bluuebunny/crossref_metadata_2025_split huggingface dataset:
Surface contact modulation of inflammatory macrophage antibody dependent cytotoxicity and prostanoid release
Authors: Gudewicz Paul W. , Frewin Mary Beth 
Published: February 2005

AbstractAdherence to extracellular matrix proteins modulates the functional and secretory activities of mononuclear phagocytes, although the mechanisms regulating these adherence‐dependent changes are poorly understood. In this study, the ability of rat inflammatory peritoneal macrophages (PM) to adhere to an endothelial cell‐derived extracellular matrix or a denatured collagen/fibronectin‐coated surface and perform antibody dependent cell cytotoxicity (ADCC) and secrete reactive oxygen intermediates was compared with PM adherent to tissue culture plastic. Prostaglandin E2 (PGE2) and thromboxane B2 (TxB2), two major cyclooxygenase products released by inflammatory macrophages, were also measured by PM adherent to the protein coated surfaces. Rat exudate PM were equally adherent to tissue culture plastic or wells coated with either endothelial cell derived matrix or denatured collagen (gelatin)/fibronectin. PM adherent to a denatured collagen/fibronectin‐coated wells demonstrated significantly less cytolytic activity (15 ± 2% lysis) when compared with either tissue culture plastic adherent PM (43 ± 7% lvsis) or PM adherent to extracellular matrix (59 ± 11% lysis). PM adherent to extracellular matrix released twofold more TxB2 than plastic adherent PM, while PM adherent to denatured collagen/fibronectin released 40% more PGE2 than cells adherent to tissue culture plastic or 80% more PGE2 than PM adherent to the extracellular matrix. PM adherent to denatured collagen/fibronectin release less superoxide anion (27 ± .9 nmoles/106 PM) than PM adherent to either tissue culture plastic (43 ± 1 nmoles/106 PM) or the extracellular matrix (60 ± 0.5 nmoles/106 PM). Furthermore, incubation of plastic adherent PM with exogenous PGE2 reduced superoxide production in a dosedependent manner. These results demonstrate that the inhibition of ADCC and secretion of reactive oxygen intermediates by PM adherent to a denatured collagen/fibronectin surface correlated with an increased release of the immunosuppressive prostanoid PGE2. Furthermore, the addition of exogenous PGE2 to plastic adherent PM reproduced the depression in ADCC and superoxide anion production observed by PM adherent to a denatured collagen/fibronectin surface. These studies suggest that the increased production and release of PGE2 by inflammatory macrophages adherent to a denatured collagen surface may act to suppress cytotoxic mechanisms and thereby constitutes part of an autocrine feedback mechanism regulating macrophage function during wound injury.
Insulin‐like growth factor 1, insulin, and progesterone induce early and late increases in Xenopus oocyte sn‐1, 2‐diacylglycerol levels before meiotic cell division
Authors: Stith Bradley J. , Kirkwood Allan J. , Wohnlich Erica 
Published: February 2005

AbstractAfter a 3 to 6 hour incubation, addition of progesterone (the most effective), insulin‐like growth factor 1 (IGF‐1; the second most effective), or insulin induces meiotic cell division in Xenopus oocytes. Measurement of an endogenous activator of protein kinase C, sn‐1,2‐diacylglycerol (DAG), by an enzymatic method recording mass demonstrates that all three hormones alter DAG levels. Five seconds after addition, only progesterone transiently reduces DAG levels by about 25%. At 15 minutes after addition, all three hormones produce a peak of DAG (115% to 160% of control values), with the more effective hormones producing a larger increase in DAG. Insulin produces the smallest DAG increase, but the DAG release is longer lasting. Finally, all three hormones induce a second peak in DAG levels just before white spot appearance (at 0.85 GVBD50, where 1.0 GVBD50 is when 50% of the cells have divided). With these data and since an activator of protein kinase C, a phorbol ester, has been found to induce meiosis, the kinase may play a role in early proliferative events at the plasma membrane and in late events at the nucleus.
Clustering of erythrocytes by fibrinogen is inhibited by carnitine: Evidence that sulfhydryl groups on red blood cell membranes are involved in carnitine actions
Authors: Fritz Irving B. , Wong Katherine , Burdzy Krystyna 
Published: February 2005

AbstractCarnitine is bound by intact red blood cells, by red blood cell ghosts, and by glutaraldehyde‐fixed human erythrocytes in a non‐saturable, temperature‐dependent manner. Binding of carnitine by these preparations is blocked by sulfhydryl reagents. Incubation or preincubation of red blood cell preparations with carnitine inhibits the aggregation of erythrocytes otherwise elicited by fibrinogen. Identical effects are obtained with red blood cell ghosts. In contrast, choline, even at high concentrations, is inactive in preventing the aggregation of erythrocytes. We discuss possible mechanisms by which carnitine favors the dispersion of red blood cells, and we present data indicating that sulfhydryl groups on erythrocyte membranes are required to permit these carnitine actions to be manifested.
Macrophages express functional receptors for calcitonin‐gene‐related peptide
Authors: Vignery Agnès , Wang Feng , Ganz Michael B. 
Published: February 2005

AbstractThe present study was designed to investigate whether non‐activated macrophages express calcitonin (CT) or calcitonin‐gene‐related peptide (CGRP) receptors. To this end, we first analyzed whether CT and CGRP induce a cAMP accumulation in macrophages. Macrophages were treated for 2 min with increasing concentrations of either CT or CGRP in the presence or absence of IBMX. A dose‐dependent cAMP accumulation was measured in response to CGRP with a half‐maximal effect attained with 1 nM CGRP. CT failed at all doses to induce an accumulation of cAMP. The effects of CT and CGRP on the activation of the Na‐H exchanger were next assessed by spectrofluorometry by using the pH‐sensitive dye 2,7 biscarboxyethyl‐5(6)‐carboxyfluorescein (BCECF). Steadystate pHi of macrophages in a 7.4, HCO3‐free solution (HEPES‐buffered) was 7.04 ± 0.08 (n = 22). pHi recovery following an NH4+/NH3 acid load was inhibited by the removal of Na+ or by the addition of the amiloride analog EIPA; therefore recovery is dependent on Na‐H exchange activity. CT had no effect on steady‐state pHi but CGRP increased pHi in a dose‐dependent fashion (10−12 to 10−6M). The pHi change induced by CGRP was due to the stimulation of the Na‐H exchanger as CGRP enhanced the rate of recovery (dpHi/dt) from an acid load from 45.3 to 77.2 μMs−1 (n = 8, P < 0.002) and was completely blocked by EIPA. These data indicate that CGRP both enhances the activity of the Na‐H exchanger and increases intracellular cAMP, thus demonstrating that macrophages express functional CGRP receptors.
Calcium transport by sarcoplasmic reticulum of vascular smooth muscle: I. MgATP‐dependent and MgATP‐independent calcium uptake
Authors: Stout Marguerit A. 
Published: February 2005

AbstractThe components of 45calcium (Ca) uptake were studied in saponin skinned rat caudal artery. The steady‐state Ca content increased when the free Ca concentration was varied from 10−8 to 10−4 M but was reduced by azide when the free Ca concentration exceeded 3.1 μM. The azide sensitivity and low affinity for Ca were consistent with functional mitochondria. The azide‐insensitive component consisted of a small bound and a larger releasable Ca fraction. After skinning in Triton X‐100, approximately 4 μmol Ca/kg wet tissue remained, which represented a tightly bound but slowly exchangeable Ca pool. The Ca content was independent of the free Ca concentration and MgATP, and it was not released with A‐23187 or Ca. The Ca content of the larger fraction was a higher order function of the free Ca concentration and was released with A‐23187, indicating it resided within a membrane‐bounded structure. Ca uptake by the releasable fraction was increased by oxalate, MgATP, phosphocreatine, temperature, phosphate, and ruthenium red and represents Ca sequestered by the sarcoplasmic reticulum (SR) with little contribution from other Ca binding or storage sites. It is described by the coefficients Umax = 96.94 μmol/kg wet tissue, K1/2 = 0.75 μM, and Hill coefficient = 1.70. The SR in this preparation regulates cytosolic Ca concentrations under physiological conditions and can accumulate Ca by MgATP‐dependent and MgATP‐independent processes. The larger, MgATP‐dependent Cauptake is described by the coefficients Umax = 72.87 μmol/kg wet tissue, K1/2 = 0.8 μM, and Hill coefficient = 2.09 and is consistent with Ca sequestered by the Ca‐transport ATPase of smooth muscle SR. The smaller, MgATP‐independent uptake is described by the coefficients Umax = 24.14 μmol/kg wet tissue, K1/2 = 0.56 μM, and Hill coefficient = 1.01 and represents Ca sequestered by an unidentified mechanism or by a subpopulation of SR.
Stimulation of large proteoglycan synthesis in cultured smooth muscle cells from pig aorta by endothelial cell‐conditioned medium
Authors: Berro Eliane , Breton Monique , Deudon Elisabeth , Picard Jacques 
Published: February 2005

AbstractWe have previously shown (Berrou et al., J. Cell. Phys., 137:430–438, 1988) that porcine endothelial cell‐conditioned medium (ECCM) stimulates proteoglycan synthesis by smooth muscle cells from pig aorta. ECCM stimulation requires protein cores for glycosaminoglycan chain initiation and is accompanied by an increase in the hydrodynamic size of proteoglycans secreted into the medium. This work investigates the mechanisms involved in the ECCM effect. (1) Control and ECCM stimulated proteoglycan synthesis (measured by a 20 min [35S]‐sulfate labeling assay) was not inhibited by cycloheximide, indicating that the proteoglycans were composed of preexisting protein cores and that ECCM stimulates glycosylation of these protein cores. (2) Whereas ECCM stimulation of [35S]‐methionine incorporation into secreted proteins only occurred after a 6 h incubation, the increase in [35S] methionine‐labeled proteoglycans was observed after 1 h, and the increase was stable for at least 16 h. (3) As analysed by electrophoresis in SDS, chondroitinase digestion generated from [14C] serine‐labeled proteoglycans 7 protein cores of high apparent molecular mass (550–200 kDa) and one of 47 kDa. The two protein cores of highest apparent molecular masses (550 and 460 kDa), but not the 47 kDa protein cores, showed increased [14C]‐serine incorporation in response to ECCM (51%, as measured by Sepharose CL‐6B chromatography). (4) Finally, incorporation of [35S]‐sulfate into chondroitinase‐generated glycosaminoglycan linkage stubs on protein cores was determined by Sepharose CL‐6B chromatography: ECCM did not modify the ratio [35S]/[14C] in stimulated protein cores, indicating that ECCM did not affect the number of glycosaminoglycan chains. The results of these studies reveal that (1) endothelial cells secrete factor(s) that preferentially stimulate synthesis of the largest smooth muscle cell proteoglycans without structural modifications and (2) the stimulation proceeds via increased glycosylation of protein core through enhancement of xylosylated protein core, followed by enhanced protein synthesis.
Gene activation mediated by protein kinase C in human macrophage and teratocarcinoma cells expressing aminoglycoside phosphotransferase activity
Authors: Maio Joseph J. , Brown Fred L. 
Published: February 2005

AbstractThe bacterial neomycin phosphotransferase gene driven by the Moloney mouse leukemia virus long terminal repeat (LTR) or SV40 early region promoter was introduced into the human promonocyte‐macrophage cell line, U937, and into the pluripotential human embryonic teratocarcinoma cell line, NT2/D1. Clonally derived cell lines capable of growing in 2–4 mg/ml of the aminoglycoside antibiotic, G418 (Geneticin), were established and transfected with pHIVCat, a plasmid expressing the bacterial chloramphenicol acetyl transferase (CAT) activity under the control of the human immunodeficiency virus (HIV‐1) LTR. All of the G418 resistant (neor) U937 cell lines and 10 of 14 neor NT2/D1 cell lines exhibited reduced basal levels of CAT expression or impaired responses to activation of the HIV‐1 LTR by phorbol 12‐myristate 13‐acetate (PMA) when compared to the parental lines. Other differences included inhibition of tat activation of the HIV‐1 LTR and increased sensitivity of U937 cells to human tumor necrosis factor α. The expression of other eukaryotic promoters including the HTLV‐1 LTR, SV40 ori sequences, and the human β‐actin gene promoter was similarly affected. However, differentiation of the neor U937 cells into macroph‐ages was neither delayed nor impaired. Because PMA is an activator of protein kinase C (PKC) and a potent inducer of HIV‐1 direcred gene expression, the amounts, sensitivity to G418, and cytosol to membrane translocation of this enzyme were determined in the wild type and neor U937 cells. G418 at concentrations too low to affect cell growth (12–150 μg/ml) inhibited PMA‐induced transactivation responses in wild type cells but did not inhibit PKC‐dependent protein phosphorylation in vitro. PKC activities in the wild type and neor cells were similar in absolute amounts and in the cytosol‐membrane distribution of the enzyme. In contrast with wild type cells, however, all of the cytosolic Ca2+‐phospholipid‐dependent form of PKC disappeared from the neor cells within 30 min after PMA induction. The results suggested that, Depending upon the cell type, gene cotransfer using aminoglycoside resistance as a selectable marker may seriously perturb important cellular control mechanisms such as the PKC pathway leading to activation of gene expression.
Cell cycle dependent alterations of chromatin structure in situ as revealed by the accessibility of the nuclear protein AF‐2 to monoclonal antibodies
Authors: Pfeffer U. , di Vinci A. , Geido E. , Vidali G. , Giaretti W. 
Published: February 2005

AbstractWe have recently described a novel nuclear antigen, AF‐2, which is related to cell cycle dependent alterations of chromatin structure. We show by two parameter flow cytometry on a cell by cell basis that the antigen is accessible to specific monoclonal antibodies only in mitotic and postmitotic early G1‐phase cells. The evaluation of nuclease susceptibility and AF‐2 antigen accessibility reveals different subcompartments of the G1 ‐phase of the cell cycle with distinct chromatin conformations.Digestion with DNase I seems to alter the chromatin structure according to concentration and this is reflected by an increase of the antigen accessibility. Chromatin in the more condensed early G1‐phase is specifically digested by lower concentrations of the enzyme than chromatin in later stages of interphase. Chromatin from cells in the late‐G1, S‐, and G2‐phases shows a higher relative resistance to DNase I and a reduced accessibility of the AF‐2 antigen to monoclonal antibodies. Nuclease S1 has a similar effect on chromatin topology, as revealed by the reaction with anti‐AF‐2 antibodies, without digestion of detectable amounts of DNA. The antigen becomes available to the antibodies in almost all cells by digestion with high concentrations of DNase I or Nuclease S1.
Karyotypic and phenotypic changes during in vitro aging of human endothelial cells
Authors: Johnson Timothy E. , Umbenhauer Diane R. , Hill Rosina , Bradt Carole , Mueller Stephen N. , Levine Elliot M. , Nichols Warren W. 
Published: February 2005

AbstractKaryotypic and phenotypic changes were found in human adult endothelial cells (EC) during aging in vitro. A trisomy of chromosome 11 was found in 11 out of 12 EC cultures examined, derived from 9 cell lines from 8 donors. The incidence of this trisomy in some cell lines increased over time from 0% to as much as 100% near the end of their in vitro life span. A number of oncogenes and other important genes are on chromosome 11. These genes might play a role in the changes observed. An increase in the percentage of polyploid cells was also found near the end of the in vitro life span in 6 lines. The cellular levels of two gene products characteristic of the EC, von Willebrand factor (vWF) or Factor VIII, and angiotensin converting enzyme (ACE) were also monitored. vWf was studied in 2 lines and was decreased in both with serial passage. ACE decreased in three out of the four lines examined. These chromosomal and phenotypic changes which occur with increasing age in vitro make the endothelial cell a suitable model to study in vitro culture‐related changes, senescence, cardiovascular disease, and tumorigenesis.
Dibutyryl‐cAMP increases functions of 5‐hydroxytryptamine2 receptors, but not of β2‐adrenergic receptors, in a clonal cell line of rat neurotumor RT4
Authors: Shigeri Yasushi , Watanabe Sachihiko , Fujimoto Masafumi 
Published: February 2005

AbstractA peripheral nervous system cell line RT4‐B, established by lmada and Sueoka (Dev. Biol., 66:97–108, 1978), was shown to respond to serotonin [5‐hydroxy‐tryptamine (5‐HT)] and catecholamines. 5‐HT induced a small and transient increase in cytosolic free Ca2+ concentration ([Ca2+]i) in the RT4‐B cells. The increase was effectively blocked by 5‐HT2 receptor antagonists (spiperone, ritanserin and mianserin), but not by a 5‐HT3 receptor antagonist (MDL72222), or a α1‐adrenergic receptor antagonist (prazosin), indicating that RT4‐B cells express 5‐HT2 receptors. On the other hand, catecholamines increased cyclic AMP production by RT4‐B. The order of potency for stimulating cyclic AMP synthesis was isoproterenol > epinephrine ≫ norepinephrine ≫ dopamine, and the stimulation was effectively inhibited by the nonselective β‐adrenergic receptor antagonist propranolol, but not by the β1‐adrenergic receptor antagonist atenolol, suggesting that RT4‐B cells express β2‐adrenergic receptors.The differentiating agent N6,2′‐O‐dibutyryladenosine 3′,5′‐monophosphate (dibutyryl‐cAMP) enhanced the 5‐HT‐induced [Ca2+]i increase, but not the catecholamine‐induced cyclic AMP production. The increase in the 5‐HT response paralleled the increase in the density of 5‐HT2 receptors. n‐Butyric acid (2 mM) and 8‐bromoadenosine 3′,5′‐monophosphate (1 mM) also increased the 5‐HT response, and the sum of these increases was nearly equal to that induced by dibutyryl‐cAMP.These results indicate that RT4‐B is a novel model cell line for the study of 5‐HT2 and β2‐adrenergic receptors and their second messenger responses and for the analysis of the mechanisms how 5‐HT2 receptor gene expression is controlled.
Sensitivity of cultured and skinned chick myotube to calcium, strontium, and barium ions examined by recording isometric contractions
Authors: Saito Koji , Yoshida Mikiharu , Tanaka Hikaru 
Published: February 2005

AbstractSensitivity of cultured chick myotubes to alkaline earth metal ions was investigated by recording contractile isometric tension through a semiconductor transducer. The myotubes were obtained by culturing myoblasts of chick embryo breast muscles, and skinned chemically before physiological experiments. Contractions developed in response to Ca2+ in a bathing medium higher than 3 × 10−7 M and reached maximum at 1 × 10−5 M. Sr2+ was less effective than Ca2+; the threshold concentration was 1 × 10−5 M and the tension reached maximum at 1 × 10−3 M. Ba2+ was the least effective among the three alkaline earth metal ions; only one fifth of the Ca2+‐induced maximum tension was attained at 1 × 10−3 M. The sensitivity was similar to that of the mature pectoral muscle fiber, a fast twitch muscle fiber, rather than that of the anterior latissimus dorsi, a slow tonic muscle fiber. The sensitivity was shown to be dependent on its troponin C by replacing it with troponin C from the mature pectoral or cardiac muscle. This indicates that TnC of a fast‐muscle type is expressed in the cultured chick myotube as in the mature pectoral muscle. The contractile apparatus was thus shown to be well developed in the cultured myotube with characteristics similar to the mature fast twitch muscle fiber.
Tumor necrosis factor stimulates DNA synthesis of mouse hepatocytes in primary culture and is suppressed by transforming growth factor β and interleukin 6
Authors: Satoh Motonobu , Yamazaki Masatoshi 
Published: February 2005

AbstractIn a previous study, we revealed that tumor necrosis factor (TNF) was secreted in mouse liver at an early phase of liver regeneration after partial hepatectomy. Here, we investigated direct actions of TNF on the in vitro DNA synthesis of adult mouse hepatocytes in primary culture. TNF enhanced both 3H‐TdR uptake and the number of 3H‐TdR‐labeled nuclei of hepatocytes. Their time courses were similar to those by epidermal growth factor (EGF) with about a 15 h lag period and a peak period of 24–48 h. This action of TNF was abrogated by DNA polymerase α inhibitor, aphidicolin and blocked specifically by anti‐TNF antibody. The actions of rm TNF were not distinguishable; ED50 was about 7.5U/ml (5ng/ml) and 30U/ml (20ng/ml) for maximal response (about 2‐fold or more of control). Other inflammatory monokines showed differential effects on in vitro DNA synthesis of hepatocyte. Neither type of interleukin 1 affected hepatocyte DNA synthesis in the range examined (up to 50 ng/ml). IL‐6 markedly inhibited the hepatocyte DNA synthesis stimulated by TNF and EGF. The action of TNF was completely suppressed by transforming growth factor β, which is known as a potent inhibitor of hepatocyte growth. Interferon γ also blocked this TNF action when added simultaneously. These results indicate that the activation of tissue macrophages and local secretion of TNF in liver after partial hepatectomy is of physiological importance in liver regeneration, in part by a direct stimulation of hepatocyte DNA synthesis. Cytokines induced by TNF may also participate in the later termination of liver regeneration.
Keratinocytes stimulate prostaglandin I2 synthesis by 3T3 cells and exhibit enhanced cornification when exposed to prostaglandin I2 analogues
Authors: Baden Howard P. , Goldaber Michael L. , Kvedar Joseph C. 
Published: February 2005

AbstractThe predominant cyclooxygenase products of keratinocytes are prostaglandin (PG)E2 and PGF2α with only trace amounts of PGI2 synthesis detected. When normal or immortal (NM1) keratinocytes were co‐cultured with mitomycin C‐treated 3T3 cells, increased synthesis of PGI2 was noted compared to mitomycin C‐treated 3T3 cells alone. The PGI2 level in co‐cultures was maximum within the first week and diminished rapidly thereafter. These results suggested keratinocytes enhance the production of PGI2 by 3T3 cells. Keratinocyte cultures incubated with lloprost and Piriprost, stable PGI2 analogues, showed evidence of increased cornification as demonstrated by staining with rhodanile blue, decreased shedding of cells into the culture medium, and more cornified material adhering to the culture surface. The cultures appeared to be responsive between the first and second weeks after plating and the inhibition of shedding could not be reversed by changing to drug‐free medium. Control and treated cultures showed identical electrophoretic protein patterns. Immunoblots showed involucrin unchanged in extracts of control and treated cultures while the 22 kd pancornulin was absent in treated cultures. The findings that keratinocytes enhance the production of PGI2 by 3T3 cells and that PGI2 analogues enhance cornification of confluent keratinocytes raise the possibility that eicosanoids may serve as autoregulatory signals together with other factors.