Immunophenotypic properties and estrogen dependency of budding cell structures in the developing mouse mammary gland

Abstract. The initial phase of growth of the parenchymal component of the mouse mammary gland is ductal clongation, which is mainly accomplished by proliferating cells in a specialized structure termed end bud. End buds are composed of multiple layers of epithelial cells (so called body cells) which are capped by a single layer of morphologically unique cells termed cap cells.
We sought to examine the interrelationship between cap cells and other epithelial cell subclasses using a variety of antibodies to different keratin proteins and also antibodies to vimentin, actin and collagen IV. An extensive immunohistochemical characterization of the epithelial components of the developing and differentiating mammary gland demonstrated that cap cells were devoid of any immunohistochemically - detectable keratins but were positive for collagen IV. In contrast, the majority of cells in the end bud along with the luminal epithelial and myoepithelial cells were keratin positive. The body cells of the end bud were the only cells which were positive for antibody to keratin 6, a keratin which previously has been reported to be expressed in proliferating mammary epithelial cells. In addition, estrogen receptor was localized only to epithelial cells of ducts, alvcoli and body cells of end buds, but not to cap cells or myoepithelial cells. We interpret these results to suggest that cap cells are not totpotent stem cells but rather cells specialized in paving the way for ductal elongation as well as serving as precursors to myoepithelial cells.     

The role of cellular migration in the repair process of gastric epithelial cells.

The epithelial cells of stomach are continuously exposed to various toxic agents that may cause mucosal injury. The epithelial lining is rapidly replaced by cells that migrate from the proliferative zone of the gastric gland, to maintain the integrity of the gastric mucosa. Thus, cell migration is an essential part of the early process of gastric mucosal repair. After various forms of gastric injury, mucosal integrity is reestablished by the rapid migration of epithelial cells. However, the cellular mechanisms of the restitution remain unclear to date. In this report, we will review the role of cellular migration in the repair process of gastric epithelial cells in culture. It has been reported that hepatocyte growth factor (HGF) has the potency of acceleration of cellular repair process. In this review, we also report that HGF plays a leading role in the mucosal repair after damage by using a novel cell culture model.     

Ultrastructure of surface epithelial cells of the normal human rectum

Luminal surface epithelial cells, excluding a few endocrine cells of the normal human rectum, were studied electron microscopically and 5 types of cells were recognized with special reference to some structure containing mucous substances. Principal-1 cells showing few tiny vesicles and Principal-2 cells containing some tiny vesicles seemed to belong to the absorptive cell group. Vesicle cells having numerous tiny vesicles, and Columnar mucous cells accompanied by numerous tiny vesicles and some round or oval mucous vacuoles, seemed to be labelled as of the secretory cell group. The common features of the epithelial columnar cells, except for the Goblet cell, were columnar shape, microvilli whose length and density had considerable variation, glycocalyceal bodies around the microvilli, and thick surface coat. Goblet cells were characterized by a goblet shape which was expanded by numerous mucous droplets. It is of special interest that 4 different types of columnar epithelial cells are recognized on the luminal surface of the normal human rectum, and that Vesicle cells and Columnar mucous cells are first observed on the luminal surface of the large intestine. Similar epithelial cells have only been reported in the crypt of the large intestine and not on the luminal surface.     

Cultured human corneal epithelial stem/progenitor cells derived from the corneal limbus

Stem/progenitor cells of the human corneal epithelium are present in the human corneal limbus, and several corneal epithelial stem/progenitor cell markers have been reported. Recently, the neurotrophin family receptors were reported to be useful markers of corneal epithelial stem/progenitor cells. Therefore, we examined an enzymatic separation method for obtaining corneal epithelial stem/progenitor cells and measuring the change in the expression of low-affinity neurotrophin receptor p75 (p75^NTR), a receptor belonging to the neurotrophin family. As a result, it was found that our separation method preserved cell viability. Furthermore, p75^NTR was mainly observed in epithelial basal cells as were the corneal epithelial stem/progenitor markers p63 and integrin β1. p75^NTR was also observed in the cultured cells, but its frequency decreased with passage. In conclusion, we propose that our culture method will enable the culture of corneal stem cells and that it is a useful tool for elucidating the molecular basis of the niche that is necessary for the maintenance of epithelial stem cells in the corneal limbus. Furthermore, we conclude that p75^NTR is a useful cell marker for evaluating the characteristics of stem/progenitor cells in culture.     

Neuroendocrinology of the thymus

The neuropeptides oxytocin (OT) and vasopressin (VP) are synthesized in the human thymus in a similar way as in the hypothalamo-neurophypophyseal system. Immunocytochemistry with polyclonal and monoclonal antibodies revealed that immunoreactive OT- and VP-producing cells are localized in the subcapsular cortex and medulla of human and murine thymuses. The epithelial nature of the neuroendocrine thymic cells is demonstrated by their immunostaining with a monoclonal antibody against cytokeratin. An original example of a neuroendocrine-immune microenvironment is given by the thymic nurse cells which are composed of a large neuroendocrine epithelial cell enclosing numerous mitotic immature thymocytes. These observations and the previously reported mitogenic and immunomodulatory properties of VP and OT upon mature T cells and thymocytes strongly support the existence of a neuroendocrine thymo-lymphoid axis and an active role of thymic VP and OT in T cell differentiation and activation.     

The glycoconjugate sugar residues of the sessile and motile cells in the thymus of normal and cyclosporin-A-treated rats: lectin histochemistry

It is well known that cell surface glycoconjugates play a determinant role in cellular recognition, cell-to-cell adhesion and serve as receptor molecules. T-lymphocytes are in strict contact with the thymic epithelial cells, which control their process of maturation and proliferation. On the other hand the normal maturation of the epithelial cells is believed to be induced by T-lymphocytes. For these reasons we have studied the glycoconjugates saccharidic moieties of the sessile and motile cells in the thymus of normal male albino Wistar rats and their changes following cyclosporin-A treatment, using a battery of seven HRP-lectins. Cytochemical controls were performed for specificity of lectin-sugar reaction. Some sections were pre-treated with neuraminidase prior to staining with HRP-lectins. Our results have demonstrated, in the control rats, a large amount and a variety of terminal and subterminal oligosaccharides within and/or on the epithelial thymic cells and in macrophages. After cyclosporin-A treatment, among the thymic epithelial cells, the subcapsular, paraseptal and perivascular cells showed the loss of some sugar residues, which characterized the same cells in the intact thymus. Some hypotheses are reported on the role played by the glycoconjugate sugar residues in control and cyclosporin-A treated rats.     

Ultrastructural localization of glucose-6-phosphatase in renal glomerulus of the adult dog

The ultrastructural distribution of glucose-6-phosphatase activity was investigated in renal glomeruli of adult dogs by electron-microscopic cytochemistry. The enzymatic activity was found mainly in the parietal epithelial cells of Bowman's capsule. Weaker activity occurred in visceral epithelial cells. No activity was found in either the endothelial or the mesangial cells. Strong activity of glucose-6-phosphatase was commonly found in the nuclear envelope, and occasionally in the rough endoplasmic reticulum. The distribution of the enzyme and its functional significance are discussed in relation to previously reported data.     

Recently identified a novel neuropeptide manserin colocalize with the TUNEL-positive cells in the top villi of the rat duodenum.

We recently isolated a novel 40 amino acid neuropeptide designated manserin from the rat brain. Manserin is derived from secretogranin II, a member of granin acidic secretory protein family by proteolytic processing, as previously reported secretoneurin and EM66. Manserin peptide are localized in the endocrine cells of the pituitary. In this study, we further investigated the manserin localization in the digestive system by immunohistochemical analysis using antimanserin antibody. In the duodenum, manserin immunostaining was exclusively observed in the nuclei of top villi instead of cytosol as observed in neurons in our previous study. Interestingly, manserin-positive cells in the duodenum are colocalized with terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) positive cells, the cells whose DNA was damaged. Since the top villi of duodenum epithelial cells are known to undergo spontaneous apoptosis during epithelial cell turn over, and since other peptides such as secretoneurin and EM66 derived from SgII have been reported to be cancer-related, these results indicated that manserin peptide may have a role in apoptosis and/or cancer pathogenesis in the digestive organ.     

Effects of cholinergic agonists on the proliferation and protein synthesis in a cultured thymic epithelial cell line

Thymic epithelial cells appear to release the humoral factors endowing precursors of T cells (thymus-dependent lymphocytes) with the capacity to differentiate and maturate into relatively mature T cells. We have separated the polypeptide fractions containing these factors from the culture supernatant of thymic epithelial cell line. Thymus is reported to be innervated by autonomic nervous system from the prenatal to the pubertal period. But the physiological significance of the nervous system in this lymphoid organ remains obscure. And the modulator of the epithelial cell functions, namely, production and release of the bioactive polypeptides have never been clarified. We show here that acetylcholine (Ach) or carbamylcholine (Cch) enhanced the proliferation of thymic epithelial cells from the TAD3 cell line at preconfluent state, and the protein synthetic activity at confluent state. This phenomenon was completely suppressed by the pretreatment of alpha-bungarotoxin (alpha-BTx). These results suggest that nicotinic Ach-receptors exist on the epithelial cell surface membrane and that the differentiation and maturation of thymic lymphocytes are indirectly regulated by the activated functions of thymic epithelial cells stimulated with cholinergic agonists.     

Requirement of ZO-1 for the formation of belt-like adherens junctions during epithelial cell polarization

The molecular mechanisms of how primordial adherens junctions (AJs) evolve into spatially separated belt-like AJs and tight junctions (TJs) during epithelial polarization are not well understood. Previously, we reported the establishment of ZO-1/ZO-2-deficient cultured epithelial cells (1[ko]/2[kd] cells), which lacked TJs completely. In the present study, we found that the formation of belt-like AJs was significantly delayed in 1(ko)/2(kd) cells during epithelial polarization. The activation of Rac1 upon primordial AJ formation is severely impaired in 1(ko)/2(kd) cells. Our data indicate that ZO-1 plays crucial roles not only in TJ formation, but also in the conversion from "fibroblastic" AJs to belt-like "polarized epithelial" AJs through Rac1 activation. Furthermore, to examine whether ZO-1 itself mediate belt-like AJ and TJ formation, respectively, we performed a mutational analysis of ZO-1. The requirement for ZO-1 differs between belt-like AJ and TJ formation. We propose that ZO-1 is directly involved in the establishment of two distinct junctional domains, belt-like AJs and TJs, during epithelial polarization.     

The development of rat palatal shelves in vitro. An ultrastructural analysis of the inhibition of epithelial cell death and palate fusion by the epidermal growth factor.

Unfused palatal shelves from day-15 rat embryos were cultured with their medial edges in contact. During the next 20 hr epithelial surfaces of apposing shelves adhered, and the epithelial cells in this area degenerated and were replaced by mesenchymal cells in a manner similar to that reported for palate fusion in vivo in the rat and in vitro in the mouse. Four hours after contact the superficial cells of apposing palates adhered with a 10–25-nm distance separating cell membranes while after 20 hr these cells were either eliminated or had lost their flattened appearance and become indistinguishable from the cuboidal-shaped cells adjacent to the basement membrane. The ratio of cytoplasm to nucleoplasm was reduced, and degenerated cells were eliminated by macrophages.Addition of epidermal growth factor (EGF) to the culture medium inhibited epithelial adhesion and degeneration. Basal and superficial cells retained their original appearance, while the ratio of cytoplasm to nucleoplasm was increased. Numerous desmosomes and tonofilaments were evident as well as elaborate membrane foldings. The appearance of these cellular elements is similar to that observed in keratinizing palatal epithelial cells from the oral region. These results suggest that EGF induces the epithelial cells to keratinize, and as a consequence the normal sequence of steps of palate fusion are inhibited.     

Expression of actin isoforms in developing rat intestinal epithelium

A minimum of six very similar but distinct actin isoforms are encoded by the mammalian genome. Developmental regulation of these genes results in a tissue-specific distribution of the isoforms in the adult. Using a panel of actin specific monoclonal antibodies (MAb), we recently reported the expression of two unique actin isoforms in adult rat intestinal brush border. In this report, we examine the developmental expression of these and other actin isoforms in rat intestinal epithelial cells. Isoforms containing the HUC 1-1 and/or C4 epitopes are present by day 15 of gestation and are continuously expressed throughout adult life. Unexpectedly, the gamma-enteric smooth muscle isoactin, defined by the B4 epitope, is transiently expressed in these non-muscle cells late in gestation. The alpha-vascular smooth muscle isoform, however, is not expressed in intestinal epithelial cells during development and, as previously reported, both smooth muscle isoforms are absent in epithelial cells of adult intestine. In addition, we demonstrate that although multiple isoforms are expressed simultaneously in these cells, they are not uniformly distributed at the subcellular level, suggesting that the cell recognizes the actin isoforms as functionally distinct entities.     

Identification of novel epithelial stem cell-like cells in human deciduous dental pulp

It is well known that interactions between epithelial components and mesenchymal components are essential for tooth development. Therefore, it has been postulated that both types of stem cells might be involved in the regeneration of dental hard tissues. Recently, mesenchymal dental pulp stem cells that have odontogenic potential were identified from human dental pulp. However, the existence of epithelial cells has never been reported in human dental pulp. In the present study, we isolated and characterized epithelial cell-like cells from human deciduous dental pulp. They had characteristic epithelial morphology and expressed epithelial markers. Moreover, they expressed epithelial stem cell-related genes such as ABCG2, Bmi-1, ΔNp63, and p75. Taken together, our findings suggest that epithelial stem cell-like cells might exist in human deciduous dental pulp and might play a role as an epithelial component for the repair or regeneration of teeth.     

Intermediate filaments of the lung

Intermediate filaments (IF), a subfamily of the cytoskeletal filaments, provide structural support to cells. Human diseases related to mutations in IF proteins in which their tissue-specific expression is reflected have been found in a broad range of patients. The properties of identified IF mutants are well-studied in vitro in cultured cells and in vivo using transgenic mice expressing IF mutants. However, the association of IF proteins with diseases of the lung is not fully studied yet. Epithelial cells in normal lung express vimentin and various keratins, and the patterns of their expression are altered depending on the progression of the lung diseases. A growing number of studies performed in alveolar epithelial cells demonstrated IF involvement in disease-related aspects including their usefulness as tumor marker, in epithelial-mesenchymal transition and cell migration. However, the lung disease-associated IF functions in animal models are poorly understood, and IF mutations associated with lung diseases in humans have not been reported. In this review, we summarize recent studies that show the significance of IF proteins in lung epithelial cells. Understanding these aspects is an important prerequisite for further investigations on the role of lung IF in animal models and human lung diseases.     

First evidence of the interaction between deleted in malignant brain tumor 1 and galectin-3 in the mammalian oviduct

The oviduct supports the transport and final maturation of gametes, and harbors fertilization and early embryo development. The oviductal epithelium is responsible for providing the correct environment for these processes. Deleted in malignant brain tumor 1 (DMBT1) is expressed by multiple organisms and several cell types, and the interaction of the rabbit ortholog of DMBT1 with galectin-3 (gal-3) modulates the polarity of epithelial cells. This interaction has not yet been shown in locations other than rabbit kidney and human-cultured endothelial cells. DMBT1 and gal-3 also protect epithelial layers from pathogens and trauma, and are innate immunity components. DMBT1 has been detected in the porcine oviduct, and gal-3 has been reported in the Fallopian tube and in the cow oviduct. Interaction between both proteins would show a probable physiological function in the female reproductive tract. This work describes the presence and co-localization of DMBT1 and gal-3 mainly in the apical region of the epithelial cells of the Fallopian tube and the porcine oviduct, and co-immunoprecipitation in membrane-enriched epithelial cell extracts from the porcine oviduct. The findings strongly support a functional interaction in the mammalian oviduct, suggestive of a role on epithelial protection and homeostasis, which might be related to epithelium–gamete interaction.     

Properties of the esophageal epithelium in relation to organizative patterns of lymphoid infiltrations in the red-eared turtle

The esophagus of the turtle, like the mucosal surfaces in other species, contains variously sized areas of lymphoid infiltration. The tunica propria and the surface epithelial layer of this area are invaded by the lymphoid cells. The features of the layer of epithelial cells which cover the lymphoid infiltrations are of a special kind: they do not possess vibratile cilia and are able to take up materials flowing into the lumen. The present paper contains further information concerning lymphoid infiltration obtained by histological and histochemical methods. The epithelial layer covering the lymphoid infiltrations is composed of cells with irregularly distributed microvilli, ciliated cells and mucous-secreting cells. After administration of silica and colloidal carbon, the microvillar epithelial cells proved to have these substances inside them, thereby accounting for the pinocytotic activity. The absorbing epithelial cells were not damaged by silica which is a macrophage-toxic agent, while the underlying macrophages are damaged. These results are compared with the features of lymphoid infiltration associated cells in various organs and animals; the hypothesis is proposed that these cells in the esophagus of turtles may originate from the covering epithelial cells.