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2020
Killing the pathogen and sparing the placenta Killing the pathogen and sparing the placenta
Indira U. Mysorekar
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Clinical Implications of Basic R ese arch
The new england journal of medicine
n engl j med 383;21 nejm.org November 19, 2020
2080
Elizabeth G. Phimister, Ph.D., Editor
Killing the Pathogen and Sparing the Placenta
Indira U. Mysorekar, Ph.D.
A healthy human pregnancy requires a careful
balancing act: the maternal immune system
must not reject the fetus — which is half foreign
— and yet must be ready to attack pathogens
such as bacteria and viruses.
1
Decidual natural
killer (NK) cells help to maintain this balance.
These cells invade the maternal side of the pla-
centa (i.e., the decidua). However, the classical
lytic mode of NK activity, releasing cytotoxic
granules that contain membrane-lysing pep-
tides, would damage the host. Crespo and col-
leagues have recently described a new, more
precise mechanism by which decidual NK cells
attack pathogens without harming the placenta
and fetus.
2
The balancing act of maintaining immune
tolerance of the fetus and protective immunity
occurs at the maternal–fetal interface of the
placenta. The placenta forms when embryo-
derived cytotrophoblasts fuse to create multinu-
cleated syncytiotrophoblasts, which mediate the
maternal–fetal exchange of nutrients and gases.
Cytotrophoblasts also make up the villi that an-
chor the embryo to the uterine wall. Moreover,
they occupy the decidual space (and thus are
extravillous trophoblasts) (Fig. 1), where they
remodel the maternal spiral arteries, increasing
the blood supply to the fetus. To evade recogni-
tion as “foreign” by maternal immune cells, ex-
travillous trophoblasts express tolerogenic HLAs
— HLA-G, HLA-E, and HLA-C. The expression
of HLA-G by extravillous trophoblasts is critical
to their ability to “educate” decidual NK cells to
promote tolerance; this is achieved through fila-
mentous projections that serve as synapses be-
tween the trophoblasts and the decidual NK
cells.
3
However, parasites, viruses, and the bac-
terium Listeria monocytogenes can subvert this
mechanism in order to infect the placenta and
fetus.
4
Thus, tolerance of extravillous tropho-
blasts by decidual NK cells permits infection of
the fetus by L. monocytogenes, resulting in fetal
loss. Fortunately, the cytotoxic activity of decid-
ual NK cells is often sufficient to kill L. monocy-
togenes. However, until now, the mechanism by
which decidual NK cells are able to kill the
bacterium without damaging placental cells was
unclear.
Crespo et al. showed that decidual NK cells
selectively kill L. monocytogenes in extravillous
trophoblasts, without killing these cells, by in-
jecting the protein granulysin into the tropho-
blasts through nanotubes (Fig. 1). The authors
first showed that supernatants from decidual
NK cells could kill intracellular L. monocytogenes
in cultures of both a trophoblast cell line and
primary extravillous trophoblasts. Treating de-
cidual NK cells with an antigranulysin antibody
Figure 1 (facing page). Protection of the Fetus by Decidual NK Cells.
The maternal–fetal interface is made up of the maternal decidua basalis and the fetally derived placenta. The decidua harbors a large
population of maternal immune cells, including decidual natural killer (NK) cells, macrophages, T cells, and dendritic cells. The placenta
is made up of trophoblasts derived from the trophoectoderm layer, which surrounds the embryo blastocyst and is entirely fetal in origin.
The trophectoderm gives rise to cytotrophoblast cells, which fuse to create multinucleated syncytiotrophoblasts; these cells mediate nu-
trient and gas exchange between the maternal and the fetal circulation. Cytotrophoblasts also make up the anchoring villi mediating the
embryo–uterine attachment; toward the end of the first trimester, cytotrophoblasts extravasate into spiral arteries and enter the decidual
space as extravillous trophoblasts. To invade the decidua while evading maternal immune surveillance, fetal extravillous trophoblasts ex-
press tolerogenic HLAs — HLA-G, HLA-E, and HLA-C. Crespo et al.
2
recently reported that decidual NK cells selectively kill Listeria mono-
cytogenes within extravillous trophoblasts without killing the cells themselves by injecting the protein granulysin, through actin-rich nano-
tubes, into the trophoblasts. Granulysin permeabilizes cholesterol-poor bacterial membranes and can thus induce microbial programmed
cell death, but it does not affect host-cell plasma membranes.
The New England Journal of Medicine
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Copyright © 2020 Massachusetts Medical Society. All rights reserved.
Clinical Implications of Basic Research
n engl j med 383;21 nejm.org November 19, 2020
2081
Placenta
Uterus
Amniotic
cavity
Umbilical
cord
Tolerogenic
HLA molecules
Decidual NK cell
Extravillous
trophoblast
Mitochondrion
reticulum
L. monocytogenes
L. monocytogenes
Actin-rich
nanotube
Cytotoxic granules
remain immobilized
owing to tolerogenic
effect of HLA expressed
by trophoblast
Fetus
UTERINE WALL
UTERINE WALL
UTERINE WALL
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Uterine spiral artery
Maternal
artery
Villous placenta
(fetal blood)
Intervillous space
(maternal blood)
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
UTERINE WALL
Maternal immune cells
Decidual NK cell
Extravillous trophoblasts
Granulysin permeabilizes cholesterol-poor
phagosome and bacterial membranes,
causing bacterial death without affecting
host-cell plasma membrane
Syncytiotrophoblast
The New England Journal of Medicine
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Copyright © 2020 Massachusetts Medical Society. All rights reserved.
Clinical Implications of Basic Research
n engl j med 383;21 nejm.org November 19, 2020
2082
or blocking their ability to secrete proteins pre-
vented bacterial killing. However, bacterial kill-
ing did not require the decidual NK cells to re-
lease cytotoxic granules. Although decidual NK
cells contain both granulysin and perforin (which
makes pores in target cells) within the granules,
they also contain cytosolic granulysin. Crespo et al.
showed that decidual NK cells kill L. monocyto-
genes in extravillous trophoblasts by directly
transferring cytosolic granulysin to the tropho-
blasts through actin-containing nanotubes. Treat-
ing the infected cells with neuraminidase, a siali-
dase that cleaves glycosidic linkages, limited
bacterial killing, which suggests that the recog-
nition of sialylated moieties on extravillous tro-
phoblasts triggers nanotube formation.
Crespo et al. showed that peripheral NK cells
could also transfer granulysin to macrophages
and dendritic cells without killing these cells.
Thus, the authors may have uncovered a general
mechanism by which NK cells selectively kill
pathogens. Although previous work had shown
that granulysin more effectively kills bacteria in
the presence of perforin or granzyme, Crespo
et al. showed that granulysin can kill L. monocyto-
genes on its own — which could be considered a
“stealth move” that destroys invading bacteria
without raising alarm.
The authors then went a step further and
asked whether this new mechanism occurs in
vivo. To address this possibility, they used mice
engineered to express human granulysin. (Al-
though most mammals have GNLY, the gene en-
coding granulysin, rodents do not.) Lieberman
and colleagues have previously shown that GNLY-
transgenic mice were able to clear intracellular
bacterial (L. monocytogenes) infections more effi-
ciently than control mice.
5
In the current study,
Crespo et al. found that pregnant GNLY-transgenic
mice were less susceptible to infection than con-
trol mice. Moreover, depletion of uterine NK cells
(the murine counterparts of decidual NK cells) led
to the resorption of fetuses in L. monocytogenes–
infected GNLY-transgenic mice. However, because
mice lack GNLY and several other genes that
mediate NK function in humans, caution is war-
ranted in considering the implications of these
data for understanding protection against bacte-
rial infection in human pregnancy.
Granulysin belongs to a family of proteins
that efficiently permeabilize cholesterol-poor bac-
terial membranes but not cholesterol-rich mam-
malian membranes (Fig. 1). In classical NK ac-
tion, granulysin and another cytotoxic molecule,
granzyme, are delivered to cells through the
action of perforin, which damages mammalian
membranes. However, granulysin itself is a po-
tent, multipronged weapon: it induces microp-
tosis — microbial programmed cell death — by
blocking microbial antioxidant defenses and
biosynthetic and metabolic pathways required
for bacterial survival. The mechanism by which
granulysin selectively cleaves the enzymes that
are essential for microbial survival is unclear,
but this ability may be evolutionarily conserved:
granulysin causes substantial mitochondrial
damage, probably because mitochondrial mem-
branes resemble bacterial membranes. Granuly-
sin can also attack the membranes of other or-
ganelles with low cholesterol content (e.g.,
endoplasmic reticulum) (Fig. 1).
Crespo and colleagues have elucidated a new
mechanism by which decidual NK cells protect
the developing fetus from infection without
damaging the placenta. Further work to delin-
eate the extent to which this mechanism is at
play in protecting human pregnancies could lead
to new strategies to treat infections during preg-
nancy. Moreover, because peripheral NK cells
and some cytotoxic T cells appear to share with
decidual NK cells the ability to secrete granu-
lysin, it will be exciting to see whether this
nanotube-mediated mechanism functions in other
tissues to kill pathogens while limiting tissue
damage.
Disclosure forms provided by the author are available at
NEJM.org.
From the Department of Obstetrics and Gynecology and the
Department of Pathology and Immunology, Washington Uni-
versity in St. Louis School of Medicine, St. Louis.
1. Rackaityte E, Halkias J. Mechanisms of fetal T cell tolerance
and immune regulation. Front Immunol 2020; 11: 588.
2. Crespo ÂC, Mulik S, Dotiwala F, et al. Decidual NK cells
transfer granulysin to selectively kill bacteria in trophoblasts.
Cell 2020; 182(5): 1125-1139.e18.
3. Tilburgs T, Crespo ÂC, van der Zwan A, et al. Human HLA-G+
extravillous trophoblasts: immune-activating cells that interact
with decidual leukocytes. Proc Natl Acad Sci USA 2015; 112: 7219-
24.
4. Cao B, Mysorekar IU. Intracellular bacteria in placental basal
plate localize to extravillous trophoblasts. Placenta 2014; 35: 139-
42.
5. Dotiwala F, Lieberman J. Granulysin: killer lymphocyte safe-
guard against microbes. Curr Opin Immunol 2019; 60: 19-29.
DOI: 10.1056/NEJMcibr2028357
Copyright © 2020 Massachusetts Medical Society.
The New England Journal of Medicine
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Copyright © 2020 Massachusetts Medical Society. All rights reserved.
