Human endometrial MUC1 carries keratan sulfate: characteristic glycoforms in the luminal epithelium at receptivity.

MUC1 is a high molecular mass, highly glycosylated epithelial apical glycoprotein that has been shown to exhibit both adhesive and anti-adhesive properties. Its expression in human glandular endometrial epithelium is transcriptionally regulated with the highest levels in the mid secretory phase, the "receptive" period during which implantation occurs. We demonstrate that endometrial MUC1 carries highly sulfated lactosaminoglycan chains recognized by monoclonal antibody (Mab) 5D4, and the sialokeratan sulfate epitope recognized by Mab D9B1. These glycans are hormonally regulated in endometrium, and show increased abundance in the secretory phase, but detailed evaluation of their distribution shows important differences. The 5D4 epitope is abundant at the luminal epithelial surface until the implantation phase, when it disappears, first from patches of cells, then altogether. D9B1 binding sites are retained in the luminal epithelium at receptivity. These data show that endometrial MUC1 carries sulfated lactosaminoglycans. They identify the luminal epithelial compartment as a site of unique MUC1 glycosylation and independent regulation. Glycosylation and the negative charge associated with sialo- and sulfoglycans may be important in the regulation of embryo attachment.

Sialyl-Lewis x and Sialyl-Lewis a are associated with MUC1 in human endometrium.

Endometrial epithelial cells express MUC1 with increased abundance in the secretory phase of the menstrual cycle, when embryo implantation occurs. MUC1 is associated with the apical surface of epithelial cells and is also secreted, being detectable in uterine fluid at elevated levels in the implantation phase. However, its physiological role is uncertain; it may either inhibit intercellular adhesion by steric hindrance or carry carbohydrate recognition structures capable of mediating cell-cell interaction. Here we show that endometrial epithelium expresses both Sialyl-Lewis x (SLex) and Sialyl-Lewis a (SLea), with a distribution and pattern of menstrual cycle regulation similar to that of MUC1. Using Western blotting and double determinant ELISA of uterine flushings, we demonstrate that SLex is associated with MUC1 core protein. The endometrial carcinoma cell lines HEC1A and HEC1B are shown to express MUC1 in a mosaic pattern, while three other cell lines express much lower amounts. HEC1A expresses both SLex and SLea while HEC1B expresses SLea only. Immunoprecipitation has been used to demonstrate that SLea is associated with MUC1 in HEC1B cells, and both SLex and SLea are associated with MUC1 in HEC1A cells.

MUC1 as a cell surface and secretory component of endometrial epithelium: reduced levels in recurrent miscarriage.

The mucin MUC1 is a large, highly glycosylated, hormonally regulated product of endometrial glandular and luminal epithelium with both cell surface-associated and secreted isoforms. The abundance of mRNA coding for MUC1 increases about sixfold from the proliferative to the early secretory phase (Hey et al., J. Clin. Endocrinol. Metab. 78:337-342, 1994). Immunohistochemical studies show intracellular deposits accumulating in the early secretory phase followed by the release of MUC1 into gland lumens. The apical surface of luminal epithelium is strongly immunopositive in the early secretory phase. We have used a two site ELISA to measure MUC1 in uterine flushings as a function of time after the luteinising hormone (LH) peak. Low levels of secretory MUC1 are observed before day LH+7, while values on days LH+7-LH+13 are much higher. Using semi-quantitative immunohistochemical methods we have shown that in women suffering recurrent spontaneous miscarriage, mid secretory phase levels of MUC1 core protein and mucin-associated glycans are reduced (Serle et al., Fertil. Steril. 62:989-996, 1994). Similarly, lower core protein levels are observed in uterine flushings after day LH+7 in these women. Reduced epithelial secretory function and a resultant change in uterine fluid composition are features of endometrium from recurrent miscarriage patients.

Laminins 2 and 4 are expressed by human decidual cells.

During pregnancy, the resident stromal cells of the endometrium differentiate to become decidual cells and produce a pericellular basement membrane. We used immunofluorescence and Western blotting with a panel of monoclonal Ab specific for various laminin subunits to examine the composition of decidual laminin. The stromal cell basement membrane contained subunits alpha 2 (M), beta 1 (B1), beta 2 (S), and gamma 1 (B2). Low levels of alpha 1 could also be detected. The glandular and vascular basement membranes of decidual tissue contained subunits alpha 1 (A), beta 1, and gamma 1. An extract was produced from decidual extracellular matrix. Western blots of nonreducing gels showed the presence of high molecular weight complexes containing alpha 2, beta 1, beta 2, and gamma 1. These data indicated that laminins 2 and 4 are coexpressed by decidual cells. Laminin 1 was present in the extract as a minor component. In contrast, cultured stromal cells expressed laminin 1 as the major secreted variant. Immunolocalization was carried out using tissue from various stages of the nonpregnant cycle. The alpha 2 chain polypeptide was absent in the proliferative phase of the cycle but present in late secretory phase in perivascular areas where predecidual differentiation occurs. Reverse transcriptase-PCR experiments confirmed the presence of alpha 2 chain mRNA in decidua but showed that this transcript is detectable throughout the nonpregnant cycle. The results showed that laminins 2 and 4 are hormonally regulated products of decidual cells. The composition of the vascular and epithelial basement membranes remained constant throughout the cycle.

MUC1 in secretory phase endometrium: expression in precisely dated biopsies and flushings from normal and recurrent miscarriage patients.

MUC1 is a cell-surface and secretory product of endometrial epithelium. Immunohistochemical studies carried out using two different antibodies to the mucin-type tandem repeat region of MUC1 indicate a cell-surface location in proliferative phase glands, with intracellular deposits accumulating in the early secretory phase. Commencing 3-4 days after the luteinizing hormone (LH) peak and continuing into the late secretory phase, secretory MUC1 appears in gland lumens. Uterine flushings were collected as a function of time after the LH peak and were analysed using a two-site enzyme-linked immunosorbent assay for MUC1. Low but measurable concentrations were observed up to day 7, while on days 7-13 much higher values were obtained. In women suffering from recurrent spontaneous miscarriage, the concentration of MUC1 in flushings was significantly lower than in the controls on day LH + 10. Lower values were observed on days 7 and 13. Reduced epithelial secretory function and a resultant change in uterine fluid composition are features of endometrium from recurrent miscarriage patients.

The endometrial cell surface and implantation. Expression of the polymorphic mucin MUC-1 and adhesion molecules during the endometrial cycle.

The cell surface mucin MUC-1 is present in endometrial epithelial cells and their associated apical glycocalyx and is also released into gland lumens as a secretory product. MUC-1 mRNA and core protein are found at low levels in the proliferative phase of the cycle, but their abundance increases after ovulation. Endometrial MUC-1 has been found to carry sialokeratan sulphate chains and these show a dramatically increased abundance in cells and secretions in the post-ovulatory phase of the cycle, reaching a maximum in secretions 6-7 days after the LH peak. The apical epithelium also contains adhesion receptor molecules of the integrin and CD44 families. MUC-1 is large and highly glycosylated and probably extends farther from the cell surface than these 'conventional' glycoprotein receptors. It has the potential to inhibit sterically receptor-mediated cell-cell adhesion. However, it is also possible that MUC-1 displays specific (e.g., glycan) recognition structures for the initial attachment of the blastocyst or that the embryo may create a specialised microenvironment in which to implant.

The polymorphic epithelial mucin MUC1 in human endometrium is regulated with maximal expression in the implantation phase.

After ovulation, progesterone stimulates a temporally regulated secretory transformation in human endometrial epithelium. Using a combination of immunohistochemistry, and Western and Northern blotting, we demonstrate that 1) the polymorphic epithelial mucin MUC1 is secreted by human endometrial epithelium; 2) low levels of both mRNA and core protein are present in the preovulatory phase of the menstrual cycle; 3) mRNA levels increase several-fold after ovulation, consistent with transcriptional regulation by progesterone; 4) there is an increase in translation product in postovulatory endometrium; and 5) the tandem repeat domain of the MUC-1 polypeptide is glycosylated in endometrium.

C-myc oncogene product P62c-myc in ovarian mucinous neoplasms: immunohistochemical study correlated with malignancy.

The monoclonal antibody Myc 1-6E10 was used to determine the cellular distribution of the c-myc oncogene product p62c-myc in 60 mucinous ovarian tumours. Three patterns of immunostaining were apparent: (i) nuclear staining alone; (ii) staining of the nucleus and basal cytoplasm; and (iii) staining of the entire cell. Of the 21 cases of mucinous cystadenoma, 11 showed nuclear staining alone, and a further case showed additional weak staining of the basal cytoplasm. Nuclear staining alone was not present in any of the 17 borderline mucinous tumours examined. Strong staining of the nucleus and basal cytoplasm was seen in 16 of these borderline cases, six of which also showed focal staining of the apical cytoplasm. All 22 cases of mucinous cystadenocarcinoma showed staining of the cell nucleus and entire cell cytoplasm. Focal staining of the apical cytoplasm in six of 17 borderline mucinous tumours produced a pattern of c-myc immunostaining similar to that of cystadenocarcinoma. Retrospective analysis of the clinical data showed that no significant differences between patients with borderline tumours of these two categories could be defined. Although immunostaining with Myc 1-6E10 can be used in the categorisation of mucinous ovarian tumours, it is concluded that standard histological criteria are more accurate indicators of tumour behaviour than is an assessment of c-myc expression.