bluuebunny/crossref_metadata_2025_split huggingface dataset:
Distribution of excitatory junction potential amplitude in cat cerebral arteries: Examination of its quantal nature and modulation by opiates
Authors: Madden Jane A. , Kalbfleisch John H. , Harder David R. 
Published: February 2005

AbstractWe used scorpion venom to release small amounts of an excitatory neurotransmitter from adventitial nerves in cat left anterior descending cerebral artery. We used glass microelectrodes to measure and record postsynaptic electrical events of minimal amplitude. These events were similar to postsynaptic spontaneous and electrically evoked excitatory junction potentials (ejp's) seen in skeletal muscle. We performed a frequency analysis of the ejp amplitudes to determine if they fit a unimodal or multimodal distribution. We also investigated the effects of phentolamine, norepinephrine, hydromorphone, and morphine on ejp amplitude and frequency in the artery. Statistical analysis of the ejp frequency and amplitude revealed a multipeaked distribution with decreasing peaks. These results were similar to the distribution reported for acetylcholine release in skeletal muscle. The ejps were inhibited by phentolamine, which suggested that these events were adrenergically mediated. Norepinephrine and the opiates, hydromorphone and morphine, reduced the frequency and amplitude of the ejp's. The vessels also constricted to increasing doses of norepinephrine both under control conditions and in the presence of opiate. These results suggest that norepinephrine blocks the ejp's by a feedback mechanism at the presynaptic membrane and that endorphins and/or enkephalins, also acting at this presynaptic site, may modulate neurotransmission in the cerebral circulation.
Induction of heat shock proteins in Chinese hamster ovary cells and development of thermotolerance by intermediate concentrations of puromycin
Authors: Lee Yong J. , Dewey William C. 
Published: February 2005

AbstractDuring 4 hr after puromycin (PUR: 20 μg/ml) treatment, the synthesis of three major heat shock protein families (HSPs: Mr = 110,000, 87,000, and 70,000) was enhanced 1.5‐fold relative to that of untreated cells, as studied by one‐dimensional gel electrophoresis. The increase of unique HSPs, if studied with two dimensional gels, would probably be much greater. In parallel, thermotolerance was observed at 10−3 isosurvival as a thermotolerance ratio (TTR) of either 2 or greater than 5 after heating at either 45.5°C or 43°C, respectively. However, thermotolerance was induced by only intermediate concentrations (3–30 μg/ml) of puromycin that inhibited protein synthesis by 15–80%; a high concentration of PUR (100 μg/ml) that inhibited protein synthesis by 95% did not induce either HSPs or thermotolerance. Also, thermotolerance was never induced by any concentration (0.01–10 μg/ml) of cycloheximide that inhibited protein synthesis by 5–94%. Furthermore, after PUR (20 μg/ml) treatment, the addition of cycloheximide (CHM: 10 μg/ml), at a concentration that reduces protein synthesis by 94%, inhibited both thermotolerance and synthesis of HSP families. Thus, thermotolerance induced by intermediate concentrations of PUR correlated with an increase in newly synthesized HSP families.This thermotolerance phenomenon was compared with another phenomenon termed heat resistance and observed when cells were heated at 43°C in the presence of CHM or PUR immediately after a 2‐hr pretreatment with CHM or PUR. Heat protection increased with inhibition of synthesis of both total protein and HSP families. Moreover, this heat protection decayed rapidly as the interval between pretreatment and heating increased to 1–2 hr, and did not have any obvious relationship to the synthesis of HSP families. Therefore, there are two distinctly different pathways for developing thermal resistance. The first is thermotolerance after intermediate concentrations of PUR treatment, and it requires incubation after treatment and apparently the synthesis of HSP families. The second is resistance to heat after CHM or PUR treatment immediately before and during heating at 43°C, and it apparently does not require synthesis of HSP families. This second pathway not requiring the synthesis of HSP families also was observed by the increase in thermotolerance at 45.5°C caused by heating at 43°C after cells were incubated for 2–4 hr following pretreatment with an intermediate concentration of PUR.
Cell cycle parameters of adult rat hepatocytes in a defined medium. A note on the timing of nucleolar DNA replication
Authors: Vintermyr Olav K. , Døskeland Stein Ove 
Published: February 2005

AbstractHepatocytes, isolated from adult (250–350 g) rats, attached and survived well in primary culture on highly diluted (< 1 μg/cm2) collagen gel in a synthetic medium without serum or hormones. About 20% of the cells “spontaneously” entered S phase during the first 4 days of culturing, and mitoses were easily demonstrated at the near physiological concentration (1.25 mM) of Ca+ + prevailing in the medium. Cultures given 9 nM epidermal growth factor (EGF) and 20 nM insulin 20 h after inoculation showed vigorous DNA synthesis and mitotic activity. Autoradiography of such cells exposed to [3H]thymidine allowed the determination of the following cell cycle parameters: Lag period from EGF/insulin stimulation till onset of increased DNA synthesis, 17 h; rate of entry into S phase (kG1/S), 0.028/h; duration of S phase, 8.4 h; duration of G2 phase, 2.7 h. The peak DNA synthesis (pulse labelling index, 24%) and peak mitotic activity (mitotic index, 1.7%) occurred 35 and 43 h, respectively, after the stimulation with EGF/insulin. These values are comparable to those reported during the in vivo compensatory hyperplasia following partial hepatectomy of adult rats. A marked variation of the intranuclear [3H]thymidine pulse labelling pattern was noted: During the first 1.5 h of the S phase, the labelling was extranucleolar and during the last 1.5 h chiefly nucleolar.The cells survived well in the absence of glucocorticoid, whose effect on cell cycle parameters therefore could be studied. Dexamethasone (25–250 nM) did not appreciably affect the durations of S phase and G2 phase or the pattern of preferential extranucleolar and nucleolar DNA synthesis within the S phase.
Possible involvement of reorganization of actin filaments, induced by tumor‐promoting phorbol esters, in changes in colony shape and enhancement of proliferation of cultured epithelial cells
Authors: Sastrodihardjo Sumadhi , Sasaki Yasuto , Shiba Yoshiki , Kanno Yoshinobu 
Published: February 2005

AbstractTumor promoters are known to induce reorganization of actin, morphological changes and enhancement of proliferation of epidermal cells in vivo. In this study, we have examined the effects of tumor promoters on these events to clarify the role played by the organization of actin filaments in the regulation of the shape and growth of colonies of epithelial cells in culture. Treatment with 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) caused a change in the shape of colonies of FL and Madin‐Darby canine kidney (MDCK) cells within 6 hr. Changes in the shape of colonies were consistent with the morphological change of individual cells and the dissociation of groups of cells in the colonies. Addition of TPA also caused reorganization of actin filaments after 2 hr, and it caused enhancement of proliferation of FL and MDCK cells after 48 hr but did not cause any such changes in KB cells. However, the binding affinities of 4β‐phorbol 12,13‐dibutyrate (PDBu) to FL and MDCK cells were similar to that of PDBu to KB cells. Related tumor promoters such as phorbol 12,13 didecanoate (PDD) and mezerein caused effects similar to those caused by TPA. In contrast, nontumor promoting phorbol esters, such as 4α‐PDD and phorbol, had no effect. Cyclic AMP blocked the TPA‐induced changes in FL and MDCK cells. These results suggest that TPA‐induced reorganization of actin filaments which can be inhibited by cyclic AMP results in changes in the shape of colonies and enhancement of proliferation.
Use of strontium to separate calcium‐dependent pathways for proliferation and differentiation in human keratinocytes
Authors: Praeger Frank C. , Stanulis‐Praeger Betzabe M. , Gilchrest Barbara A. 
Published: February 2005

AbstractThe role of extracellular calcium (Caex) in modulating keratinocyte differentiation has been well documented, but its role in proliferation has been harder to define due to the confounding effect of terminal differentiation. Because strontium (Sr) does not induce terminal differentiation in murine keratinocytes but does mimic the stimulatory effect of Caex on DNA synthesis in chick fibroblasts, experiments were undertaken to determine if Sr could be used to separate the presumably presumably opposing effects of Caex on the proliferation and differentiation of cultured human keratinocytes. In response to additions of SrCl2, keratinocytes in a serum‐free hormone‐supplemented basal medium containing 0.03 mM Ca showed a dose‐dependent increase in day 7 cell yields. Cell yield in the optimal concentration of SrCl2 (1.8 mM) was approximately twice that obtained in any concentration of CaCl2. Maximally stimulatory additions of CaCl2 varied from 0.05 to 1.8 mM, but 0.03 and 0.05 mM additional CaCl2 always increased cell yield relative to unsupplemented controls. Keratinocytes grown in low levels of CaCl2 or any level of SrCl2 have minimal contact with each other regardless of cell density in contrast to the colonies of tightly apposed and stratified cells grown in 1.8 mM CaCl2. Transmission electron micrographs of vertically sectioned confluent cultures in low or high levels of SrCl2 or low levels of CaCl2 revealed abundant ribosomes and keratin filaments but no stratification or desmosomes, while cultures in 1.8 mM CaCl2 were stratified with numerous desmosomes. These results suggest that Caex may separately stimulate keratinocyte proliferation and terminal differentiation and that Srex can substitute for Caex in the former but not the latter process.
α2HS glycoprotein is chemotactic for mononuclear phagocytes
Authors: Malone J. David , Richards Michael 
Published: February 2005

Abstractα2HS glycoprotein is a normal constituent of plasma. It has a high affinity for hydroxyapatite and is concentrated in bone 100‐fold greater than albumin, however its biological function in bone is not known. We have purified α2HS from human plasma and examined it for evidence of chemotactic activity against human peripheral mononuclear cells, polymorphonuclear leucocytes and lymphocytes in Boyden chamber assays. We observed that the protein was chemotactic for mononuclear cells and that maximal cell migration occured when its concentration in the lower compartment of Boyden chambers was 10−10 M. Chemotaxis did not occur in the absence of a concentration gradient. In addition, cell migration was blocked when the cells were preincubated with 10−10 M α2HS or when the protein was preincubated with rabbit anti‐human α2HS glycoprotein lgG. Neither polymorphonuclear leucocytes, which are responsive to a wide range of chemoattractants, or peripheral lymphocytes exhibited directed migration to α2HS in Boyden chamber assays. The glycoprotein appears therefore to act as a chemotaxin directed to monocyte recruitment, and we speculate that the protein may have an important role in monocyte recruitment to bone and possibly their subsequent fusion and differentiation into functional osteoclasts.
Proliferative potential of human fibroblasts: An inverse dependence on cell size
Authors: Angello John C. , Pendergrass William R. , Norwood Thomas H. , Prothero John 
Published: February 2005

AbstractHuman foreskin fibroblast‐like cells were separated on the basis of DNA content and cell size by fluorescence‐activated cell sorting. Subpopulations of “large” or “small” cells with the same (G1) DNA content were clonally expanded and found to contain predominantly nondividing or highly proliferative cells, respectively. From the rate of clonal growth, we deduce that small cells divide faster than large cells. Intermediate‐sized cells were found to yield primarily smaller (“attenuated”) clones. The clonal data can be incorporated into a previously reported kinetic model of clonal attenuation. This version of the model postulates that small “stem” cells yield larger daughters which have only a limited proliferative potential. We also postulate that a progressive increase in cell size can account for the decreasing concentration of DNA polymerase alpha, which has been reported in older cultures.
Relation of prostaglandin‐induced increases in cellular cAMP to stimulation of pepsinogen secretion from dispersed chief cells
Authors: Raufman Jean‐Pierre , Cosowsky Laurey 
Published: February 2005

AbstractThe actions of prostaglandins (PG) on cAMP in dispersed chief cells from guinea pig stomach were examined and compared to the actions of these agents on pepsinogen secretion. Maximal concentrations of A, B, or E prostaglandins caused a 2–5‐fold increase in pepsinogen secretion and cellular cAMP. The relative order of potency for these actions was PGEs > PGAs > PGBs. Detection of prostaglandin‐induced changes in cAMP was enhanced by adding a phosphodiesterase inhibitor to the incubation solution. The time courses for the effects of prostaglandins on pepsinogen secretion and cAMP were similar. With PGE1 an increase in cAMP and pepsinogen secretion was detected by 1 min and was maximal by 7.5 min. Although significant increases in cAMP were detected with a ten‐fold lower concentration of PGEs than PGAs, a maximal increase in cAMP was observed with the same concentration, 30 μM, of either agent. These data indicate that prostaglandins that stimulate pepsinogen secretion increase cAMP in dispersed chief cells. However, comparison of the dose‐response curves for the actions of prostaglandins on pepsinogen secretion and cAMP revealed that detectable increases in cAMP occurred with concentrations of these agents that were about tenfold greater than those needed to stimulate pepsinogen secretion. Therefore, although the similarity in the kinetics and relative potencies of prostaglandininduced changes in cAMP and enzyme secretion provides further evidence that changes in cAMP play a role in the mediation of prostaglandin‐induced pepsinogen secretion, the present data suggest the involvement of a cellular messenger in addition to cAMP.
Bicarbonate/chloride antiport in vero cells: I. Evidence for both sodium‐linked and sodium‐independent exchange
Authors: Tønnessen Tor Inge , Ludt Jannikke , Sandvig Kirsten , Olsnes Sjur 
Published: February 2005

AbstractThe effect of bicarbonate on the ability of cells to regulate the internal pH after acid and alkali loads was studied. In the presence of Na+, the normalization of the internal pH after acid loads occurred more rapidly in the presence than in the absence of bicarbonate. DIDS (4,4′‐diisothiocyano‐2,2′‐stilbene‐disulfonic acid) strongly inhibited the pH increase, whereas amiloride inhibited it to a lesser extent. The Na+‐linked, bicarbonate‐dependent pHi increase after an acid load was strongly reduced in cells depleted of Cl−. When cells were transferred to gluconate or mannitol balanced buffers containing bicarbonate, there was a rapid alkalinization of the cytosol, apparently due to influx of bicarbonate induced by chloride efflux. When the internal pH was below 7.0, the pH increase was much more rapid in the presence than in the absence of Na+, whereas at higher internal pH, there was no measurable effect of Na+. The ability of the cells to reduce the internal pH after an alkali load was increased in the presence of bicarbonate. The data indicate that both Na+‐linked and Na+‐independent bicarbonate/chloride exchange occur in Vero cells.
Endothelial cells in culture produce a vasoconstrictor substance
Authors: O'Brien Richard F. , Robbins Richard J. , McMurtry Ivan F. 
Published: February 2005

AbstractWe report that cultured vascular endothelial cells release into the culture medium a vasoconstrictor peptide, a substance we call an endotheliumderived constricting factor (EDCF). Conditioned medium from cultured bovine aortic and pulmonary artery endothelial cells caused sustained, dose‐dependent isometric constriction of vascular rings isolated from bovine coronary and pulmonary arteries and rat and guinea pig pulmonary arteries and aortas. The medium also caused vasoconstriction when infused into isolated, perfused rabbit hearts and rat kidneys. Conditioned medium from bovine aortic intimal explants also contained constrictor activity, whereas medium from denuded intimal explants, cultured microvascular endothelial cells, vascular smooth muscle cells, or lung fibroblasts did not. Constrictor activity increased progressively in the culture medium over 2–12 h of incubation. Thrombin stimulated the release of constrictor activity; hypoxia, anoxia and meclofenamate had no effect and the calcium ionophore A23187 inhibited EDCF release. The EDCF caused a characteristic slow‐onset and sustained constriction of the vascular rings that relaxed slowly over 60–90 min following removal. The constriction was not affected by inhibitors of arachidonic acid metabolism or by antagonists of serotonergic, histaminergic, aloha‐adrenergic, opioid, leukotriene, angiotensin II, or substance P receptors; constriction was reversed partly by verapamil and acetylcholine and completely by nitroprusside and isoproterenol. EDCF was heat stable, not extractable into organic solvents, and completely destroyed by trypsin and neutral protease. Cycloheximide blocked the production of EDCF. These properties and the results of polyacrylamide gel filtration experiments suggested that EDCF was a peptide with a molecular weight of 3,000 daltons. These findings show that endothelial cells in culture produce a vasoconstrictor substance and support the idea that endothelial cell products play a role in mediating vascular tone.
Evidence for two functionally different fibrinogen receptors on hemopoietic cells: The glycoprotein IIb–IIIa and the mitogenic fibrinogen receptor
Authors: Levesque Jean‐Pierre , Hatzfeld Antoinette , Hudry‐Clergeon Gilbert , Wilner George D. , Hatzfeld Jacques 
Published: February 2005

AbstractWe have previously established that the mitogenic effect of fibrinogen on hemopoietic cell lines Raji and JM is mediated via a specific receptor (Levesque, J.‐P. et al.: Proc. Natl. Acad. Sci. USA 83:6494–6498, 1986). In this study, we have further characterized the fibrinogen domain involved in the binding to the mitogenic receptor. This binding was not inhibited either by a monoclonal antibody against the C‐terminal sequence of the fibrinogen γchains or by synthetic peptides containing the Arg‐Gly‐Asp sequence. Such inhibition is specific of the platelet fibrinogen receptor, the glycoprotein IIb‐IIIa complex. Fragments containing the fibrinogen D domain were the only plasmin degradation products of fibrinogen which mitogenic. These fragments acted via direct binding on the mitogenic receptor with a Kd of 2.24 × 10−6 M. This value was similar to the K1 value of unlabeled fragments D (2.47 × 10−6 M). Our results suggest the presence of two different functional types of fibrinogen receptors: the glycoprotein IIb–IIIa receptor responsible both for platelet aggregation and leukocyte adhesion and killing, and the mitogenic receptor involved in proliferation control of hemopoietic cells.
Both protein kinase C and calcium mediate activation of the Na+/H+ antiporter in Chinese hamster embryo fibroblasts
Authors: Ober Sharon S. , Pardee Arthur B. 
Published: February 2005

AbstractChinese hamster embryo fibroblast cells (CHEF/18) possess a plasma membrane‐associated, amiloride‐sensitive Na+/H+ antiporter that affects intracellular pH (pHi) and is activated by growth factor addition. Our results using 14C‐benzoic acid distribution indicate that both epidermal growth factor (EGF) and thrombin are capable of causing rapid rises in the pHi of CHEF/18 cells. The maximal shift induced by these factors is 0.20 to 0.25 pH units above the basal unstimulated level. Distinctive differences were observed between the modes of action of these two growth factors. Sequential additions revealed that the rise in pHi due to EGF was additive with that caused by diacylglycerols (DAG), while that of thrombin was not. Furthermore, exposure of cells to the phorbol ester PMA for a prolonged period of time in order to down‐regulate protein kinase C (pkC), or treatment with the pkC inhibitor H‐7, abolished the pHi response to thrombin but not to EGF. In contrast, incubation of cells in nominally calcium‐free medium or with the calmodulin antagonists W‐7 or trifluoperazine (TFP) decreased only the ability of EGF to cause changes in pHi. These data suggest that there are two distinct mechanisms for activation of the Na+/H+ antiporter in CHEF/18 fibroblast cells and thus provide an example of the use of alternative modes for the modulation of intracellular processes.
Disparity between expression of transferrin receptor ligand binding and non‐ligand binding domains on human lymphocytes
Authors: Boldt D. H. , Phillips J. L. , Alcantara O. 
Published: February 2005

AbstractWe compared transferrin receptor (TfR) expression on human peripheral blood lymphocytes (PBL) activated by phorbol myristate acetate (PMA) or L‐phytohemagglutinin (LPHA) using two techniques: (1) 125I‐iron‐saturated transferrin (FeTf) binding, (2) reactivity with monoclonal anti‐TfR antibodies—OKT9 and B3/25. These monoclonal antibodies do not block FeTf binding, and therefore bind to TfR domains separate from the ligand binding site. Unstimulated PBL bound fewer than 1,000 molecules of 125I‐FeTf per cell, and < 5% of cells expressed TfR antigens detected by OKT9 or B3/25. 125I‐FeTf binding and antibody binding increased in parallel on LPHA‐activated PBL. After exposure to LPHA for 72 hr, 125I‐FeTf binding increased 100‐fold to 105 molecules per cell and > 50% of cells expressed TfR antigens. By contrast, PMA activation of PBI markedly increased binding of OKT9 and B3/25 but not the binding of 125I‐FeTf. Cell surface expression of TfR antigens seen by OKT9 and B3/25 did not differ between LPHA‐ and PMA‐activated PBL. However, after 72 hr with PMA, 125I‐FeTf binding increased only 6‐fold and consistently remained at < 104 molecules per cell. Therefore, PMA induced a disparity between expression of TfR ligand binding domains and immunological domains at the cell surface. Cell proliferation assessed by fluorescent DNA analysis was similar in cultures stimulated by LPHA or PMA. These data indicate that lymphoid cells may possess a mechanism for modulating TfR expression in which down‐regulation of FeTf binding occurs without receptor internalization. Alternatively, it is possible that this observation may reflect a membrane perturbation effect of PMA.
Diminished calcium influx in lectin‐stimulated T cells from old mice
Authors: Miller Richard A. , Jacobson Bruce , Weil Gary , Simons Elizabeth R. 
Published: February 2005

AbstractT lymphocytes from aged donors function poorly, but the biochemical basis for the defect remains uncertain. We tested the hypothesis that T cells from old mice had a diminished ability to transmit extracellular signals into the cytoplasm, by measuring intracellular free calcium concentrations (Cai) in T cells stimulated by the polyclonal activator concanavalin A (Con A). Using the second‐generation fluorochrome indo‐1 as a reporter of Cai, we found that the Con A‐induced elevation of Cai levels is reduced both in rate and extent in old T cells, as compared to T cells from young mice. Flow cytometric analysis showed that this age‐sensitive change represents a decline, with age, in the number of T cells that can respond to Con A by increasing their Cai above resting baseline levels (100–120 nM). These results thus show that defects in activation are manifested by T cells from old donors within the first 5 minutes of the activation process, and suggest that aging may lead to alterations either in the surface molecules that receive extracellular signals, or in the sequence of coupled events by which these extracellular signals bring about alterations in the intracellular ionic milieu.