julilili42/papers huggingface dataset:
Oligonucleotide-Targeted RNase H Protection Analysis of RNA-Protein Complexes
2003-11-15

This chapter focuses on the analysis of RNA-protein complexes (RNPs, ribonucleoprotein particles) by oligonucleotide-targeted RNase H digestion, a powerful approach to probe the domain structure of an RNP, both in crude extracts and in purified preparations. RNase H requires DNA-RNA hybrids of at least four basepairs for cleaving the RNA strand (1). Therefore, RNase H-mediated cleavage can be directed to a short, specific RNA sequence by hybridization of a complementary DNA oligonucleotide. Oligonucleotide-targeted RNase H cleavage was first used to analyze RNA structures (see, e.g., the analysis of U1 snRNA secondary structure by Lazar and Jacob [2] and Rinke et al. [3], the characterization of RNA lariat structures [4], and the structural probing of a specific mRNA by Hwang et al. [5]). The approach has been adopted to RNA-protein complexes, and an important early example is the study of the basepairing potential of the 5′ end of U1 snRNA in the U1 snRNP (6). Subsequently, oligonucleotide-targeted RNase H cleavage has been used in various systems to disrupt RNPs for functional studies and in protection assays to map RNP protein-binding domains. Examples include in vitro studies that established snRNA sequences essential for spliceosome assembly and pre-mRNA splicing in the mammalian system and in yeast (see, e.g., refs. 7–12), studies on the assembly of the yeast and mammalian spliceosome (13,14), investigations on the U7 snRNA function in histone 3′ end processing (e.g., see ref. 15), and the analysis of snRNA requirements for trans splicing in trypanosome cells (16).
Analysis of Chimerism After Stem Cell Transplantation
2003-11-15

In recent years, considerable progress has been made in the analysis of hematopoietic chimerism after allogeneic stem cell transplantation (SCT). The introduction of polymerase chain reaction (PCR)-based methods for the characterization of "variable number of tandem repeats" (VNTR) and "short tandem repeats" (STR) has made it possible to analyze donor/recipient chimerism in a sensitive and quantitative fashion in all patients except those with syngeneic donors. In turn, this has allowed the serial monitoring of engraftment and study of the kinetics of graft failure and graft rejection after allo-SCT (-). These methods also allow the identification of patients with the highest risk of relapse after allo-SCT (-). In consequence, analysis of chimerism has become a routine diagnostic instrument at many centers performing allo-SCT (-). This chapter concentrates on PCR-based techniques for the amplification of STRs, using either (a) conventional PCR and polyacrylamide gel electrophoresis (PAGE) of reaction products or (b) PCR using fluorescent labeled primers with size resolution by capillary electrophoresis. Both methods are described in detail.
Study of Mutant Proteins With Folding Defects in Cultured Patient Cells
2003-11-15

AbstractThe example used in this chapter concerns the group of autosomal recessive inherited peroxisome biogenesis disorders (PBDs). These disorders are genetically heterogeneous with at least 12 different complementation groups (CGs). CG 1 is the largest characterized by mutations in HsPEX1, a gene encoding a 143 kD AAA protein (ATPases associated with diverse cellular activities) (1–3). About 65% of the PBD patients suffering from peroxisome biogenesis disorders harbor mutations in PEX1 (4–6). Although the function of PEX1 is not defined yet, it is likely that PEX1 plays a significant role in the import of peroxisomal matrix proteins from the cytosol—where they are translated—into the peroxisomes (1,2,7,8).KeywordsSodium Dodecyl SulfateG843D MutationZellweger SyndromeFolding DefectPeroxisome Biogenesis DisorderThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.
Time course of hepatitis A viremia and viral load in the blood of human hepatitis A patients
2003-11-17

Abstract The hepatitis A virus (HAV) is the most common etiological cause of acute hepatitis infections in humans in industrialized countries. Investigations into the viral load during HAV viremia, however, are rare. Therefore, correlation studies between viral load, biochemical, and specific serological markers have been undertaken. The group of sera comprised a series of multiple consecutive blood samples drawn from 11 patients at different times after onset of the disease. During the period up to 70 days after the onset of icterus, the individual range was at 1×10 3 to 3×10 4 HAV genome equivalents/ml. From day 75 until 120 after onset of the disease, the levels traced were at 10 3 . In one case, it was possible to trace 1.25×10 4 genome equivalents/ml up to 180 days after onset of icterus and in two cases even up to 408 and 490 days viral load levels of 5×10 3 and 4×10 4 were detected, respectively. The same sera were used to measure IgM class antibodies to hepatitis A virus and the total anti‐HAV. The results demonstrate that a direct correlation to peak levels of viral load exists with peak serum transaminase levels, but neither with peak anti‐HAV IgM levels nor with total anti‐HAV. Decreasing amounts of anti‐HAV IgM tend to occur with decreasing amounts of HAV genome equivalents; and, vice versa, increasing amounts of total anti‐HAV are accompanied by decreasing amounts of HAV genome equivalents. The longest duration of viremia was found in patients infected with HAV genotype IA. J. Med. Virol. 72:10–16, 2004. © 2004 Wiley‐Liss, Inc.
Atovaquone and Proguanil versus Amodiaquine for the Treatment of Plasmodium falciparum Malaria in African Infants and Young Children
2003-11-17

Malaria-related morbidity and mortality are greatest among young children in areas with high malaria transmission intensity. An open-label, randomized study was done to evaluate the efficacy and safety of the combination of atovaquone and proguanil formulated as pediatric-strength tablets (20 and 8 mg/kg of body weight, respectively, administered once daily for 3 days), compared with amodiaquine (10 mg/kg of body weight, once daily for 3 days), among children weighing > or =5 and <11 kg in Gabon. Two hundred patients aged 3-43 months were recruited. Use of atovaquone/proguanil resulted in a cure rate on day 28 of 95% (87 of 92 children), compared with 53% (41 of 78 children) for amodiaquine (difference, 42%; 95% CI, 30%-54%; P<.001). The incidence of adverse events was similar in both groups, and no serious adverse events were attributed to the use of atovaquone/proguanil. Atovaquone/proguanil was found to be highly effective and safe for the treatment of Plasmodium falciparum malaria in infants and young children weighing 5-10 kg in Africa.
Induction of chronic lymphocytic leukemia (CLL)–specific CD4- and CD8-mediated T-cell responses using RNA-transfected dendritic cells
2003-11-18

Abstract Recently, it was demonstrated that transfection of dendritic cells (DCs) with tumor-derived RNA can elicit effective T-cell responses. This technique does not require the definition of the tumor antigen or HLA haplotype of the patients. We applied this approach to induce HLA class I– and class II–restricted T-cell responses directed against malignant cells from patients with chronic lymphocytic leukemia (B-CLL). Here, we show that DCs generated from monocytes of patients with B-CLL induce leukemia-specific cytotoxic and proliferative T-cell responses on transfection with total RNA isolated from autologous leukemic B lymphocytes. Standard 51Cr-release assays showed specific major histocompatibility complex (MHC) class I–restricted cytotoxic activity against the autologous leukemic B cells and DCs transfected with CLL-RNA, whereas nonmalignant B cells were spared. The specificity of the cytotoxic T-lymphocyte (CTL) response was confirmed using cold target inhibition assays and by blocking HLA class I molecules. Furthermore, we established a protocol for the amplification of whole B-CLL mRNA. The use of DCs transfected with in vitro amplified B-CLL mRNA elicited specific T-cell responses similar to the results obtained with native mRNA. These data suggest that vaccinations using DCs transfected with RNA might be a potent new strategy in the treatment of CLL.
A single aquaporin gene encodes a water/glycerol/urea facilitator in Toxoplasma gondii with similarity to plant tonoplast intrinsic proteins1
2003-11-19

We describe a single aquaporin gene in Toxoplasma gondii which, surprisingly, has only 28% sequence similarity to the aquaglyceroporin of another apicomplexan parasite, Plasmodium falciparum . Sequence comparisons showed 47% similarity to water‐specific plant aquaporins and the conservation of typical pore‐forming residues. We established that the Toxoplasma aquaporin protein is a bifunctional membrane pore with intermediate water and high glycerol permeability. Furthermore, we identified hydroxyurea, an antineoplastic agent with inhibitory effects on parasite proliferation, as a permeant of this channel.
Contrasting roles of DAP10 and KARAP/DAP12 signaling adaptors in activation of the RBL‐2H3 leukemic mast cell line
2003-11-19

Abstract A common feature of hematopoietic activating immunoreceptors resides in their association at the cell surface with transmembrane signaling adaptors. Several adaptors, such as the CD3 molecules, FcRγ and KARAP/DAP12, harbor intracytoplasmic immunoreceptor tyrosine‐based activation motifs (ITAM) that activate Syk‐family protein tyrosine kinases. In contrast, another transmembrane adaptor, DAP10, bears a YxxM motif that delivers signals by activation of lipid kinase pathways. We show here that the human signal‐regulatory protein SIRPβ1 can associate with both DAP10 and KARAP/DAP12 in a model of RBL‐2H3 cell transfectants. In association with KARAP/DAP12, SIRPβ1 complexes are capable of inducing serotonin release and tumor necrosis factor (TNF) secretion. By contrast,in the absence of KARAP/DAP12, engagement of SIRPβ1:DAP10 complexes does not lead to detectable serotonin release or TNF secretion by RBL‐2H3 transfectants. However, triggering of SIRPβ1:DAP10 complexes co‐stimulates RBL‐2H3 effector function induced by sub‐optimal stimulation of the endogenous FcεRI complex. Therefore, we report here a cellular model in which the association of a cell surface receptor with various signaling adaptors dictates the co‐stimulatory or the direct stimulatory properties of the complex.
Visual-tracking neurons in area MST are activated during anticipatory pursuit eye movements
2003-11-20

Here, I report that rhesus monkeys are able to generate anticipatory smooth pursuit eye movements in the transient absence of a moving target, if this target moves periodically. The eye velocity before target reappearance was significantly larger if the target trajectory was predictable compared with the control condition consisting of unpredictable target trajectory. Parallel to the registration of the eye movements, single-unit activity was recorded from neurons in the middle superior temporal (MST) area of the two monkeys. The neuronal activity of visual-tracking neurons resembled the observed eye movements, i.e. these neurons increased their activity earlier if the movement of the target was predictable compared with the unpredictable control. These results provide further evidence for the existence of extra-retinal signals in the activity of visual-tracking neurons located in area MST.
Single-Chain Fv-Based Affinity Purification of the Cellular Stress Protein gp96 for Vaccine Development
2003-11-20

Cellular stress proteins like the classical heat-shock proteins (HSPs) hsp70 (, , ), hsp90 () and hsp110 (); the glucose-regulated proteins gp96/GRP94 (,) and grp170 (); as well as the endoplasmic chaperone calreticulin (,) have been shown to induce cytotoxic T-cell responses and protective immunity when isolated from tumor or infected cells and used to vaccinate animals (, , ; reviewed in refs. 11–17). The specificity of the immune responses is owing to antigenic peptides associated with the HSPs (, , , , , , , , ). The HSP-peptide complexes are taken up by professional antigen-presenting cells (APCs) for effective presentation of the peptides to T cells (,, , , , , , ). The extensively studied endoplasmic reticulum-resident chaperone gp96 is most efficient and promising in this regard (, , , , , ); encouraging clinical studies with human cancer patients ().
Comparison of longitudinal metabolite relaxation times in different regions of the human brain at 1.5 and 3 Tesla
2003-11-21

Abstract In vivo longitudinal relaxation times of N ‐acetyl compounds (NA), choline‐containing substances (Cho), creatine (Cr), myo ‐inositol (mI), and tissue water were measured at 1.5 and 3 T using a point‐resolved spectroscopy (PRESS) sequence with short echo time (TE). T 1 values were determined in six different brain regions: the occipital gray matter (GM), occipital white matter (WM), motor cortex, frontoparietal WM, thalamus, and cerebellum. The T 1 relaxation times of water protons were 26–38% longer at 3 T than at 1.5 T. Significantly longer metabolite T 1 values at 3 T (11–36%) were found for NA, Cho, and Cr in the motor cortex, frontoparietal WM, and thalamus. The amounts of GM, WM, and cerebrospinal fluid (CSF) within the voxel were determined by segmentation of a 3D image data set. No influence of tissue composition on metabolite T 1 values was found, while the longitudinal relaxation times of water protons were strongly correlated with the relative GM content. Magn Reson Med 50:1296–1301, 2003. © 2003 Wiley‐Liss, Inc.
Proton-neutron pairing in the deformed BCS approach
2003-11-24

We examine isovector and isoscalar proton-neutron pairing correlations for the ground state of even-even Ge isotopes with mass number A=64-76 within the deformed BCS approach. For N=Z 64Ge the BCS solution with only T=0 proton-neutron pairs is found. For other nuclear systems (N>Z) a coexistence of a T=0 and T=1 pairs in the BCS wave function is observed. A problem of fixing of strengths of isoscalar and isovector pairing interactions is addressed. A dependence of number of like and unlike pairs in the BCS ground state on the difference between number of neutrons and protons is discussed. We found that for nuclei with N much bigger than Z the effect of proton-neutron pairing is small but not negligible.
Combined Application of Stable Carbon Isotope Analysis and Specific Metabolites Determination for Assessing In Situ Degradation of Aromatic Hydrocarbons in a Tar Oil-Contaminated Aquifer
2003-11-25

To evaluate the intrinsic bioremediation potential in an anoxic tar oil-contaminated aquifer at a former gasworks site, groundwater samples were qualitatively and quantitatively analyzed by compound-specific isotope analysis (CSIA) and signature metabolites analysis (SMA). 13C/12C fractionation data revealed conclusive evidence for in situ biodegradation of benzene, toluene, o-xylene, m/p-xylene, naphthalene, and 1-methylnaphthalene. In laboratory growth studies, 13C/12C isotope enrichment factors for anaerobic degradation of naphthalene (epsilon = -1.1 +/- 0.4) and 2-methylnaphthalene (epsilon = -0.9 +/- 0.1) were determined with the sulfate-reducing enrichment culture N47, which was isolated from the investigated test site. On the basis of these and other laboratory-derived enrichment factors from the literature, in situ biodegradation could be quantified for toluene, o-xylene, m/p-xylene, and naphthalene. Stable carbon isotope fractionation in the field was also observed for ethylbenzene, 2-methylnaphthalene, and benzothiophene but without providing conclusive results. Further evidence for the in situ turnover of individual BTEX compounds was provided by the presence of acetophenone, o-toluic acid, and p-toluic acid, three intermediates in the anaerobic degradation of ethylbenzene, o-xylene, and p-xylene, respectively. A number of groundwater samples also contained naphthyl-2-methylsuccinic acid, a metabolite that is highly specific for the anaerobic degradation of 2-methylnaphthalene. Additional metabolites that provided evidence on the anaerobic in situ degradation of naphthalenes were 1-naphthoic acid, 2-naphthoic acid, 1,2,3,4-tetrahydronaphthoic acid, and 5,6,7,8-tetrahydronaphthoic acid. 2-Carboxybenzothiophene, 5-carboxybenzothiophene, a putative further carboxybenzothiophene isomer, and the reduced derivative dihydrocarboxybenzothiophene indicated the anaerobic conversion of the heterocyclic aromatic hydrocarbon benzothiophene. The combined application of CSIA and SMA, as two reliable and independent tools to collect direct evidence on intrinsic bioremediation, leads to a substantially improved evaluation of natural attenuation in situ.
Efficacy of cytotoxic agents used in the treatment of testicular germ cell tumours under normoxic and hypoxic conditions in vitro
2003-11-25

Platinum-based chemotherapy is the main treatment element to achieve cure for patients with metastatic germ cell tumours. Drug resistance in testicular germ cell tumours (TGCTs) is rare and the reasons are not fully understood. While recent investigations have indicated decreased efficacy of chemotherapy in several tumour types under hypoxic conditions, this aspect has not been investigated in TGCTs so far. Furthermore, for cisplatin - the most active drug in this disease - controversial effects of hypoxia on cytotoxic efficacy have been reported. The relative efficacy of cytotoxic agents for the treatment of TGCT patients was studied in three different cell lines derived from human embryonal carcinomas (EC) in an in vitro hypoxia model. NT2, 2102 EP, and NCCIT were tested for their sensitivity towards cisplatin, etoposide, bleomycin, 4-OOH-ifosfamide, carboplatin, paclitaxel, gemcitabine, oxaliplatin, irinotecan, and mitomycin C under normoxic and hypoxic conditions using the MTT assay. Inhibitory concentrations IC(50) of the tested agents under both conditions were compared. Selected results were confirmed by flow-cytometric assessment of the apoptotic index. In all cells, doubling times were prolonged in hypoxia (NT2<NCCIT<2102 EP). All drugs were less effective under hypoxic conditions, including mitomycin C (eg, 1.6-fold increase of IC(50) in hypoxia compared to normoxia for NT2) and cisplatin (eg, NT2: two-fold increase). The relative effect of hypoxia on the IC(50) depended mainly on the cell line, and to a lesser extent on the drug. The results indicate that the reduced cell proliferation in hypoxia might be an important factor, but not the only determinant of a reduced cytotoxicity. In view of the broad spectrum of drugs with different modes of action tested, the relative resistance cannot be mediated by substance-specific resistance mechanisms like hypoxia-induced upregulation of P-glycoprotein or increased DNA-repair capacity, since many unrelated drugs were affected to a comparable extent in their efficacy by hypoxia. This study also provides the rationale to test the hypothesis whether improving tumour oxygenation by raising haemoglobin concentrations, for example, with erythropoietin in patients with TGCTs receiving chemotherapy may improve the outcome.
Investigation of Laser-Induced Choroidal Neovascularization in the Rat
2003-11-25

In the rat model of laser-induced neovascularization, newly developed endothelial cells originate from the retina and the choroid. Whereas ring-like surrounding vessels come from the retina, flat endothelial cells in deeper layers are of choroidal origin or may originate from circulating endothelial precursor cells. Dextran angiography has to be regarded critically for visualizing the choriocapillaris and CNV in laser scars. PECAM-1 immunohistochemistry is best for detection and quantification of neovascularization in laser scars.
Writhe of center vortices and topological charge: An explicit example
2003-11-25

The manner in which continuum center vortices generate topological charge\\ndensity is elucidated using an explicit example. The example vortex\\nworld-surface contains one lone self-intersection point, which contributes a\\nquantum 1/2 to the topological charge. On the other hand, the surface in\\nquestion is orientable and thus must carry global topological charge zero due\\nto general arguments. Therefore, there must be another contribution, coming\\nfrom vortex writhe. The latter is known for the lattice analogue of the example\\nvortex considered, where it is quite intuitive. For the vortex in the\\ncontinuum, including the limit of an infinitely thin vortex, a careful analysis\\nis performed and it is shown how the contribution to the topological charge\\ninduced by writhe is distributed over the vortex surface.\\n
Atypical dermatofibroma-like pattern of a melanoma on dermoscopy
2003-11-25

Department of Dermatology, University of Tuebingen, Tuebingen, Germany Correspondence to Andreas Blum, Department of Dermatology, University of Tuebingen, Liebermeisterstrasse 25, 72076 Tuebingen, Germany. Tel: +49 7071 2983766; fax: +49 7071 294561; e-mail: [email protected]
Protracted infusional 5-fluorouracil plus high-dose folinic acid combined with bolus mitomycin C in patients with gastrointestinal cancer: a phase I/II dose escalation study
2003-11-25

The aim of this study was to define the maximum tolerated dose (MTD) of bolus mitomycin C (MMC) in combination with 24 h-continuous infusion of 5-flourouracil (FU) plus folinic acid, and to assess the toxicity and activity in patients with previously treated colorectal and gastric cancer. Escalating doses of MMC starting from 6 mg m(-2) in 2 mg m(-2)-steps to a maximum of 10 mg m(-2) were applied on days 1 and 22, given to fixed doses of 5-FU (2.600 mg m(-2)) as 24 h infusion and folinic acid 500 mg m(-2) prior to 5-FU weekly for 6 weeks. At least three patients were treated at each dose level. A total of 16 patients have been included in the phase I study. At the highest dose level (MMC 10 mg m(-2)), grade III thrombocytopenia, dyspnoea, mucositis and diarrhoea were observed in one patient each (17 %). In the phase II study 45 patients, 33 with colorectal cancer and 12 with gastric cancer, 23 patients after failure of first- and 22 patients after at least second-line or subsequent chemotherapy have been treated. Seven partial responses (PR) were registered (16%), one (3%; CI(95%), 0-16) in colorectal and six (50%; CI(95%), 21-79%) in gastric cancer patients. In all, 17 (38%) achieved disease stabilisation, 15 colorectal (45%, CI(95%), 28-64%) and two gastric cancer patients (17%; CI(95%), 2-48%). The median progression-free survival was 3.1 months (range, 0.9-9.1) in colorectal and 4.6 months (range, 0.7-12.4) in gastric cancer. The median overall survival time was 6.6 months (range, 1.9-15.6) in colorectal and 7.1 months (range, 1.7-20.8) in patients with gastric cancer. This regimen was considered to be safe and well tolerated for pretreated patients with gastrointestinal adenocarcinoma. In gastric cancer,MMC plus infusional 5-FU/folinic acid may be a potential second-line regimen with promising antitumour activity.