bluuebunny/crossref_metadata_2025_split huggingface dataset:
Endothelial cells exhibiting angiogenesis in vitro proliferate in response to TGF‐β1
Authors: Iruela‐Arispe M. Luisa , Sage E. Helene 
Published: May 2005

AbstractTransforming growth factor‐β1 (TGF‐β1) has been implicated in the positive regulation of angiogenesis in vivo, whereas it inhibits the proliferation of endothelial cells in vitro. To reconcile these apparently contradictory effects, we have investigated the effect of TGF‐β1 on bovine aortic endothelial cells that exhibit spontaneous angiogenesis in vitro. We show that concentrations of TGF‐β1 which stimulate proliferation of cells that form endothelial cords and/or tubes inhibit proliferation of the same cells grown at subconfluent densities. An increase in cell number of 35% over control cultures was achieved with 0.5 ng TGF‐β1/ml. The proliferative effect was blocked by antibodies against TGF‐β. Immunological detection of BrdU‐labeled nuclei revealed an increase greater than 220% in cells treated with TGF‐β1. Moreover, a population of cells within the cords appeared to be a selective target for this cytokine. The stimulatory effect was not restricted to bovine aortic endothelial cells, as similar results were obtained with endothelial cells derived from rat microvessels. Significant levels of active TGF‐β1 were detected in cultures containing cords/tubes, whereas only latent TGF‐β1 was detected in subconfluent cultures. We show further that endothelial cells exhibiting angiogenesis in vitro secrete plasminogen activator, an enzyme that regulates activation of TGF‐β. The major increases in mRNA transcripts for extracellular matrix proteins that are typically associated with TGF‐β1 were not seen in cells exhibiting angiogenesis in vitro. Since the formation of tubular networks requires both invasion and proliferation, we propose that TGF‐β1 is a major morphoregulatory factor in angiogenesis that specifically controls endothelial cell proliferation and extracellular matrix turnover.
Studies of the Levamisole inhibitory effect on rat stromal‐cell commitment to mineralization
Authors: Klein Benjamin Y. , Gal Irena , Segal David 
Published: May 2005

AbstractThe ability of Levamisole to decrease mineralization in skeletal tissue is usually related to its effect on alkaline phosphatase (ALP). However, Levamisole is also suspected to diminish mineralization by an additional mechanism which is unrelated to the ALP control of apatite crystal growth. To delineate the time in differentiation during which Levamisole inhibits mineralization, a tissue culture model system of bone marrow stromal cells was used. Secondary cultures of stromal cells were propagated in osteoprogenitor cell (OPC) induction medium for three weeks, followed by measurement of calcium precipitation. In situ ALP assays at pH 7.6 were also performed. When cells were cultured with 0.2 mM Levamisole for three weeks, Day 20 values of calcium precipitates were lower than in controls, but Day 20 ALP values were paradoxically higher. The correlation between calcium and ALP within each group was low. The correlation slightly improved, in uninhibited cultures, when Day 21 calcium values were matched with earlier Day 12 ALP values. This suggested the existence of a Levamisole‐sensitive mechanism for mineralization inhibition effective prior to the culture's mineralization stage. To focus on this early effect on mineralization Levamisole was added to stromal cultures on different days and removed on Day 12. Levamisole decreased Day 21 mineralization when added on Days 0, 3, 5, and 7, but not when added on Day 9. The Levamisole‐induced inhibition of mineralization was accompanied by an increase in Day 12 ALP specific activity, compared to controls, when added from Day 5 and thereafter. The results indicate that part of the ability of stromal cells to mineralize is determined during the first week of culture. The early inhibitory effect of Levamisole on mineralization was associated with increased Day 12 ALP activity.
Expression and localization of elements of the plasminogen activation system in benign breast disease and breast cancers
Authors: Jankun Jerzy , Merrick Hollis W. , Goldblatt Peter J. 
Published: May 2005

AbstractThe malignant potential of solid tumors is related to the ability to invade adjacent tissue and to metastasize. These properties of cancer cells depend on the synthesis of proteolytic enzymes which are able to digest adjacent connective tissue and basement membranes. We hypothesized that all elements of the plasminogen activation system might be overexpressed in malignant human breast tumors, functioning as an essential element in tumor invasion and metastasis. As determined by histopathological methods, the malignant tumors showed statistically significantly higher expression of urokinase plasminogen activator (uPA), type‐1 plasminogen activator inhibitor (PAI‐1), and especially urokinase plasminogen activator receptor (uPAR) than benign tissues. All those elements were present in higher amounts in the cancer cells than in the cells of benign or normal breast tissues. High exhibition of tissue plasminogen activator (tPA) found in cancer seems to be random and not related to the malignant or benign state, since benign and malignant tumors show overexpression of tissue plasminogen activator with similar frequency. When the tumors express high amounts of uPA, they express a high amount of uPAR in 50% of cases and PAI‐1 in 57.3% of cases. When urokinase is expressed in low amount, the receptor is low in 28.6% and inhibitor in 21.4% of malignant breast tumors. This statistically significant consensus, 78.6% in the case of urokinase and its receptor and 78.6% in case of urokinase and its inhibitor, suggests that these activities may be the result of a unique mechanism of control, activated in the last steps of malignant transformation.
Endothelin modulates osteopontin and osteocalcin messenger ribonucleic acid expression in rat osteoblastic osteosarcoma cells
Authors: Shioide Mitsuhiro , Noda Masaki 
Published: May 2005

AbstractEndothelins (ETs) are vasoconstrictive peptides produced mainly by endothelial cells. The ET receptors are expressed in many types of cells including osteoblast‐like cells. The purpose of this study was to examine the effects of endothelin on the expression of osteoblastic phenotype‐related genes. We found that endothelin‐1 (ET‐1) enhanced approximately two‐fold the mRNA expression of both osteopontin and osteocalcin genes in rat osteoblastic osteosarcoma ROS17/2.8 cells. These effects were dose‐dependent, peaking at 10−7 M. The ET‐1 enhancement of the abundance of osteopontin and osteocalcin mRNAs was time‐dependent, with a maximal effect at 24 h. ET‐1 modulation of the expression of the two phenotype‐related gene products of osteoblasts suggests that endothelin is one of the cytokines which modulate osteoblastic functions and that this molecule may play a role in the regulation of bone metabolism.
Purification and characterization of NADPH‐cytochrome P‐450 reductase from rat epidermis
Authors: Takahara Hidenari , Zaidi Syed I. A. , Mukhtar Hasan , Handa Masafumi , Epstein William L. , Fukuyama Kimie 
Published: May 2005

AbstractNADPH‐cytochrome P‐450 oxidoreductase (P‐450 red) transfers reducing equivalents from NADPH to cytochrome P‐450 (P‐450) in the monooxygenase system. Detergent solubilized proteins from the membrane fraction of neonatal rat epidermis were purified by 2′,5′‐ADP‐agarose affinity column chromatography. The purified protein showed an apparent homogeneity on sodium dodecylsulfate‐polyacrylamide gel electrophoresis and molecular weight was estimated to be 78 kDa. NADPH‐cytochrome c reductase activity increased by 95‐fold in the purified enzyme. Epidermal P‐450 red in vitro reconstituted benzo(a)pyrene hydroxylase activity in a dose dependent manner with P‐450 purified from either rat liver or epidermis. Western blot analysis demonstrated that epidermal P‐450 red immunologically cross reacts to liver P‐450 red. Immunohistochemical staining showed that the enzyme was predominantly localized in the epidermis. The intensity of immunohistochemical staining of rat skin sections and tissue distribution did not change in the skin treated with β‐naphtoflavone, which results in a substantial increase in P‐450 1A1 activity. Quantitative assessment of P‐450 red in treated and untreated epidermis also showed no change. These findings indicate that constitutive P‐450 red, fully capable of supporting P‐450, exists in rat epidermis, and can function in metabolism of endogenous and exogenous compounds.
1α,25‐Dihydroxyvitamin D3‐induced changes in intracellular pH in osteoblast‐like cells modulate gene expression
Authors: Jenis Louis G. , Lian Jane B. , Stein Gary S. , Baran Daniel T. 
Published: May 2005

Abstract1α,25‐Dihydroxyvitamin D3 exerts rapid nongenomic effects on rat osteoblast‐like cells independent of the classic nuclear receptor. These effects include changes in phospholipid metabolism and cell calcium. Intracellular calcium itself has been proposed to regulate intracellular pH in osteoblast cell lines. The purpose of this study was to determine the effect of 1α,25‐dihydroxyvitamin D3 on intracellular pH, the relationship of changes in calcium to changes in pH, and the role of pH changes in genomic activation. 1α,25‐Dihydroxyvitamin D3 increased intracellular pH within 10 min in rat osteoblast‐like cells, an effect that was inhibited by removal of extracellular sodium and by the biologically inactive epimer 1β,25‐dihydroxyvitamin D3. The hormone increased intracellular calcium in Quin 2 loaded cells in the presence and absence of extracellular sodium. The 1α,25‐dihydroxyvitamin D3‐induced increments in osteocalcin and osteopontin mRNA levels were abolished in sodium‐free medium. The results indicate that 1α,25‐dihydroxyvitamin D3‐induced increments in cellular calcium precede cell alkalinization and that these changes in intracellular pH may modulate steady‐state mRNA levels of genes induced by vitamin D.
Extracellular annexin II is associated with divalent cation‐dependent tumor cell–endothelial cell adhesion of metastatic RAW117 large‐cell lymphoma cells
Authors: Tressler Robert J. , Updyke Timothy V. , Yeatman Timothy , Nicolson Garth L. 
Published: May 2005

AbstractUsing fixed microvessel endothelial cell monolayers the molecules involved in the adhesion of liver‐preferring murine RAW117 large cell lymphoma cells to murine liver‐derived microvessel endothelial cells were identified by affinity isolation. Detergent lysates obtained from poorly (P) or highly (H10) liver‐metastatic cells inhibited RAW117‐H10 cell adhesion to hepatic sinusoidal endothelial (HSE) cell monolayers. Allowing detergent lysates of cell surface‐labeled RAW117 cells to bind to fixed HSE cell monolayers and eluting the bound components indicated that several tumor cell surface molecules ( ∼ 70, ∼ 35, ∼ 32, ∼ 22, and ∼ 14 kDa) might be involved in RAW117 cell‐HSE cell adhesion. The ∼ 35 kDa component was cation dependent in its binding to target HSE cells. Increasing detergent concentration had no effect on binding of the ∼ 35 kDa component to HSE cell monolayers, whereas treatment with 0.5 M NaCl resulted in its selective elution from HSE cells. Incubation of the HSE cell monolayers with detergent lysates from cell surface‐labeled RAW117‐H10 cells resulted in selective depletion of the ∼ 35 kDa component, suggesting that the binding is saturable. This divalent cation‐dependent molecule is one of the major tumor cell surface components bound by several types of endothelial cells and murine hepatocytes, whereas there was poor binding of this component to unfixed or fixed human red blood cells. The purified, partially ( ∼ 40%) sequenced molecule had amino acid sequence identity with murine but not bovine annexin II, indicating that it was not bound from the bovine serum used to grow RAW117 cells. Using antibodies specific for annexin II flow cytometery indicated equivalent amounts of annexin II are expressed on RAW117 cell surfaces in the absence or presence of excess EDTA, whereas annexin I was only found in low amounts on the surfaces of RAW117 cells. Annexin II antibodies inhibited by ∼ 40–50% the adhesion of RAW117 tumor cells to live or fixed endothelial cells, and purified tumor cell surface fractions containing the ∼ 35 kDa component partially inhibited ( ∼ 35%) RAW117 cell–HSE cell adhesion. The data indicate that annexin II is expressed on the extracellular surface of RAW117 cells, and cell surface–annexin II mediates a portion of the Ca2+‐dependent RAW117 cell adhesion to liver microvessel endothelial cells.
Activities of growth factors in preimplantation embryos
Authors: Adamson Eileen D. 
Published: May 2005

AbstractThe development of the mammalian preimplantation embryo in vitro occurs more slowly and less successfully compared to development in the uterus. The fact that it can occur at all in a defined protein‐free medium suggests that the process is autonomous. Accumulated evidence indicates that a number of peptide growth factors contribute in an autocrine fashion to preimplantation development. Other growth factors are maternally derived and act in a paracrine manner on the embryo. Some of these factors such as insulin‐related factors stimulate growth preferentially, but others such as epidermal growth factor (EGF) play more important roles in differentiation. Several cytokines appear to be implicated in peri‐implantation events and in maternal‐fetal interactions. At this stage, the data are mostly descriptive. Are all these different growth factors and receptors necessary for early development? Some implications of apparent redundancy of gene expression are discussed and future studies are predicted.
Hypothalamic releasing hormones mediating the effects of interleukin‐1 on sleep
Authors: Payne Linda C. , Obál Ferenc , Krueger James M. 
Published: May 2005

AbstractThere is a substantial literature describing the interactions between the endocrine and immune systems. Although such interactions are less well known within the brain, one major brain function altered during inflammation and infection and by several endocrine hormones is sleep. Pathological disturbances, be they inflammation, infectious disease, and/or sleep deprivation, result in altered hypothalamus‐pituitary function and cytokine metabolism. In respect to hormone secretion from the pituitary, cytokines are now recognized to play an important role in modulating the neuroendocrine system. Changes in sleep provide a useful illustration of the interactions between cytokines and the hypothalamus‐pituitary axis. Evidence linking interleukin‐1 (IL‐1) to growth hormone releasing hormone and to corticotropin releasing hormone in regard to their effects on sleep is reviewed.
Regulation of angiotensin I‐converting enzyme in cultured bovine bronchial epithelial cells
Authors: Müns Günther , Vishwanatha Jamboor K. , Rubinstein Israel 
Published: May 2005

AbstractThe purpose of this study was to determine whether angiotensin I‐converting enzyme (ACE) is present in cultured bovine bronchial epithelial cells (BBECs) and whether its activity can be modulated. We found that extracts of confluent monolayers of cultured BBECs degraded [glycine‐1‐14C]hippuryl‐L‐histidyl‐L‐leucine at a rate of 843 ± 66 pmol/hr/mg protein (mean ± SEM, n = 5). In addition, we found that the enzyme was shed into the culture medium. ACE activity in BBECs was inhibited by three selective, but structurally different, ACE inhibitors (captopril, quinapril, and cisalaprilat) with an IC50 of approximately 2 nM. Increasing chloride concentration in the assay buffer resulted in an increase in BBECs ACE activity of 63%. Enzyme activity was also modulated by the presence of zinc cation in the assay buffer. Addition of dexamethasone to the culture medium was associated with a significant increase in BBECs ACE activity (P < 0.05), which was inhibited by the steroid receptor antagonist RU 38486. Western blot analysis of BBECs, tracheal and bronchial mucosal strips utilizing a cross‐reacting rabbit anti‐mouse ACE antibody, showed a faint 175 kDa band and additional strong 52 kDa and 47 kDa band. The mechanism of generation of the low M.W. bands is unknown. Our data indicate the presence of ACE in cultured BBECs and that enzyme activity can be modulated.
Further purification and characterization of a multienzyme complex for DNA synthesis in human cells
Authors: Li Congjun , Cao Long‐Guang , Wang Yu‐Li , Baril Earl F. 
Published: May 2005

AbstractThe 21 S complex of enzymes for DNA synthesis in the combined low salt nuclear extract‐post microsomal supernatant from HeLa cells [Malkas et al. (1990) Biochemistry 29:6362–6374] was purified by poly (ethylene glycol) precipitation, Q‐Sepharose chromatography, Mono Q Fast Protein Liquid Chromatography (FPLC), and velocity gradient centrifugation. The procedure gives purified enzyme complex at a yield of 45%. The 21 S enzyme complex remains intact and functional in the replication of simian virus 40 DNA throughout the purification. Sedimentation analysis showed that the 21 S enzyme complex exists in the crude HeLa cell extract and that simian virus 40 in vitro DNA replication activity in the cell extract resides exclusively with the 21 S complex. The results of enzyme and immunological analysis indicate that DNA polymerase α‐primase, a 3′,5′ exonuclease, DNA ligase I, RNase H, and topoisomerase I are associated with the purified enzyme complex. Denaturing polyacrylamide gel electrophoresis of the purified enzyme complex showed the presence of about 30 polypeptides in the size range of 300 to 15 kDa. Immunofluorescent imaging analysis, with antibodies to DNA polymerase α,β and DNA ligase I, showed that polymerase α and DNA ligase I are localized to granular‐like foci within the nucleus during S‐phase. In contrast, DNA polymerase β, which is not associated with the 21 S complex, is diffusely distributed throughout the nucleoplasm.
Catecholamine stimulated lipolysis in differentiated human preadipocytes in a serum‐free, defined medium
Authors: van de Venter Maryna , Litthauer Derek , Oelofsen Willem 
Published: May 2005

AbstractAdipocyte precursors from the stromal vascular fraction of human adipose tissue were allowed to differentiate in serum‐free defined medium, whereafter their catecholamine stimulated lipolytic response was compared to that of mature isolated human adipocytes. Seventy‐five to ninety percent of the fibroblast‐like cells accumulated lipid droplets and glycerol‐3‐phosphate dehydrogenase activities of 1,000–2,800 mU/mg protein were measured in cell homogenates of differentiated cells. Lipolysis could be stimulated by both isoproterenol and norepinephrine in both differentiated preadipocytes as well as mature adipocytes. The results obtained with β‐adrenergic agents suggested the presence of a higher affinity receptor in differentiated preadipocytes as compared to mature adipocytes. Mature adipocytes responded well to β‐adrenergic agents, but no antilipolytic α2‐adrenergic response was observed in the differentiated preadipocytes. The presence of Gi proteins in the differentiated preadipocytes was suggested by the antilipolytic effect of adenosine as well as the lipolytic activity generated by pertussis toxin. In conclusion, our medium supported the differentiation of a very high percentage of human preadipocytes which developed a sensitive β‐adrenergic lipolytic response but which lacked an α2‐adrenergic antilipolytic response.
A 17S multiprotein form of murine cell DNA polymerase mediates polyomavirus DNA replication in vitro
Authors: Wu Yan , Hickey Robert , Lawlor Kenneth , Wills Philip , Yu Fang , Ozer Harvey , Starr Robyn , Quan Jiang Yuan , Lee Marietta , Malkas Linda 
Published: May 2005

AbstractWe have identified and purified a multiprotein form of DNA polymerase from the murine mammary carcinoma cell line (FM3A) using a series of centrifugation, polyethylene glycol precipitation, and ion‐exchange chromatography steps. Proteins and enzymatic activities associated with this mouse cell multiprotein form of DNA polymerase include the DNA polymerases α and δ, DNA primase, proliferating cell nuclear antigen (PCNA), DNA ligase I, DNA helicase, and DNA topoisomerases I and II. The sedimentation coefficient of the multiprotein form of DNA polymerase is 17S, as determined by sucrose density gradient analysis. The integrity of the murine cell multiprotein form of DNA polymerase is maintained after treatment with detergents, salt, RNase, DNase, and after chromatography on DE52‐cellulose, suggesting that the association of the proteins with one another is independent of nonspecific interaction with other cellular macromolecular components. Most importantly, we have demonstrated that this complex of proteins is fully competent to replicate polyomavirus DNA in vitro. This result implies that all of the cellular activities required for large T‐antigen dependent in vitro polyomavirus DNA synthesis are present within the isolated 17S multiprotein form of the mouse cell DNA replication activities. A model is proposed to represent the mammalian Multiprotein DNA Replication Complex (MRC) based on the fractionation and chromatographic profiles of the individual proteins found to co‐purify with the complex.
c‐myc Gene expression is localized to the myocyte following hemodynamic overload in vivo
Authors: Pollack Pia S. , Houser Steven R. , Budjak Ricardo , Goldman Bruce 
Published: May 2005

AbstractExpression of the proto‐oncogene c‐myc increases in the hemodynamically overloaded heart, but expression by cardiac myocytes has not been shown. To address this issue, right ventricular overload was induced in cats by pulmonary artery banding. Expression of c‐myc and α‐skeletal actin mRNA were determined by Northern analysis. Immuno‐reactive Myc protein was identified by histochemical staining.Steady state levels of c‐myc mRNA peaked within 2 h after banding. Levels of α‐skeletal actin mRNA were maximally increased 48 h–1 week after banding and were still elevated at 1 month. Prominent staining of myocyte nuclei for immunoreactive Myc protein was detected 48 h after banding although a few interstitial nuclei were also positive.These studies show that c‐myc and α‐skeletal actin gene expression are upregulated in a large animal model of hemodynamic overload. The localization of the immunoreactive Myc protein to right ventricular myocyte nuclei after pulmonary artery banding supports the hypothesis that c‐myc induction is part of a general response in cardiac hypertrophy that is common to many mammalian species.
Transient increase in vimentin phosphorylation and vimentin‐HSC70 association in 9L rat brain tumor cells experiencing heat‐shock
Authors: Cheng Ting‐Jen , Lai Yiu‐Kay 
Published: May 2005

AbstractCharacteristic changes in vimentin were studied in 9L rat brain tumor cells treated at 45°C. During heat‐shock treatment, vimentin molecules were rapidly phosphorylated and reorganized from a filamentous form into a perinuclear higher‐order structure that was less extractable by nonionic detergent. These effects were found to be highly transient, peaked at 30 min after the onset of heat‐shock treatment, and subsided thereafter. Simultaneously, the solubility of the constitutively expressed heat‐shock protein70 (HSC70) was also temporarily decreased and the kinetics was identical to that of vimentin. The results indicated that HSC70 and vimentin were co‐insolubilized during the heat‐shock treatment. We propose that the reorganization of the intermediate filaments resulted from enhanced phosphorylation of vimentin leads to the concurrent association of HSC70 to the intermediate filaments. This process may play an essential role in regulating heat‐shock genes.