Biomedical subjects

S E Wert

Publications and source records attributed to S E Wert.

64 records · Page 4Linked to original sources

PubMed1990

Preendocytotic alterations in cumulus cell gap junctions precede meiotic resumption in the rat cumulus-oocyte complex.

Cumulus cells in the mammalian ovary are normally connected to each other and to their enclosed oocyte by an extensive network of gap junctions (GJs). We have shown that the loss of cumulus cell GJs is correlated temporally with meiotic resumption in the intact preovulatory rat follicle (Larsen et al., 1986). Here we describe morphological changes in GJ particle packing patterns (PPPs) that occur prior to GJ loss and meiotic resumption in hormonally stimulated rat cumulus-oocyte complexes (COCs). In the PMSG-primed rat, 89% of the cumulus cell GJ area detected by freeze-fracture electron microscopy consists of tightly packed junctional particles: 4% exhibit loose PPPs of randomly dispersed particles; and 7% contain a mixture of both tight and loose PPPs. One to 2 hr after stimulation with hCG, the area of GJs containing tight PPPs drops by 50%-60%, while junctions exhibiting loosely organized and mixed patterns increase concomitantly. These shifts in PPPs are accompanied by the appearance of unusual particle-free areas of puckered or ruffled nonjunctional membrane at the GJ periphery. Cumulus cell GJs from isolated COCs incubated in FSH-containing medium demonstrate a similar shift in PPPs prior to meiotic resumption. The appearance of fusing areas of particle-free nonjunctional membrane at the GJ periphery in vitro is correlated with GJ loss and is not seen in COCs treated with dihydrocytochalasin B to inhibit endocytotic removal of cumulus GJs. The structural and temporal nature of these morphological observations supports the hypothesis that interruption of junctional communication plays a role in meiotic maturation of the preovulatory oocyte.

Animals↗

PubMed1990

Cis-acting sequences from a human surfactant protein gene confer pulmonary-specific gene expression in transgenic mice.

Pulmonary surfactant is produced in late gestation by developing type II epithelial cells lining the alveolar epithelium of the lung. Lack of surfactant at birth is associated with respiratory distress syndrome in premature infants. Surfactant protein C (SP-C) is a highly hydrophobic peptide isolated from pulmonary tissue that enhances the biophysical activity of surfactant phospholipids. Like surfactant phospholipid, SP-C is produced by epithelial cells in the distal respiratory epithelium, and its expression increases during the latter part of gestation. A chimeric gene containing 3.6 kilobases of the promoter and 5'-flanking sequences of the human SP-C gene was used to express diphtheria toxin A. The SP-C-diphtheria toxin A fusion gene was injected into fertilized mouse eggs to produce transgenic mice. Affected mice developed respiratory failure in the immediate postnatal period. Morphologic analysis of lungs from affected pups showed variable but severe cellular injury confined to pulmonary tissues. Ultrastructural changes consistent with cell death and injury were prominent in the distal respiratory epithelium. Proximal components of the tracheobronchial tree were not severely affected. Transgenic animals were of normal size at birth, and structural abnormalities were not detected in nonpulmonary tissues. Lung-specific diphtheria toxin A expression controlled by the human SP-C gene injured type II epithelial cells and caused extensive necrosis of the distal respiratory epithelium. The absence of type I epithelial cells in the most severely affected transgenic animals supports the concept that developing type II cells serve as precursors to type I epithelial cells.

Animals↗

PubMed1989

Meiotic resumption and gap junction modulation in the cultured rat cumulus-oocyte complex.

Intercellular communication within the ovarian follicle has been implicated in the control of meiotic arrest and maturation in the mammalian oocyte. We have shown that a rapid down-regulation of cumulus cell gap junctions is correlated temporally with meiotic resumption in the intact rat follicle [Larsen et al., Dev Biol, 113:517-521]. Here this relationship has been analyzed further by incubating isolated rat cumulus-oocyte complexes (COCs) with agents known to influence germinal vesicle breakdown (GVBD) or that have been shown to modulate gap junction turnover in vitro. Quantitative freeze-fracture analysis revealed that cumulus cell gap junction membrane decreased significantly prior to the initiation of GVBD in COCs incubated in medium lacking serum or other additives. The addition of serum and follicle-stimulating hormone, an experimental condition that delayed GVBD, accelerated and augmented gap junction loss at both the cumulus cell and oocyte surface. The continuous elevation of cyclic adenosine monophosphate levels, which stimulates gap junction formation in other systems, maintained meiotic arrest but did not interfere with gap junction loss. Conversely, the complete inhibition of junctional loss by a microfilament destabilizing agent, dihydrocytochalasin B, did not alter the course of GVBD normally seen in its absence. Subsequent freeze-fracture analysis and dye coupling experiments confirmed that cumulus and oocyte gap junctions in these preparations were intact and functional during the period of meiotic resumption. These findings suggest that factors other than cumulus and oocyte gap junction turnover are required for the control of meiotic arrest and maturation in the isolated COC. However, these results do support our earlier suggestion that gap junction loss within the cumulus oophorus is instrumental in isolating the oocyte from the regulatory influence of its underlying membrana granulosa cells during meiotic maturation in the intact preovulatory follicle [Larsen et al., Dev Biol, 122:61-71].

Animals↗

PubMed1989

Glucocorticoids regulate surfactant protein synthesis in a pulmonary adenocarcinoma cell line.

Synthesis of pulmonary surfactant proteins SP-A, SP-B, and SP-C was demonstrated in a cell line derived from a human adenocarcinoma of the lung. The cells contained numerous lamellar inclusion bodies and formed organized groups of cells containing well-developed junctional complexes and apical microvillous membranes. Synthesis of SP-A was detected in the cells by enzyme-linked immunoabsorbent assay and by immunoprecipitation of [35S]methionine-labeled protein. SP-A was identified as an Mr 31,000-36,000 polypeptide containing asparagine-linked carbohydrate. Northern blot analysis detected SP-A mRNA of 2.2 kb. Dexamethasone (1-10 nM) enhanced the relative abundance of SP-A mRNA. Despite stimulation of SP-A mRNA, intracellular SP-A content was unaltered or inhibited by dexamethasone. SP-B and SP-C mRNAs and synthesis of the SP-B and SP-C precursors were markedly induced by dexamethasone. ProSP-B was synthesized and secreted primarily as an Mr 42,000-46,000 polypeptide. Proteolysis of the proSP-B resulted in the generation of endoglycosidase F-sensitive Mr = 19,000-21,000 and 25,000-27,000 peptides, which were detected both intra- and extracellularly. SP-C proprotein of Mr = 22,000 and smaller SP-C fragments were detected intracellularly but were not detected in the media. Mature forms of SP-B (Mr = 8,000) and SP-C (Mr = 4,000) were not detected. Glucocorticoids directly enhance the relative synthesis and mRNA of the surfactant proteins SP-A, SP-B, and SP-C. Discrepancies among SP-A mRNA, its de novo synthesis, and cell content suggest that glucocorticoid may alter both pre- and posttranslational factors modulating SP-A expression.

Adenocarcinoma↗

PubMed1988

Roles of cell junctions in gametogenesis and in early embryonic development.

Recent reviews of the role of cell junctions in development have focused primarily upon functions related to the relatively subtle physiological modulation of their subunits in relation to fundamental developmental processes in a wide variety of organisms. There is, however, considerable support from numerous laboratories that the more radical modulation of the presence and number of junctional subunits in many diverse tissues may play a pivotal role in a wide spectrum of developmental phenomena ranging from gametogenesis to organogenesis. Since a great deal of recent interest in this latter subject has concentrated upon vertebrate systems including mammals, this review will examine the functional significance of the modulation of gap junctions, tight junctions and desmosomes in a developing idealized mammalian system from gamete formation to tissue and organ differentiation during embryo-genesis.

Animals↗

PubMed1987

Differential modulation of rat follicle cell gap junction populations at ovulation.

The hypothesis proposed in the late 1970s that meiotic resumption in mammalian oocytes might result from the disruption of gap junction communication between follicle cells and the oocyte has not been supported by metabolic cooperation experiments which demonstrate that exogenous tracer transfer from the cumulus oophorus to the oocyte does not decrease until several hours after germinal vesicle breakdown (GVBD). Since these studies utilized isolated cumulus-oocyte complexes for their measurements, however, they excluded from consideration the possible effect of separation of the cumulus oophorus from the membrana granulosa which was required for this assay. We considered the possibility that the disruption of cumulus junctions within the intact follicle could mimic this experimental manipulation and previously reported that cumulus gap junctions were dramatically down-regulated during the period of GVBD in vivo. In the present study, we have utilized quantitative morphometric techniques to analyze the responses of other gap junction populations in intact preovulatory rat follicles to an ovulatory stimulus and demonstrate now that membrana granulosa, cumulus, and cumulus-oocyte gap junctions are down-regulated at different times and rates during the preovulatory period. Although membrana gap junctions are down-regulated during the period of meiotic resumption, their loss is not as rapid or as complete as in the cumulus oophorus. Cumulus-oocyte gap junctions are down-regulated after meiosis resumes but during the same period other investigators have demonstrated a reduction in metabolite transfer between the cumulus oophorus and the oocyte. Our results are interpreted to suggest that the cumulus oophorus may regulate the conduction of meiosis inhibitory signals between the membrana granulosa and the oocyte.

Animals↗

PubMed1986

A dramatic loss of cumulus cell gap junctions is correlated with germinal vesicle breakdown in rat oocytes.

Interactions between the cumulus-oophorus and the oocyte have been implicated in the regulation of meiotic maturation. Quantitative analysis of freeze-fractured rat cumulus-oocyte complexes reveals that the net area of cumulus cell gap junction membrane decreases about 15-fold, 2-3 hr following an ovulatory stimulus. This dramatic loss of gap junctions is temporally correlated with germinal vesicle breakdown and cumulus expansion, and is discussed with respect to meiotic maturation and ovulation of the mammalian oocyte.

Animals↗

PubMedDate not supplied

Expression of thyroid transcription factor-1, surfactant proteins, type I cell-associated antigen, and Clara cell secretory protein in pulmonary hypoplasia.

Pulmonary hypoplasia (PH) is a developmental abnormality characterized by diminished distal lung parenchyma. Recent studies have demonstrated that thyroid transcription factor 1 (TTF-1), a member of NKx2 family of homeodomain transcription factors, plays an important role in lung organogenesis and lung epithelial gene expression. In order to evaluate whether abnormal expression of TTF-1 contributes to the pathophysiology of PH, we studied the expression of TTF-1, as well as that of the surfactant proteins (SPs), Clara cell secretory protein (CCSP), and type I cell-associated antigen (T1 cell-Ag), in PH. Immunolocalization patterns of these proteins were evaluated in 15 cases of PH with different associated diseases and compared with those of 14 matched controls. Our study demonstrated that the concentration gradient of TTF-1 along the proximal-distal axis in normal fetal lung is disrupted in PH after 24 weeks gestational age, while the expression of the SPs, CCSP, and T1 cell-Ag seemed to be preserved. We conclude that a normal TTF-1 expression pattern might be crucial in the control of distal lung development. Failure to switch off expression of TTF-1 in PH of more than 24 weeks gestational age may be a final common pathway leading to PH associated with the disease processes investigated in this study.

Epithelial Cells↗

PubMedDate not supplied

Expression of thyroid transcription factor-1 in congenital cystic adenomatoid malformation of the lung.

Congenital cystic adenomatoid malformation (CCAM) is an abnormality of branching morphogenesis of the lung. CCAM types 1, 2, and 3 exhibit a cellular composition that is different from that of CCAM type 4 when evaluated with bronchiolar and alveolar cell markers. Thyroid transcription factor 1 (TTF-1) regulates early lung development. To evaluate the potential role of TTF-1 in the development of CCAM, TTF-1 expression in CCAM was compared to that of fetal lungs at varying gestational ages. Twenty-three CCAM cases (17 type 1, two type 2, two type 3, and two type 4) and 11 fetal lungs (3 pseudoglandular, 4 canalicular, and 4 terminal sac stages) were analyzed using a rabbit polyclonal antiserum to rat TTF-1. Nuclear staining for TTF-1 was observed in ciliated and nonciliated cells of the bronchial and bronchiolar epithelia and in cells lining the distal air spaces by 12 weeks gestational age. By mid-gestation, proximal bronchial cells were TTF-1 negative, except for the basal cells, while TTF-1 staining was maintained in distal bronchiolar and alveolar cells. TTF-1 expression decreased in both bronchial, bronchiolar, and alveolar epithelia with advancing gestational age and cytodifferentiation. At term, TTF-1 expression persisted in a few bronchial and bronchiolar basal cells and in all alveolar type II cells, whereas type I cells were negative. In CCAM, TTF-1 was detected in the nuclei of epithelial cells lining the cysts. TTF-1 was expressed in a majority of the bronchiolar-like epithelial cells of the cysts in CCAM types 1, 2, and 3, where almost 100% of the cells were TTF-1 positive. In contrast, TTF-1 expression in the alveolar-like epithelium of CCAM type 4 cysts was restricted to type II cells and only 30%-60% of the lining cells were TTF-1 positive. These results support the hypothesis that CCAM types 1, 2, and 3 reflect abnormalities in lung morphogenesis and differentiation that are distinct from those for CCAM type 4. The role played by TTF-1 in the development of CCAM, if any, is not clear.

Animals↗

PubMedDate not supplied

Absence of lamellar bodies with accumulation of dense bodies characterizes a novel form of congenital surfactant defect.

Two female sibling full-term newborns developed respiratory distress shortly after birth, which progressed to respiratory failure. Tracheal lavage demonstrated presence of surfactant protein A (SP-A), but little surfactant protein B (SP-B), without aberrant surfactant protein C (SP-C). On a lung biopsy performed in both infants, prominent type II pneumocyte hyperplasia was evident. Through ultrastructural examination an absence of normally formed lamellar bodies was determined, with numerous irregular electron dense bodies within the type II pneumocytes. These electron dense bodies could also be identified in the alveolar spaces and alveolar macrophages. No alveolar tubular myelin was present. Abnormally high immunoreactivity for surfactant proteins SP-A, proSP-B, SP-B, and proSP-C was demonstrated by light microscopy. Presence of incompletely processed immunopositive proSP-B, but not proSP-C was observed in the alveolar lumina. No mutations in either the SP-B or SP-C gene were identified by sequence analysis of amplified cDNA. We conclude that these siblings exhibit an inherited surfactant deficiency characterized by abnormal accumulations of surfactant proteins within the pneumocytes. This abnormal accumulation may be due to a primary secretory defect, a defect in surfactant phospholipids, or an abnormal interaction between the phospholipids and surfactant proteins.

Bronchoalveolar Lavage Fluid↗