In vitro and clinical feasibility study with an over-the-wire delivery system for pulsed dye laser angioplasty.

The suitability of a pulsed dye laser (504 nm) in experimental and clinical angioplasty was investigated. In an experimental study, the ablation thresholds were 3 J/cm2 +/- 8 (mean +/- standard deviation) for fibrofatty plaque and 25 J/cm2 for calcified tissues under saline. At a radiant exposure of 10 J/cm2 the etch rates were 1.7 microns per pulse +/- 0.3 for media, 2.8 microns/pulse +/- 0.4 for normal intima, and 3.9 microns/pulse +/- 1.1 for fibrofatty plaque (P less than .05). Pressure wave effects with a separation of tissue layers were predominantly localized at the internal elastic lamina. Thermal injury with vacuolations extended 15 microns +/- 6 into adjacent tissue. For clinical study, laser-assisted balloon angioplasty was performed in 10 patients (64 years +/- 14) with occlusions of peripheral arteries using a 9-F multifiber ring catheter. Lesion length ranged from 2 to 12 cm (mean, 7 cm). Laser angioplasty with an 80 mJ/pulse decreased the mean stenosis from 100% to 58% +/- 12% (P less than .005). The ankle-brachial index rose from a median of 0.48 to 0.88 (P less than .001). In 33% of patients, there were subintimal dissections after laser angioplasty. After a mean follow-up of 10.2 months, the overall clinical success was 70% with a primary patency of 78%. The over-the-wire approach with a pulsed dye laser may constitute a safe and feasible tool in laser angioplasty.     

Isolation of DNA from the centromere of human chromosome 7 by microdissection

Centromeres remain the least characterized regions of human chromosomes because they have a very high content of repetitive DNA. Here, we describe a micro-dissection library from the centromeric region of human chromosome 7 and its use for generating sequence tagged sites (STSs). The library contains about 1500 clones with an average insert size of 150 bp and only about 15% of the clones harbour repetitive human DNA. Seven clones hybridizing to alphoid DNA were found to correspond to a fragment of the D7Z2 alphoid array on chromosome 7, thus confirming the origin of the library. A number of clones not containing known repetitive DNA were used to generate STSs that identified yeast artificial chromosomes (YACs) and in turn allowed the STSs to be placed on the physical map. One STS is located between the two Genethon genetic markers closest to the centromere on the q side. Another STS was located 3–4 cM away in 7q11.2, while a third identified YACs containing both low-copy and alphoid sequences that are not yet mapped but are clearly centromeric. The library therefore comprises a collection of sequences from the centromeric region of chromosome 7 that can be used to generate STSs and to map the entire centromeric region.This revised version was published online in November 2005 with corrections to the Cover Date.     

The origin of human chromosome 2 analyzed by comparative chromosome mapping with a DNA microlibrary

Fluorescencein situ hybridization (FISH) of microlibraries established from distinct chromosome subregions can test the evolutionary conservation of chromosome bands as well as chromosomal rearrangements that occurred during primate evolution and will help to clarify phylogenetic relationships. We used a DNA library established by microdissection and microcloning from the entire long arm of human chromosome 2 for fluorescencein situ hybridization and comparative mapping of the chromosomes of human, great apes (Pan troglodytes, Pan paniscus, Gorilla gorilla, Pongo pygmaeus) and Old World monkeys (Macaca fuscata andCercopithecus aethiops). Inversions were found in the pericentric region of the primate chromosome 2p homologs in great apes, and the hybridization pattern demonstrates the known phylogenetically derived telomere fusion in the line that leads to human chromosome 2. The hybridization of the 2q microlibrary to chromosomes of Old World monkeys gave a different pattern from that in the gorilla and the orang-utan, but a pattern similar to that of chimpanzees. This suggests convergence of chromosomal rearrangements in different phylogenetic lines.     

Molecular cytogenetic characterization of the mantle cell lymphoma cell line GRANTA-519

Combining fluorescence R-banding, fluorescence in situ hybridization and spectral karyotyping allowed us to precisely define chromosomal breakpoints, gains, losses and a newly detected amplification in the human mantle cell lymphoma (MCL) cell line GRANTA-519. GRANTA-519 is characterized by the t(11;14)(q13;q32) resulting in overexpression of cyclin D1, a key player in cell cycle control. Hitherto unresolved complex rearrangements involve 1p, 1q, 3cen, 9p, 11q, 12p, 12q, 16p, 17p, and 18cen. Moreover, a 4- to 6-fold gain of sequences on 18q leads to a low-level amplification of the BCL2 gene and to an overexpression of the BCL2 protein. These results provide the basis for the identification of not only candidate oncogenes responsible for MCL in gained regions, but also for the identification of putative tumor suppressor genes in commonly deleted regions like 1p22, which would eventually enable functional studies of these genes.     

A long distance-PCR derived FISH probe detects a deletion between p15 and p16 in CML and T-ALL patients

The tumor suppressor genes p15INK4B and p16INK4A, located in the chromosomal region 9p21, are frequently inactivated by homo- or hemizygous deletions, point mutation or promotor methylation in various types of cancer. No commercial probe is yet available that allows the detection of such deletions by FISH. Long distance (LD)-PCR was successfully used to generate a FISH probe, that covers a sequence stretch of 11.68 kb, located between the tumor suppressor genes p15 and p16. The LD-PCR amplicon was cloned and biotinylated by DOP-PCR (degenerated oligonucleotide primed-PCR) or nick translation. The FISH probe was hybridized on different samples of 16 patients with leukemia (3 T-ALL, 13 CML) and normal controls. Loss of at least one FISH-signal was found in 2/3 (67%) of the T-ALL- and 2/13 (15%) of the CML-cases. The new FISH probe presented here was proven to be advantageous for the detection of deletions in chromosomal region 9p21, especially between p15 and p16.     

Identification of protein pattern in kidney cancer using ProteinChip arrays and bioinformatics

Tumor biology of renal cell carcinoma (RCC) is not very well understood, although many studies on molecular and cellular biology have been performed. It is accepted now that cancer research has to be performed also with proteomic tools, because proteins are the real actors in the genesis and progression of cancer. Therefore, we used a ProteinChip System(R) (SELDI) which is able to detect minute amounts of protein and moreover to analyze a complex protein pattern. We analyzed 37 cases of clear cell RCC as a training set including corresponding normal tissue. From all samples protein lysates were made and spotted directly on different chip surfaces (SAX2, WCX). After a washing procedure the arrays were analyzed in the ProteinChip Reader. All profiles were subjected to a bioinformatical analysis including normalization, clustering, rule extraction and rating. Defined rules (markers) were evaluated using a test set of 24 samples (13 tumor tissues and 11 normal kidney tissues). The generated rule base for the SAX2 surface showed a sensitivity of 100% and a specificity of 97.3%. For the WCX arrays the optimal rule base showed worse results. A combined rule base for SAX2 and WCX did not result in a higher sensitivity or specificity. Using the optimal rule base for the SAX2 chip in the test set, sensitivity and specificity reached 76.9% and 100%, respectively. The ProteinChip System represents a key technology for the rapid detection of cancer specific proteomic patterns. It is possible to identify clear cell renal cancer with high sensitivity and specificity from minimal amounts of cells.     

Etablierung eines Tiermodells des hepatozellul ren Karzinoms durch den Einsatz von allogenischer Implantationstechnik

Das hepatozellul re Karzinom ( HCC) ist mitmehr als 1 Million Erkrankungsf llen pro Jahrweltweit einer der h ufigsten malignen Tumoren[1] .Zweifellos ist das HCC- Tiermodell in der Strategiegegen HCC von gro er Bedeutung.Bisher stehtvielf ltige Verfahren zur Etabierung des HCC- Tier-modells zur Verf gung.Im folgenden wurde einHCC- Rattesmodell,das durch den Einsatz von allo-genischer Implantationstechnik erzeugt wurde,unddie betreffende Charaktere vorgestellt.1　 MATERIAL UND …