Isolation of Cells from the Feto-Maternal Interface

Department of Pathology, University of Cambridge, Cambridge, United Kingdom.
Current protocols in immunology / edited by John E. Coligan ... [et al.] 04/2012; Chapter 7:Unit 7.40.1-11. DOI: 10.1002/0471142735.im0740s97
Source: PubMed


The mucosal lining of the human uterus is host to a specialized population of leukocytes, which, during pregnancy, interact with invading placental cells (trophoblast) of fetal origin. Of particular interest are uterine natural killer cells, which account for around 70% of the leukocytes at this site during the first trimester of pregnancy, and seem to be specially adapted to recognize invading trophoblast cells. In order to understand the interactions between mucosal immune cells and trophoblast, and those among the immune cells themselves, it is useful to be able to isolate and culture these cells. Here, we describe protocols for the isolation of leukocytes, stromal cells, and trophoblast cells from the feto-maternal interface.

Download full-text


Available from: Ashley Moffett, Mar 23, 2015
  • Source
    • "Placental villi samples were obtained from women undergoing surgical termination of pregnancy in the first trimester. Primary human trophoblasts were isolated as previously described [18]–[19]. Briefly, the collected placental villi were minced and digested in 0.25% trypsin for 15 minutes with constant shaking. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Soluble human leukocyte antigen-G (HLA-G) is a non-classical class Ib HLA molecule that is secreted from blastocysts. Soluble HLA-G modulates the immune tolerance of the mother and can be used as a prognostic factor for the clinical pregnancy rate. However, the underlying mechanism of how soluble HLA-G5 affects pregnancy remains largely unknown. We hypothesized that soluble HLA-G5 promotes successful implantation and pregnancy by modulating trophoblast invasion through receptor binding and activation of extracellular signal-regulated protein kinase (ERK) signaling pathway. Recombinant HLA-G5 protein over-expressed in E. coli BL21 was purified to near homogeneity. We studied the expression of HLA-G5 and its receptors, the leukocyte immunoglobulin-like receptor subfamily B1 (LILRB1) and killer cell immunoglobulin-like receptor 2DL4 (KIR2DL4), in primary trophoblasts and trophoblastic (JAr and JEG-3) cell lines by florescence-labeled HLA-G5. HLA-G5 was detected in the primary trophoblasts and JEG-3 cells. The LILRB1 and KIR2DL4 receptors were expressed in both primary trophoblasts and trophoblastic cell lines. HLA-G5 stimulated cell invasion (p<0.05) and increased urokinase (uPA) and matrix metalloproteinases (MMPs) transcripts and their activity (p<0.05) in trophoblastic cells. HLA-G5 activated the ERK signaling pathway and induced ERK1/2 phosphorylation in the trophoblastic cell lines. Addition of ERK inhibitors (U0126 and PD98059) nullified the stimulatory effect of HLA-G5 on trophoblastic cell invasion. Taken together, HLA-G5 induced trophoblast invasion by binding to KIR2DL4 and LILRB1, by increasing uPA and MMPs expressions and by activating the ERK signaling pathway.
    Full-text · Article · Oct 2013 · PLoS ONE
  • [Show abstract] [Hide abstract]
    ABSTRACT: Pregnancy is a cooperative interaction between the mother and her fetus, allowing survival and normal growth of the fetus. Successful pregnancy remains a fascinating phenomenon as it resists the immunological rules of rejection. Immunological recognition of the fetus is vital for maintenance of gestation. The maternal immune system undergoes changes that lead to tolerance of the fetus. Inadequate recognition of fetal antigens may cause abortion. In fact, fetal cells express paternal alloantigens that are not recognized as foreign by the mother. A special balance between lymphocytes is present at the feto-maternal interface to control the immune response. In addition, placental trophoblasts act as a physical barrier and exert an immunoregulatory function. Trophoblast cells regulate the expression of human leucocyte antigens. Dysfunction of these cells leads to morphological and functional alterations of the feto-maternal barrier as well as to recurrent spontaneous abortions. Uterine natural killer cells are appropriate residents of the materno-fetal interface to support the adaptation of the blood vessels of the pregnant uterus and regulate trophoblast invasion into the decidua and myometrium. Cytokines are involved at the feto-placental unit by adapting normal T-cell trafficking and modulating the inflammatory process. This study discusses the complex immunological aspects of immune tolerance and the balance of immunity in pregnancy in terms of the role of the human leucocyte antigen, placental trophoblasts, maternal immunosuppression, immune cells, cytokines and immunoregulatory molecules at the feto-maternal interface.
    No preview · Article · Mar 2013 · Egyptian Journal of Histology
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: This review summarizes the potential and also some limitations of using human placentas, or placental cells and structures for toxicology testing. The placenta contains a wide spectrum of cell types and tissues, such as trophoblast cells, immune cells, fibroblasts, stem cells, endothelial cells, vessels, glands, membranes, and many others. It may be expected that in many cases the relevance of results obtained from human placenta will be higher than those from animal models due to species specificity of metabolism and placental structure. For practical and economical reasons, we propose to apply a battery of sequential experiments for analysis of potential toxicants. This should start with using cell lines, followed by testing placenta tissue explants and isolated placenta cells, and finally by application of single and dual side ex vivo placenta perfusion. With each of these steps, the relative workload increases while the number of feasible repeats decreases. Simultaneously, the predictive power enhances by increasing similarity with in vivo human conditions. Toxic effects may be detected by performing proliferation, vitality and cell death assays, analysis of protein and hormone expression, immunohistochemistry or testing functionality of signaling pathways, gene expression, transport mechanisms, and so on. When toxic effects appear at any step, the subsequent assays may be cancelled. Such a system may be useful to reduce costs and increase specificity in testing questionable toxicants. Nonetheless, it requires further standardization and end point definitions for better comparability of results from different toxicants and to estimate the respective in vivo translatability and predictive value.
    Full-text · Article · Apr 2013 · Toxicologic Pathology
Show more