bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparative study of spiculogenesis in demosponge and hexactinellid larvae
Authors: Leys S.P.
Published: October 2003

Spicule deposition was studied by electron microscopy in fixed embryos and larvae of the haplosclerid spongeRenierasp. and the hexactinellidOopsacas minuta. Spicules form in centrally located vacuoles within cells and within syncytia, as in the adult sponges. InReniera, scleroblasts differentiate from micromeres prior to gastrulation. At gastrulation the scleroblasts migrate to the periphery of the embryo and commence spicule deposition around a hexagonal axial filament. Sclerocytes have numerous pseudopodia and migrate to the posterior pole where they become aligned along the antero‐posterior axis in the free‐swimming larva.Renieralarvae possess some 40–50 oxeas, each 70–75 μm long and 1 μm wide. MatureOopsacaslarvae have up to 14 stauractin spicules, which are produced on a rectangular axial filament in tissues that lie under the smooth epithelium at the posterior pole of the larva. Young sclerocytes have many pseudopodia. The 4‐rayed spicules elongate along both the antero‐posterior and medial axes, until the longitudinal rays become anchored in a lipid‐filled cytoplasm at the anterior of the larva and the lateral rays intersect around the midline. The length of the transverse rays of the stauractins in free‐swimming larvae are 27–45 μm each, while the length of the two longitudinal rays is 40–80 μm. Although spicule deposition begins in cells with a similar morphology in both cellular and syncytial sponges, the elaboration and organization of the spicules differ markedly in cellular and syncytial sponges and appear to be an outcome of the very distinct cellular differentiation and larval morphogenesis that occur in each of these groups. Microsc. Res. Tech. 62:300–311, 2003. © 2003 Wiley‐Liss, Inc.
Some aspects of silica deposition in lithistid demosponge desmas
Authors: Pisera Andrzej
Published: October 2003

Transmitted light microscope and SEM observations of various growth stages, including very young forms, of lithistid demosponge spicules called desmas provided a wealth of new observations on silica deposition in desmas of most lithistid demosponge groups. In typical (pachastrellid) demosponges a basic feature of silica deposition in young spicules is the formation of silica granules (100–160 nm in diameter) deposited in more or less regular concentric layers. Further growth stages in typical demosponges are similar, only silica granules are smaller and more densely packed. The shape of the spicule is controlled by an organic axial filament, while features of the outer spicule surface are also determined by silicalemma. In lithistid desmas the early stage of silica deposition is controlled by an organic axial filament or, in some cases, dispersed organic molecules only. The next step, after early arrest of axial filament growth and its total encasing by silica, is the deposition of various silica granules (40–300 nm in diameter), spheres, and/or cylinders (1,300–3,330 nm), which are either the result of precipitation or the effect of templating by organic molecules (proteins and polysaccharides), without direct control by the silicalemma. The later stages of desma growth are under direct control of the silicalemma, which molds secondary branches and/or elements of sculpture of the desmas. The tips of desmas, which articulate with older desmas, are also controlled by local spatial relationships. Differentiation of morphological forms of silica in desmas, which is at least genus‐specific, clearly supports the polyphyletic nature of lithistid sponges. Microsc. Res. Tech. 62:312–326, 2003. © 2003 Wiley‐Liss, Inc.
Sponge cell reactivity to various forms of silica
Authors: Bavestrello Giorgio , Benatti Umberto , Cattaneo‐Vietti Riccardo , Cerrano Carlo , Giovine Marco
Published: October 2003

Several sponge species incorporate a wide range of foreign material. Whether such material is actively selected by the sponge is controversial. Here we compare the available suspended matter and the sediment incorporated in the tissue of the demosponge Chondrosia reniformis. Field observations and laboratory experiments indicate that this species selects and incorporates only siliceous materials, in particular quartz particles and opal sponge spicules, avoiding carbonate particles. The reaction of ectosome cells of Chondrosia depends on the forms of silica: after settlement of crystalline quartz particles on the sponge surface, the pinacocytes contract uniformly, giving rise to a ruffled surface that remains throughout the incorporation of foreign material. In contrast, the opal spicules elicit a motile response in pinacocytes, which cover the spicules as a result. After incorporation, while the opal spicules remain unaltered within sponge tissue, the engulfed quartz particles are quickly etched, reduced in size, and released from the sponge. The etching of quartz particles by C. reniformis is produced by ascorbic acid, and is the first evidence of such activity from the animal kingdom. Ascorbic acid has been found to change the quartz surface features, which leads to an increased radical production and a consequent dissolution of quartz. This process does not occur on opal spicules. Microsc. Res. Tech. 62:327–335, 2003. © 2003 Wiley‐Liss, Inc.
Physical and chemical analysis of the siliceous skeletons in six sponges of two groups (demospongiae and hexactinellida)
Authors: Sandford Floyd
Published: October 2003

The siliceous skeletons of six hexactinellids and demosponges were compared using a series of physical and chemical tests. The sponges were two hermit‐crab sponges (Class Demospongiae, family Suberitidae), one from Scotland, Suberites domuncula, and the other, Pseudospongosorites suberitoides, from the Gulf of Mexico, and four hexactinellids, Hyalonema sp., Euplectella aspergillum, Rhabdocalyptus dawsoni, and Aphrocallistes vastus. The operating hypothesis was that differences in the amorphous hydrated silica skeletons in Demosponges and Hexactinellids might prove taxonomically useful. Physical properties studied included SEM, glass density, glass transition temperature (Tg), TG/DTA to determine water content, and FTIR spectra. Chemical determinations were made using energy dispersive X‐ray fluorescence (EDXRF). Spicular skeletal material is deposited in concentric layers around the axial canal. With the exception of differences at several bands in the IR spectra, the siliceous skeletons in demosponges and hexactinellids are largely indistinguishable. Density was similar in all sponges (range 2.03–2.13 g/cc) and similar to the density of opal (SiO2 · 1.5H2O) (mean = 2.09 g/cc). IR spectra were similar, with prominent absorption bands at 460–470, 800, and 1,090–1,100 cm−1 (due to different vibrational modes of Si‐O‐Si linkages) and at 1,650 and 3,450–3,560 cm−1 (due to water). The skeletons of all six showed similar spectra to that of silica gel both before and after heating to 1,200°C. Water comprised 10–14% of the skeleton by weight, slightly higher in the demosponges. Average spicule chemical composition was 85.2% SiO2, 12.3% water, and 2.5% other elements (mainly S, Al, K, Ca, and Na). The percent amounts of Si did not differ significantly between the demosponges and the hexactinellids. Microsc. Res. Tech. 62:336–355, 2003. © 2003 Wiley‐Liss, Inc.
Molecular biology of demosponge axial filaments and their roles in biosilicification
Authors: Weaver James C. , Morse Daniel E.
Published: October 2003

For hundreds of years, the skeletal elements of marine and freshwater sponges have intrigued investigators with a diverse array of remarkably complex morphologies. Early studies of demosponge monaxonal megascleres revealed the presence of a central organic axial filament running their entire length. Until recently, however, the precise function of these axial filaments was largely unknown. The spicules from the temperate Eastern Pacific demosponge, Tethya aurantia, comprise ∼75% of the dry weight of this species, facilitating the large‐scale isolation and purification of the biosilica‐associated proteins. Silicateins, the most abundant proteins comprising the axial filaments of these spicules, prove to be members of a well‐known superfamily of proteolytic and hydrolytic enzymes and can be easily collected after silica demineralization with hydrofluoric acid. Consistent with these findings, the intact filaments are more than simple, passive templates; in vitro, they actively catalyze and spatially direct the hydrolysis and polycondensation of silicon alkoxides to yield silica at neutral pH and low temperature. Catalytic activity also is exhibited by the monomeric subunits obtained by disaggregation of the protein filaments and those produced from recombinant DNA templates cloned in bacteria. These proteins also can be used to direct the polymerization of organosilicon polymers (silicones) from the corresponding organically functionalized silicon alkoxides. Based on these observations, the silicateins are currently being used as models for the design of biomimetic agents with unique catalytic and structure‐directing properties. The presence of axial filaments in a diversity of spicule types and the evolutionary implications of these findings are also discussed. Microsc. Res. Tech. 62:356–367, 2003. © 2003 Wiley‐Liss, Inc.
Biochemistry and cell biology of silica formation in sponges
Authors: Müller Werner E.G. , Krasko Anatoli , Le Pennec Gaël , Schröder Heinz C.
Published: October 2003

The main inorganic material forming the skeletal elements in Demospongiae as well as in Hexactinellida, the spicules, is amorphous silica. The spicules occur in the cytoplasm and the extracellular space and also in the nucleus (as silicate crystals) of some sponge cells; the function in the latter compartment is unknown. Recent evidence shows that the formation of spicules is mediated by the enzyme silicatein. The cDNA as well as the gene encoding this enzyme was cloned from Suberites domuncula. The recombinant silicatein catalyzes the synthesis of amorphous silicate using tetraethoxysilane as substrate. The enzyme is dependent on ferric iron. Silicatein also has proteolytic (cathepsin‐like) activity. Incubation of primmorphs, a special form of 3D‐cell aggregates, with silicon resulted in a strong increase of their size from 1–7 mm to ∼10 mm. The morphogenetic activity of silicate is underscored by the finding that this ion increases gene expression of silicatein and collagen. Based on these findings, it is concluded that both iron and silicate stimulate the activity of silicatein. Furthermore, it is proposed that the growing spicules are surrounded by the scavenger receptor which might be considered as a docking molecule for the collagen matrix into which the spicules are embedded. Microsc. Res. Tech. 62:368–377, 2003. © 2003 Wiley‐Liss, Inc.
Fiber diffraction study of spicules from marine sponges
Authors: Croce Gianluca , Frache Alberto , Milanesio Marco , Viterbo Davide , Bavestrello Giorgio , Benatti Umberto , Giovine Marco , Amenitsch Heinz
Published: October 2003

A synchrotron radiation fiber diffraction structural study of the axial filament of siliceous spicules from two species of marine sponges (the Demosponge Geodia cydonium and the Hexactinellid Scolymastra joubini) was carried out. The sharpness of the spots in the diffraction patterns indicated that the protein units in the filament of both samples were highly organized. A possible explanation is that the arrangement of the protein units is similar to that of the pores in highly ordered siliceous mesoporous materials. Nevertheless, the diffraction patterns are quite different for the two types of spicules. The pattern of G. cydonium is consistent with a regular 2D hexagonal lattice of protein units in the direction perpendicular to the spicule axis, with a repeating distance of 5.8 nm; the units are linked to form fibers along the axis. The pattern of S. joubini indicates the presence of two different 2D lattices in which the repeating protein units are inclined by +50° and –50° with respect to the elongation axis; the distance between the units increases to 8.4 nm. This 2D model is consistent with hexagonal packing of spirally oriented cylindrical protein units elongated along the filament axis. Microsc. Res. Tech. 62:378–381, 2003. © 2003 Wiley‐Liss, Inc.
Combination with CD/5‐FC gene therapy enhances killing of human bladder‐cancer cells by radiation
Authors: Zhang Zhujun , Shirakawa Toshiro , Hinata Nobuyuki , Matsumoto Akira , Fujisawa Masato , Okada Hiroshi , Kamidono Sadao , Matsuo Masafumi , Gotoh Akinobu
Published: October 2003

Background

Resistance to radiation and chemotherapy is a significant obstacle to the treatment of advanced bladder cancer. Gene therapy combined with radiation represents a new approach to cancer treatment. In the present study, we investigated whether adenovirally directed, cytosine deaminase (CD)/5‐fluorocytosine (5‐FC) gene therapy could induce cell toxicity and radiosensitization through the intracellular production of 5‐fluorouracil (5‐FU) in bladder‐cancer cells.

Methods

Three human bladder‐cancer cell lines, KK47 (wild‐type p53+), T24 (p53 mutated) and 5637 (p53 mutated), were investigated. A recombinant adenovirus vector containing the CD gene (Ad‐RSV‐CD) was used. Cells were infected with Ad‐RSV‐CD and treated with 5‐FC. Forty‐eight hours after infection, the cells were irradiated and cytotoxicity assays performed to determine the extent of increase in in vitro cytotoxicity. A KK47 subcutaneous tumor‐xenografts model was used in an animal study to examine the tumor growth inhibitory effect of this combination therapy. Ad‐RSV‐CD was directly injected into the tumor and daily 5‐FC was intraperitoneally injected. Forty‐eight hours after injection of Ad‐RSV‐CD, the tumor was irradiated. The tumor volume was measured every day.

Results

In all three cell lines, the combination treatment enhanced the cell killing of human bladder‐cancer cells in vitro. It also enhanced the tumor‐growth inhibition in the KK47 tumor model.

Conclusions

In the present study, we demonstrated that CD/5‐FC gene therapy combined with radiation therapy enhances cell killing of human bladder‐cancer cells in in vitro and in vivo animal models. Copyright © 2003 John Wiley & Sons, Ltd.
Coxsackie‐and‐adenovirus receptor mRNA expression in human heart failure
Authors: Sasse A. , Wallich M. , Ding Z. , Goedecke A. , Schrader J.
Published: October 2003

Background

Adenoviral vectors are widely used as gene‐transfer vehicles in experimental and clinical studies. Since virus incorporation and transfection efficacy depend to a large extent on the concentration of the coxsackie‐and‐adenovirus (CAR) receptor on target cells the aim of this study was to quantify the CAR‐receptor concentration in various human cardiomyopathies.

Methods

After RNA isolation from myocardial biopsies obtained during surgical procedures, cDNA was generated by reverse transcription. The relative RNA content was analyzed by quantitative PCR using glyceraldehydes‐3‐phosphate dehydrogenase (GAPDH) as a standard reference. The cardiomyopathies (CM) analyzed were categorized according to their etiology in dilated CM (DCM, n = 28), ischemic CM (ICM, n = 52), CM in mitral valve disease (MVCM, n = 32) and aortic valve disease (AVCM, n = 32). Data were related to non‐cardiomyopathic tissue from donor hearts (non‐CM, n = 64).

Results

Compared with non‐CM hearts DCM showed a 34‐fold (±5.4) increase in CAR mRNA concentration, in ICM CAR mRNA was elevated by a factor of 12 (±4.3), in MVCM by 27 (±7) and AVCM by factor 47 (±9.3) (ANOVA p < 0.001). Compared with the expression in rat hearts CAR levels were found to be similar to those in human ICM.

Conclusions

These results show that cardiomyopathies associated with heart failure transcribe substantially higher levels – on average by a factor of 30 – of CAR‐mRNA than non‐failing control hearts. Myocardial gene transfer using adenoviral vectors should therefore be facilitated in human cardiomyopathies and may present a promising approach for therapeutic interventions. Copyright © 2003 John Wiley & Sons, Ltd.
Chemotherapy increases transgene expression in leukemic cells
Authors: Vereecque Rodolphe , Saudemont Aurore , Depil Stéphane , Quesnel Bruno
Published: October 2003

Background

Patients with acute myeloid leukemia (AML) often obtain complete remission with chemotherapy but the majority of patients relapse. Combining chemotherapy and gene therapy appears to be a promising approach; however, the effects of chemotherapy on transgene expression in leukemic cells have not yet been investigated.

Methods

DA1‐3b leukemic cells were transfected with pCDNA3 plasmids carrying GM‐CSF or LacZ cDNA. The leukemic K562 cell line and primary cultured AML cells were transduced with an Ad5.CMV‐LacZ adenoviral vector. Cells were then incubated with various concentrations of daunorubicin (DNR) and cytosine arabinoside (Ara‐C), and expression of the transgene was measured. Murine DA1‐3b‐pCDNA3/LacZ leukemic cells were also injected into syngeneic C3H/Hej mice.

Results

In the cells carrying pCDNA3, DNR and Ara‐C dramatically increased expression of the LacZ and GM‐CSF transgenes. Over‐expression depended on drug dose and was due to increased transcription. Enhancement was also observed in K562 cells and in some primary cultured AML samples transduced with the Ad5.CMV‐LacZ adenovirus. Addition of N‐acetyl‐L‐cysteine inhibited the over‐expression, suggesting that reactive oxygen species were involved in activating the CMV promoter. In the A549 lung carcinoma cell line transduced with Ad5.CMV‐LacZ, Ara‐C had only a minor effect, and DNR had a detrimental effect, suggesting that expression depends on cell type. In vivo experiments in which mice received DA1‐3b‐pCDNA3/LacZ leukemic cells, and were then treated with Ara‐C, also showed increased transgene expression in these leukemic cells.

Conclusions

In leukemic cells, chemotherapeutic agents can induce over‐expression of transgenes. This suggests a promising combined strategy for the treatment of acute leukemia. Copyright © 2003 John Wiley & Sons, Ltd.
Rational development of a HIV‐1 gene therapy vector
Authors: Anson D. S. , Fuller M.
Published: October 2003

Background

HIV‐1 provides an attractive option as the basis for gene transfer vectors due to its ability to stably transduce non‐cycling cell populations. In order to fully utilise the promise of HIV‐1 as a vector it is important that the effects of viral cis sequence elements on vector function are carefully delineated.

Methods

In this study we have systematically evaluated the effect of various cis elements from the HIV‐1 YU‐2 genome that have been implicated as either affecting vector performance, or HIV‐1 replication, on the efficiency of vector production (titre and infectivity). As a measure of the relative safety of vectors their propensity to inadvertently transfer the gagpol gene to transduced cells was assessed.

Results

Sequences that were found to increase vector titre were from the 5′ end of the gag gene, from the 5′ and 3′ ends of the env gene, from immediately upstream of the polypurine tract, and the central polypurine tract. The substitution of the HIV‐1 RRE with heterologous RNA transport elements, or the deletion of the RRE, resulted in greatly reduced vector titres. RNA analysis suggested that the role of the Rev/RRE system extends beyond simply acting as an RNA nuclear export signal. The relative safety of different vector designs was compared and an optimal construct selected.

Conclusions

Based on our results we have constructed a vector that is both more efficient, and has better safety characteristics, than the widely used pHR′ HIV‐1 vector construct. Copyright © 2003 John Wiley & Sons, Ltd.
DNA transfection of mononuclear cells in muscle tissue
Authors: Grønevik Eirik , Tollefsen Stig , Sikkeland Liv Ingunn Bjoner , Haug Terje , Tjelle Torunn Elisabeth , Mathiesen Iacob
Published: October 2003

Background

Genes encoding non‐self proteins may be injected into skeletal muscles in vivo to obtain induction of cellular and humoral immune responses against the encoded antigens (DNA vaccination). Bone marrow derived professional antigen‐presenting cells (APCs) play a key role in the induction of immunity by DNA vaccination. In the present work we have investigated whether the APCs are transfected by DNA injection into muscle.

Methods

DNA encoding green fluorescent protein (GFP) was injected into rat and mouse limb muscle and followed by electroporation. Whole mount muscle tissue with GFP‐positive mononuclear cells (MNCs) were treated with immunocytochemical markers specific for leukocytes, and studied with fluorescent microscopy. To detect transfected cells migrating to peripheral lymphoid tissue RT‐PCR was applied on RNA isolated from the draining popliteal lymph node and spleen. Lymphoid tissue was also analyzed with real‐time PCR for distribution of the injected plasmid.

Results

MNCs were transfected after intramuscular DNA injection, and, following DNA injection with electroporation, the number of GFP‐positive MNCs increased 6‐fold in rats and 14‐fold in mice. None of the GFP‐positive MNCs were stained with leukocyte‐specific antibodies. Even though GFP encoding DNA was detected in the popliteal lymph node, no RNA encoding GFP was found in the lymph node or spleen. However, MHC II‐positive cells in the muscle tissue appeared preferentially around the transfected MNCs.

Conclusions

Many MNCs in the muscle are transfected after intramuscular DNA injection. Electroporation significantly increases the number of transfected MNCs. None of the observed transfected MNCs however were leukocytes. MHC II‐positive cells accumulated around transfected MNCs; this suggests that transfer of antigen from transfected MNCs to APCs may contribute to the immune response. Copyright © 2003 John Wiley & Sons, Ltd.
Functional delivery of large genomic DNA to human cells with a peptide‐lipid vector
Authors: White Robert E. , Wade‐Martins Richard , Hart Stephen L. , Frampton Jon , Huey Bryan , Desai‐Mehta Ami , Cerosaletti Karen M. , Concannon Patrick , James Michael R.
Published: October 2003

Background

Nonviral gene transfer vectors have the potential to deliver much larger DNA constructs than current viral vectors but suffer from a low transfection efficiency. The LID vector, composed of Lipofectin (L), an integrin‐targeting peptide (I) and DNA (D), is a highly efficient synthetic vector, both in vitro and in vivo, which may allow the transfer of genomic loci for gene therapy.

Methods

Transfection efficiencies were quantitated using the green fluorescent protein (GFP) reporter. Expression of a large genomic locus (NBS1 [Nijmegen breakage syndrome], encoding nibrin) was assessed by immunofluorescence.

Results

We report a systematic study of the parameters influencing delivery of BAC‐based plasmids ranging in size from 12 to 242 kb using the LID vector. We showed 60% of cells were transfected with the smaller plasmids while plasmids up to 242 kb were consistently delivered to over 10% of cells. The number of transfected cells was related to number of plasmids in the transfection complex independent of plasmid size. Atomic force microscopy showed that LID particle size increased with plasmid size consistent with one plasmid molecule per particle. When LID vectors were used to deliver the NBS1 gene as a 143 kb construct to primary NBS cells, at least 57% of cells expressing GFP also expressed functional nibrin.

Conclusions

We show that LID vectors represent a promising tool for the transfer of complete genomic loci. Copyright © 2003 John Wiley & Sons, Ltd.
Enhancing adenovirus‐mediated gene transfer in vitro and in vivo by addition of protamine and hydrocortisone
Authors: Lin Tongyu , Gu Jian , Zhang Lidong , Davis John J. , Huang Xuefeng , Cabbini Giovanni , Ji Lin , Fang Bingliang
Published: October 2003

Background

Inclusion of positively charged polymers such as protamine in adenovector formulations has been reported to improve the efficiency of adenovirus‐mediated gene transfer in vitro and in vivo. On the other hand, corticosteroids are known to inhibit inflammation and thus might be useful in minimizing vector‐related toxicity. In this study, we evaluated the combined effect of protamine sulfate and hydrocortisone on the efficiency of adenovirus‐mediated gene transfer in vitro and in vivo.

Methods

Protamine and hydrocortisone at different concentrations were added to adenovector formulations. In vitro transgene expression with or without inclusion of protamine and hydrocortisone was evaluated in the breast cancer cell lines MDA‐MB‐231 and MCF7 and the lung cancer cell lines A549 and H460. In vivo transgene expression in the mouse lung was determined after aerosolized vector delivery.

Results

The combination of 2 µg/ml protamine and 125 ng/ml hydrocortisone significantly increased transgene expression in vitro in all the cell lines tested. Protamine is only effective when it is added to cells before or together with adenovectors, whereas hydrocortisone is effective when it is added to cells before, together with, or after adenovectors. Inclusion of protamine and hydrocortisone also augmented apoptosis induction caused by adenovectors expressing proapoptotic genes in cancer cells. Moreover, protamine and hydrocortisone dramatically enhanced transgene expression in the mouse lung after aerosolized vector delivery.

Conclusions

Inclusion of protamine and hydrocortisone in adenovector formulations can improve adenovector‐mediated gene expression and may be useful for clinical applications of current adenovirus‐mediated gene therapy. Copyright © 2003 John Wiley & Sons, Ltd.
Modulation of renal glomerular disease using remote delivery of adenoviral‐encoded solubletype II TGF‐β receptor fusion molecule
Authors: Haviv Yosef S. , Takayama Koichi , Nagi Peter A. , Tousson Albert , Cook William , Wang Minghui , Lam John T. , Naito Seiji , Lei Xiaosheng , Carey Delicia E. , Curiel David T.
Published: October 2003

Background

Systemic adenoviral (Ad) gene therapy for renal disorders is largely hampered by the unique architecture of the kidney. Consequently, currently available Ad vectors are of only limited therapeutic utility in the context of glomerular and fibroproliferative renal diseases.

Methods

The Ad vectors studied in the context of blocking renal fibrosis were AdTβ‐ExR and AdCATβ‐TR. AdTβ‐ExR encodes a chimeric soluble molecule comprising the entire ectodomain of the human type II TGF‐β receptor, genetically fused to the Fc fragment of the human IgG1 (sTβRII), while AdCATβ‐TR encodes only the dominant‐negative truncated ectodomain of the human type II TGF‐β receptor. The biologic activity of the type II TGF‐β receptor was evaluated in vitro by its ability to inhibit cellular proliferation and in vivo in a unilateral ureter obstruction fibrosis model. Renal targeting with sTβRII was evaluated immunohistochemically after intramuscular (IM) delivery of AdTβ‐ExR. The renal antifibrotic effect of the Ad vectors was evaluated in a lupus murine model with both light and electron microscopy and urinalysis.

Results

sTβRII was detected in the glomeruli after remote IM injection of AdTβ‐ExR, but not the control AdCATβ‐TR, indicating renal deposition of the heterologous soluble fusion protein after its expression in the muscle and secretion into the circulation. AdTβ‐ExR, but not AdCATβ‐TR, could transiently inhibit mesangial expansion, glomerular hypercellularity, proteinuria and cortical interstitial fibrosis in a murine lupus model. However, the autoimmune renal disease eventually surpassed the antifibrotic effect.

Conclusions

These results indicate the superiority of a soluble type II TGF‐β receptor over a dominant‐negative, non‐soluble type II TGF‐β receptor in the context of blocking renal fibrosis in murine models. Copyright © 2003 John Wiley & Sons, Ltd.
Myocardial injection of CA promoter‐based plasmid mediates efficient transgene expression in rat heart
Authors: Huang Jianhua , Ito Yoshinori , Kobune Masayoshi , Sasaki Katsunori , Nakamura Kiminori , Dehari Hironari , Takahashi Kazuhiro , Ikeda Katsuya , Uchida Hiroaki , Kato Kazunori , Hamada Hirofumi
Published: October 2003

Background

Although naked plasmid injection is the safest and most convenient method for gene delivery, a major limitation of this approach is currently poor transgene expression. The CA promoter (chicken β‐actin promoter with cytomegalovirus, CMV, enhancer) is one of the strongest transcriptional control modules found; however, it is uncertain whether a CA promoter‐based vector is efficient enough for naked gene therapy in a cardiovascular context.

Methods

The β‐galactosidase (LacZ) expression provided by CA promoter plasmid (pCAZ2) injection into the skeletal muscle or the heart of Lewis rats was compared with CMV promoter plasmid or adenoviral vector (AxCAZ3). The effect of Simian virus 40 of the replication origin (SV40ori) deletion from pCAZ2 on transgene expression was also evaluated.

Results

pCAZ2 showed the highest LacZ expression in both skeletal muscle and heart in comparison with the CMV promoter‐based vector 5 days after naked plasmid injection. LacZ expression in the heart obtained using 20 µg of pCAZ2 was almost equivalent to that shown with AxCAZ3 at 6.0 × 109 optical particle units. The time course of transgene expression driven by CMV and CA promoters in the heart were similar, with the CA promoter providing significantly higher gene expression than the CMV promoter across all time points examined. SV40ori deletion from pCAZ2 did not affect transgene expression in either skeletal muscle or heart.

Conclusions

Transgene expression mediated by naked CA promoter‐based plasmid injection was shown to be quite efficient in the heart. We propose that the CA promoter vector is suitable for myocardial gene therapy. Copyright © 2003 John Wiley & Sons, Ltd.
Enhanced antitumor activity of a combination of MBD2‐antisense electrotransfer gene therapy and bleomycin electrochemotherapy
Authors: Ivanov Marie‐Agnès , Lamrihi Badia , Szyf Moshe , Scherman Daniel , Bigey Pascal
Published: October 2003

Background

MBD2 is a methylated DNA‐binding protein that has been previously suggested to have transcriptional silencing as well as DNA demethylase activities. We have previously shown that electrotransfer of an MBD2‐antisense encoding plasmid inhibits tumor growth in vivo. In this study we tested whether a combination of MBD2‐antisense gene therapy and bleomycin chemotherapy has an augmented antitumor effect in comparison with either monotherapy.

Methods

Mice bearing human non‐small‐cell lung carcinoma line H1299 xenoplants were treated with electrotransfer of either bleomycin or MBD2‐antisense expression plasmid or a combination of both therapies and tumor growth following treatment was monitored.

Results

A combination of electrotransfer of MBD2‐antisense and bleomycin electrochemotherapy has an additive inhibitory effect on the rate of tumor growth and a synergistic effect on the number of tumor‐free animals when compared with either monotherapy.

Conclusions

Our results suggest that a combination of MBD2‐antisense electrotransfer gene therapy and chemotherapy with bleomycin is a candidate new approach to anticancer therapy. Copyright © 2003 John Wiley & Sons, Ltd.
Epidemiology of primary dystonias in Japan: Comparison with Western countries
Authors: Matsumoto Shinichi , Nishimura Masataka , Shibasaki Hiroshi , Kaji Ryuji
Published: October 2003

We performed epidemiological studies of primary dystonia in the city of Kyoto. The prevalence was at least 10.1 per 100,000 persons, which was similar to that in Western countries. Facial dystonia was more common than other types, which contrasts with that reported in Europe. Age of onset for both genders was in agreement with that in other countries. © 2003 Movement Disorder Society
Emotion recognition in patients with idiopathic Parkinson's disease
Authors: Yip James T.H. , Lee Tatia M.C. , Ho Shu‐Leong , Tsang Kin‐Lun , Li Leonard S.W.
Published: October 2003

Emotion recognition (ER) was examined in 64 patients with idiopathic Parkinson's disease (PD; 56 bilateral and 8 right‐sided) and 64 matched healthy volunteers. Participants were administered an ER battery, consisting of the following subscores: overall ER (OER), overall facial ER, facial emotion identification (FEI) and discrimination, overall prosodic ER, and prosodic emotion identification (PEI) and discrimination. Measures of visuospatial functions, auditory attention, and depression were also administered. After controlling for visuospatial functions, auditory attention and depression, results indicated that patients with bilateral PD had poorer performance on all ER subscores, regardless of the modality and type of experimental task involved, relative to healthy volunteers. However, patients with right‐sided PD had difficulty on FEI and PEI only. Whereas none of the clinical variables examined in this study predicted any of the ER subscores, visual organization and auditory attention positively predicted OER in patients with PD. In addition, visual organization also positively predicted FEI in these patients. Implications are discussed in terms of the neural substrates underlying ER. © 2003 Movement Disorser Society
Is FDG‐PET a useful tool in clinical practice for diagnosing corticobasal ganglionic degeneration?
Authors: Coulier Ilse M.F. , de Vries Joeke J. , Leenders Klaus L.
Published: October 2003

Seven consecutive patients were suspected to suffer from corticobasal ganglionic degeneration (CBGD) and were studied with [18]F‐fluorodeoxyglucose (FDG) PET imaging of the brain. At the time of their FDG‐PET scan, 4 of 7 patients fulfilled the clinical criteria of CBGD as proposed by Lang and associates [In Calne DB, 1994; Neurodegenerative disease. Philadelphia: Saunders]. For 2 of these 4 patients, however, an alternative clinical diagnosis was also considered. Three of the seven patients underwent an FDG‐PET scan when their clinical features were not yet developed sufficiently to confirm a clinical diagnosis of CBGD. Simple visual analysis of the FDG‐PET scans was carried out. All 7 patients showed an asymmetrical pattern of glucose metabolism that was demonstrated in previous studies to be characteristic in patients who had a clinical diagnosis of CBGD. The PET results helped to confirm the clinical suspicion of CBGD in 2 patients and to rule out other diagnoses in 2 other patients. For 3 patients with no sufficient symptoms to diagnose CBGD, the pattern of glucose metabolism was characteristically asymmetrical. A probable diagnosis of CBGD was made in these patients. Our results suggest that routine visual inspection of a cerebral FDG‐PET scan is a useful tool to confirm suspicion of the clinical diagnosis of probable CBGD, to differentiate from other hypokinetic‐rigid syndromes, and to support a diagnosis CBGD in patients who do not (yet) sufficiently fulfill the clinical criteria. © 2003 Movement Disorder Society
Subthalamic nucleus stimulation for Parkinson's disease preferentially improves akinesia of proximal arm movements compared to finger movements
Authors: Wenzelburger Roland , Kopper Florian , Zhang Bao‐Rong , Witt Karsten , Hamel Wolfgang , Weinert Dieter , Kuhtz‐Buschbeck Johann , Gölge Mukaddes , Illert Michael , Deuschl Günther , Krack Paul
Published: October 2003

Deep brain stimulation of the subthalamic nucleus (STN‐DBS) reduces akinesia in Parkinson's disease but its impact on fine motor functions was unknown. We assessed the effects of DBS and a levodopa (L‐dopa) test on the timing of the precision grip in 18 patients. Improvement on UPDRS‐items reflecting hand functions and the shortening of the first phases of the precision grip were more distinct in the L‐dopa test than in the pure STN‐DBS condition. Other akinesia items and the time for build‐up of lifting force were equally improved in both conditions. This suggests that routine STN‐DBS might not be equally effective on all aspects of fine motor functions. © 2003 Movement Disorder Society
Voxel‐based morphometry detects cortical atrophy in the Parkinson variant of multiple system atrophy
Authors: Brenneis Christian , Seppi Klaus , Schocke Michael F. , Müller Jörg , Luginger Elisabeth , Bösch Sylvia , Löscher Wolfgang N. , Büchel Christian , Poewe Werner , Wenning Gregor K.
Published: October 2003

To determine magnetic resonance imaging (MRI) patterns of brain atrophy in parkinsonian syndromes, we applied voxel‐based morphometry (VBM) to segmented gray matter, white matter, and cerebrospinal fluid compartments of T1‐weighted brain volumes of 12 patients with probable multiple system atrophy–parkinson variant (MSA‐P) and 12 Parkinson's disease patients, comparing them with 12 normal controls matched for age. In comparison to controls, a cortical atrophy pattern was observed in MSA‐P patients with significant clusters of volume loss in primary sensorimotor cortices bilateral, supplementary motor areas bilateral, right premotor cortex, prefrontal cortex bilateral (middle frontal gyri) and insular cortices bilateral; subcortical atrophy occurred bilaterally in caudate nuclei and putamen as well as in the midbrain. Furthermore, an enlargement of the cerebrospinal fluid compartment was found in the lateral ventricles, third ventricle, perimesencephalic and cerebellomedullar cavities. In PD patients, significant atrophy only occurred in left caudate head with enlargement of left lateral ventricle. Comparing MSA‐P to PD patients, MSA‐P showed a similar cortical pattern of atrophy as compared to controls. We conclude that VBM reveals selective cortical atrophy in patients with MSA‐P affecting primary and higher order motor areas as well as prefrontal and insular cortices. Further studies are required to determine clinical and/or subclinical correlates of cortical atrophy in MSA‐P. © 2003 Movement Disorder Society
One‐year treatment with standard and sustained‐release levodopa: Appropriate long‐term treatment of restless legs syndrome?
Authors: Trenkwalder Claudia , Seidel Victor Collado , Kazenwadel Jörg , Wetter Thomas C. , Oertel Wolfgang , Selzer Roland , Kohnen Ralf
Published: October 2003

To investigate the long‐term efficacy and safety of sustained‐release (SR) in combination with regular‐release (RR) levodopa/benserazide in the treatment of restless legs syndrome (RLS), an open‐label, prospective, extension study of a preceding double‐blind crossover trial was performed for 12 months. Twenty‐three severely disturbed RLS patients (7 men, 16 women) received a combination of RR and SR levodopa. Patients were treated on average for 10 months with a mean daily dose of 203 ± 101 mg of RR and of 185 ± 93 mg of SR levodopa. The mean daily total dose was 388 ± 162 mg levodopa. Efficacy was documented using patient's rating scales, sleep diaries, and investigator's global ratings with the Clinical Global Impressions (CGI). Ten of 23 patients completed the 1‐year extension. Between baseline of the crossover trial and endpoint of the extension study (last‐observation‐carried‐forward method, intention‐to‐treat population), quality of sleep improved (+3.5 ± 1.9, 7‐point scale), sleep latency was shortened (−131 ± 152 minutes), and total sleep time lengthened (+ 190 ± 136 minutes). Severity of RLS at time of falling asleep (−6.5 ± 3.4, 11‐point scale) and during the night (−6.0 ± 3.5) was markedly lower at the end of the extension but severity of RLS during the day (+1.9 ± 5.0) slightly increased. Of 13 dropouts, 8 patients discontinued therapy because of worsening RLS during the day. This trial shows that long‐term treatment with the combination of RR and SR levodopa/benserazide in RLS patients with late‐night problems was efficacious and not limited by tolerability problems in 40% of patients, whereas in the majority of patients, aggravating daytime problems required termination of the levodopa therapy within the 1‐year treatment period. © 2003 Movement Disorder Society
Impact of placebo assignment in clinical trials of Parkinson's disease
Authors: Goetz Christopher G. , Janko Kimberly , Blasucci Lucy , Jaglin Jean A.
Published: October 2003

Informed consent procedures in placebo‐controlled trials are developed to ensure that subjects entering studies clearly understand the possibility of placebo assignment. The extent of patient understanding and the impact of learning that they were assigned placebo treatment have not been extensively studied. By using a standardized questionnaire, we interviewed 50 consecutive placebo‐treated patients from 14 placebo‐controlled clinical trials of Parkinson's disease (PD) after completion of their involvement. All patients interviewed understood that their study contained a placebo arm. All had hoped to receive the study drug rather than placebo, and more than half the subjects believed they had improved clinically during their placebo exposure. Positive impressions of enrollment in placebo‐controlled trials were more frequent than negative, included helping to advance science (86%); liking the experience, education, and attention associated with the clinical trial (90%); and participating in research for the benefit of other patients as well as for themselves (80%). If another placebo‐controlled trial was offered, 88% expressed that they would possibly, likely, or definitely be interested in enrollment. PD patients clearly understand the concept of placebo‐controlled trials they complete them, but they are inaccurate in assessing whether they received placebo treatment. Although most wish they received the active compound being tested, the overall view of participation in placebo‐controlled trials is viewed very positively by PD patients. © 2003 Movement Disorder Society
Levodopa improves physical fatigue in Parkinson's disease: A double‐blind, placebo‐controlled, crossover study
Authors: Lou Jau‐Shin , Kearns Greg , Benice Theodore , Oken Barry , Sexton Gary , Nutt John
Published: October 2003

We quantitatively investigated the effect of carbidopa/levodopa (25/100) on physical fatigue during finger tapping and force generation in a double‐blind, placebo‐controlled crossover study. Parkinson's disease (PD) subjects were randomly assigned to carbidopa/levodopa or placebo for Visit 1 or 2 and participated in the following two studies: (1) Finger tapping. Twenty‐five PD patients used their index fingers to strike two keys 20 cm apart on a musical instrument digital interface (MIDI) keyboard. The slopes of the regression line of dwell time and movement time were used to assess the rate of fatigue development. (2) Force generation. Twelve PD patients contracted the wrist extensors maximally to obtain a baseline maximum voluntary contraction (BMVC) force. Then they repetitively contracted the wrist extensors at 50% of the BMVC for 7 seconds and rested for 3 seconds. An interval maximum voluntary contraction (IMVC) was measured every three repetitions. Fatigue was defined as an IMVC of less than 60% of the BMVC. The slope of the regression line of IMVC was used to assess the rate of force decline. These two studies were repeated 1 hour after carbidopa/levodopa (25/100) or placebo. Subjects filled out the Multidimensional Fatigue Inventory (MFI) at the beginning of the first visit. Results showed that the slope of dwell time decreased with levodopa but not with placebo (P = 0.004). The rate of force decline also decreased with levodopa but not with placebo (P = 0.01). The subscores in the dimension of physical fatigue in the MFI did not correlate with the rate changes in dwell time or the rate changes in force decline. We concluded that (1) levodopa improves physical fatigue in finger tapping and force generation, (2) physical fatigue in Parkinson's disease is at least partially related to dopamine deficiency, and (3) the MFI measures different aspects of physical fatigue compared with those measured by finger tapping and force generation. © 2003 Movement Disorder Society
Direct economic impact of Parkinson's disease: A research survey in the United Kingdom
Authors: Findley Leslie , Aujla Manjit , Bain Peter G. , Baker Mary , Beech Catherine , Bowman Clive , Holmes Jeremy , Kingdom Wendy K. , MacMahon Douglas G. , Peto Viv , Playfer Jeremy R.
Published: October 2003

The direct costs of care were evaluated prospectively in a sample of people with Parkinson's disease (PD) in the United Kingdom in 1998. The subjects were drawn from a random sample of general practitioner practices within a representative sample of 36 Regional Health Authorities and the equivalent. A total of 444 resource use questionnaires with usable data were returned (response rate, 59%). The total mean annual cost of care per patient for all patients by age was £5,993 (€9,554, n = 432). Hoehn and Yahr stage significantly (P < 0.001) influenced expenditure by stage as follows: 0 and I, £2,971 (€4,736, n = 110); II, £3,065 (€4,886, n = 89); III, £6,183 (€9,857, n = 120); IV, £10,134 (€16,155, n = 87); V, £18,358 (€29,265, n = 17). National Health Service costs accounted for approximately 38% and social services for 34% of the direct costs of care. Drug expenditure accounted for 24% of overall costs in the <65 years age group and 10% in patients aged >85 years. A move from home to residential care was associated with an approximately 500% cost increase. In conclusion, PD imposes significant direct costs on public services and on individuals. These costs should be taken into account when allocating public funds. © 2003 Movement Disorder Society
Randomized, double‐blind study of pramipexole with placebo and bromocriptine in advanced Parkinson's disease
Authors: Mizuno Yoshikuni , Yanagisawa Nobuo , Kuno Sadako , Yamamoto Mitsutoshi , Hasegawa Kazuko , Origasa Hideki , Kowa Hisayuki ,   The Japan Pramipexole Study Group
Published: October 2003

We compared the efficacy and safety of pramipexole (PPX) with placebo in the treatment of advanced Parkinson's disease (PD) as an adjunct to levodopa. A bromocriptine (BR) group was included to enable determination of the noninferiority of PPX relative to BR as the standard treatment. © 2003 Movement Disorder Society
REM sleep behavior disorder and vocal cord paralysis in Machado‐Joseph disease
Authors: Iranzo Alex , Muñoz Esteban , Santamaria Joan , Vilaseca Isabel , Milà Montserrat , Tolosa Eduardo
Published: October 2003

We evaluated the occurrence of REM sleep behaviour disorder (RBD) and vocal cord abductor paralysis (VCAP) in a group of 9 Machado‐Joseph disease (MJD) patients. RBD was diagnosed by clinical history plus audiovisual polysomnography in 4 men and 1 woman (55%). While dreaming, 4 fell out of the bed and the other injured his arms. Laryngoscopy detected bilateral VCAP in 1 patient with stridor who required emergency tracheotomy, and partial vocal cord abductor restriction in 2. RBD and VCAP are two potentially injurious conditions that should be considered part of the clinical spectrum of MJD. © 2003 Movement Disorder Society
VIM thalamic stimulation for tremor in a patient with IgM paraproteinaemic demyelinating neuropathy
Authors: Růžička Evžen , Jech Robert , Zárubová Kateřina , Roth Jan , Urgošík Dušan
Published: October 2003

We demonstrate the effect of deep brain stimulation of the ventral intermediate thalamic nucleus on intractable action tremor, in a 72‐year‐old man suffering from neuropathy associated with monoclonal gammopathy. © 2003 Movement Disorder Society
Periodic alternating nystagmus and rebound nystagmus in spinocerebellar ataxia type 6
Authors: Hashimoto Takao , Sasaki Osamu , Yoshida Kunihiro , Takei Yo‐ichi , Ikeda Shu‐ichi
Published: October 2003

We report on a family with ataxia type 6 (SCA6) showing peculiar oculomotor symptoms. The proband presented with periodic alternating nystagmus (PAN), and her 2 brothers had rebound nystagmus and gaze‐evoked nystagmus. They carried the identical mutation (the number of expanded CAG repeat, 24) in the CACNA1A gene. The intrafamilial variability of oculomotor symptoms may be ascribed to factors other than CAG repeat expansion size in SCA6. © 2003 Movement Disorder Society
Brainstem myoclonus in generalised tetanus
Authors: Warren Jason D. , Kimber Thomas E. , Thompson Philip D.
Published: October 2003

Electrophysiological studies of stimulus sensitive myoclonus in a man with generalised tetanus revealed features characteristic of hyperekplexia or brainstem myoclonus. © 2003 Movement Disorder Society