Photoimmunotherapy and ovarian cancer: an improbable fiction or a palpable hit?
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Animals↗
Biomedical subjects
S Hahn
Publications and source records attributed to S Hahn.
Recruitment of the SWI/SNF chromatin remodeling complex by transcriptional activators.
SWI/SNF is a chromatin remodeling complex that facilitates expression of a number of yeast genes. Here we demonstrate that SWI/SNF can be recruited from yeast nuclear extracts by a transcriptional activator. Recruitment is dependent on an activation domain but not on promoter sequences, TBP, or RNA polymerase II holoenzyme. We also show that acidic activation domains can target SWI/SNF remodeling activity. These results demonstrate that SWI/SNF activity can be targeted by gene-specific activators and that this recruitment can occur independently of Pol II holoenzyme.
Chromatin↗
Ageing, fitness and neurocognitive function.
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Aging↗
Intermediates in formation and activity of the RNA polymerase II preinitiation complex: holoenzyme recruitment and a postrecruitment role for the TATA box and TFIIB.
Assembly and activity of yeast RNA polymerase II (Pol II) preinitiation complexes (PIC) was investigated with an immobilized promoter assay and extracts made from wild-type cells and from cells containing conditional mutations in components of the Pol II machinery. We describe the following findings: (1) In one step, TFIID and TFIIA assemble at the promoter independently of holoenzyme. In another step, holoenzyme is recruited to the promoter. Mutations in the CTD of Pol II, Srb2, Srb4, and Srb5, and two mutations in TFIIB disrupt recruitment of all holoenzyme components tested without affecting TFIID and TFIIA recruitment. These results indicate that the stepwise assembly pathway is blocked after TFIID/TFIIA binding. (2) Both the Gal4-AH and Gal4-VP16 activators stimulate formation of active PICs by increasing the extent of PIC formation. The Gal4-AH activator stimulated PIC formation by enhancing the binding of TFIID and TFIIA, whereas Gal4-VP16 could enhance the recruitment of TFIID, TFIIA, and holoenzyme. (3) Extracts deficient in TFIIA activity showed reduced assembly of all PIC components. These and other results suggest that TFIIA acts at an early step by enhancing the stable recruitment of TFIID. (4) An extract containing the TFIIB mutant E62G, had no defect in PIC formation, but had a severe defect in transcription. Similarly, mutation of the TATA box reduced PIC formation only two- to fourfold, but severely compromised transcription. These results demonstate an involvement of TFIIB and the TATA box in one or more steps after recruitment of factors to the promoter.
Fungal Proteins↗
Vitamin E suppresses oxidative stress and glomerulosclerosis in rat remnant kidney.
Previous studies have shown that reduction of renal mass in the rat remnant kidney model induces overproduction of transforming growth factor beta1 (TGFbeta1). We investigated whether an antioxidant, vitamin E, administered before the renal mass reduction, could prevent oxidative stress, reduce the overproduction of TGFbeta1, and mitigate against the subsequent glomerulosclerosis. Our results revealed that the oxidative stress, as measured by the change in plasma malondialdehyde, is significantly reduced by prior vitamin E dietary supplementation. Finally, our data show that dietary vitamin E supplementation ameliorates the rise in TGFbeta1 secondary to renal mass reduction and inhibits the glomerular sclerosis of the remnant kidney over the time course of this experiment.
Animals↗
Task coordination and aging: explorations of executive control processes in the task switching paradigm.
A number of models of cognitive aging suggest that older adults exhibit disproportionate performance decrements on tasks which require executive control processes. In a series of three studies we examined age-related differences in executive control processes and more specifically in the executive control processes which underlie performance in the task switching paradigm. Young and old adults were presented with rows of digits and were required to indicate whether the number of digits (element number task) or the value of the digits (digit value task) were greater than or less than five. Switch costs were assessed by subtracting the reaction times obtained on non-switch trials from trials following a task switch. Several theoretically interesting results were obtained. First, large age-related differences in switch costs were found early in practice. Second, and most surprising, after relatively modest amounts of practice old and young adults switch costs were equivalent. Older adults showed large practice effects on switch trials. Third, age-equivalent switch costs were maintained across a two month retention period. Finally, the main constraint on whether age equivalence was observed in task switching performance was memory load. Older adults were unable to capitalize on practice under high memory loads. These data are discussed in terms of their implications for both general and process specific cognitive aging models.
Adolescent↗
Fetal cells in maternal blood. An update from Basel.
OBJECTIVE: The efficiency of two protocols for the enrichment of fetal cells from the blood of pregnant women was compared: a triple density gradient followed by twin magnetic separations (method A) versus a single density gradient and single magnetic separation (method B). STUDY DESIGN: Blood samples were obtained from women prior to undergoing an invasive procedure. The processed samples, 87 by method A and 332 by method B, were examined for the presence of male cells by fluorescence in situ hybridisation. RESULTS: The simpler protocol was found to be superior. The most critical component, however, is the ability of the reader to correctly evaluate the sample, where we observed large variations, with reader B attaining a sensitivity of 82.61% with a corresponding specificity of 86.96%. CONCLUSION: A simpler enrichment protocol can be used from smaller blood samples to attain detection efficiencies which are similar to or superior to current noninvasive methods.
Cell Separation↗
Attentional capture and aging: implications for visual search performance and oculomotor control.
Two studies examined potential age-related differences in attentional capture. Subjects were instructed to move their eyes as quickly as possible to a color singleton target and to identify a small letter located inside it. On half the trials, a new stimulus (i.e., a sudden onset) appeared simultaneously with the presentation of the color singleton target. The onset was always a task-irrelevant distractor. Response times were lengthened, for both young and old adults, whenever an onset distractor appeared, despite the fact that subjects reported being unaware of the appearance of the abrupt onset. Eye scan strategies were also disrupted by the appearance of the onset distractors. On about 40% of the trials on which an onset appeared, subjects made an eye movement to the task-irrelevant onset before moving their eyes to the target. Fixations close to the onset were brief, suggesting parallel programming of a reflexive eye movement to the onset and goal-directed eye movement to the target. Results are discussed in terms of age-related sparing of the attentional and oculomotor processes that underlie attentional capture.
Adult↗
Novel molecular biological approaches for the diagnosis of preeclampsia.
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DNA↗
A comparison of different density gradients and antibodies for enrichment of fetal erythroblasts by MACS.
The enrichment of fetal cells, in particular fetal erythroblasts from the blood of pregnant women offers a promising non-invasive alternative for prenatal diagnosis. The purpose of this study was to compare the retrieval of erythroblasts by different density gradients and different antibodies against erythroid surface antigens, in both a model test system and in blood samples of pregnant women. We enriched erythroblasts from artificial mixtures of cord and adult blood (1:50) and from 16 ml of peripheral blood from pregnant women at a mean gestational age of 14+2 weeks. The yield of erythroblasts was calculated and compared using Wilcoxon's matched-pairs signed-rank test. In the artificial mixture most erythroblasts were retrieved using the heaviest density gradient (specific density of 1119) followed by antibody-labelled magnetic cell sorting (MACS). With anti-GPA the yield of erythroblasts from the artificial mixture was highest (9362.5 erythroblasts) compared with anti-CD36 (5164.3), anti-i (3455.2), anti-CD71 (3055.8) and HAE9 (2364.2). The difference between anti-GPA and anti-CD71, HAE9 and anti-i was significant (p=0.0277). The enrichment of erythroblasts from peripheral blood of pregnant women showed similar results. The yield of erythroblasts using anti-GPA was the highest. These results enable us to simplify our enrichment protocols to a single density gradient of 1119 specific density followed by MACS, with anti-GPA.
Antibodies, Monoclonal↗
The role of TAFs in RNA polymerase II transcription.
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Acetyltransferases↗
[Prenatal diagnosis with fetal cells in maternal blood: report of experiences in Basal].
Currently prenatal diagnosis relies on invasive procedures such as chorion villus sampling (CVS) or amniocentesis (AC). Many parents are reluctant to expose themselves and their child to the small, but significant risk posed by these procedures to mother and child. There is, hence, a great need for a risk-free non-invasive alternative. To achieve this goal most research has been focussed on enriching fetal cells from the blood of pregnant women. The erythroblast has emerged as the target cell of choice, since it is abundant in the early fetus, rare in normal adult blood, and since it has a very short half life, there is no risk of obtaining cells from previous pregnancies. Most enrichment protocols rely either on magnetic- or fluorescent activated cell sorting (MACS and FACS) using fetal specific antibodies. These enriched cells can be examined by FISH (fluorescence in-situ hybridisation) for the presence of the most common fetal chromosomal aneuploidies (13, 18, 21, X and Y) or by polymerase chain reaction (PCR) on singly manipulated cells for genetic disorders. The efficacy in detecting fetal aneuploidies is currently being evaluated in a phase II clinical trial under the auspices of the NIH-NICHD, the so-called NIFTY Trial, in which our group is a participant. By modifying our enrichment protocols we have recently been able to obtain detection sensitivities of almost 80%, thereby renewing our optimism that this methodology provides a solid basis for an effective non-invasive prenatal diagnostic test.
Adult↗
Highly accurate analysis of heterozygous loci bysingle cell PCR.
Single cell PCR is a powerful method for determining the genetic properties of individual cells. In the instance of heterozygous loci, however, preferential amplification of one allele can lead to allele drop out (ADO) of the other allele. Fortunately ADO does not occur in all amplifications, and is usually random when it does occur, with both alleles being equally susceptible to drop out. Therefore pooling of results from multiple independently amplified cells should greatly improve the analysis of diallelic loci, and the misdiagnosis rate of diallelic loci should decrease exponentially with the number of cells analysed. We have shown that this is true and that multiplex PCR allows for the simultaneous identification of a cell in a mixture of cells using microsatellite loci known to be informative, and accurate genotyping at other loci. This approach has practical applications to non-invasive prenatal diagnosis where small numbers of fetal cells in the presence of maternal cells must be both identified and genotyped at loci involved in genetic disease.
Alleles↗
Laparoscopic-assisted application of interstitial brachytherapy for locally advanced cervical carcinoma: results of a pilot study.
PURPOSE: To assess the efficacy of diagnostic laparoscopy at the time of interstitial brachytherapy in patients with FIGO Stages IIB-IVA cervical carcinoma, who were not candidates for conventional brachytherapy after completion of whole pelvic radiation. MATERIALS AND METHODS: Six patients with FIGO Stages IIB-IVA cervical carcinoma completed whole pelvic radiation (WPR) and were assessed for the placement of conventional intracavitary brachytherapy. Three patients (Stage IIB) received 50.40 Gy WPR and three (Stage IIIA-VA) received 61.20 Gy WPR. Because distorted vaginal anatomy precluded the placement of standard intracavitary brachytherapy equipment, interstitial therapy was selected. To minimize the risk of source misplacement, needles were placed with laparoscopic guidance. 192Iridium was utilized as a source of radiation. RESULTS: A total of 98 needles were placed under direct laparoscopic guidance. The median interstitial brachytherapy tumor dose was 20.00 Gy (range 19.00-41.20 Gy). Eleven perforations in the pelvic peritoneum and/or bladder were identified intraoperatively in 5 of the 6 patients, leading to immediate repositioning of needles. No acute or short-term morbidity related to the procedure was appreciated. CONCLUSION: Interstitial brachytherapy offers an alternative for intracavitary radiation therapy for selected patients, in whom adequate placement of intracavitary afterloading equipment is precluded by abnormal pelvic geometry. Historically, the placement of interstitial sources has been a procedure that was performed without visualization of the pelvic cavity. A major concern is direct injury to pelvic viscera and intestine. The use of diagnostic laparoscopy at the time of placement of interstitial brachytherapy may avert potential complications from misapplication of interstitial sources. Longer follow-up is required to substantiate these results.
Adult↗
The tumor suppressor gene Smad4/Dpc4 is required for gastrulation and later for anterior development of the mouse embryo.
Mutations in the SMAD4/DPC4 tumor suppressor gene, a key signal transducer in most TGFbeta-related pathways, are involved in 50% of pancreatic cancers. Homozygous Smad4 mutant mice die before day 7.5 of embryogenesis. Mutant embryos have reduced size, fail to gastrulate or express a mesodermal marker, and show abnormal visceral endoderm development. Growth retardation of the Smad4-deficient embryos results from reduced cell proliferation rather than increased apoptosis. Aggregation of mutant Smad4 ES cells with wild-type tetraploid morulae rescues the gastrulation defect. These results indicate that Smad4 is initially required for the differentiation of the visceral endoderm and that the gastrulation defect in the epiblast is secondary and non-cell autonomous. Rescued embryos show severe anterior truncations, indicating a second important role for Smad4 in anterior patterning during embryogenesis.
Alleles↗
Biliary glycoprotein (CD66a), a cell adhesion molecule of the immunoglobulin superfamily, on human lymphocytes: structure, expression and involvement in T cell activation.
The biliary glycoproteins (BGP or CD66a), a group of different splice variants of a single gene, are members of the carcinoembryonic antigen family and the immunoglobulin superfamily. Recently, we detected CD66a on IL-2 activated lymphocytes. In this study we characterized the structure and the expression pattern of BGP on human lymphocytes and investigated its role in T cell activation. Lymphocytes express 2 of the 13 known splice variants, i.e. BGPa and BGPb, which are glycosylated in a lymphocyte-specific manner. Both BGPa and BGPb have the long cytoplasmic tail, which contains two immunoreceptor tyrosine-based inhibitory motif (ITIM)-like motifs, but differ in their extracellular region containing 4 and 3 immunoglobulin-like domains, respectively. On PBL BGP is expressed in small amounts only on B cells and Th cells. Stimulation with IL-2 leads to a strong up-regulation of BGP by these cells, and induces de novo BGP expression on gammabeta T cells, CD8+ and CD56+ cells, but not on CD16+ lymphocytes. This up-regulation of BGP seems to be part of the physiological process of T cell activation, since stimulation with anti-CD3 mAb is sufficient to induce BGP up-regulation. Based on the presence of the two ITIM-like motifs, one may expect that BGP inhibits T cell activation, but surprisingly, engagement of BGP enhances the proliferation of anti-CD3-stimulated T cells.
Antigens, CD↗
Disturbed feto-maternal cell traffic in preeclampsia.
OBJECTIVE: To investigate whether the transfer of fetal blood cells to the maternal circulation is perturbed in pregnancies affected by preeclampsia. METHODS: Fetal erythroblasts were isolated from eight women with clinically diagnosed preeclampsia (blood pressure values of at least 140/90 mmHg and associated proteinuria) and an equal number of matched corresponding controls. All patients in both groups were pregnant with male singleton fetuses. The presence of fetal cells was evaluated histologically and by fluorescence in situ hybridization for X and Y chromosomes. RESULTS: The number of fetal cells was higher in preeclamptic patients than in controls, with respect to both nucleated red blood cells (median per 200 cells 38 versus 7; P < .001) and the proportion of these cells that were of fetal origin (median per 2000 cells 9 versus 2; P = .001). CONCLUSION: These results suggest that the trafficking of fetal cells into the maternal periphery is disturbed in patients with preeclampsia. Because it is unlikely that such an altered flow of cells is restricted to the erythroblasts examined in this study, these findings also may lead to interesting new concepts regarding the development of preeclampsia and possibly the associated syndrome of hemolysis, elevated liver enzymes, and low platelets.
Erythroblasts↗
Allele drop-out can occur in alleles differing by a single nucleotide and is not alleviated by preamplification or minor template increments.
The ability to analyze the genetic material of single cells by the PCR opens up new prospects for diagnostics. Because only two copies of the genetic template are available for amplification, a problem that frequently arises when examining heterozygous loci in single cells is allele drop-out (ADO). ADO results from the preferential amplification of one of a pair of heterozygous alleles, in which the other allele is totally under-represented. In examining single cells from carriers heterozygous for beta-thalassemia mutations, we have found ADO to occur in alleles differing by a single nucleotide, where either the normal or the mutant genotype was absent. We have found that ADO is not overcome by either increasing the amount of DNA template to 20 pg or by primer extension preamplification (PEP), but rather that the best diagnostic accuracy is obtained by examining multiple single cells and basing a diagnosis on the combined results of such an examination.
Alleles↗