CD56bright cells increase expression of {alpha}4 integrin at ovulation in fertile cycles.

Leukocyte content of human endometrium changes rapidly after ovulation, particularly as a result of gains in CD56(bright) uterine NK (uNK) cells. We have proposed that uNK precursor cells are found within the blood CD56(bright) pool and are recruited to decidualizing endometrium through functional changes in their adhesion molecules and chemokine receptors. This study sought to quantify alterations in adhesion molecules, cytokines, chemokines, and receptors induced in circulating CD56(+) cells of fertile and infertile women by ovulation. Blood was drawn from 12 fertile volunteers and six female-infertility patients at Menstrual Cycle Day (d) 5 and on the day following the preovulatory surge of luteinizing hormone (LH). CD56(bright), CD56(dim), and CD56(+)CD3(+) cell subsets were isolated and evaluated by flow cytometry, quantitative PCR, or Western blotting. In CD56(bright) cells from fertile but not infertile women, alpha(4) integrin increased between d5 and the preovulatory LH surge. CD56(dim) and NKT cells did not show a change in alpha(4) integrin but differed significantly between fertile and infertile donors, and infertile donors had reduced homing molecule expression in CD56(dim) and NKT cells, and at ovulation, their NKT cells showed elevated cytokine production. None of the circulating CD56(+) cell subsets had transcripts for receptors for estrogen, progesterone, LH, or prolactin. Thus, immunological events associated with the LH surge induce alterations in all subsets of CD56(+) cells, and the unique induction of alpha(4) integrin in CD56(bright) cells of fertile women constitutes a potential method to promote uterine homing.

CD56+ cells are recruited to the uterus in two waves: at ovulation and during the first 2 weeks after missed menses.

PROBLEM: Uterine natural killer (uNK) cells are enriched in the post-ovulatory uterus and during pregnancy. Whether these cells arise from blood pre-cursors or from stem cells in the uterus is undefined. To support a hypothesis that precursors of uNK cells are recruited from blood, adhesive function of blood CD56+ subsets were assessed during one cycle and during pregnancy. METHOD OF STUDY: Fifteen women of proven fertility provided serial blood samples during one menstrual cycle and thirty women with a history of implantation failure or recurrent spontaneous abortion provided serial samples during infertility treatment. RESULTS: CD56(bright) cells, but not CD56(dim) cells or NKT cells, increased in ligand-binding capacity during ovulation in fertile cycles only and during the first 2 weeks from date of missed menses. CONCLUSION: Enhanced adhesive function at ovulation in CD56(bright) cells in fertile cycles and during early gestation supports a hypothesis of recruitment of pre-uNK cells from the blood CD56(bright) subset.

Decline in number of elevated blood CD3(+) CD56(+) NKT cells in response to intravenous immunoglobulin treatment correlates with successful pregnancy.

PROBLEM: Patients with elevated blood natural killer (NK) cells may be offered intravenous immunoglobulin (IVIG) treatment, but there is controversy about the utility of blood NK cell testing. Human CD56(+) NK cells include several subpopulations that include the putatively cytotoxic CD56(+) CD16(+) subset. In mouse models of pregnant failure, NKT cells appear to be important. However, a mouse model may only be pertinent to a subset of patients, as recurrent pregnancy failure is a heterogenous group. METHOD OF STUDY: An ethics-approved observational study was done to observe the effect of treatment on total blood lymphoid cells, and subsets of CD56(+) blood lymphocytes including CD56(+) CD3(+) NKT cells determined by flow cytometry, and to correlate with pregnancy outcome. Fifteen fertile women with a history of successful pregnancy and thirty-one women suffering from repeated implantation failure or recurrent spontaneous abortion provided serial blood samples during one menstrual cycle or prior to and during treatment. IVIG was administered to the latter group with or without heparin/aspirin. RESULTS: Eight of thirty infertile women presented with high numbers of CD56(+) CD3(+) NKT cells, which declined after treatment with IVIG. The elevated NKT cell group with or without concomitant autoimmunity achieved a significantly higher successful pregnancy rate over the course of the study, as compared to women with average numbers of NKT cells and no evidence of autoimmunity (P = 0.018). Elevated NKT levels alone was an independent predictor of success on treatment (P = 0.003). CONCLUSION: Elevated NKT cells in recurrent pregnancy loss or implantation failure can be ameliorated with IVIG treatment, and result in successful pregnancy. Assay of NKT cell numbers may identify patients who are more likely to benefit from IVIG therapy and merits further examination in randomized phase II studies.

A review of trafficking and activation of uterine natural killer cells.

PROBLEM: Enrichment of uterine natural killer (uNK) cells occurs during pregnancy in many species. However, functions of uNK cells and regulation of their uterine homing are not fully defined. In mice and women, uNK cells contribute to angiogenesis, a role reviewed here and now addressed in a mammal with an alternative placental type. METHODS OF STUDY: To address lymphocyte functions, RNA from murine or porcine endometrium and lymphocytes purified from endometrium were analyzed using real-time or reverse transcription PCR. To address homing potential, human blood CD56(+) lymphocytes were evaluated using both RNA and functional adhesion to endothelium presented under shear force in frozen sections of gestation day 7 C57Bl/6J implantation sites. Women were serially sampled over a menstrual cycle or a clinical preparatory cycle for embryo transfer. RESULTS: Activation of murine uNK cells is associated with much greater increases in transcription for Eomes than for T-bet (Tbx21). Lymphocytes from normal porcine implantation sites transcribe vascular endothelial growth factor, placental growth factor, interferon-gamma and hypoxia-inducible factor (HIF)-1alpha. In fertile women, increases in L-selectin- and alpha4-integrin-mediated interactions between CD56(+) cells and endothelium occur at luteinizing hormone (LH) surge (cycling women) to oocyte pick up or embryo transfer, then return to pre-LH levels. CONCLUSIONS: Uterine lymphocytes may universally promote pregnancy-associated endometrial angiogenesis. Recruitment of uNK precursor cells from blood appears to occur in a window promoted by rising plasma estrogen and LH and limited by rising progesterone.

Trafficking of peripheral blood CD56(bright) cells to the decidualizing uterus--new tricks for old dogmas?

CD56(bright) lymphocytes become abundant in the human uterus during every menstrual cycle, following the surge in pituitary-derived luteinizing hormone (LH), which initiates final oocyte maturation. While the uterus is host to some CD56(bright) cells prior to ovulation, the rapid increase is thought to be due to proliferation of the resident population, accompanied by recruitment of CD56(bright) lymphocytes from the circulation. The rapid increase in CD56(bright) cells is concurrent with the onset of decidualization, the transformation of uterine stromal cells into secretory decidual cells. Uterine CD56(bright) cells proliferate and differentiate to become the predominant lymphocytes of the post-ovulatory uterus. These distinct, tissue-specific natural killer (NK) cells either die prior to menses or increase in number during early pregnancy, and then decline toward the end of the first trimester. Since lymphocytes home to tissues from the circulation, we investigated mechanisms of NK cell traffic over the course of natural menstrual cycles by measuring functional interactions between CD56+ cells from blood and endothelial cells using the Stamper-Woodruff assay of lymphocyte adhesion to frozen tissue sections. While a baseline level of adhesion was maintained throughout the cycle, elevated l-selectin-dependent adhesion of peripheral blood CD56(bright) cells occurred during a peri-ovulatory window. However, there were no significant menstrual cycle-induced changes in the transcription of l-selectin, alpha 4 integrin or LFA-1, or in expression of these proteins by NK cells, suggesting that the enhanced adhesion was due to post-translational modifications of these molecules. Quantitative RT-PCR failed to amplify the message for LH receptor or the alpha or beta forms of progesterone or estrogen receptors from blood NK cell subsets. Thus, we conclude that the actions of LH, E(2,) and P(4) on NK cells that promote interactions with endothelium and potential uterine homing are indirectly mediated through the responsiveness of other cell types.