UittrekselMitochondrial genetics is a rapidly expanding field due to the recent
development of massive parallel sequencing, which has allowed overcoming
several limitations historically associated to the study of the mitochondrial
DNA (mtDNA). The technological advancements went along with a growing
body of evidence associating mitochondrial function to the regulation of a
wide variety of cellular processes, including embryonic development and
pluripotency and differentiation of human pluripotent stem cells (hPSC).
The goal of our study was to analyse the variants in the mtDNA of hPSC,
establishing their origin and evaluating the effects of prolonged in vitro
culture. For this, we initially developed a method for an accurate analysis of
both single nucleotide variants and large deletions in the mtDNA applicable
also to single cells.
The method proved to be more accurate and sensitive than available
softwares, and suitable for the identification of multiple deletions within the
same sample, in both bulk DNA and single cells.
The study of the mtDNA of human embryonic stem cells showed few variants
at low frequency in the early passages, and an increased presence of
variants after prolonged in vitro culture. We identified the cause of the
increased mutation load as a combination of high-grade mtDNA mosaicism
and clonal culture takeover from a single cell, with the mtDNA variants being
passenger mutations. Spontaneous mutagenesis doesn’t seem to affect
significantly the mtDNA variant load during in vitro culture. High-grade
tissue mosaicism affects also the derivation of induced Pluripotent Stem
Cells, generating lines with higher loads of variants as compared to their
embryonic counterpart. Our work might be useful to develop guidelines for
the genetic screening of human PSCs in clinical applications.
|Datum Prijs||29 mrt 2018|
|Begeleider||Claudia Spits (Promotor), Luca Gianaroli (Co-promotor), Barbara Barboni (Co-promotor), Leo van Grunsven (Jury), Thierry VandenDriessche (Jury), Luc Leyns (Jury), Alexander Gheldof (Jury), Bert Smeets (Jury) & Catherine Verfaillie (Promotor)|