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This invention concerns a process for the in vitro preparation of stem cells from ruminant somatic cells by exogenous expression of a new combinatorial gene in said somatic cells. The stem cells thus obtained can be used to expand an individual genetic background, to screen molecules or to replicate and test the virulence of certain pathogens. These stem cells have some characteristics of pluripotent stem cells, such as self-renewal, infinite proliferation, specific cell cycle and expression of particular markers.
Ruminants and in particular cattle represent a large part of livestock production in France, Europe and the world. As the life cycle of animals is particularly long, the development of any method to maintain and amplify the genetic potential of an animal in the long term, via a cell with germline potential in particular, is a priority objective for the sectors. This project to reprogram bovine somatic cells was carried out in collaboration with Allice, and is part of the programs supported by the CRB-Anim project, Centre de Ressources Biologiques pour les Animaux Domestiques (CRB-ANIM - ANR-11-INBS-0003).
The process for the in vitro preparation of stem cells according to the invention includes, in addition to exogenous expression of a new gene combinatorium, expression of at least one of the genes selected from the group consisting of OCT4, SOX2 and KLF4, preferably at least OCT4, in said somatic cells.
The reprogramming of somatic cells by exogenous expression of new reprogramming genes results in a drastic morphological change, the elongated appearance of a large fibroblastic cell with an abundant cytoplasm disappearing in favour of a small round cell with a very refractive cytoplasm. The cells thus reprogrammed express markers and cellular properties comparable to stem cells, in particular in terms of self-renewal, proliferation, cell cycle, expression and telomerase activity.
Prospects :
The stem cells obtained can allow the expansion of an individual genetic fund, or the genomic identification of this genetic fund. They are therefore potentially a tool for the conservation and management of genetic resources. In another area, their proliferation capacities can be used to screen anti-proliferative molecules or test the action of these molecules, in particular certain drugs used to treat cancer pathologies. Finally, they can also be used to replicate and test the virulence of pathogens in ruminants, including pathogenic strains of bovine tuberculosis, or to study the mechanisms of infection and spread of these pathogens.
Valuation :
European patent application filed on 25 January 2017 under No. EP 17305082.4: : : "Process for reprogramming somatic ruminant cells" (application not yet published)
Co-depositors: INRA, INSERM, ALLICE