Self-organized Pituitary-like Tissue
from Mouse ES Cells
Tuesday, 15 November 2011
The possibility that functional,
three-dimensional tissues and organs may be derived from pluripotent cells,
such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs),
represents one of the grand challenges of stem cell research, but is also one
of the fundamental goals of the emerging field of regenerative medicine.
Developmental biology has played a central role in informing such efforts, as
it has been shown that stem cell differentiation can be directed to follow a
given lineage pathway by culturing stem cells in conditions that recapitulate
the specific cellular and molecular environment from which such cells normally
emerge during embryogenesis. Intriguingly, recent work has shown that when ES
cells are cultured under the appropriate conditions, they can be driven to
self-organize into complex, three-dimensional tissue-like structures that
closely resemble their physiological counterparts, a remarkable advance for the
field.
New work by Hidetaka Suga of the Division
of Human Stem Cell Technology, Yoshiki Sasai, Group Director of the Laboratory
for Organogenesis and Neurogenesis, and others has unlocked the most recent
achievement in self-organized tissue differentiation, steering mouse ESCs to
give rise to tissue closely resembling the hormone-secreting component of the
pituitary, known as the adenohypophysis, in vitro. Conducted in collaboration
with Yutaka Oiso at the Nagoya University Graduate School of Medicine, this
work was published in Nature.
The Sasai group has complied an
impressive list of achievements in induced differentiation using an ES cell
culture technique dubbed SFEBq (shorthand for "serum-free floating culture of embryoid body-like aggregates with
quick re-aggregation"), including high-efficiency methods for the
differentiation of dopaminergic, cerebral cortex, cerebellar Purkinje, and
other neuronal cell types. In recent years, refinements of this approach have
enabled the first tantalizing insights into the developmental capacity of
pluripotent stem cells, showing their ability to give rise to multi-cell-type
populations of cortical and retinal neurons that spontaneously self-organize
into stratified tissue nearly identical to that of the developing embryo.
In the group's most recent work, Suga
sought to use SFEBq to derive the secretory component of the pituitary
(hypophysis) from mouse ES cells. In embryonic development, the pituitary
emerges from a region of the non-neural (rostral) head ectoderm, adjacent to
the anterior neural plate. Guided by molecular interactions, this placodal
region forms an indentation, known as Rathke's pouch, in the developmental
predecessor of the roof of the mouth, and eventually gives rise to the anterior
section of the pituitary, the source of hormones involved in growth,
reproduction and modulating the physiological response to stress.
The Sasai lab had previously reported
how a modified version of the SFEBq approach lacking extrinsic growth factors
could spur ES cells to give rise to hypothalamic neurons. Building on this
finding, Suga found by further tweaking the conditions he could steer
populations of such stem cells to differentiate simultaneously into the
neighboring rostral head ectoderm and hypothalamic neuroectoderm, both of which
are required to development of the adenohypophysis. Interestingly, the cells in
these clusters spontaneously organized into distinct layers resembling those of
the corresponding embryonic tissues. This effect appears to be attributable to
the increase of BMP signaling in larger cell aggregates, leading to
differentiation into non-neural ectoderm. To test whether co-culture of this
ectoderm with hypothalamic tissue would lead to the formation of Rathke's
pouch-like structures, Suga observed the ESC-derived aggregates for nearly two
weeks, and found that by adding the signaling factor Sonic hedgehog, he was
able to induce the self-directed formation of this vesicular tissue.
The next important test was whether
this in vitro anterior pituitary anlage would show similar functionality to the
physiological activity of the adenohypophysis. Of the many critically important
pituitary hormones, they chose adenocorticotropic hormone (ACTH) for their
first assay. Previous research had suggested that Notch signaling interferes
with the development of ACTH-secreting cells, so the group added a Notch
blocker to the culture medium and found that this triggered the generation of
such cells at high efficiencies. Following a similar principle, they showed
that by adding Wnt, glucocorticoid and insulin to the culture at appropriate
doses and stages, they could obtain growth hormone-secreting cells in quantity.
By varying the recipe of the growth factor cocktail, they were able to induce
other pituitary hormones as well. And, critically, the group was able to show
that in vitro hormone secretion could respond to requisite signals and engage
in regulatory feedback just as in the body.
In a final series of experiments, Suga
et al. transplanted the ESC-derived ACTH-secreting tissue into the kidneys of
adult mice whose own pituitary glands had been ablated, to see whether it would
be capable of compensating for pituitary function in this model system. Within
a week of transplantation, these mice showed strong overall survival, a marked
rise in ACTH levels and a concomitant increase in corticosterone (a
glucocorticoid hormone stimulated by ACTH) over un-transplanted controls, which
uniformly weakened and died within eight weeks of hypophysectomy.
Yoshiki Sasai, leader of the study,
commented on this most recent demonstration of the remarkable self-organizing
capabilities of embryonic stem cells in vitro.
"We
have previously shown how ES cells can give rise to self-organized,
three-dimensional neuronal and sensory tissues, and in this report we describe
for the first time how this principle can be used to generate to an endocrine
tissue, suggesting our approach is of general applicability. Suga, himself an
endocrinologist, remarks, "We currently treat pituitary deficiencies by
hormone replacement, but achieving the correct dosage is not a straightforward
problem, given the naturally fluctuating levels secreted within the body. I am
hopeful that this new finding will lead to further advances in regenerative
medicine in the endocrine system."
Source:
RIKEN
Contact:
Douglas Sipp .........
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