HLA testing involves analyzing proteins expressed on cell surfaces that regulate the immune system. These HLA proteins are encoded by genes inherited from each parent. HLA sharing between the parents helps establish a successful pregnancy by causing the mother's immune system to produce protective cytokines that support fetal development. Some HLA alleles have been associated with recurrent miscarriages, and less sharing of these alleles between partners can increase the risk of pregnancy loss.

1. HLA Testing
MR. ALI A.RADWAN
M.P.H

2. What is HLA?
 HLA stands for "Human Leucocyte Antigen". They are a group of proteins which are
expressed on the surface of almost all the cells in our body. These cell-surface proteins
are responsible for the regulation of the immune system in humans. HLA is also called as
Major Histocompatibility Complex (MHC). The genes (stretches of DNA) which encode
for HLA proteins are present in chromosome 6 (we have 46 chromosomes- 23 pairs !).
Each person receives one set of HLA genes from each parent; we inherit half of our gene
from the father and half from the mother.

3. Cont…
 HLA genes are classified into class I, class II and class III. Class I genes encode for
HLA- A, HLA- B and HLAC proteins. These proteins are present in most cells of our
body. Class I region also contains genes which give rise to HLA-E, HLA-F, HLA-G
proteins. Class II genes encode HLA-DP, HLA-DQ and HLA-DR proteins. These
proteins are present mainly in our immune system cells like B-lymphocytes,
macrophages, Tlymphocytes etc. Class III genes encode components of our
complement system (a group of proteins which function in our immune system).

4. The mother’s immune system during pregnancy and its
influence on fetal development:
 The maternal immune system plays a critical role in the establishment, maintenance, and
completion of a healthy pregnancy. Various cells and molecules of the immune system are key
players in the development and function of the placenta and the fetus. Immune cells
accumulating in the human endometrium at the time of decidualization play critical and diverse
roles at the maternal–fetal interface, including functions in implantation, placental development,
and immunity against infectious diseases. Of all decidual leukocyte populations, the most
abundant are the phenotypically unique uterine natural killer (uNK) cells.

5. Cont …
 These cells dramatically increase in number in the human endometrium 3–5 days
postovulation, accounting for 25%–40% of endometrial leukocytes prior to implantation and
accounting for ~70% of decidual leukocytes in the first trimester. In addition to uNK cells,
decidual macrophages are relatively abundant, comprising ~20% of the human decidual
leukocyte population in the first trimester. T-cells are also fairly abundant in human decidua,
comprising ~10%–20% of the human decidual leukocyte population. The main function of T-
cells in the decidua, particularly of CD4+ T-regulatory (Treg) cells, is to promote tolerance to
the fetus.

6. uNK cells regulate key early events in establishment of pregnancy:
implantation, angiogenesis, and vascular remodeling:
 1. Trophoblast Invasion: One of the earliest events of establishing a successful
pregnancy is the trophoblast invasion into maternal decidua. Several studies have
demonstrated that uNK cells play a critical rule in the trophoblas migration and
invasion into the decidua and found that uNK cells express chemokines IL-8 and
IFN-inducible protein (IP)-10.

7. Cont …
 2. Angiogenesis and vascular remodeling in early pregnancy: In humans, extensive vascular
remodeling must occur to allow for placentation and establishment of early pregnancy, as
well as to support the demands of a growing fetus. The decidual spiral arteries must be
transformed into larger-diameter vessels with low resistance and high flow, capable of
transporting nutrients and oxygen to the fetus. In addition, the endothelium of these
vessels is replaced by extravillous trophoblast cells that have migrated from the placenta,
allowing for diversion of blood flow into the space surrounding the placental villous tree
and thereby permitting nutrient and gas exchange between mother and fetus. Not only is
adequate vascular remodeling critical for the establishment of a normal pregnancy, but
abnormalities in these early events are associated with later complications of pregnancy
such as preeclampsia and intrauterine growth restriction, which can have a major impact
on fetal and neonatal health.

8. How HLA sharing (incompatibility) helps in establishing a
healthy and successful pregnancy?
 The "immunotrophic theory" explains the paradox of how the maternal immune system accepts a fetus which is
not genetically identical to hers. The theory states that recognition of the foreign antigen ( the paternal HLA) by
the mother's immune system is " good " for the fetus , because it causes the maternal immune system to
produce protective cytokines ( such as G-CSF IL-8,IFN-IP-10, IFN-γ ),which enhance implantation, promote the
growth of trophoblastic cells and help in sustaining pregnancy. Reproductive immunologists suggest that
recognition of foreign fetal antigens (mainly paternal HLA) by the mother's immune system is necessary for
eliciting a "protective" immune response ( such as the production of protective "blocking antibodies" , which
protect the fetus from immune rejection), and this actually helps in the survival and development of fetus.

9. HLA class II alleles associated with recurrent miscarriages:
 Old and recent investigations have been conducted in many countries involving several
population with different ethnic background in order to demonstrate the contribution
of some HLA alleles in pregnancy loss. These studies could identify some HLA alleles
that are described to be linked to recurrent abortion. HLA matching between the father
and the mother in terms of these HLA alleles are known to be prevalent among women
with a history of three or more recurrent pregnancy loses. The table shows the HLA
alleles that have been found prevalent among women with frequent miscarriages.

10. References:
 1. Morelli S, Mandal M, Goldsmith LT, Kashani BN, Ponzio NM. (2015). The maternal immune system during pregnancy and its influence on fetal development, DovePress.
Volume 2015:6 Pages 171—189. DOI https://doi.org/10.2147/RRB.S80652
 2. Ober C.(1999). Studies of HLA, fertility and mate choice in a human isolate. Mar-Apr;5(2):103-7.
 3. Jin, K., Ho, H. N., Speed, T. P., & Gill, T. J. (1995). Reproductive failure and the major histocompatibility complex. American Journal of Human Genetics, 56(6), 1456–1467.
 4. Ho HN1, Yang YS, Hsieh RP, Lin HR, Chen SU, Chen HF, Huang SC, Lee TY, Gill TJ 3rd. (1994). Sharing of human leukocyte antigens in couples with unexplained infertility
affects the success of in vitro fertilization and tubal embryo transfer. Am J Obstet Gynecol .170(1 Pt 1):63-71.
 5. Koyama, M., Saji, F., Takahashi, S., Takemura, M., Samejima, Y., Kameda, T., Kimura, T., Tanizawa, O. and Koyama, M. (1991), Probabilistic assessment of the HLA sharing of recurrent DRB1*01
DQA1*0101–*0104–*0105–*0107 DQB1*0501 DRB1*15 (02) DQA1*0102-0103/*0103 DQB1*0602-0602/*0601 DRB1*16 (02) DQA1*0102 DQB1*0502/*0504/*0505 DRB1*03 DQA1*05 DQB1*02
DRB1*04 DQA1*03/*04 DQB1*02/*03/*04 DRB1*11 (05) DQA1*0501 DQB1*0301 DRB1*12 (05) DQA1*0501/0601 DQB1*0602-0620/*0301 DRB1*13 (06) DQA1*0102-*0103/*0501
DQB1*0503/*0301 DRB1*14 (06) DQA1*0101/*0501 DQB1*0201/*0303 DRB1*07 DQA1*0201 DQB1*03/*04 DRB1*08 DQA1*04/*06 DQB1*0303 DRB1*09 DQA1*03 DQB1*0501 DRB1*10
DQA1*0101 spontaneous abortion couples in the Japanese population. Tissue Antigens, 37: 211–217. doi:10.1111/j.1399-0039.1991.tb01874.x.
 6. Meuleman T, Lashley LE, Dekkers OM, van Lith JM, Claas FH, Bloemenkamp KW
 7. HLA associations and HLA sharing in recurrent miscarriage: A systematic review and metaanalysis. Hum Immunol. 2015 May;76(5):362-73.
 8. Gharesi-Fard B , Askarinejad-Behbahani R, Behdin S. The effect of HLA-DRB1 sharing between the couples with recurrent pregnancy loss on the pregnancy outcome after
leukocyte therapy. Iran J Immunol. 2014 Mar;11(1):13-20. doi: IJIv11i1A2.
 9. D'Ippolito S, Gasbarrini A, Castellani R, Rocchetti S, Sisti LG, Scambia G, Di Simone N. Human leukocyte antigen (HLA) DQ2/DQ8 prevalence in recurrent pregnancy loss
women. Autoimmun Rev.2016 Jul;15(7):638-43. doi: 10.1016/j.autrev.2016.02.009. Epub 2016 Feb 13.
 10. Kruse C,Steffensen R,Varming K, Christiansen OB. A study of HLA-DR and -DQ alleles in 588 patients and 562 controls confirms that HLA-DRB1*03 is associated with
recurrent miscarriage. Hum Reprod.2004 May;19(5):1215-21.