Identification of stem/progenitor cells in the postnatal thymus
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Date
28/06/2014Author
Ulyanchanka, Sviatlana
Metadata
Abstract
The thymus is the principal site of T-cell development and maturation. Failure to
develop a functional thymus leads to severe immunodeficiency, while partially
incorrect function of the organ can lead to a variety of autoimmune diseases as well
as higher risk for infections and cancer. The thymus is organized into cortical and
medullary regions, which are functionally distinct. The diverse array of thymic
epithelial cells (TEC) are the key components of the thymic stroma, both the cortical
and medullary TEC subsets are responsible for the establishment of a self-tolerant
and self-restricted T-cell repertoire.
The thymus is most active in young individuals, and undergoes a progressive
naturally occurring involution from birth, which accelerates after puberty. Thymic
involution is characterized by loss of thymus organization and function, including an
overall reduction in the amount of functional thymic tissue. This results in decreased
production of new naïve T-cells, and contributes to the diminished capacity of the
aged immune system to adequately respond to new antigenic challenge. Involution of
the thymus, both natural and in response to different therapies such as chemotherapy,
raises interest in developing cell based treatment methods that will allow the
restoration of the thymic architecture and so elevate immune reconstitution in vivo.
The cellular mechanisms by which the postnatal thymus is maintained during
homeostasis and involution are currently unknown. The earliest thymic progenitors
in the thymus express Plet1; it has been established that from E12.5 to E15.5 these
cells when purified are able to generate all thymic epithelial cell types and initiate
thymus organogenesis. However, at least the latter capacity is reported to be lost
from E18.5. A number of papers published provide evidence for the existence of both
bipotent and unipotent TEC progenitors in the adult thymus. However the identity of
these cells remains unknown, nor has the relationship between the mature and
immature postnatal TEC compartments been established.
The aim of my research was to investigate the cellular mechanism(s) that maintain
the postnatal thymus. Specifically, I aimed to determine whether the thymus is
maintained by a stem cell mechanism or by division of terminally differentiated
thymic epithelial cells, and whether or not postnatal thymic epithelial
stem/progenitor cells express functionally relevant levels of the transcription factor
Foxn1.
To address these aims, I used two approaches: in vivo genetically heritable lineage
tracing and a novel grafting assay to assess the contribution of different lineages of
TEC.
This thesis describes the characterization of a novel mouse strain, the Foxn1CreERt2
line, which was predicted to allow conditional inducible manipulation of gene
expression in TEC. I show that this deletor strain, while thymic epithelial cellspecific,
could induce cre-mediated recombination in only in a low proportion of
TEC and thus could not be used to address the initial aim of this work as described
above. However, lineage tracing experiments using this line have provided evidence
for a persistent cortical thymic epithelial progenitor/stem cell type, that was capable
of rapid expansion within the cortical compartment over time. In parallel with
characterisation of the Foxn1CreERt2 strain, I investigated the potential of various
defined epithelial populations to contribute to the thymic environment in an assay of
TEC potency. Using this technique I have established the potential of defined TEC
subpopulations isolated from postnatal mice to generate cortical and medullary TEC.
Among the populations analysed I have identified a minor TEC subset that can
robustly contribute to both cortical and medullary TEC that coexpress Ly51 and
Plet1. I have further shown, using a limiting dilution approach, that this population
contains a postnatal common thymic epithelial stem/progenitor cells, present at a
frequency of between 87.5 and 92.5 within this population. I have also produced
evidence of a unipotent cortical progenitor population that is capable of long term
expansion in vivo.