bluuebunny/crossref_metadata_2025 huggingface dataset:
Chemical and cryo‐fixation for cadmium retention in a marine snail.
Authors: Nott J. A. , Ryan K. P. , Mavin L. J.
Published: September 2002

In the hepatopancreas of the marine snail Littorina littorea, tissue structure and elemental analysis are both adversely affected by dosage with cadmium (Cd) and preservation with an aqueous fixative saturated with hydrogen sulphide (H2S). The fixative, with and without H2S, leaches the intracellular phosphate and carbonate granules but the latter are more severely leached when H2S is added. When intracellular accumulations of cadmium are examined by X‐ray microanalysis (XRMA) cadmium retention is not enhanced by H2S. To obtain a balanced XRMA of cadmium with other labile elements like Mg and K, it is necessary to use cryo‐fixation.
Em evidence of myoblast origin in regenerating human skeletal muscle explants
Authors: Naidoo Poovalingam R.
Published: September 2002

EM study of cultured human skeletal muscle explants on 10 consecutive days after incubation made possible a record for the first time, the early events occurring during regeneration. After incubation, normal myonuclei underwent activation and dense granulation. Some myonuclei showed early transformation to presumptive myoblasts. The conclusion was that myonuclei transformed into myoblasts which developed into satellite cells (SC). These SC of myonuclear origin, proliferated, and fused forming myotubes that matured into myofibres, replacing damaged muscle. The findings have new implications for the current myoblast / cell transplant and gene transfer therapy research which may provide possible answers for muscular dystrophy in the future.
Gene regulation in intertypic heterokaryons of Solanum tuberosum and Nicotiana tabacum tissue protoplasts
Authors: van Kesteren W. J. P. , Bijmolt E. W. , Tempelaar M. J.
Published: September 2002

Activities of the ß‐glucuronidase (GUS) reporter enzyme were evaluated in transgenic plants, protoplasts, and intertypic heterokaryons of Solanum tuberosum and Nicotiana tabacum. With GUS under control of the promoter of the cauliflower‐mosaicvirus 35S RNA gene (CaMV), activities of the enzyme were nearly evenly distributed over the tissues of plants grown in vitro. Activities in microtubers of potato plants with the reporterenzyme under control of the promoter of granule‐bound starch synthase (GBSS) from S. tuberosum were higher than in leaves. The CaMV‐GUS construct present in leaf protoplasts showed an increased expression in biochemical and cytochemical assays after fusion with wildtype‐tuber protoplasts. Such an increase was not observed in the case of the GBSS‐GUS constructs. It was concluded that nuclei in plant heterokaryons are being influenced by the fusion partner, but that repressive, trans‐acting regulation factors in leaf protoplasts possibly prevent an increase of the expression of the chimeric GBSS‐gene in heterokaryons.
Microexudate‐mediated release of lipids is necessary for growth of Acanthamoeba cells.
Authors: Pigon Andrew
Published: September 2002

Synthetic medium collected from stationary phase cultures of Acanthamoeba castellanii can support limited growth of these cells if lipids are first extracted. Conditioned medium contains several lipids, both saponifiable and non‐saponifiable. Release of these lipids is probably mediated by release of microexudate, a large‐molecular complex with surface‐active properties.
Co‐culture, oviduct secretion and the function of oviduct‐specific glycoproteins.
Authors: Nancarrow Colin D. , Hill Julia L.
Published: September 2002

Co‐cultures of embryos with somatic cells, usually in the form of monolayers, or conditioned medium from these somatic cells, results in development past the early stage blocks and the formation of hatched blastocysts. Optimum rates of development are not achieved, however, and the task is to investigate components of the oviduct that are obligatory or facilitative for embryo development. Glycine and alanine are amino acids present in much higher concentrations in oviduct fluid than in serum or culture media. Glycoproteins specifically produced by the oviduct around oestrus bind to embryos and aid development but are absent from most culture media. These glycoproteins are induced by oestrogen in vivo but not in vitro. It is our contention that co‐cultures of mammalian embryos should include appropriate concentrations of amino acids and a source of embryotrophic glycoproteins as an additive or by including stromal cells in addition to epithelial cells.
Protective effects of nocardia delipidated cell mitogen on the mucosa of the small intestine after irradiation of germ‐free piglets.
Authors: Kozakova Hana , Mandel Leos , Trebichavsky Ilja , Kolinska Jirina , Barot‐Ciorbaru Rita
Published: September 2002

The radioprotective effect of the bacterial immunomodulator Nocardia delipidated cell mitogen (NDCM) on intestinal mucosa and disaccharidase activities was studied in irradiated germ‐free piglets. Three‐week‐old germ‐free (GF) piglets were intragastrically pretreated with 1 mg NDCM per 1 kg body weight. The piglets were whole‐body irradiated with 2.5 Gray five days after the NDCM pretreatment and sacrificed eight days after irradiation. In the non‐irradiated group of GF piglets, NDCM application stimulated lactase activity and markedly increased sucrase activity. This stimulatory effect of NDCM disappeared after irradiation and the piglets exhibited a normal activity of lactase in the jejunal brush‐border membrane vesicles, while the sucrase activity decreased to the level found in irradiated controls. NDCM‐pretreated intestinal mucosa contained some infrequent lymphocytes which disappeared from the control irradiated tissue. It also exhibited less injury of the epithelium and stroma cells.
GLYCOPROTEINS OF THE NUCLEAR MATRIX FROM CHICKEN LIVER CELLS
Authors: Ferraro Anna , Eufemi Margherita , Altieri Fabio , Cervoni Laura , Turano Carlo
Published: September 2002

The nuclear matrix from chicken liver cells contains a small amount of glycoproteins recognized by Concanavalin A. These proteins are present not only in the peripheral matrix, but also in the internal one. In this latter localization many glycoprotein species appear, by cross‐linking experiments, to be placed in the proximity of DNA. The effect of a partial enzymatic deglycosylation of matrix preparations suggests that these proteins contribute to the stabilization of the native matrix structure.
Identification of a macrophage‐like mSubset in the axial organ sea star Asterias rubens.
Authors: Leclerc M. , Maitre F. , Contrepois L.
Published: September 2002

The axial organ of the sea star Asterias rubens is a primitive immune organ. In addition to T and B‐like cells phagocytic cells were also identified. In this report we demonstrate that these cells can be identified using immunocytochemistry together with the mouse to human monoclonal antibody CD68 KP1, normally used as a macrophage marker.
Hydrogen peroxide induces microvilli on human retinal pigment epithelial cells in culture.
Authors: Reid G. G. , Edwards J. G. , Marshall G. E. , Sutcliffe R. G. , Lee W. R.
Published: September 2002

We have found that hydrogen peroxide (10−4 − 10−2 M) rapidly induces microvilli on separate cells and confluent sheets of human retinal pigment epithelium in culture. t‐butyl hydroperoxide and sodium arsenite do not induce microvilli. A role for hydrogen peroxide as an intercellular messenger has previously been proposed in the inflammatory response, in which hydrogen peroxide from phagocytes may signal to vascular endothelial cells. Our observations thus provide a second example of the induction of what may be a physiological response by this potentially toxic agent. In the retina, hydrogen peroxide released from illuminated photoreceptors may elongate the microvilli which extend into the spaces between them. Increased numbers of microvilli and their protrusion further into the photoreceptor layer may enhance various interactions between the two cell types, including the antioxidant functions of the epithelium.
Transforming growth factor‐beta 3.
Authors: Cox David A.
Published: September 2002

Transforming Growth Factor‐Beta (TGF‐β) is the general name for a family of naturally‐occurring polypeptides which have multiple regulatory effects on cell proliferation and differentiation. Over the last decade it has become apparent that TGF‐βs can be produced by most cell types and exert a wide range of effects in a context‐dependent autocrine, paracrine or endocrine fashion via interactions with distinct receptors on the cell surface. This review summarizes current knowledge concerning the molecular and cellular biology of TGF‐β3, the most recently described mammalian isoform, and focuses on those physiological actions which may lead to clinical applications, particularly in the indication areas of wound healing and chemoprotection.
DISTRIBUTION OF ANNEXIN I DURING NON‐PATHOGEN OR PATHOGEN PHAGOCYTOSIS BY CONFOCAL IMAGING AND IMMUNOGOLD ELECTRON MICROSCOPY
Authors: HARRICANE MARIE‐CÉCILE , CARON EMMANUELLE , PORTE FRANÇOISE , LIAUTARD JEAN‐PIERRE
Published: September 2002

Annexin I is an abundant protein in U937 cells differentiated towards a macrophagic phenotype. These cells become able to killEscherichia coli, however, the intracellular pathogenBrucella suis, known to interfere with phagosome maturation, multiply in these differentiated cells. We have analysed by confocal and electron microscopy the cellular localization of annexin I during phagocytosis of yeast, non‐pathogenicE. coliand the intracellular pathogenB. suis. Using immunocytochemical detections annexin I was found mainly as patches in the cytoplasm of uninfected cells. Upon phagocytosis of yeast orE. coliorganisms, annexin I rapidly translocated and concentrated around phagosomes. On the other hand, annexin I was never detected around liveB. suis‐containing phagosomes. However, when dead brucellae were used, annexin I did translocate to the periphagosomal region. Our results suggest that annexin I could play a role in the molecular mechanism of phagosome maturation, which is impaired by some intracellular pathogens.
ANTISENSE INHIBITION OF THE 27 kDa HEAT SHOCK PROTEIN PRODUCTION AFFECTS GROWTH RATE AND CYTOSKELETAL ORGANIZATION IN MCF‐7 CELLS
Authors: MAIRESSE N. , HORMAN S. , MOSSELMANS R. , GALAND P.
Published: September 2002

MCF‐7 cells were co‐transfected with the human HSP27 antisense cDNA and the neomycin resistance gene, included in the constitutive expression vector pSVL, and the phenotypical changes associated with decreased expression of the HSP27 protein were analysed. Three out of 10 neomycin‐resistant clones obtained proliferated normally and showed a normal HSP27 content (Western blot). The seven other clones (designated as αHSP27 clones) were characterized by a dramatic growth inhibition associated with alterations in cellular morphology. Cells became progressively hypertrophied, exhibited lamellar protrusions and tended to lose contact with each other. They also acquired characteristics of secretory cells, namely the presence of numerous refractile granules and secretory canaliculi. Among the αHSP27 clones, two were immunocytochemically analysed for HSP27 content. Both clones were immunonegative for HSP27, contrary to parental cells and neo transfectants. Actin immunostaining in one of these HSP27 negative clones revealed that microfilament organization changed from diffuse to punctate distribution. Our data support the current concept of a role for HSP27 in cell growth and differentiation and further suggests that this might occur through a control on actin polymerization‐depolymerization.
IMPRINTING EFFECTS OF THREE AMINO ACIDS (ALANINE, LYSINE AND GLYCINE) AND THEIR OLIGOPEPTIDES INTETRAHYMENA PYRIFORMIS. DATA FROM THE HORMONE AND HORMONE RECEPTOR EVOLUTION
Authors: CSABA GYÖRGY , KOVÁCS PÉTER , NOSZÁL BÉLA
Published: September 2002

Hormonal imprinting takes place at the first encounter of the hormone and receptor, and results in a changed binding capacity and reaction of the cell and its progeny generations. The imprinting effect of three amino acids and their oligopeptides is studied using fluorescent‐labelled peptides. Glycine and lysine could provoke positive imprinting (increased binding in the progeny generations) for their own peptides, but alanine could not. Mostly positive imprinting was provoked by glycine and lysine peptides for their own peptides of different chain length. The optimal chain length provoking self‐imprinting was four for glycine, two for lysine and three for alanine. Except in this case, alanine was neutral or provoked mostly negative imprinting. After reaching the optimal chain length, there is a decline in binding. Evolutionary conclusions are discussed.
IMMUNOHISTOCHEMICAL LOCALIZATION OF GABA IN CHAMELEON RETINA (CHAMALEO CHAMALEO)
Authors: QUESADA A. , CHMIELEWSKI C. , GÉNIS‐GÁLVEZ J. M. , PRADA F. A.
Published: September 2002

We used a policlonal antiserum against GABA and demonstated GABA‐immunoreactivity (GABA‐IR) in several populations of amacrine cells in the inner nuclear layer (INL), and other cells in the inner plexiform layer (IPL) of the central and peripheral retina of the chameleon. Horizontal cells do not contain GABA‐IR and the chameleon retina is therefore an exception among non‐mammals. GABA‐IR was not seen in cell bodies in the position of photoreceptor, bipolar and interplexiform cells suggesting that GABA is not involved in synaptic transmission in the outer plexiform layer of chameleon retina.
A COMPARATIVE STUDY ON THE SYNTHESIS OF THE NATRIURETIC HORMONE DOPAMINE IN OK AND LLC‐PK1CELLS
Authors: SOARES‐DA‐SILVA P. , SERRÃO M. P. , VIEIRA‐COELHO M. A.
Published: September 2002

TheVmaxvalues (in nmol/mg protein/15 min) for AAAD in OK cells (0.94±0.08) were found to be significantly (P<0.01) lower than those observed in LLC‐PK1cells (4.37±0.08). However, in both cell lines decarboxylation reaction was a saturable process with similarKmvalues (OK cells=1.1 mm(0.3, 1.8); LLC‐PK1cells=1.8 mm(1.6, 2.1)). Contrariwise to OK cells, decarboxylation ofl‐DOPA to dopamine in LLC‐PK1cells followed a linear (7.6±0.1 pmol/mg protein/min) non‐saturable kinetics till 120 min of incubation. The formation of dopamine from increasing concentrations ofl‐DOPA (10 to 500 μm) followed a non‐linear kinetics in both cell lines; the process ofl‐DOPA decarboxylation was saturated at low concentrations ofl‐DOPA with an apparentKmvalue of 11 μm(0.2, 22.6) in OK cells and 27.4 μm(11.1, 43.7) in LLC‐PK1cells. The formation of dopamine in LLC‐PK1cells (Vmax=2097±113 pmol/mg protein/6 min) was 13.7‐fold that occurred in OK cells (Vmax=153±10 pmol/mg protein/6 min). In conclusion, LLC‐PK1cells appear to be endowed with a greater ability to form dopamine from exogenousl‐DOPA when compared to OK cells.
TISSUE AND DEVELOPMENTAL SPECIFICITY OF A POLYSIALO‐GANGLIOSIDE SPECIES IN THE AMPHIBIAN XENOPUS
Authors: BERNARDINI GIOVANNI , RIZZO ANGELA M. , GORNATI ROSALBA , ROSSI FEDERICA , BERRA BRUNO
Published: September 2002

Xenopus embryos contain a considerable amount of a polysialo‐ganglioside not yet fully characterized; in this paper, we will refer to it as ganglioside X1. Preliminary experiments indicate asialo‐GM1 as the core structure of the ganglioside X1 and palmitic and oleic acid as the fatty acids of the ceramide moiety. Further analyses by comparative 2D‐TLC with adult fish and chick embryo brains indicate the pentasialilated ganglioside GP1c as the possible structure of X1. In the adult Xenopus, X1 characterizes the ganglioside pattern of the central nervous system while is absent in all the other tested tissues. At least two other more polar (presumably richer in sialic acid) bands are often visible under X1, both in embryos and in brain and spinal cord tissues of adult Xenopus. The persistence of polysialo‐gangliosides in the brain and spinal cord of adult amphibians could serve to guarantee a proper functioning of the central nervous system at low body temperature.
NUCLEAR ANTIGENS, AS DNA, OF DSB389 MAB AND SOME OTHER ANTI‐DESMIN MONOCLONAL ANTIBODIES
Authors: KAMEI HIROYA
Published: September 2002

The anti‐desmin monoclonal antibodies (MAbs) DSB389, AC54, and DSB860 recognize intermediate filaments (IFs) and nuclear antigens that appear granular, locate around chromosomes, and are insoluble following 0.5% Triton X‐100 and cetyltrimethylammonium bromide (CTAB) extraction. Nuclear antigens of the MAbs were searched for in an IFs‐deficient clone of SW13 cells. Reactive materials specific to DSB389, AC54, and DSB860 MAbs were trapped at the top of the gel of SDS—agarose—PAGE. The reactivity of the materials disappeared after treatment with DNase I. The reactivity, or trapping of this material at the top of the gel, required previous heat treatment of the sample before application to the gel. The MAbs recognized both single‐stranded and double‐stranded DNA in ELISA. These results indicate that at least the main nuclear antigens of DSB389, DSB860 and AC54 MAbs are DNA.
THE INFLUENCE OF DOSE FRACTIONATION ON RADIATION‐INDUCED HAEMOLYSIS OF HUMAN ERYTHROCYTES
Authors: ZABOROWSKI ANDRZEJ , SZWEDA‐LEWANDOWSKA ZOFIA
Published: September 2002

In this paper the radioresistance of human erythrocytes to radiation‐induced haemolysis was studied by means of the split‐dose technique. Human erythrocytes suspended in an isotonic Na‐phosphatase buffer, pH7.4 were exposed under air to gamma radiation. Erythrocytes were irradiated with a single dose of 2.7kGy and with the same total dose split into two fractions with an interval between the subsequent exposures. It seems that postradiation processes initiated by the first radiation dose of definite magnitude can lead to an increase in the resistance of erythrocytes to haemolysis under the influence of subsequent dose. The maximal effect of the fractionation (50% decrease of haemolysis) was observed when the first fraction was equal to 0.3–0.4kGy and the interval between exposures was equal to 3h.
USING ATOMIC FORCE MICROSCOPY TO INVESTIGATE PATCH‐CLAMPED NUCLEAR MEMBRANE
Authors: DANKER TIMM , MAZZANTI MICHELE , TONINI RAFFAELLA , RAKOWSKA AGNIESZKA , OBERLEITHNER HANS
Published: September 2002

Nuclear patch clamp is an emerging research field that aims to disclose the electrical phenomena underlying macromolecular transport across the nuclear envelope (NE), its properties as an ion barrier and its function as an intracellular calcium store. The authors combined the patch clamp technique with atomic force microscopy (AFM) to investigate the structure—function relationship of NE. In principle, patch clamp currents, recorded from the NE can indicate the activity of the nuclear pore complexes (NPCs) and/or of ion channels in the two biomembranes that compose the NE. However, the role of the NPCs is still unclear because the observed NE current in patch clamp experiments is lower than expected from the known density of the NPCs. Therefore, AFM was applied to link patch clamp currents to structure. The membrane patch was excised from the nuclear envelope and, after electrical evaluation, transferred from the patch pipette to a substrate. We could identify the native nuclear membrane patches with AFM at a lateral and a vertical resolution of 3nm and 0.1nm, respectively. It was shown that complete NE together with NPCs can be excised from the nucleus after their functional identification in patch clamp experiments. However, we also show that membranes of the endoplasmic reticulum can contaminate the tip of the patch pipette during nuclear patch clamp experiments. This possibility must be considered carefully in nuclear patch clamp experiments.
ADIPOCYTE DIFFERENTIATION FACTOR (ADF): A PROTEIN SECRETED BY MATURE FAT CELLS THAT INDUCES PREADIPOCYTE DIFFERENTIATION IN CULTURE
Authors: LI ZHEN‐HUA , CARRARO RAFFAELE , GREGERMAN ROBERT I. , LAU DAVID C. W.
Published: September 2002

A factor that is released into the culture medium of mature adipocytes and promotes the differentiation (adipogenic conversion) of preadipocytes has been partially characterized. The factor acts in a dose‐dependent manner on preadipocytes to produce up to a four‐fold increase in triacylglycerol (triglyceride) content and a nine‐fold increase in glycerol‐3‐phosphate dehydrogenase (GPDH) activity, a marker of the late phase of differentiation of preadipocytes. The material appears to be a protein, since it has a molecular weight (Superose‐12 gel exclusion chromatography) of about 53kDa, an isoelectric point (pl) of 4.7–4.9, and is inactivated by the proteases papain and chymotrypsin and extremes of pH (2 and 12). Considerations of molecular weight, isoelectric point, stability to specific proteases, and especially to the action of chemical agents [the adipogenic activity is not affected by either an oxidizing (KIO4) or a reducing agent (DTT)], lead to the conclusion that the differentiation factor is distinct from known cytokines. The authors suggest that the protein be designated adipocyte differentiation factor (ADF). ADF in vivo may act as a cytokine paracrine agent to regulate the differentiation of preadipocytes.
INDUCTION OF PREMATURE MITOSIS IN S‐BLOCKED ONION CELLS
Authors: Sen Rita , Ghosh Sibdas
Published: September 2002

Onion root‐tip cells were blocked at S‐phase by treating them with 5‐aminouracil (5AU). These cells were then further treated with caffeine/2‐aminopurine (Caf/2AP) or a combination of both in the presence of 5AU. These tyrosine kinase inhibitors were able to induce premature mitosis in the S‐blocked cells as evident from the breaks and gaps in the metaphase chromosomes and the presence of laggards and fragments in anaphase. Immunofluorescence showed normal spindle formation in these cells. Immunoblotting of cyclin B revealed that the level of cyclin B was slightly higher in the recovered and treated samples than the S‐blocked one. The level of p34was found to be almost equal in all three samples as expected. We failed to observe any significant difference in the level of p34containing phosphorylated tyrosine. Such premature induction of mitosis by the purine derivatives has also been reported in BHK cells. However, those cells failed to progress through mitosis. A comparative analysis indicates that the plant cells and the animal cells, perhaps, follow identical pathway for the initiation of mitosis. The possible causes for differential behaviour in mitotic progression in these cells have been discussed.
‘NON‐SYNAPTIC’ MECHANISMS IN SEIZURES AND EPILEPTOGENESIS
Authors: Dudek F. Edward , Yasumura Thomas , Rash John E.
Published: September 2002

The role of ‘non‐synaptic’ mechanisms (i.e. those mechanisms that are independent of active chemical synpases) in the synchronization of neuronal activity during seizures and their possible contribution to chronic epileptogenesis are summarized. These ‘non‐synaptic’ mechanisms include electrotonic coupling through gap junctions, electrical field effects (i.e. ephaptic transmission), and ionic interactions (e.g. increases in the extracellular concentration of K+). Several lines of evidence indicate that granule cells and pyramidal cells of the hippocampus, and probably other cortical neurons, can generate synchronized electrical activity after active chemical synaptic transmission has been blocked. This synchronized activity is sensitive to alterations in the size of the extracellular space, thus suggesting that electrical field effects and ionic mechanisms contribute to this synchronized activity. Recent studies also indicate that ‘non‐synaptic’ synchronization is quite prominent early in development. Electrophysiological data from hippocampal and neocortical slices have led to a re‐interpretation of the fast prepotentials (i.e. partial spikes) recorded in cortical pyramidal cells, suggesting that they may not be due to dendritic spike generation. Improvement in freeze‐fracture ultrastructural techniques have led to a re‐assessment of previous data on gap junctions in the nervous system and opened new approaches to the quantitative analysis and characterization of gap junctions on glia and neurons. Finally, new methods of dye/tracer coupling have the potential to provide a more rigorous basis for evaluating gap junctions and electrotonic communication between neurons in the mammalian central nervous system. Therefore, recent data continue to suggest that gap junctions and electrotonic coupling play an important role in neural integration, although additional studies using new techniques will be needed to address some of the controversial issues that have arisen over the last several decades.
D,L‐α‐DIFLUOROMETHYLORNITHINE, AN IRREVERSIBLE INHIBITOR OF ORNITHINE DECARBOXYLASE, INDUCES DIFFERENTIATION IN MEL CELLS
Authors: Choudhary Shailesh Kumar , Sharma Dipali , Dixit Aparna
Published: September 2002

In the present study, the effect of D,L‐α‐difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase (ODC), on Friend's murine erythroleukemia (MEL) cell differentiation is investigated. DFMO was able to induce differentiation of MEL cells in culture as determined by haemoglobin (Hb) content and percentage of cells synthesizing Hb detected by benzidine staining. DFMO at a concentration of 2m m resulted in about 70% benzidine‐positive cells on the fifth day. There was a time‐dependent increase in the percentage of benzidine‐positive cells starting from day three. However, only a 24h presence of DFMO in the medium was required to induce differentiation suggesting that DFMO switches on a pathway during this period leading to terminal differentiation of MEL cells. DFMO induced differentiation of MEL cells was sensitive to dexamethasone and 5‐bromo‐2′‐deoxyuridine.
APOPTOSIS INDUCED BY γ IRRADIATION IN PERIPHERAL BLOOD MONONUCLEAR CELLS IS NOT MEDIATED BY CYTOCHROME‐C RELEASE AND ONLY PARTIALLY INVOLVES CASPASE‐3‐LIKE PROTEASES
Authors: Louagie H. , Schotte P. , Vral A. , Cornelissen M. , Thierens H. , Beyaert R. , De Ridder L. , Philippe J.
Published: September 2002

Caspase 3 has been shown to be actively involved in the apoptotic process in thymocytes after γ‐irradiation. We examined caspase 3 activation in mature peripheral blood lymphocytes (PBL) after γ irradiation. Since the activation of caspase 3 is generally prceded by a decrease in mitochondrial membrane potential (ΔΨm) and cytochrome c release, these two parameters were also examined. Apoptosis in PBL after a 5‐Gy γ irradiation, is characterized by a decrease in ΔΨm, but surprisingly no release of cytochrome‐c and only a weak caspase 3 activation was noticed. In contrast, staurosporin treated PBL showed a decrease in ΔΨm with cytochrome‐c release and a clear caspase 3 activation. We were unable to block the decrease in ΔΨm with the caspase‐inhibitors zVAD‐fmk or zDEVD‐fmk after γ irradiation, but DNA fragmentation as measured by the TUNEL assay was partially inhibited. Therefore, in γ irradiated mature PBL, caspase‐dependent and ‐independent pathways, but not cytochrome c, seem to be involved in the apoptotic process.
EFFECTS ON PROTEIN KINASE C AND GENE EXPRESSION IN A HUMAN MAST CELL LINE, HMC‐1, FOLLOWING MICROWAVE EXPOSURE
Authors: Harvey Charles , French Peter W.
Published: September 2002

We used a resonant cavity which delivered a continuous wave exposure at 864.3MHz at an average specific absorption rate (SAR) of 7W/kg to determine non‐thermal biological effects of microwave exposure. A human mast cell line, HMC‐1, was used as the biological target. Cells were given three exposures each of 20‐min duration daily for 7 days. The temperature of the cell culture medium during the exposure fell to 26.5°C. Effects were seen on localization of protein kinase C, and expression of three genes of 588 screened. The affected genes included the proto‐oncogene c‐kit, the transcription factor Nucleoside diphosphate kinase B and the apoptosis‐associated gene DAD‐1. Stress response genes were variably upregulated. No significant effect on morphology or on F‐actin distribution was detected. We conclude that low‐power microwave exposure may act on HMC‐1 cells by altering gene expression via a mechanism involving activation of protein kinase C, and at temperatures well below those known to induce a heat shock response.
S100 IMMUNOREACTIVITY IS INCREASED IN REACTIVE ASTROCYTES OF THE VISUAL PATHWAYS FOLLOWING A MECHANICAL LESION OF THE RAT OCCIPITAL CORTEX
Authors: Cerutti S. M. , Chadi G.
Published: September 2002

After demonstration of the paracrine action of glial neurotrophic factors, gliosis has also been considered to be related to neuronal trophism and plasticity rather than solely a repair event following brain injury. S100 is a Ca2+binding protein, present mainly in astrocytes, that exerts paracrine trophic effects on several neuronal populations. This study analyses the presence of S100 protein by means of immunohistochemistry combined with stereology in the reactive glial cells of the rat visual pathways following a lesion of the visual cortex. Adult male Wistar rats were submitted to a unilateral aspiration of the occipital cortex or to a sham operation. One week later the rats were killed and their brain processed for immunochemistry. Single antibody immunohistochemistry was performed for the visualization of glial fibrillary acidic protein (GFAP, a marker for astrocytes), OX‐42 (a marker for microglia) and S100 protein. Double immunofluorescence procedures were applied for co‐localization of the S100/GFAP and S100/OX‐42. An optical dissector, point interceptors and rotators were used to quantify the degree of glial activation and the changes in the S100 immunoreactivity. We observed an intense microglial and astroglial reaction in addition to an increased S100 immunoreactivity in the occipital cerebral cortex, geniculate nucleus and hippocampus ipsilateral to the lesion. In the ipsilateral superior colliculus, an intense astroglial activation was accompanied by an up‐regulation of S100 immunoreactivity. Double‐immunofluoresence revealed an increased S100 immunoreactivity in reactive astrocytes, but not in the reactive microglia. Evidence has therefore been obtained that after mechanical trauma, the astroglial S100 protein participates in the trophism and plasticity of the injured visual pathways.
NEUTRAL ENDOPEPTIDASE‐24.11 (NEP) DEACTIVATES PDGF‐ AND TGF‐β‐INDUCED CELL SHAPE CHANGES IN INVERTEBRATE IMMUNOCYTES
Authors: Caselgrandi Eva , Kletsas Dimitris , Ottaviani Enzo
Published: September 2002

Using a cytofluorimetric assay, we found that immunocytes of the mollusc, Mytilus galloprovincialis, express CD10, a surface antigen known to be identical to neutral endopeptidase‐24.11 (NEP). The spectrofluorimetric analysis demonstrates that the growth factors PDGF‐AB and TGF‐β1 provoke an increase in NEP‐like activity in membrane preparations from the immunocytes, but have no effect on the soluble form in the serum. On the other hand, computer‐assisted microscopic image analysis reveals that NEP deactivates the PDGF‐AB‐ and TGF‐β1‐induced shape changes in immunocytes. However, Western blots show that, in solution, NEP does not cleave PDGF‐AB or TGF‐β1, indicating that the inactivation is not due to proteolysis. These results suggest a functional interplay in invertebrate immunocytes between growth factors and NEP, as previously shown in vertebrate cells.
IRON REGULATION OF TRANSFERRIN SYNTHESIS IN THE HUMAN HEPATOMA CELL LINE HEPG2
Authors: Barnum‐Huckins Katrina , Adrian Gwen S.
Published: September 2002

In human beings, serum transferrin levels increase during iron deficiency and decrease with iron overload. Yet, whether or not iron levels actually affect the synthesis of transferrin in human liver cells is not known. In previous studies, iron was shown to suppress the expression of chimeric human transferrin genes in livers of transgenic mice. The goal of this study was to determine if iron suppresses intact endogenous human transferrin synthesis by testing the effects of changes in iron levels on synthesis of transferrin in a human hepatoma cell line HepG2. In HepG2 cells, normalized35S‐metabolically labeled transferrin synthesis was consistently less following iron treatment with hemin or ferric citrate, than following treatment with an iron‐chelator deferroxamine. Thus, this study provides new evidence that iron can regulate synthesis of intact endogenous human transferrin.
ARREST IN METAPHASE AND ANATOMY OF MITOTIC CATASTROPHE: MILD HEAT SHOCK IN TWO HUMAN OSTEOSARCOMA CELL LINES
Authors: Erenpreisa Je , Ivanov A. , Dekena G. , Vitina A. , Krampe R. , Freivalds T. , Selivanova G. , Roach H.I
Published: September 2002

The exits from metaphase arrest and anatomy of mitotic catastrophe were studied in two human osteosarcoma cell lines, nontumorigenic HOS TE85 and its chemically transformed strain MNNG‐HOS, applying mild genotoxic damage by heat shock at 41.8°C for 24h. Under these conditions, both cell lines doubled or tripled their mitotic index entering arrest in metaphase. On return to 37°C, the arrest was either released or ended in apoptosis. The transformed strain showed a greater capacity to arrest in metaphase as well as a greater probability of developing the third pathway: to restitute this arrest in polyploid interphase. This, in turn, either entered an ‘endocycle’ or, following a delay, apoptosis. Thus, arrest in metaphase was a cross‐point of the mitotic cycle, apoptosis, and endocycle. Mitotic catastrophe can morphologically manifest combinations of elements of these three processes.
ANNEXIN I IS STORED WITHIN GELATINASE GRANULES OF HUMAN NEUTROPHIL AND MOBILIZED ON THE CELL SURFACE UPON ADHESION BUT NOT PHAGOCYTOSIS
Authors: Perretti Mauro , Christian Helen , Wheller Samantha K. , Aiello Ivana , Mugridge Kenneth G. , Morris John F. , Flower Roderick J. , Goulding Nicolas J.
Published: September 2002

Annexin I, a member of the calcium‐ and phospholipid‐binding annexin superfamily of proteins, is largely present in human neutrophils. To determine its exact intracellular distribution a combination of flow cytometry, confocal microscopy and electron microscopy analyses were performed on resting human neutrophils as well as on cells which had been activated. In resting neutrophils, annexin I was found to be present in small amounts in the nucleus, in the cytoplasm and partially also associated with the plasma membrane. The cytoplasmic pool of annexin I was predominant, and the protein was co‐localized with gelatinase (marker of gelatinase granules), but not with human serum albumin or CD35 (markers of secretory vesicles), or with lysosomes. Electron microscopy showed the presence of annexin I inside the gelatinase granules. Neutrophil adhesion to monolayers of endothelial cells, but not phagocytosis of particles of opsonized zymosan, provoked an intense mobilization of annexin I, with a marked externalization on the outer leaflet of the plasma membrane. Remaining intracellular annexin I was also found in proximity of the plasma membrane. These results provide a novel mechanism for annexin I secretion from human neutrophils, which is via a degranulation event involving gelatinase granules.
MACRONUCLEAR DIVISION AND CYTOKINESIS IN TETRAHYMENA
Authors: Numata Osamu , Fujiu Kenta , Gonda Kohsuke
Published: September 2002

Tetrahymena contains a micronucleus and a macronucleus. The micronucleus divides with typical mitosis, while the macronucleus divides amitotically. Although the mechanism responsible for macronuclear division was previously unknown, we clarified the organization of microtubules during macronuclear division. The macronuclear microtubules dynamically changed their distribution in an organized way throughout the macronuclear division. The macronuclear microtubules and the cytoplasmic microtubules cooperatively carried out the macronuclear division. When the micronuclear division was finished, p85 appeared at the presumptive division plane prior to the cytokinesis. The p85 directly interacted with calmodulin in a Ca2+‐dependent manner, and p85 and CaM colocalized to the division furrow during cytokinesis. Moreover, the Ca2+/CaM inhibitor, W7, inhibited the direct interaction between p85 and CaM, the localization of both proteins to the division plane, and the formation of the division furrow. Thus, Ca2+/CaM and p85 have important roles in initiation and progression of cytokinesis in Tetrahymena.
APHIDICOLIN INHIBITS DNA REPLICATION IN THE MICRONUCLEUS AND BLOCKS CYTOKINESIS IN PARAMECIUM CAUDATUM
Authors: Sabaneyeva Elena , Tao Weihai , Verbelen Jean‐Pierre
Published: September 2002

Aphidicolin, a selective inhibitor of DNA polymerase, totally blocks DNA replication in the micronucleus but not in the macronucleus of Paramecium caudatum. The ciliates no longer divide and after 4 days the DNA content of the macronucleus has increased by 64%. Concomitantly the cell volume has increased by 53%.
CYTOSKELETON‐INDUCED ALTERATIONS OF THE LECTIN ACTIVITY IN WINTER WHEAT UNDER COLD HARDENING AND ABSCISIC ACID (ABA)
Authors: Timofeeva Olga , Khokhlova Ludmila , Belyaeva Nella , Chulkova Yulia , Garaeva Liliana
Published: September 2002

The roots and leaves of 7‐day seedlings of three winter wheat cultivars differing in frost resistant were used to study changes in lectin activity under cytoskeleton modifiers (DMSO—7%; colchicine—1mm; oryzalin—15μm; cytochalasin B—15μm) of non‐hardened (23°C) and hardened (2–3°C, 3–7 day) plants. Plants were grown with ABA (30μm) or without ABA. Pretreatment with colchicine, oryzalin [inhibitors of microtubules (MT) polymerization], cytochalasin B [inhibitor of microfilament (MF) polymerization] increased the activity of cell wall lectins, although pretreatment with DMSO (stabilizer of microtubules) decreased the activity. Both hardening and ABA decreased the effect of the cytoskeletal modifiers. These results could be explained by the appearance of tolerant MTs with less affinity. It is probable that increase in the activity of cell wall lectins may be the compensatory mechanism which stabilizes the cytoskeleton structure in conditions tending to disrupt it. The genotype with low resistance had higher sensitivity of lectin activity to cytoskeleton modifiers than the frost resistant genotype. The results suggest that leaves have more stable MTs and MFs and stronger MT—MF binding than roots.
NITRIC OXIDE MODIFIES GLYCOLYTIC PATHWAYS IN CULTURED HUMAN SYNOVIOCYTES
Authors: Borderie D. , Le Marechal H. , Ekindjian O. G. , Hernvann A.
Published: September 2002

Nitric oxide (NO) is a free radical produced during inflammation following activation of an inducible NO synthase by pro‐inflammatory cytokines such as IL‐1β. Since both NO and IL‐1β are involved in the physiopathology of inflammatory arthropathies, we investigated the effects of exogenous NO on glycolytic pathways in cultured human osteoarthritic synovial cells. NO generated from S‐nitroso‐N‐acetyl penicillamine (SNAP) or sodium nitroprusside (SNP) inhibited glucose uptake (by 50% after 1h of incubation) and lactate production by 16% (SNAP) and 8.5% (SNP) after 3h. Both NO donors also reduced production of glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH), an enzyme of the glycolytic pathway. This effect was reversed by haemoglobin, a NO scavenger with higher affinity for the radical. In contrast, the effect on glucose uptake appeared to be irreversible.
HGF/SF‐INDUCED SPREADING OF MDCK CELLS CORRELATES WITH DISAPPEARANCE OF BARMOTIN/7H6, A TIGHT JUNCTION‐ASSOCIATED PROTEIN, FROM THE CELL MEMBRANE
Authors: Muto Samu , Sato Yasushi , Umeki Yasuhiro , Yoshida Kenji , Yoshioka Toshiaki , Nishikawa Yuji , Nakamura Toshikazu , Mori Michio , Koyama Kenji , Enomoto Katsuhiko
Published: September 2002

Changes in expression of the two tight junction‐associated proteins, barmotin/7H6 and ZO‐1, as well as the adherence junction‐associated protein, E‐cadherin, were followed during hepatocyte growth factor/scatter factor (HGF/SC)‐induced migration process of MDCK cells. Modulation of the HGF/SF‐induced migration process by staurosporine, an inhibitor of protein kinase C (PKC), was also examined. Cell migration induced by HGF/SF consisted of two distinct phases, initial cell spreading between 2 and 9h after the start of treatment, and the scattering phase which started ∼12h after treatment. Both ZO‐1 and E‐cadherin were expressed at the cell—cell border of adherent cells in the scattering phase, whereas barmotin/7H6, a barrier function‐related tight junction protein, was not seen during the early spreading phase. Confluent cultures of MDCK cells, which did not spread after HGF/SF treatment, were positive for barmotin/7H6 expression at cell—cell borders. Blocking PKC activation during HGF/SF treatment with staurosporine inhibited cell spreading, and the cells retained barmotin/7H6 expression until at 6h after HGF/SF treatment. The results indicate that disappearance of the tight junction protein, barmotin/7H6, is closely associated with cell spreading, with both barmotin/7H6 expression and cell spreading seemingly being regulated by PKC‐mediated signaling.
PRESENCE, UPTAKE AND LOCALIZATION OF AN IMMUNOREACTIVELY INTERLEUKIN 6 (IL‐6)‐LIKE MOLECULE IN TETRAHYMENA PYRIFORMIS
Authors: Kõhidai László , Kovács Péter , Lázár‐Molnár Eszter , Csaba György
Published: September 2002

The unicellular Tetrahymena and its medium contain immunoreactively interleukin 6 (IL‐6)‐like molecules (hereinafter IL‐6) in a measurable quantity in the 24 h‐old cultures. This protozoan takes up exogenously supplied IL‐6 very quickly, and this can be found in similar amounts in both the cells and the media after 1h. After 24h (48h cultures), an equal amount of IL‐6 is present in the control and IL‐6‐treated cells and their media. By 120h, cells which have not had their medium changed retained the same quantity of IL‐6 as the control; however less than half was found in IL‐6‐treated cells. In the medium of 120h‐old cultures, there was a reduction of IL‐6 content relative to the 24h content in the control; however, in the IL‐6‐treated cell culture medium, less than half of the level in the controls was found. Confocal microscopy demonstrated the localization of IL‐6 in/on the oral apparatus and basal bodies, and the nuclear envelope also showed moderate labelling. IL‐6 antibody binding was enhanced after IL‐6 pretreatment (hormonal imprinting). The experiments call attention to the presence of an IL‐6‐like molecule and its uptake at a very low level of phylogeny.
DIFFERENT PHYSIOLOGICAL MECHANISMS CONTROL ISOVOLUMETRIC REGULATION AND REGULATORY VOLUME DECREASE IN CHICK EMBRYO CARDIOMYOCYTES
Authors: Souza Marta M. , Boyle Robert T. , Lieberman Melvyn
Published: September 2002

Cultured chick embryo cardiac myocytes submitted to a 180mOsm/kg hyposmotic solution swell present a regulatory volume decrease (RVD). This RVD is mediated by a Ca2+influx followed by a 40% loss of total taurine content accompanied by the loss of lesser amounts of other osmolytes. Kidney cells respond to a gradual change in osmolality by maintaining their volume at the initial level. This is termed isovolumetric regulation (IVR), which may activate regulatory processes other than those observed with sudden changes in osmolality. When cardiac myocytes were exposed to a gradual change in osmolality, they show a partial IVR which is not dependent upon extracellular Ca2+. Potassium channel blockers, quinidine and Ba2+, and the chloride channel blocker, diphenylamine‐2‐carboxylate (DPC), compromise IVR in our model. Tritiated taurine loss and total intracellular K+contents were analyzed in cultured cardiomyocytes submitted to a gradual change in osmolality. The cultured cells lost approximately 10% of their taurine and 35% of their total K+. These findings suggest that different compensatory mechanisms are activated when cells are exposed to stepwise and gradual changes in osmolality. Inorganic osmolytes (through conductive pathways) are preferentially mobilized during the physiological and/or patho‐physiological IVR situation, perhaps reflecting energetic conservation in response to a less traumatic event for the cardiac myocytes.
2,7‐DICHLOROFLUORESCIN OXIDATION AND REACTIVE OXYGEN SPECIES: WHAT DOES IT MEASURE?
Authors: Jakubowski Witold , Bartosz Grzegorz
Published: September 2002

Intracellular 2,7‐dichlorofluorescin (H2DCF) oxidation is often used to measure the production of reactive oxygen species (ROS) within cells. The rate of H2DCF oxidation depends on the concentration of glutathione, which is an alternative target for ROS. Our results suggest that increased rate of H2DCF oxidation be interpreted as an indication of general oxidative stress due to a variety of reasons, including depletion of antioxidants, rather than as a specific proof of augmented ROS formation.
FINE STRUCTURAL MODIFICATIONS OF LIVER, PANCREAS AND BROWN ADIPOSE TISSUE MITOCHONDRIA FROM HIBERNATING, AROUSING AND EUTHERMIC DORMICE
Authors: Malatesta M. , Battistelli S. , Rocchi M.B.L. , Zancanaro C. , Fakan S. , Gazzanelli G.
Published: September 2002

An ultrastructural and morphometric study was performed on mitochondria of euthermic, hibernating and arousing hazel dormice (Muscardinus avellanarius), in order to investigate possible modifications during the seasonal cycle. Hepatocytes, pancreatic acinar cells and brown adipocytes were considered. Our results demonstrated that: (1) the general morphology of mitochondria of all cell types shows slight modifications during the seasonal cycle; (2) mitochondrial size and inner membrane length significantly increase from euthermia to hibernation and decrease upon arousal in all cell types; (3) mitochondrial matrix granules drastically increase in number during hibernation and decrease upon arousal in hepatocytes and pancreatic acinar cells, whereas they do not change in brown adipocytes. These structural modifications are probably related to the changes in cellular energy needs during the euthermia—hibernation—arousal cycle.
TERMINAL DIFFERENTIATION OF HUMAN PROMYELOCYTIC LEUKAEMIA CELLS, HL‐60, DURING THE G1 PERIOD
Authors: Matsuhisa Tadao
Published: September 2002

Human promyelocytic leukaemic cells, HL‐60, arrested in mitosis by nocodazole were released in the presence of 1α,25‐dihydroxyvitamin D3 and thymidine or hydroxyurea. Cells moved from early G1 period to the G1/S boundary and differentiated. Furthermore, cells arrested at the G1/S boundary by double thymidine block were released, with 1α,25‐dihydroxyvitamin D3 being added at the end of DNA synthesis. Under the latter conditions, differentiated cells developed, indicating that DNA synthesis is not required for cell differentiation.
EARLY MEMBRANE DEPRESSIONS IN HEPATOCYTES CULTURED ON PRIMARIA™ SUPPORTS
Authors: Griffiths B.J. Nathan , Evans Peter J.
Published: September 2002

The topography of spreading hepatocytes on positively charged Primaria™ plates was examined using scanning electron microscopy. The cells acquired a uniform morphology within 2h. The spreading was rapid and the surface of the cells showed early prominent depressions or dips. The hepatocytes had either one or two of these structures which corresponded to the frequency of mononuclear and binuclear cells, respectively. The nuclear origin of the dips was strengthened after 6h. They contained solid structures, whose number, size and shape were the same as nucleoli. The membrane dipping was independent of cell density and took place under conditions where phenotypic changes can occur. Kidney proximal tubule cells had no dips. Co‐cultures of hepatocytes and kidney proximal tubule cells showed that the cell types behave differently.