Incidence of chromosomal imbalances in advanced colorectal carcinomas and their metastases

Comparative genomic hybridization (CGH) was used to screen 54 advanced colon carcinomas. i.e., 24 primary tumors and 30 metastases, for chromosomal alterations. Using a sensitive statistical method for the determination of DNA imbalances and histograms for analysis of the incidence of changes, we identified the DNA over-representation of chromosome 20q as the most common alteration being present in 100% of cases. High incidence deletions were observed on 18q21-18q23 (96%), 4q27-4q28 (96%), 4p14 (87%), 5q21 (81%), 1p21-1p22 (72%), 21q21 (74%), 6q16 (72%), 3p12 (66%), 8p24-8p21 (66%), 9p21 (64%), 11q22 (64%), and 14q13-14q21 (64%). Further frequent over-representation was found on 7q12-7q11.2 (75%), 16p11-16p12 (70%), 19p13 (70%), 9q34 (67%), 19q13 (67%), 13q34 (64%), 13q13 (64%), 17q21 (59%), 22q11 (61%), 8q24 (57%), and 1q21 (57%). Pronounced DNA gains and losses being defined as regions in which the ratio profiles exceeded the values of 1.5 and 0.5, respectively, frequently colocalized with peaks of incidence curve. The use of difference histograms for the comparison of tumor subgroups as well as case-by-case histogram for the analysis of 15 paired tumor samples identified several of the above alterations as relevant for tumor progression and metastasis formation. The study identified additional loci and delineates more precisely those that have been previously reported. For comparative purposes, we have made our primary data (ratio profiles, clinicopathological parameters, histograms) available at the interactive web site http://amba.charite.de/cgh, where the incidence of changes can be determined at individual loci and additional parameters can be applied for the analysis of our CGH results.     

Activation kinetics of Glutamate receptor channels from wild-type Drosophila muscle

Outside-out patches from wild-type Drosophila larval muscle were exposed briefly to L-Glutamate (Glu) using a piezo-driven application system. Glu in concentrations of 0.1 to 30 mM was applied and the responses to repeated applications of a given concentration were averaged. The peak current, î, and the current rise time, t_r, from 0.1 î to 0.9 î were determined from the averages. Half-maximum activation of the channels was reached with ≈ 2 mM Glu. î increased proportional to the power n = 3.5 to n = 5.8 (average of four experiments, n = 4.4) for Glu concentrations between 0.3 and 0.5 mM. t_r increased from ≈ 0.2 ms at 10 mM Glu to a value of ≈ 3.5 ms at 0.2 mM Glu. A linear reaction scheme with five binding steps preceding the channel-opening conformational change is proposed as the kinetic mechanism of channel activation and investigated in computer simulations. A set of rate constants assuming the same affinity for each binding site is found to describe the data better than one assuming positive cooperativity. The results are very similar to those for Glu-gated channels of crayfish and locust muscle, which is evidence for a common kinetic mechanism of these channels.     

Clonal differentiation of uropathogenic Escherichia coli isolates of serotype O6:K5 by fimbrial antigen typing and DNA long-range mapping techniques

Escherichia coli isolates of serotype O6:K5 are the most common causative agents of cystitis and pyelonephritis in adults. To answer the question, as to whether strains of this particular serotype represent one special clonal group, out of a collection of 34 serotype O6:K5 isolates [Zingler et al. (1990) Zentralbl. Bakteriol Mikrobiol Hyg [A] 274:372–381] 15 strains were selected and analyzed in detail. The flagellar (H) antigen and the outer membrane protein (OMP) pattern were determined. Further serum resistance properties and the genetic presence and expression of other virulence factors, including hemolysin, aerobactin, P fimbriae, S/F1C fimbriae and type 1 fimbriae was evaluated. In addition the Xba-Imacrorestriction pattern of ten representative isolates was elaborated and the fimbrial (F) antigen type of the P fimbriae was determined, to obtain the complete O:K:H:F pattern. These analyses could clearly show that the O6:K5 isolates do not represent one clonal group. The XbaI-macrorestriction profiles were heterogeneous and marked differences in the hybridization patterns, using virulence-associated gene probes in Southern hybridization of long-range-separated genomic DNA, were observed among the strains. However, some of strains showed similarities in the genomic profiles, arguing for clonal groupings among the O6:K5 isolates. Interstingly the strains grouped together exhibited the same fimbrial F type that many indicate a coincidence of this phenotypic trait with clonality.In memoriam of Prof. G. Naumann     

The direct early diagnosis of cystic fibrosis by the detection of the deltaF508 CFTR gene mutation in a prematurely delivered boy

The suspicion of prenatal meconium ileus syndrome was raised in a pregnancy in a family with no history of cystic fibrosis because of significantly higher maternal serum alpha-fetoprotein in the 16th and 19th week of gestation, dispersed areas with increased echogenity in the fetal abdomen, slight fetal ascites in the 24th-25th weeks of gestation, decreased amniotic fluid gamma-glutamyltranspeptidase (GGT) activity and alpha-fetoprotein level in the 25th-26th weeks, and normal 46,XY karotype of the fetus. The detection of a homozygous deltaF508 cystic fibrosis transmembrane regulator (CFTR) gene mutation, by means of PCR from a small amount of white blood cells and urine sediment cells, substantiated the diagnosis of cystic fibrosis in a prematurely delivered boy in the 28th week of gestation. The repeated sweat test was unsuccessful. The autopsy examination confirmed the diagnosis of cystic fibrosis. Fetal meconium ileus syndrome was complicated by peritonitis and by formation of a meconium pseudocyst. Direct PCR typing improves postnatal diagnostic possibilities in the early neonatal period in prematurely delivered babies when the sweat test is difficult to perform.     

Utility of 123C3 monoclonal antibody against CD56 (NCAM) for the diagnosis of small cell carcinomas on paraffin sections

CD56 is immunohistochemically detectable in virtually all small cell carcinomas on frozen sections. The authors retrospectively tested the usefulness of the monoclonal antibody 123C3 against CD56 to differentiate pulmonary and extrapulmonary small cell carcinomas from nonneuroendocrine non—small cell carcinomas by paraffin-section immunohistochemistry after antigen retrieval. The study included 70 small cell carcinomas and 344 primary and metastatic nonneuroendocrine carcinomas of various primary sites. The staining results were compared with specific neuroendocrine markers (CD57, Chromogranin A, Synaptophysin). The monoclonal antibody 123C3 diffusely stained most small cell carcinomas with a strong membranous pattern (sensitivity: 0.99). The staining intensity was not diminished in areas with crush artifacts or after decalcification. The neuroendocrine markers had a combined sensitivity of only 0. 44 for small cell carcinomas. With regard to nonneuroendocrine carcinomas, the 123C3 antibody stained 7 of 28 ovarian carcinomas, 6 of 30 renal cell carcinomas, 2 of 10 endometrial carcinomas, two of three nonneuroendocrine large cell carcinomas of the lung, 1 of 38 adenocarcinomas, and 4 of 52 squamous cell carcinomas of the lung. Urothelial carcinomas, hepatocellular carcinomas, squamous carcinomas of the head/neck and cervix uteri, as well as adenocarcinomas of the breast, stomach, colon, pancreas, and prostate, showed no immunoreactivity for CD56. The specificities of 123C3 and the combined neuroendocrine markers for small cell carcinomas were 0. 94 and 0. 95, respectively. The authors conclude that monoclonal antibody 123C3 might be useful for the immunohistochemical differentiation of small cell carcinomas from nonneuroendocrine carcinomas on paraffin sections, especially in small and crushed biopsy specimens.     

Primary structure of human amphiphysin, the dominant autoantigen of paraneoplastic Stiff--Man syndrome, and mapping of its gene (AMPH) to chromosome 7p13-p14

Amphiphysin is a protein peripherally associated with synapticvesicles. It is expressed in many neurons, certain endocrinecell types, and spermatocytes. Autoantibodies against amphiphysinoccur in patients afflicted with a rare neurologic autoimmunedisease, paraneoplastic Stiff—Man syndrome. To providea basis for the understanding of antl-amphiphysin autolmmunity,we have cloned cDNAs and determined the primary structure ofhuman amphiphysin. Comparison with chicken amphiphysin definesdomains of low and high amino acid sequence conservation. Asa candidate for heritable disorders of the nervous system, endocrinetissues or male fertility, the human amphiphysin gene was mappedto chromosome 7, region p13–p14.     

Dilute solution properties of imogolite

Imogolite, a natural product in the clay fraction of Japanese soil, was characterized through its dilute solution properties. Various methods were employed for this characterization, including viscosity, sedimentation, static/dynamic light scattering, and small angle X-ray scattering. All these measurements have revealed consistently that imogolite is represented by a rigid thin rod within the accuracy of available theories, where its repeat unit is composed of twelve gibbsite units. Since the evaluation of the chain length from the observed quantities depends on the molecular weight distribution, its effect was also considered where M_w/M_n ≈ 1,2 was estimated from the sedimentation profile.     

Inhibition of leukotriene formation and IL-8 release by the PAF-receptor antagonist SM-12502

We analyzed the effect of the PAF receptor antagonist (+)-cis-3,5-dimethyl-2-(3-pyridyl) thiazolidin-4-one hydrochloride (SM-12502) on the release of leukotriene B₄ and IL-8 from human leukocytes. Peripheral blood from healthy donors was separated in two different fractions: polymorphonuclear leukocytes (PMN) and a lymphocyte, monocyte and basophil granulocyte cell fraction (LMB). After incubation of the cell population with different concentrations of SM-12502 the cells were subsequently stimulated with either the Ca ionophore A23187, the bacterial derived peptide fMLP, or with an activator of heterotrimetric G-proteins, the sodium fluoride (NaF, in the presence of Al³⁺). The PAF receptor antagonist led to a concentration and time dependent inhibition of LTB₄ formation and IL-8 release from PMN and LMB. Our data clearly indicate an inhibitory effect of the PAF receptor antagonist SM-12502 on the formation of mediators of the lipoxygenase pathway and on the release of IL-8.