bluuebunny/crossref_metadata_2025_split huggingface dataset:
Suppressive effect of endogenous regucalcin on nitric oxide synthase activity in cloned rat hepatoma H4‐II‐E cells overexpressing regucalcin
Authors: Izumi Takako , Tsurusaki Yoshinori , Yamaguchi Masayoshi 
Published: July 2003

AbstractThe role of endogenous regucalcin, which is a regulatory protein in calcium signaling, in the regulation of nitric oxide (NO) synthase activity in the cloned rat hepatoma H4‐II‐E cells was investigated. Hepatoma cells were cultured for 24–72 h in the presence of fetal bovine serum (FBS; 10%). NO synthase activity in the 5,500 g supernatant of cell homogenate was significantly increased by the addition of calcium chloride (10 μM) and calmodulin (2.5 μg/ml) in the enzyme reaction mixture. The presence of trifluoperazine (TFP; 50 μM), an antagonist of calmodulin, inhibited the effect of calcium (10 μM) addition in increasing NO synthase activity, indicating the existence of Ca2+/calmodulin‐dependent NO synthase in hepatoma cells. NO synthase activity was significantly decreased by the addition of regucalcin (10−8 or 10−7 M) in the reaction mixture without or with Ca2+/calmodulin addition. The effect of regucalcin (10−7 M) in decreasing NO synthase activity was also seen in the presence of TFP (50 μM) or EGTA (1 mM). The presence of anti‐regucalcin monoclonal antibody (10–50 ng/ml) in the reaction mixture caused a significant elevation of NO synthase activity. NO synthase activity was significantly suppressed in the hepatoma cells (transfectants) overexpressing regucalcin. This decrease was completely abolished in the presence of anti‐regucalcin monoclonal antibody (50 ng/ml) in the reaction mixture. Moreover, the effect of Ca2+/calmodulin addition in increasing NO synthase activity in the hepatoma cells (wild‐type) was completely prevented in transfectants. The present study demonstrates that endogenous regucalcin has a suppressive effect on NO synthase activity in the cloned rat hepatoma H4‐II‐E cells. J. Cell. Biochem. 89: 800–807, 2003. © 2003 Wiley‐Liss, Inc.
Nuclear distribution of Oct‐4 transcription factor in transcriptionally active and inactive mouse oocytes and its relation to RNA polymerase II and splicing factors
Authors: Parfenov Vladimir N. , Pochukalina Galina N. , Davis Donna S. , Reinbold Rolland , Schöler Hans R. , Murti Kuruganti G. 
Published: July 2003

AbstractThe intranuclear distribution of the transcription factor Oct‐4, which is specifically expressed in totipotent mice stem and germ line cells, was studied in mouse oocytes using immunogold labeling/electron microscopy and immunofluorescence/confocal laser scanning microcopy. The localization of Oct‐4 was studied in transcriptionally active (uni/bilaminar follicles) and inactive (antral follicles) oocytes. Additionally, the Oct‐4 distribution was examined relative to that of the unphosphorylated form of RNA polymerase II (Pol II) and splicing factor (SC 35) in the intranuclear entities such as perichromatin fibrils (PFs), perichromatin granules (PGs), interchromatin granule clusters (IGCs), Cajal bodies (CBs), and nucleolus‐like bodies (NLBs). It was shown that: (i) Oct‐4 is localized in PFs, IGCs, and in the dense fibrillar component (DFC) of the nucleolus at the transcriptionally active stage of the oocyte nucleus; (ii) Oct‐4 present in PFs and IGCs colocalizes with Pol II and SC 35 at the transcriptionally active stage; (iii) Oct‐4 accumulates in NLBs, CBs, and PGs at the inert stage of the oocyte. The results confirm the previous suggestion that PFs represent the major nucleoplasmic structural domain involved in active pre‐mRNA transcription/processing. The colocalization of Oct‐4 with Pol II in both IGCs and PFs in active oocytes (uni/bilaminar follicles) suggests that Oct‐4 is intimately associated with the Pol II holoenzyme before and during transcription. The colocalization of Oct‐4, Pol II, and SC 35 with coilin‐containing structures such as NLBs and CBs at the inert stage (antral follicles) suggests that the latter may represent storage sites for the transcription/splicing machinery during the decline of transcription. J. Cell. Biochem. 89: 720–732, 2003. © 2003 Wiley‐Liss, Inc.
Altered gene expression profile in the kidney of vitamin D receptor knockout mice
Authors: Li Xinmin , Zheng Wei , Li Yan Chun 
Published: July 2003

AbstractThe kidney is a primary target organ of the vitamin D endocrine system, and both vitamin D‐deficiency and vitamin D receptor (VDR) ablation lead to impaired renal functions. As an initial step to understand the molecular basis underlying the renal dysfunctions resulted from VDR inactivation, we used DNA microarray technology to search for changes in the gene expression profile in the kidney of VDR knockout mice. Three independent DNA microarray experiments were performed using Affymetrix GeneChips, which included two replicate comparisons between VDR null and wild‐type littermates, and a third comparison between 1,25‐dihydroxyvitamin D3‐treated and vehicle‐treated wild‐type mice. Based on the assumption that VDR inactivation and vitamin D stimulation cause opposite changes in the expression of vitamin D target genes, we identified 95 genes that displayed the same changes in the two VDR‐null/wild‐type comparisons but an opposite change in the third assay, of which 28 genes were up‐regulated and 67 were down‐regulated in VDR null mice. These genes can be divided into several functional categories involved in vitamin D and steroid metabolism, calcium metabolism and signaling, volume and electrolyte homeostasis, signal transduction, transcriptional regulation, cell adhesion, metabolism, immune response, and other functions. These data provide a basis for further investigations into the molecular bases underlying the physiological abnormalities associated with VDR‐ and vitamin D‐deficiency. J. Cell. Biochem. 89: 709–719, 2003. © 2003 Wiley‐Liss, Inc.
Mammalian transcription factor LSF is a target of ERK signaling
Authors: Pagon Zrinka , Volker Janet , Cooper Geoffrey M. , Hansen Ulla 
Published: July 2003

AbstractLSF is a mammalian transcription factor that is rapidly and quantitatively phosphorylated upon growth induction of resting, peripheral human T cells, as assayed by a reduction in its electrophoretic mobility. The DNA‐binding activity of LSF in primary T cells is greatly increased after this phosphorylation event (Volker et al. [1997]: Genes Dev 11:1435–1446). We demonstrate here that LSF is also rapidly and quantitatively phosphorylated upon growth induction in NIH 3T3 cells, although its DNA‐binding activity is not significantly altered. Three lines of experimentation established that ERK is responsible for phosphorylating LSF upon growth induction in both cell types. First, phosphorylation of LSF by ERK is sufficient to cause the reduced electrophoretic mobility of LSF. Second, the amount of ERK activity correlates with the extent of LSF phosphorylation in both primary human T cells and NIH 3T3 cells. Finally, specific inhibitors of the Ras/Raf/MEK/ERK pathway inhibit LSF modification in vivo. This phosphorylation by ERK is not sufficient for activation of LSF DNA–binding activity, as evidenced both in vitro and in mouse fibroblasts. Nonetheless, activation of ERK is a prerequisite for the substantial increase in LSF DNA–binding activity upon activation of resting T cells, indicating that ERK phosphorylation is necessary but not sufficient for activation of LSF in this cell type. J. Cell. Biochem. 89: 733–746, 2003. © 2003 Wiley‐Liss, Inc.
Increased expression of disintegrin‐metalloproteinases ADAM‐15 and ADAM‐9 following upregulation of integrins α5β1 and αvβ3 in atherosclerosis
Authors: Al‐Fakhri Nadia , Wilhelm Jochen , Hahn Meinhard , Heidt Martin , Hehrlein Friedrich W. , Endisch Andrea M. , Hupp Thomas , Cherian Sanjay M. , Bobryshev Yuri V. , Lord Reginald S.A. , Katz Norbert 
Published: July 2003

AbstractRegulation of αvβ3 and α5β1 integrin function plays a crucial role in atherosclerosis. Possible regulators of integrin–matrix interactions are integrin‐binding ADAMs (proteins with a disintegrin‐ and metalloproteinase‐domain), like ADAM‐15 and ADAM‐9. Molecular interactions between ADAM‐15, α5β1, and αvβ3 have been demonstrated. ADAM‐9 and ADAM‐15 were found to be interdependently regulated. This study, therefore, investigated whether the upregulation of integrins α5β1 and αvβ3 was correlated with the expression of integrin‐binding ADAMs in atherosclerotic processes. Human arterial and venous vascular smooth muscle cells (VSMCs) were incubated with PDGF over different time intervals up to a 3‐day culture period. mRNA concentrations, quantified by real‐time RT‐PCR and normalized to PBGD, of integrins αvβ3 and α5β1 were strongly increased after a 12‐h PDGF‐incubation in arterial and venous VSMC. ADAM‐15 and ADAM‐9 mRNA production showed a corresponding increase following integrin upregulation after a 24‐h incubation period. Western blot anaylsis revealed an increased protein expression of integrins and ADAMs in PDGF‐stimulated VSMC. Additionally, mRNA concentrations of atherosclerotic and normal human specimens were quantified by real‐time RT‐PCR. mRNA of ADAMs and integrins was significantly increased in atherosclerotic arteries compared to normal arteries. Immunohistochemistry of these specimens showed an increased expression and codistribution of both ADAMs and integrins in atherosclerosis. In conclusion, upregulation of ADAM‐15 and ADAM‐9 in atherosclerosis appears to follow an increase in α5β1 and αvβ3 integrins. Since α5β1 and αvβ3 are known to promote smooth muscle cell migration and proliferation, upregulation of ADAM‐15 and ADAM‐9 could balance integrin–matrix interactions and cell migration, thus modulating neointima progression. J. Cell. Biochem. 89: 808–823, 2003. © 2003 Wiley‐Liss, Inc.
Mechanisms of free‐radical induction in relation to fenretinide‐induced apoptosis of neuroblastoma
Authors: Lovat Penny E. , Ranalli Marco , Corazzari Marco , Raffaghello Lizzia , Pearson Andy D.J. , Ponzoni Mirco , Piacentini Mauro , Melino Gerry , Redfern Christopher P.F. 
Published: July 2003

AbstractThe mechanisms of fenretinide‐induced cell death of neuroblastoma cells are complex, involving signaling pathways mediated by free radicals or reactive oxygen species (ROS). The aim of this study was to identify mechanisms generating ROS and apoptosis of neuroblastoma cells in response to fenretinide. Fenretinide‐induced ROS or apoptosis of SH‐SY5Y or HTLA 230 neuroblastoma cells were not blocked by Nitro l‐argenine methyl ester (l‐NAME), an inhibitor of nitric oxide synthase. Flavoprotein‐dependent superoxide‐producing enzymes such as NADPH oxidase were also not involved in fenretinide‐induced apoptosis or ROS generation. Similarly, ketoconazole, a cytochrome P450 inhibitor, and inhibitors of cyclooxygenase (COX) were also ineffective. In contrast, inhibition of phospholipase A2 or lipoxygenases (LOX) blocked the induction of ROS and apoptosis in response to fenretinide. Using specific inhibitors of LOX, blocking 12‐LOX but not 5‐ or 15‐LOX inhibited both fenretinide‐induced ROS and apoptosis. The effects of eicosatriynoic acid, a specific 12‐LOX inhibitor, were reversed by the addition of the 12‐LOX products, 12 (S)‐hydroperoxyeicosatetraenoic acid and 12 (S)‐hydroxyeicosatetraenoic acid. The targeting of 12‐LOX in neuroblastoma cells may thus be a novel pathway for the development of drugs inducing apoptosis of neuroblastoma with improved tumor specificity. J. Cell. Biochem. 89: 698–708, 2003. © 2003 Wiley‐Liss, Inc.
Active poly(ADPribose) metabolism in DNAase‐ and salt‐resistant rat testis chromatin with high transcriptional activity/competence
Authors: Mennella Maria Rosaria Faraone , Roma Guglielmo , Farina Benedetta 
Published: July 2003

AbstractA chromatin fraction, named pP fraction, was prepared from rat testis nuclei, which had been digested with nuclease in order to separate soluble and insoluble chromatin. This fraction resembled nuclear matrix as it was highly resistant to DNAase digestion, had a high content of proteins compared to the low DNA percentage, and a noticeable transcriptional activity. Moreover, poly(ADPribosyl)ation system (i.e., poly(ADPR)polymerase, poly(ADPribose), and acceptor proteins) was still present at high levels. In order to study whether it might be identified as the protein support surrounding chromatin loops, this pP fraction was further analyzed after 3 M NaCl extraction. The 3 M NaCl extract and the highly insoluble pellet, named Nuclear Matrix Pellet, were characterized as it regards DNA, newly synthesized RNA and proteins. Furthermore, poly(ADPribose) metabolism was analyzed by measuring both poly(ADPribose) polymerase and poly(ADPribose) glycohydrolase activities, poly(ADPribose) distribution and by identifying protein acceptors. The final pellet had features of nuclear matrix containing less than 10% DNA and high percentage of proteins; 28% of newly synthesized RNA was still associated with this fraction. Long and branched polyADPribose were found in the nuclear matrix‐like pellet, although ADPribose acceptors (mainly H1 and core histones) appeared to be modified mostly with short ADPribose oligomers. Longest and branched polymers were retained on the top of protein gel, likely bound to automodified poly(ADPribose) polymerase. J. Cell. Biochem. 89: 688–697, 2003. © 2003 Wiley‐Liss, Inc.
Retinoic acid inhibits chondrogenesis of mesenchymal cells by sustaining expression of N‐cadherin and its associated proteins
Authors: Cho Seo‐Hyun , Oh Chun‐Do , Kim Song‐Ja , Kim Il‐Chul , Chun Jang‐Soo 
Published: July 2003

AbstractRetinoic acid (RA) is a well‐known regulator of chondrocyte phenotype. RA inhibits chondrogenic differentiation of mesenchymal cells and also causes loss of differentiated chondrocyte phenotype. The present study investigated the mechanisms underlying RA regulation of chondrogenesis. RA treatment in chondrifying mesenchymal cells did not affect precartilage condensation, but blocked progression from precartilage condensation to cartilage nodule formation. This inhibitory effect of RA was independent of protein kinase C and extracellular signal‐regulated protein kinase, which are positive and negative regulators of cartilage nodule formation, respectively. The progression from precartilage condensation to cartilage nodule requires downregulation of N‐cadherin expression. However, RA treatment caused sustained expression of N‐cadherin and its associated proteins including α‐ and β‐catenin suggesting that modulation of expression of these molecules is associated with RA‐induced inhibition of chondrogenesis. This hypothesis was supported by the observation that disruption of the actin cytoskeleton by cytochalasin D (CD) blocks RA‐induced sustained expression of cell adhesion molecules and overcomes RA‐induced inhibition of chondrogenesis. Taken together, our results suggest RA inhibits chondrogenesis by stabilizing cell‐to‐cell interactions at the post‐precartilage condensation stage. J. Cell. Biochem. 89: 837–847, 2003. © 2003 Wiley‐Liss, Inc.
Nuclear localization of the Hermes transposase depends on basic amino acid residues at the N‐terminus of the protein
Authors: Michel K. , Atkinson P.W. 
Published: July 2003

AbstractFor the Hermes transposable element to be mobilized in its eukaryotic host, the transposase, encoded by the element, must make contact with its DNA. After synthesis in the cytoplasm, the transposase has to be actively imported into the nucleus because its size of 70.1 kDa prevents passive diffusion through the nuclear pore. Studies in vitro using transient expression of a Hermes–EGFP fusion protein in Drosophila melanogaster Schneider 2 cells showed the transposase was located predominantly in the nucleus. In silico sequence analysis, however, did not reveal any nuclear localization signal (NLS). To identify the sequence(s) responsible for localization of Hermes transposase in the nucleus, truncated or mutated forms of the transposase were examined for their influence on sub‐cellular localization of marker proteins fused to the transposase. Using the same expression system and a GFP–GUS fusion double marker, residues 1–110 were recognized as sufficient, and residues 1–32 as necessary, for nuclear localization. Amino acid K25 greatly facilitated nuclear localization, indicating that at least this basic amino acid plays a significant role in this process. This sequence overlaps the proposed DNA binding region of the Hermes transposase and is not necessarily conserved in all members of the hAT transposable element family. J. Cell. Biochem. 89: 778–790, 2003. © 2003 Wiley‐Liss, Inc.
MPA increases idarubicin‐induced apoptosis in chronic lymphatic leukaemia cells via caspase‐3
Authors: Florio Salvatore , Crispino Luca , Ciarcia Roberto , Vacca Giovanni , Pagnini Ugo , Matteis Andrea de , Pacilio Carmen , D'Andrilli Giuseppina , Kumar Christine , Giordano Antonio 
Published: July 2003

AbstractThe caspase family of protease is speculated to have a crucial role in apoptosis. The effect of treatment with Idarubicin (IDA) and Medroxyprogesterone acetate (MPA), used alone or in combination, on the activation of Caspase‐3 in canine Chronic Lymphatic Leukaemia (CLL) cells was investigated, in order to clarify the mechanism of chemo‐ and hormone‐therapy mediated apoptosis. Caspase activity was determined by a quantitative fluorimetric assay. Apoptosis was monitored by propidium iodide (PI) and nucleosomes assay. Treatment of CLL cells for 24 h with MPA 5 μM did not significantly activate caspase‐3 but its activity was increased almost 5‐fold more with IDA 1 μM (P < 0.05) than control. Treatment of CLL cells with IDA 1 μM in equimolecular association with MPA was able to increase the activation of caspase‐3 induced by IDA of the 61.2% (P < 0.05) in comparison with IDA alone. The activation of caspase‐3 was confirmed evaluating apoptosis by PI and nucleosomes assay. Furthermore, both caspase‐3 activation and apoptosis triggered by IDA alone or in combination with MPA were significantly inhibited by specific caspase‐3 inhibitor AC‐DEVD‐CMK. These findings provide an explanation for IDA and MPA induced‐apoptosis mechanism. J. Cell. Biochem. 89: 747–754, 2003. © 2003 Wiley‐Liss, Inc.
Overexpression of interleukin‐2 receptor α in a human squamous cell carcinoma of the head and neck cell line is associated with increased proliferation, drug resistance, and transforming ability
Authors: Kuhn Deborah J. , Smith David M. , Pross Seth , Whiteside Theresa L. , Dou Q. Ping 
Published: July 2003

AbstractIt has been previously demonstrated that human carcinomas express interleukin‐2 receptor (IL‐2R) α, β, and γ chains. The β and γ chains of IL‐2R have intermediate binding affinity for IL‐2 and are responsible for the intracellular signaling cascades after IL‐2 stimulation. IL‐2Rα lacks the cytoplasmic domain, but is essential for increasing the IL‐2‐binding affinity of other receptors. Overexpression of IL‐2Rα in tumor cells is associated with tumor progression and a poor patient prognosis. To define molecular mechanisms responsible for the effects associated with IL‐2Rα expression, ex vivo experiments were performed with the squamous cell carcinoma head‐and‐neck cancer line, PCI‐13, which was genetically engineered to overexpress the IL‐2Rα chain. While IL‐2Rα‐overexpressing PCI‐13 cells were capable of forming colonies in soft agar, PCI‐13 cells transfected with the control vector or those expressing IL‐2Rγ did not. Consistently, IL‐2Rα‐expressing tumor cells proliferated more rapidly than the control or IL‐2Rγ+ cells, associated with increased levels of cyclins A and D1 and cyclin‐dependent kinase (cdk(s)) 2 and 4 proteins. In addition, IL‐2Rα‐expressing cells were significantly more resistant to apoptosis induction by a tripeptidyl proteasome inhibitor (ALLN) and two chemotherapeutic drugs (VP‐16 and taxol) than the control or IL‐2Rγ+ cells. Accompanying the drug resistance, high levels of anti‐apoptotic Bcl‐XL and Bcl‐2 proteins were found in the mitochondria‐containing fraction of IL‐2Rα‐expressing tumor cells. Treatment of IL‐2Rα‐expressing cells with a specific Janus kinase 3 (Jak3) inhibitor decreased expression of cyclin A, cyclin D1, Bcl‐XL, and Bcl‐2 proteins. Finally, high levels of ubiquitinated proteins were detected in the proliferating IL‐2Rα‐expressing cells. Our data suggest that increased proliferation rates and decreased drug sensitivity of IL‐2Rα‐expressing tumor cells are responsible for the enhanced tumor aggressiveness and poor clinical prognosis of patients whose tumors express IL‐2Rα. J. Cell. Biochem. 89: 824–836, 2003. © 2003 Wiley‐Liss, Inc.
Estrogenic endocrine disruptive components interfere with calcium handling and differentiation of human trophoblast cells
Authors: Derfoul A. , Lin F.J. , Awumey E.M. , Kolodzeski T. , Hall D.J. , Tuan R.S. 
Published: July 2003

AbstractDuring development, calcium (Ca) is actively transported by placental trophoblasts to meet fetal nutritional and the skeletal mineralization needs. Maternal exposure to estrogenic pesticides, such as 1,1‐bis(p‐chlorophenyl)‐2,2,2‐trichloroethane (DDT) and methoxychlor (MTC), has been shown to result in reproductive disorders and/or abnormal fetal development. In this study, we have examined the effects of exposure of trophoblastic cells to MTC and DTT, in comparison to 17β‐estradiol (E2) and diethylstilbestrol (DES), to test the hypothesis that cellular Ca handling is a target for these endocrine disruptive components. Treatment with DDT, MTC, DES, or E2 increased cellular Ca uptake, and the expression of trophoblast‐specific human Ca binding protein (HCaBP) was down‐regulated by both MTC and DDT. Treatment with MTC, DDT, and DES inhibited cell proliferation, induced apoptosis, and suppressed expression of several trophoblast differentiation marker genes. These effects were reversed by overexpression of metallothionein IIa, a gene highly responsive to cadmium and other metals. These results strongly suggest that trophoblast Ca handling functions are endocrinally modulated, and that their alteration by candidate endocrine disruptors, such as MTC and DDT, constitutes a possible pathway of the harmful effects of these components on fetal development. J. Cell. Biochem. 89: 755–770, 2003. © 2003 Wiley‐Liss, Inc.
Monoclonal antibody to fibulin‐1 generated by genetic immunization
Authors: Pupa S.M. , Forti S. , Invernizzi A.M. , Giovanazzi R. , Twal W.O. , Argraves W.S. , Ménard Sylvie 
Published: July 2003

AbstractFibulin‐1 (Fbln‐1) is an extracellular matrix (ECM) and plasma glycoprotein. Considering the growing evidence indicating that Fbln‐1 plays a role in cancer we sought to develop monospecific antibodies to better facilitate further studies of the function of Fbln‐1 in breast cancer. Using a plasmid expression vector encoding full‐length human Fbln‐1D as an immunogen and CpG oligodeoxyribonucleotides as adjuvant a monoclonal antibody (MAb) against Fbln‐1 was produced. This MAb, designated MEM‐2 was of IgM isotype and reacted with bacterially expressed Fbln‐1. Furthermore, MEM‐2 reacted with Fbln‐1 expressed in the ECM released by cultured human breast carcinoma SKBR‐3 cells in ELISA, and also with Fbln‐1 present in SKBR‐3 cell extract in immunoprecipitation and Western blotting. MEM‐2 also reacted with Fbln‐1 in human breast carcinoma specimens. These findings illustrate the utility of genetic immunization as a means of generating monoclonal antibodies to tumor‐related ECM proteins. MEM‐2 represents a useful new tool for the study of Fbln‐1 in breast cancer. J. Cell. Biochem. 89: 647–652, 2003. © 2003 Wiley‐Liss, Inc.