Determination of linear expansion of AlSi10Mg aluminium alloy depending on external conditions during solidification.

The article deals with the possibility of influencing the properties of aluminium alloy castings by changing the external conditions during solidification. Due to the high demands of customers, influencing the resulting microstructure is an important process in the production of castings. When casting an aluminium alloy into a metal mould, the mould must be preheated to a high temperature. For this reason, the key parameter of production is the mastery of the control of the resulting microstructure of castings and the related internal quality and mechanical properties, which are subject to high demands defined by international standards. The aim of the experiment is to evaluate the influence of different preheating of the metal mould on the resulting structure of test castings made of AlSi10Mg material and its subsequent thermal expansion. It is hypothesized that as the temperature of the mould preheating increases, the mechanical properties will deteriorate and the expansion of the alloy will increase. In the case of immediate use of castings after production, this could have a negative impact on the service life and safety of entire assembled components. All measured results will be put into comparison and direct connections between the data will be searched using the microsoft excel program.

Simple procedure for automatic detection of unstable alleles in the myotonic dystrophy and Huntington's disease loci.

Human neurodegenerative and neuromuscular disorders are associated with a class of gene mutations represented by expansion of trinucleotide repeats. DNA testing is important for the diagnosis of these diseases because clinical discrimination is complicated by their late onset and frequently overlapping symptomatology. However, detection of pathologic alleles expanded up to several thousand trinucleotides poses a challenge for the introduction of rapid, fully automatic, and simple DNA diagnostic procedures. Here we propose a simple two-step polymerase chain reaction (PCR) protocol for rapid molecular diagnostics of myotonic dystrophy, Huntington's disease, and possibly also other triplet expansion diseases. Standard PCR amplification with target repeat flanking primers is used for the detection of alleles of up to 100 repeats; next, triplet-primed PCR is applied for detection of larger expansions. Automated capillary electrophoresis of amplicons allows rapid discrimination between normal, premutated and expanded (CTG/CAG)(n) alleles. Using the suggested protocol, the expanded allele was successfully detected in all test DNA samples with known genotypes. Our experience demonstrates that the suggested two-step PCR protocol provides high sensitivity, specificity, and reproducibility; is significantly less time-consuming; is easier to perform; and provides a better basis for automation than previous methods requiring Southern analysis. Therefore, it can be used for confirmation of uncertain clinical diagnoses, for prenatal testing in at-risk families, and, generally in research on these diseases.

Amplification and overexpression of HER-2/neu in invasive ductal carcinomas of the pancreas and pancreatic intraepithelial neoplasms and the relationship to the expression of p21(WAF1/CIP1).

Overexpression of HER-2/neu was described in pancreatic intraepithelial neoplasia (PanIN) and in invasive ductal adenocarcinoma of pancreas in a variable proportion of cases. The effects of HER-2/neu overexpression on mitogenic signalling and cell cycle progression were studied in breast luminal epithelial cells and mitogen activated protein kinase-dependent induction of p21(WAF1/CIP1) was found to be necessary for G1 phase progression. Overexpression of p21(WAF1/CIP1) was described as an early event in the development of PanIN by Biankin et al. (2001) and this finding was supported by our previous study that, moreover, did not confirm the possible role of activating K-ras mutations in the induction of p21(WAF1/CIP1) overexpression. Relationship between p21(WAF1/CIP1) expression and HER-2/neu status in PanIN lesions and ductal adenocarcinoma of the pancreas was investigated in our study. Expression levels of p21(WAF1/CIP1) and HER-2/neu were examined imunohistochemically and the amplification of HER-2/neu gene was evaluated by fluorescence in situ hybridisation in HER-2/neu overexpressing adenocarcinomas. Fourty nine pancreatic resection specimens from patients with invasive adenocarcinoma were included into the study. A large spectrum of PanIN lesions adjacent to the structures of infiltrating adenocarcinoma was also examined. The possible role of HER-2/neu in an induction of p21(WAF1/CIP1) overexpression was not confirmed and p21(WAF1/CIP1) overexpression seems to be HER-2/neu independent in pancreatic ductal adenocarcinoma according to our results. Increasing levels of HER-2/neu expression were demonstrated in pancreatic intraepithelial neoplasia and in 18.75% of pancreatic adenocarcinoma. The only 2 from 9 HER-2/neu overexpressing adenocarcinomas showed the amplification of HER-2/neu gene. Based on these results, the overexpression of HER-2/neu in pancreatic adenocarcinoma seems to be a result of increased transcription rather than gene amplification. Therefore HER-2/neu represents a good target for therapy of pancreatic adenocarcinoma only in isolated cases.

Relationship between K-ras mutation and the expression of p21WAF1/CIP1 and p53 in chronic pancreatitis and pancreatic adenocarcinoma.

Overexpression of p21WAF1/CIP1 was recently described as an early event in the development of pancreatic intraepithelial neoplasia. Since activating K-ras mutations are described in more than 80% of pancreatic cancers and are known to increase intracellular levels of p21WAF1/CIP1 in experimental models, the possible role of activating K-ras mutations in an induction of the p21WAF1/CIP1 expression was investigated in our study. We examined 71 surgical specimens, 29 of chronic pancreatitis and 42 of invasive ductal adenocarcinoma both having a large spectrum of PanIN (pancreatic intraepithelial neoplasia) lesions. Expression of p53 and p21WAF1/CIP1 was examined immunohistochemically and codon 12 K-ras mutational analysis was performed using the very sensitive mutant-enriched PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) analysis. Our study demonstrated the overexpression of p21WAF1/CIP1 as an early event in the development of pancreatic intraepithelial neoplasia in the group of chronic pancreatitis and invasive adenocarcinoma as well. Overexpression of p21WAF1/CIP1 increased progressively from normal ducts through the spectrum of PanIN lesions to invasive carcinomas. The p53 overexpression increased again progressively according to the severity of the lesion and seems to be a later event in the development of pancreatic intraepithelial neoplasia if compared to p21WAF1/CIP1 expression. Our results confirmed also the possible p53 independent p21WAF1/CIP1 expression in some PanIN2, PanIN3 lesions and invasive carcinomas. K-ras mutations were not revealed in samples with only low grade PanIN lesions (PanIN1a and PanIN1b). K-ras mutations were detected in 69,4% adenocarcinomas and in only one case of chronic pancreatitis. Two codon 12 K-ras positive pancreatic carcinomas showed K-ras mutations in the surrounding normal pancreatic tissue. In adenocarcinomas, no statistically significant correlation was found between K-ras mutational status and p21WAF1/CIP1 and p53 expression, respectively. The possible role of activating K-ras mutations in an induction of p21WAF1/CIP1 expression was not confirmed in this study.

[Molecular diagnosis in pancreatic carcinoma]. / Príspevek k molekulární diagnostice pankreatického karcinomu.

Present diagnostic possibilities virtually do not make it possible to diagnose early stages of pancreatic cancer. Likewise, it is very difficult to differentiate between pancreatic cancer and chronic pancreatitis. The methods of visualization are insufficiently sensitive and the determination of certain genes could enrich our diagnostic opportunities. Considerable attention in this direction has been devoted to the determination and evaluation of the presence of oncogene K-ras. Our initial experience with the determination of K-ras in preparations from patients with pancreatic cancer or with chronic pancreatitis confirmed that K-ras in an oncomarker associated with adenocarcinoma of pancreas, whereas in patients with chronic pancreatitis it occurs in about 10% of the examined samples.

[Detection of carriers of Duchenne and Becker muscular dystrophy using the fluorescence in situ hybridization method]. / Identifikace prenasecek Duchenneovy a Beckerovy svalové dystrofie metodou fluorescencní in situ hybridizace.

Duchenne and Becker muscular dystrophies (DMD/BMD) are X-linked recessive disorders caused by mutations in the dystrophin gene. A large intragenic deletion has been described in about 65% of DMD/BMD patients. Mothers of affected males are DMD/BMD carriers in two thirds of the cases. Routine deletions detection in DMD/BMD males is performed using multiplex polymerase chain reaction (mPCR), RT-PCR with a protein truncation test (PTT) or using Southern blotting. In females the deletions detection is complicated by the presence of a normal gene copy on the second X-chromosome. We are presenting the diagnostic strategy using FISH for the deletions detection in the dystrophin gene of female DMD/BMD carriers. We have used a set of six cosmid probes for the detection of the most frequently deleted areas of the dystrophin gene from the Department of Human Genetics, Leiden University Medical Center. We have examined 14 mothers of DMD/BMD males with a deletion in the dystrophin gene identified using mPCR. Four mothers of affected males have been diagnosed as carriers of a deletion in the dystrophin gene. We have revealed no deletion mutations in the exons examined in a control group of four healthy females. No discrepancy has been found between the FISH analysis results and the results of mPCR. Our results indicate that FISH is an effective and direct method for the identification of DMD/BMD carriers and we suggest this method as a method of a first choice in the identification of DMD/BMD carriers.