SCIENCE sciencemag.org
GRAPHIC: V. ALTOUNIAN/
SCIENCE
By Violeta Silva-Vargas and Fiona Doetsch
T
he cerebrospinal fluid (CSF) is an
optically clear but molecularly com-
plex liquid that flows within the
brain ventricles. It cushions the brain
and delivers nutrients and signaling
molecules while removing others.
The CSF is produced by the choroid plexus
(ChP), an understudied epithelial barrier
that regulates the entry of factors from the
blood into CSF and is also highly secre-
tory. As a central hub with multiple
functions, the ChP is emerging as
a key contributor to normal brain
physiology and disease ( 1 ). A lack
of tools has limited exploration of
the ChP, especially in humans. On
page 159 of this issue, Pellegrini et
al. ( 2 ) establish human ChP organ-
oids, three-dimensional multicel-
lular in vitro structures. They form
compartments filled with a CSF-like
fluid and exhibit functional barrier
and secretion properties, resem-
bling those in vivo. They are a pow-
erful tool to predict drug permeabil-
ity and investigate ChP secretion
and cell diversity.
The CSF is a complex and dynamic
milieu whose composition changes
at different stages of development, in
adulthood and with aging, as well as
in different physiological states ( 1 , 3 ,
4 ). CSF composition even varies di-
urnally in adults. A variety of biologi-
cally active moieties—including signaling
and growth factors, hormones, lipoproteins,
neurotransmitters, extracellular matrix, and
extracellular vesicles—are present in the CSF
( 1 , 3 ). The ChP is a key, though not exclu-
sive, source and transport route of factors in
the CSF. The ChP and CSF together form a
highly tuned flowing milieu integrating the
delivery of local and long-range factors that
affect processes, including stem cell prolif-
eration, during development and adulthood,
as well as neural circuit plasticity and brain
physiology ( 1 , 3 , 5 – 8 ).
The ChP is a highly vascularized struc-
ture anchored to the wall of the brain ven-
tricle and floating in the CSF, with multiple
key functions ( 1 , 4 , 9 ). It consists of a single
outer epithelial cell layer surrounding an
inner layer with fenestrated blood vessels,
multiple stromal cell types, and surveilling
immune cells ( 1 ). Cuboidal choroidal epi-
thelial cells are highly polarized. Their api-
cal side contains many villi and/or cilia and
contacts the CSF. They are interconnected
by tight junctions, forming the blood-CSF
barrier, and tightly regulate access of fac-
tors from the blood into the CSF (see the
figure). Choroidal epithelial cells are meta-
bolically active and a key site for synthesis
and modification of numerous polypeptides
and detoxification of metabolites from the
CSF. They are highly secretory, with large
numbers of multivesicular bodies. Although
epithelial cells are the major source of se-
creted factors in vivo, all ChP cell types can
contribute to the CSF.
Two-dimensional cultures of primary
and embryonic stem cell–derived choroidal
epithelial cells have provided important in-
sight into ChP function. Organoids are self-
organizing, three-dimensional structures,
which more completely capture tissue com-
plexity ( 10 ). ChP tissue has previously been
induced in brain organoids ( 11 ). Pellegrini
et al. have developed a protocol for the ro-
bust generation of human ChP organoids
from pluripotent embryonic stem cells that
contain polarized epithelial cells, with tight
junctions that develop barrier properties
and secrete a clear liquid that resembles
in vivo CSF into a self-contained compart-
ment. A key issue for all organoid systems
is which developmental stage they model
and how mature the cells become. Single-
cell RNA sequencing and characterization
of the ChP organoids revealed progressively
mature epithelial cells and increasing num-
bers of stromal cells in older organoids that
resemble the cellular organization and mo-
lecular profiles of in vivo human ChP tis-
sue. Likewise, proteomics of the CSF-like
fluid over time showed that the organoid
CSF composition underwent a progressive
maturation to postnatal stages.
A major challenge in medicine is to de-
liver drugs to the brain, owing to
the blood-CSF and blood-brain bar-
riers. The blood-CSF barrier in the
ChP is a highly regulated system,
allowing the selective transport or
passage of key macromolecules and
small molecules ( 9 ). The human ChP
organoids reported by Pellegrini et
al. display barrier properties that
predict drug permeability. Sampling
the CSF-like fluid after exposing the
organoids to different drugs in the
medium revealed that the system re-
capitulates the (in)ability of known
compounds, including antidepres-
sants and chemotheraputic agents,
to cross the barrier and showed the
accumulation in organoid fluid of a
preclinical drug that induced neuro-
toxicity in patients. Thus, ChP-CSF
organoids may be a powerful screen-
ing platform for assessing and pre-
dicting blood-CSF drug permeability
in humans.
Morphologically, light and dark epithe-
lial cells have been described in vivo by
electron microscopy ( 12 ). Single-cell RNA
sequencing of human ChP organoids iden-
tified multiple epithelial cell subtypes with
distinct molecular signatures: light and
dark cells, which Pellegrini et al. show are
enriched in ciliary versus mitochondrial
genes, respectively, and an undescribed
population of myoepithelial cells, which are
contractile cells found in other secretory
organs. These subtypes of epithelial cells
showed differences in their secretory pro-
files. Future characterization of the stromal
cells found in the organoids and the addi-
tion of other cell types such as immune cells
PHYSIOLOGY
Exploring the source of
human brain fluids
Human organoid cultures can be used to probe secretion
and drug permeability into and out of the brain
Biozentrum, University of Basel, 4056 Basel, Switzerland.
Email: [email protected]
Secretion
Choroid plexus Barrier
Detoxify
Choroid ple Immune surveillance
Immune
cells
CSF
Blood
Stroma
CSF in
ventricles
Epithelium
Metabolites
Stromal cells
Choroid
plexus
Tight junction
10 JULY 2020 • VOL 369 ISSUE 6500 143
The multifunctional choroid plexus
A choroid plexus (ChP) is found in each brain ventricle and makes the
cerebrospinal fluid (CSF). Chorodial epithelial cells are interconnected
by tight junctions to form the blood-CSF barrier. Other ChP functions
include secretion, detoxification, and immune surveillance.