Elder K. Human preimplantation embryo selection

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HUMAN PREIMPLANTATION EMBRYO SELECTION

sHLA-G measurement on assisted reproductive

technology (ART) outcomes.

CHARACTERIZATION OF HUMAN

LEUKOCYTE ANTIGEN G

HLA-G was first cloned in 1987,

but not until 1990

was the atypical HLA class I molecule present in

abundant amounts at the maternal–fetal interface

determined to be HLA-G.

Alternative splicing of

the HLA-G mRNA yields different membrane-

bound and soluble variants of the HLA-G protein

and a limited number of variable sites in the DNA

sequence of the HLA-G gene have been reported.

9,10

Because HLA-G has a low level of nucleotide

sequence polymorphisms, it has been postulated as

a prerequisite for maintenance of maternal immune

tolerance. The sHLA-G molecule is strongly

expressed during the first trimester of gestation and

then decreases through the remainder of pregnancy,

suggesting a potential role in implantation and

immunoprotection of the developing embryo.

An association has been reported between

HLA-G expression in human embryos and other

factors known to influence IVF outcome. Using

non-transferred embryos at the 2–4-cell stage,

Jurisicova et al

found sibling embryos from suc-

cessful cycles were more likely to express HLA-G

than were those from non-successful cycles. Cyto-

toxicity assays determined that the primary HLA-G

gene transcript is alternatively spliced into five

main mRNA forms: HLA-G1 (full length), HLA-

G2 (minus exon 3) which codes for a membrane-

bound isoform associated with B2 microglobulin,

HLA-G3 (minus exons 3 and 4), HLA-G4 (minus

exon 4), and HLA-G5 (plus intron 4) which codes

for a soluble form of the HLA-G antigen. All HLA-G

isoforms are capable of inhibiting natural killer

(NK) cell activity. These findings suggest that

HLA-G is the public ligand for NK cell inhibitory

receptors.



HLA class Ia

HLA-G1

HLA-G2

HLA-G3

HLA-G4

L α1 α2 α3 int. 4 Tm C 3⬘UT

sHLA-G1

L α1 α3CTmint. 4 3⬘UT

sHLA-G2

α1 α2TmC3⬘UT

α1TmC3⬘UT

α3α1TmC3⬘UT

α2 α3α1TmC

3⬘UT

α2 α3α1TmC

3⬘UT

Figure 12.1 Schematic diagram of HLA class Ia and major HLA-G transcriptional isoforms. HLA-G1, -G2, -G3, and -G4 isoforms

encode potentially membrane-bound proteins, whereas sHLA-G1 and sHLA-G2 encode soluble forms. L, exon 1 encoding the leader

sequence; ␣1, ␣2, ␣3, exons 2, 3, and 4 encoding the ␣1, ␣2, ␣3 external domains, respectively; Tm, exon 5 encoding the

transmembrane domain; C, exons 6 and 7 encoding the cytoplasmic region; 3⬘UT: exon 8 encoding the 3⬘ untranscribed region. The

hexagon indicates the presence of ‘stop codons.’ Modified from Le Bouteiller et al.

HPE_Chapter12.qxp 7/13/2007 4:51 PM Page 146

ASSESSMENT OF SOLUBLE HUMAN LEUKOCYTE ANTIGEN G IN HUMAN EMBRYOS

Immunohistochemical expression of classical

and non-classical HLA class I proteins in human

placentas using monoclonal antibodies demon-

strated classical HLA class I proteins in all non-

trophoblast cells, including fetal and maternal

cells.

Syncytiotrophoblast did not show any HLA

class I staining, but extravillous cells expressed high

amounts of HLA-G.

The HLA-G gene message is alternatively spliced

into multiple transcripts, three of which encode

soluble isoforms. Morales et al

produced soluble

recombinant (r)HLA-G1, rHLA-G2 in human

embryonic kidney 293 cells and characterized the

proteins. Both isoforms were glycosylated and

formed disulfide-bonded oligomers. The rG1 anti-

body associated with B2 microglobulin, while rG2

did not. Mouse monoclonal antibodies generated to

rsG1 (specific to sG1) and to rsG2 (identifies both

soluble and membrane-bound G2) were used for

immunohistochemical isoform mapping studies on

placental tissue sections.

Soluble G1 was abundant in many subpopulations

of trophoblast cells, while membrane (m)/sG2

protein was present exclusively in extravillous

cytotrophoblast cells. Immunostaining localized

m/sG2 only to the invasive trophoblast subpopula-

tion. Analysis of function by Northern and Western

blotting demonstrated both rsG1 and rsG2 inhibit

CD8 alpha expression on peripheral blood mono-

nuclear cells (PBMC) without changing CD3 delta

expression or causing apoptotic cell death.

Collectively, these studies indicate that both sG1

and sG2 are produced in placentae and that tran-

scription and translation are linked for sG1, but not

for sG2. While expression of sG2 is exclusively associ-

ated with the invasive phenotype, the two isoforms of

sG may promote semiallogenic pregnancy by reduc-

ing expression of CD8, a molecule required for func-

tional activation of cytotoxic T lymphocytes (CTL).

The HLA-G molecule has unique properties of

low polymorphism and restricted tissue distribution

mainly to the extravillous cytotrophoblast (EVT).

These EVT cells vigorously penetrate into maternal

decidual tissues and are found in contact with mater-

nal lymphocytes, mainly with NK cells. The HLA-G

molecule inhibits effector function of maternal

NK cells via interaction with KIR2DL4 and ILT-2

inhibitory NK receptors. Efficient binding and func-

tion of ILT-2 is dependent on the presence of HLA-G

complexes on the cell surface. Their presence on the

cell surface suggests these complexes may also be

present in a soluble form.

This may be a particular

mechanism to increase the avidity of NK receptors

to HLA-G, resulting in better protection of the fetus

from maternal NK cell attack (Figure 12.2).

Soluble class Ib HLA-G glycoproteins synthe-

sized in the placenta are abundant in the pregnant

uterus and circulate in the maternal blood through-

out pregnancy. Hunt et al

established the immuno-

genicity of these proteins by testing sera of 64

women with at least one successful pregnancy, 21

women who had never been pregnant, and 54 males

for antibodies to epitopes present on recombinant

sHLA-G isoforms (G1/G2) derived from HLA 6.0

cDNA (HLA-G*0101 allele). Indirect enzyme linked

immunosorbent assay (ELISA) identified antibodies

to sHLA-G isoforms in six sera, all from multigravid

women, while all other sera were negative. These

results indicate that tolerance to HLA-G is the usual

condition, as antibodies to HLA-G were not detected

in 91% (58/64) of multigravid women. Pregnancy

stimulated the loss of tolerance in only 9% (6/64).

However, all six women delivered healthy babies,

demonstrating that antibodies to HLA-G do not

prevent successful pregnancy outcome.

Both membrane-bound and soluble HLA-G

isoforms are proposed to influence the outcome of

pregnancy. Aberrant HLA-G expression is reported

in pre-eclampsia and spontaneous abortion. Placental

hypoxia following the immature remodeling of spiral

arteries by extravillous cytotrophoblasts (CTs) has

been investigated as a pathogenic mechanism for the

development of pre-eclampsia.

18,19

HLA-G expres-

sion is also decreased at the protein and mRNA levels

in pre-eclamptic placenta.

HUMAN LEUKOCYTE ANTIGEN G AND

IMPLANTATION

Pregnancy is an immune balancing act between mater-

nal tolerance of paternal major histocompatabilty



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HUMAN PREIMPLANTATION EMBRYO SELECTION

antigens and maintaining normal immune compe-

tence. The placenta separates maternal and fetal

blood/lymphatic systems. Fetal trophoblast plays a

major role in evading recognition by the maternal

immune system. Trophoblast cells fail to express

MHC class I or II molecules and extravillous

cytotrophoblasts strongly express non-classical

MHC genes encoding HLA-G, which appear to down

regulate NK cell activity. Trophoblast expresses

Fas ligand, conferring immune privilege: maternal

immune cells expressing Fas will undergo apoptosis

at the placental–decidual interface

(Figure 12.3).

Natural killer cells in the decidua may control

trophoblast migration during implantation through

recognition of HLA-G/HLA-C by killer inhibitor/

activator receptors (KIR/KAR). Monoclonal anti-

bodies demonstrated HLA-G in extravillous tro-

phoblast expressed in both B2 microglobulin

associated form and as free heavy chains.

Lack of

classical HLA molecules in the human placenta

prevents recognition and lysis by maternal T

lymphocytes, but leaves the conceptus susceptible to

NK cell-mediated lysis. NK cells are known to express

a number of HLA-class I-specific inhibitory recep-

tors, including members of the Ig superfamily (p58,

p70, p140), characterized by a defined allele speci-

ficity and CD94/NKG2A with broad specificity for

HLA class I molecules. Analysis of NK cell clones

expressing CD94/NKG2A as identified by Z199

monoclonal antibodies, displayed markedly reduced

cytolytic activity against 221/G. Monoclonal anti-

bodies against CD94/NKG2A completely restored

target cell lysis.

Receptors mediating NK cell inhi-

bition were evaluated using monoclonal antibodies

against known NK inhibitory receptors: CD158a,

CD158b, and CD94.

In at least one-third of NK

cells inhibited by HLA-G, these antibodies alone or

in combination failed to reverse inhibition, suggest-

ing the presence of a third major unidentified

receptor for HLA-G.



Cytosol

Peptide

Golgi

β2m

-G1 -G2 -G3 -G4

HLA-G1

sHLA-G1 sHLA-G2

α1

α1α1

α1

α2

α3 α3 α3

α1

α1 α1

α2

α3

α2α1

α3 α3

β2m

Figure 12.2 Schematic of sHLA-G secretion from cytotrophoblast cells. Only the full length HLA-G1 isoform is expressed at the cell

surface whereas the HLA-G2, -G3, and -G4 forms are sequestered in the endoplasmic reticulum (ER). Both soluble HLA-G1 and -G2

isoforms can be secreted. Adapted from Le Bouteiller et al.

HPE_Chapter12.qxp 7/13/2007 4:51 PM Page 148

Class I negative syncytiotrophoblast escapes NK

lysis by maternal peripheral blood leukocytes (PBL),

while HLA-G expressing transfectants of LCL.722.221

cells are protected from lymphokine-activated killer

lysis. Extravillous cytotrophoblast cells and HLA-G

expressing choriocarcinoma cells (CC) are not pro-

tected. Avril et al

showed both JAR (HLA class I

negative) and JEG-3 (HLA-G and HLA-Cw4 posi-

tive) cells were resistant to NK cell lysis by PBL and

were equally lyzed by interleukin (IL)-2 stimulated

PBL isolated from a given donor. Down regulating

HLA-class I on JEG-3 cells by acid treatment did not

affect NK or lymphokine activated killer cell (LAK)

lysis of CC.

Soluble HLA-G produced at the maternofetal

interface by cytotrophoblasts and circulating in the

body fluids shows a capacity analogous to membrane-

bound structures to inhibit NK cells. Fuzzi et al

used a specific ELISA to investigate the presence of

sHLA-G molecules in culture supernatant of early

embryos obtained by IVF. Analysis of 285 super-

natants derived from embryos cultured in groups,

corresponding to 101 ART procedures (43 IVF/58

ICSI) identified two groups of patients based on

sHLA-G expression. No differences in clinical para-

meters were identified, but implantation occurred only

in women showing sHLA-G molecules in the culture

supernatants. They conclude that HLA-G expression

is a mandatory, but not sufficient requirement on its

own, for the development of successful pregnancy.

During pregnancy the fetus represents semi-

allograft to the mother. The relevance of sHLA-G

levels in maternal circulation in relation to the occur-

rence of characteristic pregnancy disorders were

evaluated by measuring plasma sHLA-G levels in

women with normal and pathological pregnancies.

Compared with normal pregnancy, significantly

increased sHLA-G levels were detected in women

ASSESSMENT OF SOLUBLE HUMAN LEUKOCYTE ANTIGEN G IN HUMAN EMBRYOS



Activated

T cell CD8+

β2m

α3

α1

α2

Maternal blood/decidua

Fas

Fas-L

sHLA-G1

Villous syncytiotrophoblast

Extravillous cytotrophoblast

Lysis

Apoptosis

CD8

Figure 12.3 Role of soluble HLA-G1 in induction of specific apoptosis of activated CD8⫹ T cells through CD8 interaction. Such

function is Fas/Fas ligand mediated. In vivo, soluble HLA-G1 may be produced by syncytiotrophoblast and, possibly, by extravillous

cytotrophoblast, inducing apoptosis of activated CD8⫹ T cells present in maternal blood of the intervillous space and/or in the

decidua. Modified from Solier et al.

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HUMAN PREIMPLANTATION EMBRYO SELECTION

delivered preterm due to intrauterine activation

(uncontrollable labor, rupture of fetal membranes,

cervical insufficiency, and hemolysis, elevated liver

enzymes, low platelets (HELLP) syndrome). Soluble

HLA-G levels in women with placental abruption

were more than three times lower than in normal

pregnancies. Non-parametric discriminant analysis

showed women with sHLA-G levels below 9.95 ng/mL

had a 7.1 relative risk for development of placental

abruption. These findings suggest that the occur-

rence of pregnancy-associated disease is strongly

influenced by maternal sHLA-G plasma levels.

Pre-eclampsia, a common disorder of first preg-

nancies, is thought to result from poor placentation

and may reflect abnormal maternal immune reac-

tion to the hemiallogenic fetus. HLA-G from fetal

derived extravillous cytotrophoblasts may protect

from maternal–fetal immune intolerance and allow

these cells to invade the uterus. In normal placenta,

HLA-G was shown to be expressed in the anchoring

extravillous trophoblasts with an increasing gradi-

ent of expression in the more invasive cells, while

in 9/10 pre-eclamptic placentae HLA-G expression

was absent or reduced.

Trophoblasts lacking

HLA-G may be vulnerable to attack by the maternal

immune system.

A subset of recurrent spontaneous abortions

may involve immunological mechanisms. Aberrant

T helper 1 (CD4⫹)/T helper 2 (CD8⫹) cytokine

profiles have been observed, which are not present

in uncomplicated pregnancies. Studies of classical

HLA class I and II antigens in relation to recurrent

spontaneous abortion are inconclusive. HLA-G is

expressed on invasive cytotrophoblasts and exists in

both membrane-bound and soluble forms. However,

the hypothesis of histoincompatability between pla-

centa and mother does not appear to be supported

by the data, although an altered expression profile of

HLA-G isoforms or reduced expression of certain

isoforms is still possible.

Coordinated communication between trophoblast

cells and their decidual environment is important

during the implantation process response. Numerous

studies have shown that MHC proteins play an

important role in reproduction and development by

down regulating the maternal immune response.

Initially a great deal of controversy surrounded the

issue of whether or not class I MHC antigens are

expressed on preimplantation embryos. Class I MHC

antigens have been demonstrated on preimplantation

mouse embryos through a variety of techniques,

including electron microscopy,

31,32 125

I-lactoperoxi-

dase labeling,

complement-dependent cytotoxi-

city,

cell mediated cytotoxicity assays,

and a

specific ELISA procedure.

36,37

Using RT-PCR, mRNA

for class I MHC antigens has been detected in

preimplantation mouse embryos.

A comparison of paternally and maternally

derived transcripts has shown that active transcrip-

tion of the class I MHC genes begins at the 2-cell

stage of development.

HLA-G expression has been

demonstrated during early preimplantation stages

of human development and expression of HLA-G

can be correlated with the embryo cleavage rate.

Transcripts of mRNA from both the maternal and

paternal haplotypes were demonstrated in embryos

from the 1-cell zygote to the late blastocyst stage of

development in congenic strains of mice.

These

data clearly show that both paternally and maternally

inherited MHC class I genes are transcribed from

the earliest stages of embryonic development, and

suggest that developmental regulation of expression

of their protein products is principally at the post-

transcriptional level. HLA-G is reputed to prevent

allorecognition by maternal cytotoxic lymphocytes

and to protect against NK cell mediated lysis of tar-

get cells.

In fact, HLA-G expression on extravillous

cytotrophoblast cells probably plays a compulsory

role in the development of pregnancy.

Soluble HLA-G has been identified in the culture

medium of early embryos cultured in groups. Its

concentration in the media surrounding groups of

embryos has been found to correlate with the rate of

embryo cleavage and clinical pregnancy potential.

26,42

Direct evidence supporting the role of HLA-G in

protecting cytotrophoblasts against NK cytolysis

under physiological conditions has been reported

by Rouas-Freiss et al.

In six semi-allogenic combi-

nations of maternal uterine NK cells and their own

trophoblast counterparts, as well as in 20 allogenic

combinations of maternal uterine NK cells and

trophoblasts from different mothers, they showed



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ASSESSMENT OF SOLUBLE HUMAN LEUKOCYTE ANTIGEN G IN HUMAN EMBRYOS

HLA-G-mediated protection was abolished by

treatment of cytotrophoblasts with HLA-G-specific

monoclonal antibodies. An HLA class I negative

K562 cell line transfected with the predominant

HLA-G1 isoform resulted in similar protection

from maternal uterine NK cell lysis.

The immune tolerance inducing effects of sHLA-

G1 on NK cells and T cells were investigated by Fang

et al.

A recombinant plasmid expressing sHLA-G1

was transfected into human lymphoblastoid cells.

HLA-G1 dose-dependently inhibited NK cell cyto-

toxicity as well as dose-dependently inhibited pro-

liferation of activated T cells and induced T cell

apoptosis with a dose-saturation character and

without antigen specificity.

The relationship between sHLA-G expression in

trophoblast cells and early embryo development

was studied by Wang et al

who evaluated tro-

phoblasts from 14 cases of spontaneous abortion

after IVF/ET and six cases of choriocarcinoma,

compared with ten cases of trophoblast cells from

normal placenta. In the spontaneous abortion group

only 1/14 had positive sHLA-G expression, while all

cases of choriocarcinoma had strong sHLA-G expres-

sion. All cases of normal villous were also positive

for sHLA-G expression. The expression level of

sHLA-G decreased in the spontaneous abortion

group compared with the normal placenta group. In

individual cells, staining on choriocarcinoma cells

was stronger than on normal villous cells. High

expression in normal villi and low expression in tro-

phoblast cells from spontaneous abortions, suggests

that sHLA-G from trophoblast cells does have a

relationship to early embryo development.

Differences in sHLA-G protein levels were ana-

lyzed in women with uncomplicated compared with

pre-eclamptic pregnancies. Both serum and placen-

tal HLA-G levels decreased significantly in the pre-

eclamptic group compared with the normal

pregnant women. High correlation was found

between serum and placental HLA-G levels (r ⫽

0.6). Reduced expression of placental HLA-G and

reduced release of protein into the maternal circula-

tion in pre-eclampsia may alter the maternal–fetal

immune relationship and thus be involved in the

cause of the disorder.

HUMAN LEUKOCYTE ANTIGEN G AND

ASSISTED REPRODUCTIVE TECHNOLOGIES

The role of sHLA-G in ART has only recently been

evaluated. Sher et al

report data on a retrospective

cohort of 594 embryos transferred into 201 women

aged 28–44 years. IVF/ICSI was performed with

embryo transfer (ET) of at least two 7–10-cell

embryos 72 hours after retrieval. Media from indi-

vidually cultured embryos was collected 46 hours

postICSI. Group A consisted of 159 women aged

⬍39 years. Group A1 consisted of 101 women with

at least one embryo with sHLA-G⫹ (within one SD

of the geometric mean), while group A2 consisted of

58 women with all transferred embryos below range.

Group B consisted of 42 women aged 39–44 years.

Group B1 consisted of 29 women with at least one

embryo with sHLA-G⫹, while group B2 consisted

of 13 women with all transferred embryos below

range.

The clinical pregnancy and implantation rates in

group A1 were 71% (72/101) and 38%, respectively,

compared with those in group A2 which were 22%

(13/58) and 9%, respectively. The clinical pregnancy

and implantation rates for group B1 were 52%

(15/29) and 25%, respectively, compared with those

in group B2 which were 15% (2/13) and 5%, respec-

tively. These data suggest that selecting embryos for

transfer based on individual sHLA-G expression

can maximize pregnancy rates from the transfer of

fewer embryos, which can help reduce the incidence

of high-order multiple gestation.

The same group reported the results of a pros-

pective evaluation of 107 women aged ⬍39 years,

with normal ovarian reserve, normal uterine cavity,

and two embryos transferred on day 3 scoring

GESⱖ70.

In group A (n ⫽ 51) all embryos

expressed sHLA-G within one SD of the geometric

mean optical density 0.190 ⫾ 0.006. The pregnancy

and implantation rates were 75% (38/51) and 44%

(51 sacs/116 embryos), respectively. In group B

(n ⫽ 56) all embryos transferred were sHLA-G

negative (OD: ⬍0.184). The pregnancy and implan-

tation rates were 23% (13/56) and 14% (20/143),

respectively.



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HUMAN PREIMPLANTATION EMBRYO SELECTION

In a further prospective trial, Sher et al

evalu-

ated 482 women aged 28–43 years with transfer of

7–9-cell embryos. In women ⱕ38 years (n ⫽ 387),

when all transferred embryos were HLA-G-negative

(n ⫽ 150), the pregnancy and implantation rates

were 25% and 13%, respectively. When one trans-

ferred embryo was HLA-G-positive (n ⫽ 95), the

pregnancy and implantation rates were 63% and

32%, respectively. When all transferred embryos

were HLA-G-positive (n ⫽ 142), pregnancy and

implantation rates were 69% and 36%, respectively.

In women aged 39–43 years (n ⫽ 95), when all

transferred embryos were HLA-G-negative (n ⫽ 38),

the pregnancy and implantation rates were 29% and

11%, respectively. When one transferred embryo

was HLA-G-positive (n ⫽ 21), pregnancy and

implantation rates were 38% and 15%, respectively.

When all transferred embryos were HLA-G-positive

(n ⫽ 36), pregnancy and implantation rates were

61% and 26%, respectively. Using sHLA-G meas-

urement in culture media at 46 hours postICSI to

select embryos for transfer provided a mechanism

to lower the incidence of multiple gestations.

Noci et al

evaluated sHLA-G using an ELISA

employing monoclonal antibody MEM-G9 in 318

media of single embryo culture after 48 hours of

culture. They also correlated the presence of sHLA-G

with embryo morphology and pregnancy obtained

in that cycle. No correlation between sHLA-G and

embryo morphology was noted. Pregnancy was

observed only when one embryo was sHLA-G⫹

with a 1 ng/mL limit of sensitivity (mean 4 ng/mL);

26/66 patients had no sHLA-G expression and no

pregnancy occurred (although overall, pregnancy

occurred in only 9/66). Of seven patients with no

sHLA-G expression in the first cycle who did a

second cycle, 4/5 had sHLA-G⫹ embryos and all

four became pregnant.

Yie et al

report a retrospective evaluation of

media from 386 grouped embryo cultures assayed

for sHLA-G after 72 hours culture. Soluble HLA-G

was detected in 270 samples. Secretion was inde-

pendent of embryo grade or patient age. The cleavage

rate of embryos secreting sHLA-G was significantly

higher than for those lacking it (mean 6.7 vs 5.8 cells

on day 3). The live birth rate in embryos with

sHLA-G was higher than in those without (48% vs

17%). Combining secretion of HLA-G and cleavage

rate was most predictive of pregnancy. The authors

concluded that embryonic secretion of sHLA-G is

variable, sHLA-G secretion is associated with higher

cleavage and pregnancy rates, and sHLA-G is a

better independent predictor than cleavage alone.

Finally, the combination of sHLA-G detection and

high cleavage rate was best predictor of outcome.

Taken together these studies provide strong sup-

port to the notion that sHLA-G measurement in the

supernatant of cultured embryos can be used to

predict embryo viability. In summary, the accumu-

lated literature supports the following conclusions.

(1) HLA-G is a non-classical MHC protein expressed

on extravillous cytotrophoblasts, which appears

to modulate maternal immune function by

down regulating the actions of decidual NK

cells through interaction with the CD94 killer

inhibitory receptor.

(2) Both membrane-bound and soluble forms of

HLA-G appear to be immunoprotective for the

conceptus.

(3) Specific monoclonal antibodies have been

developed to detect HLA-G expression.

(4) Measurement of sHLA-G correlates with preg-

nancy outcome. Presence of sHLA-G in embryo

culture media predicts the outcome of IVF/ET.

Combining sHLA-G secretion with other estab-

lished embryo markers may be most clinically

useful.

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logically rejected by their mothers? Biol Reprod 1988; 38: 17–29.

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during preimplantation mouse embryo development. Presented at

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HUMAN PREIMPLANTATION EMBRYO SELECTION

48. Sher G, Keskintepe L, Fisch JD et al. Soluble human leukocyte antigen G

expression in phase I culture media at 46 hours after fertilization pre-

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Steril 2005a; 83: 1410–3.

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of developmental potential in embryos. Hum Reprod 2005; 20:

138–46.

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INTRODUCTION

Preimplantation embryos express paternal as well as

maternally derived proteins, yet they are not rejected

by the maternal immune system. This enigma was first

recognized by Medawar over 50 years ago

when he

wrote ‘. . . how does the pregnant mother contrive to

nourish within itself, for many weeks or months, the

foetus that is an antigenically foreign body? Is preg-

nancy accompanied by any physiological changes

which may in some degree prevent the foetus, qua tis-

sue homograph, from immunizing the mother against

itself?’ In spite of extensive research in this area, our

knowledge of how this comes about is still incomplete.

This review focuses on immunological aspects of

the preimplantation embryo and not on later stages of

development. The reason for this is that embryos are

relevant to clinics performing assisted reproductive

technologies (ART) such as in vitro fertilization (IVF)

and intracytoplasmic sperm injection (ICSI) only at

the preimplantation stage. There are many limitations

associated with research on human embryos, includ-

ing the scarcity of material, the outbred nature of the

human population making every embryo unique, as

well as lack of federal funding in the United States and

limited funding in other countries. Therefore, animal

models are required for extensive research on preim-

plantation embryos. The mouse is a particularly good

model system because of the availability of genetically

homogeneous inbred strains, congenic strains, knock-

out strains, and transgenic strains. In addition, mice

are relatively inexpensive compared with other animal

models such as primates or domestic animal species.

For these reasons research on the mouse model

system will be highlighted in this review.

The first objective of this review is to give the

reader a brief overview of the immune system,

including a discussion of innate and adaptive immu-

nity, cells and molecules involved in the immune

system, and the fundamentals of the major histo-

compatibility complex (MHC). A brief overview of

preimplantation development is then presented in

the context of MHC expression by preimplantation

embryos and considerations of the embryo as an

allograft. A discussion of the discovery of an MHC

encoded gene, the Ped (preimplantation embryo

development) gene, follows, along with a summary

of the properties of the Ped gene product, Qa-2 in

the mouse, and its homolog, HLA-G, in humans.

Both Qa-2 and HLA-G have a role in enhancing

embryo survival. Clinically relevant properties of the

mouse Ped gene, during both the preimplantation

period and later in life, are discussed. Concluding

remarks challenge the reader to think about how

new methods in nanomedicine might be applied to

preimplantation embryos in order to gain a further

understanding of the relevance of the immune

system to reproduction.

OVERVIEW OF THE IMMUNE SYSTEM

The major function of the immune system is to

protect an organism from damage induced by

exogenously introduced foreign pathogens such as

bacteria, viruses, and fungi, and from endogenously

produced cancer cells. This is accomplished by both

immune cells and their products, called cytokines,

as well as by specialized molecules such as antibod-

ies that recognize foreign substances, termed anti-

gens. For convenience, immunologists have divided

the immune system into two major categories, the

innate immune system and the adaptive immune

system. Several excellent textbooks are available that

13. Immunological aspects of

embryo development

Carol M Warner

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