THE ROLE OF SUBPOPULATIONS OF CD8 + T LYMPHOCYTES IN THE DEVELOPMENT OF PREGNANCY

. At the present time, a broad spectrum of CD8 + T lymphocyte subsets is revealed, including na ï ve cells, memory cells and regulatory subpopulations. Along with cells with high cytolytic activity, some subsets with marked regulatory activity were found there. Each subpopulation is characterized by a set of produced mediators, surface and intracellular markers allowing to suggest their differential in vivo functional activity. The present review article proposes a classification of CD8 + Т cells which takes into account their morphological and functional features. According to conventional view, the CD8 + Т lymphocytes is a cell population exhibiting high cytotoxic ability which is of critical significance in pregnancy, under the conditions of semi-allogenic fetal cell invasion into the endometrium. The fraction of CD8 + T cells is rather high in decidual structures. The review discusses the known mechanisms of differentiation regulation, selective migration and activity of CD8 + T cells in decidual membrane and placenta in the course of pregnancy. Perforine and granzyme are the main cytotoxicity factors of CD8 + Т cells. IL-2, IL-5, IL-13, IFN γ , IL-17, TGF- β and IL-10 cytokines are considered regulatory mediators of CD8 + cells. To induce the effector properties of CD8 + T cells, an antigenic stimulation is required, which is provided by interactions between the CD8 + Т cells and activated CD4 + Т cells or dendritic cells, cytokine effects. Specific differentiation of the CD8 + T cells is determined by differences in microenvironvent. In the course of pregnancy, accumulation of CD8 + Т cells is observed in decidual membrane, but their phenotype and functional properties differ from CD8 + Т cells in peripheral blood. At present time, the mechanisms of selective CD8 + T cell migration to decidual membrane are studied. These events are suggested to be mediated by means of CXCR3 and CCR5 chemokine receptors, IL-6 and IL-15 cytokines. The features of CD8 + Т cell activities, and production of some cytokines, e.g., CSF2, IFN γ , IL-1 β , IL-2, IL-6, IL-8,IL-10, IL-12 and TNF α in decidual membrane and is of critical significance for effective invasion of trophoblast cells. In turn, the trophoblast and placental cells promote development of regulatory CD8 + Т lymphocytes in decidual membrane, being able to induce CD8 + T cell apoptosis in decidual membrane. Hence, interaction between the maternal CD8 + T cells and trophoblast in the area of uterine-placental contact is an important link during development of immunological tolerance in the maternal/fetal system.


Introduction
Cytotoxic lymphocytes are one of the key cellular elements, whose action is aimed at viral clearance. However, the spectrum of their function is much wider. It is noted that the change in metabolic pathways and function of CD8 + T lymphocytes is associated with aging of the organism [46]. In connection with the possibility of recognition of MHC I, CD8 + T lymphocytes are an important part of the antitumor immune response. In particular, activated CD8 + T lymphocytes control cell growth in solid tumours [23]. CD8 + T lymphocytes are involved in many pathological processes, for example, in nerve fibre damage in multiple sclerosis [133], cerebral vasculature in cerebral malaria [126], bronchial asthma [95], rheumatoid arthritis [16] and immunopathology in skin leishmaniasis [94]. In the host graft system, CD8 + Т-лимфоциты при беременности CD8 + T lymphocytes during pregnancy 2018, Vol. 20, 5 2018, Т. 20, № 5 the effect of CD8 + T lymphocytes is also crucial for engraftment of the graft [5,112,151]. Pregnancy is a unique example of the coexistence of genetically diverse tissues. In this case, the placenta is not an absolute barrier to the cells of the immune system of the mother: there is a mutual penetration of the cells of the mother and foetusthe phenomenon of microchimerism, which can both positively and negatively affect the outcome of pregnancy [101,108]. T lymphocytes are of great importance for the recognition and elimination of foreign genetic material in the body, however, in the case of physiological pregnancy, there is no attack from the mother's immune system -explained by the immunological tolerance for the foetus. At present, the mechanisms of the development of this tolerance are intensively studied. In the endometrium and decidua, a change in the CD8 + T lymphocyte content is observed when pregnancy occurs compared with non-pregnant women and obstetric pathologies compared to the physiological course of pregnancy, but the role of these cells has not been sufficiently studied. It has been established that along with CD8 + T lymphocytes possessing cytotoxic action, there are CD8 + T lymphocytes with regulatory properties, as well as several subpopulations of memory cells. CD8 + T lymphocytes probably play one of the most important roles in the preservation or rejection of the foetus. At the present time, however, the whole spectrum of CD8 + T lymphocyte subpopulations is insufficiently characterized and the role of these cells in maintaining a physiological pregnancy is not fully understood.
Phenotype and functional activity of CD8 + T lymphocytes For a long time, CD8 + T lymphocytes have been considered a homogeneous population. At present, it has become clear that the diversity of CD8 + T lymphocyte subpopulations is determined by their differences both in phenotype, and in functional purpose, and in the characteristics of intercellular interactions. The vast majority of CD8 + T lymphocytes express the transcription factors T-bet and STAT4 and secrete the cytokines IFNγ and TNFα [93]. The main factors of the cytotoxic effect of CD8 + T lymphocytes are perforin and granzymes [93]. In addition, CD8 + T lymphocytes express a rather wide spectrum of receptors characteristic for NK cells: KIR -CD158a/h, CD158bj, CD94, NKG2A, NKG2С [132].
Several different subpopulations of CD8 + T lymphocytes are described in the public sources. The earliest classification of these lymphocytes is based on the spectrum of cytokines secreted by them. According to this classification, CD8 + T lymphocytes can be divided into 3 subpopulations: Tc1 -producing IFNγ and not producing IL-4; Tc2 -not producing IFNγ, producing IL-4; and Tc0 -producing IFNγ and IL-4 [90,138]. The majority of CD8 + T lymphocytes of human peripheral blood (80%) belongs to Tc1 [138]. Tc1 secrete IL-2, IFNγ, small amounts of IL-5, IL-13, and express the transcription factors T-bet and Hlx. Tc2 and Tc0 secrete IL-4, IL-5, IL-10, IL-13 [138]. Tc2 express the transcription factors GATA3 and Hlx [83]. Tc0 and Tc2 are characterized by high expression of CD30, CD40L, CD28, whereas Tc1 have a low expression of these molecules [138]. Tc2 are identified by the expression of the CRTH2 marker. The cytotoxic activity of the three subpopulations of CD8 + T lymphocytes is not different [138]. At the functional level, Tc1 accumulates faster in the lymph nodes and promotes a faster development of the immune response, and they are more apoptotic after activation (the mechanism does not involve changes in Fas and FasL expression) than Tc2 [128,138]. Cytokine IL-4 stimulates the development of IL-4-producing subpopulations, suppressing production of IFNγ [138]. The effect of IL-4 results in a loss of ability of Tc1 to secrete IL-2, and hence a loss of ability to spontaneously proliferate [111]. IL-12 inhibits the production of IL-5, IL-4 and IL-10, but stimulates the production of IFNγ, stimulates the development of the Tc1 subpopulation [138]. Subpopulations of Tc0 and Tc2 are not affected by IL-12 or IL-4 [138]. Prostaglandin E2 promotes a shift in the equilibrium toward Tc2 [10]. Due to the secretion of various cytokines, Tc can change the direction of differentiation of CD4 + T lymphocytes. Currently, there are still studies using this classification.
During the immune response, CD8 + T lymphocytes form memory cells, which are divided into three types: resident, effector and central [70]. Resident memory CD8 + T lymphocytes are localized in non-lymphoid tissues (mucous membranes and reproductive tract), differ in the inability to leave tissues in circulation and are characterized by the  [70,124]. Effective CD8 + T memory lymphocytes can migrate between tissues and secondary lymphoid organs, they do not express homing molecules that determine migration to the lymph nodes [124], are characterized by the CD8 + CD45RO + CD62L -CCR7 - [70] phenotype, high expression of cytolytic enzymes [130], high expression of transcription factor GATA3, high level of expression of IL-6Rα, IL-7Rα and increased level of proliferation [74]. Under the in vitro conditions, it was shown that after activation (CD3/CD28), these cells also produce cytokines IL-2, IL-5, IL-13 and IFNγ [74]. In the experiment, it was shown that the proliferation of effector CD8 + T lymphocytes is stimulated by IFNγ, with the synergistic action of IL-6 and IL-15 [74]. Also, the proliferation of effector CD8 + T lymphocytes can support IL-2 (autocrine). Cytokines secreted by effector memory CD8 + T lymphocytes are involved in antiviral and antibacterial defence of the body, in addition, there is an increase in the content of these cells in autoimmune processes (bronchial asthma) [74]. Central memory CD8 + T lymphocytes are localized in secondary lymphoid organs, they express homing molecules that determine migration to the lymph nodes, have a CD8 + CD45RO + CD62L + CCR7 + phenotype and a high proliferative potential upon re-encounter with antigens [70,124]. By analogy with CD4 + lymphocytes, Tc17 producing IL-17 are isolated. They are characterized by the phenotype CD27 -CD28 + CD45RA -CCR5 + CCR6 + , secretion of cytokines IL-17, IFNγ [68]. A subpopulation of CD8 + non-cytotoxic IL-17-producing T lymphocytes (Tcn17) is also formed, which is formed in the presence of TGF-β and IL-6 [83]. This subpopulation is characterized by the expression of the transcription factor Th17 RORγt, reduced expression of the transcription factors GATA3, T-bet, Hlx, the lack of production of IFNγ, granzyme B, IL-10 and the absence of cytolytic activity [83].
CD8 + regulatory T lymphocytes (CD8 + Treg) are a population of cells newly identified and are currently intensively studied. Along with CD4 + Treg, CD8 + Treg are important for the formation of immunological tolerance [84]. Their presence in immunologically privileged organs, participation in many pathologies of immune genesis, as well as their key role in the hosttransplant reaction have been described. For CD8 + Treg, the expression of CTLA-4, CD25, HLA-DR, CD45RA, CCR7, CD62L, CD28, CD101, CD103, CD122, TcRα/β, ICOS, FOXO1 and HELIOS, IL-2ra, CCR4, GARP, IL-10 and TGF-β [2, 7,37,55,75,84,119]. Expression of these molecules is important both for maintaining the CD8 + Treg population and for manifesting their functional activity against other cells [55]. According to some data, CD8 + Treg does not express CTLA-4, FasL [119]. CD4 + T lymphocytes requires the expression of FoxP3 for the implementation of regulatory properties [39,121,148]. Depending on the expression of FoxP3 CD8 + Treg can be divided into CD8 + FoxP3 + and CD8 + FoxP3subpopulations. For CD8 + FoxP3 + Treg, CD62L expression is also characteristic. Phenotype CD8 + FoxP3-Treg is characterized by the expression of CD103, which is crucial for the formation and implementation of suppressive properties of CD8 + Treg lymphocytes, especially for the subpopulation of CD8 + FoxP3 cells [75,84]. There are data on intracellular expression of FoxP3, characteristic of CD8 + Treg of lymphocytes [7,55]. Expression of surface molecules is largely specifically regulated by various microRNAs [55]. Probably, the formation of two subpopulations (CD8 + FoxP3 + and CD8 + FoxP3 -Treg) is associated with the action of TGF-β (in conjunction with the activation of the T cell receptor) and IL-10 [37,84]. Both subpopulations have a low expression of perforin and granzymes, intracellular expression and secretion of IL-10, and their suppressive effect is not due to cytotoxic effects, but to the production of TGF-β and IL-10 and contact interactions involving the CTLA-4 molecule [55,84,119].
Thus, the population of CD8 + T lymphocytes is a collection of cells quite diverse in phenotype and function. The conditions for differentiation of CD8 + T lymphocytes are also different.
Differentiation of CD8 + T lymphocytes Recent thymic emigrants that have just emerged from the bone marrow are localized in the blood and spleen, and differ from mature naïve cells: they have a decreased secretion of immunoregulatory cytokines, reduced expression of CD62L, CXCR4, granzyme B, reduced cytotoxicity, the ability to rapidly proliferate with an immune response and provide antiviral protection [27]. It is believed that decidual CD8 + T lymphocytes refer to such recent thymic emigrants at the periphery, but this issue has not been studied enough.
The antigen-independent differentiation of CD8 + T lymphocytes occurs in the thymus, from where the naïve cells exit into the periphery. In addition, naïve CD8 + T lymphocytes undergo antigen-dependent differentiation in secondary lymphoid organs: in the lymph nodes [15,135] and in the spleen [11], naïve CD8 + T lymphocytes interact with dendritic cells (DC) via the XCR1 + molecule in the presence of secreted IL-15, and also in the presence of CD4 + T lymphocytes secreting IL-2 [70]. Naïve CD8 + T lymphocytes are located in the paracortical area of the lymph node [63]. After activation, CD8 + T lymphocytes can differentiate into short-living effector cells that attack infected cells and die as a result of apoptosis after performing their function, or differentiate into circulating central memory cells [9]. About 5-10% of activated CD8 + T lymphocytes are transformed into memory cells [9]. Central memory cells are localized mainly in the interfollicular region of the lymph nodes [63]. CD8 + T lymphocytes are stored in the lymph nodes due to the expression of the homing receptors CCR7 and CD62L (L-selectin) [131].
Developing the effector properties of CD8 + T lymphocytes requires a triple signal: antigen stimulation through TcR, a costimulatory signal from CD28, microenvironment -exposure to cytokines IL-12, IFNα, IFNβ [24,88]. IL-12 and IFN type I cytokines play an important role both for the formation of a pool of effector cells and for the formation of memory cells [147]. These cytokines promote differentiation mainly into the Tc1 subpopulation [90]. The effect of cytokines causes a change in the expression of hundreds of genes responsible for proliferation, cytotoxic lymphocyte effector activity, survivability, migration, a wide range of transcription factors (including T-bet and Blimp-1) [24]. Secreted on the first day after activation of naïve CD8 + T lymphocytes, IFNγ via autocrine action activates and differentiates CD8 + T lymphocytes into cytotoxic T lymphocytes, stimulates the expression of the transcription factor T-bet, granzyme B production and plays an important role in early activation [22]. The effect of IFNγ is synergistically enhanced by the action of IFNα and IFNβ, but it does not matter for the differentiation of naïve CD8 + T lymphocytes in the presence of IL-12 [22].
The interaction of CD8 + T lymphocytes with activated CD4 + T lymphocytes, DC, is important for the formation of a pool of memory CD8 + T lymphocytes, as well as the effect of cytokines. The interaction of CD8 + T lymphocytes with DC is exercised through the CD70 molecule on the DC and CD27 on the surface of CD8 + T lymphocytes, and the interaction of CD40L on activated CD4 + T lymphocytes with CD40 on DC is simultaneously observed [50]. The expansion of memory CD8 + T lymphocytes depends on the interaction with CD4 T helper cells via CD40/CD40L and minor histocompatibility molecules [109]. The kinetics of cellular interaction remains a debating point [50]. It is believed that there is also a reserve pathway for maintaining memory cell differentiation, which is relevant in the absence of IL-12 and IFN type I, but it is not yet identified [147].
In the absence of CD4 + T lymphocytes, naïve CD8 + T lymphocytes cannot activate into effector cells, whereas memory CD8 + T lymphocytes can proliferate and differentiate into effector cells [76].
The IL7R, IL2R, IL15R, microRNA (miR)-155 receptors play an important role in determining the direction of differentiation of CD8 + T lymphocytes in the direction of the effector or memory cells, as well as the nature of the activation of TcR, in particular, the impact through the molecules of diacylglycerol (DAG), DAG kinase (DGK) α and ζ [8,60,113,131,150]. Deletion of DGK α and ζ leads to a decrease in the number of memory cells. This effect can be detected by observing a decrease in CD8 + expression by T lymphocytes of CD127, CD62L expression, as well as disturbance of the homeostasis of a subpopulation of memory cells and their viability [150]. Also, the deletion of DGK α and ζ causes a decrease in CD8 + T lymphocyte expression of the chemokine receptors CCR4, CCR5 and CXCR3, responsible for migration to the lymph nodes during the immune response [150].
CD8 + T regulatory lymphocytes can differentiate from naïve or effector CD8 + T lymphocytes in both the thymus and periphery [2]. As in the case of CD4 + Treg cells, TGF-β is important in the formation of the CD8 + Treg subpopulation: it suppresses the cytotoxic activity of CD8 + lymphocytes [127] by inducing microRNAs, leading to suppression of IFNγ production [17,84]. The effect of TGF-β was also manifested in the increased expression of CTLA-4, ICOS by the CD8 + T lymphocytes, and the CD62L on the FoxP3 + subpopulation of CD8 + T lymphocytes, but molecule CD103 on the subpopulation of CD8 + FoxP3 -T lymphocytes [75,84]. TGF-β-induced CD8 + FoxP3 + and FoxP3cells reduce production of IFNγ and do not proliferate in response to IL-2 [84]. The effect of monoclonal anti-CD3 antibodies with in vitro conditions leads to the induction of differentiation and expansion of CD8 + CD25 + FoxP3 + Treg lymphocytes [32]. Mesenchymal stem cells (MSC) of the placenta promote the differentiation of the CD8 + Treg lymphocytes (CD8 + IL-10 + ) with the participation of the PDL2 molecule (antiprogrammed death ligand-2) [77]. IFNγ and TNFα stimulate the expression of PDL2 on placental MSC and differentiation of CD8 + IL-10 + lymphocytes [77].
Thus, CD8 + T lymphocytes are capable of differentiation both in the thymus and in the periphery. The direction of differentiation depends on the nature of the intercellular interactions within the antigen presentation process in conjunction with the action After differentiation, CD8 + T lymphocytes acquire the ability to specifically migrate to further their function. Migration of CD8 + T lymphocytes to the decidua CD8 + T lymphocytes are present in peripheral blood, endometrium of non-pregnant women and in the decidua during pregnancy. There is little data on migration mechanisms of CD8 + T lymphocytes. However, several specific molecules can be isolated, whose expression promotes selective migration of cytotoxic lymphocytes into the decidua. Migration of CD8 + T lymphocytes to the decidua is facilitated by an increase in the expression of CXCR3 [21] and CCR5 [63,93]. CXCR3 ligands produce decidual stromal cells: CXCL9 (MIG), CXCL10 (IP-10), CXCL11 (I-TAC) [30]. CXCL9 is expressed in the placenta and in the chorioamniotic membrane, is found in the bloodstream, and the increased production of this molecule is associated with premature delivery and low foetal weight [29,62,65,105,115]. The increased CXCL11 content in amniotic fluid in the second trimester of pregnancy is associated with premature delivery [30] and may be associated with the involvement of CD8 + T lymphocytes by enhancing their migration. CCR5 ligands are CCL3 (MIP-1α), CCL4 (MIP-1β), RANTES. CCL4 is produced by decidual stromal cells [30], and CCL3 is expressed in the placenta by trophoblast, endothelial cells [1]. RANTES is produced by the decidua [28] and the placenta [47]. Trophoblast produces MIP-1α and RANTES, CCL3 [54,99].
The decidua is observed to be the producer of IL-15 [96]. Migration of CD8 + memory cells largely depends on the binding of P-and E-selectins, the expression of which increases in the presence of IL-15 [66]. CD8 + T lymphocytes are characterized by expression of the CXCR6 receptor, which, through interaction with the chemokine CXCL16 secreted by trophoblast, attracts T lymphocytes to the decidua [52]. CD103 expression can be associated with the migration of CD8 + Treg into the inflam mation zone [56,84]. In the placenta, the CD103 ligand -E-cadherin, is expressed by the villous trophoblast and is important for the functioning of the trophoblast itself [14,78,81], as well as the decidua [33,154]. Expression by villous and extravillous trophoblast, endometrial cells of prostaglandin D2 can cause specific migration to the decidua (to the implantation site of the foetal egg) of Tc2 lymphocytes due to their expression of CRTH2 [89].
When endometrial cells are differentiated into decidual cells, the expression of CXCL9 (MIG), CXCL10 (IP10), CXCL11 (ITAC) and CCL5 (RANTES) is suppressed, which leads to a decrease in Tc1 involvement in the decidua compared to the endometrium of non-pregnant women [35]. This can be one of the mechanisms of formation of immunological tolerance in the mother-foetus system.
If there is an infection, the migration of CD8 + T lymphocytes to the decidua increases, activation of CD8 + T lymphocytes occurs and immunological aggression against the foetus without elimination of the pathogen develops due to the action of CD8 + T lymphocytes [21]. Expression of E-cadherin with syncytiotrophoblast during preeclampsia is enhanced in comparison with physiological pregnancy [53,81], which may contribute to excessive involvement of CD8 + T lymphocytes to the placenta.
Cytokine IL-6 plays an important role in the formation and development of pregnancy, is found in significant amounts in the stroma of the decidua, trophoblast and endothelium of the spiral arteries. It has been established that IL-6 promotes the proliferation and differentiation of CD8 + CD25 + FoxP3 + Treg, an increase of their share in the blood stream and, as a result, an increase in their migration to the decidua [42,56,92], which can significantly contribute to the formation of immunological tolerance of mother's immune system in relation to the foetus and the development of the placenta [102], as well as in the process of rejection of the foetal tissues during the delivery [97].
In the endometrium of non-pregnant women, the proportion of IFNγ + CD8 + T lymphocytes reaches 25% of all CD8 + T lymphocytes; in spontaneous abortions, there is a significant decrease in the content of IFNγ + CD8 + T lymphocytes in the decidua compared with the endometrium of non-pregnant women (less than 11% of CD8 + T lymphocytes) [31], which indicates the important role of CD8 + T lymphocytes in implantation and the establishment of pregnancy.
Currently, there is no consensus on the persistence of the population of cytotoxic lymphocytes in the decidua during pregnancy. Some authors claim that during the pregnancy, the amount of CD8 + T lymphocytes in the decidua of women does not change [145] and is 45-75% of the T lymphocytes of the decidua (T lymphocytes constitute 10-20% of decidual leukocytes) [34,52,93]. Other authors point out a different content of cytotoxic lymphocytes in the decidua, depending on the period of pregnancy. In the first trimester of pregnancy, the proportion of CD8 + T lymphocytes in the decidua is insignificant and is 2-7% of the CD45 + cells, while in the third trimester of pregnancy it is up to 30% [21]. Such a difference in the data obtained by the authors on the content of T lymphocytes in the decidua can be associated with the absence of standardized methods for isolating cells from the decidua. The ratio of T cell populations in the systemic blood stream and decidua is different. In the decidua, the CD8 + T lymphocyte count is 62%, and CD4 + is 33% [91] of the total number of T lymphocytes, whereas in the peripheral blood, the proportion of CD8 + lymphocytes is 21%, and 75% of CD4 + lymphocytes [91]. The number of CD8 + memory T lymphocytes in the decidua considerably exceeds their number in peripheral blood [134], and the proportion of naïve CD8 + T lymphocytes is much lower than in peripheral blood [130], which indicates the presence of predominantly differentiated cells. Decidual CD8 + T lymphocytes have a lower degree of expression of perforin and granzyme B compared to CD8 + T lymphocytes of peripheral blood, and a small number of CD8 + T lymphocytes expressing granzyme B [130] (Table 1).
With pathologies of pregnancy, there is a change in the content of CD8 + T lymphocytes in the blood and decidua. Thus, with preeclampsia, a decrease in CD8 + FoxP3 + T lymphocytes (CD8 + T regulatory cells) in peripheral blood of women has been observed [152]. Spontaneous abortion shows an increased CD8 + lymphocyte count in the decidua [6]. The development of chronic chorioamnionitis in pregnant women is accompanied by an increase in the cytotoxic activity of CD8 + T lymphocytes, characterized by the phenotype CD300a + CD8 + , in the systemic blood flow compared to women with a physiological pregnancy [64].
Thus, CD8 + T lymphocytes play one of the decisive roles in the physiological and pathological development of pregnancy. Not only the existence of specific migration mechanisms of various subpopulations of CD8 + T lymphocytes in the endometrium and decidua is observed, but their concentration in these tissues in comparison with peripheral blood. At the same time, the ratio of various subpopulations of CD8 + T lymphocytes in the decidua at various stages of development of pregnancy is substantive.

Phenotypic and functional features of CD8 + T lymphocytes of the decidua
The presence of CD8 + T lymphocytes in the endometrium is important for the implantation of the ovum. Cells of the Tc2 subpopulation predominate in the implantation site. They express CRTH2, which is an additional receptor of prostaglandin D2 and is characteristic of Th2 [89]. The cytolytic activity of CD8 + T lymphocytes is hormone-dependent: it is present in the proliferative phase of the menstrual cycle and is absent in the secretory phase [142]. Decidual CD8 + T lymphocytes differ in phenotype and functional characteristics from peripheral blood lymphocytes (Table 1). In the bloodstream, naïve CD8 + T lymphocytes predominate [131]. Decidual and placental CD8 + T lymphocytes in the vast majority are activated effector cells or effector memory Т cells [131] ( Table 1). Activated effector cells are characterized by the phenotype CD45RA + , CD45RO -, CD62L -, CCR7 -, expression of Tbet transcription factor [41, 87,131]. Effector memory cells are characterized by the phenotype CD8 + CD45RA -CCR7 -CD28 -CD45RO + CD103 + ICOS + , increased expression of CD69 and HLA-DR and lack of perforin. Whereas in the peripheral blood, naïve CD8 T cells (CD8 + CD45RA + CD45RO -CCR7 + ) predominate [91, 129,130,134]. Also for effector memory T cells, expression of NK cell receptors (KIR -CD158a/h, CD158bj, CD94, NKG2A, NKG2С) is characteristic [69,132]. NK receptor expression is controlled by the cytokines IL-2, IL-4, IL-23, TGF-β, IL-15, IL-6, IL-10, IL-21, and also depends on the nature of the antigen (due to the interaction with CD28) [132]. Expression of one of the NK receptors -CD158a/h, is increased in CD8 + T lymphocytes of the decidua compared to CD8 + T lymphocytes of peripheral blood; the expression of the remaining NK receptors on CD8 + T lymphocytes of the decidua and peripheral blood is the same [132]. At the same time, decidual CD8 + T lymphocytes are characterized by reduced cytotoxic activity compared with peripheral CD8 + T lymphocytes. One of the proposed mechanisms for this phenomenon may be the reduced expression of FasL, as well as perforin and granzyme B. At the same time, high expression of mRNA perforin and granzyme B was established, which indicates a post-translational mechanism of blocking the production of these proteins [130,134]. Also, reduced cytotoxicity may be associated with the effect of the locus molecules HLA-G and HLA-C expressed by the trophoblast. The functions of NK receptors on CD8 + T lymphocytes have not been adequately studied at present.
The decidual CD8 + T lymphocytes of the first tri mester of pregnancy secrete in vitro CSF2, IFNγ, IL-1β, IL-2, IL-6, IL-8, IL-10, IL-12, and TNF (especially IFNγ and IL-8), that stimulates trophoblast invasion [91,114]. A high level of TGF-β provides a high threshold level of cytolytic activity of CD8 + T lymphocytes in the decidua [21], which also contributes to the successful invasion of trophoblast. This effect can be associated with suppression of CD8 molecule expression by cytotoxic T lymphocytes in the presence of TGF-β [98].
An ICOS molecule is expressed on 96% of decidual CD8 + T lymphocytes [91], which belongs to the family of CD28-B7 costimulatory molecules. Expression of ICOS is inducible. ICOS binds to ICOSL, expressed on professional APC (B cells, macrophages, DC), epithelial and endothelial cells [143], including on the trophoblast [91]. It was noted that as a result of ICOS binding to ligand, decidual CD8 + T lymphocytes secrete more IFNγ and IL-10 than CD8 + T lymphocytes of peripheral blood [91]. Such stimulation of secretory activity of CD8 + T lymphocytes upon contact with trophoblast through ICOS can be directed to stimulation of trophoblast invasion. It was shown that the ICOS molecule is important for the realization of the immunological response by T helper cells (cell proliferation, activation), T regulatory cells (differentiation, proliferation) [146] and Th17 lymphocytes [61,91,143]. The important role of the ICOS molecule expressed by CD8 + T lymphocytes in the antigen-dependent response in tissue transplantation has been shown. In particular, in the "graft versus host" system, the deletion of ICOS on CD8 + T lymphocytes leads to a fatal outcome, caused by excessive expansion and secretory activity of CD8 + T lymphocytes [153]. In peripheral CD8 + T lymphocytes, early expression of ICOS results in migration to non-lymphoid organs, increased cytolytic activity and decreased ability of CD8 + T lymphocytes to participate in the development of a secondary immune response [82]. Probably, decidual CD8 + T lymphocytes thus contribute to the formation of a favourable cytokine microenvironment and a decrease in the cytotoxicity of the cells of the immune system of the mother during the trophoblast implantation. On the other hand, decidual CD8 + T lymphocytes of the first trimester of pregnancy in vitro have the ability for extracellular cytolysis [114]. When decidual CD8 + T lymphocytes are exposed to proinflammatory cytokines and activation of their TcR, their expression of perforin and granzyme B, as well as their degranulation of cytotoxic granules, is enhanced, which distinguishes these cells from decidual NK cells characterized by greatly reduced cytolytic activity [21]. Simultaneously, decidual CD8 + T lymphocytes produce more granulin as compared to a similar subpopulation of peripheral blood, which can provide local protection against intracellular and extracellular pathogens [21].
The proteins Tim-3 (T cell immunoglobulin mucin-3) and PD-1 (programmed cell death-1) CD8 + Т-лимфоциты при беременности CD8 + T lymphocytes during pregnancy 2018, Vol. 20, 5 2018, Т. 20, № 5 are important signalling molecules of CD8 + T lymphocytes. Their functions have not been sufficiently studied. The high level of expression of PD-1 and Tim-3 CD8 + T lymphocytes correlates with reduced antiviral activity of these cells, reduced proliferative activity of CD8 + T lymphocytes and reduced secretion of pro-inflammatory cytokines CD8 + T lymphocytes, while their blockade leads to restoration of proliferation and synthetic activity cells [57,100]. There is also evidence that the interaction of PD-1 and B7-H1 determines the cytotoxic pathway of differentiation of CD8 + T lymphocytes [40]. Signals from PD-1 and Tim-3 synergistically reinforce each other [57,140]. It has been shown that Tim + PD-1 + CD8 + T lymphocytes are concentrated in the decidua in a physiological pregnancy and participate in the establishment of pregnancy in the early stages [57,140,149]. Tim + PD-1 + CD8 + T lymphocytes show an increased level of proliferative activity in comparison with Tim-3-PD-1-CD8 + lymphocytes, have greater expression of activation markers CD69 and HLA-DR and greater secretion of Th2 cytokines (in particular, IL-4 and IL-10) [140]. In the case of spontaneous abortions, both the amount of Tim + PD-1 + CD8 + T lymphocytes in the decidua, and the production of anti-inflammatory cytokines are reduced. At the same time, increased production of proinflammatory cytokines IFNγ and TNFα is observed [149]. The mouse model shows that the blockade of Tim-3 and PD-1 leads to a significant decrease in the weight of the foetus and the number of offspring, which indicates the important role of these molecules in providing foetal protection and normal pregnancy [140]. In women with spontaneous abortions, an association of a decreased number of decidual CD8 + T lymphocytes with the PD-1 + CD8 + phenotype and a decreased proliferative activity has been observed [140]. PD-1L is expressed on syncytiotrophoblast cells directly in contact with the parent cells [137], which is one of the mechanisms of formation of immunological tolerance for trophoblast cells. Expression of PD-1L with trophoblast is stimulated by EGF and IFNγ [48]. It is likely that increased production of IFNγ trophoblast in early pregnancy [3,4] and its autocrine effect may be one of the mechanisms of survival of a semi-allogenic foetus as a result of the interaction of PD-1 on T lymphocytes and PD-1L on the trophoblast. From the first to the second trimester, the expression of PD-1L in the placenta is increased, the regulation of expression is probably carried out depending on oxygenation (insufficient oxygenation in the uteroplacental blood flow zone causes a decrease in expression of PD-1L) [49]. Decreased expression of PD-1L in the decidua is associated with habitual miscarriage [77].
Thus, Tim and PD-1 are important in establishing the tolerogenic nature of CD8 + T lymphocyte differentiation in pregnancy. Factors secreted by the placenta and decidua, significantly affect the formation of immunological tolerance in pregnancy, in particular, affect the apoptosis of the mother's cells. It was noted that the level of apoptosis of CD3 + T lymphocytes in peripheral blood is higher in pregnancy than in nonpregnant women [25]. At the same time, the level of apoptosis of CD3 + CD8 + T lymphocytes at the onset of pregnancy is lower than in non-pregnant women, and the maximum level of apoptosis of this subpopulation of T lymphocytes is observed in the first trimester of pregnancy [25], which can be aimed at ensuring foetal survival in early pregnancy and elimination of alloreactive cells. The decidua produces significant amounts of glycodelins A (GdA), which inhibits the cytotoxic activity of activated CD8 + T lymphocytes and their proliferation [123] using CD7 as a receptor [125]. GdA suppresses the synthesis of IL-2R, which interferes with the action of IL-2 on T lymphocytes and the formation of functionally mature cytotoxic cells [122]. The placenta secrets the preimplantation factor (PIF), which binds to the proteins of the CD8 + T lymphocytes cytoskeleton and suppresses the immune response of the mother [12]. Interaction with soluble HLA-C, whose concentration in the peripheral blood rises in pregnancy, causes apoptosis of CD8 + T lymphocytes by enhancing its production of FasL [19,104].
CD8 + T lymphocytes active against paternal antigens, including anti-minor histocompatibility antigens (mHag-peptides derived from polymorphic proteins, recognized as heterogenous) have been described in the literature, but they do not exert any influence on the outcome of pregnancy [131]. This may be due to the formation of subpopulations of regulatory CD4 and CD8 lymphocytes, levelling their influence [131].
In pregnancy, virus-specific CD8 + T lymphocytes accumulate in the decidua [134]. But since decidual CD8 + T lymphocytes acquire more regulatory properties than cytotoxic ones, while retaining the ability to recognize the antigen present in the HLA locus molecule, including the viral antigen -there is no complete elimination of the infected cells [21,134]. Most likely, the activity of CD8 + T lymphocytes can be aimed at resisting a viral infection in the context of maintaining pregnancy.
Thus, decidual CD8 + T lymphocytes differ in phenotype, secretory and cytolytic activity from CD8 + T lymphocytes of peripheral blood, which is aimed at supporting the invasion of trophoblast in the early stages of development of pregnancy, as well as the formation of immunological tolerance for the halfallogenic foetus. The disturbance of the homeostasis of decidual CD8 + T lymphocytes has serious consequences that affect the outcome of pregnancy. The mechanisms of maintaining the balance of the functioning of CD8 + T lymphocytes within the framework of maintaining the physiological course of pregnancy and at the same time effective protection against infectious agents remain insufficiently studied. The diverse interaction of CD8 + T lymphocytes with microenvironment cells in the utero-placental blood flow zone has a significant effect on the nature of the functional activity of all interacting cell types.

Intercellular interactions of CD8 + T lymphocytes
In pregnancy, trophoblast cells play a central role in the development of immunological tolerance. In the regulation of the cytotoxicity of immune system cells from the trophoblast side, the expression of the locus molecules of the main histocompatibility complex HLA-G and HLA-C is important. Receptors for the molecules of the HLA-G locus on CD8 + T lymphocytes are the molecules CD8, LILRB1 and CD160 [72]. HLA-G locus molecules are expressed on the trophoblast in combination with endoplasmic reticulum aminopeptidase 1 (ERAP1), which generates peptides capable of forming a stable complex with HLA-G [106]. This contributes to the successful invasion of the trophoblast. Expression of this complex on trophoblast cells is enhanced by exposure to LIF and VEGF [106]. The extravillous cytotrophoblast produces both a membrane and a soluble form of HLA-G [59,67]. The soluble form is found not only in the amniotic fluid, but also in the mother's bloodstream [13,45,120]. It was shown that the soluble form of the HLA-G locus molecule promotes immunological tolerance, causing apoptosis of CD8 + T lymphocytes [73], including through interaction with the CD8 molecule [72]. HLA-G locus molecules can also inhibit the proliferation of CD4 + lymphocytes, thereby reducing the risk of developing a mixed lymphocyte response from both the mother and the foetus. The low level of production of HLA-G by the trophoblast is associated with complications, including the development of preeclampsia [45,85]. At the same time, some of the latest studies have revealed an increased level of HLA-G mRNA in the decidua in preeclampsia [103].
According to the latest data described in the literature, several stages can be distinguished in the interaction of trophoblast and CD8 + T lymphocytes. First, direct recognition of HLA-C by the T cell receptor of CD8 + T lymphocytes. Further, a peptide (viral or allogenic) is presented in the MHC I complex, activation of CD8 lymphocytes. The mouse model shows that the presentation of foetal antigens occurs exclusively in an indirect way, due to maternal APC, and antigen presentation occurs only in the late stages of pregnancy, which is associated with the late onset of the expression of MHC I molecules on foetal cells [18,21,36]. At the same time, it has been repeatedly shown that macrophages and DC of the placenta and decidua are of a tolerogenic nature [26,38,79,116,118,139,144]. The literature also describes the presentation of minor and major histocompatibility antigens in the maternal APC after the capture of trophoblast debris by CD4 + T lymphocytes, which in turn secrete cytokines that affect the differentiation of CD8 + T lymphocytes.
Antigen-presenting cells and trophoblast produce significant amounts of IL-27 [20,43,107]. In naïve CD8 + T cells (CD8 + CD45RA + CCR7 + ), IL-27 enhances the expression of the T-bet transcription factor, which indicates the polarization of CD8 + T lymphocytes in the Tc1 direction, stimulates proliferation, IFNγ secretion, granzyme B and cytotoxicity of these cells [117]. However, the mechanisms of influence of IL-27 on the development of pregnancy remain unclear. The IL-27 receptor is expressed on CD8 + T lymphocytes to a greater extent than on CD4 + lymphocytes, mainly on CD8 + CD45RO + lymphocytes (i.e., memory cells) [117]. Intracellular expression of the IL-27 receptor (TCCR/gp130) is characteristic of naïve cells, with activation the surface expression of the receptor is enhanced [117]. The mouse model shows that the production of IL-27 by the decidua is enhanced before implantation [86]. In women with habitual miscarriages, much lower expression of IL-27 in the decidua is observed compared to women with spontaneous abortion and the physiological course of pregnancy [141].
The extravillous cytotrophoblast expresses the B7H2 molecule (ICOS ligand). The specific interaction of ICOS/B7H2 contributes to the secretion of T cells by cytokines IFNγ, IL-10 and IL-2 [91]. According to other data, foetal formation causes strong proliferation of CD8 + T lymphocytes in the uterine lymph nodes during pregnancy, but they are simultaneously susceptible to clonal deletion [36]. The remaining activated CD8 + T lymphocytes are characterized by reduced secretion of IFNγ, IL-2 and IL-4 and inability to cytolytic activity [36]. Previously, it was also noted that IL-4, secreted by Th2, leads to a decrease in the production of IFNγ, TNFα and IL-10 by Tc1 cells [110].
It has been established that trophoblast stimulates the formation of a subpopulation of CD8 + CD8 + Т-лимфоциты при беременности CD8 + T lymphocytes during pregnancy 2018, Vol. 20, 5 2018, Т. 20, № 5 T regulatory cells [77,119]. Since activation of CD8 + T lymphocytes involves CD4 + T lymphocytes, they will have a significant effect on the functioning of cytotoxic T lymphocytes. CD4 + Treg cells by secretion of IL-10 contribute to the formation of CD8 + cell memory in viral infection [71]. CD4 + Treg lymphocytes suppress CD8 + T cell response: CD4 + Treg cause apoptosis of CD8 + cytotoxic T lymphocytes and reduce their expression of granzymes A and B [44]. CD4 + Th1 lymphocytes are involved in the induction and maintenance of viability of CD8 + lymphocytes (Tc1), both effector and memory cells, through IL-2 secretion, activation of DC by CD40/ CD40L interactions, and also using pMHC I [51]. CD8 + Treg stimulate the differentiation of CD4 + Treg [75]. Subpopulations of CD8 Tc1 and Tc2 lymphocytes differently influence the differentiation of CD4 + T lymphocytes: in the presence of Tc1, Th1lymphocytes predominate, and in the presence of Tc2, Th2-lymphocytes predominate [138]. It has been shown that CD8 + T lymphocytes stimulate the expansion of cells of a specific subset of CD4 + Treg expressing the TCR Vβ5 chain whose proliferation regulation mechanism differs significantly from other Tregs and does not depend on IL-2 [58]. The enhancement of the surface expression of TNFα, observed with the activation of CD8 + lymphocytes, promotes the expansion of Vβ5 + Treg lymphocytes [58]. Simultaneously, the expression of TNFR2 on Vβ5 + Treg plays a leading role in inducing the proliferation of these cells [58]. The action of Treg is aimed at limiting the cytotoxic activity of CD8 + T lymphocytes, therefore the revealed regulation mechanism is a system of mutual influence of T cell subpopulations during the antiviral response. The results were obtained on a mouse model, but there is reason to assume that such a mechanism exists in humans [58]. The secretory products of decidual CD8 + lymphocytes of the first trimester of pregnancy stimulate trophoblast invasion [114], which can be mediated by the action of IFNγ. Secreted by CD8 + lymphocytes, IL-8 can play an important role in vascular remodelling in the first trimester of pregnancy by stimulating MMP production. Thus, CD8 + lymphocytes contribute to the formation of the placenta in the early stages of pregnancy.
The trophoblast cells (including due to the production of the locus molecules HLA-C and HLA-G), T helper lymphocytes, APC of the placenta and decidua, have a significant effect on the formation of the subpopulation composition of CD8 + T lymphocytes of the decidua. In turn, decidual CD8 + T lymphocytes affect trophoblast cells and CD4 + T lymphocytes; however, the molecular mechanisms of these cell-cell interactions have not been sufficiently studied.

Conclusion
The population of CD8 + T lymphocytes is a collection of cells quite diverse in phenotype and function. Great importance is attributed to subpopulations of memory CD8 + T lymphocytes and CD8 + Treg, identified relatively recently. Differentiation of CD8 + T lymphocytes depends on the cytokine environment and antigen presentation involving surface molecules. Not only differentiation is possible in the primary lymphoid organs, but also in the secondary ones; however, the molecular mechanisms for the regulation of differentiation of CD8 + T lymphocytes have not been adequately studied. Migration of CD8 + T lymphocytes to the decidua is caused both by changes in expression of surface molecules of CD8 + T lymphocytes under the action of soluble products in the utero-placental contact zone, and by changes in the production of soluble and surface molecules by cells of the decidua. Molecular mechanisms of specific migration of CD8 + T lymphocytes in pregnancy are poorly understood. An important role of the concentration of CD8 + T lymphocytes in the endometrium and decidua during the whole pregnancy is distinguished. Violation of the quantitative content or functional activity of CD8 + T lymphocytes in the decidua can lead to serious complications of pregnancy. CD8 + T lymphocytes in the utero-placental contact zone interact with a wide range of cells. Their functioning in these tissues is aimed at stimulating the invasion of trophoblast and the formation of immunological tolerance, accompanied by a decrease in its own cytotoxic activity and an increase in regulatory function. However, the molecular mechanisms of intercellular interactions are subject to further detailed study.