bluuebunny/crossref_metadata_2025 huggingface dataset:
Ovarian follicle cells in torpedo marmorata synthesize vitellogenin
Authors: Marina Prisco , Salvatore Valiante , Maurizio Romano , Loredana Ricchiari , Annamaria Liguoro , Vincenza Laforgia , Ermelinda Limatola , Piero Andreuccetti
Published: February 2004

The pattern of vitellogenesis is similar in all non‐mammalian vertebrates: the liver, under oestrogenic stimulus, synthesizes vitellogenin (VTG) that, via the maternal circulation, is delivered to the oocyte and here internalized by receptor‐mediated endocytosis (Wallace, 1985: Development Biology. A comprehensive synthesis. Vol. 1 Oogenesis:127–177; Schneider, 1996: Int Rev Cytol 166:103–134; LaFleur, 1999: Encyclopedia of Reproduction Vol. 4:985–992). The contribution to vitellogenesis of different components of the ovarian follicle has also been reported in amphibians (Wallace, 1985), squamate reptiles (Ghiara and Limatola, 1980: Acta Embryol Morphol Exper 1:5–6; Andreuccetti, 1992: J Morphol 212:1–11), and recently, supporting previous reports (Chieffi and Pierantoni, 1987: Hormones and Reproduction in Fishes, Amphibians and Reptiles Single vol.:117–144), in Torpedo marmorata (Prisco et al., 2001: Perspective in comparative endocrinology: Unity and diversity Single vol.:1197–1201; Prisco et al., 2002b: Gen Comp Endocrinol 128:171–179). The present investigation, performed with immunoblotting, immunohistochemical, and in situ hybridization techniques during different stages of follicular growth in T. marmorata, shows that, as previously supposed (Prisco et al., 2002b), granulosa cells in both previtellogenic and vitellogenic phases actively synthesize VTG. This is the first time among vertebrates that the synthesis of this protein has been found to occur also within the ovarian follicle. The present data also demonstrate that the contribution of granulosa cells becomes particularly evident during vitellogenesis. Indeed, in vitellogenic follicles, small, intermediate, and pyriform‐like cells cross‐react with an anti‐VTG antibody and are positive to a hybridization signal with a VTG mRNA probe. By contrast, in previtellogenesis only the enlarged cells, i.e., intermediate and pyriform‐like cells, are involved in VTG synthesis. Mol. Reprod. Dev. 67: 424–429, 2004. © 2004 Wiley‐Liss, Inc.
GPI‐anchored aminopeptidase is involved in the acrosome reaction in sperm of the mussel mytilusedulis
Authors: Togo Tatsuru , Morisawa Masaaki
Published: February 2004

The sperm of the mussel Mytilus had hydrolytic activities against substrates for aminopeptidase. Acrosome reaction (AR) was suppressed in the presence of aminopeptidase substrate, Phe‐4‐methylcoumaryl‐7‐amide (MCA), and an aminopeptidase inhibitor, bestatin. Treatment of sperm with phosphatidylinositol‐specific phospholipase C (PI‐PLC) released aminopeptidase activity from sperm and suppressed AR. These results suggest that the enzyme is located on the sperm surface via glycosylphosphatidylinositol (GPI)‐anchor and is involved in the AR. Immunoblot analysis showed that tyrosine residues of 40, 59, 68, and 72 kDa proteins were phosphorylated during induction of the AR. The 40 kDa protein was also recognized by anti‐c‐Src antibody by immunoblotting. The tyrosine phosphorylation of these proteins was inhibited when sperm were inseminated in the presence of Phe‐MCA, and by PI‐PLC treatment. Treatment of sperm with tyrosine kinase activator, 9,10‐dimethyl‐1,2‐benzanthracene, induced AR, and its inhibitor, genistein, suppressed AR. These results suggest that tyrosine phosphorylation of 40, 59, 68, and 72 kDa proteins, induced by the interaction of GPI‐anchored aminopeptidase with oocyte surface, triggers AR in Mytilus sperm. Mol. Reprod. Dev. 67: 465–471, 2004. © 2004 Wiley‐Liss, Inc.
In vitro effect of gamma‐aminobutyric acid on bovine spermatozoa capacitation
Authors: Ritta M.N. , Bas D.E. , Tartaglione C.M.
Published: February 2004

Sperm capacitation is defined as the maturational changes that render a sperm competent for fertilization and occurs in the female reproductive tract. Identification of the factor/s that regulate sperm capacitation would allow the understanding of these phenomena. Among these factors, gamma‐aminobutyric acid (GABA) has recently become as a putative modulator of sperm function. The aim of this study was to explore the presence of a GABAergic regulation of bovine sperm capacitation as well as the possible intracellular mechanisms involved. GABA was detected in fresh semen by a sensitive radioreceptor assay (spermatozoa, 0.064 ± 0.003 nmoles/106 cells; seminal plasma, 23.21 ± 1.16 nmoles/ml). Scatchard analysis of [3H]‐muscimol binding to sperm membranes yielded a linear plot consistent with a single population of binding sites (Kd = 3.87 nM, Bmax = 417 fmol/mg prot.). [3H]‐muscimol specific binding to sperm membranes was significantly inhibited by the GABA A receptor (GABA A‐R) antagonist bicuculline and by the agonists muscimol and isoguvacine. Addition of GABA to the incubation medium resulted in a concentration‐dependent increase in the percentage of capacitated spermatozoa (chlortetracycline assay). We observed a significant increment on intracellular calcium and cyclic 3′,5′ adenosine monophosphate (cAMP) concentrations induced by GABA, being the cation influx abolished when the cell suspensions were coincubated with the antagonists bicuculline or picrotoxin. It is concluded that GABA induces sperm capacitation through an intracellular mechanism dependent on calcium influx and cAMP accumulation mediated by a specific GABA A‐R. Mol. Reprod. Dev. 67: 478–486, 2004. © 2004 Wiley‐Liss, Inc.
Phospholipid hydroperoxide glutathione peroxidase: expression pattern during testicular development in mouse and evolutionary conservation in spermatozoa
Authors: Nayernia Karim , Diaconu Mihaela , Aumüller Gerhard , Wennemuth Gunther , Schwandt Iris , Kleene Kenneth , Kuehn Hartmut , Engel Wolfgang
Published: February 2004

Phospholipid hydroperoxide glutathione peroxidase (PHGPx) is a selenoprotein belonging to the family of glutathione peroxidases and has been implicated in antioxidative defense and spermatogenesis. PHGPx accounts for almost the entire selenium content of mammalian testis. In an attempt to verify the expression pattern of PHGPx, testes of mouse mutants with arrest at different stages of germ cell development and testes of mice at different ages were subjected to immunostaining with a monoclonal anti‐PHGPx antibody. PHGPx was detected in Leydig cells of testes in all developmental stages. In the seminiferous tubuli, the PHGPx staining was first observed in testes of 21‐day‐old mice which is correlated with the appearance of the first spermatids. This result was confirmed when the testes of mutant mice with defined arrest of germ cell development were used. An immunostaining was observed in the seminiferous tubuli of olt/olt and qk/qk mice which show an arrest at spermatid differentiation. In Western blot analysis of proteins extracted from testes of mutant mice and from developing testes, two signals at 19‐ and 22‐kDa were observed which confirm the existence of two PHGPx forms in testicular cells. In mouse spermatozoa, a subcellular localization of PHGPx and sperm mitochondria‐associated cysteine‐rich protein (SMCP) was demonstrated, indicating the localization of PHGPx in mitochondria of spermatozoa midpiece. For verifying the midpiece localization of PHGPx in other species, spermatozoa of Drosophila melanogaster, frog, fish, cock, mouse, rat, pig, bull, and human were used in immunostaining using anti‐PHGPx antibody. A localization of PHGPx was found in the midpiece of spermatozoa in all species examined. In electronmicroscopical analysis, PHGPx signals were found in the mitochondria of midpiece. These results indicate a conserved crucial role of PHGPx during sperm function and male fertility. Mol. Reprod. Dev. 67: 458–464, 2004. © 2004 Wiley‐Liss, Inc.
Expression of epidermal growth factor receptor in the goat cumulus–oocyte complex
Authors: Gall Laurence , Chene Nicole , Dahirel MichÈle , Ruffini Sylvie , Boulesteix Claire
Published: February 2004

It has been previously reported that epidermal growth factor (EGF) influences meiotic maturation and development competence of oocytes in various mammalian species. The present study was undertaken to analyze the expression of the gene encoding the EGF‐receptor (EGF‐R) in the goat cumulus–oocyte complex during meiotic competence acquisition. Expression of EGF‐R mRNA was evaluated by PCR on reverse transcribed mRNA from follicular cells and oocytes, using EGF‐R specific primers designed from human cDNA. The presence of the EGF‐R transcript was evidenced in follicular cells as well as in meiotically competent and incompetent oocytes. Western blot analysis performed with specific anti EGF‐R antibody revealed in meiotically competent and incompetent oocytes and in follicular cells a 170 kD polypeptide corresponding to the goat EGF‐R protein. In oocytes the amount of EGF‐R increased with meiotic competence acquisition. EGF‐R distribution was examined by indirect immunofluorescence on frozen sections of cumulus–oocyte complexes (COCs). EGF‐R immunoreactivity was observed in cumulus cells and in oocytes. Staining appeared to be confined to the periphery of the cells for both oocytes and cumulus cells. In this study, we identified the main component required for signaling via EGF‐R in the goat oocyte and in follicular cells. These results suggest a possible involvement of EGF in the regulation of follicular growth and oocyte maturation in goat. Mol. Reprod. Dev. 67: 439–445, 2004. © 2004 Wiley‐Liss, Inc.
The fragile X mental retardation protein, FMRP, recognizes G‐quartets
Authors: Darnell Jennifer C. , Warren Stephen T. , Darnell Robert B.
Published: February 2004

Fragile X mental retardation is a disease caused by the loss of function of a single RNA‐binding protein, FMRP. Identifying the RNA targets recognized by FMRP is likely to reveal much about its functions in controlling some aspects of memory and behavior. Recent evidence suggests that one of the predominant RNA motifs recognized by the FMRP protein is an intramolecular G‐quartet and that the RGG box of FMRP mediates this interaction. Searching databases of mRNA sequence information, as well as compiled sequences of predicted FMRP targets based on biochemical identification, has revealed that many of these predicted FMRP targets contain intramolecular G‐quartets. Interestingly, many of the G‐quartet containing RNA targets encode proteins involved in neuronal development and synaptic function. Defects in the metabolism of this set of RNAs, presumably in the translation of their protein products, is likely to underlie the behavioral and cognitive changes seen in the disease. MRDD Research Reviews 2004;10:49–52. © 2004 Wiley‐Liss, Inc.
The fragile X syndrome: From molecular genetics to neurobiology
Authors: Willemsen Rob , Oostra Ben A. , Bassell Gary J. , Dictenberg Jason
Published: February 2004

Since the identification of the FMR1 gene basic research has been focused on the molecular characterization of the FMR1 gene product, the fragile X mental retardation protein (FMRP). Recent developments in fragile X research have provided new insights and knowledge about the physiological function of FMRP in the cell and the nerve cell in particular. Currently, compelling evidence suggests a role for FMRP in the transport/translation of dendritically localized mRNAs. In addition, the identification of some of the target mRNAs of FMRP have led to an increased interest in the neurobiology of the syndrome. This review highlights the role of FMRP in dendritic mRNA transport/translation in relation to synaptic plasticity, a molecular mechanism implicated in learning and memory. MRDD Research Reviews 2004;10:60–67. © 2004 Wiley‐Liss, Inc.
The coupled radial–axial deformations analysis of flexible pipes conveying fluid
Authors: Monprapussorn T. , Chucheepsakul S. , Huang T.
Published: February 2004

This paper presents large deformation analysis of pipes conveying fluid in which two complicated behaviours are taken into consideration. The first is the coupling between radial and axial deformations of pipe wall, and the other is the interaction between a deformed pipe and transported fluid having the variable internal flow velocity. The coupled radial–axial deformation theory of the pipes and the continuity theory of flow inside the moving deformed pipes are developed to undertake these coupling behaviours. All strong and weak forms of governing equations are obtained by carrying out the virtual work formulation. The hybrid‐finite element method is used to solve the highly non‐linear static problems, which configure the initial large deflection and large strain conditions of the pipes. The state‐space finite element model for use in analyses of non‐linear vibration and system stability is established as well as the suggested numerical solution procedures. The numerical studies of the pipes under circumstances of intense radial loads such as deep‐water risers demonstrate that even a slight change of the radial deformation has a significant effect in increasing non‐linear responses, and reducing stabilities of the pipes. Copyright © 2004 John Wiley & Sons, Ltd.
A coupling procedure of FE and SBFE for soil–structure interaction in the time domain
Authors: Yann Junyi , Zhang Chuhan , Jin Feng
Published: February 2004

A coupling procedure of finite element (FE) and scaled boundary finite element (SBFE) is presented for three‐dimensional (3D) dynamic analysis of unbounded soil–structure interaction in the time domain. The procedure is implemented with the following efficient techniques: (1) Based on the concepts of linear system theory, the acceleration unit‐impulse response matrix of the unbounded soil calculated by the SBFE method is converted into a group of time‐independent matrices; thus, the time history of the unit‐impulse response function of the unbounded medium can be replaced by an equivalent state‐variable description. (2) The interaction forces between the unbounded soil and the structure are evaluated by a system of linear equations instead of time‐consuming convolution integrals. (3) Since only the partial information of the unit‐impulse response function is sufficient to capture the main character of the unbounded medium and the history of response function can be truncated at the cut‐off time tc, the computational effort spent in the SBFE method can be further reduced. In addition, the accuracy of the procedure can be controlled with prescribed parameters, thus the main advantage of the SBFE method as a highly accurate procedure is retained. Three numerical examples of foundation response demonstrate that good agreement can be achieved when compared with previous results, and high efficiency is also evident compared with the direct convolution integral method. Copyright © 2004 John Wiley & Sons, Ltd.
Effect of haloperidol on the numeric density of neurons and nuclear height in the rat hippocampus: A stereological and histopathological study
Authors: Unal B. , Özbek M.E. , Aydın M.D. , Aydın N. , Bulucu Z. , Vuraler Ö. , Odaci E. , Sahin B. , Kaplan S.
Published: February 2004

In recent studies, some neuroleptics have shown neurotoxic activities. Clinical and experimental studies have been carried out to investigate the effects of different neuroleptic drugs considered to affect the central nervous system. The aim of this study was to evaluate neurotoxic effects of halperidol on hippocampal neurons. The drug was given in daily doses of either 1 or 3 mg/kg for 6 weeks to adult male guinea pigs. After treatment, all animals were anaesthetized via short inhalation of ether, and then were fixed by a mixture of 2% glutaraldehyde and +2% paraformaldehyde in 0.1 M phosphate buffer. Brains were removed from the cranium and stored in the same fixative overnight. On the following day, the CA1 region of the hippocampus was dissected out. After embedding in araldite resin and obtaining semi‐thin sections, the tissues were stained with toluidine blue. The physical dissector was used for measurements of nuclear height and numerical density of neurons and the sections were also evaluated histopathologically. The numerical density of neurons and nuclear height in the hippocampus for the low‐dose (1 mg/kg) and high‐dose (3 mg/kg) experimental groups were 12.4 mm3 and 3.6 μm and 7.14 mm3 and 3.56 μm, respectively. In contrast, the control group had a neuronal numerical density of 16.55 mm −3 and a nuclear height of 4.09 μm. There was a significant difference in both the mean density of neurons and the mean height of nuclei between haloperidol‐treated and control groups (p < 0.05). There was also a statistical difference in the mean density of neurons (but not in nuclear height) when comparing the dosage of haloperidol (p < 0.05). These findings suggest that haloperidol treatment may lead to a loss of neurons as well as a decrease in the height of nuclei in the hippocampus.
Effects of low‐dose oxcarbazepine administration on developing cerebellum in newborn rat: A stereological study
Authors: Odaci E. , Kaplan S. , Sahin B. , Bas O. , Gevrek F. , Aygün D. , Ünal B. , Sönmez O.F. , Colakoglu S. , Bilgiç S.
Published: February 2004

Oxcarbazepine (OXC) is a widely used novel antiepileptic drug that has been available for routine prescription for 10 years. To examine low dose OXC‐induced neurotoxic effects on cerebellar development, we administered 25 mg/kg OXC orally to newborn Wistar rats once a day on postnatal days 2–14. Microscopic processed cerebellar sections of the control and treated groups were examined by volumetric analysis. Volume estimations were obtained using the Cavalieri's principle using a computerized stereological image analyzer (CAST‐GRID). The total volume of the cerebellum, white matter and the various cerebellar layers (except extragranular layer) were significantly increased in the treated animals. These data may provide useful implications for the management of OXC‐induced developmental neurotoxicity in children exposed to OXC during the late fetal period. Our findings suggest that women suffering from epilepsy should be given OXC carefully only at the lowest effective doses during pregnancy.
A long‐term locomotor function and cognition study following middle cerebral artery occlusion in rats
Authors: Jung Hyuk‐Sang , Lee Hyun‐Sam , Shin Jung‐Won , Lee Doo‐Jae , Won Ran , Sohn Nak‐Won
Published: February 2004

A long‐term study of a cerebral ischemia model, considered valuable in developing better treatments of ischemia, was performed to understand the nature of the dysfunctions associated with stroke. This study focused on the behavioral recovery and cognitive learning and memory of rats over a 17‐week period following middle cerebral artery occlusion (MCAO). The present study involved the design of a comprehensive guideline for clinical behavior evaluation. The behavior of ischemic rats was examined over 17 weeks from 4 different aspects: posture & balance tests (4 subtests), reflex tests (6 subtests), muscle‐tone tests (3 subtests), and a foot‐fault test. After the tests, the results were analyzed to reveal the relationship between hemispheric atrophy and functional evaluation scores. Each battery of behavioral tests revealed multiple recovery phases in the time courses. Also, in the passive avoidance test, the MCAO group differed significantly from the sham group until 8 weeks, at which point the latencies to the sham group levels had occurred and after which there was no significant difference until 17 weeks. Thus, the present study provides a valuable protocol in long‐term neuroprotection studies and presents an important profile of motor function that will supplement standard histopathological analysis at a later stage of ischemia.
The effects of niguldipine pretreatment on cerebral vasospasm after subarachnoid haemorrhage in rabbits
Authors: Gürelik Mustafa , Bolat Mehmet , Öztürk Mustafa , Polat Serkan , Göksel Hakan Murat
Published: February 2004

Several different mechanisms contribute to the pathogenesis of cerebral vasospasm after subarachnoid haemorrhage. Whatever these mechanisms are, the vessel contraction is ultimately induced by an increase of free intracellular Ca2+ in the arterial wall. In this study, we investigated the effect on cerebral blood flow of a new dihydropyridine derivative Ca‐channel blocker Niguldipine, in a rabbit experimental cerebral vasospasm model. The cerebral blood flow was significantly decreased in the saline treatment group when compared with control group. Niguldipine pretreatment caused a significant increase in cerebral blood flow. This effect should be investigated further.
Immunophenotypic analysis of inflammatory breast cancers: identification of an ‘inflammatory signature’
Authors: Charafe‐Jauffret Emmanuelle , Tarpin Carole , Bardou Valérie‐Jeanne , Bertucci François , Ginestier Christophe , Braud Anne‐Chantal , Puig Brigitte , Geneix Jeannine , Hassoun Jacques , Birnbaum Daniel , Jacquemier Jocelyne , Viens Patrice
Published: February 2004

Inflammatory breast cancer (IBC) is a rare but very aggressive form of breast cancer. Its definition is based on clinical criteria, but a molecular definition could be useful when data are incomplete or features are missing. Recently, the identification of overexpression of E‐cadherin in IBC has improved understanding of the molecular basis of this disease. Consequently, the aim of this study was to try to determine an immunophenotypic ‘signature’ of IBC. A series of 80 cases of IBC were compared with 552 non‐IBC control cases and a model was elaborated to evaluate the probability of an inflammatory carcinoma being present in any clinical situation. Tissue microarrays (TMAs) were used to determine the immunohistochemical profile of eight proteins including E‐cadherin, EGFR, oestrogen and progesterone receptor (ER and PR), MIB1, ERBB2, MUC1, and P53. All the parameters tested were differentially expressed between IBC and control cases in univariate analysis (p < 0.001). The five variables that were significantly associated with IBC in multivariate analysis were E − cadherin ≥ 300 [HR = 5.64 (2.92–10.87)], ER negative [HR = 3.00 (1.67–5.51)], MIB1 > 20 [HR = 3.54 (1.87–6.71)], MUC1 cytoplasmic staining [HR = 2.72 (1.49–4.96)], and ERBB2 positive 2+ or 3+ [HR = 2.46 (1.26–4.78)]. The probability that a breast cancer with this full phenotype at diagnosis was an IBC was 90.5%. If any one of the five parameters was missing, this probability dropped to 75% and was less than 50% when one, two, or three parameters were present. The 5‐year overall survival (OS) and 5‐year disease‐free survival (DFS) of patients with IBC were not significantly different from those of the non‐IBC control group that expressed four or five parameters (nIBC‐1), but this nIBC‐1 control group had a significantly worse outcome than the non‐IBC control group (nIBC‐2) with only 0–3 parameters (p = 0.0049 for OS and p < 0.0001 for DFS). In conclusion, an immunophenotypic signature was suggested for IBC. This could help to determine the worst cases, independent of clinical criteria. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Emmprin, a cell surface inducer of matrix metalloproteinases (MMPs), is expressed in T‐cell lymphomas
Authors: Nabeshima Kazuki , Suzumiya Junji , Nagano Mitsuyuki , Ohshima Koichi , Toole Bryan P , Tamura Kazuo , Iwasaki Hiroshi , Kikuchi Masahiro
Published: February 2004

Degradation of the extracellular matrix by matrix metalloproteinases (MMPs) is a crucial step in tumour invasion and metastasis. In human carcinomas, tumour cell–fibroblast interactions (TFIs) have been demonstrated to play a role in the up‐regulation of MMP levels in tumours, and emmprin is a surface molecule on tumour cells that stimulates nearby fibroblasts to produce MMP‐1, 2, and 3. T‐cell lymphomas frequently show extranodal organ involvement and skin invasion, but a role for TFIs in their invasion has not been examined in detail. This study investigated TFIs in T‐cell lymphomas with special reference to emmprin expression and MMP production. Immunohistochemically, only germinal centre cells and some histiocytes expressed emmprin in non‐neoplastic lymph nodes (ten cases), while all T‐cell lymphomas [14 cases of adult T‐cell leukaemia/lymphoma (ATLL), six cases of lymphoblastic lymphoma, seven cases of anaplastic large cell lymphoma, and nine cases of angio‐immunoblastic T‐cell lymphoma] expressed emmprin strongly and diffusely. FACS analysis of peripheral blood from normal individuals revealed that small fractions of B‐cells, T‐cells, and monocytes expressed emmprin, whereas emmprin‐expressing T‐cells were much increased in number, and expressed this protein to a higher level, in ATLL patients. In vitro co‐cultures of emmprin‐positive HTLV‐1‐transformed lymphocytes (MT‐2) and emmprin‐negative human fibroblasts enhanced the production of pro‐MMP‐2 (gelatinase A) and active MMP‐2, compared with cultures of either cell type alone. This stimulation was inhibited by an activity‐blocking peptide against emmprin. Moreover, in histopathological sections from patients with ATL skin involvement, MMP‐2 was demonstrated in fibroblasts around infiltrating ATL cells, but not in fibroblasts in non‐diseased areas. In conclusion, emmprin is overexpressed by T‐lymphoma cells, when compared with normal counterparts, and facilitates MMP‐2 production via interactions with fibroblasts, which could play a role in stromal invasion by lymphoma cells. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Increased expression of Met protein is associated with up‐regulation of hypoxia inducible factor‐1 (HIF‐1) in tumour cells in papillary carcinoma of the thyroid
Authors: Scarpino Stefania , Cancellario d'Alena Francesca , Di Napoli Arianna , Pasquini Anna , Marzullo Antonella , Ruco Luigi P
Published: February 2004

Met protein, the high affinity receptor for hepatocyte growth factor (HGF), was highly expressed by the tumour cells of 64 well‐differentiated papillary carcinomas of the thyroid. The p145 mature form and the p170 precursor form of the protein were both isolated from the tumours. Enhanced expression of Met protein was associated with a 9.5 ± 5‐fold increase in MET RNA transcript levels, suggesting increased transcription of the gene. In the same tumours, the levels of RNA transcripts for hypoxia inducible factor‐1 (HIF‐1), a potent stimulator of met gene transcription, were 4.5 ± 3‐fold higher than those present in the surrounding normal thyroid tissues. HIF‐1 is generally induced by hypoxia. Histological features suggestive of a hypoxia were observed in 37 of 50 tumours and included coagulative necrosis, psammoma bodies, cystic changes, intratumoural haemorrhage, and hyalinization of the fibrous stroma. Immunostaining for Met protein was particularly intense in some cells located at the tumour periphery which were characterized by an invasive phenotype. Microdissection of tumour cell nests from the invading front revealed that the levels of RNA transcripts for MET/HIF were higher than in the centre of the tumour in four of nine cases. Taken together, the findings of this study suggest that HIF‐1, perhaps driven by hypoxia, may be one of the factors leading to the increased transcription of met gene in papillary carcinoma and that this event is often more pronounced at the tumour periphery. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Reduced selenium‐binding protein 1 expression is associated with poor outcome in lung adenocarcinomas
Authors: Chen Guoan , Wang Hong , Miller Charles T , Thomas Dafydd G , Gharib Tarek G , Misek David E , Giordano Thomas J , Orringer Mark B , Hanash Samir M , Beer David G
Published: February 2004

The effects of selenium, an essential nutrient with anti‐carcinogenic properties, are mediated by selenium‐binding proteins. The protein expression status of human selenium‐binding protein 1 (SBP1) in human tumours and the exact function of this protein are not known. In this study, quantitative two‐dimensional polyacrylamide gel electrophoresis (2‐D PAGE) was used on 93 lung adenocarcinomas and ten uninvolved lung samples. Two likely isoforms of a 56 kD protein that showed a significantly decreased abundance in lung adenocarcinomas were observed. Tandem mass spectrometry and 2‐D western blot analysis identified these two proteins as human SBP1. Tumour tissue microarrays were utilized to examine the cellular expression patterns of SBP1 using immunohistochemistry. The same tissue samples were examined for SBP1 mRNA expression using oligonucleotide microarrays. Two major SBP1 isoforms were detected, with an acidic isoform (457) being significantly down‐regulated in lung adenocarcinomas compared with normal lung (p = 0.02). Two additional more acidic SBP1 isoforms were only observed in normal lung. SBP1 protein isoforms and SBP1 mRNA levels were significantly decreased in poorly differentiated (versus moderately and well‐differentiated), T2–T4 (versus T1), and bronchus‐derived (versus bronchioloalveolar) tumours. Low levels of SBP1 protein (native form, 460) correlated significantly with poor survival (p = 0.007). The lack of SBP1 expression was not due to gene deletion. Treatment of A549 lung adenocarcinoma cells with the methylation inhibitor 5‐azacytidine did not affect expression of the SBP1 protein. Analysis of the tumour proliferation status using Ki‐67 suggests that down‐regulated expression of SBP1 may reflect increased cell proliferation and decreased differentiation in lung adenocarcinomas. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Western blot analysis of a limited number of cells: a valuable adjunct to proteome analysis of paraffin wax‐embedded, alcohol‐fixed tissue after laser capture microdissection
Authors: Martinet Wim , Abbeloos Vanessa , Van Acker Nathalie , De Meyer Guido RY , Herman Arnold G , Kockx Mark M
Published: February 2004

In recent years, laser capture microdissection (LCM) has been used successfully to obtain distinct populations of cells for subsequent molecular analysis. Because of the limited sample availability and the absence of in vitro amplification steps for proteins, the use of LCM for proteome analysis largely depends on highly sensitive protein detection methods. In this study, a western blot protocol was developed and validated for the detection of β‐actin and the moderately expressed cell death protein caspase‐3 in small numbers of cells. Initially, cultured human U937 monocytes and whole sections of paraffin wax‐embedded, alcohol‐fixed human tonsils were used to optimize protein electrophoresis and western blotting conditions. High‐performance NuPAGE Bis‐Tris gels in combination with high‐quality transfer membranes, optimized antibody concentrations, and a sensitive chemiluminescent substrate provided a strong signal for β‐actin with ∼500 U937 cells. In the same way, procaspase‐3 could be identified with ∼1000 cells. Similar results were obtained with germinal centre cells that were procured from paraffin wax‐embedded, alcohol‐fixed human tonsils by LCM. Treatment of U937 cells with etoposide rapidly induced cell death and allowed the detection of active caspase‐3 with ∼2500 cells (0.8 pg of protein). The findings of this study suggest that western blotting is a valuable adjunct to proteome analysis of LCM procured cells. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Linkage of angiotensin I‐converting enzyme gene insertion/deletion polymorphism to the progression of human prostate cancer
Authors: Medeiros Rui , Vasconcelos André , Costa Sandra , Pinto Daniela , Lobo Francisco , Morais Antonio , Oliveira Jorge , Lopes Carlos
Published: February 2004

Angiotensin‐converting enzyme (ACE) degrades vasodilator kinins and generates angiotensin II (Ang II). It has been reported that ACE is synthesized by the prostate and that the AT‐1 receptor subtype is the predominant prostatic Ang II receptor. A polymorphism in the human ACE gene has been described and the highest levels of circulating and tissue ACE activity are found in carriers of the DD genotype. In the present study, ACE genotypes were determined in 170 patients with prostate cancer and their association with disease progression was analysed. It was found that the DD genotype was present in 31 of 78 (39.8%) patients with advanced disease and in 19 of 82 (23.2%) with localized disease: this difference was statistically significant (OR = 2.18, 95% CI = 1.11–4.03; p = 0.024). Step‐wise logistic regression analysis was used to identify predictive parameters of advanced disease and it was observed that the DD genotype (p = 0.002, OR = 5.4, 95% CI = 1.84–16.06), high‐grade tumour (p < 0.001, OR = 8.04, 95% CI = 3.03–21.33), and high serum PSA (p < 0.001, OR = 10.87, 95% CI = 4.06–29.13) were significantly associated with advanced disease. The results of this study support the hypothesis that genetic factors related to ACE may influence the behaviour of human prostate cancer. Copyright © 2004 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Arzneimittelwechselwirkungen mit H1‐Antihistaminika: Worauf sollte bei der Abgabe von Antihistaminika geachtet werden?
Authors: Haen Ekkehard
Published: February 2004

H1‐Antihistaminika sind im allgemeinen gut verträgliche Substanzen, die auch nur zu wenigen Arzneimittelinteraktionen Anlass geben. Am relevantesten sind die pharmakodynamische Verstärkung der sedierenden Wirkung anderer ZNS‐aktiver Substanzen und der pharmakokinetisch bedingte Anstieg der Wirkstoffkonzentration bei Hemmung des Cytochrom‐P450‐Enzymsystems durch Arznei‐ und Lebensmittel mit der Folge potenziell lebensbedrohlicher Herzrhythmusstörungen. Werden H1‐Antihistaminika zur Behandlung von Allergien eingenommen, so sollten β‐Blocker nur mit großer Vorsicht angewendet werden.
H1‐Antihistaminika‐Medikation: Wie sieht die richtige Pharmazeutische Betreuung aus?
Authors: Teerling Ulrike
Published: February 2004

H1‐Antihistaminika verfügen über das Potenzial, die Lebensqualität von Patienten zu verbessern [31]. Durch die sedierenden Eigenschaften, insbesondere der älteren H1‐Antihistaminika, ergeben sich oftmals schwerwiegende Folgen: So zeigten sich H1‐Antihistaminika sowohl als Ursache von Autounfällen als auch als eine der häufigsten Arzneimittel in Autopsieberichten von Piloten, deren Flugzeug abgestürzt war. Auch wenn Patienten sich nicht schläfrig fühlen, kann die Gehirnfunktion gelegentlich beeinträchtigt sein [32]. Aber auch die anderen diskutierten Neben‐ und Wechselwirkungen machen diese Substanzen zu einer sehr beratungsintensiven Arzneimittelgruppe.

Die Auswahl des jeweils günstigsten H1‐Antihistaminikums sollte unter Berücksichtigung des Zulassungsstatuses (Indikation, Kinder, Schwangere) und der individuellen Erfordernisse des jeweiligen Individuums Patient (Sedierung erwünscht, kardiale Risikofaktoren, Begleitmedikation, Alter, Leber‐ oder Niereninsuffizienz) erfolgen.

Das ideale H1‐Antihistaminikum existiert zur Zeit noch nicht, aber neue Substanzen befinden sich in der klinischen Erprobung [12].

Zur richtigen Zeit, am richtigen Ort mit den richtigen Hinweisen eingenommen stellen sie ohne Frage eine wichtige, sichere Therapieoption bei den verschiedenen, genannten Krankheitsbildern dar.

Dem Fazit von Herrn Prof. Mutschler bleibt auch in diesem Fall nichts hinzuzufügen:

Eine unkritische Empfehlung für die Selbstmedikation ist entschieden abzulehnen! (H1‐Antihistaminika als freiverkäufliche Hypnotika
Ausgezeichnet!
Authors: Dingermann Theo , Zündorf Ilse
Published: February 2004

Gleich zwei pharmazeutische Hochschullehrer wurden Ende 2003 mit hoch‐dotierten Preisen ausgezeichnet: Prof. Dr. Bernd Clement aus Kiel erhielt für sein In‐vitro‐Modell zum Fremdstoffmetabolismus den mit 10.000 Euro dotierten Tierschutzpreis Schleswig‐Holstein und Prof. Dr. Michael Karas, Frankfurt, wurde zusammen mit Prof. Dr. Franz Hillenhamp, Münster, der mit 30.000 Euro dotierte Beckurts‐Preis für ihre revolutionierenden Arbeiten auf dem Gebiet der Massenspektrometrie verliehen.
Kampf der Birkenpollenallergie
Authors: Zündorf Ilse
Published: February 2004

Ca. 100 Millionen Menschen leiden unter einer Birkenpollenallergie. Und immerhin 70 % der Birkenpollen‐Allergiker entwickeln auch Typ‐1‐Hypersensitivitätsreaktionen gegenüber diversen Früchten und Gemüsetypen.
Cadmium ist ein Metalloestrogen
Authors: Götz Regina
Published: February 2004

Cadmium wirkt wie Estrogen — das ist das Ergebnis einer tierexperimentellen Studie, die im Wissenschaftsmagazin Nature Medicine veröffentlicht wurde. Die Studie wirft die Frage nach einer Neubewertung der Toxizität von Cadmium auf.
The inheritance of wealth and the commonwealth: The ideal of paideia in an age of affluence
Authors: Schervish Paul G.
Published: February 2004

The intergenerational transfer of philanthropy involves teaching and learning that enables subsequent generations, like the disciples in all spiritual traditions, to become generative bearers and propagators rather than simple receivers and distributors of the moral citizenship of financial care.
Transforming philanthropy: Generativity, philanthropy, and the reflective practitioner
Authors: Hodge James M.
Published: February 2004

Development professionals exist in the gap between their institutions and their missions and benefactors and their values systems, providing a vital connection. Along with their benefactors, they too can be transformed in the development process and become more and more human through living well‐examined lives.
Paved with good intentions
Authors: Smith David H.
Published: February 2004

Giving takes work, and it is at its best when it is part of an attitude of gratitude. We give philanthropically because we need help in adequately showing that gratitude.
Sweet Substitute: A software tool for in silico fragmentation of peptide‐linked N‐glycans
Authors: Clerens Stefan , Van den Ende Wim , Verhaert Peter , Geenen Lieve , Arckens Lutgarde
Published: February 2004

A software tool, Sweet Substitute, is described, which assists tandem mass spectrometry (MS/MS)‐based glycosylation characterization from within a tryptic digest. The algorithm creates a virtual nanoelectrospray‐quadrupole time‐of‐flight style‐MS/MS spectrum of any user‐defined N‐linked glycan structure. An empirical peak height modeling routine is implemented in the program. By comparing the theoretical MS/MS data with the deconvoluted and deisotoped experimental MS/MS data, the user is able to quickly assess whether a proposed candidate oligosaccharide structure is a plausible one.
Identification of chromosomally encoded membranal polypeptides of Bacillus anthracis by a proteomic analysis: Prevalence of proteins containing S‐layer homology domains
Authors: Chitlaru Theodor , Ariel Naomi , Zvi Anat , Lion Menahem , Velan Baruch , Shafferman Avigdor , Elhanany Eytan
Published: February 2004

Bacillus anthracis is the causative agent of anthrax disease. Improvement of existing anthrax vaccines, which are currently based on the administration of Protective Antigen (the highly immunogenic nontoxic subunit of the bacterial toxin) may entail other bacterial immunogenic elements, part of which are predicted to reside on the surface of bacterial cells. In the present study, membranal proteins extracted from a stationary‐phase culture of a nonvirulent B. anthracis strain, devoid of the native virulence plasmids pXO1 and pXO2, were separated by two‐dimensional electrophoresis (2‐DE) and a characteristic protein map was defined. The proteomic analysis allowed matix‐assisted laser desorption/ionization‐time of flight mass spectrometry‐assisted identification of 86 protein spots which represent the product of 30 individual open reading frames (ORF). Among these, a prevalent class of proteins was the S‐layer proteins (which were found to represent more than 75% of the B. anthracis membranal fraction) and proteins containing S‐layer homology (SLH)‐membranal localization domains. Five novel SLH proteins, previously inferred only from bioinformatic ORF analysis (draft genome sequence), were identified and one was shown to be a highly abundant membranal protein. Western blots of the 2‐DE gels were probed with sera from convalescent rabbits and guinea pigs infected with virulent B. anthracis (Vollum strain). This analysis revealed that B. anthracis immune animals exhibit antibodies against at least 14 distinct membranal proteins present in the 2‐DE map, establishing that these proteins are expressed in vivo and are able to elicit an immune response. The identification of the protein components of the B. anthracis membranal fraction, as well as the establishment of their potential immunogenicity, underscore the strength of the proteomic approach for identifying molecules which may serve for further analysis of immune and protective abilities.
Profiling the activity of G proteins in patient‐derived tissues by rapid affinity‐capture of signal transduction proteins (GRASP)
Authors: Berman David M. , Shih Ie‐Ming , Burke Lorri‐Anne , Veenstra Timothy D. , Zhao Yingming , Conrads Thomas P. , Kwon Sung Won , Hoang Van , Yu Li‐Rong , Zhou Ming , Kurman Robert J. , Petricoin Emanuel F. , Liotta Lance A.
Published: February 2004

The next phase in molecular medicine will require the ability to identify signal transduction events inside a cell, in the biologic context of the disease‐host interface and at a given point in time. New technologies are needed to profile the activity of these signaling pathways in patient tissue rather than cultured cell lines since the tumor‐host microenvironment influences the cellular proteome. We introduce such a technology, rapid affinity capture of signaling proteins (GRASP), to investigate the activity of signaling pathways from patient‐derived carcinomas and benign epithelial surfaces and apply it to studying important signaling events in ovarian carcinoma. During the progression from benign ovarian epithelium to invasive carcinoma, there is loss of repression of Rho A as evidenced by its dissociation from its inhibitor, Rho Guanine Nucleotide Dissociation Inhibitor (RhoGDI). GRASP is more informative than simply profiling transcript or protein levels. Furthermore, GRASP coupled with mass spectrometry allowed us to identify a protein‐binding partner of RhoGDI, demonstrating the power of this technology in the discovery of potentially novel protein‐protein interactions. GRASP represents an advance in the field of proteomics as it detects protein interactions present in cells as they exist in their native tissue microenvironment.
Database search post‐processing by neural network: Advanced facilities for identification of components in protein mixtures using mass spectrometric peptide mapping
Authors: Lokhov Petr G. , Tikhonova Olga V. , Moshkovskii Sergei A. , Goufman Eugene I. , Serebriakova Marina V. , Maksimov Boris I. , Toropyguine Ilya Yu. , Zgoda Victor G. , Govorun Vadim M. , Archakov Alexander I.
Published: February 2004

Database search post‐processing by neural network was employed in peptide mapping experiments. The database search was performed using both the known algorithms and score functions, such as Bayesian, MOWSE, Z‐score, correlations between calculated and actual peptide length fractional abundance, and, in addition, the probability of protein digest pattern in peptide fingerprint, all embedded in locally developed program. The new signal‐processing algorithm based on neural network improves signal‐noise separation and is acceptable for automatic protein identification in mixtures. Its power was tested on Helicobacter pylori protein inventory after preceding protein separation by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE). Increase in protein identification success rate was observed, and about 100 proteins were identified with no need of human participation in database search estimation.
A two‐dimensional liquid‐phase separation method coupled with mass spectrometry for proteomic studies of breast cancer and biomarker identification
Authors: Hamler Rick L. , Zhu Kan , Buchanan Nathan S. , Kreunin Paweena , Kachman Maureen T. , Miller Fred R. , Lubman David M.
Published: February 2004

A two‐dimensional liquid‐phase separation scheme coupled with mass spectrometry (MS) is presented for proteomic analysis of cell lysates from normal and malignant breast epithelial cell lines. Liquid‐phase separations consist of isoelectric focusing as the first dimension and nonporous silica reverse‐phase high‐performance liquid chromatography (NPS‐RP‐HPLC) as the second dimension. Protein quantitation and mass measurement are performed using electrospray ionization‐time of flight MS (ESI‐TOF MS). Proteins are identified by peptide mass fingerprinting using matrix‐assisted laser desorption ionization‐time of flight MS (MALDI‐TOF MS) and MALDI‐quadrupole time of flight (QTOF)‐tandem mass spectrometry (MS/MS). Two pH regions with 50–60 unique proteins in each pH range were chosen for analysis. Mass maps were created that allowed visualization of protein quantitation differences between normal and malignant breast epithelial cells. Of the approximately 110 unique proteins observed from mass mapping experiments over the limited pH range, 40 (36%) were positively identified by peptide mass fingerprinting and assigned to bands in the mass maps. Of these 40 proteins, 22 were more highly expressed in one or more of the malignant cell lines. These proteins represent potential breast cancer biomarkers that could aid in diagnosis, therapy, or drug development.
Photoaptamer arrays applied to multiplexed proteomic analysis
Authors: Bock Chris , Coleman Mike , Collins Brian , Davis Jody , Foulds Glenn , Gold Larry , Greef Chad , Heil Jim , Heilig Joseph S. , Hicke Brian , Nelson Hurst Michele , Husar Gregory M. , Miller Darcey , Ostroff Rachel , Petach Helen , Schneider Dan , Vant‐Hull Barry , Waugh Sheela , Weiss Allison , Wilcox Sheri K. , Zichi Dominic
Published: February 2004

Multiplexed photoaptamer‐based arrays that allow for the simultaneous measurement of multiple proteins of interest in serum samples are described. Since photoaptamers covalently bind to their target analytes before fluorescent signal detection, the arrays can be vigorously washed to remove background proteins, providing the potential for superior signal‐to‐noise ratios and lower limits of quantification in biological matrices. Data are presented here for a 17‐plex photoaptamer array exhibiting limits of detection below 10 fM for several analytes including interleukin‐16, vascular endothelial growth factor, and endostatin and able to measure proteins in 10% serum samples. The assays are simple, scalable, and reproducible. Affinity of the capture reagent is shown to be directly correlated to the limit of detection for the analyte on the array.
Proteomic analysis of selected prognostic factors of breast cancer
Authors: Roberts Kylie , Bhatia Kiron , Stanton Peter , Lord Roger
Published: February 2004

The incidence of breast cancer is on the rise but as yet there is no guaranteed beneficial treatment for many of the sufferers. The treatments specific for breast and other hormone‐sensitive cancers work well at times, however, the population of women that they will benefit is relatively small. Many are limited to surgical, chemotherapy, and radiotherapy options. Here, using two‐dimensional electrophoresis (2‐DE) in conjunction with a silver stain and Western blotting approach, we attempt to locate selected known prognostic markers for breast cancer. With these results, we can exclude these proteins from the future search for potential pharmaceutical targets, using the same techniques. The proteins that were located include the estrogen receptor‐α, β‐casein, cytokeratin 7, calponin and bax. For each protein an estimated Mr and pI was gained. Each protein was found in multiple variants. By locating these proteins the number of unknown proteins found on the 2‐DE gel has been reduced, helping the future search for novel markers that are shown as being differentially expressed between healthy and cancerous tissue samples.