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At least 1,441 records · Page 80Linked to original sources
PubMed1990

Human tumor fluorouracil trapping: clinical correlations of in vivo 19F nuclear magnetic resonance spectroscopy pharmacokinetics.

We previously reported that fluorouracil (5FU) accumulation and metabolism in human livers and tumors can be studied by in vivo nuclear magnetic resonance spectroscopy (NMRS). We have extended these observations by evaluating the pharmacokinetics of 5FU in the tumors of 11 patients with carcinoma of the breast, colon, endometrium, cervix, and kidney, using 19F-NMRS in a 1.5 Magnetom (Siemens Medical Systems, Cerrito, CA) magnetic resonance imaging unit (MRI). These NMRS measurements detected a long-lived tumor pool of 5FU in six of 11 tumors in our patients including carcinomas in the pelvis, breast, lung, and liver. The half-life (T1/2) of this tumor pool of "trapped" 5FU was 0.33 to 1.3 hours (20 to 78 minutes), much longer than the T1/2 of 5FU in blood (5 to 15 minutes). Neither the anabolites of 5FU (fluorinated nucleosides, nucleotides, 5FU-RNA, or 5FU-thymidylate synthase) nor the catabolites (eg, fluorobetaalanine [FBAL]) were detectable by 19F NMRS. Patient response to chemotherapy appeared to correlate with the extent of trapping of free 5FU in the human tumors: in the seven patients receiving 5FU, or 5FU or FUdR plus leucovorin, four of four patients whose tumors trapped 5FU responded to fluorinated pyrimidine chemotherapy, whereas three patients in whom there was a failure to detect tumor trapping were resistant to 5FU. We conclude that NMRS is clinically feasible, and enables investigators to study 5FU pharmacokinetics and metabolism in tumors in vivo. 19F-NMRS of 5FU allows for in vivo evaluation of 5FU metabolic modulation and might be able to guide therapeutic decisions.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1999

C-C chemokines in allergen-induced late-phase cutaneous responses in atopic subjects: association of eotaxin with early 6-hour eosinophils, and of eotaxin-2 and monocyte chemoattractant protein-4 with the later 24-hour tissue eosinophilia, and relationship to basophils and other C-C chemokines (monocyte chemoattractant protein-3 and RANTES).

The relationship of expression of the C-C chemokines eotaxin, eotaxin 2, RANTES, monocyte chemoattractant protein-3 (MCP-3), and MCP-4 to the kinetics of infiltrating eosinophils, basophils, and other inflammatory cells was examined in allergen-induced, late-phase allergic reactions in the skin of human atopic subjects. EG2+ eosinophils peaked at 6 h and correlated significantly with eotaxin mRNA and protein, whereas declining eosinophils at 24 h correlated significantly with eotaxin-2 and MCP-4 mRNA. In contrast, no significant correlations were observed between BB1+ basophil infiltrates, which peaked at 24 h, and expression of eotaxin, eotaxin-2, RANTES, MCP-3, and MCP-4 or elastase+ neutrophils (6-h peak), CD3+ and CD4+ T cells (24 h), and CD68+ macrophages (72 h). Furthermore, 83% of eosinophils, 40% of basophils, and 1% of CD3+ cells expressed the eotaxin receptor CCR3, while eotaxin protein was expressed by 43% of macrophages, 81% of endothelial cells, and 6% of T cells (6%). These data suggest that 1) eotaxin has a role in the early 6-h recruitment of eosinophils, while eotaxin-2 and MCP-4 appear to be involved in later 24-h infiltration of these CCR3+ cells; 2) different mechanisms may guide the early vs late eosinophilia; and 3) other chemokines and receptors may be involved in basophil accumulation of allergic tissue reactions in human skin.

Adolescent↗
PubMed1997

Update on techniques in the diagnosis of Chlamydia trachomatis infections.

There are two major advances in the laboratory diagnosis of Chlamydia trachomatis, one lies in the use of nucleic acid amplification techniques and the second in the evaluation of urine as an alternative to invasive sampling of urethral and cervical specimens. There is however, the problem of inhibitors in urine that needs to be addressed, in order for this method to achieve 100% sensitivity. The Q-beta (Q beta) replicase test, Gen-Probe transcription-mediated amplification (TMA) test and the nucleic acid sequence-based amplification (NASBA) test are some of the newer nucleic acid amplification methods being evaluated for the detection of C. trachomatis. These are RNA-based amplification techniques that can potentially achieve very high levels of sensitivity because of the presence of multiple RNA copies in microorganisms and may also be useful for detecting active infection. Q-beta has been found to be less subjected to inhibitory substances in urine than polymerase chain reaction (PCR). Cell culture remains the gold standard for the legal diagnosis of C. trachomatis infections and is the method of choice for the detection of infection at uncommon sites. It forms part of the expanded gold standard for the evaluation of nonculture methods that do not involve nucleic acid amplification, and is also a confirmatory test for such techniques. Notwithstanding, clinicians must remember the basic tenet of laboratory tests, that is, good specimen collection and handling, for any laboratory test to yield accurate information to guide their management of patients.

Cells, Cultured↗
PubMed2000

Expression profile viewer (ExProView): a software tool for transcriptome analysis.

A software tool, Expression Profile Viewer (ExProView), for analysis of gene expression profiles derived from expressed sequence tags (ESTs) and SAGE (serial analysis of gene expression) is presented. The software visualizes a complete set of classified transcript data in a two-dimensional array of dots, a "virtual chip," in which each dot represents a known gene as characterized in the transcript databases Expressed Gene Anatomy Database or UniGene. The virtual chip display can be changed between representations of different conceptual systems for gene/protein classification and grouping. Four alternative projections are currently available: (i) cellular role, (ii) subcellular compartment, (iii) chromosome localization, and (iv) total UniGene display. However, the chip can be adapted to any other desired layout. By selecting dots, further information about the represented genes is obtained from the local database and WWW links. The software thus provides a visualization of global mRNA expression at the descriptive level and guides in the exploration of patterns of functional expression, while maintaining direct access to detailed information on each individual gene. To evaluate the software, public EST and SAGE gene expression data obtained from the Cancer Genome Anatomy Project at the National Center for Biotechnology Information were analyzed and visualized. A demonstration of the software is available at http://www.biochem.kth. se/exproview/.

Data Display↗
PubMed2002

Preoperative blood reverse transcriptase-PCR assays for prostate-specific antigen and human glandular kallikrein for prediction of prostate cancer progression after radical prostatectomy.

The association of the results reverse transcription-PCR (RT-PCR) assays for prostate-specific antigen (PSA) with prostate cancer stage and progression remains controversial. We have previously shown that RT-PCR amplification of a native human glandular kallikrein fragment (hK2-L), but not a splice-variant hK2 transcript (hK2-U), performed preoperatively on peripheral blood before radical prostatectomy is an independent predictor of metastases to lymph nodes [K. M. Slawin, et al., Cancer Res., 60: 7142-7148, 2000]. We evaluated the ability of preoperative peripheral blood RT-PCR for PSA and hK2 to predict prostate cancer progression after surgery. The study group consisted of 224 consecutive patients who underwent radical prostatectomy for clinically localized disease (median follow-up: 52.9 months). Also evaluated were 14 healthy men and 8 patients with hormone-naïve, prostate cancer metastatic to bone. RT-PCR for PSA, hK2-L, and hK2-U were positive in 24, 25, and 26%, respectively, of prostatectomy patients; 88, 71, and 86%, respectively, of patients with metastases to bones; 7, 14, and 36%, respectively, of healthy men. Preoperative RT-PCR-hK2-L positivity was associated with higher final Gleason sum (P = 0.028), with an increased risk of metastases to lymph nodes (P = 0.019) and with overall and aggressive disease progression (P = 0.0231 and P = 0.028, respectively). Preoperative blood RT-PCR-PSA and RT-PCR-hK2-U were not associated with characteristics or outcomes of prostate cancer. Preoperative blood RT-PCR for PSA and hK2-U are neither therapy-guiding staging tools nor prognostic indicators in patients with clinically localized prostate cancer. In contrast, preoperative RT-PCR-hK2-L status is associated with established markers of aggressive prostate cancer.

Adult↗
PubMed2001

Virologic and regimen termination surrogate end points in AIDS clinical trials.

Suppression of plasma human immunodeficiency virus (HIV) RNA levels has been widely accepted as an appropriate surrogate end point for HIV disease progression, and it is currently used as the primary end point to determine efficacy in many antiretroviral trials. However, this end point does not always measure other important effects of treatment, such as inducement of multidrug resistance, which depletes future therapy options, and toxic effects. An alternative that directly factors in these treatment costs is a composite regimen termination end point, defined as a protocol-determined change in regimen due to either virologic failure or treatment-related toxic effects. Pros and cons for using purely virologic vs various composite primary end points are discussed. Conclusions include (1) a trial's clinical objective guides the choice of primary end point, (2) a purely virologic end point is often preferable, (3) it may be important to analyze both end point types in interpreting study results, and (4) long-term clinical outcome studies are needed for identifying the most predictive surrogate end points.

Acquired Immunodeficiency Syndrome↗
PubMed2004

The Drosophila ARF6-GEF Schizo controls commissure formation by regulating Slit.

The CNS of bilateral symmetric organisms is characterized by intensive contralateral axonal connections. Genetic screens in Drosophila have identified only a few genes required for guiding commissural growth cones toward and across the midline. Two evolutionarily conserved signaling molecules, Netrin and Slit, are expressed in the CNS midline cells. Netrin acts primarily as an attractive signaling cue, whereas Slit mediates repulsive functions. Here, we describe a detailed analysis of the Drosophila gene schizo, which is required for commissure formation. schizo leads to a commissural phenotype reminiscent of netrin mutant embryos. Double-mutant analyses indicate that Netrin and Schizo act independently. The schizo mutant phenotype can be suppressed by either expressing netrin in the CNS midline cells or by a reduction of the slit gene dose, indicating that the balance of attractive and repulsive signaling is impaired in schizo mutants. Overexpression of the schizo RNA in the CNS midline using the GAL4/UAS system leads to a slit phenocopy, suggesting that schizo primarily antagonizes Slit signaling. This is further supported by cell type-specific rescue experiments. The schizo gene generates at least two proteins containing a conserved Sec7 and a pleckstrin homology domain (PH) characteristic for guanine nucleotide exchange factors (GEF) acting on ARF GTPases, which are known to regulate endocytosis. In support of the notion that schizo regulates Slit expression via endocytosis, we found that block of endocytosis leads to a schizo-like phenotype. We thus propose that the balance of the two signaling cues Netrin and Slit can be regulated, controlling membrane dynamics.

ADP-Ribosylation Factor 6↗
PubMed2004

Fatal West Nile virus encephalitis in a renal transplant recipient.

West Nile virus (WNV), a mosquito-transmitted single-stranded RNA flavivirus, causes human disease of variable severity. We report clinical and pathologic findings of fatal encephalitis from the transmission of WNV from an organ donor to a kidney transplant recipient. The patient developed a febrile illness 18 days after transplantation, which progressed to encephalitis. Postmortem examination demonstrated extensive viral encephalopathic changes. Immunohistochemical studies highlighted WNV antigens within neurons, especially in the cerebellum and brainstem. Flavivirus virions were detected ultrastructurally within the cerebellum, and WNV was isolated from the brain and the brainstem. Thus, this case demonstrates the first death in the first solid organ transplant-associated transmission of WNV. Immunosuppression of the transplant recipient might have been responsible for the fulminant viral effects. The pathologic diagnosis helped guide subsequent epidemiologic and laboratory studies.

Adult↗
PubMed2002

Epstein-Barr virus-associated smooth muscle tumour: a distinctive mesenchymal tumour of immunocompromised individuals.

immunosuppressed patients are predisposed to the development of smooth muscle tumours which show near consistent association with Epstein-Barr virus (EBV). This report describes a 37-year-old patient with acquired immunodeficiency syndrome who initially presented with two masses in the liver. Image-guided core biopsy revealed a spindle cell tumour with histological and immunological features of smooth muscle neoplasm which was shown by in situ hybridisation for EBV early RNAs to be EBV-associated. The literature on this uncommon entity is critically reviewed and the differential diagnosis is also discussed.

Acquired Immunodeficiency Syndrome↗
PubMed1998

Detection and analysis of cytokine mRNA in tissues and cell lines.

Many normal and pathological processes have been shown to be influenced by the quantitative and qualitative nature of cytokine production. Cytokines have also been shown to be central in modulating host responses to invasion by pathogens. Therefore detection and analysis of cytokine production are critical in the understanding of host responses to a variety of immunological insults. The detection of cytokines by bioassay or immunoassay provides data from in vitro experiments and can be used to determine the circulating plasma levels of particular cytokines. However, many investigations require the detection of cytokines in small numbers of cells or directly from tissue samples. It is in this particular area that detection and analysis of cytokine gene expression by use of the sensitive and specific reverse transcription-polymerase chain reaction (RT-PCR) has application. The purpose of these protocols is to provide a practical guide to the detection of cytokine gene expression using RT-PCR techniques. Procedures for analysing mRNA expression in cell culture, cell suspensions and tissue samples will be described and pitfalls and caveats concerning the techniques will be discussed. These protocols should provide a starting point for the development of RT-PCR technology in laboratories with only limited molecular experience.

Cytokines↗
PubMed1996

Effects of SKF 108922, an HIV-1 protease inhibitor, on retrovirus replication in mice.

Rationally designed synthetic inhibitors of retroviral proteases inhibit the processing of viral polypeptides in cultures of human T lymphocytes infected with human immunodeficiency virus type 1 (HIV-1) and therefore suppress the infectivity of HIV-1 in vitro. We have previously reported the antiviral activity in vitro of HIV-1 protease inhibitors against the C-type retrovirus Rauscher murine leukemia virus (RMuLV) and the lentivirus simian immunodeficiency virus (SIV). The same compounds which blocked the infectivity of HIV-1 also inhibited the infectivity of RMuLV and SIV in vitro. This report extends these findings by testing the antiviral activity of HIV-1 protease inhibitors in vivo in the RMuLV model. RMuLV-infected mice were treated twice a day (bid) with either an active (SKF 108922) or inactive (SKF 109273) compound for fourteen days by the intraperitoneal (i.p.) route. Compared with excipient control, SKF 108922, formulated with hydroxypropyl-beta-cyclodextrin (HPB), reduced virus-induced splenomegaly, viremia, and serum reverse transcriptase (RT) levels, while SKF 109273 was inactive. The HPB vehicle by itself enhanced replication of RMuLV. The effects of changing the formulation and the route of administration were examined. SKF 108922, formulated in HPB, had similar antiviral activity when administered by the i.p. or subcutaneous (SC) routes. However, SKF 108922 administered as a colloidal suspension in cholesterol sulfate (CS) had no detectable antiviral effect. Measurements of the circulating levels of the protease inhibitor in plasma explained this result. Plasma concentrations of SKF 108922 exceeded 1000 nM within 10 min after SC administration of the compound solubilized in HPB, but SKF 108922 was not detected in plasma after SC administration of the same dose formulated with CS. Information on optimal conditions for administering these agents should prove useful in guiding their clinical application Therefore, RMuLV should provide a good model for the preclinical evaluation and development of this class of agents for the treatment of HIV.

2-Hydroxypropyl-beta-cyclodextrin↗
PubMed2005

Crystallization and preliminary X-ray characterization of the nitrile reductase QueF: a queuosine-biosynthesis enzyme.

QueF (MW = 19.4 kDa) is a recently characterized nitrile oxidoreductase which catalyzes the NADPH-dependent reduction of 7-cyano-7-deazaguanine (preQ0) to 7-aminomethyl-7-deazaguanine, a late step in the biosynthesis of the modified tRNA nucleoside queuosine. Initial crystals of homododecameric Bacillus subtilis QueF diffracted poorly to 8.0 A. A three-dimensional model based on homology with the tunnel-fold enzyme GTP cyclohydrolase I suggested catalysis at intersubunit interfaces and a potential role for substrate binding in quaternary structure stabilization. Guided by this insight, a second crystal form was grown that was strictly dependent on the presence of preQ0. This crystal form diffracted to 2.25 A resolution.

Bacillus subtilis↗
PubMed2004

Systems biology of the 2-cell mouse embryo.

The transcriptome of the 2-cell mouse embryo was analyzed to provide insight into the molecular networks at play during nuclear reprogramming and embryonic genome activation. Analysis of ESTs from a 2-cell cDNA library identified nearly 4,000 genes, over half of which have not been previously studied. Transcripts of mobile elements, especially those of LTR retrotransposons, are abundantly represented in 2-cell embryos, suggesting their possible role in introducing genomic variation, and epigenetic restructuring of the embryonic genome. Analysis of Gene Ontology of the 2-cell-stage expressed genes outlines the major biological processes that guide the oocyte-to-embryo transition. These results provide a foundation for understanding molecular control at the onset of mammalian development.

Animals↗
PubMed1998

Evidence for the participation of nerve growth factor and its low-affinity receptor (p75NTR) in the regulation of the myogenic program.

We have studied expression and function of neurotrophins and their receptors during myogenic differentiation of C2C12 cells, a clonal cell line derived from mouse muscle that is capable of in vitro differentiation. The genes coding for nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and their common low-affinity receptor p75(neurotrophin receptor) (p75NTR) were shown to be expressed in C2C12 myoblasts and downregulated during myogenic differentiation and fusion into myotubes. Cocultures with dorsal root ganglia from day 8 chick embryos revealed neurite-promoting activities of C2C12 cells that ceased with myogenic differentiation. These data suggest a temporal and developmental window for the effect of myogenic cell-derived neurotrophins on neuronal as well as on myogenic cell populations. NGF was shown to increase DNA synthesis and cell growth of C2C12 myoblasts and to enhance myogenic differentiation in this cell line. We present evidence that NGF-mediated processes take place at stages preceding myogenic differentiation. Enhanced muscle differentiation was also seen in p75NTR-overexpressing C2C12 myoblasts which maintained high levels of receptors but ceased to produce NGF during differentiation. In contrast, when exogenous NGF was present at the onset of myogenic differentiation of receptor-overexpressing cells, muscle cell development was strongly repressed. This indicates that downregulation of p75NTR is necessary for guiding myogenic cells towards terminal differentiation. Since none of the trk high-affinity neurotrophin receptors could be demonstrated in C2C12 cells, we conclude that NGF mediates its nonneurotrophic effect via its low-affinity receptor in an autocrine fashion.

Animals↗
PubMed1990

All subgenomic mRNAs of equine arteritis virus contain a common leader sequence.

During the replication of equine arteritis virus (EAV) six subgenomic mRNAs are synthesized. We present evidence that the viral mRNAs form a 3'-coterminal nested set and contain a common leader sequence of 208 nucleotides which is encoded by the 5'-end of the genome. The leader is joined to the bodies of mRNA 5 and 6 at positions defined by the sequence 5' UCAAC 3'. The part of the leader sequence flanking the UCAAC motif is very similar to the 5'-splice site of the Tetrahymena pre-rRNA. A possible internal guide sequence has been identified 43 nucleotides downstream of the leader sequence on the genome. Hybridization analysis shows that all EAV intracellular RNAs contain the leader sequence. These data imply that the viral subgenomic mRNAs are composed of leader and body sequences which are non-contiguous on the genome.

Animals↗
PubMed2002

Molecular recognition by the Candida albicans group I intron: tertiary interactions with an imino G.A pair facilitate binding of the 5' exon and lower the KM for guanosine.

A G.A pair at position -5 in the P1 helix of the Candida albicans ribozyme contributes to tertiary binding of the 5' exon substrate [Disney, M. D., Haidaris, C. G., and Turner, D. H. (2001) Biochemistry 40, 6507-6519]. Here, the G in the G.A pair is replaced with inosine (I) in both semisynthetic ribozymes and oligonucleotide mimics of the internal guide sequence. Comparisons of oligonucleotide binding affinity for these and other sequences indicate that the G.A pair is in an imino conformation where the exocyclic amine of G contributes approximately 1.4 kcal/mol to tertiary interactions that help dock the ribozyme's P1 helix. Furthermore, replacement of the G.A pair with a G-C pair produces less favorable interactions with the 2'-hydroxyl group at the -3 position and a less favorable K(M) for pG in a ribozyme-catalyzed transesterification reaction. These results are also consistent with the G.A pair promoting docking of the P1 helix into the catalytic core. Evidently, tertiary interactions with the exocyclic amino group of a G in a single G.A pair can increase the equilibrium constant for tertiary folding of RNA by roughly 10-fold at 37 degrees C. Results with a G.U or G.G pair replacing the G.A pair at the -5 position suggest similar tertiary interactions with these pairs.

Adenine↗
PubMed2002

Manipulating gene expressions by electroporation in the developing brain of mammalian embryos.

One of the goals of developmental neuroscience in the post-genomic era is to clarify functions of a huge number of anonymous genes of which only DNA sequences are identified. More convenient methods for genetic manipulation in vertebrates, especially mammals, could help us to identify functions of the novel genes. Here we introduce a novel gene transfer technology using electroporation (EP), which is a simple and powerful strategy for genetic analysis. We have applied this method to cultured mammalian embryos in order to understand the function of specific genes in the developing brain. We have also performed EP in developing fetuses in utero guided by ultrasound image. The combination of these techniques in addition to analysis of genetic mutants will clarify functions of individual genes, gene interactions, and the molecular mechanisms underlying the brain development.

Animals↗
PubMed2000

The tmRNA website.

The tmRNA Website collects all available tmRNA sequences into a single public resource, along with alignments and a guide to searching for new sequences. Over the last year, several sequences have been updated or newly found by monitoring ongoing genome sequencing projects; tmRNA sequence data from 70 species are now available. New features include: color-coding of sequences to mark suggested base-paired regions, a list of the literature concerning tmRNA, careful crediting of tmRNA sequence identifications, and a split browser window. Updates are very frequent. The tmRNA Website has a new URL: http:www.indiana.edu/tmrna

Internet↗