bluuebunny/crossref_metadata_2025 huggingface dataset:
Governance for human development: the role of external partners
Authors: Cheema G. Shabbir , Maguire Linda
Published: January 2005

There is a positive correlation between democratic governance and human development. Democratic governance is governance infused with the principles of, inter alia, equity, participation, rule of law, transparency and accountability. Human development is at its very core the expansion of people's choices and their increased participation in decisions that affect their lives. Therefore democratic governance reinforces human development, and human development reinforces democratic governance. External partners—those actors outside of the national development process—can positively contribute to a country's transition to and consolidation of democratic governance (and, thereby, its human development) in a number of ways. External partners act through democratic governance entry points to deliver strategic services and use indicators to measure their relative progress. Indicators can be valuable tools in informing external partners and recipients of assistance of what works, what does not work and why. Copyright © 2001 John Wiley & Sons, Ltd.
Polyaniline and its modification for electroresponsive material under applied electric fields
Authors: Cho M. S. , Kim J. W. , Choi H. J. , Jhon M. S.
Published: January 2005

Polyaniline (PANI) and its various copolymers were synthesized using different monomers such as o‐methyl‐, o‐methoxy‐, o‐ethyl‐, o‐ethoxyaniline and sodium diphenylamine sulfonate by the chemical oxidation polymerization as air‐stable organic conducting polymers, and adopted as one of the most potential materials of electrorheological (ER) fluids, especially for the anhydrous system. A relatively low density, a controllable conductivity, and thermal stability are advantages of the PANI based ER system compared with other ER materials. An FT‐IR analysis was adopted to confirm their synthesis and a scanning electron microscopy (SEM) analysis indicated the shape of PANI derivatives was irregular. The effect of functional group of PANI on electric and electroresponsive properties of poly(aniline‐co‐ethoxyaniline) (COPA) particles were examined. A universal scaling equation of the yield stress was applied to these ER fluids and it was found that all data were collapsed successfully onto a single curve regardless of monomer type of the polyaniline synthesized. Copyright © 2005 John Wiley & Sons, Ltd.
Reversible control of micellization induced by hydrogen bond crosslinking for a nonamphiphilic diblock copolymer with an α,ω‐diamine
Authors: Yoshida Eri , Ohta Masayo , Terada Yoshikazu
Published: January 2005

The micelle formation in a non‐selective solvent has great advantages in molecular designing. It has already been revealed that a poly(vinylphenol)‐block‐polystyrene diblock copolymer (PVPh‐b‐PSt) formed micelles in the presence of α,ω‐diamine in 1,4‐dioxane, a non‐selective solvent through hydrogen bond crosslinking. In this article it is reported that the dissociation‐reconstruction of the PVPh‐b‐PSt micelles with 1,4‐butanediamine (BDA) is reversibly controlled by the variation in temperature, the copolymer concentration, and the diamine concentration. Light scattering studies demonstrated that the micellization of PVPh‐b‐PSt with Mn(PVPh‐b‐PSt) of 10K‐b‐120K occurred at a higher BDA concentration and produced micelles with a lower aggregation number as the temperature increased. The micelles were thermally unstable and dissociated into unimers by increasing the temperature, however, the micelles were completely reconstructed by decreasing the temperature. This dissociation‐reconstruction of the micelles was perfectly controlled by the variation in temperature. An increase in the BDA concentration prevented the dissociation of the micelles but promoted the reconstruction. The increase in the BDA concentration causes an increase in the crosslinking points for hydrogen bonding in the cores of the micelles, thus preventing the dissociation of the micelles by the strong hydrogen bond crosslinking. Copyright © 2005 John Wiley & Sons, Ltd.
Recent developments in polymeric phase change materials for energy storage: poly(ethylene oxide)/stearic acid blends
Authors: Pielichowski Krzysztof , Flejtuch Kinga
Published: January 2005

The thermal behavior of a series of poly(ethylene oxide) (PEO)/stearic acid blends was investigated by means of differential scanning calorimetry (DSC), modulated temperature DSC, Fourier transform infrared spectroscopy (FT‐IR) and scanning electron microscopy (SEM) for application as thermal energy storage materials. It has been found that PEO/stearic acid blends are efficient polymer‐based phase change materials that can be applied in the temperature range 55.7–75.2°C with the enthalpy of phase transition values in the range of 202.8–252.3 J/g. The PEO/stearic acid blend (1:3 w/w) exhibits a senergistic effect—its value of enthalpy of melting and crystallization is higher than those of pure components. The origin of this promising effect was investigated by FT‐IR and SEM techniques to ascertain the influence of hydrogen bonds on the sample's supramolecular morphology. Copyright © 2005 John Wiley & Sons, Ltd.
Asymmetric PS‐block‐(PS‐co‐PB)‐block‐PS block copolymers: morphology and deformation behavior of star block copolymer/PS blends
Authors: Adhikari R. , Buschnakowski M. , Lebek W. , Godehardt R. , Michler G. H. , Calleja F. J. Baltá , Knoll K.
Published: January 2005

The morphology and deformation behavior of blends comprising an asymmetric styrene/butadiene star block copolymer and polystyrene homopolymer (hPS) were investigated by means of transmission electron microscopy (TEM) and uniaxial tensile testing. The hPS samples used had a wide range of molecular weights. In spite of the high polystyrene (PS) content, the star block copolymer exhibited a morphology consisting apparently of a rubbery matrix, attributable to its asymmetric architecture and presence of polystyrene/polybutadiene random copolymer (PS‐co‐PB) as soft rubbery blocks. In the blends, depending on the molecular weight and modality of the added hPS and on the composition of the mixtures, different morphologies (ranging from microphase‐separated and macrophase‐separated ones to a combination of both of them) were observed. Consequently, it was possible to tailor the tensile mechanical properties based on the nature of the nanoscopic morphology of the star block copolymer/PS blends. Copyright © 2005 John Wiley & Sons, Ltd.
Morphology and micromechanical properties of ethylene/1‐octene copolymers and their blends with high density polyethylene
Authors: Adhikari R. , Godehardt R. , Lebek W. , Frangov S. , Michler G. H. , Radusch H.‐J. , Calleja F. J. Baltá
Published: January 2005

The relationship between morphology and deformation behavior of selected ethylene/1‐octene copolymers (EOCs) and their blends with high density polyethylene (HDPE) was investigated. The copolymers showed, depending on the 1‐octene content, different morphologies ranging from lamellar to worm‐like crystalline domains. The binary HDPE/EOC blends studied, which showed well phase‐separated structures consistent with individual melting and crystallization behavior of the blend components, were characterized by a wide range of mechanical and micromechanical properties. The study of strain induced structural changes in an HDPE/EOC blend revealed that at large strains, the extensive stretching of the soft EOC phase is accompanied by rotation of lamellar stack along the strain axis and subsequent fragmentation of the crystals forming beaded‐string‐like structures. A significant depression in microhardness was observed in the copolymers. In their blends with HDPE, a deviation in microhardness behavior from the additivity law was observed. Copyright © 2005 John Wiley & Sons, Ltd.
Effect of gamma irradiation on HDPE/HA (80:20) composites
Authors: Albano C. , Karam A. , González G. , Domínguez N. , Sánchez Y. , Manzo F. , Guzmán‐García C.
Published: January 2005

Effect of gamma irradiation on high density polyethylene/hydroxyapatite (HDPE/HA) composites prepared by solution was studied. Melt flow index (MFI) showed a strong increase with radiation due to some decalin remnant. The evaluation of the mechanical behavior showed an increase in the Young's modulus and resistance at break values attributed to the HA addition. Elongation at break decrease strongly. Radiation did not produce any significant changes on crystallization temperatures. Crystallinity of the polymer decreases when HA is added. The activation energy values allowed for the estimation of the optimum radiation dose. Copyright © 2005 John Wiley & Sons, Ltd.
Oncogene lineages of human papillomavirus type 16 E6, E7 and E5 in preinvasive and invasive cervical squamous cell carcinoma
Authors: Hu Xinrong , Pang Tianyun , Guo Zhongmin , Pontén Jan , Nistér Monica , Bernard Afink Gijs
Published: January 2005

Human papillomavirus (HPV)16 accounts for about 60% of the HPV infections in invasive cervical cancer (ICC). There are many sequence variations within HPV16, some of which have been associated with different biological properties, although no definite correlations have yet been established. However, the definition ‘variant’ has been a source of confusion in research and diagnosis, since it is based on all sequence deviations from a randomly selected prototype. This study has sequenced the HPV16 oncogenes E6, E7 and E5 from 61 Swedish cases with cervical intraepithelial neoplasia grade III (CIN III) or ICC. Clustering the sequence variations at the three common sites of variation (nucleotide 350 in E6, which has previously been associated with the progression from CIN III to ICC, and nucleotides 3979 and 4042 in E5) resulted in the distinction of three major oncogene lineages encompassing more than 95% of the cases, and two minor oncogene lineages. Simple comparison of the distribution of the individual variations or oncogene lineages between CIN III and ICC showed no significant difference, but the number of variations in addition to the three common ones was significantly higher in ICC. This novel classification scheme, based on the variations in the E6, E7 and E5 region, is considered to be a major improvement over the classical ‘prototype‐variant’ classification, and can help to clarify the interpretation of HPV sequence data in relation to the progression of cervical cancer. Copyright © 2001 John Wiley & Sons, Ltd.
p53 over‐expression identifies a subset of nodal peripheral T‐cell lymphomas with a distinctive biological profile and poor clinical outcome
Authors: Pescarmona Edoardo , Pignoloni Patrizia , Puopolo Maria , Martelli Maurizio , Addesso Maria , Guglielmi Cesare , Baroni Carlo D.
Published: January 2005

Peripheral T‐cell lymphomas (PTCL) are usually characterized by aggressive clinical behaviour and poor clinical outcome, but their biological background has not been extensively investigated to date, due to their low incidence, about 10% of all non‐Hodgkin's lymphoma cases in Western countries, and also to the paucity of specific molecular‐genetic abnormalities. Neverthless, there is increasing biological and clinical evidence that primary nodal PTCL should be considered separately from extra‐nodal cases, but little is known about biological factors of possible clinical and prognostic impact. This immunohistochemical study has analysed the expression of p53, Mdm2, p21WAF1, BCL‐2 and p‐glycoprotein (MDR‐1 gene product) in a series of 45 cases of nodal peripheral T‐cell lymphomas (PTCL) with ‘high‐grade’ histology. The immunohistochemical findings were then correlated with proliferative activity and clinical outcome. p53 was over‐expressed in 13 cases (28.9%). p53 positive cases showed significantly higher proliferative activity (p<0.01), more frequent expression of Bcl‐2 (p<0.01) and less frequent expression of p21WAF1 than p53 negative cases. Mdm2 and p‐glycoprotein were expressed in 4/13 (30.8%) and 8/13 (61.5%) p53 positive cases respectively, and in none (0%) of the p53 negative cases (p<0.01). Analysis of the survival curves showed that p53 positive cases were associated with a significantly poorer clinical outcome than p53 negative cases, in terms of both overall survival (p=0.0032) and event‐free survival (p=0.0004). Furthermore, multivariate analysis showed that p53 expression was the most important independent prognostic variable. These findings indicate that p53 over‐expression identifies a subset of nodal PTCL cases with a distinctive biological profile (higher proliferative activity, less frequent expression of p21WAF1 and more frequent expression of Bcl‐2, Mdm2 and p‐glycoprotein than p53 negative cases) and poor clinical outcome. The immunohistochemical analysis of p53 expression is a simple, rapid and low‐cost method which may provide information of potential clinical and prognostic value in nodal peripheral T‐cell lymphomas. Copyright © 2001 John Wiley & Sons, Ltd.
Decline in the expression of the serine proteinase inhibitor maspin is associated with tumour progression in ductal carcinomas of the breast
Authors: Maass Nicolai , Teffner Martin , Rösel Frank , Pawaresch Reza , Jonat Walter , Nagasaki Koichi , Rudolph Pierre
Published: January 2005

Maspin is an inhibitor of serine proteinases with tumour suppressor activity. Its expression appears to be reduced in advanced stages of breast cancer. A large series of archival breast tissue specimens has been examined, including normal glands (n=7), fibrocystic change (n=22), ductal carcinoma in situ (DCIS, n=12), infiltrating carcinomas (n=128) and their lymph node metastases (n=65), using a specific monoclonal antibody. Myoepithelium invariably showed strong maspin expression. In epithelial cells, the strongest expression was found in normal breast and fibrocystic change. A significant stepwise decrease in maspin expression (p<0.0001) occurred in the sequence DCIS – invasive cancer – lymph node metastasis. However, a subset of infiltrating carcinomas showed strong maspin expression, significantly associated with a lower rate of lymph node metastasis at the time of diagnosis (p<0.01). This was independent of tumour size and grade. The in vivo observations presented here are in keeping with data obtained in prior in vitro experiments. Maspin emerges as an indicator of tumour progression and metastatic potential, and might be exploited to predict breast cancer prognosis. According to in vitro data, its tumour suppressor activity is likely to involve both the modulation of cell motility/invasiveness and the inhibition of angiogenesis. Copyright © 2001 John Wiley & Sons, Ltd.
Heavy metals of relevance to human health induce genomic instability
Authors: Coen Natasha , Mothersill Carmel , Kadhim Munira , Wright E. G.
Published: January 2005

Heavy metals used in medical prostheses or present in water supplies or tobacco can build up in tissues and blood and are well known to produce toxic effects. Normally, legislative controls on the levels of these substances are determined by reference to the acute toxicity data. This paper shows that cadmium and nickel can produce delayed effects in human cells in vitro, which are characteristic of genomic instability. The effects occur even at levels where no acute toxic effects can be demonstrated. Genomic instability can be demonstrated by persistent induction of cytogenetic abnormalities and delayed cell death in progeny of cells many generations after exposure. Formerly, this syndrome has only been definitively proven to occur following irradiation, but in these experiments cell populations exposed for only 1 or 24 hours were expanded over several months, involving eight passages, and the yield of chromosomal aberrations and cell loss due to lethal mutations did not decrease. The consequences of this genomic instability are not yet known but it is possible that many of the systemic symptoms associated with exposure to low concentrations of these metals could involve delayed expression of cellular damage. It is also clear that these effects cannot be predicted from acute toxicity data. Copyright © 2001 John Wiley & Sons, Ltd.
Differential expression of the chromosome 11 mucin genes in colorectal cancer
Authors: Sylvester Paul A. , Myerscough Neil , Warren Bryan F. , Carlstedt Ingemar , Corfield Anthony P. , Durdey Paul , Thomas Michael G.
Published: January 2005

The four secretory mucin genes clustered on chromosome 11, MUC2, MUC5AC, MUC5B and MUC6, were screened in 37 patients with cancers in the left hemi‐colon or rectum and 10 normal rectal controls. The mucin genes were detected by in situ hybridization using oligonucleotide probes to the variable number tandem repeat (VNTR) sequences, while the proteins were stained with non‐VNTR (MUC2, MUC5AC and MUC5B) or VNTR (MUC6) antibodies. Low levels of MUC2 mRNA were detected in non‐mucinous adenocarcinomas (5/27) while a higher proportion of mucinous carcinomas (4/9) was positive. All 25 cases of adjacent normal tissue expressed MUC2 mRNA. No transcripts for MUC5AC, MUC5B or MUC 6 were detected in any of these specimens. MUC2 protein product was detected immunohistochemically in 34/36 carcinoma specimens, with no change from normal controls. There was de novo expression of MUC5AC in 23/36 carcinomas. No MUC5B or MUC6 protein was detected. No difference in MUC2 and MUC5AC protein was found between mucinous and non‐mucinous carcinomas. The level of MUC2 was increased in moderately differentiated cancers compared with normal controls and decreased in the poorly differentiated group. Decreased MUC2 was found in poorly differentiated compared with moderately differentiated tumours. More MUC5AC protein was detected in well and moderately differentiated tumours than in poorly differentiated tumours and in all tumours relative to controls. The pattern of MUC2 staining in cancers was different from control tissue, with strong staining in the perinuclear region and none in goblet cell vesicles. MUC5AC staining was mainly detected in the cytoplasm. Poor detection of MUC2 and MUC5AC mRNA and associated strong staining for the total protein suggests altered biosynthesis and processing, leading to the characteristic subcellular distribution. Hence, change in the synthesis of MUC2 and the de novo appearance of MUC5AC in colorectal carcinomas may be significant events in the adenoma‐carcinoma sequence, with possible implications for tumour prognosis. Copyright © 2001 John Wiley & Sons, Ltd.
Molecular classification of breast cancer patients by gene expression profiling*
Authors: Ahr André , Holtrich Uwe , Solbach Christine , Scharl Anton , Strebhardt Klaus , Karn Thomas , Kaufmann Manfred
Published: January 2005

For many tumuors, pathological subclasses exist which have to be further defined by genetic markers to improve therapy and follow‐up strategies. In this study, cDNA array analyses of breast cancers have been performed to classify tumuors into categories based on expression patterns. Comparing purified normal ductal epithelial cells and corresponding tumour tissues, the expression of only a small fraction of genes was found to be significantly changed. A subset of genes repeatedly found to be differentially expressed in breast cancers was subsequently employed to perform a classification of 82 normal and malignant breast specimens by cluster analysis. This analysis identifies a subgroup of transcriptionally related tumours, designated class A, which can be further subdivided into A1 and A2. Correlation with classical clinicopathological parameters revealed that subgroup A1 was characterized by a high number of node‐positive tumours (14 of 16). In this subgroup there was a disproportionate number of patients who had already developed distant metastases at the time of diagnosis (25% in this subgroup, compared with 5% among the rest of the samples). Taken together, the use of these differentially expressed marker genes in conjunction with sample clustering algorithms provides a novel molecular classification of breast cancer specimens, which facilitates the identification of patients with a higher risk of recurrence. Copyright © 2001 John Wiley & Sons, Ltd.
Cathepsin K – a marker of macrophage differentiation?
Authors: Bühling F. , Reisenauer A. , Gerber A. , Krüger S. , Weber E. , Brömme D. , Roessner A. , Ansorge S. , Welte T. , Röcken Ch.
Published: January 2005

Cathepsin K is a cysteine protease with high matrix‐degrading activity. Initially, cathepsin K was described as being expressed exclusively by osteoclasts. It was suggested that cathepsin K expression is a specific feature of cells involved in bone remodelling. The aim of this study was to investigate the hypothesis that cathepsin K is expressed not only in bone‐resorbing macrophages, but also more generally in specifically differentiated macrophages, such as epithelioid cells and multinucleated giant cells in soft tissues. Specimens obtained from different organs and anatomical locations of patients suffering from sarcoidosis, tuberculosis, granulomas caused by foreign materials, and sarcoid‐like lesions were investigated for the expression of cathepsins B, K, and L. Immunohistochemistry and in situ hybridization showed cathepsin K in epithelioid cells and multinucleated giant cells irrespective of the pathological condition and anatomical location, but not in normal resident macrophages. By immunoelectron microscopy, cathepsin K was discovered in cytoplasmic granules of multinucleated giant cells. In contrast, cathepsin B and cathepsin L were expressed ubiquitously in CD68‐positive tissue macrophages, epithelioid cells, and multinucleated giant cells. The results demonstrate that cathepsin K, but not cathepsin B or cathepsin L, differentiates specific phenotypes of macrophages independently of the anatomical site. Its enzymatic characteristics, particularly its high matrix‐degrading activity, suggest that cathepsin K‐positive epithelioid cells and multinucleated giant cells are characterized by an enhanced specific proteolytic capability. Copyright © 2001 John Wiley & Sons, Ltd.
Collagen VII expression in glomerular sclerosis
Authors: Onetti Muda Andrea , Ruzzi Laura , Bernardini Silvia , Teti Anna , Faraggiana Tullio
Published: January 2005

Glomerular sclerosis is the final stage of a variety of kidney diseases and matrix molecules not normally expressed in the extracellular matrix are synthesized and accumulate during the sclerotic process. Collagen type VII is the major component of the anchoring fibrils at the dermal–epidermal junction, but it is usually not present in normal glomeruli. The aim of this study was to investigate whether this type of fibrillary collagen, different from types I and III, is expressed in chronically diseased glomerular extracellular matrix. The presence and distribution of collagen VII have been examined in 50 renal biopsies by indirect immunofluorescence staining, standard electron microscopy, and immuno‐electron microscopy. In selected cases, collagen VII mRNA expression was also measured by RT‐PCR on isolated glomeruli. Cases included focal segmental glomerulosclerosis, minimal change disease, membranous glomerulonephritis, IgA nephropathy, SLE nephritis, diabetic glomerulosclerosis, ischaemic renal disease, extracapillary glomerulonephritis, and end‐stage renal disease. Collagen VII protein and mRNA expression was absent or present in trace amounts in normal kidneys or in disorders with only a mild increase of mesangial matrix, without scarring of the tuft. Maximal expression was evident in the presence of adhesions between the glomerular tuft and Bowman's capsule or fibrous crescents. The results showed that collagen VII is actively synthesized and laid down in areas of glomerular and/or tubular scarring, irrespective of the underlying disease, confirming the de novo expression of fibrillary collagens in diseased renal extracellular matrix. The appearance of an anchoring collagen may be a response to support mechanical stress and it takes part in the process of cell proliferation and tissue repair. Copyright © 2001 John Wiley & Sons, Ltd.
Abnormal nucleotide repeat sequence in the TGF‐βRII gene in hepatocellular carcinoma and in uninvolved liver tissue
Authors: Enomoto Akiya , Esumi Mariko , Yamashita Kazuya , Takagi Keiko , Takano Seigo , Iwai Shigetomi
Published: January 2005

Replication error (RER)‐related genetic alterations are associated with a subset of hepatocellular carcinomas (HCCs) with multiple primary cancers. This study investigated whether mutations in the nucleotide repeats of three putative target genes of RER are associated with hepatocarcinogenesis. The genes examined were those encoding transforming growth factor β type II receptor (TGF‐βRII), BCL‐2‐associated X protein (BAX), and insulin‐like growth factor II receptor (IGF‐IIR). Tumour and non‐tumour hepatic tissues were examined in 48 HCC patients, 34 with solitary HCC and 14 who had double cancer with gastric cancer. Four double‐cancer cases showed an abnormal signal in the single nucleotide repeat (A)10 of the TGF‐βRII gene. These four were among the six RER‐positive cases in this series. The genotypes of the poly A tract of the TGF‐βRII gene in the liver tumour tissue of the four cases with an abnormal signal were (A)9/10, (A)9/10, (A)9/10, and (A)9/9. Five uninvolved liver tissue specimens from these four patients showed (A)9/10 and (A)9/9, (A)9/10, (A)10/10 and (A)9/9, respectively. The genotype in the stomach cancer specimens of these four patients was (A)10/10, indicating no germline mutation of the TGF‐βRII gene. There were no mutations in the nucleotide repeats of the BAX and IGF‐IIR genes in any of the liver tissue specimens. Abnormality of the nucleotide repeat in the TGF‐βRII gene occurred in the uninvolved liver tissue as well as the HCC tissue in some HCC patients. Such genetic instability may be gene‐specific and tissue‐specific in carcinogenesis. Copyright © 2001 John Wiley & Sons, Ltd.
Co‐localization of multiple ErbB receptors in stratified epithelium of oral squamous cell carcinoma
Authors: Bei Roberto , Pompa Giorgio , Vitolo Domenico , Moriconi Enrica , Ciocci Luciano , Quaranta Manlio , Frati Luigi , H Kraus Matthias , Muraro Raffaella
Published: January 2005

The expression of all four ErbB receptors was compared by immunohistochemistry, using receptor‐specific polyclonal antisera, in 32 invasive, 11 in situ carcinomas, six benign lesions, and 22 samples of histologically normal mucosa adjacent to specimens of carcinoma originating from oral cavity epithelium. Among invasive and in situ carcinoma, EGFR expression was the most prevalent (in 29/32 and 8/11 cases, respectively) followed by ErbB2 (17/32 and 2/11) and ErbB4 (9/32 and 1/10), while ErbB3 was only detected in invasive tumours (12/32). Specific patterns included invasive tumours with expression of EGFR (8/32) or ErbB4 (1/32) alone, as well as different receptor combinations (EGFR+ErbB2, EGFR+ErbB4, EGFR+ErbB2+ErbB3, EGFR+ErbB2+ErbB4, and all four receptors). Simultaneous expression of three or four ErbB receptors correlated with tumour invasion (p=2.2×10−4) and localized in the intermediate epithelial cell layer of well and moderately differentiated tumours. No other significant correlation with clinico‐pathological features was noticed. Some benign lesions and histologically normal mucosa adjacent to carcinomas showed weak immunostaining of EGFR (10/28), ErbB2 (4/28) or ErbB4 (3/28). By comparison, overexpression, as indicated by increased staining intensity, was observed in invasive tumours for EGFR (18/32), ErbB2 (8/32), ErbB4 (3/32), and ErbB3 (3/32). Statistical evaluation demonstrated a significant association of EGFR or ErbB2 overexpression with invasive carcinoma when compared with benign lesions and apparently normal epithelium (p=5.2×10−7 and p=5×10−3, respectively). Tumour‐specific overexpression of ErbB receptors and their co‐expression, most frequently involving EGFR and ErbB2, in the same cell layer of neoplastic epithelium, implicate receptor heterodimers in the pathogenesis of oral squamous carcinoma. Copyright © 2001 John Wiley & Sons, Ltd.
Caspase activation during spontaneous and radiation‐induced apoptosis in the murine intestine
Authors: Marshman Emma , Ottewell Penelope D. , Potten Christopher S. , Watson Alastair J. M.
Published: January 2005

The aim of this study was to characterize the activation of caspase‐3 along the crypt/villus axis in the normal and irradiated intestine and to compare active caspase‐3 expression with existing apoptosis detection techniques. Small and large intestine were removed from mice at various time points after exposure to 8 Gy γ‐radiation. Positive apoptotic cells stained with an antibody against active caspase‐3, haematoxylin and eosin (H&E) or TUNEL were scored in histological sections of small and large intestinal crypts and villi. In the control intestine, active caspase‐3 expression was rarely observed; however, expression was markedly increased following exposure to radiation and was predominantly confined to apoptotic bodies. Measurement of apoptosis in intestinal crypts using active caspase‐3 expression gave similar results to apoptosis detected from H&E‐stained sections. In the normal villus, active caspase‐3 expression was observed infrequently and did not significantly increase following radiation, consistent with a lack of apoptotic body formation from H&E sections. This study indicates that caspase‐3 is activated in intestinal crypts but not in villi following γ‐radiation. Active caspase‐3 detection compared favourably with existing immunological techniques, suggesting that it is a suitable alternative method for apoptosis quantification. Copyright © 2001 John Wiley & Sons, Ltd.
Research using human tissues – a crisis of supply?
Authors: Furness Peter N.
Published: January 2005

It is a paradox that the pursuit of ethical practice can sometimes produce distinctly unethical results. Pathologists seek to study diseased human tissues for the benefit of mankind, but in the UK there has been a sudden restriction in the availability of this basic resource, supposedly on ethical grounds. This was triggered by adverse publicity about the inappropriate retention of whole organs at paediatric post‐mortem, but the reaction has spread to influence the use of ‘surgical waste’ left over after routine analysis of therapeutic resections, diagnostic biopsies, and even blood samples. The use of such material for research is being restricted, and its use in teaching and laboratory quality control is being questioned. This has occurred despite the facts that public opinion favours using such ‘waste’ material for research; that the tissues would otherwise be incinerated; and that until very recently, such tissues were regarded as having been ‘abandoned’ by the patient. This review attempts to chart how this dramatic change has occurred. It then considers some of the ethical problems in using such ‘surplus’ tissues and proposes mechanisms by which this valuable resource can remain available for most biomedical research, while maintaining or enhancing the autonomy of individual patients. This crisis has shown a tendency for regulatory authorities to apply very restrictive, oversimplified rules, without due consideration of the character of each project. The motivation is to avoid the possibility of media criticism, even though such rules block ethically sound work which is of value to all, including the individuals whose protection is supposedly being sought. The moral need for logical ethical arguments instead of such ‘one size fits all’ regulations is emphasized. Copyright © 2001 John Wiley & Sons, Ltd.
Viewing apoptosis through a ‘TUNEL’
Authors: Walker J. A. , Quirke P.
Published: January 2005

Apoptosis has an important role in carcinogenesis and cancer treatment. The end result of this complex pathway is the formation of apoptotic bodies. These can be difficult to quantify accurately, but quantitation is important if we wish to study this process. Several techniques are available which can help. ‘TUNEL’ is discussed, with its potential drawbacks, and newer antibody techniques, such as M30 and caspase 3, are then reviewed. Copyright © 2001 John Wiley & Sons, Ltd.
High‐risk HPV testing in women with borderline and mild dyskaryosis: long‐term follow‐up data and clinical relevance
Authors: Denise Zielinski G. , Snijders Peter J. F. , Rozendaal Lawrence , Voorhorst Feja J. , Runsink Arnold P. , de Schipper Frits A. , Meijer Chris J. L. M.
Published: January 2005

In The Netherlands and most other European countries, women with two serial cervical smears with borderline or mild dyskaryosis (BMD) within 6 months are referred for colposcopy‐directed biopsies. Only about 10% of these women have high‐grade cervical intraepithelial neoplasia (CIN). This study therefore investigated whether human papillomavirus (HPV) testing could identify which women with smears read as BMD are most likely to have high‐grade CIN, either at referral or during follow‐up and the relationship was determined between clearance of high‐risk HPV and regression of abnormal cytology. Women with smears read as BMD (n=278) were referred to the gynaecologist for colposcopy. They were subdivided into two groups; group A comprised women with a single smear (n=172) and group B women with two sequential smears (n=106) read as BMD before referral. High‐risk HPV detection with Hybrid Capture II (HC II) was performed on a cervical scrape taken at the first visit before colposcopy (i.e. baseline smear) and during follow‐up. Biopsies were taken when lesions suspected for CIN were seen at colposcopy. High‐risk HPV DNA was present in the baseline smears of 126 (45.0%) women; 26 (20.6%) of them had histologically confirmed CIN 2/3 at the first visit and another 14 (11.1%) during follow‐up. Only one of the 152 women (0.7%) with a negative high‐risk HPV test had a CIN 2 lesion at the first visit and no CIN lesions were detected during follow‐up of these women. After exclusion of women who were treated for prevalent high‐grade CIN, the median follow‐up times were 1.3 years (range 0.0–4.3 years) and 1.6 years (range 0.0–4.5 years) for women with HPV‐negative and HPV‐positive baseline smears, respectively. The sensitivity of a positive high‐risk HPV test for CIN 2/3 at the first visit was 96.3%, the specificity 60.2%, the positive predictive value 20.6%, and the negative predictive value 99.3%. These values did not change markedly when stratified for group A or group B. Thus, a high‐risk HPV positive test was strongly associated with the presence at the first visit and the development of CIN 2/3 lesions during follow‐up. Moreover, regression of abnormal cytology in women with a positive high‐risk HPV test at baseline was strongly associated with viral clearance and occurred 0.3 years (range −1.2 to 1.7 years) later than HPV clearance. This study establishes the value of a high‐risk HPV positive test for women at risk of high‐grade CIN, with virtually no risk for missing CIN 2/3. Addition of a test on high‐risk HPV in women with BMD could prevent 55% of the referrals and/or repeat smears. Copyright © 2001 John Wiley & Sons, Ltd.
Identiﬁcation by HPLC‐PAD‐MS and quantiﬁcation by HPLC‐PAD of phenylethanoid glycosides of five Phlomis species
Authors: Kırmızıbekmez Hasan , Montoro Paola , Piacente Sonia , Pizza Cosimo , Dönmez Ali , Çalış İhsan
Published: January 2005

The qualitative and quantitative determination of phenylethanoid glycosides in methanolic extracts of ﬁve species of the genus Phlomis (Lamiaceae) has been investigated using a new reversed‐phase HPLC method combined with photodiode‐array detection and electrospray/MS analysis. Forsythoside B, verbascoside, samioside, alyssonoside, isoverbascoside, leucosceptosides A and B and martynoside were detected. The quantiﬁcation of the above constituents was performed using echinacoside as internal standard. Knowledge of the content of phenylethanoid glycosides contributes to the chemotaxonomy of the genus. Copyright © 2005 John Wiley & Sons, Ltd.
Batch solvent extraction of flavanolignans from milk thistle (Silybum marianum L. Gaertner)
Authors: Wallace Sunny N. , Carrier Danielle J. , Clausen Edgar C.
Published: January 2005

Seeds of milk thistle (Silybum marianum L. Gaertner) contain silymarins and ca. 25% (w/w) of oil. A pre‐treatment step involving reﬂuxing with petroleum ether is usually performed before extraction of the silymarins using organic solvents. This paper compares the extraction of whole and defatted milk thistle seeds in various solvents as a function of temperature. The extraction of whole seeds of milk thistle with water at 50, 70 and 85°C was also examined: the yield of silymarin increased with increasing water temperature. In most cases, ethanol at 60°C recovered the largest quantities of silymarins. However, boiling water proved to be an efﬁcient extraction solvent for the more polar silymarins such as taxifolin and silychristin, even when using whole seeds. Extractions of defatted seed meal with boiling ethanol returned maximum yields of 0.62, 3.89, 4.04, and 6.86 mg/g defatted seed of taxifolin, silychristin, silybinin A and silybinin B, respectively. When extracting defatted seed meal with ethanol, yields of taxifolin, silybinin A and silybinin B were, respectively, 6.8‐, 0.95‐, 1.7‐ and 1.6‐fold higher than when extracting whole seeds. When extracting with boiling water, the yields of silychristin, silybinin A, and silybinin B were 380, 47 and 50% higher for whole seeds compared with defatted seeds. Copyright © 2005 John Wiley & Sons, Ltd.
Quantitative determination of the major saponin mixture bacoside A in Bacopa monnieri by HPLC
Authors: Deepak M. , Sangli G. K. , Arun P. C. , Amit A.
Published: January 2005

Bacoside A, the putative bioactive component of the Indian medicinal plant Bacopa monnieri, was found to be a mixture of saponins with bacoside A3 (1), bacopaside II (2), jujubogenin isomer of bacopasaponin C (3) and bacopasaponin C (4) as major constituents. An HPLC method together with an optimised extraction procedure was developed for the estimation of 1–4 in B. monnieri to enable standardisation of the latter. Concentration ranges of the analytes in samples of B. monnieri collected from different regions of India were 0.14–0.85% (w/w) (1), 0.12–0.69% (2), 0.05–0.72% (3) and 0.05–0.44% (4). The importance of using bacoside A, with known concentrations of 1–4, as a reference standard for the routine analysis of B. monnieri is highlighted. Two common ﬂavonoids, luteolin and apigenin, were present in all samples of B. monnieri. Copyright © 2005 John Wiley & Sons, Ltd.
Discriminating the xanthones in an extract of Swertia franchetiana by retention parameters
Authors: Longxing Wang , LeFeng Zhang , Shisheng Wang , Hongbin Xiao , Xinmiao Liang
Published: January 2005

The quantitative structure—retention relationship is one of the most actively studied topics in the ﬁeld of chromatography. In this paper, retention parameters of components were used to discriminate the xanthones in a methanol extract of Swertia franchetiana. The extract was analysed by HPLC under two different multistage linear gradient conditions and the retention parameters calculated from these retention data. It was found that the retention parameters of xanthones are in a speciﬁc region in the plot of log kw vs. S and the xanthones in the extract could be distinguished from other components by this feature. Furthermore, xanthone aglycones and xanthone glucosides could also be discriminated by retention parameters. Copyright © 2005 John Wiley & Sons, Ltd.
Application of multidimensional gas chromatography to the enantioselective characterisation of the essential oil of Eupatorium buniifolium Hooker et Arnott
Authors: Lorenzo Daniel , Paz Daniel , Davies Philip , Villamil José , Vila Roser , Cañigueral Salvador , Dellacassa Eduardo
Published: January 2005

The qualitative and quantitative composition of the essential oil from aerial parts of Eupatorium buniifolium Hooker et Arnott (Asteraceae) has been investigated for the ﬁrst time. The essential oils were obtained by steam distillation of leaves and analysed by GC‐MS; 44 components were identiﬁed. Monoterpene and sesquiterpene hydrocarbons, especially α‐pinene (14.7%), β‐elemene (12.2%), germacrene D (11.5%), trans‐β‐guaiene (6.5%) and (E)‐caryophyllene (4.3%), were the major constituents found in the oils. The characterisation of E. buniifolium by enantioselective GC was performed by evaluation of the enantiomeric ratios of α‐pinene, sabinene, β‐pinene, limonene, terpinen‐4‐ol and germacrene D. Copyright © 2005 John Wiley & Sons, Ltd.
Quality control of roots of Eleutherococcus senticosus by HPLC
Authors: Apers Sandra , Naessens Tania , Van Miert Sabine , Pieters Luc , Vlietinck Arnold
Published: January 2005

An HPLC method based on several known methods for the determination of eleutherosides B and E was developed, optimised and validated in terms of linearity, precision (repeatability and intermediate precision on different days and at different concentration levels) and accuracy (recovery). The extraction procedure, the extraction solvent and the extraction yield were evaluated and optimised. A reversed‐phase RP‐18 column gradient eluted with a two‐phase system consisting of phosphoric acid:water (0.5:99.5) and acetonitrile was used to evaluate the samples; detection was at 220 nm. Although eleutherosides B and E are commercially available, they are very costly, and therefore ferulic acid was chosen as external standard. The correction factors for the response of ferulic acid against both eleutherosides were determined and validated. This method, accepted by the European Pharmacopoeia Commission, will be included in the monograph on Eleutherococcus senticosus roots to assay the content of eleutherosides B and E. Copyright © 2005 John Wiley & Sons, Ltd.
Isolation and identiﬁcation of neo‐clerodane diterpenes from Ajuga remota by high‐performance liquid chromatography
Authors: Coll Josep , Tandrón Yudelsy
Published: January 2005

In order to explore the suitability of reversed‐phase HPLC for the rapid isolation, identiﬁcation and semipreparative fractionation of neo‐clerodane diterpenes from Ajuga remota, an extract from aerial parts of the plant was examined using a C18 column with UV detection and gradient elution with water:methanol. In addition to the known diterpenes ajugarins I, II, IV and V and clerodin, the presence and chromatographic behaviours of ajugapitin, dihydroajugapitin, dihydroclerodin and deacetylajugarin IV, not previously isolated from A. remota, were established. Presence of the epimeric mixture of 14‐hydro‐15‐hydroxyclerodin (scutecyprol A) was also demonstrated together with that of 14‐hydro‐15‐hydroxyajugapitin, the latter only in trace amounts. The structures of all isolated diterpenes were determined from NMR data. The anti‐feedant activities of the isolated compounds against Spodoptera littoralis are also reported here. Copyright © 2005 John Wiley & Sons, Ltd.
Inaccurate estimation of risk in second trimester serum screening for Down syndrome among women who have already had first trimester screening
Authors: Hackshaw A. K. , Wald N. J.
Published: January 2005

Problems can arise in prenatal screening for Down syndrome when tests are performed in the first and second trimester and some women who have a negative first trimester test have a second trimester serum test. The second test result does not usually take account of the previous one being negative. Even if it does, it is often inaccurate. Using published data the extent of the error was examined. The age‐specific risk of an affected pregnancy in such women will be lower than if no first trimester test had been performed. The distributions of the screening markers in affected and unaffected pregnancies will be different from those in unscreened women. If the appropriate age‐specific risk and marker distributions are not used, error will arise. For example, a 35‐year‐old woman with nuchal translucency (NT), pregnancy‐associated plasma protein‐A (PAPP‐A) and free β‐human chorionic gonadotrophin (hCG) levels at the normal median would have a risk of 1 in 6500. If she then had the Triple Test with alpha‐fetoprotein (AFP), unconjugated oestriol, and hCG levels of 0.7, 0.7 and 1.5 multiples of the median (MoM), respectively, her risk, ignoring the previous result, would be overestimated (1 in 95 compared with the correct estimate of 1 in 705). If the previous result was included, but the age‐specific risk and second trimester marker distributions were not revised, her risk would be underestimated (1 in 820). If the correct age‐specific risk and screening marker distributions were used, risk estimates would be accurate, but two tests would be less efficient than integrating all the screening information into a single test. The practice of offering second trimester serum screening to women who have already been screened is best avoided. Copyright © 2001 John Wiley & Sons, Ltd.
Novel feto‐specific mRNA species suitable for identification of fetal cells from the maternal circulation
Authors: Boye Kit , Hougaard David M. , Ebbesen Peter , Vuust Jens , Christiansen Michael
Published: January 2005

Feto‐specific markers are necessary for genetic diagnostics on fetal cells isolated from maternal blood. Differential display reverse transcription‐polymerase chain reaction (DDRT‐PCR) was used to identify mRNA species preferentially expressed in trophoblast‐enriched primary cell cultures compared to female peripheral mononuclear blood cells (FPMBCs). We obtained 15 different cDNA clones expressed predominantly in placenta cells using a commercial kit with oligo‐(dT) anchor primers and arbitrary upstream primers for differential display analysis of total RNA. Reverse Northern dot‐blot analysis and semi‐quantitative RT‐PCR confirmed the different expression levels. Seven mRNA species were exclusively expressed in the placenta: the mRNAs of ε‐globin, a renowned marker of fetal erythroblasts, early placenta insulin‐like peptide (EPIL), Yes‐associated protein (YAP65) and osteopontin were expressed at high levels, whereas the mRNAs of PP14, JM27 protein, and an unidentified expressed sequence tag (EST) were moderately expressed. A further eight mRNAs with low expression in FPMBCs and higher expression in first trimester placenta were identified. Some mRNAs were expressed in a trophoblast cell line, e.g. YAP65, whereas others were not, e.g. EPIL. The new mRNA markers may be used to construct DNA/RNA probesto identify fetal cells in maternal circulation by hybridization techniques, or to identify proteins selectively expressed in fetal cells and amenable to immunocytochemical detection. Copyright © 2001 John Wiley & Sons, Ltd.
Fetal nucleated red blood cells from CVS washings: an aid to development of first trimester non‐invasive prenatal diagnosis
Authors: Voullaire Lucille , Ioannou Panayiotis , Nouri Sara , Williamson Robert
Published: January 2005

Fetal nucleated red blood cells (n‐rbc) occur in the maternal circulation from 7 weeks of pregnancy. The enrichment of these cells from maternal blood will depend upon their stage of differentiation, which changes during development. We characterised the fetal n‐rbc from chorionic villus sample (CVS) washings and used them to model first trimester non‐invasive prenatal diagnosis. The ratio of ε‐ to γ‐globin‐producing cells declined rapidly from 10 to 13 weeks, as did the ratio of nucleated to non‐nucleated rbc. By 13 weeks the great majority of cells containing γ‐ or ε‐globin are anucleate. The fetal n‐rbc were highly variable in size and density and sedimented over a wide density range with a high proportion (>80%) at a density overlapping with that of maternal rbc. We have devised an enrichment procedure using Orskoff lysis to differentially lyse the maternal cells followed by density centrifugation and separation using magnetic beads. This simple protocol allowed recovery of 70% (69±22%) of fetal cells when added at approximately 10 fetal cells/ml maternal blood. When 1 fetal cell/ml millilitre maternal blood was added (total volume 10 ml) the recovery was more variable but remained at approximately 70% (72±47%), with at least one fetal cell recovered in all cases. Copyright © 2001 John Wiley & Sons, Ltd.