Secretory Morphology Research Articles

Comparison of the structural and ultrastructural characteristics of the female prostate between pregnant and non-pregnant plains viscacha (Lagostomus maximus)

Some years ago, our group reported the presence of the female prostate in all the studied females of the plains viscacha (Lagostomus maximus). The goal of the present study was to characterize and compare the female prostate gland between adult pregnant and non-pregnant plains viscacha using histochemical, lectin-histochemical and immunohistochemical techniques, as well as optic and electron microscopy. Structurally, alveoli are lined by a simple epithelium formed by different cell types: basal cells, secretory cells in different stages of the secretory cycle and cells of clear cytoplasm. Secretory cells are the most abundant cell type, differing between them depending on the quantity and electron-density of their granules. The basal cells are less abundant and are responsible for the renewal of the alveolar epithelium. Likewise, other cells with secretory morphology were found in all the studied females; these have a clear cytoplasm, few granules and mitochondria. It could be considered that they are degranulated secretory cells or that they have partially released their granules. The stroma of the organ is formed by connective tissue and smooth muscle fibers, which are immunohistochemically evidenced against desmin. Histochemical and lectin-histochemical analysis revealed the presence of different glucidic residues in the different cell types. No structural, histochemical, lectin-histochemical, and ultrastructural differences were observed between pregnant and non-pregnant females of plain viscachas, except for the expression of some lectins. The paraurethral gland of Lagostomus maximus can be used as a model for studying the gland in other species since its structural and ultrastructural characteristics do not depend on the hormonal status of the female.

Chronic hyperandrogenemia in the presence and absence of a western-style diet impairs ovarian and uterine structure/function in young adult rhesus monkeys.

Does chronic hyperandrogenemia beginning at menarche, in the absence and presence of a western-style diet (WSD), alter ovarian and uterine structure-function in young adult rhesus monkeys? Phenotypic alterations in ovarian and uterine structure/function were induced by exogenous testosterone (T), and compounded in the presence of a WSD (T+WSD). Hyperandrogenemia is a well-established component of PCOS and is observed in adolescent girls, indicating a potential pubertal onset of disease symptoms. Obesity is often associated with hyperandrogenemia and it is hypothesized that metabolic dysfunction exacerbates PCOS symptoms. Macaque females (n = 40) near the onset of menarche (~2.5 years of age) were assigned to a 2 by 2 factorial cohort design. Effects on reproductive characteristics were evaluated after 3 years of treatment. Rhesus macaques (Macaca mulatta) were fed either a normal balanced diet (n = 20) or a WSD (n = 20). Additionally, implants containing cholesterol (n = 20) or T (n = 20) were implanted subcutaneously to elevate serum T approximately 5-fold. This resulted in treatment groups of controls (C), T, WSD and T+WSD (n = 10/group). Vaginal swabbing was performed daily to detect menses. After 3 years of treatment, daily serum samples from one menstrual cycle were assayed for hormone levels. Ovarian structure was evaluated in the early follicular phase by 3D/4D ultrasound. Uterine endometrial size and ovarian/luteal vascular function was also evaluated in subgroups (n = 6/group) in the late follicular and mid-luteal phases by 3D/4D ultrasound and contrast-enhanced ultrasound, respectively. Expression of steroid hormone receptors and markers of decidualization and endometrial receptivity were assessed in endometrial biopsies at mid-luteal phase. Approximately 90% of menstrual cycles appeared ovulatory with no differences in frequency or duration between groups. Serum estradiol (E2) levels during the early follicular phase were greatest in the T alone group, but reduced in T+WSD (P < 0.02). Serum LH was elevated in the T group (P < 0.04); however, there were no differences among groups in FSH levels (P > 0.13). Ovarian size at menses tended to be greater in the WSD groups (P < 0.07) and antral follicles ≥1 mm were more numerous in the T+WSD group (P < 0.05). Also, females in T and T+WSD groups displayed polycystic ovarian morphology (PCOM) at greater frequency than C or WSD groups (P < 0.01). Progesterone (P4) levels during the luteal phase were reduced in the T+WSD group compared to C and T groups (P < 0.05). Blood volume (BV) and vascular flow (VF) within the corpus luteum was reduced in all treatment groups compared to C (P < 0.01, P = 0.03), with the WSD alone group displaying the slowest BV and VF (P < 0.05). C and WSD groups displayed endometrial glands at mid-luteal phase with low estrogen receptor 1 (ESR1) and progesterone receptor (PGR) mRNA and immunohistochemical staining in the functionalis zone, but appreciable PGR in the stroma. In contrast, T and T+WSD treatment resulted in glands with less secretory morphology, high ESR1 expression in the glandular epithelium and low PGR in the stroma. Endometrial levels of TIMP3 and MMP26 mRNA and immunostaining were also decreased in the T and T+WSD groups, whereas AR expression was unchanged. None. Females are young adults, so effects could change as they reach prime reproductive age. The T level generated for hyperandrogenemia may be somewhat greater than the 3-4-fold increase observed in adolescent girls, but markedly less than those observed in male monkeys or adolescent boys. Alterations to ovarian and uterine structure-function observed in T and, in particular, T+WSD-treated female macaques are consistent with some of the features observed in women diagnosed with polycystic ovary syndrome (PCOS), and suggest impaired fertility. Research reported in this publication was supported by the Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD) of the National Institutes of Health (NIH) under Award Number P50HD071836 (to RLS). The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Additional funding was provided by Office of the Director, NIH under Award Number P51OD011092 (Support for National Primate Research Center). Authors declare no competing interests.

A single transcription factor is sufficient to induce and maintain secretory cell architecture.

We hypothesized that basic helix-loop-helix (bHLH) MIST1 (BHLHA15) is a "scaling factor" that universally establishes secretory morphology in cells that perform regulated secretion. Here, we show that targeted deletion of MIST1 caused dismantling of the secretory apparatus of diverse exocrine cells. Parietal cells (PCs), whose function is to pump acid into the stomach, normally lack MIST1 and do not perform regulated secretion. Forced expression of MIST1 in PCs caused them to expand their apical cytoplasm, rearrange mitochondrial/lysosome trafficking, and generate large secretory granules. Mist1 induced a cohort of genes regulated by MIST1 in multiple organs but did not affect PC function. MIST1 bound CATATG/CAGCTG E boxes in the first intron of genes that regulate autophagosome/lysosomal degradation, mitochondrial trafficking, and amino acid metabolism. Similar alterations in cell architecture and gene expression were also caused by ectopically inducing MIST1 in vivo in hepatocytes. Thus, MIST1 is a scaling factor necessary and sufficient by itself to induce and maintain secretory cell architecture. Our results indicate that, whereas mature cell types in each organ may have unique developmental origins, cells performing similar physiological functions throughout the body share similar transcription factor-mediated architectural "blueprints."

Cellular Evidence of Exosomes in the Reproductive Tract of Chinese Soft-Shelled Turtle Pelodiscus sinensis.

The oviduct is a location of egg production, fertilization, and sperm storage. While its secretions have broadly attributes toward different physiological functions. We examined the ultrastructure of oviduct epithelium and glands in relation to the secretions, particularly with exosomes origin in Chinese soft-shelled turtle Pelodiscus sinensis using immunohistochemistry and transmission electron microscopy. The ciliated epithelial and gland cells were involved in the release of exosomes and secretions into lumen throughout the year. The exosomes were either released directly from epithelium or in relation with multivesicular body (MVB). The average size of the particles varies between 50 and 130 nm. These exosomes were also widely distributed in the epithelial ciliated cells and pericytoplasm of glands lumen. Intracellular MVB was characterized by membrane-bounded exosomes of different sizes. Exosomes were also found in close contact with the cilia and sperm membrane in the lumen, which is suggestive of their fusogenic properties. Immunohistochemistry results showed strong to moderate positive expression of exosomes, in ciliated and gland cells, during January, September, and December, as it is the time of sperm storage in this turtle, whereas they showed moderate to weak level of expression during breeding season (May). This is first study about identification of the exosomes in female turtles. Epithelial and glandular exosomes, intracellular MVB, secretions, and secretory vesicles give this turtle specie a unique secretory morphology and a potential model for investigating the secretory nature of the oviduct.

Abstract 2130: The RNA-binding protein tristetraprolin controls intestinal cell differentiation and tumorigenesis through the Notch signaling pathway

Abstract Colorectal cancer (CRC) is the 2nd leading cause of cancer death in U.S. With lifetime incidence rates as high as 1 in 20, it is critical to better define the molecular events controlling intestinal homeostasis and tumor initiation/progression. In normal intestinal homeostasis, the Notch pathway plays a fundamental role in controlling epithelial cell fate through juxtacrine signaling between membrane-bound ligands to the Notch receptor present on contacting cells. This complex promotes intestinal epithelial cell fate through transcription factors Hes1 and Atoh1/Math1, allowing for differentiation of absorptive enterocyte lineage by Hes1 or secretory (goblet, enteroendocrine and Paneth) cell lineage controlled by Atoh1. In CRC however, constitutive Notch activation is observed allowing for enhanced Hes1 expression associated with tumor progression and self-renewal of tumor-initiating cells. Yet, a clear mechanism for the constitutive activation of this pathway in CRC is not known. Here, we investigated the regulation of Notch signaling by the mRNA decay factor Tristetraprolin (TTP) in CRC and TTP knockout mice. TTP is a member of the TIS11 family of RNA-binding proteins, and the major factor promoting decay of mRNAs bearing AU-rich element (ARE) motifs in their 3′UTR. TTP plays an essential role in controlling pro-inflammatory and oncogenic gene expression in the GI tract, however during CRC progression, TTP expression is silenced in tumors. In this study, TTP-inducible CRC cells were used to show that expression of TTP suppresses proliferation and anchorage-independent growth, impacts cell cycle, and reduces xenograft tumor growth. Interestingly, TTP expression altered CRC cells such that cells now displayed a secretory cell-like morphology, consistent with studies showing effects of Notch signaling attenuation. Alcian blue staining of acidic mucins was significantly increased in TTP-expressing CRC cells and in xenograft tumors, suggesting a role for TTP in directing secretory cell differentiation. Furthermore, gene expression arrays and reporter gene assays showed that levels of Notch signaling genes were significantly impacted by TTP and TTP preferentially promoted a 3-fold downregulation of Notch1 receptor as compared to Notch2-4. The ability of TTP to target Notch1 mRNA for rapid decay occurred through its ARE-containing 3′UTR as observed using 3′UTR reporter and RNA-binding assays. To determine if these effects were specific to tumor cells, intestinal cell lineage was examined in TTP knockout mice with a remarkable reduction in goblet cell number observed as compared to wild-type mice. These findings demonstrate a novel role for TTP in controlling intestinal cell differentiation via post-transcriptional regulation of Notch1 and provide mechanistic insights into constitutive Notch activation observed in CRC occurring through loss of TTP expression. Citation Format: Shufei Zhuang, Liangyan Hu, Sandhya Sanduja, Shahid Umar, Shrikant Anant, Dan A. Dixon. The RNA-binding protein tristetraprolin controls intestinal cell differentiation and tumorigenesis through the Notch signaling pathway. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2130. doi:10.1158/1538-7445.AM2015-2130

Postpartum breast involution reveals regression of secretory lobules mediated by tissue-remodeling

IntroductionA postpartum diagnosis of breast cancer is an independent predictor of metastases, however the reason is unknown. In rodents, the window of postpartum mammary gland involution promotes tumor progression, suggesting a role for breast involution in the poor prognosis of human postpartum breast cancers. Rodent mammary gland involution is characterized by the programmed elimination of the secretory lobules laid down in preparation for lactation. This tissue involution process involves massive epithelial cell death, stromal remodeling, and immune cell infiltration with similarities to microenvironments present during wound healing and tumor progression. Here, we characterize breast tissue from premenopausal women with known reproductive histories to determine the extent, duration and cellular mechanisms of postpartum lobular involution in women.MethodsAdjacent normal breast tissues from premenopausal women (n = 183) aged 20 to 45 years, grouped by reproductive categories of nulliparous, pregnant and lactating, and by time since last delivery were evaluated histologically and by special stain for lobular area, lobular type composition, apoptosis and immune cell infiltration using computer assisted quantitative methods.ResultsHuman nulliparous glands were composed dominantly of small (approximately 10 acini per lobule) and medium (approximately 35 acini per lobule) sized lobules. With pregnancy and lactation, a >10 fold increase in breast epithelial area was observed compared to nulliparous cases, and lactating glands were dominated by mature lobules (>100 acini per lobule) with secretory morphology. Significant losses in mammary epithelial area and mature lobule phenotypes were observed within 12 months postpartum. By 18 months postpartum, lobular area content and lobule composition were indistinguishable from nulliparous cases, data consistent with postpartum involution facilitating regression of the secretory lobules developed in preparation for lactation. Analyses of apoptosis and immune cell infiltrate confirmed that human postpartum breast involution is characterized by wound healing-like tissue remodeling programs that occur within a narrowed time frame.ConclusionsHuman postpartum breast involution is a dominant tissue-remodeling process that returns the total lobular area of the gland to a level essentially indistinguishable from the nulliparous gland. Further research is warranted to determine whether the normal physiologic process of postpartum involution contributes to the poor prognosis of postpartum breast cancer.

Ki-67 Index as an Ancillary Tool in the Differential Diagnosis of Proliferative Endometrial Lesions With Secretory Change

"Secretory change" can accompany a variety of proliferative endometrial lesions, ranging from hyperplasia to carcinoma. It is characterized by subnuclear or supranuclear vacuolization, mimicking early secretory endometrium (SEM). As an additional diagnostic challenge, mitotic activity and cytologic atypia are often low in hyperplastic lesions with secretory change. As mitotic activity in lesions with secretory change is decreased, the mitotic index may not be useful to distinguish SEM with glandular crowding from hyperplasia with secretory change. We therefore hypothesized that Ki-67 immunohistochemistry, an alternative marker of proliferative activity, might be useful in this setting. Forty-four endometrial lesions with secretory change and 30 controls were stained for Ki-67. Three "hot spot" areas per case were photographed and 200 to 300 cells were manually counted to obtain the ratio of Ki-67-positive cells versus total cells. A second pathologist performed an independent review of the same preselected fields and estimates without preselection. There was an incremental increase in the Ki-67 index from 2.6% in SEM to 17% in nonatypical hyperplasia, 36% in atypical hyperplasia, and 60% in endometrial carcinoma. The Ki-67 index for SEM was significantly (P<0.01) lower than hyperplastic lesions and carcinoma with secretory change. Similar, statistically significant results were obtained by independent estimates of Ki-67 immunopositivity. In the setting of secretory morphology, the Ki-67 index was highly sensitive and specific (>90%) for the differential diagnosis of SEM with crowding versus nonatypical hyperplasia, atypical hyperplasia, and endometrial carcinoma. In summary, the Ki-67-labeling index is a useful technique to distinguish SEM with crowding, an exaggerated physiological condition, from cancer precursors.

Mitochondria-targeted plastoquinone derivative SkQ1 decreases ischemia-reperfusion injury during liver hypothermic storage for transplantation

The ability of the mitochondria-targeted plastoquinone derivative 10-(6'-plastoquinonyl)decyl triphenylphosphonium (SkQ(1)) to decrease ischemia-reperfusion injury in isolated liver during hypothermic storage (HS) was studied. Rat liver was stored for 24h at 4°C without or in the presence of 1μM SkQ(1) with following reperfusion for 60min at 37°C. The presence in the storage medium of SkQ(1) significantly decreased spontaneous production of reactive oxygen species and intensity of lipid peroxidation in the liver during HS and reperfusion. The GSH level after HS in solution with SkQ(1) was reliably higher, but reperfusion leveled this effect. At all stages of experiment the presence of SkQ(1) did not prevent the decrease of antioxidant enzyme activities such as catalase, GSH peroxidase, GSH reductase, and glucose-6-phosphate dehydrogenase. The addition of SkQ(1) to the storage medium improved energetic function of the liver, as was revealed in increased respiratory control index of mitochondria and ATP level. SkQ(1) exhibited positive effect on the liver secretory function and morphology after HS as revealed in enhanced bile flow rate during reperfusion and partial recovery of organ architectonics and state of liver sinusoids and hepatocytes. The data point to promising application of mitochondria-targeted antioxidants for correction of the ischemia-reperfusion injury of isolated liver during long-term cold storage before transplantation.

ER, PR and Ki‐67 expression status in granulomatous and chronic non‐specific endometritis

To study the changes in the histological pattern, distribution and intensity of sex steroid receptors (estrogen and progesterone) and cell proliferation by Ki-67 expression by semi-quantitative scores in granulomatous and chronic non-specific endometritis in the premenstrual phase. A retrospective study was conducted on 20 cases of granulomatous endometritis, 10 of chronic non-specific endometritis and 30 age matched (2 years) controls with no endometrial lesions. Morphological changes were noted on histological examination and semi-quantitative scoring of Estrogen Receptor (ER), Progesterone Receptor (PR) and Ki-67 expression was done by immunohistochemistry. There was significantly higher ER, PR and Ki-67 expression in endometrial glandular and stromal cells in inflamed endometria as compared with the controls (all P-values < 0.02) regardless of the character of the inflammation. The cases with morphology not conforming to the secretory phase at which biopsy was taken had significantly higher ER, PR and Ki-67 expression in both endometrial and stromal cells indicating a lag in the endometrial maturation (all P-values < 0.02). Interestingly, all parameters except PR expression in glandular cells had a significantly higher expression even in cases with secretory morphology indicating disturbances in local milieu. Endometrial inflammation interferes with local expression of ER, PR and Ki-67. This may contribute to infertility regardless of other factors and other endometrial dysfunctional states.

Effect of hypo-, iso- and hypertonic saline irrigation on secretory mucins and morphology of cultured human nasal epithelial cells

Conclusion. We speculate that isotonic saline is the most physiological irrigation solution in terms of mucus secretion and the cellular morphology of nasal epithelial cells. Objective. To determine the most physiologic and effective saline concentration for nasal irrigation in terms of mucus secretion and cellular morphology by using normal human nasal epithelial cells treated with hypo-, iso- and hypertonic saline. Material and methods. Fully differentiated passage-2 normal human nasal epithelial cells were treated with pure water and with 0.3% (hypotonic), 0.9% (isotonic) and 3% (hypertonic) saline solutions. mRNA expression levels of MUC5AC and MUC5B, which are known to be major airway mucins, were analyzed after 30 min using reverse transcriptase polymerase chain reaction. Total mucin and MUC5AC and MUC5B mucin secretions were analyzed using dot-blotting. Cellular morphology was observed with light microscopy after hematoxylin–eosin staining and with scanning electron microscopy. Results. MUC5AC and MUC5B mRNA levels did not change after treatment with pure water and various concentrations of saline. Total mucin and MUC5AC mucin secretions only increased following pure water treatment, while MUC5B mucin secretion increased with pure water, hypo- and hypertonic saline treatment. Morphologic analysis revealed that pure water severely damaged normal human nasal epithelial cells and that only isotonic saline did not affect their morphology.

Ultrastructure of Clara cells stimulated by isoproterenol

The incidence of peripheral pulmonary adenocarcinoma has increased in recent years. Clara cell has been known as target for carcinogens and source of pulmonary tumors. One of the presumed roles of the bronchiolar Clara cell is the secretion of pulmonary surfactant into the bronchiolar lumen. To establish the secretory morphology of Clara cell, a well-defined secretory agonist, isoproterenol (500 mg/kg) and the antagonist, propranolol (20 mg/kg), were administered into five-week old mice. The secretory response was examined at 1 hour and 4 hours after injection. Ultrastructural morphometry was used to quantitate the secretory response by measuring area of apical cap of the Clara cells. Isoproterenol caused a significant increase in area of apical cap of Clara cells 1 and 4 hours after injection (p < 0.0001), while pretreatment with propranolol prevented this effect at 4 hours. Propranolol alone significantly decreased the area of Clara cells (p < 0.0001). Clara cells secretory granules disappeared 1 hour after propranolol plus isoproterenol administration, and the granules reappeared at 4 hours. The accelerated secretion of Clara cells by isoproterenol provides evidence of their secretory mechanism controlled by beta-adrenergic agonists. The study has confirmed the secretory role of Clara cells. The secretion is both apocrine and merocrine type.

Distribution of Eisenia tetradecapeptide immunoreactive neurons in the nervous system of earthworms

A detailed mapping of Eisenia-tetradecapeptide-immunoreactive neurons in the central and peripheral nervous system combined with quantitative morphological measurements was performed in Eisenia fetida and Lumbricus terrestris. In Eisenia, most labelled neurons were observed in the ganglia of the ventral cord (20.38% of the total cell number of the ganglion) and 15.67% immunoreactive cells occurred in the brain, while 6% of the neurons could be shown in the subesophageal ganglion. In the case of Lumbricus, most immunoreactive cells were found in the subesophageal ganglion (16.17%) and in the ventral ganglia (12.54%). The brain contained 122 ETP-immunoreactive cells (5.6%). The size of the immunoreactive cells varied between 35-75 microm. A small number of Eisenia-tetradecapeptide immunoreactive fibres were seen to leave the ventral ganglia via segmental nerves, and labelled processes could also be observed in the stomatogastric system and the body wall. Labelled axon branches originating from the segmental nerves formed an immunoreactive plexus both between the circular and longitudinal muscle layer and on the inner surface of the longitudinal muscle layer. This inner plexus was especially rich in the setal sac. Among the superficial epithelial cells the body wall contained a significant number of immunoreactive cells. Only a few Eisenia-tetradecapeptide immunoreactive neurons and fibres occurred in the stomatogastric ganglia. In the enteric plexus the number of immunoreactive neurons and fibres decreased along the cranio-caudal axis of the alimentary tract. Eisenia-tetradecapeptide immunoreactive cells were also present among the epithelial cells in the alimentary canal. Some of these cells resembled sensory neurons in the foregut, while others showed typical secretory cell morphology in the midgut and hindgut.

Modulation of mammary development and programmed cell death by the frequency of milk removal in lactating goats.

1. Unilateral changes in mammary cell number are elicited when one gland is milked more or less frequently than the contralateral gland in lactating goats. These changes were investigated using histochemical and immunocytochemical markers of mammary cell types, and the degree of mammary apoptosis was determined by end-labelling of fragmented DNA. 2. Histological analysis confirmed that unilateral cessation of milking initiated involution and cell loss preferentially in the unmilked gland. The presence of fragmented DNA and morphological characteristics consistent with apoptosis demonstrated that these changes in mammary cell number in unmilked glands were, in part, the result of programmed alveolar cell death. 3. De-differentiation of the remaining secretory cells to ductal epithelial cells occurred with an increase in staining of cytokeratin markers and decreased staining by peanut lectin and casein antisera. 4. Differential once- and thrice-daily milking of lactating goats was also associated with unilateral changes in mammary cell number and milk yield. Milk yield and alveolar size were reduced after 4 weeks of infrequent milking. The latter was due to the increased loss of secretory cells by apoptosis, as indicated by a higher degree of fragmented DNA laddering. 5. After 10 weeks of differential milking, a homogeneous secretory morphology, albeit with smaller alveoli, was maintained in thrice-daily milked glands. Once-daily milked glands possessed a heterogeneous composition of terminal structures, resulting in the simultaneous presence of secretory and involuting alveoli as well as resting ductules. 6. The differences in programmed cell death and mammary morphology between unmilked and twice-daily milked glands, and between once- and thrice-daily milked glands, suggests that mammary apoptosis is subject to modulation by intra-mammary mechanisms sensitive to the frequency of milk removal.

Secretory and Morphological Changes in the Baboon (Papio Anubis) Uterus and Placenta during Early Pregnancy1

The study was undertaken to characterize changes in uterine endometrial secretory activity and morphology in the baboon between Days 18 and 32 of pregnancy. Proteins synthesized by the implantation site endometrium (RI) and non-implantation site endometrium (RIII) and developing placenta were analyzed by two-dimensional PAGE and fluorography. The electrophoretic mobility and localization of chorionic gonadotropin (CG) in the baboon placenta, and insulin-like growth factor binding protein-1 (IGFBP-1) and retinol binding protein (RBP) in the baboon uterus were confirmed by Western blot analysis and immunocytochemistry. Placental protein synthesis was primarily characterized by the presence of radiolabeled CG at Days 18 and 25, which decreased by Day 32. Peak CG synthetic activity, immunolocalized to the syncytiotrophoblast, occurred during the formation of tertiary villi. Also associated with the development of mature villi was a corresponding increase in a basic protein (M(r) 23,000). The overall secretory pattern of the baboon endometrium was similar to that observed during the midluteal stage; however, the synthetic activity was enhanced during pregnancy. Radiolabeled and immunoreactive IGFBP-1 and RBP were detectable in the Day 32 functionalis and basalis, respectively, and their synthesis was higher in RI compared to RIII. The immunolocalization of IGFBP-1 in RI appeared to be directly related to the degree of decidualization. In addition, a novel group of low molecular-weight (M(r) 27,000-30,000) isoelectrovariant proteins was secreted predominantly by the basalis at Days 18 and 25 of pregnancy and disappeared by Day 32 in all but one animal. The synthesis of this group of proteins did not show regional specificity. These studies indicate that the baboon endometrium undergoes morphological and secretory changes during pregnancy that are associated with the development of a functional placenta and decidualized endometrium. Furthermore, these studies suggest that the conceptus/placenta may mediate specific changes in IGFBP-1 and RBP expression at the implantation site.

Progesterone Induced Expression of Alkaline Phosphatase Is Associated with a Secretory Phenotype in T47D Breast Cancer Cells

In our previous work we reported on stimulation of a tissue unspecific alkaline phosphatase (liver/bone/kidney, L/B/K) in the human breast cancer cell line T47D by progestins. Here we show that in these cells the synthetic progestin R5020 (Promegestone) induces transcription of a 2.7-kilobase ALP mRNA. In this cell line, maximal induction is reached after 24 hours, decreases to 50 % after 72 hours and is sensitive to inhibitors of protein synthesis. The induction of ALP mRNA and enzyme activity is specific for R5020 and dexamethasone and is completely inhibited in the presence of 10 −7 M RU486. When treated with R5020 for 48 hours, T47D cells exibit a substantially altered phenotype, lipid vacuoles accumulate all over the cytoplasm, conferring to the cells a secretory morphology. This phenotype is associated with increased ALP enzyme activity which is also maximal at 48 hours. This effect is progestin specific since other steroids do not lead to the same macroscopical changes.

Effects of Acute and Chronic Treatments with Atrial Natriuretic Factor (ANF) on the Leydig Cells of the Rat Testis

Acute ANF bolus administration (40 micrograms.kg-1) did not affect secretory activity and morphology of rat Leydig cells. Prolonged (7-day) ANF infusion (20 micrograms.kg-1.h-1), on the contrary, elevated both basal and hCG-stimulated testosterone blood concentration, and caused a notable hypertrophy of rat Leydig cells. Leydig-cell hypertrophy was due to increases in the volume of all the organelles involved in cholesterol and testosterone synthesis (i.e. mitochondria, smooth endoplasmic reticulum and peroxisomes). These findings suggest that ANF, when chronically administered, is able to stimulate the growth and steroidogenic capacity of rat Leydig cells.

A morphologically distinct candidate for an epithelial stem cell in mouse mammary gland.

Transplantation studies demonstrate that an epithelial stem cell component must exist in the mouse mammary gland throughout life. Samples taken from any portion of the mammary gland at any age and at any developmental stage, including full functional differentiation, give rise to mammary epithelial outgrowths with complete developmental capacity. Cytological examination of mouse mammary gland explants revealed the presence of morphologically distinct cells distributed sporadically among the mammary epithelium, whose behaviour in vivo and in vitro suggested that they might represent a latent epithelial stem cell population. These pale-staining cells possessed large spherical nuclei, a clear cytoplasm and a round smooth-contoured shape. Electron microscopy confirmed their pale-staining characteristics and revealed a cytoplasm sparsely populated with organellar structures, such as mitochondria and endoplasmic reticulum. Their epithelial genealogy was demonstrated by the presence of terminal bars and tight junctions formed with their epithelial cell neighbours. In vivo, these cells were found among mammary epithelial cell populations in 16-day-old embryos onward in both ductal or lobular structures during all stages of pregnancy, lactation and involution. In explant cultures, these cells did not undertake a secretory morphology in the presence of lactogenic hormones, although occasionally they became immunologically positive for casein. They did not incorporate [3H]-thymidine into their nuclei under any of the experimental conditions used; however, they appeared to undergo mitosis within 4 h regardless of the presence or absence of hormone(s). At 24 h increased numbers of pale cells were found in pairs or in groups. At 72 h in the presence of IFPrl (medium containing insulin, hydrocortisone and prolactin), the pairs and groups of pale cells observed at 24 h were not found. Instead, individual pale cells were seen among groups of cytologically and functionally differentiated secretory epithelial cells. When lactogenic hormones were not present, groups of pale cells were still present in the explants at 72 h. These findings suggest that the pale cells are arrested at G2 phase of the cell cycle and that they give rise by mitosis to daughter cells capable of differentiating in the presence of lactogenic stimuli. Inhibition of DNA synthesis in the explants did not alter these cellular events.

Secretory morphology of the intermediate lobe of the rat pituitary incubated in vitro.

The ultrastructure of the incubated intermediate lobe of the rat pituitary and the morphological effect of isoproterenol stimulation on its cells were studied under in vitro conditions. The general structure of isolated neurointermediate lobes maintained for 2-3 h in vitro was well preserved, and the presence of intact nerve terminals establishing synaptic contacts with the glandular cells of the intermediate lobe was confirmed. Removal of the intermediate lobe from central inhibition leads to increased hormonal secretion, which was reflected by large Golgi areas and the appearance of secretory images. However, no obvious degranulation or peripheral migration of the secretory granules after 2-3 h in vitro was seen. The secretory granules varied in electron density; totally electron-lucent granules were regularly observed and exocytotic phenomena were shown. In addition, more extensive invaginations suggesting secretion by compound exocytosis were seen. A three-fold increase in the beta-endorphin secretion during a 4-min stimulation with 10(-6) M isoproterenol did not induce any morphometrically detectable changes in the incubated cells. This indicates that only a minor fraction of the total granule content is mobilized during an acute increase in secretory activity.

Morphogenesis of mouse mammary epithelial cells growing within collagen gels: ultrastructural and immunocytochemical characterization

Mammary epithelial cells from adult virgin mice have been cultured within collagen gels in totally serum-free medium containing either epidermal growth factor or the mammogenic hormones, progesterone and prolactin, or prolactin alone. The cellular organization, differentiation and cell-type composition of the colonies from the three culture conditions were assessed by transmission electron microscopy and light-microscope immunocytochemistry. The epithelial cells form branching duct-like structures and, when exposed to mammogenic hormones, assume a secretory morphology (including casein micelles) similar to that seen in the early to mid-pregnant mouse.

Fine structure of the Caenorhabditis elegans secretory—excretory system

The secretory-excretory system of C. elegans, reconstructed from serial-section electron micrographs of larvae, is composed of four cells, the nuclei of which are located on the ventral side of the pharynx and adjacent intestine. (1) The pore cell encloses the terminal one-third of the excretory duct which leads to an excretory pore at the ventral midline. (2) The duct cell surrounds the excretory duct with a lamellar membrane from the origin of the duct at the excretory sinus to the pore cell boundary. (3) A large H-shaped excretory cell extends bilateral canals anteriorly and posteriorly nearly the entire length of the worm. The excretory sinus within the cell body joins the lumena of the canals with the origin of the duct. (4) A binucleate, A-shaped gland cell extends bilateral processes anteriorly from cell bodies located just behind the pharynx. These processes are fused at the anterior tip of the cell, where the cell enters the circumpharyngeal nerve ring. The processes are also joined at the anterior edge of the excretory cell body, where the excretory cell and gland are joined to the duct cell at the origin of the duct. Secretory granules may be concentrated in the gland near this secretory-excretory junction. Although the gland cells of all growing developmental stages stain positively with paraldehyde-fuchsin, the gland of the dauer larva stage (a developmentally arrested third-stage larva) does not stain, nor do glands of starved worms of other stages. Dauer larvae uniquely lack secretory granules, and the gland cytoplasm is displaced by a labyrinth of large, transparent spaces. Exit from the dauer stage results in the return of active secretory morphology in fourth-stage larvae.