bluuebunny/crossref_metadata_2025 huggingface dataset:
Prediction of lobe growth and decay in centreless grinding based on geometric considerations
Authors: Harrison A J L , Pearce T R A
Published: September 2002

The growth and decay of lobes during centreless grinding have been studied by previous researchers using physical tests, time-domain simulation and identification of the roots of the Laplace transform of the characteristic equation. In this paper, the authors have extended these latter two methods to generate complete stability diagrams encompassing the entire practical range of machine set-up angles. These diagrams indicate that by varying the set-up angles in a prescribed manner during grinding, rapid rounding of arbitrarily lobed components can be achieved. This is verified via time-domain simulation.

Secondly, a novel and arguably more intuitive method of predicting the lobe growth and decay during centreless grinding is presented. The method considers the locations of the three points of contact between a lobed workpiece and the regulating wheel, the support plate and the grinding wheel. Axial symmetry is assumed. A unique circle can be drawn through these three points. The centre and radius of this circle vary continually as the workpiece rotates, in a manner dependent upon the workpiece's profile and the set-up angles. An above-average instantaneous radius leads, via machine stiffness, to a correspondingly larger grinding force and so to an increased instantaneous depth of cut. If this occurs when the trough of a lobe is being ground, the trough will become deeper and lobe growth will result. By contrast, if the instantaneous radius is below average when the trough is being ground, the lobe will decay. From this simple geometric consideration, the authors have calculated the rates of decay and growth of a range of numbers of lobes, across a wide range of set-up angles. The results are shown to agree well with those given using the previous methods.
On the removal of material along a polishing path by fixed abrasives
Authors: Zhang L , Tam H Y , Yuan C-M , Chen Y-P , Zhou Z-D , Zheng L
Published: September 2002

Material is removed when a surface is polished. This paper presents a model of the removal of material along a polishing path when a surface is polished by fixed abrasives. The proposed model assumes that the pressure distribution is Hertzian at the contact between the tool and the surface and the abrading rate follows the Archard wear law. According to the model, the profile of material removal is parabolic, and the depth and width of the profile may depend on the tool orientation. Polishing experiments are presented for the validation of the proposed model. Examples are also given to illustrate the applications of the model.
On-line monitoring of the keyhole welding pool in variable polarity plasma arc welding
Authors: Wang H , Kovacevic R
Published: September 2002

This paper presents a technique to monitor the weld penetration in real time by the front-side image sensing of the keyhole welding pool in variable polarity plasma arc welding (VPPAW) of a 6061 aluminium alloy. A novel optical system is developed to acquire the keyhole welding pool images from two sides of the plasma arc welding torch. The geometric sizes of the visible keyhole can be accurately obtained by using the developed image-processing algorithm. The variation in sizes of the visible keyhole has a close relationship with the real diameter of the keyhole. An artificial neural network (ANN) model is established to describe the relationship between the keyhole diameter and the geometrical sizes of the keyhole welding pool obtained from the image.
Methodology for bringing the intuitive design method's framework into design activities
Authors: Cavallucci D , Lutz P , Thiébaud F
Published: September 2002

Today, virtually all designers acknowledge that of the four fundamental activities which make up their daily tasks, the area that remains deficient is creativity, which they cannot systematize. Indeed, while the collection, construction and growth factors can be estimated and scheduled in terms of both deadlines and relevance, designers cannot guarantee the same objectives for creativity. The present investigations into the relevance of the assistance that a method can offer designers have led to the conclusion that TRIZ can provide a definite contribution, since it increases the relevance of the creative phase in design through systematization. This article describes how this methodological system, stemming from TRIZ, was introduced into the design process in the company.
Modifying therapeutic homework for patients with personality disorders
Authors: Freeman Arthur , Rosenfield Brad
Published: September 2002

Generalizing therapeutic gains made during therapy sessions to patients' daily lives is the most fundamental goal of psychotherapy. Assigning between‐session homework is an efficient method to teach new skills and to generalize those skills to the patient's natural environment. Given the persistent, chronic, and pervasive nature of the problems presented by the individual with a personality disorder, homework is not merely recommended but is an important and essential part of the therapeutic work. This article discusses modifying homework for personality‐disordered patients in the context of a literature review and case examples from the various Personality Disorder Clusters. Also offered are a number of techniques to overcome patient “resistance.” © 2002 Wiley Periodicals, Inc. J Clin Psychol/In Session 58: 513–524, 2002.
Schizosaccharomyces pombe spPABP, a homologue of Saccharomyces cerevisiae Pab1p, is a non‐essential, shuttling protein that facilitates mRNA export
Authors: Thakurta Anjan G. , Ho Yoon Jin , Dhar Ravi
Published: September 2002

Poly(A)‐binding proteins play important roles in mRNA metabolism in eukaryotic cells. We examined the role of the Schizosaccharomyces pombe homologue of the Saccharomyces cerevisiae poly(A)‐binding protein, Pab1p, in cellular growth and mRNA export. In contrast to PAB1, the sppabp gene is not essential for cellular viability. Like the human hPABP1 protein, spPABP is cytoplasmically localized and can shuttle between the nucleus and the cytoplasm. We found that a spPABP–GFP fusion protein expressed from a multicopy plasmid could suppress the growth and mRNA export defect of rae1‐16 7 nup184‐1 synthetic lethal mutations. However, about 20–25% of cells in the population exhibited a pronounced nuclear accumulation of poly(A)+ RNA. The same cells also localized the spPABP–GFP fusion to the nucleus, suggesting that the shuttling ability of spPABP is related to its function in mRNA export. When a heterologous nuclear export activity from spMex67p was fused to spPABP–GFP fusion protein, it overcame the nuclear retention but did not increase nuclear mRNA export. We discuss the implications of these observations in relation to how spPABP could function in mRNA export. Published in 2002 by John Wiley & Sons, Ltd.
Interdecadal variability in north‐east Australian tropical cyclone formation
Authors: Grant Andrew P. , Walsh Kevin J. E.
Published: September 2002

Interdecadal variability in north‐east Australian regional tropical cyclone formation is investigated. It is found that mean tropical cyclone formation is significantly higher during positive phases of the Interdecadal Pacific Oscillation (IPO) than during negative phases. This is related to changes in vertical wind shear in the north‐east Australian region. Copyright © 2003 Royal Meteorological Society.
Immunoelectron microscopic localization of cytochrome P‐450 isoenzyme CYP4A1 in liver, ileum and kidney of nafenopin treated male rats.
Authors: Persohn Elke , Thomas Helmut , Waechter Felix
Published: September 2002

A monoclonal antibody raised against and specific for cytochrome P‐450 isoenzyme CYP4A1 was used to investigate the subcellular distribution of this enzyme in the liver, kidney and ileum of nafenopin treated rats by means of immunoelectron microscopy. In the liver and kidney, labelling was restricted to peroxisomes and mitochondria of hepatocytes and proximal tubular epithelial cells whereas in ileum, immunolabelling was exclusively detected in mitochondria of absorptive cells.
Inward‐directed current generated by the Na+,K+ pump in Na+‐ and K+‐free medium
Authors: Efthymiadis Athina , Rettinger Jürgen , Schwarz Wolfgang
Published: September 2002

In Na+‐ and K+‐free solution, an inward‐directed current can be detected in Xenopus oocytes, which is inhibited by cardic glycosides and activated by ATP. Therefore, it is assumed to be generated by the Na+, K+ pump. At negative membrane potentials, the pump current increases with more negative potentials and with increasing [H+] in the external medium. This current is not observed when Mg2+ instead of Ba2+ is the only divalent cation present in the bath medium, and it does not depend on whether Na+ or K+ is present internally. At 5 to 10 mM Na+ externally, maximum pump‐generated current is obtained while no current can be detected in presence of physiological [Na+]. It is suggested that in low‐Na+ and K+‐free medium the Na+, K+ pump molecule can either form a conductive pathway that is permeable to Ba2+ or protons or operate in its conventional transport mode accepting Ba2+ as a K+ congener. A reversed pump mode or an electrogenic uncoupled Na+‐efflux mode is excluded.
Inhibition of an endogenous growth‐related proteinase enhances the recovery of a negative growth regulator from cultured human cells.
Authors: Manilal Sushila , Scott G. Kenneth , Tse Cynthia A.
Published: September 2002

The possibility that a growth‐related proteinase may act by degrading a negative growth regulatory protein has been investigated. Proteinase inhibitors which inhibit the enzyme also enhance the accumulation of the growth regulator by human fibroblasts. The negative growth regulator shows a similar specificity of inhibition of cellular growth to inhibitors of the growth‐related proteinase.
Induction of cell expansion of goldfish melanocytoma cells (GMM‐1) by epinephrine and dexamethasone requires external calcium.
Authors: Shih Yung‐Luen , Lo Szecheng J.
Published: September 2002

Treatment of GMM‐1 (a goldfish melanocytoma cell line) cells with epinephrine induced a rapid cell expansion (flattening of cells, extension and broadening of cellular processes) similar to the effect of dexamethasone reported previously (Shih et al., 1990). Studies on the possible involvement of secondary messengers in cell expansion indicated that (i) both 8‐bromo‐CAMP and forskolin caused cell shrinking (the opposite of cell expansion); (ii) TPA also caused cell shrinking; (iii) phospholipid derivatives, such as 1,2‐dioctanoyl‐sn‐glycerol, lysophosphatidic acid, and arachidonic acid caused cell expansion; and (iv) EGTA (calcium chelator) and nifedipine (calcium channel blocker) inhibited the effect of epinephrine. Together with the previous findings, these observations indicate that epinephrine and dexamethasone may share a common pathway in triggering an external calcium influx to cause cell expansion. The results of the effects of epinephrine agonists and antagonists, together with those of other workers, also show that there are multiple isoforms of adrenoceptor in the goldfish.
Studies on bcl‐2 protein in cultured lymphoid cells.
Authors: Filip Agata , Rożynkowa Danuta
Published: September 2002

Bc1‐2 protein was analysed in cell cultures derived from non‐Hodgkin lymphomas and chronic lymphocytic leukaemia patients. The presence of bc1‐2 proto‐oncogene protein product has been demonstrated on cytospin preparations utilising immunochemical methods. The correlation between the expression of the bc1‐2 protein and the appearance of proliferation‐ or differentiation related figures was studied. The reported bc1‐2 up regulation in malignant lymphoid cells was confirmed in haemo‐ and lymphopoietic progenitor cells as well as in TPA/PHA activated peripheral blood lymphocytes. In comparison to neoplastic cells lower intensity of bc1‐2 positive staining in freshly isolated cells from non‐leukaemic tissue was observed.
Macronuclear DNA demethylation is involved in the encystment process of the ciliate Colpoda inflata.
Authors: Palacios Gemma , Martin‐Gonzalez A. , Gutierrez Juan C.
Published: September 2002

Ciliate encystment is an eukaryotic cell differentiation process which involves a specific gene expression, to form the resting stage. In this study, we investigate, for first time, the DNA methylation pattern changes during encystment in the ciliate Colpoda inflata, and the 5‐azacytidine effect on growing cells and encystment. Results indicate that 5‐methylcytosine is present in macronuclear DNA of this ciliate and the 5‐azacytidine treatment induces encystment in growth conditions. From restriction enzyme digestion and 5‐azacytidine experiments, we conclude that a specific DNA demethylation is probably involved in the encystment gene expression of this ciliate.
Adhesion molecule expression and infiltrating maternal leucocyte phenotypes during blastocyst implantation in the pig
Authors: Whyte Anthony , Binns Richard M.
Published: September 2002

During the implantation period (days 14 to 30 post coitum) in Large White pigs substantial numbers of endometrial sub‐epithelial leucocytes were observed from about day 18 p.c. These were predominantly MHC Class II+, CD45+ and reactive for an accessory cell determinant carried by polymorphs and macrophages. Maternal lymphocytes of the CD2+, CD4−, CD8−, and a γδTcR+ subtype identified by the mAb 86D, were also present. Few CD4+, CD8+, or double‐positive lymphocytes were seen. CD2−CD4−CD8− γδTcR+ "Null" cells were seen in stroma adjacent to endometrial glands, as were CD2+ lymphocytes. There was no apparent upregulation of CD18, VLA‐4 or CD25. E‐selectin could not be detected on endothelium within uterine tissues or on trophectoderm, nor was a ligand for L‐selectin detectable. The luminal aspect of the uterine epithelium was strongly reactive with anti‐CD15 mAb, which was specifically blocked by lacto‐N‐fucopentaose III, indicating the presence of the LewisX determinant.
Polyamines and terminal deoxynucleotidyl transferase expression In KM 3 pre‐B cell line during phorbol ester induced differentiation.
Authors: Stabellini Giordano , Rapino Monica , Di Primio Roberto , Trubiani Oriana
Published: September 2002

The aliphatic polyamines, putrescine, spermine and spermidine belong to a category of molecules implicated in DNA replication. Their synthesis is strongly activated during the G1 period and they have been implicated in the regulation of cell proliferation and differentiation. Terminal transferase is a DNA polymerase present in pre‐T and pre‐B cells and its expression can be modulated by phorbol ester treatment. In this study we have monitored the relationship of intracellular polyamine levels with terminal deoxynucleotidyl transferase down‐regulation induced by 12‐0‐tetradecanoyl phorbol myristate 13‐acetate treatment in the human pre‐B KM‐3 cell line. Phorbol myristate acetate can cause an increase, at 4 and 8 hours of differentiation, of intracellular levels of putrescine as well as a decrease in terminal deoxynucleotidyl transferase synthesis showing the probable involvement that polyamines have in the differentiation process.
Differences in the motility of Amoeba proteus isolated fragments are determined by F‐actin arrangement and cell nucleus presence
Authors: Greȩbecka Lucyna , Pomorski Pawel , Łopatowska Anna
Published: September 2002

Isolated fragments produced by bisection of Amoeba proteus differ by their position in the original cell and by the presence or absence of the cell nucleus. Immediately after the operation, both types of anterior fragments preserve the former motory polarity, and do not interrupt locomotion. In the same time, all posterior fragments stop, round up and fail to react stimuli. In the second phase of experiment, these anterior fragments, which had no nucleus ceased to move, whereas the nucleated posterior ones resumed locomotion.

It was demonstrated, that the behaviour of a fragment is primarily determined by the peripheral F‐actin distribution, which is different depending on the origin of the fragment either from the anterior or from the posterior cell region. Later, the "inherited" F‐actin distribution may be stabilized or reorganized in the presence of the nucleus, or desorganized in its absence.
Leucocyte involvement in lectin‐induced deciduomata formation
Authors: Shaw Timothy J. , Murphy Christopher R.
Published: September 2002

Intra‐uterine injection of the lectin Concanavalin A (ConA) on day 5 of PSP induced a rapid and persistent infiltration of leucocytes into the rat uterine stroma. Although the infiltration of leucocytes was witnessed along the entire length of the uterine horn, areas of stromal oedema, indicative of decidualisation (as indicated by the positive Pontamine Sky Blue reaction), were only associated with regions in which the movement of leucocytes across the uterine epithelium was evident. Epithelial disruption and trauma was frequently noted within these regions. We believe that ConA may initiate decidualisation through indirectly causing epithelial trauma.
Effects of retinoic acid and dimethylsulfoxide on the morphogenesis of the sea urchin embryo
Authors: Sciarrino Serafina , Matranga Valeria
Published: September 2002

Sea urchin embryos of the species Paracentrotus lividus were treated continuously with different concentrations of all‐trans retinoic acid (RA) or dimethylsulfoxide (DMSO) at different developmental stages. A delay in embryonic development was observed when embryos were cultured in the presence of 2x10−5 M RA, between 1 and 12 hours of development. Hence, at 48 hours of development, while control embryos had reached the pluteus stage, RA‐treated embryos were at the prism stage. At 72 hours of development RA‐treated embryos recovered and continued normal development reaching the pluteus stage. No effect was observed when treatment was performed before 1 hour or after 12 hours of developmet. DMSO treatment had no effect on normal sea urchin embryo development, although we observed that pigment cells, clearly visible at the pluteus stage, become visible earlier with respect to control embryos. This report confirms the advantages that the sea urchin embryo offers for the study of problems in cellular and developmental biology.
The subcellular localization of newly synthesized cytochrome b5.
Authors: Carlsen Jens , Christiansen Kirsten
Published: September 2002

The fate of newly synthesized cytochrome b5 was studied in rat hepatocytes. Using an antibody specific for microsomal cytochrome b5, we found newly synthesized microsomal cytochrome b5 in both mitochondria and a mitochondria associated membrane fraction as well as in microsomes. Newly synthesized cytochrome b5 was quickly removed from the site of synthesis on free ribosomes and inserted into membranes at random. No migration of newly synthesized cytochrome b5 between cellular compartments was observed and therefore the assembly of the apoprotein with the heme moiety is apparently not taking place in any particular cellular compartment.
Proliferative capacity of embryonic chick notochord: a comparaison between PCNA positivity versus 3H‐thymidine incorporation.
Authors: Ghanem E. , Cornelissen M. , De Ridder L.
Published: September 2002

The proliferation rate of the embryonic chick notochord is measured during its functional period. Chick embryos of 4 different age groups respectively, 3.5, 4.5, 5.5 and 6.5 days of development, are selected. To measure proliferation activity in the notochords, two different methods are used. 1. Notochords are immunostained for proliferating cell nuclear antigen (PCNA). 2. After in vivo incorporation of 3H‐thymidine (3H‐TdR), DNA synthesis is visualized by autoradiography. After embedding in paraffin, 5 μm sections are cut and stained with PC10 monoclonal antibody and avidine biotin peroxidase complex (ABC) technique. Corresponding chick embryos of the same age groups are injected in ovo with 3H‐thymidine. After fixation 2 μm sections are made and exposed for autoradiography. Results obtained by PCNA and autoradiography labelling techniques for the evaluation of the proliferative capacity are different. Both however show a regression of the proliferative activity. The regression of the number of cells that went through S‐phase, as shown by autoradiography, is lower than that of PCNA positive cells. So, a number of notochordal cells stops in G1 phase. Our results moreover, indicate that the proliferating activity of the notochord of chick embryos is higher at young developmental stages than at later stages.
EARLY STEPS OF SPERM—EGG INTERACTIONS DURING MAMMALIAN FERTILIZATION
Authors: BARROS C. , CROSBY J. A. , MORENO R. D.
Published: September 2002

Mammalian eggs are surrounded by two egg coats: the cumulus oophorus and the zona pellucida, which is an extracellular matrix composed of sulfated glycoproteins. The first association of the spermatozoon with the zona pellucida occurs between the zona glycoprotein, ZP3 and sperm receptors, located at the sperm plasma membrane, such as the 95kDa tyrosine kinase‐protein. This association induces the acrosome reaction and exposes the proacrosin/acrosin system. Proacrosin transforms itself, by autoactivation, into the proteolytical active form: acrosin. This is a serine protease that has been shown to be involved in secondary binding of spermatozoa to the zona pellucida and in the penetration of mammalian spermatozoa through it. The zona pellucida is a specific and natural substrate for acrosin and its hydrolysis and fertilization can be inhibited by antiacrosin monoclonal antibodies. Moreover, inin vitrofertilization experiments, trypsin inhibitors significantly inhibits fertilization. The use of the silver‐enhanced immunogold technique has allowed immunolocalization of the proacrosin/acrosin system in spermatozoa after the occurrence of the acrosome reaction. This system remains associated to the surface of the inner acrosomal membrane for several hours in human, rabbit and guinea‐pig spermatozoa while in the hamster it is rapidly lost. In the hamster, the loss of acrosin parallels the capability of the sperm to cross the zona pellucida. Rabbit perivitelline spermatozoa can fertilize freshly ovulated rabbit eggs and retain acrosin in the equatorial and postacrosomal region. These spermatozoa also show digestion halos on gelatin plates that can be inhibited by trypsin inhibitors. This evidence strongly suggests the involvement of acrosin in sperm penetration through the mammalian zona. Recently it was shown, however, that acrosin would not be essential for fertilization. It is likely, then, that such an important phenomenon in the mammalian reproductive cycle would be ensured though several alternative mechanisms.
EPITHELIAL CELL MIGRATION IN THE INTESTINE
Authors: HEATH JULIAN P.
Published: September 2002

Despite significant advances in our understanding of the roles of the cytoskeleton and matrix receptors in cell locomotion, derived largely fromin vitrostudies on the movement of epithelial cell sheets and isolated cells, the mechanism of epithelial cell migration in the adult intestine remains an enigma. The primary function of the epithelial cell cytoskeleton seems to be in the maintenance of the apical region of the epithelium facing the gut lumen. There we find the brush border, with its associated enzymes, and the intercellular adhesion complexes that give the epithelium its cohesiveness and its barrier function. Curiously, there is little in the way of an organized cytoskeleton in the basal region of the epithelium adjacent to the basement membrane on which the epithelium is presumed to migrate. In this short review, I focus on what is known about epithelial migration from our understanding of the structure of the epithelium and from studies on wound healing, and indicate some avenues for future study.
TRANSCELLULAR BIOSYNTHESIS OF CYSTEINYL LEUKOTRIENES BY KUPFFER CELL—HEPATOCYTE COOPERATION IN RAT LIVER
Authors: FUKAI FUMIO , SUZUKI YOSHITAKA , NISHIZAWA YOSHINORI , KATAYAMA TAKASHI
Published: September 2002

We previously proposed that an enzymatic cooperation between Kupffer cells and hepatocytes may play an important role in cysteinyl leukotriene (LT) production in rat liver. Anin vitrotranscellular synthesis cysteinyl LTs by a Kupffer cell—hepatocyte coculture system was characterized here. Kupffer cells alone, with A23187 stimulation, did not generate cysteinyl LTs until supplemented either with isolated hepatocytes or with LTC4synthase and glutathione, indicating that Kupffer cells can synthesize LTA4but not convert it into LTC4. In contrast, hepatocytes converted the LTA4into cysteinyl LTs and further degraded the cysteinyl LTs. Cysteinyl LT production by the Kupffer cell—hapatocyte coculture system was optimized by addition of 1–3% serum albumin to the culture and by bringing the cell—cell distance closer to less than 3μ. Tumour necrosis factor also stimulated cysteinyl LT production by the coculture system. From these results, it is expected that the Kupffer cell—hepatocyte transcellular system for cysteinyl LT production actually functionsin vivo.
LEUCOCYTE BINDING TO THE UTERINE EPITHELIAL CELL SURFACE DURING LECTIN‐INDUCED DECIDUALIZATION
Authors: SHAW TIMOTHY J. , MURPHY CHRISTOPHER R.
Published: September 2002

Intra‐uterine injection of the lectin Concanavalin A (ConA) on day 5 of pseudopregnancy induced a rapid and persistent infiltration of leucocytes into the rat uterine stroma. Although the infiltration of leucocytes was seen along the entire length of the uterine horn, areas of stromal oedema, indicative of decidualization (as indicated by a positive Pontamine Sky Blue reaction), were only associated with regions in which leucocytes had crossed the uterine epithelium and were present in the uterine lumen. Ultrastructural evaluation of the interaction of the luminal leucocytes with the apical surface of the uterine epithelium appeared strikingly similar to that of the blastocyst and the uterine epithelium during normal implantation. It is proposed that leucocytes, induced by ConA, may initiate a decidual response in a manner analogous to that of the blastocyst through surface epithelial interaction.
BASEMENT MEMBRANE ASSOCIATED CHANGES IN THE RAT VENTRAL PROSTATE FOLLOWING CASTRATION
Authors: DE CARVALHO FAUSTINO HERNANDES , LINE SÉRGIO R. P.
Published: September 2002

This study focuses on the basement membrane associated modifications that take place after androgen blockade, by studying some of its main components, through histochemical, immuno‐histochemical and Western blotting tests, and its ultrastructural aspects. It was demonstrated that laminin and collagen type IV remain associated with a thickened basement membrane and that there is an apparent increase in heparan sulfate content 21 days after castration. Ultrastructurally, basal lamina appeared extensively folded and pleated. It was also observed that detachment of epithelial cells is not dependent of basal lamina degradation and that the free basal lamina surfaces are folded by the action of adjacent cells. We have also observed some aspects of smooth muscle cell degeneration and death, that lead to modifications of the associated basal lamina. In this case, residual basal lamina also shows extensive folding. The results suggested that degradation of excess basement membrane does not occur or is a very slow process within the period examined, and that basement membrane is left re‐organized but ultrastructurally and compositionally unaffected.
RICIN‐BINDING PROTEINS ALONG THE ENDOCYTIC PATHWAY: THE MAJOR ENDOSOMAL RICIN‐BINDING PROTEIN IS ENDOSOME‐SPECIFIC
Authors: ALAMI MÉRIEM , TAUPIAC MARIE‐PIERRE , BEAUMELLE BRUNO
Published: September 2002

Ricin is internalized after binding at the cell surface via lectin activity of the B‐chain recognizing terminal galactose residues. Ricin‐A chain is then translocated to the cytosol from various endocytic structures. Cell death is the result of catalytic inactivation of protein synthesis. Using 125I‐ricin overlays, we examined the distribution of ricin binding‐proteins within highly purified preparations of plasma membrane vesicles, endosomes and lysosomes from lymphocytes. All compartments of the endocytic pathway had distinct profiles; some ricin‐binding proteins were present throughout the pathway; others were restricted to the plasma membrane and endosomes. The major endosomal protein recognized by 125I‐ricin, a 166kDa glycoprotein, was endosome‐specific. When endosomal proteins were solubilized before chromatography onto ricin‐agarose this protein was also by far the major specifically‐bound glycoprotein. This 166 kDa glycoprotein might be involved in ricin translocation from this compartment.
INTERCELLULAR COMMUNICATION IN RAPIDLY PROLIFERATING AND DIFFERENTIATED C6 GLIOMA CELLS IN CULTURE
Authors: GHOSH PARAMITA , SINGH UDAI N.
Published: September 2002

Glial cells in the brain are known to provide structural and functional supports to neurons. To sustain such a supportive role, they have developed cell‐to‐cell communicating gap junctional channels. The authors studied the effect of dbcAMP on gap junctional channels mediated communication in C6 cells, a rat glioma cell line. Quantitative assessment of coupled cells under microscope after microinjection of a fluorescent dye was taken as a measure of junctional permeability. An enhanced coupling between cells was observed following dbcAMP treatment and this elevated coupling was found to be dependent on the duration of exposure of cells to dbcAMP. The studies have focused on a subtle shift in the spatial organization of the functional channels to the processes of dbcAMP induced differentiated cells from the cell cytoplasms and membranes of dbcAMP untreated cells. Immunofluorescence study with affinity purified antibody against gap junction further confirmed the spatial distribution of gap junctional protein(s) in the processes and also showed an increase in the density of the protein at the intercellular spaces in dbcAMP induced differentiated C6 glioma cells.
MODULATION OF OSTEOGENIC DIFFERENTIATION IN HUMAN SKELETAL CELLS IN VITRO BY 5‐AZACYTIDINE
Authors: LOCKLIN RACHEL M. , OREFFO RICHARD O.C , TRIFFITT JAMES T.
Published: September 2002

Cellular differentiation is controlled by a variety of factors including gene methylation, which represses particular genes as cell fate is determined. The incorporation of 5‐azacytidine (5azaC) into DNA in vitro prevents methylation and thus can alter cellular differentiation pathways. Human bone marrow fibroblasts and MG63 cells treated with 5azaC were used as models of osteogenic progenitors and of a more mature osteoblast phenotype, respectively. The capacity for differentiation of these cells following treatment with glucocorticoids was investigated. 5azaC treatment led to significant expression of the osteoblastic marker alkaline phosphatase in MG63 osteosarcoma cells, which was further augmented by glucocorticoids; however, in human marrow fibroblasts alkaline phosphatase activity was only observed in glucocorticoid‐treated cultures. MG63 cells represent a phenotype late in the osteogenic lineage in which demethylation is sufficient to induce alkaline phosphatase activity. Marrow fibroblasts are at an earlier stage of differentiation and require stimulation with glucocorticoids. In contrast, the expression of osteocalcin, an osteoblastic marker, was unaffected by 5azaC treatment, suggesting that regulation of expression of the osteocalcin gene does not involve methylation. These models provide novel approaches to the study of the control of differentiation in the marrow fibroblastic system.
DETECTION OF INTRANUCLEOLAR CHROMATIN USING AN ULTRASTRUCTURAL IMMUNOLABELLING TECHNIQUE
Authors: Ghosh Sibdas , Paweletz Neidhard
Published: September 2002

The intranucleolar chromatin has been detected by feeding HeLa cells with bromodeoxyuridine and then using an immunocytochemical technique for its detection at the ultrastructural level. Labelling was mainly observed on the dense fibrillar component (DFC), which surrounds the fibrillar centres (FC). Labelling could also be seen on the network within the granular component (GC) of the nucleoli. Ring shaped nucleoli with large FCs showed labelling, in addition, on the FCs. Nucleoli from spread preparations of cells treated with hypotonic salt solution for different periods revealed a nucleolonemal network, consisting of units comparable to rDNA transcriptional units in length and associated with tufts of fibrils and granules. Thread‐like axes could be detected running through the units and in between two units. Chromatin could also be identified within these units by the immunolabelling technique. The observations strongly indicate that these units are identical with the rDNA transcriptional units. A dynamic nature of the nucleolar chromatin and nucleolar organization has been discussed.
MULTINUCLEATION AND PRESERVATION OF NUCLEOLAR INTEGRITY OF MACROPHAGES
Authors: SORIMACHI KENJI , NAORA HONAMI , AKIMOTO KAZUMI , NIWA AKIRA , NAORA HIROTO
Published: September 2002

Rat bone marrow‐derived macrophages formed multinucleated giant cells spontaneously when cultured in slide glass chambers or when induced with the polyanion acetyl lignin. Nuclei in such cells tended to cluster in distinct rings. DNA fragmentation appeared to occur in multinucleated cells, as detected by 3′ end‐labeling. Southern blot analyses, using probes specific for nucleolar and non‐nucleolar genes, indicated that chromatin DNA was fragmented whereas nucleolar DNA was relatively intact. Autoradiography revealed preservation, in multinucleated cells, of nucleoli into which radiolabeled uridine was incorporated. Multinucleated macrophages appeared to eventually fragment. Preserved integrity of nucleoli seems to be a feature of macrophage multinucleation, a process which apparently culminates in cell death.
A METASTATIC BREAST TUMOR CELL LINE, GI‐101A, IS ESTROGEN RECEPTOR POSITIVE AND RESPONSIVE TO ESTROGEN BUT RESISTANT TO TAMOXIFEN
Authors: MORRISSEY JOSEPH J. , RANEY SHULA
Published: September 2002

The progression of human breast cancer is often associated with a loss of estrogen dependence for growth, a resistance to estrogen antagonists such as tamoxifen, and the metastatic spread of the disease to secondary sites. Cell lines developed from such advanced breast tumors are often metastatic in athymic mice, show a loss of estrogen receptor mRNA and protein (ER−), and do not respond to 17β‐estradiol. However many advanced human breast tumors do express significant amounts of ER transcript, especially when analyzed by more sensitive methods of detection including RT‐PCR and Ribonuclease Protection Assay (RPA). No metastatic, ER+breast tumor cell line has previously existed to examine the role of ER in metastatic progression and acquired drug (tamoxifen) resistance. The GI‐101A cell line was recently developed from a metastatic breast tumor xenograft and is both tumorogenic and metastatic to the lungs and lymph node when injected into athymic mice, a pattern similar to that seen in patients. While Western blot analysis initially indicated that GI‐101A was ER−, analysis of ER mRNA by RT‐PCR and RPA have demonstrated the expression of ER (as well as EGF receptor and neu oncogene) transcripts. Functional ER in GI‐101A was confirmed by a clear growth response to 17β‐estradiol in culture. Optimal 17β‐estradiol concentrations were significantly lower for GI101A than for MCF‐7 (1nmas opposed to ≥10nm), and GI‐101A growth was inhibited at 17β‐estradiol concentrations above 10nm. Unlike MCF‐7 cells, GI‐101A shows constitutive expression of pS2 protein in hormone depleted media with no apparent induction by 17β‐estradiol supplimentation, as well as a resistance to the anti‐estrogen tamoxifen at concentrations up to 10nm. Finally, ER transcripts which likely represent an alternately spliced ER variant which has previously been shown to encode a constitutively active ER protein have been detected in GI‐101A at levels similar to the wild type transcript, and offer a possible mechanism for estrogen independence, tamoxifen resistance, and constitutive pS2 expression.
EFFECTS OF cAMP ON ERK MITOGEN‐ACTIVATED PROTEIN KINASE ACTIVITY IN HEPATOCYTES DO NOT PARALLEL THE BIDIRECTIONAL REGULATION OF DNA SYNTHESIS
Authors: Thoresen G. Hege , Johasen Ellen Joanne , Christoffersen Thoralf
Published: September 2002

Previous studies have indicated that cAMP has bidirectional effects on epidermal growth factor (EGF)‐induced DNA synthesis in cultured hepatocytes, acting to stimulate soon after plating (early G1) and to inhibit at later stages (nearer the G1/S transition). In this study we examined the role of the extracellular signal‐regulated kinase (ERK) subgroup (p42/p44) of the mitogen activated protein (MAP) kinases both at growth‐stimulatory and growth‐inhibitory conditions. When added at low concentrations early during culturing, glucagon and 8‐chlorophenylthio‐cAMP (8‐CPT‐cAMP) did not increase MAP kinase activity, but enhanced the subsequent DNA synthesis. However, when administered at 24h, glucagon and 8‐CPT‐cAMP decreased basal and EGF‐induced MAP kinase activity and also inhibited EGF‐induced DNA synthesis. Thus, although MAP kinase might play a role in the growth‐inhibitory effect, it does not seem to be involved in growth‐promoting regulation by cAMP in hepatocytes.
INHIBITION OF ANTIGEN AND CALCIUM IONOPHORE INDUCED SECRETION FROM RBL‐2H3 CELLS BY PHOSPHATASE INHIBITORS
Authors: Ludowyke Russell I. , Warton Krystina , Scurr Lyndee L.
Published: September 2002

The role of serine/threonine protein phosphatases PP1 and PP2A in mast cell secretion was investigated using the phosphatase inhibitors okadaic acid and calyculin A. Calyculin A (5–25n m) inhibited antigen‐induced secretion from a rat mucosal mast cell line (RBL‐2H3) when added in conjunction with the activator. Okadaic acid (250–1000n m) inhibited secretion only when added before activation and did so in a time‐ and concentration‐dependent manner. Both inhibitors caused the cells to become rounder, but only calyculin A induced membrane blebbing and a loss of adherence. Okadaic acid also inhibited secretion induced by the calcium ionophore A23187, in the presence or absence of PMA, indicating that the phosphatase inhibitors act on a component of the secretory pathway downstream of calcium mobilization. Okadaic acid increased the phosphorylation of a number of proteins, as did an analogue methyl okadaate, which also inhibited secretion, but less effectively. Okadaic acid induced the phosphorylation of triton‐insoluble proteins of 55, 18 and 16kDa. The 55kDa protein was identified as vimentin and okadaic acid induced its partial translocation to the triton‐soluble fraction. Our data indicate that full secretory function in mucosal mast cells requires phosphatase activity.
ULTRASTRUCTURE, HISTOLOGICAL DISTRIBUTION, AND FREEZE‐FRACTURE IMMUNOCYTOCHEMISTRY OF GAP JUNCTIONS IN RAT BRAIN AND SPINAL CORD
Authors: Rash John E. , Yasumura Thomas , Dudek F. Edward
Published: September 2002

The historical development of concepts of gap junctions as sites for electrical, ionic, and metabolic coupling is reviewed, from the initial discovery of gap junctions linking heart cells, to the current concepts that gap junctions represent ‘electrotonic synapses’ between neurons. The ultrastructure and immunocytochemistry of gap junctions in heart, brain, and spinal cord of adult rats is examined using conventional thin sections, negative staining, grid‐mapped freeze‐fracture replicas, and immunogold‐labeled freeze‐fracture replicas. We review evidence for neuronal gap junctions at ‘mixed’ (combined electrical and chemical) synapses throughout adult rat spinal cord. We also show immunogold labeling of connexin43 in astrocyte and ependymocyte gap junctions and of connexin32 in oligodendrocyte gap junctions. Ultrastructural and freeze‐fracture immunocytochemical methods have provided for definitive determination of the number, size, histological distribution, and connexin composition of gap junctions between neurons in all regions of the central nervous systems of vertebrate species.
CELL‐TO‐CELL COMMUNICATION VIA PLANT ENDOMEMBRANES
Authors: Cantrill L.C , Overall R.L , Goodwin P.B
Published: September 2002

Cell‐to‐cell communication was investigated in epidermal cells cut from stem internodal tissue of Nicotiana tabacum and Torenia fournieri. Fluorescently labelled peptides and dextrans were microinjected using iontophoresis into the cytoplasm andcortical endomembrane network of these cells. The microinjected endomembrane network was similar in location and structure to the endoplasmic reticulum (ER) as revealed by staining with 3,3′‐dihexyloxacarbocyanine iodide (DiOC6). No cell‐to‐cell movement of dextrans was observed following cytoplasmic injections but injection of dextrans into the endomembrane network resulted in rapid diffusion of the probes to neighbouring cells. It is proposed that the ER acts as a pathway for intercellular communication via the desmotubule through plasmodesmata.
PERMEABILIZATION BY STREPTOLYSIN‐O REVEALS A ROLE FOR CALCIUM‐DEPENDENT PROTEIN KINASE C ISOFORMS ALPHA AND BETA IN THE RESPONSE OF CULTURED CARDIOMYOCYTES TO HYPOSMOTIC CHALLENGE
Authors: Boyle Robert T. , Lieberman Melvyn
Published: September 2002

Immunocytochemical techniques indicate that the uninhibited activity of protein kinase C alpha and protein kinase C beta are necessary for a normal regulatory volume decrease (RVD) response of cultured chick embryo cardiomyocytes subjected to a hyposmotic environment. Antibodies against protein kinase C isoforms alpha, beta, gamma and epsilon were introduced into the cultured myocytes using a developed streptolysin‐O (SLO) permeabilization technique that allows the targeted cells to accumulate large biomolecules without perturbing their normal physiological state. The loaded cells were then tested for their ability to RVD when submitted to hyposmotic stimulus. Results show that exposing the cultured cells to SLO in the presence of antibodies against protein kinase C alpha and beta, prior to volume challenge, significantly slows the RVD rate. Additional experiments that combined anti‐alpha and anti‐beta antibodies in the same exposure media did not result in a significantly different rate than the anti‐alpha or anti‐beta rates alone. The evidence gained in this study is in agreement with previous work in the cultured chick embryo cardiomyocyte that report the involvement of a calcium dependent protein kinase C in the signal transduction pathway of the RVD.
CHANGES IN LIVER GANGLIOSIDES IN STREPTOZOTOCIN‐INDUCED DIABETIC RATS
Authors: Sánchez S. S. , Abregú A. V. , Aybar M. J. , Sánchez Riera A. N.
Published: September 2002

Diabetes mellitus is associated with various structural and functional liver abnormalities that affect the glycogen and lipid metabolisms. The effects of streptozotocin‐induced diabetes and of insulin supplementation to Sprague—Dawley diabetic rats on ganglioside patterns in liver were determined. Diabetic livers showed a tendency to hepatomegaly 3 weeks after STZ‐induction of diabetes. The concentration of total gangliosides in diabetic and non‐diabetic livers was similar, but the concentration of total gangliosides in the liver of insulin‐stabilized rats was slightly increased. Bidimensional TLC chromatographic analysis of gangliosides isolated from normal diabetic and insulin‐stabilized diabetic livers showed quantitative and qualitative changes. In comparison with normal controls, the densitometric analyses of diabetic liver ganglioside patterns had increased amounts of GM3, GM1, GD1b, and GT1b gangliosides, while GM2 could not be detected. The hepatic ganglioside pattern of insulin‐stabilized diabetic rats was partially restored, resembling the profile of normal rats. The activity of GalNAcT, GalT‐2 and SialT‐4 transferases was measured in liver microsomal fractions of the different groups of animals. Diabetic rats showed an increased activity of GalNAcT and a decrease in the activity of GalT‐2 and SialT‐4 compared with the controls. The enzymatic activities found in insulin‐treated rats showed a tendency to return to the values observed in normal control animals. The results evidenced that streptozotocin‐induced diabetes affects the liver ganglioside pattern and the ganglioside synthesis enzyme activity. The alterations found in ganglioside metabolism could represent one of the earliest changes associated with the diabetic pathology.
ABNORMALITIES OF SPINDLE AND CYTOKINE BEHAVIOR LEADING TO THE FORMATION OF MEIOTIC RESTITUTION NUCLEI IN INTERGENERIC CEREAL HYBRIDS
Authors: Shamina Natalia , Dorogova Natalia , Goncharov Nicolay , Orlova Albina , Trunova Svetlana
Published: September 2002

Mobile stages of meiosis have been analysed by visualizing the spindle in fertile cereal F1 hybrids. We describe four different mechanisms of the formation of restitution nuclei in meiotic division: (1) centripetal migration of telophase chromosome groups from the poles of a curved spindle at early telophase; (2) centripetal migration of the chromosome groups at late telophase when cell plate formation has failed; (3) preferable migration of univalents to one of the poles although spindle appearance is morphologically normal; and (4) in the absence of chromosome segregation where kinetochore fibers have failed to form.
PARTIAL CHARACTERIZATION OF MOSQUITO LARVAE EXTRACT INDUCING DNA SYNTHESIS ALTERATIONS ON HUMAN MONONUCLEAR CELLS
Authors: Ronderos J. R. , Salas M. A. , Rimoldi O. J. , Finarelli G.
Published: September 2002

A crude mosquito larvae and dialysed extract alters the mitotic rate of several epithelial cell populations in normal young and adult hepatectomized mice. A crude extract also showed a biphasic effect on the proliferation of human mononuclear cells (MNCs), either stimulating or inhibiting them depending on the dose applied. In the present paper, we assayed the effect of the dialysed mosquito larvae extract and two different protein fractions on human MNCs. Analysis of cell viability after culture indicated that the extract did not have toxic effects. Our results show a dual response of the MNCs to the dialysed, as well as to the protein fraction, with the highest molecular weight inhibiting or stimulating proliferation, depending on the dose applied. The protein fraction with the lowest molecular weight (range between 12–80kDa) showed only an inhibitory effect on cell proliferation.
MITOCHONDRIAL KINETICS DURING AMPHIBIAN ERYTHROPOIESIS RELATED TO HAEME SYNTHESIS
Authors: Cianciarullo Aurora M. , Bertho Álvaro L. , Meirelles Maria De Nazareth L.
Published: September 2002

Flow cytometry, light and epifluorescence microscopies and transmission electron microscopy were used to follow the mitochondrial kinetics during amphibian erythropoiesis. A similar behaviour in response to the induction of anaemia was observed in the diploid Bufo ictericus and the tetraploid Odontophrynus americanus. A high cellular activity was observed ten days after haemolytic anaemia induced by phenylhydrazine, based on the higher Rhodamine 123 uptake by the erythroid cells. In addition, the more intense expression of the mitochondrial enzyme cytochrome oxidase, isocitrate and succinic dehydrogenases were cytochemically detected at this stage. This suggests that erythroid cell mitochondria, at this time, could be in a more active functional state than at other stages. In both species, mitochondrial plasticity was observed during cell maturation. A progressive loss of oxidation—reduction enzyme expression seemed to follow changes at the mitochondrial cristae morphology, from transverse to longitudinal form, mainly at the 20th day of recovery from anaemia, possibly related to a natural loss of function. The presence of these mitochondrial enzymes in mitochondrion‐like organelles also favours their participation in the haeme synthesis, although with a reduced expression, since this suggests the presence of a complete and active enzymatic complex in these modified organelles. This also supports the idea that all these organelles are mitochondria in distinct metabolic stages, and not mitochondrion‐like organelles or haemosomes, as proposed by some authors.