The metrial gland in the rat and its similarities to granular cell tumors.

Metrial glands are normal structures located in the mesometrial triangle of the pregnant rat uterus from gestational day (GD) 8 through termination of pregnancy. Metrial glands are composed of a dynamic mixed cell population of granulated metrial gland (GMG) cells, endometrial stromal cells, trophoblasts, blood vessels, and fibroblasts. Collections of similar cells may be seen in association with pseudopregnancy and other hormonal disturbances. Granulated metrial gland cells are the hallmark cell of the metrial gland. They are bone-marrow-derived, perforin-positive, natural killer cells that proliferate in the pregnant uterus. Understanding the normal histogenesis of the metrial gland and recognizing the possible existence of GMG cells and a reactive metrial gland in the nonpregnant state are important when examining any uterine lesion that contains granulated cells. This report demonstrates that the cellular composition, morphology, and immunohistochemical staining profile of normal metrial glands are similar to reported granular cell neoplasms in rats and mice. The possibility of a non-neoplastic lesion involving the metrial gland should be considered when proliferative lesions involving granulated cells are observed in the uterus of mice and rats from nonclinical toxicity studies. Positive immunohistochemical staining for perforin and S100 would assist in the classification of such lesions as a reactive metrial gland or decidual reaction.

Mode of cell death in the rat metrial gland during peripartum regression.

In the rodent uterus, the metrial gland develops during midpregnancy and undergoes regression prior to parturation. The involution of the gland is reported to be accompanied by the loss of gland cells due to their death in situ. Cell death has been classified by using morphological criteria into two types: necrosis and apoptosis. To study the mechanism involved in the peripartum regression of the rat metrial gland, we examined the mode of cell death in the gland during the last week of gestation. We identified apoptotic cells in the regressing metrial gland by using DNA fragmentation, in situ DNA 3'-end labeling, and electron microscopy. Expression of progesterone receptor (PR) and estrogen receptor (ER) was also demonstrated by immunohistochemistry in the gland. The mean weight of metrial gland nodes decreased after day 18 of pregnancy. The apoptotic granulated metrial gland (GMG) cells that were detected by using the in situ DNA 3'-end labeling method were observed on day 16 of pregnancy, and they increased in number after day 20 of pregnancy. Intense fragmentation of DNA was also found from day 20 to day 22 of pregnancy. Electron microscopy demonstrated apoptotic GMG cells in the regressing metrial glands, confirming the results of the labeling studies. Immunohistochemical study revealed that expression of PR and ER, which were localized mainly in fibroblast-like stromal cells but not in GMG cells, was almost unchanged during late pregnancy. Apoptotic cell death is the major mode of rat metrial gland cell death in the peripartum loss of metrial gland cells.

Lectin histochemical studies of mouse granulated metrial gland cells.

A lectin histochemical study has been carried out on mouse granulated metrial gland cells, the major leucocyte population that differentiates in the uterine wall in pregnancy. The binding characteristics of 26 lectins were examined using light microscopical methods. Fourteen of the lectins, with affinities ranging through N-acetylgalactosamine, galactose, N-acetylglucosamine, mannose and sialic acid residues, bound to the cytoplasmic granules of granulated metrial gland cells, and each appeared to bind to the limiting membrane of the granules. The binding characteristics of three of these lectins (Wheat germ agglutinin, Concanavalin A and Helix pomatia agglutinin) were examined using electron microscopical methods. These showed a different binding pattern to the cytoplasmic granules of granulated metrial gland cells compared with that found using light microscopical methods, as they appeared to bind evenly across the granule's matrix. This binding pattern corresponds to the reactivity of the granule matrix in the periodic acid-Schiff technique. Six lectins bound to the cell membranes of granulated metrial gland cells. These included the E and L isoforms of Phaseolus vulgaris agglutinin, with affinities for complex carbohydrates, whose binding differences were related to the stage of differentiation of the granulated metrial gland cells. The lectin binding described presents additional markers of granulated metrial gland cells and tools for investigating carbohydrate moieties in the functional activities of granulated metrial gland cells.

Lectin histochemistry of pregnant rat uterine tissues.

Glycoconjugate residues were examined in the rat uterus at days 10, 12 and 15 of pregnancy using 17 biotinylated lectins with specificities for a variety of carbohydrate moieties. A wide variety of glycoconjugate residues were detected in the cytoplasm of some antimesometrial and mesometrial decidual cells with lectins from Canavalia ensiformis (Con A), Lens culinaris (LcH) culinaris (LcH), Pisum sativum (PSA), Griffonia simplicifolia II (GS-II), Triticum vulgaris (WGA), Griffonia simplicifolia I (GS-I), Maclura pomifera (MPA) and Phaseolus vulgaris (PHA-E and PHA-L). Reactivity with some of these lectins was also pericellular and the glycoconjugate residues detected may be related to extracelluar matrix produced by decidual cells. Reactivity with PHA-E, demonstrating complex carbohydrates, was most intense in the mesometrial decidua closest to the trophoblastic giant cells and may demonstrate glycoconjugates involved in maintaining integrity at this maternotrophoblast interface. Lectins derived from Helix pomatia (HPA), Vicia villosa (VVA), Glycine max (SBA) and Arachis hypogaea (PNA) reacted only with occasional small cells in antimesometrial or mesometrial decidua, probably demonstrating alpha and/ or beta-linked N-acetylgalactosamine residues on lymphocytes or monocytes. The glycoprotein granules of granulated metrial gland (GMG) cells in the decidua basalis and the metrial gland reacted strongly with WGA, MPA and PHA-L demonstrating complex carbohydrates including sialic acid residues and tri/tetra-antennary, nonbisected N-linked glycans. GMG cell cytoplasm reacted diffusely with Con A, LcH, PSA, WGA, PHA-E and PHA-L. In some GMG cells reactivity with WGA was apparently localised to the Golgi region indicating involvement in granule formation. Pericellular reactivity of some GMG cells with WGA may relate to migratory activity. The lectin reactivity of fibroblast-like stromal cells in the material gland was similar to that of the extracellular matrix and it is likely that many matrix molecules are produced by the stromal cells. A range of lectins (Con A, LcH, PSA, GS-II GS-I and PHA-L) reacted with some blood vessels in the metrial gland at day 12 of pregnancy: this may indicate activation changes associated with the transendothelial passage of GMG cells at this stage. Lectins derived from Dolichos biflorus (DBA), Bauhinia purpurea (BPA), Lotus tetragonolobus (LTA) and Ulex europaeus-I (UEA-I) did not react with decidual and metrial gland regions. Lectin binding profile studies can provide further information on the events occurring in the uterus in pregnancy: investigations at the ultrastructural level are required to resolve further intra and extracellular changes.

Perforin-expressing cells during spontaneous abortion.

The metrial gland of pregnant rodents contains an abundant population of natural killer (NK)-like cells called granulated metrial gland (GMG) cells. Since GMG cells express the cytolytic protein, perforin, and since cells with NK activity have been implicated in spontaneous abortions, we have studied the distribution of perforin-containing cells in the uterus of mice undergoing normal pregnancy (Swiss mice) and spontaneous abortions (CBA/J x DBA/2 mice). The distribution of perforin-positive GMG cells was essentially the same near healthy and aborting conceptuses, suggesting that GMG cells are not involved in most cases of spontaneous abortion in this abortion model. Small perforin-positive cells were observed near the aborting conceptus in about 5% of the cases. However, it is not known whether their presence was the result or the cause of abortion.

Perforin-expressing granulated metrial gland cells in murine deciduoma.

It has previously been shown that granulated metrial gland (GMG) cells of the pregnant uterus express abundant quantities of the lymphocyte pore-forming protein, perforin. No perforin was present before implantation of the embryo, but large numbers of perforin-producing GMG cells were observed after implantation, which coincides with decidualization of the uterus. The possible source of the activation factors responsible for perforin gene induction in GMG cells was studied here with the pseudopregnancy model, in which cervical stimulation of mice during estrus leads to a series of hormonal changes resembling those seen in pregnancy, but in the absence of an embryo. Subsequent stimulation of the uterus of pseudopregnant mice with oil causes the stimulated portion of the endometrium to differentiate into decidual tissue. Perforin-containing GMG cells were in fact present in the deciduomata, but not in adjacent nondecidualized tissues of the same mice. These results suggest that maternal factors associated with decidual tissue are responsible for the local expression of perforin in GMG 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.