bluuebunny/crossref_metadata_2025 huggingface dataset:
Anti‐CD30‐scFv‐Fc‐IL‐2 antibody‐cytokine fusion protein that induces resting NK cells to highly efficient cytolysis of Hodgkin's lymphoma derived tumour cells
Authors: Heuser Claudia , Guhlke Stefan , Matthies Alexander , Bender Hans , Barth Stefan , Diehl Volker , Abken Hinrich , Hombach Andreas
Published: March 2004

The pathogenesis of Hodgkin's disease (HD) is associated with the accumulation of functionally anergic T cells in the near vicinity of the malignant Hodgkin/Reed‐Sternberg (H/RS) cell. To stimulate locally the anti‐tumour immunity in Hodgkin's disease, we generated an anti‐CD30‐antibody‐interleukin‐2 fusion protein (HRS3‐scFv‐Fc‐IL‐2) that binds to CD30 constitutively expressed on H/RS cells. The fusion protein is composed of a CD30 binding domain (HRS3‐scFv) that is linked via the human IgG hinge‐CH2/CH3 domain to human IL‐2. The HRS3‐scFv‐Fc‐IL‐2 fusion protein is expressed as a 140 kDa homodimer, has binding specificities to both the CD30 antigen and the IL‐2 receptor and stimulates proliferation of preactivated T cells in vitro, demonstrating its IL‐2 bioactivity. After binding to CD30+ Hodgkin lymphoma cells, HRS3‐scFv‐Fc‐IL‐2 moreover induces resting NK cells, but not T cells, to lyse the lymphoma cells with high efficiency. Recruitment of resting NK cells towards a cytolytic immune response against CD30+ lymphoma cells has the potential to build up an effective anti‐tumour response despite of Hodgkin's disease associated T‐cell anergy and makes the HRS3‐scFv‐Fc‐IL‐2 fusion protein suitable for the specific immunotherapy of Hodgkin's lymphoma. © 2004 Wiley‐Liss, Inc.
Pigmentary characteristics, sun sensitivity and non‐Hodgkin lymphoma
Authors: Hughes Ann Maree , Armstrong Bruce K. , Vajdic Claire M. , Turner Jennifer , Grulich Andrew , Fritschi Lin , Milliken Sam , Kaldor John , Benke Geza , Kricker Anne
Published: March 2004

We report on pigmentary characteristics, sun sensitivity and some other possible risk factors for non‐Hodgkin lymphoma (NHL) in people 20–74 years of age. A statewide case‐control study was conducted in New South Wales, Australia, with population‐based sampling of cases (n = 704) and controls (n = 694). Risk of NHL was increased in subjects with hazel eyes (OR = 1.48; 95% CI = 1.07–2.04), very fair skin (OR = 1.44; 95% CI = 1.01–2.07) and poor ability to tan (OR = 1.70; 95% CI = 1.06–2.71). Risk with mild facial freckling as a child (OR = 0.77; 95% CI = 0.59–0.99) was reduced relative to that with no or moderate to severe freckling. Smokers were not at increased risk of NHL. A past history of treatment for skin cancer was associated with a slight nonsignificant increase in risk. Previous radiotherapy and chemotherapy were associated with 1.5‐ to 2‐fold increases in risk but with wide confidence intervals. These results provide weak support for the possibility that sun sensitivity or perhaps sun exposure increases risk of NHL. © 2004 Wiley‐Liss, Inc.
Hyperparathyroidism and subsequent incidence of breast cancer
Authors: Michels Karin B. , Xue Fei , Brandt Lena , Ekbom Anders
Published: March 2004

Preliminary data are available on the coexistence of primary hyperparathyroidism and breast carcinoma. To further understand the association between hyperparathyroidism and breast cancer, we conducted a record‐linkage study in Sweden using the Swedish Cancer Registry from 1958–1997. A total of 9,835 women who underwent surgery for primary parathyroid adenoma were followed to evaluate the hypothesis that a history of primary hyperparathyroidism increases the risk of subsequent breast cancer. During 99,929 person‐years of follow‐up, 331 cases of newly diagnosed breast cancer were reported. The number of expected breast cancers in this population was 260.0. This resulted in a standardized incidence ratio of 1.27 (95% confidence interval [CI] = 1.14–1.41). The relation persisted over time after the surgical removal of the parathyroid adenoma. Possible explanations for the observed association are a shared etiology including genetic and environmental factors such as early life radiation, and hypercalcemia after the overproduction of parathyroid hormone, which may increase breast cancer incidence. © 2004 Wiley‐Liss, Inc.
GIS tools and the design of irrigation canals
Authors: Depeweg Herman , Urquieta Elga Rocabado
Published: March 2004

The previous decades have shown an incredible progress in information and computer technology for among others the optimal planning and use of water and soil resources, especially after the introduction of geographic information systems (GIS), satellite remote sensing and global positioning systems. GIS is able to integrate spatial and non‐spatial geographical data for the management and planning of land and water resources. These recent developments provide new ways to collect, store and retrieve, evaluate and analyse a broad variety of data for a wide range of irrigation management aspects. They also may assist in specific planning and decision‐making processes in irrigation through the input, spatial analysis and output of relevant information. Furthermore, if GIS is employed together with other computer applications it becomes more powerful in various fields, for example in the design and construction of irrigation and drainage canals.

This article describes a procedure to design an irrigation canal and to optimise this design in view of earthwork. The procedure uses specific data processing and functions in GIS to facilitate the canal layout based on physical conditions and economic considerations. Digital information on topography, soils and natural drainage is used to identify the most suitable location for the canal. Digital elevation models (DEMs) are applied to prepare the horizontal and vertical canal alignment. Additionally, a computer model in Visual Basic has been developed to obtain an optimal hydraulic design by an iterative process. Next, the results of both packages are linked in a spreadsheet to optimise the canal layout in view of minimal earthwork. Copyright © 2004 John Wiley & Sons, Ltd.
Agricultural changes affecting water availability: from abundance to scarcity (Fucino Plain, central Italy)
Authors: Burri Ezio , Petitta Marco
Published: March 2004

The study reported in this paper analysed the significant agricultural changes that the Fucino Plain (central Italy) has experienced in the past few years and their impact on water availability. In the past, the Fucino Plain (surface area: over 200 km2) hosted the largest lake of central Italy. In the late 1800s, the lake was drained to be used as farmland. Recently, however, the fast change in agricultural activities (with the expansion of water‐intensive horticultural crops) and climate (decreasing precipitation and increasing temperature) has broken the environmental equilibrium of the plain, causing groundwater lowering, spring exhaustion, decreased canal discharge in summer and groundwater pollution risks connected with fertiliser and pesticide use.

New data on water usage and farming made it possible to develop a new “integrated” water budget of the plain, taking into account the impact of human activities on the water cycle. Monitoring of the main well fields, of crop distribution and of natural discharge in canals and springs, will enable better management of local water resources, reducing their exploitation during the irrigation season. Copyright © 2004 John Wiley & Sons, Ltd.
Mutagenicity of textile dye products
Authors: Schneider Klaus , Hafner Christoph , Jäger Ismene
Published: March 2004

Within an EU‐funded research project, 281 textile dye products in use at nine textile finishing companies from eight European countries were assessed for potential mutagenic properties. Most of the dyes belonged to the so‐called existing substances. Data sources considered were data published in the literature, unpublished industrial data provided by dye producing companies, and laboratory testing. Data on mutagenicity are virtually absent for many of the dyes. Unpublished test results performed on behalf of the dye manufacturing industry proved to be an important data source that is not accessible under usual circumstances. Four dye stuffs contained in seven dye products in use at the textile finishing companies were judged to be mutagenic, based on published data from the literature. Mutagenicity testing using Salmonella typhimurium, strains TA98 and TA100, revealed positive results for about 28% (15 out of 53) of the dye products investigated. Upon further testing with the mouse lymphoma assay (L5178Y/TK+/‐) 67% (6 out of 9) of Ames‐positive dyes proved to be mutagenic in this mammalian cell test. All data sources combined led to an overall assessment of 14 dye products out of 281 being mutagenic. For 16 there is a suspicion of mutagenicity due to positive responses in one test but 71 of the dye products are without any data on mutagenicity. This paper describes the data aggregation process, evaluation criteria and the overall assessment, and exemplifies controversial evaluations. Copyright © 2004 John Wiley & Sons, Ltd.
Cytogenetic risk assessment of etoposide from mouse bone marrow
Authors: Choudhury Ramesh C. , Palo Anil K. , Sahu Prajyoti
Published: March 2004

Increased clinical applications of the anticancer drug etoposide (a non‐intercalative epipodophyllotoxin derivative) and the frequent induction of a second malignancy, particularly leukaemia, in post‐etoposide‐treated cancer survivors warrant detailed genotoxicity testing of etoposide. The genotoxicity test results available on etoposide are either primarily in in vitro test systems or in lower organisms after treatment with unusually high doses, or after chronic exposures, having little extrapolative value to humans. Therefore, a cytogenetic risk assessment study on etoposide in mouse in vivo was undertaken after a low dose (in accordance with the human therapeutic dose) single exposure. The cytogenetic toxicity of etoposide was assessed from bone marrow of mouse at three separate endpoints: chromosomal aberration and mitotic index studies at 24 h post‐treatment and the micronucleus test (MNT) at 30 h post‐treatment. The flame drying technique using colchicine, hypotonic sodium citrate, methanol‐glacial acetic acid and Giemsa was followed for the preparation of slides for the metaphase chromosomal aberration and mitotic index studies and a simple technique was followed for the MNT. Although induction of chromosomal aberrations, excluding gaps, per 100 metaphases by 10 and 15 mg kg−1 etoposide was not significant statistically, 20 mg kg−1 of etoposide induced a significantly higher number of chromosomal aberrations in female (P ≤ 0.01) and male (P ≤ 0.05) mice. There was no significant change in the induced percentages of dividing cells by any of the doses of etoposide tested. The micronucleus induction also was not significant statistically with the lowest dose but it was significant in female (P ≤ 0.01) and male (P ≤ 0.05) mice that received 15 mg kg−1 etoposide and was highly significant (P ≤ 0.01) in both female and male mice that received 20 mg kg−1 etoposide. The results indicated the clastogenic action of etoposide in mouse bone marrow after a single treatment with such low doses. However, the drug did not interfere with cell cycle progression. Although it is a DNA‐non‐intercalating agent, etoposide is known for its interference in the activity of DNA topoisomerase IIα enzyme, particularly in the proliferative cells where the concentration and activity of the enzyme are greater. This might be the reason for the induction of leukaemia in post‐etoposide‐treated cancer survivors. Therefore, it has become absolutely necessary to make etoposide target‐specific, i.e. specific to the topoisomerase II enzymes of cancerous cells. Copyright © 2004 John Wiley & Sons, Ltd.
Effect of milk on intestinal fluid accumulation and renal injury following mercuric chloride ingestion in rats
Authors: Kamijo Yoshito , Hayashi Izumi , Yoshimura Kuniko , Soma Kazui
Published: March 2004

To investigate the effect of milk on intestinal fluid accumulation and renal injury following mercuric chloride (HgCl2) ingestion, 10 ml kg−1 of saline or 10 mg kg−1 of HgCl2 dissolved in 10 ml kg−1 of water or raw milk was administered enterally to rats and the mercury content in biological samples was determined by cold vapour atomic absorption spectrometry. Three hours after administration, the intestinal water content in rats that received HgCl2 in water (group S2) was significantly higher than in rats that received saline (group S1) (P < 0.01) or HgCl2 in milk (group S3) (P < 0.01). The amount of mercury detected per gram dry weight of small intestine was higher in group S2 than in group S3 (P < 0.05). Seventy‐two hours after administration, both the serum urea nitrogen and creatinine concentrations in rats that received HgCl2 in milk (group L3) were significantly higher than in rats that received saline (group L1) (P < 0.05) or HgCl2 in water (group L2) (P < 0.05). Mercury levels in many of the biological samples in group L3 were higher than in group L2 (P < 0.05). Milk may reduce the intestinal cytotoxicity of mercury but it promotes its absorption, which may lessen intestinal fluid accumulation but worsen renal injury. Copyright © 2004 John Wiley & Sons, Ltd.
Pulmonary toxicity of wollastonite in vivo and in vitro
Authors: Tátrai E. , Kováčiková Z. , Brózik M. , Six É.
Published: March 2004

Pulmonary toxicity of wollastonite has been studied in both in vivo long‐term sequential and in vitro methods in Sprague‐Dawley rats. Wollastonite was administered by intratracheal instillation and the lungs were examined after 1, 3 and 6 months by morphological methods. UICC crocidolite was applied as the positive control. In addition, the effects of both ﬁbres were examined in primary cultures of pulmonary alveolar macrophages and type II pneumocytes to determine the effects of the ﬁbres on the membranes of these cells, the activity of Cu,Zn/superoxide dismutase and the redox system and the release of proinﬂammatory peptides: macrophage chemoattractant protein‐1 (MCP‐1) and macrophage inhibitory protein‐1α (MIP‐1α). By the end of six months wollastonite had induced mild pulmonary interstitial ﬁbrosis, whereas crocidolite induced progressive interstitial ﬁbrosis as a function of time. The membranes of macrophages and pneumocytes were disrupted at the lowest concentration of crocidolite. The activity of enzymes of the redox system and cytoplasmic superoxide dismutase signiﬁcantly decreased with crocidolite. Wollastonite decreased only the activity of gamma‐glutamyl transpeptidase and glutathione peroxidase. Crocidolite induced expression of the proinﬂammatory peptides at the lowest concentration (1 µg ml−1) but wollastonite increased production of these peptides only at medium and high concentrations (5 and 10 µg ml−1). These results underline the importance of further human epidemiological studies and the need for the determination of a hygienic standard. Copyright © 2004 John Wiley & Sons, Ltd.
Development of acute inhalation reference exposure levels (RELs) to protect the public from predictable excursions of airborne toxicants
Authors: Collins James F. , Alexeeff George V. , Lewis David C. , Dodge Daryn E. , Marty Melanie A. , Parker Thomas R. , Budroe John D. , Lam Richard H.F. , Lipsett Michael J. , Fowles Jefferson R. , Das Rupali
Published: March 2004

Uniform guidelines have been developed for the derivation of 1‐h acute inhalation reference exposure levels (RELs) applicable to the general public exposed routinely to hazardous substances released into the environment. Existing acute exposure guidance values developed by other organizations have been examined, and strengths and weaknesses in these existing guidelines have been identiﬁed. The results of that examination have led to the development of a reproducible and resource‐intensive methodology to calculate acute inhalation RELs for 41 prioritized chemicals. Approaches to estimating levels protective against mild and severe acute effects are discussed in this report. The default methodology is the no‐observed‐adverse‐effect level (NOAEL)/uncertainty factor (UF) approach using mainly reports in the peer‐reviewed toxicological and medical literature. For two well‐studied chemicals, ammonia and formaldehyde, the data allowed a benchmark dose (or concentration) methodology, as a departure from the default options, to be used. However, better human dose–response data from, for example, improved workplace monitoring correlated with symptoms, and more extensive epidemiological studies are needed before the departure from default approaches can be expanded to more substances. Copyright © 2004 John Wiley & Sons, Ltd.
Quantitative analysis of three‐dimensional fluid flow in rotating bioreactors for tissue engineering
Authors: Botchwey Edward A. , Pollack Solomon R. , Levine Elliot M. , Johnston Eric D. , Laurencin Cato T.
Published: March 2004

Tissue engineering has emerged as a viable alternative to the problem of organ and tissue shortage. Our laboratory has developed matrices for bone tissue engineering based on sintered spherical particles and, using bioreactor technology, has demonstrated the ability to produce highly mineralized matrices in vitro. In this study, porous microcapsule scaffolds were developed for bone tissue engineering in the high aspect ratio vessel rotating bioreactor. The motion of individual microcapsules as well as scaffolds in the bioreactor were studied by numerical simulation and in situ imaging analysis. Results show that spherical microcapsules with density less than the surrounding fluid exhibited two motions: (1) a periodic circular orbit with tangential speed equal to the free fall speed of the particle, and (2) an inward radial migration of the circular orbit toward the center of the bioreactor vessel. Lighter‐than‐water scaffolds were fabricated by sintering poly(lactic‐co‐glycolic acid) hollow microcarriers with diameter from 500 to 860 μm into a fixed three‐dimensional geometry with approximately 30% pore volume and 180 to 190 μm median pore size. Scaffolds were fabricated with aggregate densities ranging from 0.65 g/mL and 0.99 g/mL by appropriate combinations of hollow and solid microcarriers within the scaffold. Scaffold velocity in the bioreactor for the above range of densities was accurately predicted by numerical simulation and ranged from 100 mm/s to 3 mm/s. Maximum shear stress estimation due to media flow over the exterior of the scaffold ranged from 0.3 N/m2 to 0.006 N/m2. Internal perfusion velocity through scaffolds also was calculated and ranged from 13 mm/s to 0.2 mm/s. Estimates of maximum interior shear stress ranged from 0.03 to 0.0007 N/m2. These analytical methods provide an excellent vehicle for the study of bone tissue synthesis in three‐dimensional culture with fluid flow. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 69A: 205–215, 2004
Preparation and in vitro bioactivity of zinc containing sol‐gel–derived bioglass materials
Authors: Oki Aderemi , Parveen Bushra , Hossain Safiyyah , Adeniji S. , Donahue Henry
Published: March 2004

Bioactive glass material of the type CaOP2O5SiO2ZnO was obtained by the sol‐gel processing method. This material was produced both in powder and in disks form by compression of the powder. The obtained material was characterized by X‐ray powder diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and surface electron micrograph (SEM). The bioactivity was examined in vitro with respect to the ability of hydroxyapatite layer to form on the surface as a result of contact with simulated body fluid (SBF). The XRD and FTIR studies were conducted before and after contact of the material with SBF. The gel‐derived materials were amorphous as shown by XRD, but were able to crystallize calcium phosphates on their surfaces when exposed to SBF. We also examined the alkaline phosphatase (AP) activity of osteoblasts, using human fetal osteoblastic cells (hFOB 1.19) cultured on the zinc bioglass, and compared it with the polystyrene plates. The bone cells consistently expressed higher AP activity in the zinc bioglass materials compared with the polystyrene plates, which indicates the zinc containing composition stimulates bone cells production of AP. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 69A: 216–221, 2004
Optimization of bone‐tissue engineering in goats
Authors: Kruyt Moyo C. , Dhert Wouter J. A. , Oner Cumhur , van Blitterswijk Clemens A. , Verbout Abraham J. , de Bruijn Joost D.
Published: March 2004

Successful bone‐tissue engineering (TE) has been reported for various strategies to combine cells with a porous scaffold. In particular, the period after seeding until implantation of the constructs may vary between hours and several weeks. Differences between these strategies can be reduced to (a) the presence of extracellular matrix, (b) the differentiation status of the cells, and (c) the presence of residual potentially immunogenic serum proteins. These parameters are investigated in two types of calcium phosphate scaffolds in a goat model of ectopic bone formation. Culture‐expanded bone‐marrow stromal cells from eight goats were seeded onto two types of hydroxyapatite granules: HA60/400 (60% porosity, 400‐μm average pore size) and HA70/800. Scaffolds seeded with cells and control scaffolds were cultured for 6 days in medium containing autologous or semisynthetic serum, in the presence or absence of dexamethasone. Other scaffolds were seeded with cells just before implantation in medium with or without serum. All conditions were implanted autologously in the paraspinal muscles. After 12 weeks, bone had formed in 87% of all TE constructs, as demonstrated by histology. Histomorphometry indicated significantly more bone in the HA70/800 scaffolds. Furthermore, a significant advantage in bone formation was found when the constructs had been cultured for 6 days. In conclusion, both scaffold characteristics (porosity) and TE strategy (culturing of the constructs) were demonstrated to be important for bone TE. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 69B: 113–120, 2004
Stripe wear rates in alumina THR—Comparison of microseparation simulator study with retrieved implants
Authors: Manaka Masakazu , Clarke Ian C. , Yamamoto Kengo , Shishido Takaaki , Gustafson Allen , Imakiire Atsuhiro
Published: March 2004

Alumina‐on‐alumina hip implants with microseparation were run in a hip simulator for comparison of the nonseparation simulator mode and retrievals. The 28‐, 32‐, and 36‐mm Biolox‐forte implants were run to 5 million cycles with the use of 50% newborn calf serum. Howmedica Osteonics Trident cups with titanium backing were used in all sets. In standard (STD) and microseparation (MSX) mode, the typical biphasic wear trend was evident, but the MSX test mode had much higher magnitudes. There was a 5‐fold increase for run‐in wear and up to a 35‐fold increase in steady‐state wear. The stripe wear on the ball formed early, but did not progress in grade beyond 0.6 Mc. The locations of the stripes were similar in retrieved and simulator balls. However, the stripes from the simulator were narrower than short‐term retrievals and much narrower than some long‐term retrievals. The long‐term retrieved balls had a grade of wear greater than the simulators. In vivo a broader range of motion occurs and this may lead to the wider stripe observed on the retrievals. These observations suggested that simulators could produce the loading and kinematics similar to a patient walking but not necessarily the variety of motions possible in the in vivo situation. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 69B: 149–157, 2004
Alpha‐crystallin‐incorporated collagen matrices as an aid for dermal wound healing
Authors: Ahmed M. Rafiuddin , Gopinath D. , Gomathi K. , Sehgal P. K. , Jayakumar R.
Published: March 2004

This study evaluated the effects of noncovalently incorporated crystallin into the collagen matrix for dermal wound‐healing processes in rats. Crystallin‐incorporated collagen matrix (CIC) showed better healing when compared to wounds treated with collagen matrix (CS) and without collagen (CR). Biochemical parameters and histological analysis revealed that increased wound contraction enhanced cell proliferation and efficient radical scavenging in the CIC group. The higher shrinkage temperature of CIC films when compared to CS groups suggested increased hydrothermal stability for the former material. An in vitro release study of CIC has showed sustained and time‐dependent release of crystallin from the collagen matrix. These results demonstrate the possibility of using crystallin as therapeutic protein in the wound‐healing process. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 69B: 241–248, 2004
Self‐reinforced polylactide/polyglycolide 80/20 screws take more than 1½ years to resorb in rabbit cranial bone
Authors: Tiainen Johanna , Soini Ylermi , Törmälä Pertti , Waris Timo , Ashammakhi Nureddin
Published: March 2004

The aim of this study was to assess tissue reactions to bioabsorbable self‐reinforced polylactide/polyglycolide (SR‐PLGA) 80/20 miniscrews in rabbit cranial bone. One PLGA screw was implanted on one side and one titanium screw on the other side of the sagittal suture (n = 21). Three animals were sacrificed after 2, 4, 8, 16, 24, 54, and 72 weeks. In histological examination the numbers of macrophages, giant cells, active osteoblasts, and fibrous tissue layers were assessed and degradation of the bioabsorbable screws was evaluated. After 2 weeks, macrophages were seen near the heads of both screws. After 4 and 8 weeks, the bioabsorbable screws were surrounded by fibrous tissue. Osteoblastic activity and groups of several giant cells were seen. After 24 weeks, a significant change in the morphology of the PLGA screws had occurred. Osteoblastic activity and the amount of giant cells had decreased. After 1 year, some PLGA biomaterial was still present. PLGA screws had been replaced by adipose tissue, fibrous tissue, and “foamy macrophages” that had PLGA particles inside them. After 1½ years, the amount of biomaterial remaining had decreased remarkably. The particles of biomaterial were inside foamy macrophages. SR‐PLGA 80/20 screws are biocompatible and have no clinically manifested complications when used in the cranial bone of rabbits. No contraindications as regards their clinical use in craniofacial surgery was found when these screws were studied in the cranial bones of rabbits. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 70B: 49–55, 2004
Release of anthracyclines adsorbed on copper‐treated hydroxylapatites
Authors: Bacha Rosana Ahamad , Fiallo Marina M. L. , Sharrock Patrick
Published: March 2004

Anthracyclines are antibiotics widely used for the treatment of leukemia and solid tumors. Perfusion is the standard clinical administration form. Unfortunately, this does not allow treatment of gastric cancers. A new galenic presentation, consisting of anthracyclines adsorbed onto hydroxylapatite granules, has been developed. It was found to be possible to modulate the release of the drug as a function of the pH by using copper(II) ions. At neutral pH, the drug remains on the mineral support, but at pH 3, the drug is released into solution. Further investigations demonstrate that copper‐containing hydroxylapatite can retain a determined quantity of antibiotic by surface complexation. This result could contribute to new designs for drug carriers targeted for gastric cancers. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 70B: 103–105, 2004
Measuring mechanical properties of fine‐wire cross‐sections used in medical devices
Authors: Yu Ning , Polycarpou Andreas A. , Wagoner Johnson Amy J.
Published: March 2004

In this study, it was demonstrated that the nanoindentation technique can be used to measure the elastic modulus and hardness of fine‐wire cross‐sections with diameters ranging from 100–200 μm. Using miniature optics and a specially developed micrometer positioning system, measurements were successfully performed on the wire cross‐sections with an accuracy of 1 μm. Homogeneous wire cross‐sections, such as MP35N (high‐strength nickel–cobalt alloy), composite wires such as MP35N with a silver core (MP35N/Ag‐core), and platinum‐clad tantalum (Pt‐clad Ta) were studied. From the nanoindentation measurements, the elastic modulus and hardness at different positions and depths were calculated. It was found that both elastic modulus and hardness reached a constant value below the hardened surface, which resulted from mechanical polishing. Material properties obtained from the measurements were compared with the literature values of bulk materials as well as available results on fine wires. The data from the wire cross‐sections, especially the uniform material samples, showed that both the elastic modulus and hardness were significantly lower near the outer 10–15 μm of the wire as compared to the rest of the wire, which is due to surface weakening during manufacturing. In addition, both modulus and hardness showed a small but consistent “sawtooth” variation across the diameter of the wires, excluding the outer 10–15 μm, that is postulated to be induced by the distribution of residual stress. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 70B: 106–113, 2004
Differential response to keratinocyte growth factor receptor and epidermal growth factor receptor ligands of proliferating and differentiating intestinal epithelial cells
Authors: Visco Vincenzo , Belleudi Francesca , Marchese Cinzia , Leone Laura , Aimati Laura , Cardinali Giorgia , Kovacs Daniela , Frati Luigi , Torrisi Maria Rosaria
Published: March 2004

The expression of the keratinocyte growth factor receptor (KGFR) has been analyzed on intestinal epithelial Caco‐2 cells upon confluence‐induced spontaneous differentiation. Western blot and immunofluorescence analysis showed that the expression of functional KGFRs, differently from that of epidermal growth factor receptor (EGFR), was up‐modulated in post‐confluent differentiated cultures compared with the pre‐confluent cells. Confocal microscopy and immunoelectron microscopy revealed that the up‐regulated KGFRs displayed a basolateral polarized distribution on the cell surfaces in the monolayer. In vivo immunohistochemical analysis on normal human colon tissue sections showed that KGFRs, differently from EGFRs, were mostly distributed on the more differentiated cells located on the upper portion of the intestinal crypt. Bromodeoxyuridine incorporation assay and Ki67 labeling indicated that the differentiated cells were able to proliferate in response to the two ligands of KGFR, KGF and FGF‐10, whereas they were not stimulated by the EGFR ligands TGFα and EGF. Western blot and quantitative immunofluorescence analysis of the expression of carcinoembryonic antigen (CEA) in post‐confluent cells revealed that incubation with KGF induced an increase of cell differentiation. Taken together these results indicate that up‐modulation of KGFR may be required to promote proliferation and differentiation in differentiating cells and that, among the cells componing the intestinal epithelial monolayer, the target cells for KGFR ligands appear to be different during differentiation from those responsive to EGFR ligands. © 2004 Wiley‐Liss, Inc.
Expression and regulation of the insulin‐like growth factor axis components in rat liver myofibroblasts
Authors: Novosyadlyy Ruslan , Tron Kyrylo , Dudas Jozsef , Ramadori Giuliano , Scharf Jens‐Gerd
Published: March 2004

Apart from hepatic stellate cells (HSC), liver myofibroblasts (MF) represent a second mesenchymal cell population involved in hepatic fibrogenesis. The IGF system including the insulin‐like growth factors I and II (IGF‐I, ‐II), their receptors (IGF‐I receptor, IGF‐IR; IGF‐II/mannose 6‐phosphate receptor, IGF‐II/M6‐PR), and six high affinity IGF binding proteins (IGFBPs) participate in the regulation of growth and differentiation of cells of the fibroblast lineage, possibly contributing to the fibrogenic process. The aim of this work was to study the expression and regulation of the IGF axis components in rat liver MF. Methods: Cultures of MF from passages 1 to 4 (P1–4) were studied. IGFBP secretion was analyzed by [125I]‐IGF‐I ligand and immunoblotting. IGF‐I, IGF‐IR, IGF‐II/M6‐PR, and IGFBP messenger RNA (mRNA) expression was assessed by Northern blot hybridization. DNA synthesis was evaluated by 5‐bromo‐2′‐deoxyuridine (BrdU) incorporation assay. Results: MF from P1 to 4 constitutively expressed mRNA transcripts specific for IGF‐I, IGF‐IR, and IGF‐II/M6‐PR. In MF, biosynthesis of IGFBP‐3 and ‐2 was observed that was stimulated by IGF‐I, insulin, and transforming growth factor β (TGF‐β), whereas platelet‐derived growth factor (PDGF‐BB) revealed inhibitory effects. IGF‐I and to a lesser extent insulin increased DNA synthesis of MF. Simultaneous addition of recombinant human IGFBP‐2 or ‐3 with IGF‐I diminished the mitogenic effect of IGF‐I on MF whereas preincubation of MF with IGFBP‐2 or ‐3 further potentiated the IGF‐I stimulated DNA synthesis. In conclusion, the present study demonstrates that the IGF axis may play a role in the regulation of MF proliferation in vitro which might be relevant in vivo for the process of fibrogenesis during acute and chronic liver injury. © 2004 Wiley‐Liss, Inc.
Vinculin, VASP, and profilin are coordinately regulated during actin remodeling in epithelial cells, which requires de novo protein synthesis and protein kinase signal transduction pathways
Authors: Quinlan Margaret P.
Published: March 2004

Transformation progression of epithelial cells involves alterations in their morphology, polarity, and adhesive characteristics, all of which are associated with the loss and/or reorganization of actin structures. To identify the underlying mechanism of formation of the adhesion‐dependent, circumferential actin network, the expression and localization of the actin binding and regulating proteins (ABPs), vinculin, VASP, and profilin were evaluated. Experimental depolarization of epithelial cells results in the loss of normal F‐actin structures and the transient upregulation of vinculin, VASP, and profilin. This response is due to the loss of cell–cell, and not cell–substrate interactions, since cells that no longer express focal adhesions or stress fibers are still sensitive to changes in adhesion and manifest this in the altered profile of expression of these ABPs. Transient upregulation is dependent upon de novo protein synthesis, and protein kinase‐, but not phosphatase‐sensitive signal transduction pathway(s). Inhibition of the synthesis of these proteins is accompanied by dephosphorylation of the ribosomal S6 protein, but does not involve inhibition of the PI3‐kinase‐Akt‐mTOR pathway. Constitutive expression of VASP results in altered cell morphology and adhesion and F‐actin and vinculin structures. V12rac1 expressing epithelial cells are constitutively nonadhesive, malignantly transformed, and constitutively express high levels of these ABPs, with altered subcellular localizations. Transformation suppression is accompanied by the restoration of normal levels of the three ABPs, actin structures, adhesion, and epithelial morphology. Thus, vinculin, VASP, and profilin are coordinately regulated by signal transduction pathways that effect a translational response. Additionally, their expression profile maybe indicative of the adhesion and transformation status of
epithelial cells. J. Cell. Physiol. 200: 277–290, 2004. © 2004 Wiley‐Liss, Inc.
Significance of N‐methyl‐d‐aspartate (NMDA) receptor‐mediated signaling in human keratinocytes
Authors: Nahm Walter K. , Philpot Benjamin D. , Adams Michelle M. , Badiavas Evangelos V. , Zhou Linda H. , Butmarc Janet , Bear Mark F. , Falanga Vincent
Published: March 2004

Increasing data suggest that glutamate might act as a cell‐signaling molecule in non‐neuronal tissues such as the skin. Here we demonstrate the presence of functional N‐methyl‐d‐aspartate (NMDA)‐type glutamate receptors in human keratinocytes. NMDA receptor expression strongly reflects the degree of cell‐to‐cell contact. Wounding polarizes the expression of NMDA receptors in keratinocytes involved in re‐epithelialization, and the process of re‐epithelialization is inhibited by NMDA receptor activation. We also demonstrate that squamous cell carcinomas lack NMDA receptors. Our data suggest that Ca2+ entry through NMDA receptors influences the cycle of keratinocyte proliferation, differentiation, and migration during epithelialization. Moreover, NMDA receptor activation might play a role in contact‐mediated inhibition of growth, a process that is absent during neoplastic pathology. This receptor may serve as a pharmacological target for modulating keratinocyte behavior and treating cutaneous disorders. © 2003 Wiley‐Liss, Inc.
Role of RB and RB2/P130 genes in marrow stromal stem cells plasticity
Authors: Jori Francesco P. , Napolitano Marco A. , Melone Mariarosa A.B. , Cipollaro Marilena , Cascino Antonino , Giordano Antonio , Galderisi Umberto
Published: March 2004

Marrow stromal cells (MSCs) are stem‐like cells having a striking somatic plasticity. In fact, besides differentiating into mesenchymal lineages (bone, cartilage, and fat), they are capable of differentiating into neurons and astrocytes in vitro and in vivo. The RB and RB2/P130 genes, belonging to the retinoblastoma gene family, play a key role in neurogenesis, and for this reason, we investigated their role in neural commitment and differentiation of MSCs. In MSCs that were either uncommitted or committed toward neural differentiation, we ectopically expressed RB and RB2/P130 genes and analyzed their role in regulating the cell cycle, apoptosis and differentiation. In uncommitted MSCs, the activity of RB and RB2/P130 appeared limited to negatively regulating cell cycle progression, having no role in apoptosis and differentiation (toward either mesenchymal or neural lineages). On the other hand, in MSCs committed toward the neural phenotype, both RB and RB2/P130 reduced cell proliferation rate and affected the apoptotic process. RB protected differentiating cells from programmed cell death. On the contrary, RB2/P130 increased the percentage of cells in apoptosis. All of these activities were accomplished mainly in an HDAC‐independent way. The retinoblastoma genes also influenced differentiation in neural committed MSCs. RB2/P130 contributes mainly to the induction of generic neural properties, while RB triggers cholinergic differentiation. These differentiating activities are HDAC‐dependent. Our research shows that there is a critical temporal requirement for the RB genes during neuronal differentiation of MSCs: they are not required for cell commitment but play a role in the maturation process. For the above reasons, RB and RB2/P130 may have a role in neural differentiation but not in neural determination. © 2004 Wiley‐Liss, Inc.
Temperature‐sensitive polymer‐conjugated IFN‐γ induces the expression of IDO mRNA and activity by fibroblasts populated in collagen gel (FPCG)
Authors: Sarkhosh Kourosh , Tredget Edward E. , Uludag Hasan , Kilani Ruhangiz T. , Karami Ali , Li Yunyuan , Iwashina Takashi , Ghahary Aziz
Published: March 2004

Indoleamine 2,3‐dioxygenase (IDO) is an intracellular tryptophan‐catabolizing enzyme possessing various immunosuppressive properties. Here, we report the use of this enzyme to suppress the proliferation of peripheral blood mononuclear cells (PBMC) co‐cultured with IDO‐expressing fibroblasts of an allogeneic skin substitute in vitro. Fetal foreskin fibroblasts populated within collagen gel (FPCG) were treated with interferon‐gamma (IFN‐γ) conjugated with a temperature‐sensitive polymer to induce the expression of IDO mRNA and protein. SDS–PAGE showed successful conjugation of IFN‐γ with the temperature‐sensitive polymer. Expression of IDO mRNA was evaluated by Northern analysis. IDO enzyme activity was evaluated by the measurement of kynurenine levels. The results of Northern blot analysis showed an induction of IDO mRNA expression when treated with polymer‐conjugated IFN‐γ. Kynurenine levels, as a measure of IDO bioactivity, were significantly higher in IFN‐γ‐treated fibroblasts than in controls (P < 0.001). In a lasting effect experiment, the expression of IDO mRNA in FPCG treated with polymer‐conjugated IFN‐γ was significantly longer than in those treated with free (non‐conjugated) IFN‐γ (P < 0.001). IFN‐γ radiolabeling showed a prolonged retention of IFN‐γ within collagen gel in its polymer‐conjugated form, compared to its free form. Presence of IDO protein in FPCG was demonstrated by Western analysis even 16 days after removal of the conditioned medium (containing released IFN‐γ). To demonstrate the immunosuppressive effects of IDO on the proliferation of PBMC, IDO‐expressing FPCG treated with polymer‐conjugated IFN‐γ were co‐cultured with PBMC for a period of 5 days. The results showed a significant reduction in proliferation of PBMC co‐cultured with IFN‐γ‐treated IDO‐expressing fibroblasts, compared to those co‐cultured with non‐IDO‐expressing fibroblasts (P < 0.001). The addition of an IDO inhibitor (1‐methyl‐D‐tryptophan) reversed the suppressive effects of IDO on PBMC proliferation. In conclusion, IDO expression in FPCG suppresses the proliferation of immune cells in vitro. The use of a temperature‐sensitive polymer further prolongs the effect of IFN‐γ on the expression of IDO. Therefore, modulating IDO levels in situ might be an alternative for prolonging the survival of skin allografts. J. Cell. Physiol. 201: 146–154, 2004. © 2004 Wiley‐Liss, Inc.
Uncoupling protein‐3 sensitizes cells to mitochondrial‐dependent stimulus of apoptosis
Authors: Dejean Laurent , Cámara Yolanda , Sibille Brigitte , Solanes Gemma , Villarroya Francesc
Published: March 2004

The mitochondrial uncoupling protein‐3 is a member of the mitochondrial carrier protein family. As a homologue of the thermogenic brown fat uncoupling protein‐1, it possesses a mitochondrial uncoupling activity and thus can influence cell energy metabolism but its exact biological function remains unclear. In the present study, uncoupling protein‐3 was expressed in 293 cells using the tetracycline‐inducible system and its impact on cell bioenergetics and responsiveness to the apoptotic stimulus was determined. The induction of uncoupling protein‐3 expression in mitochondria did not lead to uncontrolled respiratory uncoupling in intact cells. However, it caused a GDP‐inhibition of state 4 respiration and a GDP‐induced re‐polarization of the inner mitochondrial membrane in the presence of fatty acids, in agreement with its expected physiological behavior as an uncoupling protein (UCP). Uncoupling protein‐3 expression did not cause apoptosis per se but increased the responsiveness of the cells to a mitochondrial apoptotic stimulus (i.e., addition of staurosporine in the culture medium). It enhanced caspase 3 and caspase 9 activation and favored cytochrome c release. Moreover, cells in which uncoupling protein‐3 expression had been induced showed a higher mitochondrial Bax/Bcl‐2 ratio essentially due to enhanced translocation of Bax from cytosol to mitochondria. Finally, the induction of uncoupling protein‐3 also increased the sensitivity of mitochondria to open the permeability transition pore in response to calcium. It is concluded that the presence of uncoupling protein‐3 in mitochondria sensitizes cells to apoptotic stimuli involving mitochondrial pathways. © 2004 Wiley‐Liss, Inc.
Mitogenic signalling by B2 bradykinin receptor in epithelial breast cells
Authors: Greco S. , Muscella A. , Elia M.G. , Romano S. , Storelli C. , Marsigliante Santo
Published: March 2004

The kinin peptides are released during inflammation and are amongst the most potent known mediators of vasodilatation, pain, and oedema. A role in the modulation or induction of healthy breast tissue growth has been postulated for tissue kallikrein present in human milk. Moreover, tissue kallikrein was found in malignant human breast tissue and bradykinin (BK) stimulates the proliferation of immortalised breast cancer cells. Aim of the present article was to investigate whether BK also exerts mitogenic activity in normal breast epithelial cells and partially characterise the signalling machinery involved. Results show that BK increased up to 2‐fold the 24 h proliferation of breast epithelial cells in primary culture, and that the BK B2 receptor (not B1) inhibitor alone fully blocked the BK response. Intracellular effects of B2 stimulation were the following: (a) the increase of free intracellular Ca2+ concentration by a mechanism dependent upon the phospholipase C (PLC) activity; (b) the cytosol‐to‐membrane translocation of conventional (PKC)‐α and ‐β isozymes, novel PKC‐δ, ‐ε, and ‐η isozymes; (c) the phosphorylation of the extracellular‐regulated kinase 1 and 2 (ERK1/2); and (d) the stimulation of the expression of c‐Fos protein. EGF, a well known stimulator of cell proliferation, regulated the proliferative response in human epithelial breast cells to the same extent of BK. The effects of BK on proliferation, ERK1/2 phosphorylation, and c‐Fos expression were abolished by GF109203X, which inhibits PKC‐δ isozyme. Conversely, Gö6976, an inhibitor of PKC‐α and ‐β isozymes, and the 18‐h treatment of cells with PMA, that led to the complete down‐regulation of PKC‐α, ‐β, ‐ε, and ‐η, but not of PKC‐δ, did not have any effect, thereby indicating that the PKC‐δ mediates the mitogenic signalling of BK. Phosphoinositide 3‐kinase (PI3K), tyrosine kinase of the epidermal growth factor receptor (EGFR), and mitogen activated protein kinase kinases (MEK) inhibitors were also tested. The results suggest that EGFR, PI3K, and ERK are required for the proliferative effects of BK. In addition, the BK induced cytosol‐to‐membrane translocation of PKC‐δ was blocked by PI3K inhibition, suggesting that PI3K is upstream to PKC‐δ. In conclusion, BK has mitogenic actions in cultured human epithelial breast cells; the activation of PKC‐δ through B2 receptor acts in concert with ERK and PI3K pathways to induce cell proliferation. J. Cell. Physiol. 201: 84–96, 2004. © 2004 Wiley‐Liss, Inc.
Impact of novel histone deacetylase inhibitors, CHAP31 and FR901228 (FK228), on adenovirus‐mediated transgene expression
Authors: Taura Kojiro , Yamamoto Yuzo , Nakajima Akio , Hata Koichiro , Uchinami Hiroshi , Yonezawa Kei , Hatano Etsuro , Nishino Norikazu , Yamaoka Yoshio
Published: March 2004

Background

Histone deacetylase inhibitors (HDIs) are known to enhance adenovirus (Ad)‐mediated transgene expression. Recently, novel HDIs, including cyclic hydroxamic‐acid‐containing peptide 31 (CHAP31) and FR901228 (FK228), have been developed.

Methods

The effects of these two novel HDIs on Ad‐transduced or endogenous gene expression were investigated. Acetylation of core histones and the expression of the coxsackie and adenovirus receptor (CAR) in HDI‐treated cells were examined using Western blot and a quantitative reverse transcription polymerase chain reaction (TaqMan RT‐PCR), respectively. Their in vivo effect on adenoviral gene expression was investigated in BALB/c mice.

Results

Both compounds enhanced and prolonged Ad‐mediated β‐galactosidase expression more effectively than did trichostatin A, a classic HDI. The same effect was observed in Ad‐transduced heat shock protein 72 (HSP72), but not in hyperthermia‐induced endogenous expression of HSP72, suggesting that the effect is specific for transduced gene expression. Hyperacetylation of core histones induced by HDIs was considered responsible for the augmentative effects of gene expression. Intravenous administration of either CHAP31 or FR901228 enhanced β‐galactosidase expression in mice infected with AdLacZ.

Conclusions

CHAP31 and FR901228 amplified Ad‐mediated transgene expression. The enhancement of transgene expression by HDIs may result in fewer vector doses for necessary gene expression, helping to alleviate disadvantages caused by Ad vectors. This could be a useful tool in overcoming current limitations of gene therapy using adenovirus vectors. Copyright © 2004 John Wiley & Sons, Ltd.