bluuebunny/crossref_metadata_2025_split huggingface dataset:
Casein accumulation in distended rough endoplasmic reticulum of collagen gel‐cultivated mouse mammary epithelia
Authors: Hurley David , Hwang Soo‐In , Rocha Victor 
Published: February 2005

AbstractMouse mammary epithelial cells cultivated on collagen gels synthesize and secrete casein in a hormone‐dependent manner. Fine‐structure electron microscopy of secretory cultures revealed numerous cytoplasmic structures surrounded by membrane that is studded with ribosomes. The structures appear to be distended rough endoplasmic reticulum (RER). Electron microscope protein A‐colloidal gold immunolocalization showed casein antiserum‐specific deposition of gold particles over the RER cytoplasmic vesicles in cells provided insulin, prolactin, and hydrocortisone (IPF). Nonimmune antiserum showed no gold particle deposition over these cytoplasmic structures. Epithelia provided only insulin showed no such cytoplasmic vesicles nor any specific deposition of gold particles. Immunoblot analysis of cell lysate and culture medium showed casein only in IPF‐treated cultures. It appears that the casein secretory pathway in collagen gel cultured mammary epithelia is blocked at the step that fuses RER vesicles to Golgi membrane. The data raise questions regarding the processing and maturation of casein and the mechanism of casein secretion in these cultures.
Membrane‐associated hyaluronate‐binding activity of chondrosarcoma chondrocytes
Authors: McCarthy Mary T. , Toole Bryan P. 
Published: February 2005

AbstractThe association of hyaluronate with the surface of chondrocytes was examined by several approaches using primary cultures of chondrocytes derived from the Swarm rat chondrosarcoma. In culture, chondrosarcoma chondrocytes produced large pericellular coats, which can be visualized by particle exclusion, and which can be removed by Streptomyceshyaluronidase. Exposure of chondrocytes, which had been metabolically labelled with 3H‐acetate, to exogenous hyaluro‐nate or to Streptomyceshyaluronidase resulted in the release of 36–38% of the endogenous, labelled chondroitin sulfate from the cell layer into the incubation solution. These results imply that at least 37% of the cell layer chondroitin sulfate proteoglycan is retained there by an interaction with hyaluronate. Thus membranes were prepared from cultured chondrocytes and examined for sites which bind3H‐hyaluronate. Binding was observed and found to be saturable, specific for hyaluronate, of high affinity (Kd =∼10−10M),and destroyed by treating the membranes with trypsin. The 3 H‐hyaluronate‐binding activity was inhibited competitively by hyaluronate decasaccharides but not by hexasaccharides or octasaccharides, indicating that the binding sites recognize a sequence of hyalu‐ronate composed of five disaccharide repeats. The binding activity was partially purified from a detergent extract of chondrocyte membranes by ion exchange chromatography on DEAE‐cellulose, followed by affinity chromatography on wheat germ agglutinin‐agarose. Analysis of the partially purified binding activity by SDS‐PAGE revealed five protein bands of 48,000–66,000 daltons in silver‐stained gels. SDS‐PAGE followed by Western blotting and exposure to mono‐clonal antibodies which recognize epitopes present in link protein and in the hyaluronate‐binding region of cartilage proteoglycan revealed no immunoreac‐tive protein bands in the partially purified material. We conclude that one mechanism by which hyaluronate associates with the chondrocyte surface may be via interaction with a membrane‐bound hyaluronate‐binding protein which is distinct from link protein and proteoglycan.
Possible role of prostaglandins as negative regulators in growth stimulation by tumor necrosis factor and epidermal growth factor in human fibroblasts
Authors: Hori Takamitsu , Kashiyama Satoshi , Hayakawa Makio , Shibamoto Sayumi , Tsujimoto Masafumi , Oku Naoto , Ito Fumiaki 
Published: February 2005

AbstractRecombinant tumor necrosis factor (TNF), epidermal growth factor (EGF), and transforming growth factor β (TGF‐β) stimulated growth of confluent human diploid fibroblasts (FS‐4 cells) in the presence of fetal calf serum. TGF‐β synergistically enhanced both the TNF‐ and EGF‐stimulated cell growth, whereas synergism between the mitogenic action of EGF and that of TNF was not observed. When indomethacin or acetylsalicylic acid, an inhibitor of prostaglandin production, was added to FS‐4 cells, cell growth stimulated by EGF or TNF was increased, suggesting that prostaglandins induced by these mitogens antagonize their growth stimulatory actions. In contrast, neither indomethacin nor acetylsalicylic acid had a significant effect on the TGF‐β‐induced growth of FS‐4 cells. Mitogenic responses of indomethacin‐treated cells to EGF, TNF, and TGF‐β were similarly suppressed by the addition of exogenous prostaglandin D2 (PGD2). Other prostaglandins such as PGE2 and PGF2α produced less inhibition of the cell growth.
Enucleation of the rat pheochromocytoma clonal cell line, PC12: Effect on neurite outgrowth
Authors: Nichols Robert A. , Chandler Charles E. , Shooter Eric M. 
Published: February 2005

AbstractThe effect of removal of PC12 cell nuclei on neurite outgrowth was studied. Enucleation (80–90%) was accomplished in the presence of cytochalasin B using a centrifugation technique that exploited the very tight adhesivity of PC12 cells for a substratum composed of an extracellular matrix secreted by bovine corneal endothelial cells in response to epidermal growth factor treatment. Neither nucleated nor enucleated PC 12 cells showed significant neurite outgrowth on this particular matrix in the absence of nerve growth factor. In the presence of nerve growth factor both PC12 cell types initiated neurite outgrowth, but whereas neurites from nucleated cells grew continuously for two days, those from enucleated cells reached a maximum length after one day. The results suggest that neurite initiation but not continued neurite growth or stabilization can occur in the absence of transcription.
Role of protein kinase c in interleukin 1, anti‐T3, and mitogenic lectin‐induced interleukin 2 secretion
Authors: Mills Gordon B. , May Christopher , Hill Mary , Gelfand Erwin W. 
Published: February 2005

AbstractActivation of T‐lymphocytes by antigen, mitogenic lectins, or antibodies against the T‐cell receptor complex, particularly in the presence of IL1, induces the secretion of the T‐cell growth factor IL2. IL2 then has a major role in regulating the duration and magnitude of the immune response. Interaction of antigen, antibodies against the T‐cell receptor complex, or mitogenic lectins with T‐lymphocytes also induces hydrolysis of membrane phospholipids, leading to the production of diacylglycerol, an activator of the Ca2+‐ and phospholipid‐dependent phosphotransferase, protein kinase C (PKC). Phorbol esters, potent activators of PKC, augment secretion of the T‐cell growth factor, interleukin 2 (IL2). Activation of PKC may therefore serve as an important early event in the production and secretion of IL2. We have determined whether IL2 secretion can be induced in the murine cell T‐lymphocyte line LBRM 331A5, where PKC is inhibited by staurosporine or sphingosine or in cells where PKC is depleted by prolonged incubation with high concentrations of phorbol esters. In cells in which PKC was either inhibited or depleted, antibodies against the T3 portion of the T‐cell receptor complex and the mitogenic lectin phytohemagglutinin (PHA) still triggered IL2 secretion. In addition, the monokine Ill augmented this IL2 secretion irrespective of whether PKC was inhibited or depleted. These data indicate that activation of PKC is not an obligatory step for IL2 secretion in LBRM 331A5 murine T‐lymphocytes.
Analysis of effect of age on synthesis of specific proteins by hepatocytes
Authors: Butler James A. , Heydari Ahmad R. , Richardson Arlan 
Published: February 2005

AbstractThe effect of age on the synthesis of specific proteins by hepatocytes was studied in Fischer F344 rats using two‐dimensional polyacrylamide gel electrophoresis. Almost all proteins synthesized by hepatocytes from young rats were synthesized by hepatocytes isolated from old rats. Of over 500 proteins visually compared by two‐dimensional polyacrylamide gel electrophoresis, only 11 proteins were observed to disappear and/or appear consistently with increasing age. The rates of synthesis of 36 randomly chosen proteins were quantified. Interestingly, the synthesis of 35 of the 36 proteins decreased between 5 and 30 months of age. The decrease in protein synthesis varied (15% to 70%) from one protein to another; i.e., a heterogeneity was observed in the age‐related decrease in the synthesis of proteins. The age‐related decrease in protein synthesis was statistically significant for 53% of the proteins studied. The total decrease in the rate of synthesis of all 36 proteins studied was 40% between 5 and 30 months of age, which is essentially the same as the decrease in total protein synthesis by suspension of hepatocytes isolated from 5‐ and 30‐month‐old rats. The results of this study demonstrate that the mechanism underlaying aging is different from development, which is characterized by a major change in the species of proteins synthesized by a cell.
Contribution of endosomes to intrahepatic distribution of apolipoprotein B and apolipoprotein E
Authors: Wong Laurence 
Published: February 2005

AbstractThe endosomal contribution of apolipoprotein B and apolipoprotein E to the intrahepatic distribution of these 2 apolipoproteins was studied. The endosomal contribution of the apolipoproteins was estimated by comparing the specific radioactivities of the injected apolipoproteins with that of apolipoproteins in the liver. The endosomal contribution is further validated by using a least‐squares linear combination method that has been described previously (Wong and Pino: European Journal of Biochemistry 164: 357–367, 1987). The results using these two independent analyses were similar. There was no difference in endosomal contribution of the apolipoproteins between the fasted and fed rats. In both cases, total endosomal contribution in the microsomal fraction of apolipoprotein B was estimated at 7–10% and that for apolipoprotein E was estimated at 62–66%. The molecular weights of the two apolipoproteins in the endosomes closely resembles those of the injected apolipoproteins in the very low density lipoproteins, indicating little or no apolipoprotein loss during receptor‐mediated endocytosis. It is concluded that while there was no substantial contribution of apolipoprotein B to the intrahepatic distribution of that protein, the contribution of extrahepatic apolipoprotein E to intrahepatic apolipoprotein E distribution is substantial.
Calcium‐independent growth of human ovarian carcinoma cells
Authors: Chan Thomas C. K. 
Published: February 2005

AbstractEvidence in the literature suggests that cancer cell growth in vitro is generally not sensitive to external calcium. A human ovarian carcinoma cell line (SKOV3) retained 60% of its normal growth in Dulbecco modified Eagle's medium (DME) when the calcium concentration was reduced from 3 mM to 10 μM. Chinese hamster ovary cells (CHO) were growth‐arrested in media containing less than 500 μM calcium. In low‐calcium (10 μM) DME, 10 μM of a calmodulin antagonist W7 inhibited the growth of SKOV3 cells by more than 90%, while 100 μM of its inactive analog W5 was mildly inhibitory (20%). The growth inhibition by W7 was antagonized by increasing calcium concentrations in the culture media, while the inhibition by W5 was calcium‐independent. The phorbol ester TPA was also effective in antagonizing W7's growth inhibition in low‐calcium DME, suggesting that the W7 effect is mediated via protein kinase C inhibition. SKOV3 total cellular protein kinase C activity was 1.6 times higher than CHO cells when incubated in normal DME. When incubated in low‐calcium DME, a large drop in protein kinase C activity in the CHO cells was observed while the enzyme activity was unchanged in the SKOV3 cells. Our data suggest that these human ovarian tumor cells have altered cellular calcium regulatory processes associated with the defective down‐regulation of protein kinase C. This defect may confer these cells the ability to proliferate independently of the external calcium concentration. Targeting the cellular signal transduction components may be useful in cancer chemotherapy.
Heat protectors and heat‐induced preferential redistribution of 26 and 70 kDa proteins in chinese hamster ovary cells
Authors: Lee Yong J. , Armour Elwood P. , Borrelli Michael J. , Corry Peter M. 
Published: February 2005

AbstractAn overall increase of 40% in nuclear‐associated protein has been shown to be one of the sequellae of exposure of eukaryotic cells to elevated temperatures. Several investigators have shown that the increased protein/DNA ratios correlated well with the degree of cytotoxicity. In previous investigations, we have shown that cycloheximide, which protects the cell from the killing effects of heat, produces a dramatic reduction of the bulk nuclear‐associated proteins after heating. In this investigation, we studied a previously unobserved efflux of a 26 kDa protein after heat shock and the preferential accumulation of the 70 kDa protein. The 26 kDa protein was shown not to be a member of previously described heat shock protein families. Preferential reduction of a 26 kDa protein and accumulation of a 70 kDa protein was observed in nuclei isolated from Chinese hamster ovary cells after heating at 43°C. After heat treatment, the 26 kDa protein in the nucleus was decreased to a level 0.1–0.3 times the original amount in unheated cells, and the 70 kDa protein in the nucleus increased by a factor of 1.6–1.8. The normal levels of these two proteins were restored when cells were incubated at 37°C following heat shock. Cells treated with heat protectors, cycloheximide and histidinol, demonstrated approximately the same redistribution in nuclear 26 and 70 kDa proteins immediately after heating as those not exposed to these drugs. On the other hand, restoration to control levels was much faster in the protector‐treated cells, suggesting that “repair” of heat‐induced damage is an important factor in the cells ability to survive this insult. Return to normal protein levels did not require new protein synthesis.
Effects of D‐threo‐PDMP, an inhibitor of glucosylceramide synthetase, on expression of cell surface glycolipid antigen and binding to adhesive proteins by B16 melanoma cells
Authors: Inokuchi Jin‐Ichi , Momosaki Keiko , Shimeno Hiroshi , Nagamatsu Atsuo , Radin Norman S. 
Published: February 2005

AbstractIncubating B16 melanoma cells with an inhibitor of glucosylceramide (GlcCer) synthetase, D‐threo‐1‐phenyl‐2‐decanoylamino‐3‐morpholino‐1‐propanol (D‐threo‐PDMP), led to a considerable decrease in the levels of GlcCer and lactosylceramide (LacCer). The content of ganglioside GM3 was little affected, but the ability to bind a monoclonal antibody against the ganglioside (M2590) was greatly reduced, suggesting that the reduction in the simple glycolipids led to encryption of the membrane antigen. This interpretation is supported by the observation that permeabilization of the treated cells with Triton X‐100 restored immunological reactivity. Specificity of the PDMP effect was shown by its lack of effect on the reactivity of two other surface antigens to anti‐melanoma monoclonal antibodies M562 and M622, and of the major histocompatibility antigens to anti‐H‐2KbDb monoclonal antibody. The ability of the treated cells to attach to laminin or type IV collagen was lost but that to fibronectin was not. The effects of the enzyme inhibitor were counteracted by including GlcCer in the culture medium. This indicates that the lipid was absorbed by the cells and utilized like endogenously‐formed GlcCer. Cells preattached to laminin or collagen could be induced to round up by addition of inhibitor. In contrast, L‐threo‐PDMP (which does not block the synthesis of GlcCer) had no effect on the immunologic reactivity of GM3 or the adhesion properties of the cells. However, it did produce considerable accumulation of LacCer. These data suggest that the simple glycolipid, GlcCer, is an essential factor for antigenic expression of the more complex glycolipids on cell surfaces and that there is a close association and interaction between glycolipids and adhesive receptors on the cell surface.
Hemin enhances differentiation and maturation of cultured regenerated skeletal myotubes
Authors: Funanage Vicky L. , Schroedl Nancy A. , Moses Priscilla A. , Smith Susan M. , Kirwin James J. , Hartzell Charles R. 
Published: February 2005

AbstractSatellite cells, isolated from marcaine‐damaged rat skeletal muscle, differentiate in culture to form contracting, cross‐striated myotubes. Addition of 20 μM hemin (ferriprotoporphyrin IX chloride) to the culture medium resulted in increases in the number, size, and alignment of myotubes; in the number of myotubes that exhibited cross‐striations; and in the strength and frequency of myotube contractions. Hemin increased satellite cell fusion by 27%, but decreased cell proliferative rate by 30%. Hemin increased the specific activity of creatine kinase (CK), a sensitive indicator of muscle differentiation, by 157%. Separation of CK isoenzymes by agarose gel electrophoresis showed that hemin increased only the muscle‐specific CK isoenzymes (MM‐CK and MB‐CK). Thus, hemin seems to duplicate some of the effects of innervation on cultured myotubes by increasing contraction frequency and strength, appearance of cross‐striations, and muscle‐specific isoenzymes. In contrast, 3‐amino‐1,2,4‐triazole, an inhibitor of heme biosynthesis, decreased the number of cross‐striated myotubes, the strength and frequency of myotube contractions, and CK activity. These inhibitory effects were reversed by hemin. Collectively, these results demonstrate a physiologically significant role for heme in myotube maturation.
Reversible forskolin‐induced impairment of sucrase‐isomaltase mRNA levels, biosynthesis, and transport to the brush border membrane in Caco‐2 cells
Authors: Rousset Monique , Chantret Isabelle , Darmoul Dalila , Trugnan Germain , Sapin Catherine , Green Fiona , Swallow Dallas , Zweibaum Alain 
Published: February 2005

AbstractHybridization analysis of mRNA with a cDNA probe for human sucrase‐isomaltase, pulse‐chase experiments with L‐[35S]‐methionine followed by SDS‐PAGE, and immunofluorescence detection of sucrase‐isomaltase were used to analyze the level(s) at which forskolin interferes with the expression of the enzyme in Caco‐2 cells in culture. Three effects are observed in Caco‐2 cells treated with forskolin: (1) a marked decrease in the level of sucrase‐isomaltase mRNA, (2) a marked decrease in the biosynthesis of the enzyme without any alteration of its stability, and (3) an almost total inhibition of its transport to the brush border membrane. All three effects are reversible when the drug is removed from the culture medium, though this reversibility is asynchronous: transport to the brush border membrane resumes after 24 h, sucrase‐isomaltase mRNA levels are back to the normal after 5 days, whereas the biosynthesis of the enzyme, although increasing progressively, remains lower than in control cells, even 10 days after removal of the drug. The possibility that some effects are directly dependent on cAMP and others a consequence of changes in glucose metabolism is discussed.
Pulmonary microvascular endothelial cell contractility on silicone rubber substrate
Authors: Morel Nicole M. L. , Dodge Andrea B. , Patton Wayne F. , Herman Ira M. , Hechtman Herbert B. , Shepro David 
Published: February 2005

AbstractEndothelial cell (EC) motility may contribute to the regulation of microvascular perfusion and/or paracellular permeability. The experiments reported herein demonstrate that bovine pulmonary microvessel EC can reversibly deform a silicone substrate in response to agents known to contract and relax smooth muscle cells. Contracting pulmonary microvessel EC exerted a tension that created wrinkles in the underlying deformable substrate. Relaxation and loss of tension were revealed by the disappearance of these wrinkles without loss of cell adhesion to the substratum. Angiotensin II (Ang II) and bradykinin stimulated pulmonary microvessel EC to contract within 3 to 8 min in a Ca2+‐dependent fashion. The peak of contraction at 10 to 20 min was followed by relaxation. Forskolin and sodium nitroprusside (SNP) initiated relaxation of the microvessel EC within 3 to 10 min respectively. Relaxed EC contracted following the addition of Ang II, also within 3 min. Dibutyryl cAMP, dibutyryl cGMP, and the photoactivated internalized “caged” cAMP and cGMP promoted EC relaxation in a manner similar to forskolin and SNP. Increases in the intracellular concentration of inositol triphosphate (IP3) with the photoactivated IP3 complex promoted EC contraction in 2 min with a peak at 7 min. The contraction was followed by relaxation, which occurred at 20–25 min. Neither bovine pulmonary artery nor retinal microvessel EC, used as controls, contracted under these experimental conditions. One could speculate that this unique contractile property of pulmonary microvessel EC as observed in vitro may play a regulatory role in vivo, in local perfusion and/or in intercellular gap regulation.
Secretory character of a group of isoproterenol‐induced polypeptides in mouse parotid glands
Authors: Solís Remigio O. López , Miranda Dante , Alliende Cecilia , González Julieta , Díaz Nelson , Díaz Héctor , Ruiz Fernando 
Published: February 2005

AbstractThe secretory nature of the isoproterenol‐induced mouse parotid polypeptides C, D, E, F, and G (molecular weights 64,000, 61,000, 51,500, 38,000, and 37,000, respectively) is documented. Polypeptides C, D, E, F, and G, accumulated in response to successive daily stimulations with isoproterenol, were detected in a fraction enriched in hypertrophic parotid acinar cells. These cells, characterized by an increased content of cytoplasmic granules, maintain a secretory responsiveness to isoproterenol, which has been evidenced by light microscopy, enzymatic analysis, and unidimensional SDS‐polyacrylamide gel electrophoresis. Thus, a parallelism in the loss and recovery of both secretory granules, α‐amylase and polypeptides C, D, E, F, and G, was observed. Moreover, after secretion stimulation, polypeptides C, D, E, F, and G were detected in the fluid collected directly from parotid gland cannulation. Given the secretory character of polypeptides C, D, E, F, and G, mechanisms explaining both their progressive accumulation along the chronic administration of isoproterenol, as well as their progressive disappearance observed after suspending that treatment, are discussed.