bluuebunny/crossref_metadata_2025_split huggingface dataset:
An efficient gene transfer method mediated by ultrasound and microbubbles into the kidney
Authors: Koike Hiromi , Tomita Naruya , Azuma Haruhito , Taniyama Yoshiaki , Yamasaki Keita , Kunugiza Yasuo , Tachibana Katsuro , Ogihara Toshio , Morishita Ryuichi 
Published: November 2004

AbstractBackgroundSafety issues are of paramount importance in clinical human gene therapy. From this point of view, it would be better to develop a novel non‐viral efficient gene transfer method. Recently, it was reported that ultrasound exposure could induce cell membrane permeabilization and enhance gene expression.MethodsIn this study, we examined the potential of ultrasound for gene transfer into the kidney. First, we transfected rat left kidney with luciferase plasmid mixed with microbubbles, Optison, to optimize the conditions (duration of ultrasound and concentration of Optison). Then, 4, 7, 14 and 21 days after gene transfer, luciferase activity was measured. Next, localization of gene expression was assessed by measuring luciferase activity and green fluorescent protein (GFP) expression. Expression of GFP plasmid was examined under a fluorescence microscope at 4 and 14 days after gene transfer. Finally, to examine the side effects of this gene transfer method, biochemical assays for aspartate aminotransferase (AST), alanine aminotransferase (ALT), blood urea nitrogen (BUN) and creatinine (Cre) were performed.ResultsOptison and/or ultrasound significantly enhanced the efficiency of gene transfer and expression in the kidney. Especially, 70–80% of total glomeruli could be transfected. Also, a significant dose‐dependent effect of Optison was observed as assessed by luciferase assay (Optison 25%: 12.5 × 105 relative light units (RLU)/g tissue; 50%: 31.3 × 105 RLU/g tissue; 100%: 57.9 × 105 RLU/g tissue). GFP expression could be observed in glomeruli, tubules and interstitial area. Results of blood tests did not change significantly after gene transfer.ConclusionsOverall, an ultrasound‐mediated gene transfer method with Optison enhanced the efficiency of gene transfer and expression in the rat kidney. This novel non‐viral method may be useful for gene therapy for renal disease. Copyright © 2004 John Wiley & Sons, Ltd.
Chronic nicotine and smoke treatment modulate dopaminergic activities in ventral tegmental area and nucleus accumbens and the γ‐aminobutyric acid type B receptor expression of the rat prefrontal cortex
Authors: Li Shu‐Peng , Park Moon Seok , Kim Jong Hun , Kim Myeong Ok 
Published: November 2004

AbstractDopaminergic afferents from the mesencephalic areas, such as ventral tegmental area (VTA), synapse with the γ‐aminobutyric acid (GABA)‐ergic interneurons in the prefrontal cortex (PFC). Pharmacological and electrophysiological data show that the reinforcement, the dependence‐producing properties, as well as the psychopharmacologic effects of nicotine depend to a great extent on activation of nicotinic receptors within the mesolimbocortical dopaminergic projection. To explore further the relationship between the mesencephalic dopaminergic neurons and PFC GABAergic neurons, we investigated the effects of nicotine and passive exposure to cigarette smoke on the regulation of tyrosine hydroxylase (TH) in VTA and substantia nigra (SNC) and dopamine (DA) D1 receptor levels in nucleus accumbens (NAc) and caudate‐putamen (CPu). Also, the simultaneous changes in GABAB receptors mRNAs in the PFC were studied. The results showed that chronic nicotine and smoking treatment differentially changed the levels of TH protein in VTA and SNC and DA D1 receptor levels in Nac and CPu. GABAB1 and GABAB2 receptor mRNA levels also showed different change patterns. Ten and thirty minutes of smoke exposure increased GABAB1 receptor mRNA to a greater extent than that of GABAB2, whereas GABAB2 was greatly enhanced after 1 hr of smoke exposure. The TH levels in VTA were closely related to DA D1 receptor levels in NAc and with GABAB receptor mRNA changes in PFC. These results suggest that the mesolimbic pathway and GABAB receptor mRNA in PFC are modulated by nicotine and cigarette smoke, implying an important role in nicotine's psychopharmacological effects. © 2004 Wiley‐Liss, Inc.
In vivo optical imaging of tone‐evoked activity in the dorsal cochlear nucleus with a voltage sensitive dye
Authors: Kaltenbach J.A. , Zhang J.S. 
Published: November 2004

AbstractWe investigated the use of optical imaging for observing the spatial patterns of neural activation in the dorsal cochlear nucleus (DCN) of hamsters during tonal stimulation. The patterns of activation were studied in the DCN, in vivo, following application of a voltage sensitive dye, Di‐2‐ANEPEQ, to the DCN surface. Beginning 60–90 min following dye application, tones were presented to the ipsilateral ear. Electrophysiological recordings after dye application revealed no significant toxicity of Di‐2‐ANEPEQ that affected the frequency‐tuning properties of DCN neurons. We examined areas of activation in response to each of a series of test stimuli consisting of pure tones ranging in frequency from 2 to 20 kHz. For each stimulus condition, images were collected over a stimulus interval of 400 msec and averaged over 32 stimulus repetitions. These images revealed areas of activation with definable epicenters. The epicenters shifted from lateral to more medial locations on the DCN surface with increases in stimulus frequency. Comparison with electrophysiological data indicated a close parallel between the tonotopic gradient defined by optical imaging and that defined by the distribution of characteristic frequencies. The principal temporal and spatial features of these optical responses are described. © 2004 Wiley‐Liss, Inc.
Climate dynamics of atmosphere and ocean in the equatorial zone: a synthesis
Authors: Hastenrath Stefan , Lamb Peter J. 
Published: November 2004

AbstractA synopsis is offered of circulation mechanisms in the oceanic regions of the equatorial zone. Over the eastern Atlantic and Pacific, and especially in boreal summer, cross‐equatorial flow from the Southern Hemisphere is strong and induces a tongue of cold surface waters, centred to the south of the equator. Upon crossing the equator in these sectors, owing to the Coriolis effect and a kinetic energy imbalance, the airstream speeds up and divergence develops, producing the Intertropical Divergence Zone (ITDZ). Once these processes result in the wind recurving from southeasterly to southwesterly, the flow slows down and becomes convergent, manifest in the Intertropical Convergence Zone, with a maximum to the south of the wind confluence. By contrast, over the western Atlantic and central Pacific and especially in boreal winter, winds in the equatorial band are predominantly from the east, upper‐ocean Ekman transport is directed away from the equator, and the upwelling and cold tongue are centred on the equator. Cross‐equatorial flow is insufficient to produce recurvature, the ITDZ is narrower and weaker, the divergence maximum is at the equator rather than in low northern latitudes, and the convergence maximum straddles the wind confluence.Over the Indian Ocean, the wind field is dominated by the alternation between the predominantly meridional flow of the winter and summer monsoons. Equatorial westerlies are limited to the short monsoon transition seasons. Essential for their origin is an eastward pressure gradient along the equator and weak southern trade winds, allowing recurvature somewhat south of the equator. Because the zonal pressure gradient is strongest in boreal summer and the southern trade winds are weakest in austral summer, the equatorial westerlies peak in spring and autumn. The boreal autumn equatorial westerlies are the surface manifestation of a powerful zonal–vertical circulation cell along the Indian Ocean equator. Equatorial zonal–vertical circulation cells require well‐developed zonal flow in the lower troposphere along the equator and, therefore, appear confined to the oceanic longitudes and certain seasons. Thus, they are found over the Atlantic only in boreal winter and over the Indian Ocean only in boreal autumn, whereas over the Pacific they prevail all year round. Copyright © 2004 Royal Meteorological Society
Expression of growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15), and BMP receptors in the ovaries of goats
Authors: Silva J.R.V. , van den Hurk R. , van Tol H.T.A. , Roelen B.A.J. , Figueiredo J.R. 
Published: November 2004

AbstractThe process of ovarian folliculogenesis is composed of proliferation and differentiation of the constitutive cells in developing follicles. In goats, relatively little information is available on the local factors that regulate this process. We studied the presence and distribution of growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15), and BMP receptors types 2 (BMPR2), 1A (BMPR1A), and 1B (BMPR1B) in goat ovaries to find evidence for their possible roles in folliculogenesis. Ovaries of cyclic goats were collected and fixed in paraformaldehyde for immunohistochemical localization of GDF9 and BMP15 proteins or used to collect follicles and luteal tissue to study the mRNA expression of GDF9, BMP15, and BMP receptors using reverse transcriptase polymerase chain reaction (RT‐PCR). GDF9 and BMP15 proteins were found in oocytes of all types of follicles and granulosa cells of primary, secondary, and antral but not primordial follicles. The mRNAs for GDF9, BMP15, BMPR2, BMPR1A, and BMPR1B were detected in primordial, primary, and secondary follicles as well as in oocyte and granulosa cells of antral follicles. Transcripts for BMPR2, BMPR1A, BMPR1B, and GDF9, and GDF9 protein were furthermore found in corpora lutea. It is concluded that the mRNAs and proteins of GDF9 and BMP15 and the mRNAs of BMP receptors are expressed in goat ovarian follicles at all stages of their development, and that they form a complex intrafollicular regulatory system during folliculogenesis. Expression of all BMP receptor mRNAs and GDF9 mRNA and protein in luteal tissue additionally points to a role of GDF9 in corpus luteum function. Mol. Reprod. Dev. 70: 11–19, 2004. © 2004 Wiley‐Liss, Inc.
Mouse epiblasts change responsiveness to BMP4 signal required for PGC formation through functions of extraembryonic ectoderm
Authors: Okamura Daiji , Hayashi Katsuhiko , Matsui Yasuhisa 
Published: November 2004

AbstractMouse primordial germ cells (PGCs) are initially identified as a cluster of alkaline phosphatase (AP)‐positive cells within the extraembryonic mesoderm near the posterior part of the primitive streak at embryonic day (E) 7.25. Clonal analysis of epiblast cells has revealed that the putative precursors of PGCs are localized in the proximal epiblast, and we demonstrated that the conditions required for PGC formation are induced in the proximal region of epiblasts by extraembryonic ectoderm. Bone morphogenetic protein (BMP) 4 and BMP8b, which belong to the transforming growth factor‐β (TGF‐β) superfamily, might generate induction signals from extraembryonic ectoderm. Smad1 and Smad5, which are intracellular signaling molecules for BMP4, might also play a critical role in stimulating epiblasts to form PGC. However, how pluripotential epiblasts temporally and spatially respond to BMP signals to form PGCs remains unclear. The present study examines changes of responsiveness to BMP4 for PGC formation in epiblasts and their molecular mechanisms. We initially examined the effect of recombinant human (rh) BMP4 upon cultured epiblasts at different developmental stages, and found that they acquire the ability to respond to BMP4 signals for PGC formation between E5.25 and E5.5. In addition, such competence was conferred upon epiblasts by the extraembryonic ectoderm. We also showed that the increased expression of Smad1 and the onset of Smad5 expression induced by extraembryonic ectoderm might be responsible for quick acquisition of this competence. Furthermore, we show that only proximal epiblast cells maintain responsiveness to BMP4 for PGC formation at E6.0, and that this is associated with the proximal epiblast‐specific expression of Smad5. These results explain why only the proximal region of epiblasts can sustain the ability to form PGCs. Mol. Reprod. Dev. 70: 20–29, 2004. © 2004 Wiley‐Liss, Inc.
Oxygen‐regulated expression of GLUT‐1, GLUT‐3, and VEGF in the mouse blastocyst
Authors: Kind Karen L. , Collett Rachael A. , Harvey Alexandra J. , Thompson Jeremy G. 
Published: November 2004

AbstractThe oxygen concentration used in the incubation atmosphere during embryo culture influences embryo development rates and embryo quality. In somatic cells, oxygen levels can influence the expression of a range of genes, including glucose transporters, glycolytic enzymes, and angiogenic growth factors. Many of these oxygen‐regulated genes have important roles in embryonic development and metabolism. The aim of this study was to determine whether oxygen regulates gene expression in the preimplantation mouse blastocyst. Mouse embryos were cultured from the 1‐cell to morula stage under 7% oxygen, followed by culture under 20, 7, or 2% oxygen to the blastocyst stage. Expression of glucose transporter (GLUT)‐1, GLUT‐3, and vascular endothelial growth factor (VEGF) in blastocysts was measured by real‐time reverse transcription PCR. Development from morula to blastocyst was not altered by culture under different oxygen conditions. Expression of GLUT‐1, GLUT‐3, and vascular endothelial growth (VEGF) was increased by 2‐ to 4‐fold in embryos cultured under 2% oxygen, when compared to embryos cultured under 20 or 7% oxygen, and when compared to embryos developed in vivo (all P < 0.001). These results suggest that the preimplantation mouse embryo has the capacity to detect and respond to low oxygen availability with changes in expression of oxygen‐regulated genes. Mol. Reprod. Dev. 70: 37–44, 2004. © 2004 Wiley‐Liss, Inc.
TESSP‐1: A novel serine protease gene expressed in the spermatogonia and spermatocytes of adult mouse testes
Authors: Takano Naoharu , Matsui Hitoshi , Takahashi Takayuki 
Published: November 2004

AbstractA cDNA encoding a novel type of serine protease, designated testis‐specific serine protease 1 (TESSP‐1), was cloned using mRNA isolated from the adult mouse testis. The open reading frame of this cDNA codes for a protein of 322 amino acids, which includes a hydrophobic signal peptide of 18 amino acids and an N‐terminal activation peptide of 34 amino acids. The protein has an additional hydrophobic amino acid sequence at the C‐terminus. Expression of the TESSP‐1 gene was restricted to the testis. TESSP‐1 mRNA expression initiated in the mouse testis at 2 weeks after birth, and its level increased steadily with sexual maturation of the animal. In situ hybridization analysis revealed that TESSP‐1 mRNA was expressed in type B spermatogonia and spermatocytes at stages between preleptotene and pachytene. The testis contained at least five distinct forms of TESSP‐1 transcript, which presumably resulted from alternative splicing of the mRNA, but only one of these transcripts encodes a complete, functional enzyme. Expression experiments using COS‐7 cells showed that TESSP‐1 was synthesized as a glycoprotein with N‐glycosylated carbohydrates. Tests also showed that the C‐terminal hydrophobic region of TESSP‐1 was important upon its binding to the membrane by anchoring through glycosylphosphatidylinositol (GPI). Mol. Reprod. Dev. 70: 1–10, 2004. © 2004 Wiley‐Liss, Inc.
Detection and expression of hepatitis B virus X gene in one and two‐cell embryos from golden hamster oocytes in vitro fertilized with human spermatozoa carrying HBV DNA
Authors: Ali Bahy‐Ahmed , Huang Tian‐Hua , Xie Qing‐Dong 
Published: November 2004

AbstractThe main objectives of this study were to introduce motile human sperm carrying Hepatitis B virus (HBV) DNA to golden hamster oocytes in a co‐culture environment and to detect the replication and expression of the HBx gene in early embryonic cells. Zona‐free hamster oocytes were inseminated with human sperm carrying pBR322‐HBV DNA plasmid using the in vitro fertilization (IVF) technique. Both the one‐ and two‐cell stages of early embryonic development were studied. PCR, RT‐PCR, and Dot hybridization were performed to observe the HBx gene and its expression in these stages. “Fluorescence in situ hybridization” (FISH) was carried out to confirm the integration of HBV into the pronucleus, nucleus, and the chromosomes of embryos. The results showed that we have the ability to obtain a fertilization rate of 80%. RT‐PCR showed that the HBx gene could be expressed in both one‐ and two‐cell stages of embryonic development. The data suggested the possibility of sperm as a vector for the vertical transmission of HBV DNA to the next generation. Mol. Reprod. Dev. 70: 30–36, 2004. © 2004 Wiley‐Liss, Inc.
Morphological abnormalities in the spermatozoa of fertile and infertile men
Authors: Kubo‐Irie Miyoko , Matsumiya Kiyomi , Iwamoto Teruaki , Kaneko Satoru , Ishijima Sumio 
Published: November 2004

AbstractThe morphological analysis of the spermatozoa from fertile and infertile men was performed using light and electron microscopy to clarify the relationship between sperm morphology and fertility. Semen samples obtained from 22 partners of pregnant women were prepared according to the protocol standardized in an international collaborative study. Semen samples from 17 patients with asthenozoospermia or varicocele were collected in a hospital. Abnormalities in the spermatozoa were classified into three types for the tails, two for the midpieces, and six for the heads according to the criteria adapted from WHO guidelines (World Health Organization, 1999: WHO laboratory manual for the examination of human semen and semen–cervical mucus interaction (4th edition)). Approximately 14% of the spermatozoa from the fertile men had abnormal tails at the light microscopic level while approximately 44% had abnormal heads. Most types of abnormalities found in the spermatozoa from the asthenozoospermic and varicocele patients were encountered in those from the fertile men, although the semen from the fertile men contained a higher percentage of normal spermatozoa than that from the patients. These results were also confirmed at the ultrastructural level. Most abnormal cell types are encountered in semen from fertile men, although the incidence of abnormalities is low. Mol. Reprod. Dev. 70: 70–81, 2004. © 2004 Wiley‐Liss, Inc.
Implications of structural genomics target selection strategies: Pfam5000, whole genome, and random approaches
Authors: Chandonia John‐Marc , Brenner Steven E. 
Published: November 2004

AbstractStructural genomics is an international effort to determine the three‐dimensional shapes of all important biological macromolecules, with a primary focus on proteins. Target proteins should be selected according to a strategy that is medically and biologically relevant, of good value, and tractable. As an option to consider, we present the “Pfam5000” strategy, which involves selecting the 5000 most important families from the Pfam database as sources for targets. We compare the Pfam5000 strategy to several other proposed strategies that would require similar numbers of targets. These strategies include complete solution of several small to moderately sized bacterial proteomes, partial coverage of the human proteome, and random selection of approximately 5000 targets from sequenced genomes. We measure the impact that successful implementation of these strategies would have upon structural interpretation of the proteins in Swiss‐Prot, TrEMBL, and 131 complete proteomes (including 10 of eukaryotes) from the Proteome Analysis database at the European Bioinformatics Institute (EBI). Solving the structures of proteins from the 5000 largest Pfam families would allow accurate fold assignment for approximately 68% of all prokaryotic proteins (covering 59% of residues) and 61% of eukaryotic proteins (40% of residues). More fine‐grained coverage that would allow accurate modeling of these proteins would require an order of magnitude more targets. The Pfam5000 strategy may be modified in several ways, for example, to focus on larger families, bacterial sequences, or eukaryotic sequences; as long as secondary consideration is given to large families within Pfam, coverage results vary only slightly. In contrast, focusing structural genomics on a single tractable genome would have only a limited impact in structural knowledge of other proteomes: A significant fraction (about 30–40% of the proteins and 40–60% of the residues) of each proteome is classified in small families, which may have little overlap with other species of interest. Random selection of targets from one or more genomes is similar to the Pfam5000 strategy in that proteins from larger families are more likely to be chosen, but substantial effort would be spent on small families. Proteins 2005. © 2004 Wiley‐Liss, Inc.
Associative mechanism for phosphoryl transfer: A molecular dynamics simulation of Escherichia coli adenylate kinase complexed with its substrates
Authors: Krishnamurthy Harini , Lou Hongfeng , Kimple Adam , Vieille Claire , Cukier Robert I. 
Published: November 2004

AbstractThe ternary complex of Escherichia coli adenylate kinase (ECAK) with its substrates adenosine monophosphate (AMP) and Mg‐ATP, which catalyzes the reversible transfer of a phosphoryl group between adenosine triphosphate (ATP) and AMP, was studied using molecular dynamics. The starting structure for the simulation was assembled from the crystal structures of ECAK complexed with the bisubstrate analog diadenosine pentaphosphate (AP5A) and of Bacillus stearothermophilus adenylate kinase complexed with AP5A, Mg2+, and 4 coordinated water molecules, and by deleting 1 phosphate group from AP5A. The interactions of ECAK residues with the various moieties of ATP and AMP were compared to those inferred from NMR, X‐ray crystallography, site‐directed mutagenesis, and enzyme kinetic studies. The simulation supports the hypothesis that hydrogen bonds between AMP's adenine and the protein are at the origin of the high nucleoside monophosphate (NMP) specificity of AK. The ATP adenine and ribose moieties are only loosely bound to the protein, while the ATP phosphates are strongly bound to surrounding residues. The coordination sphere of Mg2+, consisting of 4 waters and oxygens of the ATP β‐ and γ‐phosphates, stays approximately octahedral during the simulation. The important role of the conserved Lys13 in the P loop in stabilizing the active site by bridging the ATP and AMP phosphates is evident. The influence of Mg2+, of its coordination waters, and of surrounding charged residues in maintaining the geometry and distances of the AMP α‐phosphate and ATP β‐ and γ‐phosphates is sufficient to support an associative reaction mechanism for phosphoryl transfer. Proteins 2005. © 2004 Wiley‐Liss, Inc.