bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cell density‐dependent decrease in cytoskeletal actin and myosin in cultured osteoblastic cells: Correlation with cyclic AMP changes
Authors: Egan John J. , Gronowicz Gloria , Rodan Gideon A. 
Published: May 2005

AbstractDuring bone development, osteoblasts form a contiguous layer along recently deposited osteoid and their morphology changes from fibroblast‐like to cuboidal. In culture, similar changes occur with increased cell density. We examined the possible role of cyclic AMP in this process since cyclic AMP was reported to increase in fibroblasts with increased cell density and similar shape changes were seen in response to parathyroid hormone, which also increases cellular cyclic AMP in osteoblastic cells. Osteoblast‐enriched rat calvaria cells were seeded at increasing density. The distribution between Triton X‐100 extractable and nonextractable actin and myosin was estimated by polyacrylamide gel electrophoresis. Intracellular cyclic AMP was estimated by prelabeling the cellular ATP pool with 3H‐adenine, followed by extraction and separation of 3H‐cAMP by high‐performance liquid chromatography. We found that osteoblastic cells contain about 40 pg actin and 5.3 pg myosin per cell. Around 60% of the actin and 70% of the myosin were in the nonextractable (crosslinked) form at cell densities of 10,000 to 50,000 cells per cm2. Above 50,000 cells/cm2, there was a cell density‐dependent reduction in crosslinked actin and myosin and a concomitant increase in cellular cyclic AMP. A comparable rise in cyclic AMP, produced by incubation with phosphodiesterase inhibitors, and treatment with other agents that increase cyclic AMP produced a similar decrease in the level of cytoskeletal actin and myosin. Cytochalasin B treatment, through its effect on actin polymerization, produced similar changes in cell shape and cytoskeletal actin. The findings suggest that an elevation in intracellular cyclic AMP may play a role in the density‐dependent changes in cell shape and microfilament organization observed in osteoblasts.
Effects of soluble factors and extracellular matrix components on vascular cell behavior in vitro and in vivo: Models of de‐endothelialization and repair
Authors: Madri Joseph A. , Marx Martin , Merwin June R. , Basson Craig , Prinz Christian , Bell Leonard 
Published: May 2005

AbstractVessel walls are comprised of several different cell populations residing in and on complex extracellular matrices. Each of the vascular cell types has diverse and sometimes unique functions and morphologies, and each has roles in repair processes following injury. Large vessel endothelial cells are known to respond to denudation injury by sheet migration and proliferation. This is in contrast to the migration through soft tissues with tube formation and subsequent lumen formation exhibited by microvascular endothelial cells in response to injury. Vascular smooth muscle cells of larger vessels respond to injury by migration from the arterial media into the intima, proliferation, and matrix biosynthesis, ultimately causing intimal thickening. Both these cell types exhibit “dysfunctional” phenotypes during their responses to injury. Microvascular cell responses to injury, while extremely variable, are less well documented. Specifically, responses to injury by microvascular endothelial vascular cells appear to be modulated, in part, by the composition and organization of the surrounding matrix as well as by the various soluble factors and cytokines found at sites of injury, suggesting that the extracellular matrix and soluble factors modulate each other's effects on local vascular cell populations following injury.
Lectin cell adhesion molecules (LEC‐CAMs): A new family of cell adhesion proteins involved with inflammation
Authors: Lasky Laurence A. 
Published: May 2005

AbstractThe means by which leukocytes, including lymphocytes, monocytes, and neutrophils, migrate from the circulation to sites of acute and chronic inflammation is an area of intense research interest. Although a number of soluble mediators of these important cellular interactions have been identified, a major site of great importance to the inflammatory response is the physical interface between the white cell and the endothelium. This critical association is mediated by an array of cell surface adhesion molecules. Previous data have demonstrated that the integrin subfamily of heterotypic adhesion molecules was a major component of these adhesive interactions, although it was clear that other, non‐integrin‐like molecules of unknown identity also seemed to be involved during the inflammatory process. A number of these other cell‐surface glycoproteins which may be involved with inflammation have recently been characterized by molecular cloning. These glycoproteins, including the peripheral lymph node homing receptor (pln HR), the endothelial cell adhesion molecule (ELAM), and PADGEM/gmp140, are all members of a family of proteins which are unified by the inclusion of three characteristic protein motifs: a lectin or carbohydrate recognition domain, an epidermal growth factor (egf) domain, and a variable number of short concensus repeats (scr) which are also found in members of the complement regulatory proteins. The appearance of lectin domains in all of these adhesion molecules is consistent with the possibility that these glycoproteins function by binding to carbohydrates which are expressed in a cell and/or region specific manner, and the members of this adhesion family have been given the generic name LEC‐CAM (lectin cell adhesion molecules). The discovery of adhesive proteins which appear to utilize the recognition of carbohydrate moieties for restricted cell adhesion may present new opportunities for the development of carbohydrate based anti‐inflammatory drugs.
Progesterone augments proliferation induced by epidermal growth factor in a feline mammary adenocarcinoma cell line
Authors: Modiano Jaime F. , Kokai Yasuo , Weiner David B. , Pykett Mark J. , Nowell Peter C. , Lyttle C. Richard 
Published: May 2005

AbstractSteroid hormones and peptide growth factors promote growth and development of normal mammary tissues and some types of breast cancer. Ovarian steroids may influence mammary growth directly or indirectly. The epidermal growth factor (EGF) family of proteins may also regulate mammary growth. These two pathways may function independently of each other or they may act in concert, with steroids inducing transcription of genes that encode growth factors or growth factor receptors. We used a feline mammary adenocarcinoma cell line (K12) to address whether there was an interrelation between progesterone (PGN) and EGF‐associated growth pathways. K12 cells responded to EGF by a dose‐dependent increase in proliferation. PGN or promegestone (R5020, a synthetic progestagen) alone did not stimulate K12 growth, but when EGF and PGN, or EGF and R5020 were combined, they were synergistic. This synergistic response was abrogated by the PGN receptor antagonist RU486 or by antibodies that blocked binding of EGF to its receptor. K12 cells expressed characteristic double‐affinity EGF receptors, as well as p185 (a functionally and structurally related protein, product of the neu gene) on their surface. PGN receptors were also found on intact cells and in cleared cytosols. Stimulation of K12 cells by PGN or by R5020 induced a two‐ to threefold increase in the number of high‐affinity surface EGF receptors after 24 h. Stimulation of these cells by PGN also affected the relative levels of phosphorylation of the EGF receptors and p185 within minutes, but not of other cellular phosphoproteins. Our results show that PGN enhances the EGF‐induced growth of K12 cells and suggest that this effect may be mediated at least partly via an increase in the number or function of high‐affinity EGF receptors.
Epidermis generated in vitro: practical considerations and applications
Authors: Parenteau Nancy L. , Nolte Cynthia M. , Bilbo Patrick , Rosenberg Mireille , Wilkins Leon M. , Johnson Eric W. , Watson Stephen , Mason Valerie S. , Bell Eugene 
Published: May 2005

AbstractThe technology for culture of epidermis is one of the most advanced to date for generation of a tissue in vitro. Cultured epidermis is already used for a number of applications ranging from use as a permanent skin replacement to use as an organotypic culture model for toxicity testing and basic research. While simple epidermal sheets have been grafted successfully, more advanced models for skin replacement consisting of both dermal and epidermal components are in development and being tested in a number of laboratories. One of the most advanced in vitro models is the living skin equivalent, an organotypic model consisting of a collagen lattice contracted and nourished by dermal fibroblasts overlaid with a fully formed epidermis.
Expression and function of interleukin‐6 in epithelial cells
Authors: Krueger James , Ray Anuradha , Tamm Igor , Sehgal Pravinkumar B. 
Published: May 2005

AbstractEpithelial cells both produce and are affected by interleukin‐6 (IL‐6). Experiments with an adenocarcinoma‐derived cell line (HeLa) reveal that activation of the transfected human IL‐6 promoter occurs largely through two partially overlapping second messenger (cAMP, phorbol ester)‐ and cytokine (IL‐1, TNF, serum)‐responsive enhancer elements (MRE I, −173 to −151 and MRE II, −158 to −145). MRE I contains the typical GACGTCA cAMP and phorbol ester‐responsive (CRE/TRE) motif, whereas MRE II defines a new CRE/TRE motif that contains an imperfect dyad repeat. The mechanism of dexamethasone‐mediated repression of IL‐6 gene expression in epithelial cells involves occlusion of the entire MRE enhancer region and of the core‐promoter elements (TATA‐box and RNA start site) by ligand‐activated glucocorticoid receptor. Enhanced levels of IL‐6 expression are observed in many solid tumors and in the hyperprolifer active (and glucocorticoid‐suppressible)lesions of psoriasis. In cell culture, IL‐6 enhances, inhibits, or has no effect on the proliferation of epithelial cells depending upon the cell‐type examined IL‐6 enhances proliferation of keratinocytes but inhibits that of breast carcinoma cell lines ZR‐75‐1 and T‐47D. In these breast carcinoma cells, IL‐6 elicits a major change in cell phenotype which is characterized by a fibroblastoid morphology, enhanced motility, increased cell‐cell separation, and decreased adherens type junctions (desmosomes and focal adhesions). The new data identify IL‐6 as a regulator of epithelial cell growth and of cell‐cell association.
Genetics of the malignant progression of astrocytoma
Authors: Mikkelsen Tom , Cairncross J. Gregory , Cavenee Webster K. 
Published: May 2005

AbstractThe tendency of human cancers to progress towards a more malignant state over time is a well described biological phenomenon. Recent investigations elucidating the genetic nature of malignancy and the possible mechanisms responsible for this evolution have suggested that a sequential pathway may exist whereby a cell population accumulates a nested set of genetic aberrations which endow it with the ability to overwhelm other populations and dominate the tumor. Human astrocytomas are a dramatic case in point, where specific genetic events of amplification and deletion are seemingly related to the stages of malignancy. The identification of these aberrations represents the first stage in the dissection of the temporal process of this cancer. Its augmentation with functional analyses will likely allow a fuller genetic description of in vivo transformation.
Regulation of the sarcoplasmic reticulum Ca2+‐release channel requires intact annexin VI
Authors: Hazarika P. , Sheldon A. , Kaetzel M. A. , Díaz‐Muñoz M. , Hamilton S. L. , Dedman J. R. 
Published: May 2005

AbstractAnnexin VI has eight highly conserved repeated domains; all other annexins have four. Díaz‐Muñoz et al. (J Biol Chem 265:15894, 1990) reported that annexin VI alters the gating properties of the ryanodine‐sensitive Ca2+‐release channel isolated from sarcoplasmic reticulum. To investigate the domain structure of rat annexin VI (67 kDa calcimedin) required for this channel regulation, various proteolytic digestions were performed. In each case, protease‐resistant core polypeptides were produced. Annexin VI was digested with V8 protease and two core polypeptides were purified by Ca2+‐dependent phospholipid binding followed by HPLC. The purified fragments were shown to be derived from the N‐ and C‐terminal halves of annexin VI, and demonstrated differential immunoreactivity with monoclonal antibodies to rat annexin VI. While both core polypeptides retained their ability to bind phospholipids in a Ca2+‐dependent manner, they did not regulate the sarcoplasmic reticulum Ca2+‐release channel as did intact annexin VI.
From growth arrest to growth suppression
Authors: Philipson Lennart , Sorrentino Vincenzo 
Published: May 2005

AbstractSince the introduction of the cell cycle concept two approaches to study growth regulation of cells have been proposed. One claims that cells are naturally quiescent, requiring a stimulatory encounter with growth factors for induction of cell division. The other considers cellular multiplication as the natural steady‐state; cessation of multiplication is thus a restriction imposed on the system. In the latter case emphasis is mainly on the signals involved in arrest of multiplication. This Prospect focuses on specific events occurring in mammalian cells at growth arrest, senescence, and terminal differentiation, specifically emphasizing the growth inhibitory factors, tumor suppressor genes, and other signals for growth suppression.
Affinity isolation of active murine erythroleukemia cell chromatin: Uniform distribution of ubiquitinated histone H2A between active and inactive fractions
Authors: Dawson Barbara A. , Herman Tim , Haas Arthur L. , Lough John 
Published: May 2005

AbstractThis laboratory recently reported the development of a biotin‐cellulose/streptavidin affinity chromatography method based on the DNase I sensitivity of active chromatin to isolate a DNA fraction from murine erythroleukemia (MEL) cells that is more than 15‐fold enriched in active genes (Dawson et al.: Journal of Biological Chemistry 264:12830–12837, 1989). We now report the extension of this technique to isolate and characterize chromatin that is enriched in active genes. In this approach, DNA in nuclei isolated from MEL cells was nicked with DNase I at a concentration that does not digest the active beta‐globin gene, followed by repair of the nicks with a cleavable biotinylated nucleotide analog, 5‐[(N‐biotin‐amido)hexanoamido‐ethyl‐1,37apos;‐dithiopropionyl‐3‐aminoallyl]‐2' ‐deoxyuridine 5'‐triphosphate (Bio‐19‐SS‐dUTP), during a nick‐translation reaction. After shearing and sonication of the nuclei to solubilize chromatin, chromatin fragments containing biotin were separated from non‐biotinylated fragments by sequential binding to streptavidin and biotin cellulose. The bound complex contained approximately 10% of the bulk DNA. Reduction of the disulfide bond in the biotinylated nucleotide eluted approximately one‐half of the affinity isolated chromatin. Hybridization analysis of DNA revealed that whereas inactive albumin sequences were equally distributed among the chromatin fractions, virtually all of the active beta‐globin sequences were associated with chromatin fragements which had bound to the affinity complex. Western blot assessment for ubiquitinated histones revealed that ubiquitinated histone H2A (uH2A) was uniformly distributed among active (bound) and inactive (unbound) chromatin fractions.
Fluorescent tetradecanoylphorbol acetate: A novel probe of phorbol ester binding domains
Authors: Balázs Margit , Szöllösi Jánòs , Lee William C. , Haugland Richard P. , Guzikowski Anthony P. , Fulwyler Mack J. , Damjanovich Sandor , Feuerstein Burt G. , Pershadsingh Harrihar A. 
Published: May 2005

AbstractProtein kinase C (PKC) has a prominent role in signal transduction of many bioactive substances. We synthesized the fluorescent derivative, phorbol‐13‐acetate‐12‐N‐methyl‐N‐4‐(N,N′‐di(2‐hydroxyethyl)amino)‐7‐nitrobenz‐2‐oxa‐1,3‐diazole‐aminododecanoate (N‐C12‐Ac(13)) of 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) to monitor the location of phorbol ester binding sites and evaluate its potential use as a probe of PKC in viable cells. The excitation maximum wavelength of N‐C12‐Ac(13) is close to 488 nm, facilitating its use in argon‐ion laser flow and imaging cytometry. When incubated with 100 nM N‐C12‐Ac(13) at 25°C, P3HR‐1 Burkitt lymphoma cells accumulated the dye rapidly, reaching maximum fluorescence within 25 min, 20‐fold above autofluorescence. Addition of unlabeled TPA significantly decreased the fluorescence of N‐C12‐Ac(13) stained cells in a dose‐dependent manner indicating specific displacement of the bound fluoroprobe. Competitive displacement of [3H]‐phorbol‐12,13‐dibutyrate ([3H]‐PBu2) from rat brain cytosol with N‐C12‐Ac(13) gave an apparent dissociation constant (Kd) of 11 nM. N‐C12‐Ac(13) possessed biological activity similar to TPA. Like TPA (final concentration 65 nM) N‐C12‐Ac(13), at a lower concentration (51 nM), induced expression of Epstein‐Barr viral glycoprotein in P3HR‐1 cells, differentiation of promyelocytic HL60 cells, and caused predicted changes in the mitotic cycle of histiocytic DD cells. Microspectrofluorometric images of single cells labeled with N‐C12‐Ac(13) showed bright fluorescence localized intracellularly and dim fluorescence in the nuclear region, consistent with dye binding mainly to cytoplasmic structures and/or organelles and being mostly excluded from the nucleus. Because of the high level of non‐specific binding of N‐C12‐Ac(13), this probe is not ideal for visualizing PKC in intact cells, but would be a valuable fluoroprobe to investigate the kinetic properties of purified PKC. Also, knowledge gained from these studies allows us to predict structures of fluorescent phorbols likely to have less non‐specific binding and, consequently, be potentially useful for monitoring PKC in viable cells.
Quantitative variations in gene expression: Possible role in cellular diversification and tumor progression
Authors: Nicolson Garth L. 
Published: May 2005

AbstractChanges in the quantitative expression of certain genes or in the amounts of their products can quickly stimulate progression to the metastatic phenotype. This has been done experimentally by transferring dominantly acting oncogenes such as c‐H‐rasEJ into susceptible cells or more recently by interfering with metastasis suppressor genes. In vivo such rapid qualitative changes in dominantly acting oncogenes or suppressor genes occur only rarely, and progression to highly metastatic phenotypes is thought to occur through a process involving the slow stepwise progression of a subpopulation of neoplastic cells to more malignant states. Such slow changes can be reversible and need not involve known dominantly acting oncogenes or metastatic suppressor genes, consistent with clinical and experimental observations on naturally occuring, highly advanced metastatic tumors. An important element in the natural progression of tumors to more malignant states may be their ability to circumvent host environmental controls that regulate growth and cellular diversity. They also evolve into heterogeneous cellular phenotypes, a process that appears to mainly involve quantitative changes in gene expression but can be rapidly stimulated in cell culture by the introduction of a dominantly acting oncogene or inhibited by the introduction of a suppressor gene. The oncogenes and suppressor genes that affect malignancy may control important steps in the quantitative regulation of sets of genes that are ultimately responsible for the cellular alterations seen in adhesion receptors, cell motility responses, cell‐cell communication components, degradative enzymes and their inhibitors, growth factor receptors, components that aid in escape from host surveillance mechanisms and others that are important in malignancy. Highly malignant cells that have slowly evolved in vivo may contain only a few qualitative gene changes but have undergone extensive cycles of diversification and accumulation of quantitative changes in the expression of genes that encode products that are related to malignancy and metastasis. Thus highly malignant cells can arise quickly due to specific qualitative changes in critical controlling genes or more slowly by less critical qualitative genetic changes together with cycles of cellular diversification and accumulation of quantitative changes in gene expression.
Association of vanadate‐sensitive Mg2+‐ATPase and shape change in intact red blood cells
Authors: Xu Y.‐H. , Lu Z.‐Y. , Conigrave A. D. , Auland M. E. , Roufogalis B. D. 
Published: May 2005

AbstractIntact human erythrocytes, initially depleted of Mg2+ by EDTA incubation in the presence of A23187, exhibit Mg2+‐dependent phosphate production of around 1.5 mmol per liter cells · h, half‐maximally activated at around 0.4 mM added free Mg2+. This appears to correspond to Mg2+‐stimulated adenosine triphosphatase (Mg2+‐ATPase) activity found in isolated membranes, which is known to have a similar activity and affinity for Mg2+. Vanadate (up to 100 μM) inhibited Mg2+‐dependent phosphate production and ATP breakdown in intact cells. Over a similar concentration range vanadate (3–100 μM) transformed intact cells from normal discocytes to echinocytes within 4–8 h at 37°C, and more rapidly in Mg2+‐depleted cells. The rate of Ca2+‐induced echinocytosis was also enhanced in Mg2+‐depleted cells. These results support previous studies in erythrocyte ghosts suggesting that vanadate‐induced shape change is associated with inhibition of Mg2+‐ATPase activity localized in the plasma membrane of the red blood cell.
Facile platelet adhesion to collagen requires metabolic energy and actin polymerization and evokes intracellular free calcium mobilization
Authors: Smith J. Bryan , Dangelmaier Carol , Selak Mary A. , Daniel James L. 
Published: May 2005

AbstractThe attachment of platelets to collagen‐coated microtiter plates at 20°C was inhibited strongly by depletion of metabolic energy or by addition of cytochalasins and was slightly inhibited by the intracellular Ca2+ chelator BAPTA. In keeping with their respective potencies as inhibitors of actin polymerization, cytochalasins D and H were the most potent inhibitors of adhesion, while cytochalasin B was the least potent. Energy depletion, cytochalasin D or, to a much lesser extent, BAPTA also inhibited platelet adhesion to collagen in a suspension assay system at 37°C. Collagen‐induced platelet cytosolic Ca2+ mobilization was inhibited up to 70% by cytochalasin D and abolished by energy depletion or BAPTA. Elevation of intracellular platelet calcium by treatment with ionomycin had little effect on platelet adhesion to collagen. We propose that rapid platelet spreading along collagen fibers is both energy‐ and actin‐dependent and necessary to produce maximal adhesion needed to elicit Ca2+ mobilization required for subsequent responses.