The effect of atrophy, hyperplasia, and keratinization accompanying the estrous cycle on Langerhans' cells in mouse vaginal epithelium

The morphology and numbers of Langerhans' cells vary in epithelia with different patterns of hyperplasia and keratinization. Langerhans' cells stained for ATPase were compared at five phases of the estrous cycle in murine vaginal epithelium. The cells were more dendritic and sparsely distributed with hyperplasia and were less dendritic and more densely distributed with atrophy. Greater numbers of the cells did not accompany keratinization at estrus. Ultrastructurally, three types of Langerhans' cells were discriminated. The first type, active in protein synthesis and phagocytosis, was commonest in sloughing and atrophic epithelium. The second type, containing accumulated and dispersed, electron-dense bodies presumed to be lysosomes, predominated in hyperplastic epithelium. The third, a mature resting cell, was found only after keratinization was complete. This study shows that Langerhans' cells in murine vaginal epithelium vary in morphology and numbers with the epithelial changes of the estrous cycle which may relate to their immunological role, but does not support the contention that their distribution is important for keratinization.     

Some hormones involved in the nesting behaviour of hens

Fourteen hens belonging to two breeds and with nesting experience were ovariectomized, and injected daily with oestrogen for periods of 6 weeks. Thereafter they were given progesterone and oestrogen injections daily for a week alternating with a week of oestrogen injections only; at any time during this period half were receiving progesterone and oestrogen and the remainder oestrogen only. Birds of both breeds showed nest examination and nest entry following progesterone and oestrogen treatment, but some Brown Leghorns showed these behaviour patterns under the influence of oestrogen.     

Langerhans cells in murine vaginal epithelium affected by oestrogen and topical vitamin A

Langerhans cells vary in their morphology and distribution in the vaginal epithelium of ovariectomized mice stimulated to hyperplasia and keratinization by oestrogen. When the stratum corneum was removed by topical vitamin A application, the shape and distribution of Langerhans cells were unaffected. It was concluded that Langerhans cell morphology and distribution depend on the configuration of the lower strata of the epithelium and not on the presence of a stratum corneum.     

Ovarian hormones regulate expression of the focal adhesion proteins,talin and paxillin,in rat uterine luminal but not glandular epithelial cells

During early pregnancy in the rat, focal adhesions disassemble in uterine luminal epithelial cells at the time of implantation to facilitate their removal so that the implanting blastocyst can invade into the underlying endometrial decidual cells. This study investigated the effect of ovarian hormones on the distribution and protein expression of two focal adhesion proteins, talin and paxillin, in rat uterine luminal and glandular epithelial cells under various hormone regimes. Talin and paxillin showed a major distributional change between different hormone regimes. Talin and paxillin were highly concentrated along the basal cell surface of uterine luminal epithelial cells in response to oestrogen treatment. However, this prominent staining of talin and paxillin was absent and also a corresponding reduction of paxillin expression was demonstrated in response to progesterone alone or progesterone in combination with oestrogen, which is also observed at the time of implantation. In contrast, the distribution of talin and paxillin in uterine glandular epithelial cells was localised on the basal cell surface and remained unchanged in all hormone regimes. Thus, not all focal adhesions are hormonally dependent in the rat uterus; however, the dynamics of focal adhesion in uterine luminal epithelial cells is tightly regulated by ovarian hormones. In particular, focal adhesion disassembly in uterine luminal epithelial cells, a key component to establish successful implantation, is predominantly under the influence of progesterone.     

Hormonal control of enzyme activity during the plasma membrane transformation of uterine epithelial cells

Ultrastructural and light microscopic catalytic histochemical methods were used to study the distribution and changes in distribution of four phosphatase enzymes; alkaline phosphatase, 5'-nucleotidase, thiamine pyrophosphatase and adenosine triphosphatase in uterine epithelial cells in response to the ovarian hormones, oestrogen, progesterone or a combination of both used in different regimes on ovariectomised rats. Reaction product for all four enzymes was clearly localised in the epithelial cells, especially with oestrogen priming. However, the four enzymes showed markedly different patterns of organisation of reaction product in response to other hormonal treatments.Our findings clearly show that the expression of these enzymes is under ovarian hormonal control. However, while all of the enzymes are upregulated by oestrogen, the response to progesterone is variable, which can upregulate or downregulate different enzymes. The findings are particularly obvious at the electron microscopic level on the apical plasma membrane of the uterine epithelial cells, which was the main focus of our study.     

Antigen recognition in the female reproductive tract. II. Endocytosis of horseradish peroxidase by Langerhans cells in murine vaginal epithelium.

Previous studies have shown that dendritic cells in the murine vaginal epithelium at diestrus and metestrus can endocytose intravaginally administered soluble protein tracers, but the identity of the dendritic cells was not established. In the investigation reported here, we used a combination of histochemistry and transmission electron microscopy to study the endocytosis of exogenous horseradish peroxidase by vaginal dendritic cells and to identify these cells as Langerhans' cells on the basis of their cellular associations, ultrastructural morphology, and the presence of Langerhans' cell granules.     

DEVELOPMENT OF VAGINAL EPITHELIUM SHOWING IRREVERSIBLE PROLIFERATION AND CORNIFICATION IN NEONATALLY ESTROGENIZED MICE: AN ELECTRON MICROSCOPE STUDY *

Irreversible proliferation and cornification of the mouse vaginal epithelium were induced by 10 daily injections of 20μg estradiol-17β starting on the day of birth. Development of the irreversible vaginal epithelium during the period of estrogen injections in early postnatal life was observed under light and electron microscopes. Small electron-dense cells (A-cells) in clusters were present in the columnar vaginal epithelium of newborn mice. A-cells were proliferated by 2 daily estrogen injections. At the sites of A-cell clumps, large electron-dense cells (B-cells) characterized by long winding cytoplasmic processes appeared in mice given 3 daily injections, forming nodules which then fused together to form a layer under the columnar epithelium after 4 daily injections. In mice given 7 daily injections, the primary epithelium was shed by the superficial cornification of the newly formed layer. The B-cell membrane bore fewer desmosomes than in the basal and intermediary cells of the vaginal epithelium of ovariectomized ‘normal’ adult mice after 5 daily injections of 100μg estradiol-17β. Hyperplastic epithelial downgrowths in old ovariectomized mice given neonatal estrogen injections contained another type of cells with reduced density which formed much fewer processes and only a few desmosomes (C-cells).     

Distribution of oestrogen and androgen receptors between the stroma and epithelium of the guinea-pig prostate

A procedure is described for separating guinea-pig prostatic tissue into viable epithelial and stromal fractions. Epithelial cells were isolated using 0.1% protease, but this method resulted in significant damage to the stromal cells. However, using a mechanical tissue fractionation technique, a viable stromal matrix consisting predominantly of confluent sheets of smooth muscle cells and intervening collagen fibres was obtained. Although this method selectively spilled-out the epithelial cells, the majority were non-viable and therefore not suitable for receptor studies. Electron microscopy confirmed that cell architecture and organelle morphology were well preserved in both the enzymatic epithelial preparation and the mechanically prepared stroma. Saturation analysis studies indicated that the concentration of high affinity (Kd approximately 0.15 nM) oestrogen receptors was approx. 10-times greater in the separated stroma than in the epithelial fraction. In contrast, the concentration of androgen receptors (Kd approximately 2.2nM) was almost 2-fold greater in the epithelial than in the stromal fraction. These findings suggest that oestrogen, either independently or in association with androgen, may play a role in the growth and development of the stromal component of the guinea-pig prostate.     

Oestrogen receptor content of normal breast cells and breast carcinomas throughout the menstrual cycle

To examine the presence and distribution of oestrogen receptors in the normal breast during the menstrual cycle cytological samples obtained by fine needle aspiration from 69 premenopausal women with normal breasts were analysed immunocyto-chemically with a monoclonal antibody to oestrogen receptor; samples from 15 postmenopausal women were also analysed. The receptor content of breast cancers from 83 premenopausal women was also determined in relation to when during the menstrual cycle excision was performed. In the normal premenopausal women oestrogen receptors were detected in the nuclei of epithelial cells in 21 out of 68 (31%) assessable samples. All 21 of these samples were obtained from the 35 women who were studied during the first half of their menstrual cycle (days 28 to 14). None of the 33 samples obtained during the second half of the cycle contained oestrogen receptors. Samples were assessable in eight of the postmenopausal women, six giving a positive result for oestrogen receptor. Fifty one of the 83 carcinomas were positive for oestrogen receptor, 24 having been excised during the first half of the cycle and 27 during the second half.Production of oestrogen receptor protein is suppressed at the time of ovulation in the normal breast epithelium of premenopausal women. In contrast, breast carcinoma cells either synthesise this protein continuously throughout the cycle or fail to express it despite fluctuations of serum hormone concentrations.     

Immunocytochemical analysis of oestrogen receptors and progesterone receptors in the human uterus throughout the menstrual cycle and after the menopause.

To obtain more insight into the relationship between cyclic and regional changes in steroid receptor expression and function-related changes in the various types of cell of the normal human uterus, we performed an immunocytochemical study on paraffin-embedded sections. The distribution and intensity of immunostaining for the oestrogen receptor and the progesterone receptor in the various types of cell were semiquantitatively scored. The data were statistically compared for the different phases of the menstrual cycle and after the menopause, and for the different regions of the corpus and (endo)cervix uteri. During the menstrual cycle, significant changes in oestrogen receptor score were observed in glandular and stromal cells of endometrium basalis and functionalis and in smooth muscle cells of the myometrium. In all types of cell, oestrogen receptor expression reached a maximum in the late proliferative phase. During the early secretory phase, oestrogen receptor staining declined sharply in stromal and smooth muscle cells, whereas, in glandular epithelium, oestrogen receptor expression decreased more gradually. During mid- and late-secretory phases, an increase in oestrogen receptor staining was also observed in predecidualizing stromal cells and smooth muscle cells. Progesterone receptor numbers changed significantly in glandular epithelium but not in stromal and smooth muscle cells. Glandular progesterone receptor expression reached a maximum in the early secretory phase and was then drastically reduced. During mid- and late-secretory phases stromal cells were moderately stained for progesterone receptor in contrast to epithelial gland cells which showed no or very weak staining. No regional variations in steroid receptor distribution in endometrium and myometrium were found.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regulation of gene expression and cellular localization of prostaglandin synthase by oestrogen and progesterone in the ovine uterus

Expression of the gene for prostaglandin synthase (PGS) was examined in whole endometrial tissue derived from ewes during the oestrous cycle (Days 4-14), on Day 15 of pregnancy and following ovariectomy and treatment with ovarian steroid hormones. Whilst no significant differences were seen in PGS mRNA concentrations analysed by Northern blot analysis in endometrial tissue during the oestrous cycle or in early pregnancy, treatment of ovariectomized (OVX) ewes with oestradiol-17 beta markedly reduced endometrial PGS mRNA concentration. There was no difference in PGS mRNA concentration in ewes treated with progesterone, either alone or in conjunction with oestrogen, from that in OVX controls. In contrast, differences in immunolocalization of PGS observed in uterine tissue from OVX-steroid-treated ewes were much more marked and reflected similar changes seen previously in the immunocytochemical distribution of endometrial PGS during the oestrous cycle. In OVX ewes and those treated with oestrogen, immunocytochemical staining for PGS was seen in stromal cells, but little immunoreactive PGS was located in the endometrial epithelial cells. However, in ewes treated with progesterone alone or with oestrogen plus progesterone, PGS was found in luminal and glandular epithelial cells and in stromal cells. Intensity of immunostaining for PGS in endothelial cells and myometrium did not differ between the treatments. Thus, whilst oestrogen lowers PGS mRNA in the endometrium, presumably in stroma, it may also increase the stability of the enzyme itself in the stromal cells. Although oestradiol-17 beta has no effect on PGS in endometrial epithelium, progesterone stimulates the production of PGS in endometrial epithelial cells without altering the overall abundance of PGS mRNA in the endometrium as a whole. Conceptus-induced changes in PGF-2 alpha release by ovine endometrium would not appear to be mediated via effects on PGS gene expression or protein synthesis.     

DNA synthesis in Langerhans' cells of mouse ear epithelium revealed by tritiated thymidine autoradiography and histochemical staining for non-specific esterase and beta-glucuronidase activity

The proportion of Langerhans' cells in DNA synthesis in normal mouse skin was assessed by combining tritiated thymidine [3H]TdR autoradiography with enzyme histochemistry. After injection of [3H]TdR, ear skin was treated in two ways. Epithelial sheet preparations were stained for the presence of non-specific esterase and cytospin preparations of epithelial cell suspensions were stained for beta-glucuronidase activity. The labelling index (+/- SE mean) for cytospins, 40 min after injecting [3H]TdR, was 1.6 +/- 0.15%, doubling to 3-4% from 7-17 days after injection. The sheet preparations showed the proportion of label attributable to paired Langerhans' cells rising from 18% at 40 min after injection, to approximately 45%, on days 1-4 after injection. These results suggest that the proliferation of Langerhans' cells in normal mouse skin might be higher than was previously thought to be the case.     

The multifaceted role of oestrogen in enhancing Chlamydia trachomatis infection in polarized human endometrial epithelial cells

The oestrogen receptor (ER) α-β+ HEC-1B and the ERα+β+ Ishikawa (IK) cell lines were investigated to dissect the effects of oestrogen exposure on several parameters of Chlamydia trachomatis infection. Antibody blockage of ERα or ERβ alone or simultaneously significantly decreased C. trachomatis infectivity (45-68%). Addition of the ERβ antagonist, tamoxifen, to IK or HEC-1B prior to or after chlamydial infection caused a 30-90% decrease in infectivity, the latter due to disrupted eukaryotic organelles. In vivo, endometrial glandular epithelial cells are stimulated by hormonally influenced stromal signals. Accordingly, chlamydial infectivity was significantly increased by 27% and 21% in IK and HEC-1B cells co-cultured with SHT-290 stromal cells exposed to oestrogen. Endometrial stromal cell/epithelial cell co-culture revealed indirect effects of oestrogen on phosphorylation of extracellular signal-regulated kinase and calcium-dependant phospholipase A2 and significantly increased production of interleukin (IL)-8 and IL-6 in both uninfected and chlamydiae-infected epithelial cells. These results indicate that oestrogen and its receptors play multiple roles in chlamydial infection: (i) membrane oestrogen receptors (mERs) aid in chlamydial entry into host cells, and (ii) mER signalling may contribute to inclusion development during infection. Additionally, enhancement of chlamydial infection is affected by hormonally influenced stromal signals in conjunction with direct oestrogen stimulation of the human epithelia.     

Hormonal control of pinocytosis in the uterine epithelium of the rat.

Ovariectomized rats were treated with oestradiol-17 beta and/or progesterone to mimic the hormonal parameters inducing uterine sensitivity for implantation. The degree of pinocytosis of trypan blue and ferritin in the endometrial cells was examined. Significant epithelial pinocytosis of trypan blue occurred after a 3-day treatment of progesterone, and uptake was independently increased by priming with oestrogen and by oestradiol given on the 3rd day of progesterone treatment. Progesterone treatment caused uptake of ferritin by the epithelial cells; in control animals epithelial and stromal cells were involved. Oestrogen priming enhanced ferritin absorption, while 'nidatory' oestrogen had no effect. Oestradiol given alone completely blocked pinocytosis of both intraluminally injected substances.     

Modification of cell evagination and cell differentiation in quail oviduct hyperstimulated by progesterone

Quail oviduct development is controlled by sex steroid hormones. Estrogen (E) induce cell proliferation, formation of tubular glands by epithelial cell evagination and cell differentiation. Progesterone (P) strongly increases the secretory process in E-treated quails, but inhibits cell proliferation, cell evagination and differentiation of ciliated cells. The balance between E and P is critical for harmonious development of the oviduct. After 6 daily injections of two doses of estradiol benzoate (10 or 20 micrograms/d) and high doses of P (4 mg/d), tubular gland formation by epithelial cell evagination was inhibited, while epithelial cell proliferation occurred, as shown by the height of the villi and the increase in DNA. Secretory processes were strongly stimulated. Ovalbumin, a tubular gland cell marker and avidin, a mucous cell marker, were localized by immunofluorescence and immunogold labeling. Ovalbumin was localized only in the rudimentary tubular glands, whereas avidin was dispersed throughout the secretory cells. High doses of progesterone inhibited tubular gland cell proliferation, disturbed the distribution of avidin and inhibited differentiation of ciliated cells. Ovalbumin synthesis occurred only in epithelial cells which were evaginated despite the hyperstimulation. Ovalbumin gene expression appeared highly dependent upon the cell position.     

Langerhans cells phagocytose vaginal epithelial cells undergoing apoptosis during the murine estrous cycle.

Langerhans' cells (LCs) have been studied extensively in the epidermis, where they function as antigen-presenting cells. LCs are also present in the stratified epithelia of the murine vagina and cervix, but their function at these sites is not known. Recent reports noted the association of LCs with vaginal epithelial cells undergoing apoptosis and suggested that LCs might be involved in phagocytosis of dead cells. The present study describes the ultrastructural details of this process. The results demonstrate that LCs in murine vaginal epithelium during late metestrus and early diestrus phagocytose apoptotic epithelial cells and may thereby contribute to the normal turnover of the vaginal epithelium during the estrous cycle.     

Cellular changes in the prostatic stroma of glucocorticoid-treated rats

Glucocorticoid hormones (GCs) have been widely used for the treatment of prostate cancer because of their inhibitory property against tumour growth. However, their mechanism of action in the prostate has received little attention. Excess GCs can lead to peripheral insulin resistance resulting in hyperglycaemia and hyperinsulinaemia. Insulin plays an important role as a cellular stimulant and high levels are related to low levels of androgens. Our objective has been to describe the effects of insulin resistance induced by dexamethasone treatment on the morphology of rat ventral prostate. Male adult Wistar rats received daily intraperitoneal injections of dexamethasone or saline for five consecutive days after which the rats were killed and the ventral prostate was removed, weighed and prepared for conventional and transmission electron microscopy (TEM). Dexamethasone treatment resulted in atrophy and decreased proliferative activity of prostatic epithelial cells. TEM analysis revealed changes in the epithelium-stroma interface, with some interruptions in the basement membrane. Fibroblasts showed a secretory phenotype with dilated endoplasmic reticulum. Smooth muscle cells exhibited a contractile pattern with 50% atrophy, an irregular membrane and twisted nuclei. Mitochondrial alterations, such as enlarged size and high electron density in the mitochondrial matrix, were also detected in smooth muscle cells. Insulin resistance induced by dexamethasone is thus associated with epithelial atrophy similar to that described for diabetic rats. However, GCs are responsible for morphological changes in the stromal cell population suggesting the activation of fibroblasts and atrophy of the smooth muscle cells.     

Oestrogen‐induced angiogenesis promotes adenomyosis by activating the Slug‐VEGF axis in endometrial epithelial cells

Adenomyosis is an oestrogen‐dependent disease characterized by the invasion of endometrial epithelial cells into the myometrium of uterus, and angiogenesis is thought to be required for the implantation of endometrial glandular tissues during the adenomyotic pathogenesis. In this study, we demonstrate that compared with eutopic endometria, adenomyotic lesions exhibited increased vascularity as detected by sonography. Microscopically, the lesions also exhibited an oestrogen‐associated elevation of microvascular density and VEGF expression in endometrial epithelial cells. We previously reported that oestrogen‐induced Slug expression was critical for endometrial epithelial–mesenchymal transition and development of adenomyosis. Our present studies demonstrated that estradiol (E2) elicited a Slug‐VEGF axis in endometrial epithelial cells, and also induced pro‐angiogenic activity in vascular endothelial cells. The antagonizing agents against E2 or VEGF suppressed endothelial cells migration and tubal formation. Animal experiments furthermore confirmed that blockage of E2 or VEGF was efficient to attenuate the implantation of adenomyotic lesions. These results highlight the importance of oestrogen‐induced angiogenesis in adenomyosis development and provide a potential strategy for treating adenomyosis through intercepting the E2‐Slug‐VEGF pathway.     

CD43 is relocated from the basal to the apical plasma membrane of rat uterine epithelial cells by progesterone

Adhesion molecules play an important part in preparing uterine epithelial cells for receptivity to the implanting embryo, and their rearrangement is crucial in allowing successful implantation. CD43 is an adhesion molecule which has previously been suggested to take part in implantation in mice. Indirect immunofluorescence microscopy localising CD43 was performed on uterine tissue during early pregnancy, and tissue obtained from ovariectomised rats administered with ovarian hormones. Western blotting was performed during early pregnancy on isolated epithelial cells and ovariectomised rats for comparison of the amount of CD43. Immunofluorescence microscopy showed CD43 was situated basally in uterine luminal epithelial cells on day 1 of pregnancy and during oestrogen administration, corresponding to a 95-kDa band of CD43 seen in western blotting. At the time of implantation, and during progesterone or progesterone plus oestrogen combined treatment, CD43 is apical in uterine luminal epithelial cells, resulting in an 85-kDa form of CD43. We suggest that a de-glycosylated form of CD43 moves from basally to apically at the time of implantation, thus facilitating blastocyst attachment to uterine epithelial cells as well as their removal.     

Progesterone-dependent and -independent avidin production in chick oviduct organ culture

Progesterone-dependent and -independent avidin inductions were found in chick oviduct culture. The effects of oestrogen priming and actinomycin D on these inductions were studied. Priming comprised daily diethylstilboestrol (DES) injections in vivo and one day's incubation in vitro with DES before experimental treatment. After 0 or 1 day of DES pretreatment no measurable amount of avidin was found. Avidin was present in progesterone-untreated incubated tissues after 4 days' DES pretreatment. Incubation with progesterone for 24 h increased avidin levels only after 4 or more days of oestrogen priming. No induction of avidin by actinomycin D similar to that found in vivo was observed in vitro. Actinomycin D inhibited both progesterone-dependent and -independent avidin production, but this effect diminished as oestrogen pretreatment was prolonged. Actinomycin D also significantly reduced total oviductal RNA synthesis. It is concluded that oestrogen priming enhances both progesterone-dependent and -independent avidin production in vitro and that both inductions are partially dependent on new RNA synthesis.