The macrophage tetraspan MS4A4A enhances dectin-1-dependent NK cell-mediated resistance to metastasis.

The plasma membrane tetraspan molecule MS4A4A is selectively expressed by macrophage-lineage cells, but its function is unknown. Here we report that MS4A4A was restricted to murine and human mononuclear phagocytes and was induced during monocyte-to-macrophage differentiation in the presence of interleukin 4 or dexamethasone. Human MS4A4A was co-expressed with M2/M2-like molecules in subsets of normal tissue-resident macrophages, infiltrating macrophages from inflamed synovium and tumor-associated macrophages. MS4A4A interacted and colocalized with the ß-glucan receptor dectin-1 in lipid rafts. In response to dectin-1 ligands, Ms4a4a-deficient macrophages showed defective signaling and defective production of effector molecules. In experimental models of tumor progression and metastasis, Ms4a4a deficiency in macrophages had no impact on primary tumor growth, but was essential for dectin-1-mediated activation of macrophages and natural killer (NK) cell-mediated metastasis control. Thus, MS4A4A is a tetraspan molecule selectively expressed in macrophages during differentiation and polarization, essential for dectin-1-dependent activation of NK cell-mediated resistance to metastasis.

Human Decidual CD1a+ Dendritic Cells Undergo Functional Maturation Program Mediated by Gp96.

Heat shock proteins (hsps), in certain circumstances, could shape unique features of decidual dendritic cells (DCs) that play a key role in inducing immunity as well as maintaining tolerance. The aim of the study was to assess the binding of gp96 to Toll-like receptor (TLR) 4 and CD91 receptors on decidual CD1a+ DCs present at the maternal-fetal interface in vitro as well as the influence of CD1a+ DCs maturation status. Immunohistology and immunofluorescence of paraffin-embedded first-trimester decidua tissue sections of normal and pathological (missed abortion MA and blighted ovum BO) pregnancies were performed together with flow cytometry detection of antigens in CD1a+ DCs after gp96 stimulation of decidual mononuclear cells. Gp96 efficiently bound CD91 and TLR4 receptors on decidual CD1a+ DCs in a dose-dependent manner and increased the expression of CD83 and HLA-DR. The highest concentration of gp96 (1000 ng/mL) increased the percentage of Interferon-γ (INF-γ) and IL-15 expressing gp96+ cells. Gp96 binds CD91 and TLR4 on decidual CD1a+ DCs, which causes their maturation and significantly increases INF-γ and IL-15 in the context of Th1 cytokine/chemokine domination, which could support immune response harmful for ongoing pregnancy.

Differences in tolerogenic status of NK cells between luminal A type, luminal B type, and triple-negative breast cancer.

Globally, breast cancer is the main cause of death among female cancer patients. The tumor-infiltrating lymphocytes (TILs) in breast cancer are associated with a more favorable outcome of a disease. Natural killer (NK) cells are important cytotoxic cells involved in tumor immunosurveillance, causing the direct killing of tumor cells. In solid tumors, peripheral NK cells and tumor-infiltrating NK cells display an altered phenotype characterized by reduced cytotoxicity or anergy. The goal of this study was to investigate the NK cells' phenotype and activation status in order to get into the pathological process of breast cancer subtypes. In our study, the normal tissues and tumoral breast tissue were fixed in formalin, embedded in paraffin, and the phenotypic marker CD56 and proinflammatory cytokine IL-15 were identified by immunohistology. The distribution and expression of receptors repertoire (NKG2A, NKG2C, NKp46, CD94, CD69, and CD107a) were investigated in peripheral NK cells of mononuclear cells by flow cytometry. mRNA of cytolytic mediators was determined by real-time PCR. The frequency of CD56+ and IL-15+ cells were significantly higher in triple-negative breast cancer tissue. The frequency of NK cell activating receptors was decreased in investigated breast cancer subtypes while the inhibitory NKG2A receptor was increased. Decreased percentage of CD69+/CD107a+ in NK cells could indicate lower cellular activation and cytotoxicity. In luminal B breast cancer, the mRNA of cytolytic mediators was upregulated. In conclusion, modulation of activation status in tumor-infiltration NK cells could be involved in the pathogenesis of molecular breast cancer subtypes. This highlights the importance of NK cells as an appropriate target for potent anti-tumor response in the immunosuppressive tumor microenvironment of breast cancer.

Monocyte-macrophage polarization and recruitment pathways in the tumour microenvironment of B-cell acute lymphoblastic leukaemia.

B-cell acute lymphoblastic leukaemia (B-ALL) reprograms the surrounding bone marrow (BM) stroma to create a leukaemia-supportive niche. To elucidate the contribution of immune cells to the leukaemic microenvironment, we investigated the involvement of monocyte/macrophage compartments, as well as several recruitment pathways in B-ALL development. Immunohistochemistry analyses showed that CD68-expressing macrophages were increased in leukaemic BM biopsies, compared to controls and predominantly expressed the M2-like markers CD163 and CD206. Furthermore, the "non-classical" CD14+ CD16++ monocyte subset, expressing high CX3CR1 levels, was significantly increased in B-ALL patients' peripheral blood. CX3CL1 was shown to be significantly upregulated in leukaemic BM plasma, thus providing an altered migratory pathway possibly guiding NC monocyte recruitment into the BM. Additionally, the monocyte/macrophage chemoattractant chemokine ligand 2 (CCL2) strongly increased in leukaemic BM plasma, possibly because of the interaction of leukaemic cells with mesenchymal stromal cells and vascular cells and due to a stimulatory effect of leukaemia-related inflammatory mediators. C5a, a macrophage chemoattractant and M2-polarizing factor, further appeared to be upregulated in the leukaemic BM, possibly as an effect of PTX3 decrease, that could unleash complement cascade activation. Overall, deregulated monocyte/macrophage compartments are part of the extensive BM microenvironment remodelling at B-ALL diagnosis and could represent valuable targets for novel treatments to be coupled with classical chemotherapy.

Combined upper limb and breathing exercise programme for pain management in ambulatory and non-ambulatory multiple sclerosis individuals: part II analyses from feasibility study.

PURPOSE: The present small semi-controlled feasibility study investigated a possible efficacy of a combined upper limb and breathing exercise programme in managing pain in ambulatory and non-ambulatory patients with EDSS from 0.0-8.0. METHOD: People with MS (N = 19) were enrolled in this single-blind randomized controlled study and divided into 2 groups: exercise group (5 ambulatory, 5 non-ambulatory; Expanded Disability Status Scale (EDSS), 1.0-8.0) and related control group that performed no exercise (4 ambulatory, 5 non-ambulatory; EDSS, 1.0-7.5). The exercise group performed combined upper limb and breathing exercises in a group led by a physiotherapist (2 days/week, 60 min/session) accompanied by independent home exercises (3 days/week, ≥ 20 min/session). Participants underwent measures of pain level (visual analogue scale) for physical pain, functional independence of daily activities (Barthel index) and handgrip strength (HGS) for dominant (D) and non-dominant (ND) hand evaluated by a dynamometer before and after the 4-week period by the blinded assessor. RESULTS: The VAS for pain showed statistically significant group-by-time interaction only in non-ambulatory (p = .049) individuals, but with large intervention effects on both subgroups (ambulatory, p = .159; non-ambulatory, d = 0.97). Functional independence in daily activities (Barthel index) showed statistically non-significant group-by-time interaction in ambulatory (p = .195, d = 0.89) and non-ambulatory (p = .102, d = 1.64) individuals, but despite the absence of statistical significance, there were large intervention effects. Handgrip strength was significantly improved for both hands in ambulatory (D, p = .012; d = 2.07; ND, p = .025, d = 1.77) and only non-dominant hand in non-ambulatory individuals (D, p = .288, d = 0.83; ND, p = .012, d = 2.21). CONCLUSION: This small pilot study provides preliminary proof-of-concept data supporting low-intensity upper limb and breathing exercise programme for potential reduction of pain and improvement of functional independence in both ambulatory and non-ambulatory individuals with MS in a larger sample and that strengthening the upper limbs might be an additional pain relief mechanism. TRIAL REGISTRATION: NTC03222596.

Cell death mechanisms at the maternal-fetal interface: insights into the role of granulysin.

During mammal pregnancy, a sensitive balance between hormones, cytokines, humoral factors, and local cellular interactions must be established. Cytotoxic cells infiltrating the decidua are heavily equipped with cytolytic molecules, in particular perforin and granulysin. Granulysin is especially abundant in NK cells which are able to spontaneously secrete high quantities of granulysin. Besides being a potent bactericidal and tumoricidal molecule, granulysin is also found to be a chemoattractant and a proinflammatory molecule. The precise role(s) of granulysin at the maternal-fetal interface has not been elucidated yet. It is possible that it behaves as a double-edged sword simultaneously acting as an immunomodulatory and a host defense molecule protecting both the mother and the fetus from a wide spectrum of pathogens, and on the other hand, in case of an NK cell activation, acting as an effector molecule causing the apoptosis of semiallograft trophoblast cells and consequently leading to various pregnancy disorders or pregnancy loss.

HSP70 In triple negative breast cancer: Prognostic value and clinical significance.

BACKGROUND: Triple-negative breast cancer (TNBC) has the worst prognosis and the highest immunogenic potential of all breast cancer subtypes. It is characterized by a lack of estrogen and progesterone receptors as well as HER2. A major component of the tumor microenvironment (TME) of TNBC is tumor-infiltrating lymphocytes (TILs). A chaperone heat shock protein 70 (HSP70) is involved in several pathways that enable tumour growth and progression, as well as in immune modulation. METHODS: Immunohistochemical analysis of HSP70 expression in immune cells, as well as expression of immunosuppressive markers CTLA4 and PD-L1 and major TILs components: CD8, CD4 and Tregs were analyzed in the superficial and deep tumor layer of primary TNBC and compared with established clinicopathological parameters. Clinical data and surgical tissue samples from 68 TNBC patients who underwent initial surgery were included in the analysis and 36 control samples from benign breast tissue biopsies. RESULTS: A higher expression of TILs, CD4, CD8 and PD-L1 was found in the invasive tumor front (ITF), as compared to the tumor center (TC) (p < 0001). HSP70 positive immune cells (HSP70(+) IC) in TC were associated with adverse clinical and pathological markers: higher stage of disease (P = 0.013), higher grade (P = 0.05) and a higher pN status (P < 0.001). In addition, higher expression of HSP70(+) IC from TC was correlated with the higher expression of FOXP3(+)T cells both in ITF (N = 61, rho=0.42, p < 0.001) and in metastatic tissue from the draining lymph nodes (N = 13, rho=0.61, P = 0.026). CONCLUSION: Correlations between HSP70 immune cells expression and individual TILs components support the hypothesis of its active role in inducing immunosuppression and tumor progression. Routine determination of HSP70 expression, in immune cells of TC, may be of added value in the clinical decision-making process concerning axillary surgery.

COVID-19 and pregnancy: are they friends or enemies?

OBJECTIVES: Novel coronavirus disease (COVID-19) is rapidly spreading all over the world. Although in many cases the infection causes very weak symptoms, it can be severe in patient with diverse chronical diseases and immunological compromising patients. Pregnancy is a unique condition in which mother and fetus peacefully collaborate. Diverse endocrine-immune mechanisms, mostly under progesterone control work together to protect the fetus from maternal immunocompetent cell activation driven rejection. The physiological shift to Th2 dominant environment, while favourable for fetus, it makes mothers susceptible to infective pathogens, making pregnancy during COVID-19 pandemic challenging. MATERIALS AND METHODS: Studies involving COVID-19 in pregnancy and those analysing changes of immune system induced by COVID-19 were searched in databases such as PubMed, Scopus, Google Scholar and ScienceDirect. Databases were searched using a keyword COVID-19/coronavirus, that was combined with following terms: immune system, pregnancy, oestrogen, or progesterone. Search included studies published up to 01.07.2020. Almost 1,500 articles were found, but only 18 met criteria. RESULTS: Most frequent symptoms of COVID-19 in mothers infected in the late pregnancy were fever and cough accompanied with lymphopenia and elevated C-reactive protein. Mothers reported to have severe disease had comorbidities and were obese. Low rate of neonatal complications of maternal Sars-Coc-2 infection without neonatal mortality was observed. CONCLUSIONS: Currently available data didn't show significant relationship between COVID-19 severity and pregnancy and there is no strong evidence that mother's infection can lead to adverse pregnancy outcome, but further studies are needed to determinate the possible effects of COVID-19 gained during earlier pregnancy.

The Role of Innate Immunity in the Pathogenesis of Breast Cancer.

BACKGROUND: Breast carcinoma is the most common malignant disease in the female population and one of the leading causes of death among women worldwide. One crucial hallmark of cancer is chronic inflammation where the immunosuppressive environment is dominant. The immunosuppressive environment is largely achieved by the interaction of tumor cells and infiltrating leukocytes. SUMMARY: Usually, human macrophages and natural killer cells are involved in antitumor immunity. The therapeutic potential of this population against cancers has stimulated their study and led to the discovery of several different tumor-associated macrophages and natural killer cell subsets, each of which is endowed with different immunoregulatory functions. Both heterogeneity and plasticity of the tumor-associated macrophages and natural killer cell compartment, which are both tightly linked to the tumor microenvironment of different breast cancer types. KEY MESSAGES: The identification of specific tumor-associated macrophages and natural killer cell subsets endowed with particular functional capabilities might help monitor tumor-mediated responses in breast cancer patients. Currently, one of the most used strategies for breast cancer of newly diagnosed patients is neoadjuvant chemotherapy.

The involvement of the progesterone receptor in PIBF and Gal-1 expression in the mouse endometrium.

PROBLEM: The progesterone-regulated genes, PIBF and Gal-1, are key players in the feto-maternal immunological interaction. This study aims to investigate the expression of PIBF and Gal-1 in WT and progesterone receptor KO models as well as subsequent effects of PIBF on decidualization of stromal cells. METHOD OF THE STUDY: PRAKO, PRBKO and PRKO BALB/c mice were used for assessing the role of PR isoforms in PIBF induction. PIBF- and Gal-1 mRNA expression in the uterus was tested by real-time PCR. The effect of PIBF on decidualization of endometrial stromal cells was verified by anti-desmin immunofluorescence. Immunohistochemistry was used for testing PIBF expression in the uterus. Gal-1, ERα and PR positive decidual NK cells were detected by immunofluorescence. RESULTS: PIBF mRNA was significantly increased in progesterone-treated WT mice, but not in PRKO and PRAKO mice. PIBF protein expression was reduced in the endometria of PRKO and PRAKO, but not in PRBKO mice. During a 6-day culture, PIBF induced decidual transformation of endometrial stromal cells. PIBF expression in the mouse uterus was highest during the implantation window, while Gal-1 mRNA expression continuously increased between day 2.5 and day 11.5 of gestation. Decidual NK cells express Gal-1 and ERα, but not PR at day 7.5 murine pregnancy. CONCLUSION: PIBF produced via engagement of PRA, is highly expressed in the endometrium during the implantation window, and plays a role in decidualization. The concerted action of PIBF and Gal-1 might contribute to the low cytotoxic activity of decidual NK cells.

Assessing whether progesterone-matured dendritic cells are responsible for retention of fertilization products in missed abortion.

We hypothesize that progesterone causes tolerogenic maturation of myeloid dendritic cells (DCs) in human decidua of threatened miscarriage or missed abortion characterized by a distinct phenotype and cytokine production, including reduction of the main NK cell proliferation and cytotoxic factor interleukin (IL)-15. During DC/NK cell interaction, progesterone-shaped DCs cannot efficiently multiply or equip NK cells with the cytotoxic mediators peforin and granulysin, which might harm trophoblasts and induce abortion. We propose that the presence, and maturation stage of decidual myeloid DCs be investigated using semi-quantitative immunohistological analyses and/or double-color immuno-fluorescent labeling of DC lineage and activation markers. The spatial arrangement of granulysin+ cells, NKp46+ NK cells, DCs, and trophoblasts might provide information about their mutual interactions in vivo. Multiple flow cytometry analyses of NK-receptors would provide insight into NK cell activation status. NK cell activation status could be also assessed by cytotoxicity assays against trophoblast cell lines, or isolated cognate extra-villous trophoblast cells. A correlation between decidual progesterone concentration or IL-15 expression, and the degree of DC maturation or the frequency of granulysin+ cells, might help to elucidate the mechanism of abortion retention in utero.

Granulysin-mediated apoptosis of trophoblasts in blighted ovum and missed abortion.

PROBLEM: Granulysin (GNLY) is a cytotoxic molecule mostly present in decidual natural killer (NK) cells. Blighted ovum (BO) and missed abortion (MA) represent the early pathological pregnancies with hindered development of the embryoblast or a dead embryo. We investigated the GNLY-mediated apoptotic mechanism potentially responsible for delayed termination of pregnancy. METHOD OF STUDY: We performed immunohistological and immunofluorescence labeling of decidual tissues (GNLY, Apaf-1, NF-κB). NKG2A expression was analyzed by flow cytometry and GNLY mRNA by RT-qPCR. RESULTS: The GNLY labeling intensity (H score) was lower in the nuclei of trophoblast cells in BO and MA. GNLY gene levels were inversely detected in BO and MA. A decreased decidual NK cell percentage was found in MA. NK cells from pathological pregnancies expressed lower NKG2A levels. The highest frequency of Apaf-1 was found in trophoblast cells of MA. NF-kB was highly expressed in decidual cells of BO. CONCLUSION: The reduced activation of GNLY-mediated killing might be implicated in the slower rejection of trophoblast cells in BO and MA. A decreased authentic decidual NK cell number could be responsible for low cytotoxicity against trophoblast cells in MA. In BO, trophoblast cells have a higher survival potential due to increased NF-kB expression.

Potential role of heat-shock protein 70 and interleukin-15 in the pathogenesis of threatened spontaneous abortions.

PROBLEM: The role of HSP70 and both its constitutive (Hsc) and inducible (Hsp) forms in the pathogenesis of threatened spontaneous abortions was investigated. METHOD OF STUDY: Immunohistology and/or immunofluorescence was used to analyze paraffin-embedded tissue sections, and reverse transcriptase-quantitative polymerase chain reaction and flow cytometry were used for analyses of decidual mononuclear cells (DMCs) and confocal microscopy for the detection of perforin, granulysin, and lysosome-associated membrane protein-1 (LAMP-1) in decidual lymphocytes (DLs). RESULTS: The percentage of single Hsp70(+) , Hsc70(+) , and IL-15(+) cells and mRNA levels of HSP70, CD91, and TLR4 were lower in the decidua basalis in cases of threatened miscarriages compared to that in cases of normal pregnancy. In a suspension of normal DMCs, IL-15 significantly decreased the HSP70 members and TLR4 in dendritic cells, T cells, and NK cells while increasing CD91 in NK cells alone. CONCLUSION: Downregulation of Hsc70, Hsp70, and IL-15 expression at gene and/or protein levels might support the retention of fertilization products in cases of missed abortion and blighted ovum.

Heat-Shock Proteins 70 Induce Pro-Inflammatory Maturation Program in Decidual CD1a(+) Dendritic Cells.

PROBLEM: The aim of the study was to assess possible binding of a mixture of constitutive Hsc70 and inducible Hsp70 forms (HSP70) to Toll-like receptor (TLR) 4 and CD91 receptors on decidual CD1a(+) dendritic cells (DCs) and their influence on DCs maturation status. METHOD OF STUDY: Immunohistology and immunofluorescence of paraffin-embedded first trimetester and term pregnancy decidua were performed together with flow cytometry detection of antigens in DCs after stimulation of decidual mononuclear cells with HSP70. RESULTS: Hsc70 and Hsp70 labeling revealed intracellular and nuclear staining in trophoblast cells. The numbers of Hsc70(+) and Hsp70(+) cells of decidual tissue were higher in early pregnancy decidua than in decidua at term. HSP70 binds CD91 and TLR4 receptors on CD1a(+) DCs and increased the expression of CD83, HLA-DR, CD80, and CD86, but decreased CC receptor (CCR) 5. HSP70 increased CC ligand (CCL) 3 and CCL22. HSP70 in the concentration of 1 µg/mL increased the percentage of interferon-γ and interleukin (IL)-15-expressing cells over the cells expressing IL-4. CONCLUSION: HSP70 binds CD91 and TLR4 on decidual CD1a(+) DCs, causes their maturation, and increases IL-15 in the context of Th1 cytokine/chemokine domination, which could support immune response harmful for ongoing pregnancy.

Granulysin expression and the interplay of granulysin and perforin at the maternal-fetal interface.

Granulysin (GNLY) is a cytolytic/apoptotic molecule highly expressed in immune cells, particularly NK cells, at the maternal-fetal interface. The primary function of GNLY is to carry out lysis or apoptosis induction in target cells, tumor cells or cells infected by intracellular pathogens. To exert some of its functions GNLY needs to collaborate with perforin. The purpose of this study was to determine: (a) the expression of GNLY at the gene and protein levels at the maternal-fetal interface, (b) the relationship(s) between GNLY and perforin, and (c) GNLY secretion by NK cells stimulated by the NK-sensitive K562 cell line and its HLA-C and HLA-G transfectants. GNLY and perforin genes were found to be highly activated at the interface. GNLY mRNA was present at significantly higher levels compared with other cytolytic/apoptotic molecules. Confocal microscopy analysis showed that most first trimester pregnancy decidual lymphocytes simultaneously contained both GNLY and perforin protein in their cytoplasm, with a punctuate pattern consistent with granule localization. In contrast to peripheral blood, in unstimulated decidual lymphocytes GNLY and perforin rarely co-localized (10% of GNLY-positive cells and 20% of perforin-positive cells were positive for both proteins). Contact between decidual lymphocytes and K562 cells caused GNLY and perforin to be expressed in the same granules (approximately 50% co-localization), i.e., to attain the pattern seen in peripheral blood lymphocytes. The abundant GNLY secretion by decidual NK cells compared with peripheral blood NK cells after 2h of contact with the NK-sensitive K562 cells and K562 transfectants was striking.

The significance of heat-shock protein gp96 and its receptors' CD91 and Toll-like receptor 4 expression at the maternal foetal interface.

PROBLEM: Differences in the expression of gp96 and its receptors were analysed in normal and pathological human pregnancy. MATERIAL AND METHODS: Immunohistology and immunofluorescence of sections from decidual part of term placenta, first trimester normal decidua, missed abortion and blighted ovum decidua were performed together with reverse transcriptase-quantitative polymerase chain reaction and flow cytometry. RESULTS: In missed abortion, gp96 was intensively stained, when compared to normal early pregnancy. The intensity of CD91 and TLR4 was higher in the first trimester pregnancy and blighted ovum, when compared to missed abortion. Decidual part of the term placenta is invaded with gp96⁺ , CD91⁺ and TLR4+ trophoblast. Progesterone-induced blocking factor (PIBF) decreased the frequency of TLR4⁺ T lymphocytes, CD91⁺ T, natural killer (NK) and mature dendritic cells after an 18-h culture. Decidual mononuclear cells (DMCs) treated with PIBF down-regulated CD91, TLR4 and gp96 gene expression. CONCLUSION: The presence of gp96, CD91 and TLR4 at the maternal-foetal interface provides a molecular basis for their interaction, particularly in the absence of PIBF.

First trimester pregnancy decidual natural killer cells contain and spontaneously release high quantities of granulysin.

PROBLEM: Granulysin (GNLY) is a novel cytolytic protein lytic against a variety of tumor cells and microbes. The role of GNLY during pregnancy has not been extensively explored. The aim of this study is to examine GNLY expression and distribution in the first trimester pregnancy peripheral blood (PB) and decidua, the ability of decidual and PB natural killer (NK) cells to secrete GNLY spontaneously, and the role of antigen-presenting cells (APC) in the regulation of GNLY expression in decidual NK cells. METHOD OF STUDY: GNLY expression was analyzed using cell permeabilization method, flow cytometry, and immunohistochemistry. GNLY secretion by purified NK cells was detected by ELISA method. RESULTS: GNLY is abundantly expressed at the maternal-fetal interface in the first trimester pregnancy. Decidual T lymphocytes express significantly higher levels of GNLY (58%) then PB T lymphocytes (11%). Over 85% of decidual CD56(+) cells express GNLY and when cultured spontaneously release high quantities of GNLY. Decidual APC participate in the control of GNLY expression in CD56(+) cells. CONCLUSION: Abundant expression of GNLY in the decidual immunocompetent cells and the capacity of decidual CD56(+) cells to spontaneously secrete high quantities of GNLY point to important protective and immunomodulatory role that this molecule could play at the maternal-fetal interface.

Phenotype of NK cells and cytotoxic/apoptotic mediators expression in ectopic pregnancy.

PROBLEM: The expression of cytotoxic/apoptotic mediators and the phenotype characteristics of uterine NK cells (uNK) in tubal ectopic pregnancy (EP) were investigated. METHOD OF STUDY: Samples of uterine decidua and tubal mucosa as well as peripheral blood (PB) of the same women with EP were used for phenotype characterization of NK cells and detection of cytotoxic/apoptotic mediators and IL-15. RESULTS: In tubal mucosa, perforin, FasL, granulysin and IL-15 were almost completely absent, but they were present in normal and EP uterine deciduas. TRAIL was present on trophoblast and tubal mucosa, contrary to its lack in normal and EP uterine decidua. CD16⁻ CD56(dim) NK cells, mostly CD94⁻ and NKG2A⁻, predominate in tubal mucosa, whereas CD16⁻ CD56(bright) NK cells, predominantly CD94(+) and NKG2A(+) prevail in EP uterine decidua. NK cells at the EP implantation site express lower percentages of perforin and granulysin, but they express a higher percentage of TRAIL than do EP uterine decidual and PB NK cells. Lower percentage of TNF-α-expressing and IL-4-expressing NK cells were found at the implantation site compared to EP uterine decidua. CONCLUSIONS: Authentic uNK cell population seems to be insufficient to restrict trophoblast invasion because of low expression of cytotoxic/apoptotic mediators.