The pig as a research model

New therapies, drug testing, disease models and even the promise of organ generation for human use; these represent some examples of the unlimited potential of pigs in research. Laboratory of Dr. Vilceu Bordignon, McGill University

Although mice are the most common animal models in research, pigs have the advantage of anatomical, physiological, metabolic and genetic similarities with humans.
Since animal domestication, human intervention through breeding programs has allowed the establishment of various breeds of pig. Apart from breeds specialized for food production, smaller sized breeds (miniature- and micro-pigs) have also been selected. Their use in biomedical research has been increasing considerably in recent years, largely due to advantages they exhibit related to size and maintenance, when compared to regular sized breeds.

Building the models

Pigs offer many exciting applications, including stem cell research, tissue engineering and xenotransplantation. The combination of new molecular biology methods, genome editing tools and reproductive technology, such as in vitro embryo production and somatic cell nuclear transfer (SCNT or cloning), can now be applied to create genetically engineered pigs (both regular and minipigs).

Indeed, several research groups have successfully used these methods in regular and minipigs. Genome editing tools, including clustered, regularly interspaced, short palindromic repeats (CRISPR) together with CRISPR associated (Cas) nucleases (CRISPR/Cas), allow us to do precise manipulations in specific genes in cells, zygotes and developing embryos.

Dr Bordignon team is investigating somatic cell reprogramming, oocyte and embryo development, and using SCNT, in vitro embryo technologies and the CRISPR/Cas system to create unique models to study development, stress, metabolism and degenerative conditions.

The CRISPR/Cas system (Figure 1) includes the Cas9 nuclease, which cleaves DNA, and a single guide RNA (sgRNA), which determines the specific site in the genome to be cleaved by the Cas9. The system can be deliver to zygotes via cytoplasmic (through the membrane) or pronuclear (in the nucleus) microinjection. In embryos produced by cloning, it is possible to modify the genome either by delivering the CRISPR/Cas system in the nuclear donor cell before nuclear transfer or in the reconstructed zygotes (after SCNT).

Xenotransplantation

The main concerns surrounding xenotransplantation revolve around the transmission of infectious agents, as well as tissue/organ rejection. However, these challenges may be addressed through editing and engineering the pig genome. Porcine endogenous retroviruses (PERVs) are present in the pig’s DNA as inactive proviruses that can become active in other species. In 2015, researchers were able to inactivate 62 PERVs in pig embryos using the CRISPR/Cas system. Furthermore, many pig models have already been created to address tissue/organ rejections, which includes the inactivation of the α-Gal epitope, one of the main causes of tissue rejection. New studies combining the inactivation of both infectious agents and the mitigation of hyperacute rejection reactions have pushed xenotransplantation one step closer to successful application.

Farming human organs in pigs

Despite the great promise of xenotransplantation, the generation of pig-derived organs for use in humans is still a debatable issue. Meanwhile, researchers have been exploring the creation of non-immunogenic donor organs in pigs using human induced pluripotent stem cells (hiPSCs). This technology may become a reality in the coming years. Indeed, recent discoveries have shown chimera formation using hiPSCs injected into pig embryos. This may open a new perspective for circumventing the shortage of cell/organs for transplant.

Ethical concerns

The greatest limitation may not be the ability to generate efficient genetically modified animal models for research, but finding an adequate balance between research objectives and avoiding unnecessary animal suffering or distress. This is even more important when considering specific genome manipulations that are known to result in significant animal disease and/or discomfort.

Conclusion

The pig is a remarkable biomedical model that is gaining approval and recognition due to its similarities with humans. Innumerable applications can be derived from swine models. Despite all the potential benefits, many obstacles, both biological and ethical in nature, remain to be addressed.

References

Dooldeniya, M.D., and Warrens, A.N. (2003). Xenotransplantation: where are we today? J R Soc Med 96, 111-117.

Gutierrez, K., Dicks, N., Glanzner, W.G., Agellon, L.B., and Bordignon, V. (2015). Efficacy of the porcine species in biomedical research. Front Genet 6, 293.

Prather, R.S., Lorson, M., Ross, J.W., Whyte, J.J., and Walters, E. (2013). Genetically engineered pig models for human diseases. Annual review of animal biosciences 1, 203-219.

Walters, E.M., Wolf, E., Whyte, J.J., Mao, J., Renner, S., Nagashima, H., Kobayashi, E., Zhao, J., Wells, K.D., Critser, J.K., et al. (2012). Completion of the swine genome will simplify the production of swine as a large animal biomedical model. BMC Med Genomics 5, 55.

Wu, J., Platero-Luengo, A., Sakurai, M., Sugawara, A., Gil, M.A., Yamauchi, T., Suzuki, K., Bogliotti, Y.S., Cuello, C., Morales Valencia, M., et al. (2017). Interspecies Chimerism with Mammalian Pluripotent Stem Cells. Cell 168, 473-486 e415.

Yang, L., Guell, M., Niu, D., George, H., Lesha, E., Grishin, D., Aach, J., Shrock, E., Xu, W., Poci, J., et al. (2015). Genome-wide inactivation of porcine endogenous retroviruses (PERVs). Science 350, 1101-1104.

RQR 24 Hours of Science – Genome Code modification : How far can we go?

The 24 Hours of Science is a day of science and technology activities for audiences of all ages, taking place throughout Quebec and varying from one hour to 24 hours. The objective is to foster meetings between researchers and the general public, stimulate the general interest in science and technology, and promote scientific careers for youth.

This year’s theme is « Science and Fiction ». Cloning, gene editing, gene therapy: that appeared to be science fiction just 50 years ago may have already come true!

Therefore, the RQR invite you to its activity entitled « Genome Code Modification: How far can we go? » to discuss about gene editing with Dr. Vardit Ravitsky, RQR member and professor at the Université de Montréal. What are the techniques? What are the applications in reproductive biology? What are the ethical issues raised by this new technology?

The event is free and will take place at the Café des Beaux-Arts of Montréal on Saturday, May 13th from 13:30 to 15:00. For more information or to register, please contact the network manager by email at j.blouin@umontreal.ca or by phone at (450) 773-8521 ext. 8286.

RQR Knowledge TransferAward

During the 9th Symposium in November, the RQR presented awards of $400 to two members who distinguished themselves for their engagement with the end users. The awardees are Dr. Géraldine Delbès from the INRS and Phanie L. Charest, MSc student at the Université Laval.

By knowledge transfer, it is implied that the activities are intended for end users and not only to the scientific community. Thus we speak of scientific popularization to make the reproductive biology accessible to end users, including:

• the general public,
• pharmaceuticaland biotech companies,
• veterinarians,
• clinicians,
• government
• and other appropriate targets.

Dr. Delbès distinguished herself through her participation in activities for the general public such as the Pint of Science Canada Festival. She also participated to RQR video entitled « Au coeur de l’expertise en reproduction humaine et animale » on the TV program « Quoi de neuf chercheur » at the Canal Savoir. Phanie has been involved in giving conferences and workshops for veterinary pathologists and
toxicologists as well as animal health technicians.

On behalf the RQR members, congratulations to both recipients.