bluuebunny/crossref_metadata_2025 huggingface dataset:
Groundwater–surface‐water interactions in a braided river: a tracer‐based assessment
Authors: Rodgers P. , Soulsby C. , Petry J. , Malcolm I. , Gibbins C. , Dunn S.
Published: March 2004

Natural tracers (alkalinity and silica) were used to infer groundwater–surface‐water exchanges in the main braided reach of the River Feshie, Cairngorms, Scotland. Stream‐water samples were collected upstream and downstream of the braided section at fortnightly intervals throughout the 2001–2002 hydrological year and subsequently at finer resolution over two rainfall events. The braided reach was found to exert a significant downstream buffering effect on the alkalinity of these waters, particularly at moderate flows (4–8 m3 s−1/≅Q30–70). Extensive hydrochemical surveys were undertaken to characterize the different source waters feeding the braids. Shallow groundwater flow systems at the edge of the braided floodplain, recharged by effluent streams and hillslope drainage, appeared to be of particular significance. Deeper groundwater was identified closer to the main channel, upwelling through the hyporheic zone. Both sources contributed to the significant groundwater–surface‐water interactions that promote the buffering effect observed through the braided reach. Their impact was less significant at higher flows (>15 m3 s−1/>Q10) when acidic storm runoff from the peat‐covered catchment headwaters dominated, as well as under baseflow conditions (<4 m3 s−1/<Q70), when upstream alkalinity was already buffered owing to headwater groundwater sources assuming dominance. The significant temporally and spatially dynamic influence of these groundwater–surface‐water interactions was therefore seen to have important implications for both catchment functioning and instream ecology. Copyright © 2004 John Wiley & Sons, Ltd.
Hydrological influences on hyporheic water quality: implications for salmon egg survival
Authors: Malcolm I. A. , Soulsby C. , Youngson A. F. , Hannah D. M. , McLaren I. S. , Thorne A.
Published: March 2004

The spatial and temporal variability of groundwater–surface‐water (GW–SW) interactions was investigated in an intensively utilized salmon spawning riffle. Hydrochemical tracers, were used along with high‐resolution hydraulic head and temperature data to assess hyporheic dynamics. Surface and subsurface hydrochemistry were monitored at three locations where salmon spawning had been observed in previous years. Temperature and hydraulic head were monitored in three nests of three piezometers located to characterize the head, the run and the tail‐out of the riffle feature. Hydrochemical gradients between surface and subsurface water indicated increasing GW influence with depth into the hyporheic zone. Surface water was characterized by high dissolved oxygen (DO) concentrations, low alkalinity and conductivity. Hyporheic water was generally characterized by high levels of alkalinity and conductivity indicative of longer residence times, and low DO, indicative of reducing conditions. Hydrochemical and temperature gradients varied spatially over the riffle in response to changes in local GW–SW interactions at the depths investigated. Groundwater inputs dominated the head and tail of the riffle. The influence of SW increased in the area of accelerating flow and decreasing water depth through the run of the riffle. Temporal GW–SW interactions also varied in response to changing hydrological conditions. Gross changes in hyporheic hydrochemistry were observed at the weekly scale in response to changing flow conditions and surface water inputs to the hyporheic zone. During low flows, caused by freezing or dry weather, hyporheic hydrochemistry was dominated by GW inputs. During higher flows hyporheic hydrochemistry indicated that SW contributions increased. In addition, high‐resolution hydraulic head data indicated that rapid changes in GW–SW interactions occurred during hydrological events. The spatial, and possibly the temporal, variability of GW–SW interactions had a marked effect on the survival of salmon ova. It is concluded that hyporheic dynamics and their effect on stream ecology should be given increased consideration by fisheries and water resource managers. Copyright © 2004 John Wiley & Sons, Ltd.
Estimating the distribution of snow water equivalent and snow extent beneath cloud cover in the Salt–Verde River basin, Arizona
Authors: Molotch N. P. , Fassnacht S. R. , Bales R. C. , Helfrich S. R.
Published: March 2004

The temporal and spatial continuity of spatially distributed estimates of snow‐covered area (SCA) are limited by the availability of cloud‐free satellite imagery; this also affects spatial estimates of snow water equivalent (SWE), as SCA can be used to define the extent of snow telemetry (SNOTEL) point SWE interpolation. In order to extend the continuity of these estimates in time and space to areas beneath the cloud cover, gridded temperature data were used to define the spatial domain of SWE interpolation in the Salt–Verde watershed of Arizona. Gridded positive accumulated degree‐days (ADD) and binary SCA (derived from the Advanced Very High Resolution Radiometer (AVHRR)) were used to define a threshold ADD to define the area of snow cover. The optimized threshold ADD increased during snow accumulation periods, reaching a peak at maximum snow extent. The threshold then decreased dramatically during the first time period after peak snow extent owing to the low amount of energy required to melt the thin snow cover at lower elevations. The area having snow cover at this later time was then used to define the area for which SWE interpolation was done. The area simulated to have snow was compared with observed SCA from AVHRR to assess the simulated snow map accuracy. During periods without precipitation, the average commission and omission errors of the optimal technique were 7% and 11% respectively, with a map accuracy of 82%. Average map accuracy decreased to 75% during storm periods, with commission and omission errors equal to 11% and 12% respectively. The analysis shows that temperature data can be used to help estimate the snow extent beneath clouds and therefore improve the spatial and temporal continuity of SCA and SWE products. Copyright © 2004 John Wiley & Sons, Ltd.
Evidence for influence of mineral weathering on stream water sulphate in Vermont and New Hampshire (USA)
Authors: Bailey S. W. , Mayer B. , Mitchell M. J.
Published: March 2004

Mass balance studies in forested catchments in the northeastern USA show that S losses via streamwater SO42− exceed measured atmospheric S inputs. Possible sources of the excess S loss include underestimated dry deposition, mineralization of organic S in soils, desorption of soil sulphate, oxidation of recently formed sulphides and mineral weathering. Evaluating the relative contribution of these sources and processes to SO42− export is important to our understanding of S cycling as well as to policy makers in their evaluation of the efficacy of S emission controls. In order to evaluate the potential for mineral weathering contributions to SO42− export, we measured concentration and isotopic composition (δ34S and δ18O) of SO42− in stream water, and concentration and δ34S values of four S fractions in bedrock and soil parent material in catchments of varying geological composition. Geological substrates with low S concentrations were represented by catchments underlain by quartzite and granite, whereas geological substrates with high S concentrations were represented by catchments underlain by sulphidic slate, schist and metavolcanic rocks. Catchments with S‐poor bedrock had stream‐water SO42− concentrations <100 µeq L−1 and isotopic values consistent with those of atmospheric SO42− that had been cycled through the organic soil pool. Catchments with S‐rich bedrock had stream‐water SO42− concentrations ranging from 56 to 229 µeq L−1. Isotopic values deviated from those of SO42− in atmospheric deposition, clearly indicating a mineral weathering source in some cases, whereas in others spatial variability of mineral δ34S values precluded the isotopic detection of a weathering contribution. These results, along with evidence suggesting formation of secondary sulphate minerals in bedrock weathering rinds, indicate that mineral weathering may be an important source of S in the surface waters of some forested catchments in the northeastern USA. Copyright © 2004 John Wiley & Sons, Ltd.
Application of spectral analysis to determine hydraulic diffusivity of a sandy aquifer (Pingtung County, Taiwan)
Authors: Shih David Ching‐Fang , Lin Gwo‐Fong
Published: March 2004

The study demonstrates spectral relationships in the time–frequency domain for one‐dimensional groundwater flow in aquifers bounded by fluctuating boundaries. By nature, the solutions of spectral equations are non‐linear complex functions. To determine hydraulic diffusivity in the governing equations, it is required that the data are collected from the spectra of water levels at the fluctuating boundaries and observation wells. Hydraulic diffusivity thus can be obtained by an iterative inverse approach. This paper presents an application in Pingtung County of Taiwan to determine the hydraulic diffusivity of a sandy aquifer under confined conditions. Spectral density function of water level obtained from tidal boundaries and observation wells are used to approximate hydraulic diffusivity, which yields an averaged value of 1·26 × 106 m2/h. Copyright © 2004 John Wiley & Sons, Ltd.
HERV‐K(HML‐2) GAG/ENV antibodies as indicator for therapy effect in patients with germ cell tumors
Authors: Kleiman Anna , Senyuta Natalia , Tryakin Alexey , Sauter Marlies , Karseladze Apollon , Tjulandin Sergei , Gurtsevitch Vladimir , Mueller‐Lantzsch Nikolaus
Published: March 2004

Germ cell tumors (GCT) are strictly associated with the expression of HERV‐K(HML‐2) proviruses, and the majority of GCT patients produce antibodies to structural proteins of these proviruses. The objective of our study was to determine the significance of the serological response to HERV‐K(HML‐2) Gag and Env proteins for diagnosis, management of GCT patients and estimation of the therapy success. The data document a strong association of HERV‐K(HML‐2) antibodies and the clinical manifestation of the disease and therapy success. HERV‐K(HML‐2) antibodies seem to have an important diagnostic value as well as indicator of chemotherapy success. © 2004 Wiley‐Liss, Inc.
Limited relevance of the CHEK2 gene in hereditary breast cancer
Authors: Dufault Michael R. , Betz Beate , Wappenschmidt Barbara , Hofmann Wera , Bandick Katrin , Golla Astrid , Pietschmann Andrea , Nestle‐Krämling Caroline , Rhiem Kerstin , Hüttner Christine , von Lindern Celia , Dall Peter , Kiechle Marion , Untch Michael , Jonat Walter , Meindl Alfons , Scherneck Siegfried , Niederacher Dieter , Schmutzler Rita K. , Arnold Norbert
Published: March 2004

To establish the importance of CHEK2 mutations for familial breast cancer incidence in the German population, we have screened all 14 of the coding exons in 516 families negative for mutations in both the BRCA1 and BRCA2 genes. We found 12 distinct variants in 30 unrelated patients (5.81%), including 5 that are novel and an additional 4 found for the first time in breast cancer. These aberrations were evaluated in 500 healthy women aged over 50 years and in the case of the 2 exon 10 mutations, 1100delC and 1214del4bp, in 1315 randomized healthy controls. According to our results, a statistically significant association for the exon 10 mutations was observed (p = 0.006). The prevalence of the 1100delC mutation in the German population, however, is significantly lower than those reported for other Caucasian populations both in familial breast cancer patients (1.6%) and controls (0.5%), and shows independent segregation with breast cancer in 2 of 4 families analyzed. The remaining 10 variants were more abundant in patients (21) compared to the controls (12) although the difference was not statistically significant. Interestingly, we found no increased breast cancer risk associated with the splice site mutation IVS2+1G→A or the most common missense mutation I157T, which account for more than half (12/21) of the variants observed in patients. The low prevalence and penetrance of the exon 10 deletion mutations together with no, or an uncertain elevation in risk for other CHEK2 mutations suggests a limited relevance for CHEK2 mutations in familial breast cancer. Further evaluation of the unique variants observed in breast cancer is required to determine if they may play a role in a polygenic model of familial breast cancer. Nevertheless, it seems premature to include CHEK2 screening in genetic testing. © 2004 Wiley‐Liss, Inc.
The use of digitized endoscopic imaging of 5‐ALA‐induced PPIX fluorescence to detect and diagnose oral premalignant and malignant lesions in vivo
Authors: Zheng Wei , Olivo Malini , Soo Khee Chee
Published: March 2004

5‐aminolevulinic acid (ALA)‐induced protoporphyrin IX (PPIX) fluorescence has shown an outstanding sensitivity for the assessment of oral lesions, but its application was hampered by low specificity due to the high false‐positive rates. The purpose of our study was to explore the feasibility of quantifying PPIX fluorescence images to improve the diagnostic specificity for detecting early oral lesions in vivo. A digitized 5‐ALA‐mediated endoscopic imaging system was utilized to acquire PPIX fluorescence images from in vivo oral tissues. Forty‐nine patients (118 biopsies) with known or suspected premalignant or malignant oral lesions were recruited for ALA‐PPIX fluorescence endoscopic imaging. The red and blue channels of PPIX fluorescence images were digitized and stored for fluorescence quantification. The red‐to‐blue intensity ratios were calculated from the fluorescence images to correlate with histologic findings of the biopsies. The results showed that normal oral mucosa exhibited blue color of the back‐scattered excitation light in the fluorescence images, whereas the suspicious lesions displayed bright reddish fluorescence. Applying the red‐to‐blue intensity ratio (IR/IB) as a diagnostic algorithm yielded a sensitivity of 92% and 98%, and specificity of 96% and 96%, for separating benign tissue from dysplasia, and cancer tissue, respectively, and a sensitivity and specificity of 98% and 92%, respectively, for differentiating cancer tissue from dysplasia in the oral cavity. Our study demonstrates that quantifying ALA‐PPIX fluorescence endoscopic images associated with the red‐to‐blue intensity ratio as a diagnostic algorithm can provide good differentiation between the different stages of oral premalignancy and malignancy (p<0.0001, unpaired 2‐sided Student's t‐test), and thus has a potential to significantly improve the noninvasive diagnosis and evaluation of early oral neoplasia in vivo. © 2004 Wiley‐Liss, Inc.
Smoking and the risk of breast cancer among carriers of BRCA mutations
Authors: Ghadirian Parviz , Lubinski Jan , Lynch Henry , Neuhausen Susan L. , Weber Barbara , Isaacs Claudine , Baruch Ruth‐Gershoni , Randall Susan , Ainsworth Peter , Freidman Eitan , Horsman Douglas , Tonin Patricia , Foulkes William D. , Tung Nadine , Sun Ping , Narod Steven A.
Published: March 2004

The effect of cigarette smoking on the risk of breast cancer is controversial, although most studies show little or no effect. It has been suggested that smoking may reduce the risk of developing hereditary breast cancer. We completed a case‐control study on 1,097 women with breast cancer who were BRCA1 or BRCA2 mutation carriers and 1,097 age‐matched controls with a mutation in the same gene but without breast cancer. There were no statistically significant differences between the cases and controls in terms of the number of current and ex‐smokers (41.2% and 40.4%, respectively) or the age at smoking commencement (18.2 years and 18.5 years, respectively). There were no statistically significant differences between cases and controls regarding beginning smoking within 5 years of menarche (OR = 1.03; 95% CI 0.83 to l.28) or before the first pregnancy (OR = 1.09; 95% CI = 0.90 to 1.33). In conclusion, contrary to our previous report, smoking does not appear to be a risk factor for breast cancer among carriers of BRCA mutations. © 2004 Wiley‐Liss, Inc.
CNTO 95, a fully human monoclonal antibody that inhibits αv integrins, has antitumor and antiangiogenic activity in vivo
Authors: Trikha Mohit , Zhou Zhao , Nemeth Jeffrey A. , Chen Qiming , Sharp Celia , Emmell Eva , Giles‐Komar Jill , Nakada Marian T.
Published: March 2004

Integrins of the αv family, such as αvβ3 and αvβ5, are implicated in tumor‐induced angiogenesis; but their role in tumor growth has not been fully explored. CNTO 95 is a fully human antibody that recognizes the αv family of integrins and is likely to be less immunogenic in humans compared to chimeric or humanized antibodies. CNTO 95 bound to purified αvβ3 and αvβ5 with a Kd of approximately 200 pM and to αv integrin–expressing human cells with a Kd of 1–24 nM. In vitro, CNTO 95 inhibited human melanoma cell adhesion, migration and invasion at doses ranging 7–20 nM. In a rat aortic ring sprouting assay, CNTO 95 (approx. 70 nM) completely inhibited sprouting. Using a human melanoma xenograft model in nude mice wherein CNTO 95 recognized αvβ3 and αvβ5 on human tumor cells but not mouse angiogenic integrins, CNTO 95 (10 mg/kg, 3 times/week) inhibited growth of human melanoma tumors in nude mice by approximately 80% (p = 0.0005), suggesting that CNTO 95 inhibited human tumor growth independently of its antiangiogenic activity. In a nude rat human xenograft model where CNTO 95 binds and blocks both tumor and host integrins, this antibody (10 mg/kg once/week) reduced final tumor weight by >99% (p < 0.0001). Based on these preclinical data, a dose‐escalating phase I clinical trial in cancer patients has been initiated. To our knowledge, CNTO 95 is the first fully human MAb to αv integrins that has potent antitumor and antiangiogenic properties in in vivo preclinical models. © 2004 Wiley‐Liss, Inc.
Immortalization without neoplastic transformation of human mesenchymal stem cells by transduction with HPV16 E6/E7 genes
Authors: Hung Shih‐Chieh , Yang Den‐Mei , Chang Ching‐Fang , Lin Ruey‐Jen , Wang Jih‐Shiuan , Low‐Tone Ho Larry , Yang Wen K.
Published: March 2004

hMSCs derived from bone marrow are useful as a species‐specific cell culture system for studying cell lineage differentiation and tissue remodeling. However, hMSCs usually have a short in vitro life span due to replicative senescence. We therefore used a high dose of retroviral vector LXSN‐16E6E7 to transduce hMSCs of an aging donor and obtained an actively proliferating cell line, designated KP‐hMSCs, which expressed HPV16 E6/E7 mRNA. Whereas parental hMSCs ceased to grow after 30 PDs, KP‐hMSCs could be propagated beyond 100 PDs. With culture procedures to avoid selection pressure and crowded cell growth, KP‐hMSCs showed no signs of neoplastic transformation as examined by soft‐agar anchorage‐independent growth and NOD‐SCID mouse tumorigenicity assays. KP‐hMSCs gave similar cytofluorimetric profiles of 31 CD markers to those of the parental primary hMSCs, except with some morphologic changes and expansion of an originally very minor CD34dimCD38+CD50+ cell population. Upon exposure to specific stimulating conditions in vitro, KP‐hMSCs could respond and differentiate along the mesenchymal (bone, fat and cartilage) and nonmesenchymal (neuron) cell lineages. Our results indicated that hMSCs could be immortalized by transduction with HPV16 E6/E7, maintained without neoplastic transformation by careful culture procedures and thus useful for stem cell research and clinical application. © 2004 Wiley‐Liss, Inc.
Reduced levels of XPA, ERCC1 and XPF DNA repair proteins in testis tumor cell lines
Authors: Welsh Carey , Day Roger , McGurk Claire , Masters John R.W. , Wood Richard D. , Köberle Beate
Published: March 2004

Over 80% of patients with advanced metastatic testis tumors can be cured using cisplatin‐based combination chemotherapy. This is unusual as metastatic cancer in adults is usually incurable. Cell lines derived from testis tumors retain sensitivity to cisplatin in vitro. We previously investigated 2 testis tumor cell lines with a low capacity to remove cisplatin‐induced DNA damage and found that they had low levels of the DNA nucleotide excision repair proteins XPA, ERCC1 and XPF. To determine whether low levels of XPA, ERCC1 and XPF proteins are characteristic of testis tumor cell lines, we investigated 35 cell lines derived from cancers to determine whether groups of cell lines from diverse tissue origins differ from one another in constitutive levels of these NER proteins. Quantitative immunoblotting was used to compare groups of cell lines representing prostate, bladder, breast, lung, cervical, ovarian and testis cancers. Only the 6 testis tumor cell lines showed significantly lower mean levels of XPA (p = 0.001), XPF (p = 0.001) and ERCC1 (p = 0.004) proteins from the other groups. Our results encourage further investigation of the possibility that low levels of these nucleotide excision repair proteins could be related to the favorable response of testis tumors to cisplatin‐based chemotherapy. © 2004 Wiley‐Liss, Inc.
Alterations in tropomyosin isoform expression in human transitional cell carcinoma of the urinary bladder
Authors: Pawlak Geraldine , McGarvey Terence W. , Nguyen Trang B. , Tomaszewski John E. , Puthiyaveettil Raghunath , Malkowicz S. Bruce , Helfman David M.
Published: March 2004

Previous studies of transformed rodent fibroblasts have suggested that specific isoforms of the actin‐binding protein tropomyosin (TM) could function as suppressors of transformation, but an analysis of TM expression in patient tumor tissue is limited. The purpose of our study was to characterize expression of the different TM isoforms in human transitional cell carcinoma of the urinary bladder by immunohistochemistry and Western blot analysis. We found that TM1 and TM2 protein levels were markedly reduced and showed >60% reduction in 61% and 55% of tumor samples, respectively. TM5, which was expressed at very low levels in normal bladder mucosa, exhibited aberrant expression in 91% of tumor specimens. The Western blot findings were confirmed by immunohistochemical analysis in a number of tumors. We then investigated the mechanism underlying TM expression deregulation, in the T24 human bladder cancer cell line. We showed that levels of TM1, TM2 and TM3 are reduced in T24 cells, but significantly upregulated by inhibition of the mitogen‐activated protein kinase‐signaling pathway. In addition, inhibition of this pathway was accompanied by restoration of stress fibers. Overall, changes in TM expression levels seem to be an early event during bladder carcinogenesis. We conclude that alterations in TM isoform expression may provide further insight into malignant transformation in transitional cell carcinomas of the bladder and may be a useful target for early detection strategies. © 2004 Wiley‐Liss, Inc.
Using modelling to improve operational performance in the Cu Chi irrigation system, Vietnam
Authors: George Biju A. , Malano Hector M. , Tri Vo Khac , Turral Hugh
Published: March 2004

The Cu Chi irrigation system in Vietnam forms part of the Dau Tieng reservoir system in the Dong Nai basin. Water competition in this system is intensifying as a result of increasing urban and environmental demand. System operation modelling using the Irrigation Main System OPeration (IMSOP) model was applied to the Cu Chi irrigation system, Vietnam, to improve its operational performance. The operation improvement process involved the comparison of supply and demand during two years, 2001 and 2002, revealing that the system's operation was carried out without consideration of the crop water demand and proper canal regulation. The retrospective operational analysis revealed a substantial amount of oversupply and inequity of distribution. Two new operation strategies were selected after a simulation analysis and evaluated in the field. The new scenarios showed a more equitable distribution of water in the system. The average pre‐intervention supply–demand ratio which ranged from 1.68 to 2.51 was reduced to 1.20 for continuous flow and 1.17 for rotational operation. A comprehensive analysis of water reallocation to Ho Chi Minh City for urban and industrial purposes and its effect on the security of supply on the Cu Chi system was also undertaken. Copyright © 2004 John Wiley & Sons, Ltd.
l‐Arginine ameliorates oxidative stress in alloxan‐induced experimental diabetes mellitus
Authors: El‐Missiry M. A. , Othman A. I. , Amer M. A.
Published: March 2004

Oxidative stress occurs in diabetic patients and experimental models of diabetes. The ability ofl‐arginine to ameliorate the oxidative stress and metabolic changes after treatment with alloxan was investigated in rats. Adult male rats were injected intraperitoneally with 100 mg kg−1of alloxan to produce experimental oxidative stress characteristic of diabetes mellitus. Hyperglycaemia and hypercholesterolaemia were observed in serum after 7 days of alloxan treatment. This was associated with a depression of glutathione (GSH) concentration as well as superoxide dismutase (SOD) and catalase (CAT) activities in the liver and brain. In addition, the thiobarbituric acid‐reactive substances (TBARS) were significantly elevated, indicating increased lipid peroxidation and oxidative stress in the same tissues. Administration of 100 mg kg−1l‐arginine for 7 days either before or after alloxan injection significantly ameliorated the oxidative stress evidenced by a lower TBARS and a higher level of the endogenous GSH concentration and SOD and CAT activities than alloxan‐treated rats. These effects were paralleled by marked protection and partial prophylaxis against alloxan‐induced hyperglycaemia and cholesterolaemia. Thus, these results showed that exogenously administeredl‐arginine might improve the clinical manifestation of diabetes mellitus and decrease the oxidative stress in the liver and brain. In addition, the study supports the beneficial effect ofl‐arginine, which might be attributed to its direct, NO‐dependent antioxidant capacity and/or NO‐independent pathways. Copyright © 2004 John Wiley & Sons, Ltd.
Glioma cell antioxidant capacity relative to reactive oxygen species produced by dopamine
Authors: Mazzio Elizabeth A. , Soliman Karam F. A.
Published: March 2004

The pathology of Parkinson's disease involves oxidative damage to dopaminergic neurons of the substantia nigra. Oxidation of the dopamine (DA) neurotransmitter itself may contribute to the generation of a reactive oxygen species (ROS) and subsequent neurodegeneration. Glia cells can either exacerbate injury or exert protective properties on local neurons in the brain. We investigate glial antioxidant enzyme systems relative to ROS generated during cytokine activation, monoamine oxidase (MAO) activity and autoxidation of DA in glioma cells. Rat C6 glioma cells stimulated with lipopolysaccharide Escherichia coli 0111:B4 and interferon gamma (LPS/IFN‐g) produced high levels of nitric oxide (241 nmol mg−1 protein 24 h−1) but not superoxide (O−2) or hydrogen peroxide (H2O2). Basal C6 cells exhibited a rapid and robust capacity to remove exogenous H2O2 within minutes. Preincubation with sodium azide but not buthionine‐[S, R]‐sulfoximine attenuated this response, indicating catalase as the primary enzyme responsible for this effect. The glioma catalase reaction rate was slightly attenuated by exposure to LPS/IFN‐g for 24 h. However, the reduction in catalase activity was not due to nitric oxide, because both the supernatant and sodium nitroprusside had no effect on isolated catalase enzyme activity. Hydrogen peroxide was produced only through substrate‐driven MAO activity in prepared lysate. However, the quantity of H2O2 produced per unit time (0.46 nmol mg−1 protein min−1) was negligible compared with the enormous capacity for its removal by catalase (213.9 nmol mg−1 protein min−1) (≥462× greater). Similarly, H2O2 generated by DA autoxidation per unit time (0.28 nmol mg−1 protein equiv. min−1), was rapidly dissolved by glioma cells at high capacity (≥750× greater). In conclusion, C6 cells produce nitric oxide under cytokine/endotoxin‐stimulated conditions. Moreover, C6 cells exhibit a dynamic H2O2 scavenging capacity, with ample facility to dispose of the peroxide generated by both MAO activity and spontaneous DA autoxidation. Copyright © 2004 John Wiley & Sons, Ltd.
Amelioration of sulfur mustard skin injury following a topical treatment with a mixture of a steroid and a NSAID
Authors: Dachir Shlomit , Fishbeine Eliezer , Meshulam Yakov , Sahar Rita , Chapman Shira , Amir Adina , Kadar Tamar
Published: March 2004

The ability to ameliorate sulfur mustard (HD)‐induced oedema by treatment with anti‐inflammatory drugs was reported previously after screening four steroids and four non‐steroidal anti‐inflammatory drugs (NSAIDs) using the mouse ear vesicant model. Following the screening study, one steroid and one NSAID (Adexone and Voltaren) were selected as the most effective, and a mixture of the two was chosen for the present more extensive research. The effect of the combined treatment on clinical, biochemical and histopathological parameters following HD insult was studied. Mice ears were exposed to 0.2 µl of HD for 10 min to produce a moderate skin injury. Oedema development peaked ca. 48 h following exposure, as determined by weighing ear biopsies. Histological observations at that time exhibited damage to the epidermis and dermis. An increase in prostaglandin E (PGE) was measured in skin homogenates, starting 8 h following exposure and lasting at least up to 48 h post‐exposure.

A topical treatment using the above anti‐inflammatory mixture significantly reduced inflammatory parameters when applied up to 4 h following exposure. These parameters included extent of oedema, levels of PGE, area of clinical damage and extent of cytotoxic injury (vesications and damaged epithelial cells).

Thus, a combination of a steroid and NSAID was found to be effective in reducing the intensity of HD skin injury and possibly shortening the time to full recovery. The treatment, however, did not prevent completely the ensuing cytotoxic processes in the epithelial layer. Copyright © 2004 John Wiley & Sons, Ltd.
In vivo exposure of Dreissena polymorpha mussels to the quinones menadione and lawsone: menadione is more toxic to mussels than lawsone
Authors: Osman A. M. , Rotteveel S. , den Besten P. J. , van Noort P. C. M.
Published: March 2004

The principal aim of this study was to assess whether the two quinones, menadione (2‐methyl‐1,4‐naphthoquinone) and lawsone (2‐hydroxy‐1,4‐naphthoquinone), elicit differential toxicity in mussels as has been reported for higher organisms. Therefore, the effects of short‐term (48 h) and long‐term (20 days) exposure of the two quinones at concentrations of 0.56 and 1 mg l−1 to zebra mussels, Dreissena polymorpha, under laboratory conditions were studied. After the short‐term exposure, the specific activities of the two‐electron quinone oxidoreductase (DT‐diaphorase) and the one‐electron catalysing quinone reductases NADPH‐cytochrome c reductase and NADH‐cytochrome c reductase were determined in the gills and the rest of the soft tissues (soft mussel tissues minus the gills) of both treated and control mussels. At the higher concentrations of menadione and lawsone used, a significant reduction of the activity of NADPH‐cytochrome c reductase in the gills and in the rest of the soft mussel tissues (by 33–34% and 31–43%, respectively) was observed. The activities of DT‐diaphorase and NADH‐cytochrome c reductase were not significantly affected.

Interestingly, DT‐diaphorase was observed in the gills, an organ requiring protection against antioxidants. Furthermore, a single‐cell electrophoretic assay (comet assay) performed with gill cells to assess DNA damage by the quinones did not show any significant difference between the treated and the control organisms. This indicates that the formation of reactive species by the quinone metabolism in vivo in the mussels was possibly suppressed through the concerted action of DT‐diaphorase and antioxidant enzymes. The results of in vitro experiments with gill extracts confirmed the protective role of DT‐diaphorase. The rate of the two‐electron quinone reduction was found to be five times that of the one‐electron quinone reduction.

The results of the long‐term exposure unambiguously demonstrated that in mussels menadione, unlike in higher organisms, is more toxic than lawsone. The lack of detectability of xanthine oxidase in the mussel tissues could explain the comparatively lower toxicity of lawsone in the invertebtrate, lending support to a previous suggestion that xanthine oxidase might be responsible for the mechanism of toxicity of lawsone in higher organisms in vivo. Copyright © 2004 John Wiley & Sons, Ltd.
In vitro protection of plasma cholinesterases by metoclopramide from inhibition by mipafox
Authors: Petroianu G. , Kühn F. , Arafat K. , Zuleger K. , Missler A.
Published: March 2004

Metoclopramide (MCP) is a dopamine receptor antagonist and serotonin receptor agonist widely used as an antiemetic and gastric prokinetic drug. In addition MCP is a reversible inhibitor of cholinesterases from human central nervous system and blood. Metoclopramide may have a cholinesterase protective effect against inhibition by organophosphates. The purpose of the study was to quantify in vitro, by means of the ic50 shift, the extent of MCP conferred protection, using mipafox (MPFX) as an inhibitor. Mipafox is a neuropathic organophosphate. Cholinesterase activities (with acetylthiocholine [ChE‐A] and butyrylthiocholine [ChE‐B] as substrates) in human plasma were measured photometrically in the presence of different MPFX concentrations and the ic50 was calculated. Determinations were repeated in the presence of increasing MCP concentrations. It appears that the shift induced by the presence of MCP increases with the MCP concentration in a linear manner. In the presence of a clinically easily achievable plasma concentration of 1 µM MCP, the ic50 of MPFX for cholinesterase ‘shifts’ by a factor of ca. 3–6. The protective effect of MCP on cholinesterase could be of practical relevance in the treatment of organophosphate poisoning. We conclude that in vivo testing of MCP as an organophosphate protective agent is warranted. Copyright © 2004 John Wiley & Sons, Ltd.
Existence question for difference families and construction of some new families
Authors: Martínez Luis , Ðoković Dragomir Z̆. , Vera‐López Antonio
Published: March 2004

We establish some properties of mixed difference families. We obtain some criteria for the existence of such families and a special kind of multipliers. Several methods are presented for the construction of difference families by using cyclotomy and genetic algorithms. © 2004 Wiley Periodicals, Inc. J Combin Designs 12: 256–270, 2004.
Drug‐resistant breast carcinoma (MCF‐7) cells are paradoxically sensitive to apoptosis
Authors: Chen Jack S.K. , Konopleva Marina , Andreeff Michael , Multani Asha S. , Pathak Sen , Mehta Kapil
Published: March 2004

The purpose of this study was to determine whether expression of tissue transglutaminase (TG2) and caspase‐3 proteins in drug‐resistant breast carcinoma MCF‐7/DOX cells would render these cells selectively susceptible to apoptotic stimuli. Despite high resistance to multidrug resistance (MDR)‐related drug, doxorubicin (≥150‐fold), the MCF‐7/DOX cells were extremely sensitive to apoptotic stimuli. Thus, calcium ionophore, A23187 (A23187) and the protein kinase C inhibitor staurosporine (STS) each induced rapid and time‐dependent apoptosis in MCF‐7/DOX cells. The apoptosis induced by either agent was accompanied by caspase‐3 activation and other downstream changes that are typical of cells undergoing apoptosis. The alterations upstream of caspase‐3 activation, however, such as loss in mitochondrial membrane potential (ΔΨ), release of cytochrome c, and activation of caspase‐8, and caspase‐9, were detected only in STS‐treated cells. The A12387 failed to induce any of the caspase‐3 upstream changes, implying that A23187‐induced apoptosis may utilize one or more novel upstream pathways leading to the activation of caspase 3. In summary, these data demonstrate that MCF‐7/DOX cells are much more sensitive to apoptotic stimuli than previously thought and that A23187‐induced apoptosis may involve some novel, yet unidentified, upstream pathway that leads to the activation of caspase‐3 and other downstream events. © 2004 Wiley‐Liss, Inc.
Excision of Sleeping Beauty transposons: parameters and applications to gene therapy
Authors: Liu Geyi , Aronovich Elena L. , Cui Zongbin , Whitley Chester B. , Hackett Perry B.
Published: March 2004

A major problem in gene therapy is the determination of the rates at which gene transfer has occurred. Our work has focused on applications of the Sleeping Beauty (SB) transposon system as a non‐viral vector for gene therapy. Excision of a transposon from a donor molecule and its integration into a cellular chromosome are catalyzed by SB transposase. In this study, we used a plasmid‐based excision assay to study the excision step of transposition. We used the excision assay to evaluate the importance of various sequences that border the sites of excision inside and outside the transposon in order to determine the most active sequences for transposition from a donor plasmid. These findings together with our previous results in transposase binding to the terminal repeats suggest that the sequences in the transposon‐junction of SB are involved in steps subsequent to DNA binding but before excision, and that they may have a role in transposase–transposon interaction. We found that SB transposons leave characteristically different footprints at excision sites in different cell types, suggesting that alternative repair machineries operate in concert with transposition. Most importantly, we found that the rates of excision correlate with the rates of transposition. We used this finding to assess transposition in livers of mice that were injected with the SB transposon and transposase. The excision assay appears to be a relatively quick and easy method to optimize protocols for delivery of genes in SB transposons to mammalian chromosomes in living animals. Copyright © 2004 John Wiley & Sons, Ltd.
Genetic engineering of cytolytic T lymphocytes for adoptive T‐cell therapy of neuroblastoma
Authors: Gonzalez Sergio , Naranjo Araceli , Serrano Lisa M. , Chang Wen‐Chung , Wright Christine L. , Jensen Michael C.
Published: March 2004

Background

Disease relapse is the leading cause of mortality for children diagnosed with disseminated neuroblastoma. The adoptive transfer of tumor‐specific T cells is an attractive approach to target minimal residual disease following conventional therapies. We describe here the genetic engineering of human cytotoxic T lymphocytes (CTL) to express a chimeric immunoreceptor for re‐directed HLA‐independent recognition of neuroblastoma.

Methods

The CE7R chimeric immunoreceptor was constructed by PCR splice overlap extension and is composed of a single‐chain antibody extracellular domain (scFv) derived from the L1‐CAM‐specific murine CE7 hybridoma fused to human IgG1 hinge‐Fc, the transmembrane portion of human CD4, and the cytoplasmic tail of huCD3‐ζ chain (scFvFc:ζ). Primary human T cells were genetically modified by naked DNA electrotransfer of plasmid expression vector CE7R‐pMG then analyzed by Western blotting, flow cytometry for CE7R expression and cell surface trafficking, 4‐h chromium release assay for re‐directed neuroblastoma lysis, and ELISA for tumor‐specific activation of cytokine production.

Results

CE7R is expressed as an intact chimeric protein that trafficks to the cell surface as a type I transmembrane protein. Primary human CE7R‐expressing CD8+ CTL clones specifically recognize human neuroblastoma tumor cells and are activated for tumor cell lysis and Tc1 cytokine production.

Conclusions

These data demonstrate the utility of CE7R for re‐directing the effector function of CTL to neuroblastoma and have provided the rationale to initiate a FDA‐authorized (BB‐IND#9149) pilot clinical trial to establish the feasibility and safety of adoptive transfer of autologous CE7R+CD8+ CTL clones to children with recurrent/refractory neuroblastoma. Copyright © 2004 John Wiley & Sons, Ltd.
In vivo gene electrotransfer into skeletal muscle: effects of plasmid DNA on the occurrence and extent of muscle damage
Authors: Durieux Anne‐Cécile , Bonnefoy Régis , Busso Thierry , Freyssenet Damien
Published: March 2004

Background

Understanding the mechanisms underlying gene electrotransfer muscle damage can help to design more effective gene electrotransfer strategies for physiological and therapeutical applications. The present study investigates the factors involved in gene electrotransfer associated muscle damage.

Methods

Histochemical analyses were used to determine the extent of transfection efficiency and muscle damage in the Tibialis anterior muscles of Sprague‐Dawley male rats after gene electrotransfer.

Results

Five days after gene electrotransfer, features of muscle degeneration and regeneration were consistently observed, thus limiting the extent of transfection efficiency. Signs of muscle degeneration/regeneration were no longer evident 21 days after gene electrotransfer except for the presence of central myonuclei. Neither the application of electrical pulses per se nor the extracellular presence of plasmid DNA per se contributed significantly to muscle damage (2.9 ± 1.0 and 2.1 ± 0.7% of the whole muscle cross‐sectional area, respectively). Gene electrotransfer of a plasmid DNA, which does not support gene expression, increased significantly muscle damage (8.7 ± 1.2%). When plasmid DNA expression was permitted (gene electrotransfer of pCMV‐β‐galactosidase), muscle damage was further increased to 19.7 ± 4.5%. Optimization of cumulated pulse duration and current intensity dramatically reduced gene electrotransfer associated muscle damage. Finally, mathematical modeling of gene electrotransfer associated muscle damage as a function of the number of electrons delivered to the tissue indicated that pulse length critically determined the extent of muscle damage.

Conclusion

Our data suggest that neither the extracellular presence of plasmid DNA per se nor the application of electric pulses per se contributes significantly to muscle damage. Gene electrotransfer associated muscle damage mainly arises from the intracellular presence and expression of plasmid DNA. Copyright © 2004 John Wiley & Sons, Ltd.
Recombinant bispecific antibodies for the targeting of adenoviruses to CEA‐expressing tumour cells: a comparative analysis of bacterially expressed single‐chain diabody and tandem scFv
Authors: Korn Tina , Nettelbeck Dirk M. , Völkel Tina , Müller Rolf , Kontermann Roland E.
Published: March 2004

We have generated two distinct recombinant bispecific antibody molecules for the retargeting of adenoviral vectors to CEA‐expressing tumour cells. These antibody molecules were produced by combining the antigen‐binding sites of a neutralising anti‐fibre knob scFv (S11) and an anti‐CEA antibody either in a single‐chain diabody format (scDb CEA‐S11) or a tandem scFv format (taFv CEA‐S11). In order to facilitate expression of taFv CEA‐S11 in bacteria we selected from a phage display library taFv molecules with an optimised linker that connects the two scFv fragments. ScDb CEA‐S11 and taFv CEA‐S11 were expressed and purified in soluble form from the bacterial periplasm with yields of approximately 100 µg per litre of bacterial culture. In vitro, both bispecific molecules mediated selective and enhanced transduction of CEA‐expressing tumour cells by recombinant adenoviruses. These assays did not reveal any differences in efficiency of adenoviral transduction by the two antibody formats. However, compared with taFv CEA‐S11, scDb CEA‐S11 exhibited a 2‐ to 3‐fold increased stability in human plasma at 37 °C. In summary, we could demonstrate that both formats are suitable for adenovirus targeting to tumour cells with similar biological activity in vitro. Copyright © 2004 John Wiley & Sons, Ltd.
Encapsulated transgene cells attenuate hypertension, cardiac hypertrophy and enhance renal function in Goldblatt hypertensive rats
Authors: Chen Li‐Guo , Wang Zheng‐Rong , Wan Chao‐Min , Chao Lee , Chao Julie , Xing Hao‐Yang
Published: March 2004

Background

The success of any gene‐therapy approach depends on the survival of the genetically engineered cells that are implanted in the patient to deliver the therapeutic product. Immunoisolation of nonautologous cells within a microcapsule is a unique approach for gene therapy.

Methods

We employed an immunoisolation device that protects nonautologous cells from destruction, to implant human atrial natriuretic peptide (hANP)‐producing Chinese hamster ovary (CHO) cells in two‐kidney, one‐clip (2K1C) hypertensive rats. CHO cells transfected with the plasmid CMV‐cANP were encapsulated in biocompatible polycaprolactone (PCL) capsules, and then the PCL capsules were implanted into 2K1C hypertensive rats intraperitoneally.

Results

The implantation of encapsulated hANP‐producing cells caused a significant delay of blood pressure (BP) increase 2 weeks post‐implantation and the effect lasted for more than 5 months. The implantation of encapsulated hANP‐producing cells also caused significant increases in renal blood flow (RBF), glomerular filtration rate (GFR), sodium output, urine excretion, and urinary cGMP levels. These beneficial effects were reflected morphologically by an attenuation of the glomerular sclerotic lesions, reduction in cardiomyocyte size, tubular injury and renal arterial thickening. Immunoreactive hANP can be detected in the blood of 2K1C rats after implantation of the PCL capsules containing hANP‐producing cells.

Conclusions

This study demonstrates the usefulness of encapsulated ANP gene transfected cells as a new tool for ANP gene delivery in studying renovascular hypertension and cardiovascular diseases. Thus, our results may have important implications for clinical use of transgene cells as therapeutic agents in the treatment of cardiovascular diseases. Copyright © 2004 John Wiley & Sons, Ltd.
Synthesis of deuterium‐labelled 6‐[5‐(4‐amidinophenyl)furan‐2‐yl]nicotinamidine and N‐alkoxy‐6‐{5‐[4‐(N‐alkoxyamidino)phenyl]‐ furan‐2‐yl}‐nicotinamidines
Authors: Ismail Mohamed A. , Boykin David W.
Published: March 2004

6‐[5‐(4‐Amidinophenyl)furan‐2‐yl]nicotinamidine‐d4 (5) was synthesized from 6‐[5‐(4‐cyanophenyl)furan‐2‐yl]nicotinonitrile‐d4 (3), through the bis‐O‐acetoxy‐amidoxime followed by hydrogenation. Compound 3 was prepared from 6‐(furan‐2‐yl)‐nicotinonitrile by a Heck coupling reaction with 4‐bromobenzonitrile‐d4, a product of selective cyanation reaction of 1,4‐dibromobenzene‐d4 with Cu(1)CN. Deuterium‐labelled N‐methoxy‐6‐{5‐[4‐(N‐methoxy‐amidinophenyl]‐furan‐2‐yl}‐nicotinamidines were prepared via methylation of their respective amidoximes with dimethyl sulfate‐d6 in aqueous sodium hydroxide in good yields. Copyright © 2004 John Wiley & Sons, Ltd.