Granulated metrial gland cells in `minor'' species

Granulated metrial gland cells, also known as uterine natural killer cells or large granular lymphocytes, are pregnancy associated leucocytes of granular phenotype. They are well characterised in mice and humans in terms of their structure, origin and distribution although the function of these cells has yet to be determined. In this review, granulated metrial gland cells in ‘minor' species of rodents, insectivores, primates and species with epitheliochorial placentae are described. Emphasis is given to the comparative structure and distribution of granulated metrial gland cells in these minor species and to their possible functional association with trophoblast. Comparative studies of granulated metrial gland cells in minor species complements other approaches such as can be provided using mutant mice.     

Leucocyte membrane antigens on mouse granulated metrial gland cells

An indirect immunofluorescence technique has been used to study the expression of leucocyte membrane antigens on mouse granulated metrial gland (GMG) cells. GMG cells isolated from cultured metrial gland explants and GMG cells in cryostat sections of uterine implantation sites were examined. GMG cells were found to express both the asialo-GM1 antigen and the Thy-1 antigen. GMG cells did not express the Lyt-1, Lyt-2, Mac-1, L3T4 or IgM antigens. These results provide new evidence that GMG cells are a type of NK cell.     

The killing of mouse trophoblast cells by granulated metrial gland cells in vitro

Mouse placental cell preparations have been maintained in culture, and the types of cell that attached to the culture dish were classified according to morphological criteria. However, these morphological criteria were insufficient to determine from which trophoblast layer in the placenta all of the types of cell found in the cultures originated. Some placental cell preparations were co-cultured with granulated metrial gland (GMG) cells and these cultures were studied using time-lapse video. Various responses to contacts between GMG cells and trophoblast cells were observed. These responses included the killing of trophoblast cells by GMG cells.     

Receptors for IgG2b on cells of the mouse metrial gland

Single cells prepared from metrial glands of mice killed at days 10, 13 and 17 of pregnancy were assayed for the expression of Fc gamma receptors in a standard rosetting assay using sheep red blood cells sensitised with a mouse monoclonal IgG2b antibody. Rosettes, indicating Fc gamma receptors, were found on both granulated metrial gland (GMG) cells and non-GMG cells, comprising mainly stromal cells, from each stage of pregnancy. Some animals were given an intravenous injection of horseradish peroxidase 2 h before they were killed in order to identify endocytic cells. No GMG cells were found to have endocytosed the horseradish peroxidase. Non-GMG cells which showed endocytic activity all expressed Fc gamma receptors but these receptors were also found on some of the non-GMG cells which had not exhibited endocytosis. The finding of Fc gamma receptors on GMG cells provides further evidence that these cells may be related to NK cells.     

The killing of rat placental cells by rat and mouse granulated metrial gland cells in vitro

Small round cells which migrated from explant cultures of rat metrial gland were identified as granulated metrial gland (GMG) cells. They contained large amounts of glycoprotein and displayed the leucocyte common antigen. Other cells which migrated from the explants were probably derived from the fibroblast-like stromal cells of the metrial gland. The asialo-GM1 antigen was found on rat GMG cells in culture and in cryostat sections of rat metrial gland. The rat GMG cells in culture exhibited locomotion and, when co-cultured with placental cells, made numerous contacts with the placental cells. A small number of these contacts (less than 1 per cent) were followed rapidly by the death of the placental cell. Mouse GMG cells which had migrated from explant cultures of mouse metrial gland were also co-cultured with rat placental cells. The migratory activity of the mouse GMG cells also involved numerous contacts being made with rat placental cells and a small number (less than 1 per cent) of these contacts were cytotoxic for the rat placental cells. The observations support previous suggestions that GMG cells are a type of killer cell. The cytotoxic activity of rat and mouse GMG cells against co-cultured rat placental cells is discussed in relation to the nature of the target molecule involved.     

Immunoglobulin G in the decidua basalis and metrial gland of the pregnant mouse uterus

The distribution of immunoglobulin G (IgG) in the decidua basalis and metrial gland of the mouse uterus at days 10 and 14 of pregnancy has been investigated using immunohistochemical methods. IgG was found in the intercellular spaces but none was found in decidual cells, stromal cells of the metrial gland or granulated metrial gland cells. These results differ from those of other studies which have localised IgG in the cytoplasm of rat granulated metrial gland cells.     

Immunological characterisation of surface receptors on rat metrial gland cells

Cells from metrial glands of pregnant rats were examined for surface receptors. No E, EA mu or EAC receptors were demonstrated. Fc gamma receptors were detected by EA gamma rosette formation, using sheep red blood cells sensitised with the IgG fraction of rabbit or rat antisera. Significantly more of the metrial gland cells, and of the rat peritoneal exudate and spleen cells examined as controls, formed rosettes with red cells sensitised with rabbit IgG than with those sensitised with rat IgG. The proportion of metrial gland cells forming EA gamma rosettes decreased significantly between day 12 and day 15 of pregnancy but increased by day 19. Metrial gland cells from deciduomata formed EA gamma rosettes, and the proportions varied during pseudopregnancy. At day 13 of pregnancy a greater proportion of metrial gland cells displayed Fc gamma receptors in multiparous rats than in primigravid rats. The binding affinity of the Fc gamma receptors was characterised by inhibition studies with homologous and heterologous IgGs. Maximal inhibition occurred when the inhibitory IgG was homologous to the IgG used to sensitise the red cells. EA gamma rosette formation by cells from the metrial gland was inhibited by both monomeric and heat-aggregated IgGs.     

Evaluation of the murine metrial gland for immunological function

The metrial gland (MG) is a transient uterine structure associated with rodent pregnancy. The gland is a complex structure consisting of stromal and vascular elements, as well as a population of histologically distinctive, large, granulated metrial gland (GMG) cells. The functions of the MG and of the GMG cells, as well as their relationship to the success of pregnancy, are unknown. Based upon morphological and morphometric studies it has been proposed that the MG might be involved in the immunology of pregnancy and that GMG cells could be immunocompetent. Explant cultures of MG have therefore been evaluated for immunological function. Lytic activity against the NK sensitive target cell line YAC and mitogen responsiveness could not be detected. MG tissue and medium conditioned by overnight culture of MG tissue (MG-CM) suppressed the response of murine spleen cells to Con A. MG-CM also reduced the lytic activity of splenic NK cells against YAC target cells. However, uptake of [3H]thymidine was elevated when YAC cells were cultured in MG-CM. The response of embryonic and uterine cells to growth in MG-CM was complex. MG-CM had little effect on isotope incorporation by decidual cells recovered at 6.5 days or by embryonic cells recovered from 12.5 day embryos. However, thymidine incorporation was less in MG-CM than in control medium for 12.5 day placental cells, 6.5 day embryonic sac, 6.5 day ectoplacental cone and 3.5 day blastocysts. Cytotoxicity and cytostasis accounted for reduced uptake of isotope in cultures of 3.5 day blastocysts and 6.5 day embryonic tissues. Loss of viability could not be detected in any other assays. Both YAC cells and unstimulated splenocytes showed altered morphology and improved viability when cultured in MG-CM. This study suggests that the only immunological role the MG might have during normal pregnancy is that of non-specific intra-uterine suppression. Alternatively, differential regulation of cell proliferation might be a function of the MG, within the pregnant uterus. The latter mechanism could also account for the apparent observation of non-specific immunosuppression.     

Localisation of immunoglobulin (IgG) in granulated metrial gland cells of deciduomata of the pseudopregnant rat

Using an immunoperoxidase technique, after fixation in saturated alcoholic mercuric chloride, it was possible to detect cytoplasmic immunoglobulin (IgG) in granulated metrial gland cells from deciduomata of pseudopregnancy in the rat. The extent of the reaction for IgG was variable but did not appear to be related to the day of pseudopregnancy or to the extent of the decidual reaction. Examination of single cell suspension enabled quantification of IgG-containing cells, but no significant differences were detected in the numbers of positive cells at the days of pseudopregnancy which were examined. Surface IgG was detected on a small proportion of the cells, and they were distinguished from the Fcgamma receptor-bearing cells in the metrial gland of deciduomata of pseudopregnancy.     

The characterisation of Fc receptors on cells from the metrial gland of the pregnant rat uterus

Using an EA rosetting technique on cells from the rat metrial gland at days 12--15 of pregnancy, approximately 30% of the cells were found to possess immunoglobulin (Fc) receptors. At dat 14 a comparison was made between cells from the metrial gland and cells from the peritoneal exudate; binding affinity of each for homologous and heterologous immunoglobulins was assessed by the resulting inhibition of rosette formation. Cells from the metrial gland were inhibited to a greater degree than cells from the peritoneal exudate by low concentrations of rabbit, human and rat immunoglobulin. The Fc receptors observed on cells from the metrial gland show some resemblance to those described by other workers on yolk sac cells and certain foetal placental cells, but their function is at present unknown.     

Localisation of immunoglobulin (IgG) within the rat metrial gland

An immunohistological method was used to assess the IgG content of the rat metrial gland at different stages of pregnancy. The result apparently varied according to the type of fixative used. Saturated alcoholic mercuric chloride was found to produce the most consistent demonstration, with the IgG located in cells which also contained diastase-fast, PAS-positive granules.Using single cell suspensions prepared from metrial glands, significantly more cells were shown to contain cytoplasmic IgG at day 13, compared to day 14 of pregnancy. However, there were no other significant differences at other stages of pregnancy examined. Surface IgG was detected on a small proportion of the cells and the findings are discussed in relation to the hypothesis of a lymphocytic origin for the granulated metrial gland cells.     

A monoclonal antibody, 4H12, recognizes a surface antigen found on granulated metrial gland cells in the murine decidua

A monoclonal antibody (MAb), designated 4H12, was selected for reactivity to a surface antigen on PYS-2 teratocarcinoma cells. 4H12 was the product of a fusion of lymphoid cells of a non-immunized pregnant C57BL/6 mouse to NS-1 myeloma cells. Initial studies utilizing immunohistochemistry revealed that MAb 4H12 bound to an antigen found on cells in the decidua basalis of 7-, 8- and 10-day pregnant mice. Antigen-positive cells of 11--19-day pregnant mice were also found predominantly in the decidua. A few antigen-positive cells were found in the labyrinth of the placenta and up against Reichert's membrane. Antigen-positive cells were morphologically and spatially distinct, oval to round with large periodic acid Schiff positive granules. Indirect immunofluorescent (IIF) labeling of decidual cultures showed antigen on the surface of cells that were small, oval to round and adherent. The antigen recognized by MAb 4H12 was removed from tissue sections with trypsin and protease and therefore is suggested to be a protein. We conclude that MAb 4H12 recognizes a surface antigen found on cells historically described as granulated metrial gland (GMG) cells. This MAb should greatly facilitate the further analysis of the life history and function of GMG cells during pregnancy.     

Tracing the glycogen cells with protocadherin 12 during mouse placenta development

Among the different trophoblast subtypes of the mouse placenta, the glycogen cells (GC) are one of the trophoblast subtypes that invade the decidua. We previously established that GC specifically expressed protocadherin 12 (PCDH12). In this paper, we investigated the origin of the PCDH12-positive cells and we characterized their fate in the maternal tissues. Our data indicate that they directly originate from the central part of the ectoplacental cone at embryonic day (E) 7.5. PCDH12-positive cells start to accumulate glycogen from E10.5 and the first migrating cells could be observed from this age. Unlike other placental and decidual cells, GC do not express P-cadherin, which may explain their migration properties in this organ. In the decidua, GC settle in the vicinity of the maternal vascular sinuses but do not incorporate in the endothelium. By the end of gestation (E17.5), most GC islets of the decidua enter into a lytic phase and form large lacunae. These lacunae, filled with glycogen, may provide a substantial source of energy at the end of gestation or during delivery. Our data suggest that spongiotrophoblasts and GC are two independent lineages and we bring insights into GC migration and fate.