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PubMed2002

Complexities in ETS-domain transcription factor function and regulation: lessons from the TCF (ternary complex factor) subfamily. The Colworth Medal Lecture.

The ETS-domain transcription factor family can be divided into a series of subfamilies. Elk-1 represents the founding member of the ternary complex factor (TCF) subfamily. By focusing on the TCF subfamily, we can demonstrate the complexities that exist in the function and regulation of ETS-domain transcription factors. This article focuses on Elk-1 in detail and summarizes the functions of other TCFs. The key themes covered include the domain structure of the TCFs, the mechanisms of complex formation with serum response factor, regulation of TCFs by mitogen-activated protein kinase cascades, and transcriptional regulatory properties of the TCFs. Finally, the emerging role of the TCFs in vivo is discussed. A picture is developing indicating that, while these proteins exhibit significant sequence and functional conservation, key differences in their structure and regulation are being identified which may relate to unique functions of these proteins in vivo.

Amino Acid Sequence↗
PubMed1992

Update on tumor vaccines.

Vaccination against tumor has always been an attractive idea for the treatment of patients bearing tumor. By harnessing the host's own immune response the attack on tumor cells would act on a continuing basis, with emerging tumor cells stimulating their own destruction. However, the approach has been hampered by our poor understanding of the nature of tumor antigens and of the pathways by which immune cells might operate against tumor growth. Recent developments in molecular biology and immunology are remedying this deficiency and bringing vaccination to the forefront of new approaches to treatment of a range of tumors. Results obtained in B-cell tumors, where the idiotypic immunoglobulin at the cell surface provides a well-defined tumor antigen, are already indicating exciting possibilities as well as delineating problems. There is considerable clinical evidence that patients have some intrinsic ability to control tumor growth and that certain tumors remain dormant for long periods. Attempts to understand and perhaps stimulate the mechanisms involved are being made through the use of biological modifiers and by manipulating potential effector cells in vitro. Ideally this approach, which may include non-specific and specific elements, could be combined with specific vaccination in order to combat the apparent ability of many tumor cells to evade host defences.

Antibodies, Neoplasm↗
PubMed2000

New diagnostic methods for esophageal carcinoma.

The increasingly severe problem of esophageal carcinoma on world public health merits the application of new endoscopic methods to assist in early detection and screening. Older methods, such as tissue staining, combined with magnification endoscopy, have shown promising results, while newer techniques capitalize on measurements that discriminate benign from malignant cells based on a wide array of different attributes, ranging from the molecular to the macroscopic level. Instrumentation based on laser-induced fluorescence spectroscopy, ratio fluorescence imaging, elastic scattering spectroscopy, Raman spectroscopy, and optical coherence tomography is presently being tested and compared with standard endoscopic techniques. Using pathologic interpretation of pinch biopsies as the "gold standard," these techniques have shown the ability to identify dysplastic or malignant regions of tissue that would not be visible to the unassisted endoscopist and offer increased sensitivity for detection compared to rigorous random biopsy protocols. The rapid speed of the instruments allows the provision of information to the endoscopist almost instantaneously, potentially allowing therapeutic decisions to be conducted within the confines of the same endoscopic procedure, thereby achieving gains in efficiency and reductions in overall cost. Large, multicenter trials will be necessary to determine the sensitivity and specificity of individual and combined techniques, as well as their ability to favorably influence the early detection, management, and overall outcome of this disease.

Esophageal Neoplasms↗
PubMed1979

In vitro killing of Pasteurella multocida: the effect of rabbit granulocyte and specific antibody source.

In vitro granulocyte-killing assays were performed to examine the ability of granulocytes from pasteurella-free or immunized rabbits, the in combination with specific immune serum, to kill Pasteurella multocida. Granulocytes from healthy rabbits and from rabbits with P multocida infections were equally competent. Granulocyte source, serum source, and specific antibody titer had no effect on granulocyte phagocytic activity. Moreover, serum containing specific antibody and complement supported the growth of the bacterium. These data suggest that chronic P multocida infections are not attributable to defective granulocytes or lack of serum antibody production.

Animals↗
PubMed1994

Nerve-dependent induction of AChR epsilon-subunit gene expression in muscle is independent of state of differentiation.

To determine if the expression of the epsilon-subunit of the acetylcholine receptor by the subsynaptic nuclei in skeletal muscle is dependent on the state of differentiation of the muscle, we have compared the spatiotemporal distribution of epsilon-subunit transcripts during synapse formation in fetal and adult muscle. Both during "ontogenic" synaptogenesis in the fetus and during "ectopic" synaptogenesis in the adult animal the motor nerve induced focally the expression of the epsilon-subunit mRNA in subsynaptic nuclei. The temporal expression patterns at both types of developing synapses were similar. The results support the view that in muscle developing in vivo epsilon-subunit gene transcription and its stabilization in subsynaptic nuclei is exclusively controlled by the motor neuron, independently of the developmental state of the muscle nuclei. Thus, both nerve and muscle remain plastic in their respective abilities to induce and express the synapse-specific combination of AChR subunit genes.

Age Factors↗
PubMed2003

Clinical value of FDG-PET in the follow up of post-operative patients with endometrial cancer.

OBJECTIVE: The clinical usefulness of FDG-PET in the follow up of post-operative patients with endometrial cancer was retrospectively evaluated. METHODS: Twenty-one post-operative patients with endometrial cancer received 30 FDG-PET examinations to evaluate recurrence or response to treatment. The findings of FDG-PET were compared with their serum levels of tumor markers, CT and/or MRI findings, and the final outcome. Results of FDG-PET were also correlated with the clinical course of each patient. RESULTS: In detecting recurrent lesions and evaluating treatment responses, FDG-PET, with the help in anatomic information by CT/MRI, showed better diagnostic ability (sensitivity 100.0%, specificity 88.2%, accuracy 93.3%) compared with combined conventional imaging (sensitivity 84.6%, specificity 85.7%, accuracy 85.0%) and tumor markers (sensitivity 100.0%, specificity 70.6%, accuracy 83.3%). FDG-PET had no false-negative results, suggesting the possibility of its use as the first-line examination in a patient's follow-up. FDG-PET could detect unknown lesions in 4 cases, and, as reported for other malignancies, FDG-PET affected the patient management in one-third of the cases. Furthermore, the results of FDG-PET correlated well with the clinical outcome of the patients, with patients with negative PET results tending to show disease-free courses. CONCLUSIONS: These results suggest that, despite the limited number of patients studied, FDG-PET was accurate in detecting recurrence and evaluating therapeutic response, and could afford important information in the management of post-operative patients with endometrial cancer. FDG-PET also appeared to have a possibility to predict the outcome of each patient.

Adult↗
PubMed1985

Hormonal effects on the estrogen receptor system in the epididymis and accessory sex organs of sexually immature rabbits.

The epididymis and male accessory sex organs (vesicular gland, prostate, and bulbourethral gland) of sexually immature rabbits contain a functional estrogen receptor system which is regulated in an organ-specific manner by various hormones. In both intact and castrated animals, acute estrogen challenge causes depletion of estrogen receptor from the cytosolic fraction and its appearance in the nuclear fraction of these tissues. A considerable amount of unoccupied nuclear receptor was detected both before and after estrogen challenge. An estrogen-activated, receptor-processing mechanism is operable in these organs since chronic treatment (daily for 14 days) with estradiol benzoate modified the levels of total estrogen receptor, and altered the relative amounts of occupied to unoccupied nuclear receptor present following estrogen challenge. Chronic treatment with estradiol benzoate, Tamoxifen, and testosterone propionate (alone and in combination) had differential, organ-specific effects on the ability of subsequent estrogen challenge to cause accumulation of nuclear receptor. The vesicular gland was the most responsive to estrogen treatment and the bulbourethral gland the least responsive.

Androgens↗
PubMed2003

Quantitative RT-PCR ELISA to determine the amount and ratio of positive- and negative strand viral RNA synthesis and the effect of guanidine in poliovirus infected cells.

Quantitative reverse transcription-polymerase chain reaction enzyme linked immunosorbent assay (RT-PCR ELISA) is the method of choice to study positive- and negative strand viral RNA synthesis during poliovirus replication. In comparison with other methods used for this purpose, it fulfils all necessary requirements to accurately determine RNA of different polarity. It combines high specificity, high sensitivity, safety, speed, and the ability to perform quantitative analysis. The enterovirus specific RT-PCR ELISA method described in this work, was used to determine quantitatively the amount of de novo poliovirus positive- and negative strand RNA synthesis at different time-points in the viral replication cycle, both in presence and absence of the viral RNA synthesis inhibitor guanidine hydrochloride.

Biotin↗
PubMed1998

Translocation of calmodulin to the nucleus supports CREB phosphorylation in hippocampal neurons.

Activation of the transcription factor CREB is thought to be important in the formation of long-term memory in several animal species. The phosphorylation of a serine residue at position 133 of CREB is critical for activation of CREB. This phosphorylation is rapid when driven by brief synaptic activity in hippocampal neurons. It is initiated by a highly local, rise in calcium ion concentrations near the cell membrane, but culminates in the activation of a specific calmodulin-dependent kinase known as CaMK IV, which is constitutively present in the neuronal nucleus. It is unclear how the signal is conveyed from the synapse to the nucleus. We show here that brief bursts of activity cause a swift (approximately 1 min) translocation of calmodulin from the cytoplasm to the nucleus, and that this translocation is important for the rapid phosphorylation of CREB. Certain Ca2+ entry systems (L-type Ca2+ channels and NMDA receptors) are able to cause mobilization of calmodulin, whereas others (N- and P/Q-type Ca2+ channels) are not. This translocation of calmodulin provides a form of cellular communication that combines the specificity of local Ca2+ signalling with the ability to produce action at a distance.

Biological Transport↗
PubMed1968

Automated procedure for measuring antigenicity of extracted and intact influenza virus.

An automated serum-blocking (S-B) technique was developed in an attempt to find an in vitro test for the determination of the antigenicity of extracted influenza vaccines. The S-B test depends on the ability of an antigen to combine with (or block) specific (in this case, hemagglutination-inhibiting) antibodies. After mixing the test virus with a constant amount of specific antiserum and hemagglutinating virus in an Auto Analyzer, chicken erythrocytes were pumped into the system and the mixture was incubated by passing through coils. The hemagglutinated cells were removed and the residual cells were lysed. The optical density was read and recorded automatically. The S-B test was much more reproducible than the chicken cell agglutination (CCA) test. There was good correlation between the S-B and CCA titers of intact influenza virus, but not of ether-extracted influenza virus. The CCA titer of influenza strains of type A was reduced significantly during ether-extraction. The S-B titers indicated that there was no significant loss in specific antigenicity when influenza strains of types A and B were extracted with ether and Tween-80 according to the described procedure. The S-B test seemed to be a true measurement of the total antigens present in influenza vaccines.

Animals↗
PubMed2006

Cj1496c encodes a Campylobacter jejuni glycoprotein that influences invasion of human epithelial cells and colonization of the chick gastrointestinal tract.

Campylobacter jejuni has an N-linked protein glycosylation pathway that is required for efficient cell invasion and chick gastrointestinal colonization by the microbe. In this study, we constructed insertion mutants of 22 putative glycoprotein genes and examined the ability of each to invade the human intestinal epithelial cell line INT-407. Among the mutants tested, one carrying an insertion in Cj1496c was defective for invasion into INT-407 cells; this defect was also observed in an in-frame deletion mutant of Cj1496c (delta Cj1496c). The delta Cj1496c mutant C. jejuni also showed a reduced ability to colonize chick ceca. Site-specific mutagenesis combined with Western blot analysis suggested that the Cj1496c protein is glycosylated at N73 and N169. However, the delta Cj1496c mutant expressing a nonglycosylated form of Cj1496c exhibited levels of invasion and colonization equivalent to those of the parent strain, suggesting that glycans are not directly involved in the function of Cj1496c.

Animals↗
PubMed2000

Cid1, a fission yeast protein required for S-M checkpoint control when DNA polymerase delta or epsilon is inactivated.

The S-M checkpoint is an intracellular signaling pathway that ensures that mitosis is not initiated in cells undergoing DNA replication. We identified cid1, a novel fission yeast gene, through its ability when overexpressed to confer specific resistance to a combination of hydroxyurea, which inhibits DNA replication, and caffeine, which overrides the S-M checkpoint. Cid1 overexpression also partially suppressed the hydroxyurea sensitivity characteristic of DNA polymerase delta mutants and mutants defective in the "checkpoint Rad" pathway. Cid1 is a member of a family of putative nucleotidyltransferases including budding yeast Trf4 and Trf5, and mutation of amino acid residues predicted to be essential for this activity resulted in loss of Cid1 function in vivo. Two additional Cid1-like proteins play similar but nonredundant checkpoint-signaling roles in fission yeast. Cells lacking Cid1 were found to be viable but specifically sensitive to the combination of hydroxyurea and caffeine and to be S-M checkpoint defective in the absence of Cds1. Genetic data suggest that Cid1 acts in association with Crb2/Rhp9 and through the checkpoint-signaling kinase Chk1 to inhibit unscheduled mitosis specifically when DNA polymerase delta or epsilon is inhibited.

Amino Acid Sequence↗
PubMed1991

Analysis of anti-tumor antibodies in mice and rabbits induced by monoclonal anti-idiotope antibodies.

Eight different mouse monoclonal anti-idiotope antibodies (mAb2) generated against a mouse monoclonal anti-human melanoma proteoglycan Ag (MPG) antibody (mAb1), MEM136, were tested for their ability to induce anti-MPG responses in mice and rabbits. All Ab2 were idiotypically cross-reactive and combining site-specific as demonstrated by competitive cross-inhibition studies and their ability to inhibit the binding of MEM136 to the melanoma cells, Colo38. However, only two Ab2, IM32 and IM06, were able to induce specific anti-TAA-specific (Ab1') responses in rabbits. When IM32 and IM06 were tested in allogeneic stains of mice for the induction of anti-MPG responses, only IM32 produced an Ab1' response. In mice, the Ab3 response induced by IM32 is idiotypically cross-reactive with its Ab1. Furthermore, the IM32-induced murine Ab3 and MEM136 recognized a similar MPG epitope on the melanoma cells because the Ab3 inhibited the binding of MEM136 to melanoma cells. The Ab3 induced by IM32 and IM06 in rabbits also recognized a similar epitope as the Ab1. In rabbits, the Ab3 response induced by IM32 and IM06 were idiotypically cross-reactive with each other. However, additional studies indicated that the majority of Ab3 induced by IM32 were IM32 Id-specific and lacked IM06 idiotopes. Further experimentation indicated that IM32-induced rabbit Ab3 were biologically active as demonstrated by the ability of the Ab3 to inhibit melanoma cell invasion in a Matrigel assay.

Animals↗
PubMed2004

Structural basis of diverse substrate recognition by the enzyme PMM/PGM from P. aeruginosa.

Enzyme-substrate complexes of phosphomannomutase/phosphoglucomutase (PMM/PGM) reveal the structural basis of the enzyme's ability to use four different substrates in catalysis. High-resolution structures with glucose 1-phosphate, glucose 6-phosphate, mannose 1-phosphate, and mannose 6-phosphate show that the position of the phosphate group of each substrate is held constant by a conserved network of hydrogen bonds. This produces two distinct, and mutually exclusive, binding orientations for the sugar rings of the 1-phospho and 6-phospho sugars. Specific binding of both orientations is accomplished by key contacts with the O3 and O4 hydroxyls of the sugar, which must occupy equatorial positions. Dual recognition of glucose and mannose phosphosugars uses a combination of specific protein contacts and nonspecific solvent contacts. The ability of PMM/PGM to accommodate these four diverse substrates in a single active site is consistent with its highly reversible phosphoryl transfer reaction and allows it to function in multiple biosynthetic pathways in P. aeruginosa.

Binding Sites↗
PubMed2004

Enhancement of DNA vaccine potency by coadministration of a tumor antigen gene and DNA encoding serine protease inhibitor-6.

Serine protease inhibitor 6 (SPI-6), also called Serpinb9, inhibits granzyme B and thus may provide a method for delaying apoptotic cell death in dendritic cells. We have previously enhanced DNA vaccine potency by targeting antigen to MHC antigen presentation pathways, using proteins such as Mycobacterium tuberculosis heat shock protein 70, calreticulin, domain II of Pseudomonas aeruginosa exotoxin A, or the sorting signal of the lysosome-associated membrane protein type 1. In this study, we explored intradermal coadministration of DNA encoding SPI-6 with DNA constructs encoding human papillomavirus type 16 E7 linked to these intracellular targeting molecules for its ability to generate E7-specific CD8+ T-cell immune responses and E7-specific antitumor effects. This combination of strategies resulted in significantly increased E7-specific CD8+ T-cell and CD4+ Th1-cell responses, enhanced tumor treatment ability, and stronger tumor protection when compared with vaccination without SPI-6. Among these targeting strategies tested, mice vaccinated with Sig/E7/lysosome-associated membrane protein type 1 mixed with SPI-6 showed the greatest fold increase in E7-specific CD8+ T cells ( approximately 5-fold). Vaccination with a nonfunctional mutant of SPI-6 did not result in immune enhancement, indicating that enhancement was dependent on the antiapoptotic function of SPI-6. Our results suggest that DNA vaccines combining strategies that enhance MHC class I and II antigen processing with SPI-6 have potential clinical implications for control of viral infection and neoplasia.

Animals↗
PubMed2006

Rapid screening for major depression in post-myocardial infarction patients: an investigation using Beck Depression Inventory II items.

OBJECTIVE: To determine the ability of three questions from the Beck Depression Inventory II (BDI-II) to detect major depressive disorder (MDD) in a cohort of patients hospitalised for acute myocardial infarction (MI). DESIGN: Prospective observational study. SETTING: Coronary care unit and cardiac step-down unit of an urban academic medical centre. PATIENTS: 131 post-MI patients within 72 h of symptom onset. INTERVENTIONS: Patients were administered the BDI-II and participated in a structured diagnostic interview for MDD. Three individual BDI-II items (regarding sadness, loss of interest and loss of pleasure) were examined individually and in two-question combinations to determine their ability to screen for MDD. MAIN OUTCOME MEASURES: Sensitivity, specificity, negative and positive predictive values and proportion of patients with MDD correctly identified. RESULTS: The individual items and two-question combinations had good sensitivity (76-94%), specificity (70-88%) and negative predictive values (97-99%). Item 1 (sadness) performed the best of the individual items (48% with a positive response to the item had MDD; 3% with a negative response had MDD; over 80% of patients with MDD were correctly identified). A combination of questions about sadness and loss of interest performed best among the two-question combinations (37% with positive response had MDD v 1% with a negative response; 94% of patients with MDD were identified). CONCLUSIONS: One to two questions regarding sadness and loss of interest serve as simple and effective screening tools for post-MI depression.

Cohort Studies↗
PubMed1989

Stimulation of macrophage phagocytic but not bactericidal activity by colony-stimulating factor 1.

The ability of mouse peritoneal cells to phagocytose and lyse Listeria monocytogenes was measured after the cells were incubated with purified murine macrophage-specific colony-stimulating factor (CSF-1). Activation of combined phagocytic and bacteriolytic ability required 24 h, with an optimal dose of 1,000 U of CSF-1 per ml. No activation was achieved with a shorter period of incubation, known to be sufficient for GM-CSF to stimulate phagocytosis by granulocytes, and there was no advantage in longer exposure. After 24 h in 1,000 U of CSF-1, macrophages showed visibly increased spreading on the plastic petri dish. Activated cells examined microscopically showed an increase in the number of phagocytic cells and in the numbers of bacteria per phagocytic cell. This increased phagocytic ability was evident also in the increase in the amount of radioactivity associated with the cells following a 30-min incubation with radiolabeled bacteria. When these cells were carefully washed, the percentage of this initial uptake released during the next 2 h was not increased by pretreatment of the cells with CSF-1, showing that the effect of this growth factor was on phagocytosis of the bacteria not on the killing mechanisms per se.

Animals↗
PubMedDate not supplied

Immunomodulating efficacy of combined administration of galactoside-specific lectin standardized mistletoe extract and sodium selenite in BALB/c-mice.

The immunomodulating abilities of commercially available mistletoe extract standardized for the galactoside-specific lectin (mistletoe lectin-1, ML-1) and sodium selenite (Se) were evaluated in BALB/c-mice and compared to non-treated control animals. Following the optimal schedule of administration (ML-1: 1ng/kg BW; days 1, 2, 3, 5 and Se: 3,5 micrograms/kg BW for 7 subsequent days before evaluation) yielded enhanced counts (thymocytes; peritoneal macrophages, MO; peripheral blood leukocytes; lymphocytes, PBL; monocytes, PBM) and activities (CD-25 positive PBL, MAC-3 positive PBM) of desired immune cells reaching statistical significance for most parameters. However, combined administration of ML-1 and Se proved to be superior to monotherapy since immune cell counts (thymocytes, leukocytes, PBL, PBM) and activities (CD-25 positive PBL) exceeded values obtained after monotherapy. These data are in favour of therapeutical strategies in complementary oncology and suggest that the combination of defined immunomodulating substances might positively enhance the efficacy.

Animals↗