource. The technologies developed have wider applications studying further viruses and other pathogens, including bacteria and parasitic pathogens, and could also be applied to other livestock species.

African Swine Fever Virus (ASFV) and Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) are currently responsible for unparalleled financial losses and welfare concerns in commercial pig farming across the world. ASFV causes a lethal haemorrhagic fever that, in the absence of an effective vaccine, necessitates strict containment and mass culling to eradicate disease. PRRSV is less virulent but is widespread in pig herds and causes infertility and respiratory problems that make PRRS the most costly viral disease in commercial pig farming. Both viruses preferentially infect pig macrophages and studying these viruses until now has relied on primary macrophage cultures or transformed cell lines: both of which have major limitations. To overcome this limitation we have developed a stem cell-based culture system for producing normal pig
macrophages that are readily infected with ASFV and PRRSV. We propose that macrophages generated from pig stem cell lines provide a biologically relevant and scalable system for ASFV/PRRSV research and vaccine development, significantly reducing the requirement for pigs as either a source of ex vivo macrophages, or as subjects in viral challenge experiments.

The aim of this programme is to maximise the efficiency of generating pig macrophages in culture by 1) Optimising macrophage differentiation using haematopoietic/macrophage reporter cell lines to characterise, monitor and refine the differentiation process, and 2) Deriving conditionally immortalised macrophage progenitor cell lines. Reporter activity and transcriptional profiling of key stages in stem cell-macrophage differentiation will inform rational development of optimised protocols and facilitate the establishment of new pig macrophages progenitor cell lines. This novel stem cell based platform will reduce the number of pigs required in ASFV/PRRSV research and will illustrate the potential for applying stem cell-based technology more generally in livestock research.

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Project grant

Status

Pending start

Institution

University of Edinburgh

Grant reference number

NC/V001140/1

Award date

Nov 2020 - Oct 2022

Grant amount

£350,989