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Biomedical subjects

J Mulholland

Publications and source records attributed to J Mulholland.

At least 19 recordsLinked to original sources

Nursing, humanism and transcultural theory: the 'bracketing-out' of reality.

This paper addresses the emergence of a humanist discourse within nursing and questions the extent to which it represents the panacea implied within much of the humanist nursing literature. Particular attention will be given to whether it represents an ontological and epistemological framework capable of understanding the social, economic and political dynamics formative in the structuring of nurse-client relations. It will be argued that the humanist analyses extant within much nursing literature are vague, idealistic, inconsistent and inadequate in the sense that they offer little by way of a meaningful analysis of power. A critique will also be made of the methodological individualism implicit within much humanist analyses. The paper will go on to identify the influence of humanist approaches on transcultural theory, and the manner in which the epistemological foundations of the latter have shared the limitations of the humanistic nursing approach generally. As such, the transcultural nursing literature is often vague, inconsistent in its use of terminology, lacking in any rigorous analysis of power, and suspect in its conceptualizations of culture. Its capacity for enabling nurses to examine critically the socio-economic and political dynamics of nurse-client relations and develop strategies for addressing racisms, considered by many to be endemic within nursing and health care system generally, is seriously undermined.

Cultural Diversity

Morphological and immunohistochemical differentiation patterns of rabbit uterine epithelium in vitro.

We describe morphological and immunohistochemical changes of uterine epithelium from immature rabbits in vitro in response to hormonal treatments, using a matrix-coated semipermeable filter. These investigations were compared to in vivo studies of uterine epithelium from immature rabbits treated with estrogen and/or progesterone. In vitro, polarization of the epithelium seems to be best developed under progesterone dominance, and the pattern of cell organelles is similar to those seen in vivo. Two types of apical protrusions could be observed in cultures treated with progesterone, some shaped like domes, containing cell organelles, and some irregular in shape with small lucent vesicles. Both types of apical differentiation are typical for the in vivo situation. In vitro, estrogen leads to a more pseudostratified growth pattern of the cells. They develop apical protrusions with big vesicles probably containing mucin, as in vivo. Treatment with both steroid hormones leads to a heterogeneous response of the uterine epithelial cells in culture, some cells responding more to the estrogen, others to the progesterone whereas in vivo the progesterone-dominant features are obvious. Immunohistochemistry of uteroglobin in monensin-treated cultures gives evidence for uteroglobin secretion in all cultures, but to a lesser extent in the untreated, and this is strongly increased in cultures treated with estrogen and progesterone. These results correspond to observations made in vivo. This in vitro cell culture method seems therefore to provide a useful model for investigating the regulatory mechanisms of sexual steroid hormones and the cell biology of uterine receptivity.

Animals

Ultrastructure of the yeast actin cytoskeleton and its association with the plasma membrane.

We characterized the yeast actin cytoskeleton at the ultrastructural level using immunoelectron microscopy. Anti-actin antibodies primarily labeled dense, patchlike cortical structures and cytoplasmic cables. This localization recapitulates results obtained with immunofluorescence light microscopy, but at much higher resolution. Immuno-EM double-labeling experiments were conducted with antibodies to actin together with antibodies to the actin binding proteins Abp1p and cofilin. As expected from immunofluorescence experiments, Abp1p, cofilin, and actin colocalized in immuno-EM to the dense patchlike structures but not to the cables. In this way, we can unambiguously identify the patches as the cortical actin cytoskeleton. The cortical actin patches were observed to be associated with the cell surface via an invagination of plasma membrane. This novel cortical cytoskeleton-plasma membrane interface appears to consist of a fingerlike invagination of plasma membrane around which actin filaments and actin binding proteins are organized. We propose a possible role for this unique cortical structure in wall growth and osmotic regulation.

Actins

Regulation of connexin26 and connexin43 expression in rat endometrium by ovarian steroid hormones.

A distinct spatial and temporal pattern of connexin26 and connexin43 (cx26 and cx43) expression was observed in the rat endometrium in response to embryo implantation; however, connexin expression was suppressed during the preimplantation period. Pseudopregnant rats did not show connexin mRNA, while artificial decidualization induced by a scratch led to a strong expression of cx26 and cx43 in the endometrium of these animals. In order to examine the regulatory effects of ovarian steroid hormones on connexin expression, ovariectomized rats were treated with progesterone (P) and/or estradiol-17 beta (E2). Untreated, ovariectomized animals expressed mRNA for cx43, but not for cx26. Endometrial expression of mRNA for both connexins was strongly enhanced by E2 treatment; immunolabeling revealed protein for cx26 in the uterine luminal epithelial cells and for cx43 in the uterine stromal cells. P treatment, either alone or in combination with E2, suppressed expression of connexin mRNA. P suppression in the presence of E2 was reversible when P was withdrawn. When administered on Days 0-2 of pregnancy, the antiprogestin onapristone inhibited the effect of P and gave rise to strong expression of both connexin transcripts. These results demonstrate that expression of cx26 and cx43 in the rat uterine endometrium is differentially regulated by E2 and P during early pregnancy.

Animals

Competency-based learning applied to nursing management.

The ability to manage change has become an essential skill for all managers, particularly those employed within the Health and Social Services. It is recognized that managers may have received an introduction to management concepts and skills within their professional education but require more specific management development. The government approach to adult education with an increased emphasis on vocational training as opposed to professional education (Hyland 1991) has had an important impact upon the theory and practice of nurse management. Nurses are now required to demonstrate, not only the appropriate knowledge and theory of management, but also the competencies necessary to fulfil their role. This is becoming an important dimension of management development programmes. This paper discusses aspects of competency based learning and considers how these concepts are applied to a higher education nurse management programme on 'Managing Change'. The difficulties experienced in applying vocational standards to professional education within the culture of a higher education will be explored, difficulties identified and some alternatives offered.

Competency-Based Education

Heparin-binding epidermal growth factor-like growth factor is differentially regulated by progesterone and estradiol in rat uterine epithelial and stromal cells.

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) cDNA was isolated from a subtracted cDNA library that selected for progesterone-induced transcripts from rat uterine stromal cells. In this study, the effects of progesterone and estradiol on the expression of HB-EGF in mature rat uterine epithelial and stromal cells have been examined. RNase protection assays and in situ hybridization demonstrated that progesterone stimulated expression of HB-EGF in rat uterine stromal cells, but repressed levels of HB-EGF mRNA in luminal and glandular epithelial cells. In contrast, estradiol treatment strongly enhanced HB-EGF expression in epithelial cells, but had no effect on mRNA levels for this growth factor in stromal cells. Progesterone treatment followed by estradiol injection stimulated HB-EGF expression in stromal cells and repressed expression in luminal and glandular epithelium. Stimulation of HB-EGF expression in stromal cells by progesterone was not inhibited by treatment with cycloheximide, demonstrating that HB-EGF mRNA expression is a primary response of stromal cells to progesterone. These results reveal that expression of HB-EGF is stimulated in epithelial and stromal cells in vivo under the same hormonal conditions that induce cell proliferation in each of these cell types and strongly suggest that HB-EGF may mediate the mitogenic effects of steroid hormones in the rat uterus.

Animals

Progesterone regulation of heparin-binding epidermal growth factor-like growth factor gene expression during sensitization and decidualization in the rat uterus: effects of the antiprogestin, ZK 98.299.

The ability of uterine stromal cells to respond to embryo attachment or an artificial decidual stimulus and the development and maintenance of decidual cells are primarily regulated by progesterone (P), yet few P-induced genes have been described. Previously, we reported that P stimulated the expression of heparin-binding epidermal growth factor-like growth factor (HB-EGF) messenger RNA (mRNA) in rat uterine stromal cells. We now present evidence that the effects of P on HB-EGF expression are blocked by treatment with ZK98299 (onapristone), indicating that they are receptor mediated. Additionally, animals treated with ZK98299 alone or together with P failed to respond to a decidual stimulus, suggesting that stromal cell sensitivity was inhibited. These results provide further evidence that HB-EGF plays a role in the development of stromal cell sensitivity to decidual stimuli. Expression of HB-EGF mRNA was also investigated in differentiating stromal cells after an artificial decidual stimulus. HB-EGF mRNA levels increased from 8-48 h after stimulation, the same interval in which stromal cells exhibit maximal mitotic activity. In situ hybridization revealed that stromal and decidual cells were the major sources of HB-EGF mRNA during this period. These results suggest that HB-EGF acts as an autocrine/paracrine factor in stimulating stromal cell proliferation after the induction of decidualization.

Animals

Growth factor effects on endometrial epithelial cell differentiation and protein synthesis in vitro.

OBJECTIVE: To develop a baseline for projected studies of a rat endometriosis model. DESIGN: We investigated the effects of two macrophage-related growth factors, platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta), on proliferation, in vitro differentiation, and protein secretion of uterine epithelial cells from immature rats. Uterine epithelial cells grown on matrix covered filters were treated with growth factors (GFs) or estrogen and/or P. Incorporation of [35S]methionine by polarized uterine epithelial cell proteins and secretion of labeled proteins into apical and basal culture medium were examined. SETTING: Department of Cell Biology, Baylor College of Medicine, Houston, Texas. MAIN OUTCOME MEASURES: Cell associated and secreted proteins were resolved by gel electrophoresis, fluorography, and immunoblotting. Proliferation was quantified by cell counts in parallel cultures by hemocytometer. RESULTS: Estrogen and P increase protein synthesis by uterine epithelium. Transforming growth factor-beta depressed protein synthesis and secretion in uterine epithelial cells. Platelet-derived growth factor appears to have no effect on epithelial protein synthesis or secretion and does not modulate the effect of TGF-beta. Estrogen and P increase complement component 3 (C3) production by epithelial cells. CONCLUSION: Macrophage-secreted GFs may play a role in the development and maintenance of ectopic endometrial tissue. Both TGF-beta and ovarian steroids may participate in the dynamic regulation of protein synthesis by ectopic uterine epithelium. These molecules may indirectly affect the macrophage-stromal axis through nonspecific modulation of C3 secretion. Platelet-derived growth factor appears to have no direct effect on uterine epithelial cells. The recognized effect of PDGF on ectopic endometrial tissue is most likely mediated via the stromal component.

Analysis of Variance

Receptivity is a polarity dependent special function of hormonally regulated uterine epithelial cells.

Useful knowledge of the mechanisms which regulate ovoreceptivity and implantation remains elusive in spite of increasing efforts to apply the technology of biochemistry and to a lesser extent, cellular and molecular biology to the analysis of the problem. Existing models used to analyze interactions of the blastocyst and endometrial cells of the uterus have been unable to account for nongenotypic embryonic losses, particularly those following in vitro fertilization and embryo transfer. Separation of endometrial uterine epithelial (UE) and uterine stromal (US) cells was used to demonstrate that each cell type responds independently and interdependently to the same regulatory signals. Cultured by classical techniques UE cells proved unable to respond to steroid hormone signals. For this reason UE cell cultures could not be used to develop an experimental cell system that mimicked growth and development of UE cells in utero. The failure of classical UE cell cultures derived from their inability to maintain epithelial cell polarity. Polarity, the spatial asymmetry of plasma membrane domains, is intrinsic to the structure and function of an epithelial cell. Apical and basolateral surfaces have different lipid and protein compositions which are correlates of the special functions of that epithelial cell. As epithelial cells differentiate they must, in response to regulatory cues, direct the flux of membrane components moving into and out of each surface in order to establish the polarity characteristic of each stage specific expression. The acquisition of receptivity by the apical surface of the UE cell may be considered to be such a special function. To prove this hypothesis polarized cultures of primary UE cells had to be developed that were hormonally responsive. Such an experimental cell system could serve as a model for in vitro implantation. This essay describes such a culture system in which blastocysts cocultured with UE cells in the presence of estrogen, will as predicted, fail to attach. This polarized UE cell system provides a functional in vitro model to study ovoreceptivity. It is now feasible to initiate studies of hormonal regulation of the composition and function of UE cell plasma membranes as they reflect the nonreceptive, receptive, and refractory nature of its apical surface.

Animals

Characteristics of uterine aging.

Uterine aging is in part responsible for a decline in fecundity which begins in middle age in most mammals. Characteristics of uteri from a variety of animals in middle age and old age are reviewed and the factors which may be responsible for this decline discussed. These include age-related changes in the hypothalamus, pituitary, and ovaries; loss of number or function of steroid hormone receptors; morphological changes in the uterine epithelium; the accumulation of collagen fibrils in the uterine stroma; and loss or impairment of the decidual response. The ultrastructural morphology of uterine epithelial, stromal, and decidual tissue from 4 month old and 18 month old Fischer 344 rats is presented and compared.

Aging

Subcellular localization of Cdc42p, a Saccharomyces cerevisiae GTP-binding protein involved in the control of cell polarity.

The Saccharomyces cerevisiae Cdc42 protein, a member of the Ras superfamily of low-molecular-weight GTP-binding proteins, is involved in the control of cell polarity during the yeast cell cycle. This protein has a consensus sequence (CAAX) for geranylgeranyl modification and is likely to be associated, at least in part, with cell membranes. Using cell fractionation and immunolocalization techniques, we have investigated the subcellular localization of Cdc42p. Cdc42p was found in both soluble and particulate pools, and neither its abundance nor its distribution varied through the cell cycle. The particulate form of Cdc42p could be solubilized with detergents but not with NaCl or urea, suggesting that it is tightly associated with membranes. An increase in soluble Cdc42p was observed in a geranylgeranyltransferase mutant strain (cdc43-2ts) grown at the restrictive temperature. In addition, Cdc42p from a cdc42C188S mutant strain (that has an alteration at the prenylation consensus site) was almost exclusively in the soluble fraction, suggesting that membrane localization is dependent on geranylgeranyl modification at Cys-188. Immunofluorescence and immunoelectron microscopy experiments demonstrated that Cdc42p localizes to the plasma membrane in the vicinity of secretory vesicles that were found at the site of bud emergence, at the tips and sides of enlarging buds, and within mating projections (shmoo tips) in alpha-factor-arrested cells. These results indicate that Cdc42p is localized to the bud site early in the cell cycle and suggest that this localization is critical for the selection of the proper site for bud emergence and for polarized cell growth.

Amino Acid Sequence

Patterns of sialylation in differentiating rat decidual cells as revealed by lectin histochemistry.

Lectin histochemistry was used to demonstrate changes in the surface glycan distribution of uterine stromal cells as they differentiate to form decidual cells. Decidualization was induced in hormone-treated, ovariectomized rat uteri by needle scratch. Uterine tissue from days 2 to 8 of deciduoma development was examined with a panel of lectins specific for terminal non-reducing structures in N- and O-linked classes of glycoprotein glycan, including alpha 2,3- and alpha 2,6-linked sialic acid residues. Immunostaining for desmin was used to identify decidual cells. An increase in N-linked glycans associated with the cell surface and recognized by lectins from Phaseolus vulgaris (leukoagglutinin) (l-PHA), Pisum sativum (PSA) and Triticum vulgaris (WGA) was found during the early growth of decidual cells. As decidualization progressed regionally from the antimesometrial to mesometrial uterus, an increase in alpha 2,3-linked sialic acid residues was followed by a loss of the alpha 2,6-linked form. The results suggest that as stromal cells differentiate, glycoprotein biosynthesis and glycosyl transferase activity are altered. These changes in patterns of glycosylation may give rise to altered decidual cell-matrix and cell-cell interactions during differentiation and play a role in the modulation of decidual cell interactions with trophoblast during early placentation.

Animals

The VPH1 gene encodes a 95-kDa integral membrane polypeptide required for in vivo assembly and activity of the yeast vacuolar H(+)-ATPase.

Yeast vacuolar acidification-defective (vph) mutants were identified using the pH-sensitive fluorescence of 6-carboxyfluorescein diacetate (Preston, R. A., Murphy, R. F., and Jones, E. W. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 7027-7031). Vacuoles purified from yeast bearing the vph1-1 mutation had no detectable bafilomycin-sensitive ATPase activity or ATP-dependent proton pumping. The peripherally bound nucleotide-binding subunits of the vacuolar H(+)-ATPase (60 and 69 kDa) were no longer associated with vacuolar membranes yet were present in wild type levels in yeast whole cell extracts. The VPH1 gene was cloned by complementation of the vph1-1 mutation and independently cloned by screening a lambda gt11 expression library with antibodies directed against a 95-kDa vacuolar integral membrane protein. Deletion disruption of the VPH1 gene revealed that the VPH1 gene is not essential for viability but is required for vacuolar H(+)-ATPase assembly and vacuolar acidification. VPH1 encodes a predicted polypeptide of 840 amino acid residues (molecular mass 95.6 kDa) and contains six putative membrane-spanning regions. Cell fractionation and immunodetection demonstrate that Vph1p is a vacuolar integral membrane protein that co-purifies with vacuolar H(+)-ATPase activity. Multiple sequence alignments show extensive homology over the entire lengths of the following four polypeptides: Vph1p, the 116-kDa polypeptide of the rat clathrin-coated vesicles/synaptic vesicle proton pump, the predicted polypeptide encoded by the yeast gene STV1 (Similar To VPH1, identified as an open reading frame next to the BUB2 gene), and the TJ6 mouse immune suppressor factor.

Amino Acid Sequence

Characterization of the Saccharomyces Golgi complex through the cell cycle by immunoelectron microscopy.

The membrane compartments responsible for Golgi functions in wild-type Saccharomyces cerevisiae were identified and characterized by immunoelectron microscopy. Using improved fixation methods, Golgi compartments were identified by labeling with antibodies specific for alpha 1-6 mannose linkages, the Sec7 protein, or the Ypt1 protein. The compartments labeled by each of these antibodies appear as disk-like structures that are apparently surrounded by small vesicles. Yeast Golgi typically are seen as single, isolated cisternae, generally not arranged into parallel stacks. The location of the Golgi structures was monitored by immunoelectron microscopy through the yeast cell cycle. Several Golgi compartments, apparently randomly distributed, were always observed in mother cells. During the initiation of new daughter cells, additional Golgi structures cluster just below the site of bud emergence. These Golgi enter daughter cells at an early stage, raising the possibility that much of the bud's growth might be due to secretory vesicles formed as well as consumed entirely within the daughter. During cytokinesis, the Golgi compartments are concentrated near the site of cell wall synthesis. Clustering of Golgi both at the site of bud formation and at the cell septum suggests that these organelles might be directed toward sites of rapid cell surface growth.

Biological Transport

Loss of collagen type VI from rat endometrial stroma during decidualization.

The expression of collagen type VI in the extracellular matrix of rat uterine endometrial stroma after a decidual stimulus was examined by immunolocalization and immunoblotting. The intermediate filament protein, desmin, was used as a marker to identify decidual cells. Tissue was examined from pregnant animals and from ovariectomized, hormone-treated rats in which decidualization had been induced artificially. In undifferentiated tissue from both groups of animals, collagen type VI was abundant, and desmin was present only in vascular smooth muscle cells. By 72 h after a decidual stimulus, however, collagen type VI had essentially disappeared from the matrix of the antimesometrial stromal compartment, and desmin was highly expressed in the decidualizing cells. During regression of the decidual tissue, collagen type VI began to reappear in the stromal matrix, whereas desmin expression declined as decidual cells degenerated. These results indicate that remodeling of the uterine extracellular matrix in response to embryo implantation is a function of the differentiating decidual cell.

Animals

Structure of the yeast endoplasmic reticulum: localization of ER proteins using immunofluorescence and immunoelectron microscopy.

The endoplasmic reticulum (ER) and other secretory compartments of Saccharomyces cerevisiae have biochemical functions that closely parallel those described in higher eukaryotic cells, yet the morphology of the yeast organelles is quite distinct. In order to associate ER functions with the corresponding cellular structures, we localized several proteins, each of which is expected to be associated with the ER on the basis of enzymatic activity, biological function, or oligosaccharide content. These marker proteins were visualized by immunofluorescence or immunoelectron microscopy, allowing definition of the S. cerevisiae ER structure, both in intact cells and at the ultrastructural level. Each marker protein was most abundant within the membranes that envelop the nucleus and several were also found in extensions of the ER that frequently juxtapose the plasma membrane. Double-labeling experiments were entirely consistent with the idea that the marker proteins reside within the same compartment. This analysis has permitted, for the first time, a detailed characterization of the ER morphology as yeast cells proceed through their growth and division cycles.

Cell Cycle

Developmental expression of adenosine deaminase during decidualization in the rat uterus.

Adenosine deaminase (ADA) is expressed in high concentrations at the fetal-maternal interface during postimplantation stages of gestation in the mouse. The experiments reported here were designed to identify the specific uterine cells that express ADA subsequent to implantation in the rat and to determine if embryonic cells contribute to ADA expression. The results of biochemical analysis demonstrate that ADA-specific activity increases to very high levels in implantation sites, beginning approximately 72 h after blastocyst attachment. Immunocytochemical analysis localized this ADA expression to the decidualized stromal cells in the antimesometrial region of the pregnant uterus. In experimentally induced deciduoma, these cells were capable of synthesizing high levels of both ADA and mRNA for ADA in the absence of embryos. The enzyme first appeared in decidual cell cytoplasm, approximately 72 h after induction of decidualization, and later was localized in the decidual cell nuclei. Since the expression of ADA and its mRNA in decidual cells follows the appearance of desmin, a protein marker for decidualization, by at least 48 h, ADA appears to be involved in the functioning of mature decidual cells rather than in stromal cell differentiation. The expression of ADA, but not desmin, was restricted to the antimesometrial decidual cells and decreased when these cells regressed. At mid-gestation ADA activity increased and was localized principally in the fetal placenta. The results presented here demonstrate that ADA is localized to the antimesometrial decidual cell and that its expression is consequent to differentiation of the uterine stromal cell and independent of any embryonic stimulus.

Adenosine Deaminase