bluuebunny/crossref_metadata_2025_split huggingface dataset:
Thiol oxidation of cell signaling proteins: Controlling an apoptotic equilibrium
Authors: Cross Janet V. , Templeton Dennis J. 
Published: July 2004

AbstractStudies of cell signal transduction have predominantly focused on regulation of protein function by phosphorylation. However, recent efforts have begun to uncover another layer of regulation mediated by direct oxidation of cysteine residues in signaling proteins. Typically induced during signaling responses accompanied by generation of reactive oxygen species, these thiol modifications have a variety of functional consequences for target proteins. Using specific signaling protein targets as examples, we discuss how thiol oxidation generally activates pro‐apoptotic signaling pathways while inhibiting pathways that promote cell survival. We propose a model in which thiol oxidation acts to control the equilibrium between survival and apoptosis, fine tuning cellular responses that play a central role in the apoptotic decision‐making process. We identify areas of focus for future work, including a better understanding of specificity in thiol oxidation events, and a critical need for approaches to examine these modifications under physiologically relevant signaling conditions. © 2004 Wiley‐Liss, Inc.
Simultaneous detection of DsRed2‐tagged and EGFP‐tagged human β‐interferons in the same single cells
Authors: Maruyama Masato , Nishio Teruko , Yoshida Toyokazu , Ishida Chisaki , Ishida Kayo , Watanabe Yoshihiko , Nishikawa Makiya , Takakura Yoshinobu 
Published: July 2004

AbstractThe red fluorescent protein DsRed2 is a useful fusion tag for various proteins, together with the enhanced green fluorescent protein (EGFP). These chromoproteins have spectral properties that allow simultaneous distinctive detection of tagged proteins in the same single cells by dual color imaging. We used them for tagging a secretory protein, human interferon‐β (IFN‐β). Expression plasmids for human IFN‐β tagged with DsRed2 or with EGFP at the carboxyl terminal were constructed and their coexpression was examined in Mardin–Darby canine kidney epithelial cells. Although maturation of DsRed2 for coloration was slow and the color intensity was weak compared with EGFP, low temperature treatment (20°C) allowed DsRed2‐tagged human IFN‐β to be detected in the cells using color imaging. Consequently, the two chimeric proteins were shown to be colocalized in the same single cells by dual color confocal microscopy. This approach will be useful for investigating subcellular localization of not only cell resident proteins but also secretory proteins. © 2004 Wiley‐Liss, Inc.
Effects of phosphorylation by protein kinase CK2 on the human basal components of the RNA polymerase II transcription machinery
Authors: Cabrejos María Eugenia , Allende Catherine C. , Maldonado Edio 
Published: July 2004

AbstractWe have investigated the role of phosphorylation by vertebrate protein kinase CK2 on the activity of the General Transcription Factors TFIIA, TFIIE, TFIIF, and RNAPII. The largest subunits of TFIIA, TFIIE, and TFIIF were phosphorylated by CK2 holoenzyme. Also, RNA polymerase II was phosphorylated by CK2 in the 214,000 and 20,500 daltons subunits. Our results show that phosphorylation of TFIIA, TFIIF, and RNAPII increase the formation of complexes on the TATA box of the Ad‐MLP promoter. Also, phosphorylation of TFIIF increases the formation of transcripts, where as phosphorylation of RNA polymerase II dramatically inhibits transcript formation. Furthermore, we demonstrate that CK2β directly interacts with RNA polymerase II, TFIIA, TFIIF, and TBP. These results strongly suggest that CK2 may play a role in regulating transcription of protein coding genes. © 2004 Wiley‐Liss, Inc.
Multiple functions of BMPs in chondrogenesis
Authors: Yoon Byeong S. , Lyons Karen M. 
Published: July 2004

AbstractThe ability of bone morphogenetic proteins (BMPs) to promote chondrogenesis has been investigated extensively over the past two decades. Although BMPs promote almost every aspect of chondrogenesis, from commitment to terminal differentiation is well known, the mechanisms of BMP action in discrete aspects of endochondral bone formation have only recently begun to be investigated. In this review, we focus on in vivo studies that have identified interactions between BMP signaling pathways and key downstream targets of BMP action in chondrogenesis. We also discuss evidence regarding the potential roles of BMP receptors in mediating distinct aspects of chondrogenesis, and studies investigating the intersection of BMP pathways with other pathways known to coordinate the progression of chondrocytes through the growth plate. These studies indicate that both Smad‐dependent and ‐independent BMP pathways are required for chondrogenesis, and that BMPs exert essential roles via regulation of the Indian hedgehog (IHH)/parathyroid hormone‐related protein (PTHrP) and fibroblast growth factor (FGF) pathways in the growth plate. © 2004 Wiley‐Liss, Inc.
Down‐regulation of p21WAF1 promotes apoptosis in senescent human fibroblasts: Involvement of retinoblastoma protein phosphorylation and delay of cellular aging
Authors: Huang Ying , Corbley Michael J. , Tang Zuoqin , Yang Long , Peng Yong , Zhang Zong Yu , Tong Tan Jun 
Published: July 2004

AbstractIt has been suggested that genes which exercise checkpoint control during cell cycle traverse are equally important to the process of apoptotic cell death. In this study, we show that the key cell cycle regulatory gene p21WAF1 is also involved in the execution of apoptosis. p21WAF1 expression was down‐regulated during NaBu‐induced apoptosis of senescent normal diploid human 2BS fibroblasts. Conversely, when p21WAF1 expression was actively suppressed in 2BS cells by a stably transfected antisense p21WAF1 construct, apoptosis was accelerated and senescence was delayed, as shown by several markers of cell aging. Down‐regulation of p21WAF1 by antisense caused an increase in the phosphorylation and inactivation of pRb. Phosphorylation of pRb was further enhanced upon induction of apoptosis by NaBu. Our results suggest that p21WAF1, acting through the phosphorylation of pRb, regulates whether 2BS cells cease to proliferate and become senescent but resistant to apoptosis, or whether they accelerate proliferation while becoming more susceptible to apoptotic stimuli. © 2004 Wiley‐Liss, Inc.
Bone morphogenetic protein‐2 suppresses collagenase‐3 promoter activity in osteoblasts through aruntdomain factor 2 binding site
Authors: Varghese Samuel , Rydziel Sheila , Canalis Ernesto 
Published: July 2004

AbstractTransforming growth factor‐β (TGFβ) superfamily of growth factors, which include bone morphogenetic proteins (BMPs), have multiple effects in osteoblasts. In this study, we examined the regulation of collagenase‐3 promoter activity by BMP‐2 in osteoblast‐enriched (Ob) cells from fetal rat calvariae. BMP‐2 suppressed the activity of a −2 kb collagenase‐3 promoter/luciferase recombinant in a time‐ and dose‐dependent manner. The BMP‐2 effect on the collagenase‐3 promoter was further tested in several collagenase‐3 promoter deletion constructs and it was narrowed down to a −148 to −94 nucleotide segment of the promoter containing aruntdomain factor 2 (Runx2) site at nucleotide −132 to −126. The effect of BMP‐2 was obliterated in a collagenase‐3 promoter/luciferase construct containing a mutated Runx2 (mRunx2) sequence indicating that the Runx2 site mediates the BMP‐2 response. Electrophoretic mobility shift assays, using nuclear extracts from control and BMP‐2‐treated Ob cells, indicated that the Runx2 protein is a component of the specific DNA–protein complex formed on the Runx2 site and that the BMP‐2 effect may be associated with minor protein modifications rather than major changes in the composition of specific proteins interacting with the Runx2 site. We confirmed that other members of the TGFβ family can down‐regulate the collagenase‐3 promoter by showing that TGFβ1 also suppresses the promoter activity in a time‐ and dose‐dependent manner. In conclusion, this study demonstrates that BMP‐2 and TGFβ1 suppress collagenase‐3 promoter activity in osteoblasts and establishes a link between BMP‐2 action and collagenase‐3 expression via Runx2, a major regulator of osteoblast formation and function. © 2004 Wiley‐Liss, Inc.
KCNQ1/KCNE1 channels during germ‐cell differentiation in the rat: Expression associated with testis pathologies
Authors: Tsevi Irini , Vicente Rubén , Grande Maribel , López‐Iglesias Carmen , Figueras Agnès , Capellà Gabriel , Condom Enric , Felipe Antonio 
Published: August 2004

AbstractKCNQ1/KCNE1 channels are responsible for the Jervell–Lange–Nielsen cardiac syndrome, which is also characterized by congenital deafness. KCNQ1/KCNE1 is crucial for K+ transport in the inner ear. We show that KCNQ1 and KCNE1 are associated in testis and that their expression is closely regulated during development. Both genes were expressed in undifferentiated germ cells in 21‐day‐old rats and mostly confined to basal immature germ cells in adulthood. Leydig and Sertoli cells were negative. KCNQ1 and KCNE1 were also studied in various germ‐cell pathologies. First, in spontaneous unilateral rat testis atrophy, hematoxylin–eosin analysis revealed massive germ‐cell aplasia with only Sertoli cells and groups of interstitial Leydig cells. In these samples, KCNQ1 and KCNE1 were not expressed. In human seminoma samples characterized by a proliferation of undifferentiated germ cells, KCNQ1/KCNE1 protein levels were higher than in healthy samples. Our results demonstrate that the expression of KCNQ1 and KCNE1 is associated with early stages of spermatogenesis and with the presence of undifferentiated healthy or neoplastic germ cells. The presence of a K+ rich‐fluid in the seminiferous tubule suggests that KCNQ1/KCNE1 is involved in K+ transport, probably during germ‐cell development. © 2004 Wiley‐Liss, Inc.
Key role of the achievement of an appropriate ribosomal RNA complement for G1‐S phase transition in H4‐II‐E‐C3 rat hepatoma cells
Authors: Derenzini Massimo , Montanaro Lorenzo , Chillà Alessandra , Tosti Elena , Vici Manuela , Barbieri Stefania , Govoni Marzia , Mazzini Giuliano , Treré Davide 
Published: August 2004

AbstractCell growth is closely related to cell proliferation and an adequate ribosome biogenesis appears to be necessary for cell duplication. In the present study, we have investigated the relationship between rRNA synthesis and cell cycle progression. For this purpose, in a first set of experiments, we evaluated the effect of rRNA synthesis variation on cycle duration in asynchronously growing H4‐II‐E‐C3 rat hepatoma cells. Cells were either treated with insulin or insulin plus actinomycin D (AMD). The hormone stimulated ribosome biogenesis, which was later followed by an increased synthesis of DNA and a shortening of cell doubling time (DT). Bivariate flow cytometry indicated that the reduced length of the cell cycle was mainly due to the shorter G1‐phase. AMD, at the concentration of 0.04 μg/ml, hindered ribosome biogenesis without affecting heterogeneous RNA production. A 12‐h reduction in ribosome biogenesis level by AMD caused a lowering of DNA synthesis and a lengthening of cell DT with a longer G1‐phase. In a second set of experiments, we analyzed the cell content variations of 28S and 18S rRNA transcripts during G1 phase in H4‐II‐E‐C3 cells, synchronized by serum deprivation, and then stimulated by serum, serum plus insulin, and serum plus insulin and AMD. In control cells, a progressive increase in rRNA content occurred until the highest value of rRNA content was reached 21 h after serum stimulation. In insulin‐treated cells, the highest rRNA value was reached at 12 h whereas in AMD‐treated cells, the rRNA quantity was constantly low until 18 h and then sharply increased at 21 h. In the three experimental conditions, the highest values of rRNA amount were reached at the end of G1 phase and were quite similar to one another. We also evaluated, by real‐time RT‐PCR, cyclin E mRNA expression, which appeared to sharply increase at those times in which the maximum increase in the rRNA content was observed. Our results indicated that the achievement of an appropriate amount of rRNA allows G1/S phase transition, probably by modulating the expression of cyclin E mRNA. © 2004 Wiley‐Liss, Inc.
Differences in X‐ray absorption due to cadmium treatment in Saponaria officinalis leaves
Authors: Reale Lucia , Lai Antonia , Tucci Adele , Poma Anna , Faenov Anatoly , Pikuz Tania , Flora Francesco , Spano Laura , Limongi Tania , Palladino Libero , Ritucci Antonio , Tomassetti Giuseppe , Petrocelli Giovanni , Francucci Massimo , Martellucci Sergio 
Published: July 2004

AbstractA method for detecting cadmium uptake in leaves of Saponaria officinalis doped with a solution of cadmium acetate is described. The technique based on the exposure of dried leaves to X‐rays of a wavelength close to that of the metal K‐edge could be useful for phytoremediation studies as it could reveal the bioaccumulation in plants due to the treatment either in vivo or in vitro with heavy metals. X‐ray microradiography measurements are in agreement with those from peroxidase enzyme assay utilized to follow the oxidative damage induced by heavy metals. At present, as we will see in this report, microradiography has still poorer sensitivity in comparison with enzyme assay, but it has the advantage of being faster, not destructive, and usable even at very high doping levels, where the enzyme assay technique results are fully saturated. Further analysis of the optical density values could lead to a quantitative measurement of the heavy metal in the sample. Thus, the technology developed in this article could be useful for tracing the intake in phytoremediation studies. Microsc. Res. Tech. 64:21–29, 2004. © 2004 Wiley‐Liss, Inc.
3‐D morphological characterization of the liver parenchyma by atomic force microscopy and by scanning electron microscopy
Authors: Melling Mahmoud , Karimian–Teherani Daniela , Mostler Sascha , Behnam Mark , Hochmeister Sonja 
Published: July 2004

AbstractA comparative study of atomic force microscopy (AFM) and scanning electron microscopy (SEM) imaging of the healthy human liver parenchyma was carried out to determine the similarities and the differences. In this study, we compared the fine hepatic structures as observed by SEM and AFM. Although AFM revealed such typical hepatic structures as bile canaliculi and hepatocytes, it also showed the location of the nucleus and chromatin granules in rough relief structure, which was not visible by SEM. By contrast, SEM visualized other structures, such as microvilli, the central vein, and collagenous fibers, none of which was visualized by AFM. For better orientation and confirmation of most of the structures imaged by SEM and AFM, Congo Red–stained specimens were also examined. Amyloid deposits in the Disse's spaces were shown especially clearly in these images. The differences between the SEM and AFM images reflected the characteristics of the detection systems and methods used for sample preparation. Our results reveal that more detailed information on hepatic morphology is obtained by exploiting the advantages of both SEM and AFM. Microsc. Res. Tech. 64:1–9, 2004. © 2004 Wiley‐Liss, Inc.
Glycogen autophagy
Authors: Kotoulas Othon B. , Kalamidas Stefanos A. , Kondomerkos Dimitrios J. 
Published: July 2004

AbstractGlycogen autophagy, which includes the sequestration and degradation of cell glycogen in the autophagic vacuoles, is a selective process under conditions of demand for the massive hepatic production of glucose, as in the postnatal period. It represents a link between autophagy and glycogen metabolism. The formation of autophagic vacuoles in the hepatocytes of newborn animals is spatially and biochemically related to the degradation of cell glycogen. Many molecular elements and signaling pathways including the cyclic AMP/cyclic AMP‐dependent protein kinase and the phosphoinositides/TOR pathways are implicated in the control of this process. These two pathways may converge on the same target to regulate glycogen autophagy. Microsc. Res. Tech. 64:10–20, 2004. © 2004 Wiley‐Liss, Inc.
Healthy and pre‐eclamptic placental basal plate lining cells: Quantitative comparisons based on confocal laser scanning microscopy
Authors: Smith R.K. , Ockleford C.D. , Byrne S. , Bosio P. , Sanders R. 
Published: July 2004

AbstractImmunocytochemical confocal laser scanning microscope images of the monolayer of cells lining the intervillus space at the basal plate of term placentae were analysed using stereology. Immunoreactively‐distinct regions of this mosaic layer were measured. In basal plate from healthy pregnancies, trophoblast epithelium occupied 18.91% of the surface area and endothelium 60.81%. In pre‐eclampsia the equivalent areas were 15.57% and 67.63%. Acellular fibrinoid covers the remaining area and this component decreases in area in pre‐eclampsia. The statistically significant increase in the cellular endothelial compartment may be relevant to the hypertensive pathology of pre‐eclampsia as endothelial signalling plays a major role in regulation of blood pressure. Microsc. Res. Tech. 64:54–62, 2004. © 2004 Wiley‐Liss, Inc.
Confocal laser scanning microscope study of cytokeratin immunofluorescence differences between villous and extravillous trophoblast: Cytokeratin downregulation in pre‐eclampsia
Authors: Ockleford C.D. , Smith R.K. , Byrne S. , Sanders R. , Bosio P. 
Published: July 2004

AbstractPre‐eclampsia is a disease characterized by failures in interstitial implantation. One product of the implantation process is the basal plate; a structure whose complexity makes it hard to fully appreciate the pathological changes in significant diseases of pregnancy. This article describes our use of CLSM immunofluorescence to examine the cytokeratin composition of the cells of trophoblastic origin in the term placental basal plate. Large differences in the content of the structural polymeric protein were compared using analysis of digital images. We show that greater pancytokeratin immunofluorescence is observed in extravillous cytotrophoblast cells as compared with villous trophoblast. There is a >30‐fold difference in the mean area percent of the most intensely immunofluorescent pixels in the tissue containing these cells. This is a very high, statistically significant difference as defined by the Wilcoxon Signed Ranks Test Asym. Sig. (two‐tailed): P < 0.001. The most invasive population of cells of the trophoblast lineage (the extravillous trophoblast) exhibits a significant reduction in cytokeratin immunofluorescence when comparisons of healthy and pre‐eclamptic pregnancies are made. This ratio was 2.4:1. It was tested using the Mann‐Whitney U‐test. From healthy to pre‐eclamptic the reduction was from mean rank 83.42(healthy) to 51.13(pre‐eclamptic). The difference was very highly statistically significant (n = 53 + 75 = 128; U = 984.500; Z = –4.852; P < 0.001). There was also less cytokeratin‐related immunofluorescence in villous trophoblast when healthy villi were compared with pre‐eclamptic villi. The observed alterations in trophoblastic cytoskeletal components are expected to damage the anchorage and motility of cells. The extravillous trophoblast is known to be necessary for implantation. This leads to a cellular hypothesis of the failure of implantation resulting in reduced depth of uterine invasion and failure to adapt the spiral arterioles for low‐pressure perfusion of the intervillus space, two well‐known features of pre‐eclampsia. The reduction in cytokeratin‐related immunofluorescence in the villus trophoblast seen on comparing healthy term placentae with those from pre‐eclamptics implies that the trophoblast is a weaker epithelial layer in the hypertensive pregnancy. This could account for the rise in deported trophoblast associated with pre‐eclampsia. Deported trophoblast has been invoked as the systemic messenger that leads to generalized maternal hypertension seen in this condition. Microsc. Res. Tech. 64:43–53, 2004. © 2004 Wiley‐Liss, Inc.
Expression of brain natriuretic peptide in the rat heart studies during heart growth and in relation to sympathectomy
Authors: Hansson Magnus , Forsgren Sture 
Published: July 2004

AbstractBrain natriuretic peptide (BNP) might be of importance during heart development and is described to be increasingly expressed in congestive heart failure and to affect the progress of this condition. However, details in the normal expression of BNP are still unclear in various parts of the adult and growing heart, including the conduction system. In this study, we investigated the expression of BNP in relation to that of atrial natriuretic peptide (ANP) in the growing as well as in the adult rat heart. The effects of chemical sympathectomy in adult rats were also examined. Contrary to previous BNP immunohistochemical studies, the BNP antiserum was preabsorbed with an excess of ANP before staining to abolish the crossreactivity with ANP. There was a pronounced BNP immunoreaction in the auricles, the trabeculated ventricular walls, and the peripheral parts of the conduction system at 0–1 days postnatally. The degree of immunoreaction gradually decreased with increasing age. A similar developmental pattern was seen concerning ANP expression, but the magnitude of the latter clearly exceeded that for BNP. Immunoreaction for BNP was never detected in the atrioventricular (AV) node and AV bundle at any stage. In contrast to the situation for ANP previously observed, no obvious changes in BNP immunoreaction patterns were observed in response to sympathectomy. This is the first study to thoroughly demonstrate the expression of BNP in the various regions of the rat heart during growth and in the normal and sympathectomized adult stage. The observations are related to possible functions of natriuretic peptides in the growing and adult heart. Microsc. Res. Tech. 64:30–42, 2004. © 2004 Wiley‐Liss, Inc.
Ultrastructural study of apoptotic U7 cells treated with different pro‐apoptotic substances
Authors: Abbro Luigi , Lanubile Remigio , Dini Luciana 
Published: July 2004

AbstractHuman monocytic leukemia U937 cells readily undergo apoptosis when exposed to various stimuli, including inhibition of protein synthesis, oxidative stress, antitumoral agents, etc. The sequential, step‐by‐step morphological changes in U937 cells that occur during the apoptotic program are largely determined by the activation of a specific class of proteases, the caspases. The action of these proteases were followed at the ultrastructural level. From our observations 1) no unique morphological feature exists during apoptosis, even in the same cell type; 2) the extent of the morphological modifications are inducer‐ and dose‐dependent; 3) double or triple treatments amplify the morphological modifications with a single inducer, but not the rate of apoptosis; and 4) in the case of double treatment the second inducer has to have a cytoplasmatic target because damage to the cytoplasm occurs before nuclear modifications become visible. These data should facilitate a more objective evaluation of apoptosis in conditions where antiproliferative drugs, like antiblastic or immunosuppressive molecules, are used to monitor the efficiency of treatment. Microsc. Res. Tech. 64:77–85, 2004. © 2004 Wiley‐Liss, Inc.