bluuebunny/crossref_metadata_2025 huggingface dataset:
Engraftment of sorted/expanded human central nervous system stem cells from fetal brain
Authors: Tamaki Stanley , Eckert Karl , He Dongping , Sutton Richard , Doshe Monika , Jain Gitanjali , Tushinski Robert , Reitsma Michael , Harris Brent , Tsukamoto Ann , Gage Fred , Weissman Irving , Uchida Nobuko
Published: October 2002

Direct isolation of human central nervous system stem cells (CNS‐SC) based on cell surface markers yields a highly purified stem cell population that can extensively expand in vitro and exhibit multilineage differentiation potential both in vitro and in vivo. The CNS‐SC were isolated from fetal brain tissue using the cell surface markers CD133+, CD34–, CD45–, and CD24–/lo (CD133+ cells). Fluorescence‐activated cell sorted (FACS) CD133+ cells continue to expand exponentially as neurospheres while retaining multipotential differentiation capacity for >10 passages. CD133–, CD34–, and CD45– sorted cells (∼95% of total fetal brain tissue) fail to initiate neurospheres. Neurosphere cells transplanted into neonatal immunodeficient NOD‐SCID mice proliferated, migrated, and differentiated in a site‐specific manner. However, it has been difficult to evaluate human cell engraftment, because many of the available monoclonal antibodies against neural cells (β‐tubulin III and glial fibrillary acidic protein) are not species specific. To trace the progeny of human cells after transplantation, CD133+‐derived neurosphere cells were transduced with lentiviral vectors containing enhanced green fluorescent protein (eGFP) expressed downstream of the phosphoglycerate kinase promoter. After transduction, GFP+ cells were enriched by FACS, expanded, and transplanted into the lateral ventricular space of neonatal immunodeficient NOD‐SCID brain. The progeny of transplanted cells were detected by either GFP fluorescence or antibody against GFP. GFP+ cells were present in the subventricular zone‐rostral migrating stream, olfactory bulb, and hippocampus as well as nonneurogenic sites, such as cerebellum, cerebral cortex, and striatum. Antibody against GFP revealed that some of the cells displayed differentiating dendrites and processes with neurons or glia cells. Thus, marking human CNS‐SC with reporter genes introduced by lentiviral vectors is a useful tool with which to characterize migration and differentiation of human cells in this mouse transplantation model. © 2002 Wiley‐Liss, Inc.
Effects of gamma irradiation on physical and chemical properties of chickpeas (Cicer arietinum)
Authors: Graham John A , Panozzo Joe F , Lim Patrick C , Brouwer Jan Bert
Published: October 2002

Changes in the physical and chemical properties of chickpeas gamma irradiated with 60Co at doses of 0–50 kGy were investigated. Irradiation between 0 and 20 kGy had no significant effect on the hydration capacity of the chickpeas; however, increasing the dose from 20 to 50 kGy significantly decreased the hydration capacity owing to leaching of soluble compounds from the cotyledon to the water. There was an improvement in cooking quality (defined as degree of softness) with increased irradiation. Gelatinisation properties of the starch (measured with a Rapid Visco‐Analyser) exhibited a deterioration with increasing irradiation dose, while reversed phase HPLC identified a sequential decrease in low‐molecular‐weight protein composition with increasing irradiation dose.

© 2002 Society of Chemical Industry
In vitro availability of minerals from oat products
Authors: Skibniewska Krystyna A , Kozirok Witold , Fornal Lucja , Markiewicz Kazimierz
Published: October 2002

The content of macroelements P, Mg and Ca and microelements Mn, Fe, Zn and Cu was determined in 10 commercially sold oat products made by different technological processes (dehulling, instantinising, extrusion, flaking). Phosphorus was the most prevalent of the macroelements (from 240.8 ± 2.2 to 845.5 ± 8.1 mg per 100 g), followed by magnesium (from 73.2 ± 0.7 to 271.9 ± 2.7 mg per 100 g) and calcium (from 30.69 ± 0.01 to 112.7 ± 0.3 mg per 100 g). The Ca/P ratio ranged from 1:5.3 in crushed oat to 1:8.2 in oat flakes. Regarding the microelements, manganese was present at the highest concentrations (from 2.62 ± 0.02 to 8.69 ± 0.01 mg per 100 g). The content of iron was similar and that of zinc not much lower, whereas the amount of copper was considerably smaller (from 0.23 ± 0.002 to 0.59 ± 0.002 mg per 100 g). The highest concentrations of mineral elements were found in instant oat bran flakes and the lowest in extruded oat and corn crisps containing 50% corn grouts. Samples of the products analysed were subjected to in vitro enzymatic digestion, simulating the digestive process occurring in the human alimentary tract. The supernatants thus obtained were analysed for their content of the previously determined mineral components; the percentage of minerals released from the products was calculated. The following sequence of mineral components released was observed: Cu (57.2–95.6%) > P (39.7–60.9%) > Ca (18.2–39.5%) > Mg (16.4–39.8%) > Mn (6.4–24.7%) > Fe (6.5–29%) > Zn (11–17.2%). The Ca/P ratio in the supernatant worsened from 1:8.2 in crushed oat to 1:23.3 in extruded oat and corn crisps. Crushed oat released the highest amounts of mineral elements during enzymatic hydrolysis, with oat grouts coming second. As regards the other products, it is difficult to establish their relative sequence in the release of minerals.

© 2002 Society of Chemical Industry
Effect of minimal processing on the textural and structural properties of fresh‐cut pears
Authors: Soliva‐Fortuny Robert C , Grigelmo‐Miguel Nuria , Hernando Isabel , Lluch M Ángeles , Martín‐Belloso Olga
Published: October 2002

The feasibility of minimal processing and modified atmosphere packaging to preserve the original textural quality of fresh‐cut pears was evaluated throughout storage under refrigeration. Fresh‐cut pear firmness could be extended up to several weeks with low‐O2 atmospheres. A packaging atmosphere of 100% N2 combined with the use of plastic bags with an O2 permeability of 15 cm3 m−2 bar−1 day−1 maintained cell structure and partially avoided membrane breakdown and exudate accumulation in intercellular spaces. Under the other conditions studied, pear texture underwent dramatic deterioration, which could be related to complete inundation of the extracellular environment. Firmness decreased linearly throughout the storage period studied, with rate constants ranging from 0.0097 to 0.040 day−1. The consideration of other textural variables also gave valuable information, similar to that provided by firmness data.

© 2002 Society of Chemical Industry
Dehydroepiandrosterone as an enhancer of functional recovery following crush injury to rat sciatic nerve
Authors: Gudemez Eftal , Ozer Kagan , Cunningham Brian , Siemionow Krzysztof , Browne Earl , Siemionow Maria
Published: October 2002

This study was designed to investigate the effect of dehydroepiandrosterone (DHEA) on the recovery of the rat sciatic nerve following crush injury. A standard hemostat system was used to create the injury, with a length of 1.5 mm in three groups of 18 animals each. In group I, the crush injury was applied without any treatment. In groups II and III, vehicle (ethylene glycol) and DHEA solutions were injected subepineurally 30 min following the crush injury. Sciatic function index (SFI), toe contracture measurement, gastrocinemius muscle weight, total number of myelinated fibers, fiber diameters, myelin thickness, and axon/fiber cross‐sectional ratio were measured at 3, 6, and 12 weeks. The SFI values in the DHEA group showed a faster return to normal values confirmed at 3 and 6 weeks (P < 0.05). The number of myelinated fibers and fiber diameters at 6 and 12 weeks were significantly higher in the DHEA group (P < 0.05). In this study, the subepineural injection of DHEA following crush injury was found to enhance functional recovery of the rat sciatic nerve. © 2002 Wiley‐Liss, Inc. MICROSURGERY 22:234–241 2002
Repeated upper limb salvage in a case of severe traumatic soft‐tissue and brachial artery defect
Authors: Noaman Hassan Hamdy , Shiha Anis Elsayed
Published: October 2002

We present the case of a 9‐year‐old male patient who suffered a gunshot injury to the right arm. The patient arrived in shock, his right arm severely traumatized, with soft‐tissue loss involving the anterior surface and both sides of the right arm. The humerus was exposed. There was brachial artery defect and damage to the lateral fibers of the median nerve. The mangled extremity severity score (MESS) was 8 points. The patient was treated with general resuscitation, blood transfusion, and debridement. A venous graft, 12 cm in length, to bridge the brachial artery defect, and tendon transfer, triceps to the biceps, was performed in one step. Postoperatively, there was a normal radial pulse, normal skin color, normal temperature, and normal movement of the fingers without pain. Unfortunately, the patient then sustained a second trauma to the right arm 3 weeks later, rupturing the graft. This time he lost 1,500 cc of blood. After another blood transfusion, we performed a second reverse saphenous vein graft. The patient stayed at the hospital for 3 weeks. At follow‐up 12 months later, the limb has good function and, except for the presence of a scar and skin graft, is equal in appearance to the left side. © 2002 Wiley‐Liss, Inc. MICROSURGERY 22:249–253 2002
Transient deformation of a poroelastic channel bed
Authors: Hsieh P.C. , Shih W.P. , Huang L.H.
Published: October 2002

The coupled transient response of a poroelastic bed form due to stream flow and non‐linear water waves is investigated numerically. The theory of potential flow is applied to channel flow while Biot's theory of poroelasticity (J. Appl. Phys. 1962; 33(4):1482) is adopted to deal with the deformable porous bed. A boundary‐fitted co‐ordinate system is used to calculate the variation in the bed form. The result of a simple periodic wave form over a soft poroelastic bed agrees well with the analytical solution of Hsieh et al. (J. Eng. Mech., ASCE 2000; 126(10):1064). However, due to the rapidly damping second dilatational wave inside the soft poroelastic bed, the solution for transient bed form near the interface is not easy to compute accurately. In order to overcome this difficulty, a simplified numerical model based on the boundary layer correction concept proposed by Hsieh et al. (2000) is established, which neglects Darcy's terms. The transient deformation of an irregular poroelastic bed that includes a trench and a downward step at the channel bed is simulated successfully. Copyright © 2002 John Wiley & Sons, Ltd.
A theoretical framework for constructing elastic/plastic constitutive models of triaxial tests
Authors: Collins Ian F. , Hilder Tamsyn
Published: October 2002

Modern ideas of thermomechanics are used to develop families of models describing the elastic/plastic behaviour of cohesionless soils deforming under triaxial conditions. Once the form of the free energy and dissipation potential functions have been specified, the corresponding yield loci, flow rules, isotropic and kinematic hardening rules as well as the elasticity law are deduced in a systematic manner. The families contain the classical linear frictional (Coulomb type) models and the classical critical state models as special cases. The generalized models discussed here include non‐associated flow rules, shear as well as volumetric hardening, anisotropic responses and rotational yield loci. The various parameters needed to describe the models can be interpreted in terms of ratio of the plastic work, which is dissipated, to that which is stored. Non‐associated behaviour is found to occur whenever this division between dissipated and stored work is not equal. Micro‐level interpretations of stored plastic work are discussed. The models automatically satisfy the laws of thermodynamics, and there is no need to invoke any stability postulates. Some classical forms of the peak‐strength/dilatancy relationship are established theoretically. Some representative drained and undrained paths are computed. Copyright © 2002 John Wiley & Sons, Ltd.
Making ‘modernising government initiatives’ work: culture change through Collaborative Inquiry (CI)
Authors: Kakabadse Andrew , Kakabadse Nada
Published: October 2002

The purpose of this article is to share experience of a strategic intervention adopting a collaborative inquiry (CI) process that took place in a formal institutional setting and which evolved into a community‐based inquiry. The article illustrates the CI process through a twelve‐month exploration with a group of senior managers from within the UK Civil Service. The form of CI demonstrated is similar to what has become known as community based research, in that all participants collaboratively pursued the inquiry. A critical element of this collaborative inquiry is that although it started formally and did not spontaneously emerge from the inquiry community, its members shaped the inquiry, conveyed the message, acted as advocates and evaluated the outcomes of the inquiry. The impact of this inquiry is now being positively felt within the UK Civil Service as the members of the inquiry community apply their learning within their own departments. Copyright © 2002 John Wiley & Sons, Ltd.
Building civic infrastructure: implementing community partnership grant programmes in South Africa
Authors: Adams Charles F. , Bell Michael E. , Brown Trevor
Published: October 2002

This article examines recent efforts to establish Community Partnership Grant Programmes (CPG) in six South African communities. CPG programmes provide the financial and organizational infrastructure to support citizen‐initiated neighbourhood projects. We review our efforts to disseminate the CPG programme model in three different environments in South Africa—a large metropolitan area, three smaller municipalities and two tribal villages—and analyse why preliminary implementation results were positive in some communities but not others. Our findings form the basis of a model of CPG programme implementation based on the mode of implementation and the breadth of stakeholder involvement. We use this model to make recommendations to increase the likelihood of implementation success for similar programmes. The article concludes with observations about the applicability of the CPG programme idea as a way to develop civic infrastructure, and on the political lessons learned from our implementation experience in South Africa. Copyright © 2002 John Wiley & Sons, Ltd.
Gains and losses of adhesion molecules (CD44, E‐cadherin, and β‐catenin) during oral carcinogenesis and tumour progression
Authors: Bánkfalvi Agnes , Kraßort Melanie , Buchwalow Igor B. , Végh Andras , Felszeghy Endre , Piffkó Jozsef
Published: October 2002

The aim of this study was to define whether or not the impaired expression of CD44, E‐cadherin (E‐cad), and β‐catenin (β‐cat) correlates with the clinical evolution and prognosis of oral cancer. Ninety‐three primary oral squamous cell carcinomas (OSCCs) with tumour‐adjacent normal and/or dysplastic mucosa, 30 associated metastases, and 12 recurrences were immunostained for CD44s, ‐v3, ‐v4, ‐v5, ‐v6, ‐v7, ‐v9, E‐cad, and β‐cat. In non‐neoplastic epithelium, all molecules investigated were constitutively expressed in the basal layers. In the majority of dysplasias, immunoreactivity for all adhesion molecules was increased, but there was restricted loss for CD44s, E‐cad, and β‐cat in a few cases. In carcinomas, a striking accumulation of CD44s, v3, v4, v9 and a loss of E‐cad/β‐cat were observed at the invasive tumour front. In metastases and recurrences, besides a loss of CD44s, v4, v7, and E‐cad, a significant increase of v9 was recorded, whereas CD44v5 and v6 remained unchanged. Clinically, reduced expression of CD44v3, E‐cad, and changes of CD44v9 phenotype within the primary tumours correlated significantly with poor prognosis; decreased β‐cat expression was a predictive marker for nodal metastases. These findings indicate that there is some perturbed expression of adhesion molecules during the stepwise course of oral carcinogenesis and tumour progression. Distinct phenotypic alterations project poor prognosis, while others predict metastasis. Some of these restricted molecular changes may serve as potential targets for future antibody‐based tumour therapy. Copyright © 2002 John Wiley & Sons, Ltd.
Therapeutic effect of sulphated hyaluronic acid, a potential selectin‐blocking agent, on experimental progressive mesangial proliferative glomerulonephritis
Authors: Matsuda Mitsuhiro , Shikata Kenichi , Shimizu Fujio , Suzuki Yasuo , Miyasaka Masayuki , Kawachi Hiroshi , Kawashima Hiroto , Wada Jun , Sugimoto Hikaru , Shikata Yasushi , Ogawa Daisuke , Tojo Shinichiro J. , Akima Kazuo , Makino Hirofumi
Published: October 2002

The initial event in the process of leukocyte infiltration is characterized by leukocyte rolling on the surface of the endothelium, which is mediated by selectins. P‐ and L‐selectin bind to the sulphated sugar chains of their natural ligands, including sulphated glycolipids such as sulphatide. Recently, it has been demonstrated that sulphated glycolipids and sulphated oligosaccharides interfere with selectin binding pathways. This study synthesized sulphated hyaluronic acid (SHA), which is a potential selectin‐blocking agent, and examined its therapeutic effect on the experimental progressive mesangial proliferative glomerulonephritis induced by anti‐Thy‐1 monoclonal antibody (1‐22‐3 MAb) after unilateral nephrectomy. The selectin‐inhibitory effect of SHA in vitro was confirmed. SHA inhibited the binding of P‐ and L‐selectin to sulphatide, which is a glycolipid ligand for P‐ and L‐selectin, at a concentration of 1.5 µg/ml and 100 µg/ml. Immunohistochemical examination showed that P‐selectin was up‐regulated in the glomeruli in the 1‐22‐3 MAb nephritis model, while the ligands for L‐selectin were not detected in the glomerular tufts. A single administration of SHA ameliorated proteinuria and glomerular leukocyte infiltration in 24 h after the injection of anti‐Thy‐1 MAb. Anti‐P‐selectin MAb, but not anti‐L‐selectin MAb, inhibited proteinuria and glomerular leukocyte infiltration. To examine further the therapeutic effect of SHA on chronic glomerulonephritis, SHA was administered daily from day 3 to day 14 in this model. Proteinuria and glomerular leukocyte infiltration were significantly diminished in SHA‐treated rats on day 14. These results suggest that SHA ameliorated rat progressive mesangial proliferative glomerulonephritis by inhibiting P‐selectin‐dependent leukocyte infiltration in glomeruli. Sulphated oligosaccharides may be beneficial for the therapy of mesangial proliferative glomerulonephritis. Copyright © 2002 John Wiley & Sons, Ltd.
Alterations of the INK4a‐ARF gene locus in pleomorphic adenoma of the parotid gland
Authors: Weber Anette , Langhanki Larissa , Schütz Alexander , Wittekind Christian , Bootz Friedrich , Tannapfel Andrea
Published: October 2002

Pleomorphic adenomas of the parotid gland are benign tumours composed of epithelial and mesenchymal cells. The INK4a‐ARF (CDKN2A) locus on chromosome 9p21 encodes two tumour suppressor proteins, p16INK4a and p14ARF, which act as upstream regulators of the Rb‐CDK4 and p53 pathways. To study the contribution of each pathway in pleomorphic adenomas, this study analysed alterations of p14ARF, p16INK4a, p53, and pRb in these tumours. After microdissecting the different histological components, 42 pleomorphic adenomas of the parotid gland were analysed for INK4a‐ARF inactivation by DNA sequence analysis, methylation‐specific PCR (MSP), restriction enzyme‐related polymerase chain reaction (RE‐PCR), mRNA expression, microsatellite analysis, and immunohistochemistry. In addition, microdeletion of p14ARF and p16INK4a were assessed by differential PCR. The status of p53 and Rb was examined by direct sequencing and immunohistochemistry. Using microdissection, it was possible to examine the tumour components, i.e. epithelial, mesenchymal, and transitional, separately after immunohistochemical identification. Methylation of p14ARF was found in 1/42 cases and alterations of p16INK4a occurred in 12/42 of pleomorphic adenomas, which correlated with loss of mRNA transcription. Microdeletions or specific mutations of either exon were not detected. Methylation was detected exclusively in the epithelial and transitional components and not within the mesenchymal part of the tumour. p53 mutations were detected in 4/42 adenomas, also occurring solely in the epithelial components of the tumours. pRb was detected immunohistochemically in 40/42 adenomas. In normal, corresponding parotid tissue, p14ARF, p16INK4a, p53, and pRb alterations were not observed. The observation that alterations of p14ARF and p16INK4a, and also p53 mutations, occurred exclusively in the epithelial and transitional components of pleomorphic adenoma supports the theory that these areas are prone to malignant transformation to carcinoma in adenoma. Copyright © 2002 John Wiley & Sons, Ltd.
Morphometry of dermal collagen orientation by Fourier analysis is superior to multi‐observer assessment
Authors: van Zuijlen Paul PM , de Vries Henry JC , Lamme Evert N , Coppens Joris E , van Marle Jan , Kreis Robert W , Middelkoop Esther
Published: October 2002

In human dermis, collagen bundle architecture appears randomly organized, whereas in pathological conditions, such as scar tissue and connective tissue disorders, collagen bundle architecture is arranged in a more parallel fashion. Histological examination by one or two observers using polarized light is the most common method to determine collagen orientation. The hypothesis on which this study is based is that an objective image analysis technique, Fourier analysis, would improve the reliability (are the measurements reproducible?) and the accuracy (does the method measure what it is supposed to measure?) of collagen orientation assessment, compared with observer ratings. Fourier analysis was applied to 271 images of scar tissue and normal skin that were acquired by confocal laser‐scanning microscopy. Observers rated the same areas using polarized light as well as the confocal microscopy images. Computer images consisting of different types of ellipses were generated with a fixed orientation. Observers and Fourier analysis evaluated the images to evaluate accuracy. The inter‐observer reliability was acceptable when at least three observers rated polarized light images (r > 0.69), whereas two observers were sufficient for rating confocal microscopy images (r > 0.71). Fourier analysis correlated better with observer ratings of confocal microscopy images (r = 0.69) than with polarized light microscopy images (r = 0.42). Fourier analysis was more accurate than four observers for the evaluation of the ‘true’ orientation for almost all types of computer‐generated images. For the first time it is shown that Fourier image analysis is suitable for the morphometry of dermal collagen orientation and leads to a superior measurement of collagen orientation compared with subjective histological evaluation by several experts. If an evaluation is performed by conventional light microscopy, at least three observers are required to attain an acceptable inter‐observer reliability. Copyright © 2002 John Wiley & Sons, Ltd.
Antimicrobial peptides are expressed and produced in healthy and inflamed human synovial membranes
Authors: Paulsen Friedrich , Pufe Thomas , Conradi Lenard , Varoga Deike , Tsokos Michael , Papendieck Jann , Petersen Wolf
Published: October 2002

The objective of this study was to determine the expression and production of antimicrobial peptides by healthy and inflamed human synovial membranes. Deposition of the antimicrobial peptides lysozyme, lactoferrin, secretory phospholipase A2 (sPA2), matrilysin (MMP7), human neutrophil alpha‐defensins 1–3 (HNP 1–3), human beta‐defensin 1 (HBD‐1), and human beta‐defensin 2 (HBD‐2) was determined by immunohistochemistry. Expression of mRNA for the antimicrobial peptides bactericidal permeability‐increasing protein (BPI), heparin binding protein (CAP37), human cationic antimicrobial protein (LL37), human alpha‐defensin 5 (HD5), human alpha‐defensin 6 (HD6), HBD‐1, HBD‐2, and human beta‐defensin 3 (HBD‐3) was analysed by reverse transcription polymerase chain reaction (RT‐PCR). RT‐PCR revealed CAP37 and HBD‐1 mRNA in samples of healthy synovial membrane. Additionally, HBD‐3 and/or LL37 mRNA was detected in synovial membrane samples from patients with pyogenic arthritis (PA), osteoarthritis (OA) or rheumatoid arthritis (RA). BPI, HD5, HD6, and HBD‐2 mRNAs were absent from all samples investigated. Immunohistochemistry identified lysozyme, lactoferrin, sPA2, and MMP7 in type A synoviocytes of all samples. HBD‐1 was only present in type B synoviocytes of some of the samples. Immunoreactive HBD‐2 peptide was only visible in some inflamed samples. HNP1‐3 was detected in both healthy and inflamed synovial membranes. The data suggest that human synovial membranes produce a broad spectrum of antimicrobial peptides. Under inflammatory conditions, the expression pattern changes, with induction of HBD‐3 in PA (LL37 in RA; HBD‐3 and LL37 in OA) as well as down‐regulation of HBD‐1. HBD‐3 holds therapeutic potential in PA as it has a broad spectrum of antimicrobial activity and accelerates epithelial healing. However, caution is appropriate since defensins also promote fibrin formation and cell proliferation — key elements in joint infection. Clarification of the role of antimicrobial peptides in OA and RA will require further investigation. Copyright © 2002 John Wiley & Sons, Ltd.
The framework of pathology: good laboratory practice by quantitative and molecular methods
Authors: Baak Jan PA
Published: October 2002

Combined confocal laser scan microscopy (CLSM) and Fourier analysis (FA) by non‐pathologists of dermal collagen bundle orientation recently gave results superior to subjective evaluation by experts. According to Good Laboratory Practice (GLP) criteria, combined CLSM/FA has not yet been adequately tested to replace current collagen evaluation, but this will not take long. Non‐pathologists (clinicians) will then have taken over a laboratory test historically belonging to pathology. A general trend in this direction may develop, because pathologists seem not always to care enough about clinical significance, reproducibility and prognostic value, and new demands for innovative methods. Quantitative image analysis (QIA) and molecular methods are reproducible, inexpensive, and easy to perform; they often have greater value than classical evaluations and their cost–benefit ratio is good. However, their acceptance is not as widespread as one would expect and theoretical reasons which have been advanced do not provide a satisfactory explanation. A formal implementation study was therefore performed, in which an attempt was made to modernize a classical pathology laboratory. An external customer satisfaction investigation showed that 96% of the clinicians were ‘very satisfied’ (the highest rating possible) with the completed innovations, contrasting with low satisfaction at the beginning. Lack of primary innovative leadership among pathologists was judged to be the dominant cause preventing implementation. Pathologists should focus on carefully reacting to new clinical needs, using GLP criteria. Reproducibility and predictive accuracy should be major themes in any pathology practice. Copyright © 2002 John Wiley & Sons, Ltd.
A copula‐based model for multivariate non‐normal longitudinal data: analysis of a dose titration safety study on a new antidepressant
Authors: Lambert Philippe , Vandenhende François
Published: October 2002

A new model for multivariate non‐normal longitudinal data is proposed. In a first step, each longitudinal series of data corresponding to a given response is modelled separately using a copula to relate the marginal distributions of the response at each time of observation. In a second step, at each observation time, the conditional (on the past) distributions of each response are related using another copula describing the relationship between the corresponding variables. Note that there is no need to consider the same family of distributions for these response variables. The technique is illustrated in a dose titration safety study on a new antidepressant. The haemodynamic effect on diastolic blood pressure, systolic blood pressure and heart rate is studied. These three responses are measured repeatedly over time on ten healthy volunteers during the dose escalation. The available covariates are sex and the concentration of drug in the plasma at time of measurement. Copyright © 2002 John Wiley & Sons, Ltd.
Proportional hazards models with frailties and random effects
Authors: O'Quigley John , Stare Janez
Published: October 2002

We discuss some of the fundamental concepts underlying the development of frailty and random effects models in survival. One of these fundamental concepts was the idea of a frailty model where each subject has his or her own disposition to failure, their so‐called frailty, additional to any effects we wish to quantify via regression. Although the concept of individual frailty can be of value when thinking about how data arise or when interpreting parameter estimates in the context of a fitted model, we argue that the concept is of limited practical value. Individual random effects (frailties), whenever detected, can be made to disappear by elementary model transformation. In consequence, unless we are to take some model form as unassailable, beyond challenge and carved in stone, and if we are to understand the term ‘frailty’ as referring to individual random effects, then frailty models have no value. Random effects models on the other hand, in which groups of individuals share some common effect, can be used to advantage. Even in this case however, if we are prepared to sacrifice some efficiency, we can avoid complex modelling by using the considerable power already provided by the stratified proportional hazards model. Stratified models and random effects models can both be seen to be particular cases of partially proportional hazards models, a view that gives further insight. The added structure of a random effects model, viewed as a stratified proportional hazards model with some added distributional constraints, will, for group sizes of five or more, provide no more than modest efficiency gains, even when the additional assumptions are exactly true. On the other hand, for moderate to large numbers of very small groups, of sizes two or three, the study of twins being a well known example, the efficiency gains of the random effects model can be far from negligible. For such applications, the case for using random effects models rather than the stratified model is strong. This is especially so in view of the good robustness properties of random effects models. Nonetheless, the simpler analysis, based upon the stratified model, remains valid, albeit making a less efficient use of resources. Copyright © 2002 John Wiley & Sons, Ltd.
A semi‐parametric accelerated failure time cure model
Authors: Li Chin‐Shang , Taylor Jeremy M. G.
Published: October 2002

A cure model is a useful approach for analysing failure time data in which some subjects could eventually experience, and others never experience, the event of interest. A cure model has two components: incidence which indicates whether the event could eventually occur and latency which denotes when the event will occur given the subject is susceptible to the event. In this paper, we propose a semi‐parametric cure model in which covariates can affect both the incidence and the latency. A logistic regression model is proposed for the incidence, and the latency is determined by an accelerated failure time regression model with unspecified error distribution. An EM algorithm is developed to fit the model. The procedure is applied to a data set of tonsil cancer patients treated with radiation therapy. Copyright © 2002 John Wiley & Sons, Ltd.
A simple confidence interval for meta‐analysis
Authors: Sidik Kurex , Jonkman Jeffrey N.
Published: October 2002

In the context of a random effects model for meta‐analysis, a number of methods are available to estimate confidence limits for the overall mean effect. A simple and commonly used method is the DerSimonian and Laird approach. This paper discusses an alternative simple approach for constructing the confidence interval, based on the t‐distribution. This approach has improved coverage probability compared to the DerSimonian and Laird method. Moreover, it is easy to calculate, and unlike some methods suggested in the statistical literature, no iterative computation is required. Copyright © 2002 John Wiley & Sons, Ltd.
Parametric models for accelerated and long‐term survival: a comment on proportional hazards
Authors: Frankel Paul , Longmate Jeffrey
Published: October 2002

The Cox proportional hazards model (CPH) is routinely used in clinical trials, but it may encounter serious difficulties with departures from the proportional hazards assumption, even when the departures are not readily detected by commonly used diagnostics. We consider the Gamel–Boag (GB) model, a log‐normal model for accelerated failure in which a proportion of subjects are long‐term survivors. When the CPH model is fit to simulated data generated from this model, the results can range from gross overstatement of the effect size, to a situation where increasing follow‐up may cause a decline in power. We implement a fitting algorithm for the GB model that permits separate covariate effects on the rapidity of early failure and the fraction of long‐term survivors. When effects are detected by both the CPH and GB methods, the attribution of the effect to long‐term or short‐term survival may change the interpretation of the data. We believe these examples motivate more frequent use of parametric survival models in conjunction with the semi‐parametric Cox proportional hazards model. Copyright © 2002 John Wiley & Sons, Ltd.
The phage Mu repressor c and IS30 transposase proteins are significantly related
Authors: Dalrymple Brian
Published: October 2002

The IS30 transposase exhibits significant amino acid sequence homology to the phage Mu repressor c in the amino‐ and carboxy‐terminal regions of the proteins. The conserved sequences include the proposed Mu repressor DNA binding site, which is also related to the proposed Mu and D108 transposase DNA binding sites. The carboxy‐terminal homologies are characterised by two almost complete, and one partial, somewhat diverged amino acid sequence repeats. Only weak homologies to this domain are present in the Mu transposase (Mu A). Nevertheless, a clear link between an insertion sequence and a bacteriophage has been established.
A computer graphics model of frog γ‐crystallin based on the three‐dimensional structure of calf γ‐II crystallin
Authors: Summers L.J. , Blundell T.L. , Gause G.G. , Tomarev S.I.
Published: October 2002

Molecular models for Rana γ‐1 and γ‐2 crystallins have been constructed using computer graphics on the basis of the protein primary structure derived from the complementary DNA sequence and the three‐dimensional structure of calf γ‐II crystallin that has been defined at high resolution by X‐ray analysis. The models show that the cores of the two domains are conserved as hydrophobic, with the polypeptide chain arranged as a four Greek‐key motif structure. Although many lysines replace arginines at equivalent positions in mammalian proteins, the Rana crystallins also have an extensive series of ion pairs on their surface; these are strongly implicated in their function as stable structural molecules, which are highly conserved in the evolution of the vertebrate eye lens.
Involvement of three intracellular messenger systems, protein kinase C, calcium ion and cyclic AMP, in the regulation of c‐fos gene expression in Swiss 3T3 cells
Authors: Tsuda Terutaka , Hamamori Yasuo , Yamashita Takayuki , Fukumoto Yasuo , Takai Yoshimi
Published: October 2002

In quiescent cultures of Swiss 3T3 cells, platelet‐derived growth factor or fibroblast growth factor known to induce both protein kinase C activation and Ca2+ mobilization raised c‐fos mRNA. This action of the growth factors was mimicked by the specific activators for protein kinase C, such as phorbol esters and a membrane‐permeable synthetic diacylglycerol, and also by the Ca2+ ionophores, such as A23187 and ionomycin. Prostaglandin E1 known to elevate cyclic AMP also raised c‐fos mRNA, and this action was mimicked by 8‐bromo‐cyclic AMP, dibutyryl cyclic AMP and forskolin. These results suggest that expression of the c‐fos gene is regulated by three different intracellular messenger systems, protein kinase C, Ca2+ and cyclic AMP, in Swiss 3T3 cells.
Structure of rubredoxin from the bacterium Desulfovibrio desulfuricans
Authors: Sieker Larry C. , Stenkamp Ronald E. , Jensen Lyle H. , Prickril Ben , LeGall Jean
Published: October 2002

The X‐ray crystallographic structure of rubredoxin from Desulfovibrio desulfuricans strain 27774 is described. This molecule is 15% smaller than previously studied rubredoxins, lacking a seven‐residue loop of chain but containing a histidine and a free‐sulfhydryl cysteine. Except for solvent exposure of the single invariant tryptophan, no other major difference occurs in the molecule.
Primary structure of elongation factor 1β from Artemia
Authors: Maessen G.D.F. , Amons R. , Maassen J.A. , Möller W.
Published: October 2002

cDNA complementary to mRNA coding for the elongation factor EF‐1β has been cloned. A γgt 11 cDNA library has been screened with an antiserum against EF‐1β which exchanges GDP bound to EF‐1α with exogenous GTP during protein synthesis. The derived amino acid sequence corresponds to 208 amino acids including the N‐terminal methionine which is absent in the mature protein. About sixty percent of the protein was sequenced by direct protein sequence analysis. Comparison of Artemis salina EF‐1β with Escherichia coli EF‐Ts shows no evident homology.
Identification of granzyme A isolated from cytotoxic T‐lymphocyte‐granules as one of the proteases encoded by CTL‐specific genes
Authors: Masson Daniéle , Zamai Moreno , Tschopp Jürg
Published: October 2002

A serine esterase called granzyme A, which is specifically expressed in cytolytic lymphocytes has been characterized. It is a disulfide‐linked dimer and exhibits a trypsin‐like specificity cleaving best after Arg. N‐terminal sequence analysis revealed that granzyme A is identical to a protease recently predicted from a cloned CTL‐specific gene.
Assignment of the 31P and 1H resonances in oligonucleotides by two‐dimensional NMR spectroscopy
Authors: Sklenár̆ Vladimir , Miyashiro Hirotsugu , Zon Gerald , Todd Miles H. , Bax Ad
Published: October 2002

The use of new 1H‐detected heteronuclear 1H‐31P shift correlation experiments is demonstrated for oligonucleotides of 12 and 40 base pairs. The methods give unambiguous assignments of the 31P resonances and also permit identification of the C4′ and C5′ sugar protons. Use of the new methods enables one to make sequence‐specific resonance assignments without reference to a known or assumed conformation of the DNA fragment.
The binding of skeletal muscle C‐protein to regulated actin
Authors: Yamamoto Katsuhiro
Published: October 2002

The binding of C‐protein, a component of thick filament of myofibrils, to regulated actin filaments in the presence or absence of CA2+ was studied. The amount of C‐protein bound to regulated actin filaments in the presence of CA2+ was higher than those in the absence of Ca2+. The addition of C‐protein to regulated actin caused an increase in turbidity, especially in the presence of Ca2+, and this was found to result from side‐by‐side association of actin filaments into bundles. In the absence of Ca2+, no actin filament bundles were formed.
The evolution of placental mammals
Authors: Harris J.R.
Published: October 2002

Based on morphological, virological, biochemical and molecular biological data, it is proposed that the presence of endogenous retrovirus particles in the placental cytotrophoblasts of many mammals is indicative of some beneficial action provided by the virus in relation to cell fusion, syncytiotrophoblast formation and the creation of the placenta. Further, it is hypothesised that the germ line retroviral infection of some primitive mammal‐like species resulted in the evolution of the placental mammals.
Subunit separation in reversed micelle system reveals the existence of active centers both on light and heavy γ‐glutamyltransferase subunits
Authors: Kabanov Alexander V. , Nametkin Sergey N. , Chernov Nikolai N. , Klyachko Natalya L. , Levashov Andrey V.
Published: October 2002

Regulation of supra‐macromolecular composition and catalytic activity of a heterodimeric enzyme, γ‐glutamyltransferase, in the system of Aerosol OT (sodium bis(2‐ethylhexyl) sulfosuccinate) reversed micelles in octane were studied. Variation of the surfactant hydration degree (parameter, determining dimensions of the polar inner cavity of the micelle) causes a reversible dissociation of the enzyme to light and heavy subunits. Both enzyme subunits possess catalytic activity. The light and heavy subunits of the enzyme were separated on a preparative scale in a reversed micelle system using ultracentrifugation. The active centers of γ‐glutamyltransferase were studied using its irreversible inhibitor — AT‐125 (L·(αS.5S)‐α‐amino‐3‐chloro‐4,5‐dihydro‐5‐isoxazoleacetic acid). Separation of the γ‐glutamyltransferase subunits results in the ‘opening’ of a new active center located at the heavy subunit. In the dimer form of the enzyme this center is masked and it is not accessible to both substrate and inhibitor molecules.
Effects of the membrane potential upon the Ca2+‐ and cumene hydroperoxide‐induced permeabilization of the inner mitochondrial membrane
Authors: Novgorodov Sergey A. , Gudz Tatjana I. , Kushnareva Yulia E. , Eriksson Ove , Leikin Yury N.
Published: October 2002

A protonophore‐induced ΔΨ decrease in a 180‐140 mV range causes an increase in the lag‐period of Ca2+‐induced mitochondrial permeabilization but has little effect on the cumene hydroperoxide‐induced permeability transition of mitochondria. Suppression of the non‐specific permeability induction seems to be mediated by an increase in [ADP] in the mitochondrial matrix. A further decrease in ΔΨ leads to additional suppression of the non‐specific permeability as a result of a partial ruthenium red‐sensitive efflux of the previously accumulated Ca2+. On the other hand, complete dissipation of ΔΨ causes immediate induction of the non‐specific permeability. It is concluded that only complete dissipation of ΔΨ caused by H+ leakages may act as a trigger for non‐specific permeability induction.
Molecular cloning and expression of the cDNA coding for a new member of the S100 protein family from porcine cardiac muscle
Authors: Ohta Hisataka , Sasaki Toshiya , Naka Michiko , Hiraoka Osamu , Miyamoto Chikara , Furuichi Yasuhiro , Tanaka Toshio
Published: October 2002

We isolated a new calcium‐binding protein from porcine cardiac muscle by calcium‐dependent hydrophobic and dye‐affinity chromatography. It showed an apparent molecular weight of 11 000 on SDS‐PAGE. Amino acid sequence determination revealed that the protein contained two calcium‐binding domains of the EF‐hand motif. The cDNA gene coding for this protein was cloned from the porcine lung cDNA library. Sequence analysis of the cloned cDNA showed that the protein was composed of 99 amino acid residues and its molecular weight was estimated to be 11 179. Immunological and functional characterization showed that the recombinant S100C protein expressed in Escherichia coli was identical to the natural protein. Homologies to calpactin light chain, S100α and β protein were 41.1%, 40.9% and 37.5%, respectively. The protein was expressed at high levels in lung and kidney, and low levels in liver and brain. The tissue distribution was apparently different from those of the other S100 protein family. These results indicate that this protein represents a new member of the S100 protein family, and thus we refer to it as S100C protein.
EPR‐spectroscopy of reduced oxyferrous‐P450cam
Authors: Davydov R. , Kappl R. , Hüttermann J. , Peterson J.A.
Published: October 2002

X‐irradiation of the ternary complex of P450:substrate:O2 at 77 K produces a reduced intermediate by electron addition to the Fe:O2 complex which can be studied by EPR‐spectroscopy. The EPR spectrum of the new species exhibits rhombic symmetry with g‐factors of 2.27, 2.17 and 1.95, respectively. Increasing the temperature of the sample to 190 K results in loss of intensity of the intermediate signals. X‐irradiation of oxymyo‐ and oxyhemoglobin produces similar EPR signals indicating that the added electron is resident on the Fe:O2 complex (Kappl, R., et al. (1985) Biochim. Biophys. Acta 870, 20–30).
Cloning and functional characterization of a cocaine‐sensitive dopamine transporter
Authors: Giros Bruno , El Mestikawy Salah , Bertrand Lucie , Caron Marc G.
Published: October 2002

We report the cloning of a rat cDNA encoding a functional dopamine transporter. This cDNA, derived from an intron‐containing gene, encodes a protein of 620 amino acids. Hydropathicity analysis of the protein sequence suggests the presence of 12 putative transmembrane domains. The protein displays considerable identity with transporters for noradrenaline and GABA (64 and 30%, respectively). Transient expression of the cDNA in COS7 cells directs the expression of dopamine uptake activity with appropriate pharmacology and in a sodium‐dependent fashion. In situ hybridization reveals that the mRNA for this transporter is expressed in the substantia nigra and ventral tegmental area, regions that contain dopaminergic cell bodies.
Fluorine‐19 NMR studies of the thermal unfolding of 5‐fluorouracil‐substituted Escherichia coli valine transfer RNA
Authors: Chu Wen-Chy , Horowitz Jack
Published: October 2002

19F NMR spectroscopy was used to monitor the thermal unfolding of E. coli tRNAVal labeled by incorporation of 5‐fluorouracil (FUra). With rising temperatures, resonances in the 19F NMR spectrum of (FUra)tRNAVal gradually shift towards the central region of the spectrum and merge into a single broad peak above 85°C. FU55 and FU12 are the first to shift, beginning at temperatures below 40°C, which suggests that the initial steps of thermal denaturation of tRNAVal involve disruption of the tertiary interactions between the D‐ and T‐arms. The acceptor stem and the FU64‐G50 wobble base pair in the T‐stem are particularly stable to thermal denaturation. A temperature‐dependent splitting of the 19F resonance assigned to FU64, at temperatures above 40°C, suggests that the T‐arm of (FUra)tRNAVal exists in two conformations in slow exchange on the NMR time scale.
Phosphorylation‐activated 6‐phosphofructo‐2‐kinase from mantle tissue of marine mussels
Authors: Vázquez-Illanes M.D. , Ramos-Martinez J.I.
Published: October 2002

PKF‐2 from mussel mantle was phosphorylated by cAMP‐dependent protein kinase. The phosphorylation does not change the enzyme activity at neutral pH values, but at acid pH the activity of the phosphorylated form is higher than the native PFK‐2. With respect to the native enzyme, the activation consisted of a reduction in the K
fm for Fru‐6‐P and a decrease in the inhibitory effect of PEP. These results are in keeping with the stabilized concentration of Fru‐2,6‐P2 found in the mussel mantle during the physiological hypoxia caused by the closure of the valves.