Detection of point mutations in KRAS oncogene by real-time PCR-based genotyping assay in GIT diseases.

OBJECTIVES: The determination of gene mutations is important for the diagnosis and prognosis of various gastrointestinal cancers. The aim of our study was to develop a new procedure for the analysis of KRAS gene mutation by application of the real-time PCR method. BACKGROUND: The detection process requires discriminate trace amount of mutant allele in a large excess of wild-type DNA in various samples. METHODS: The real-time PCR based technique using hybridization probes for five most frequently KRAS codon 12 mutations and WT specific peptide nucleic acid (PNA) was performed. Our multiplex detection system was tested in various DNA samples (tissue, bile, pancreatic juice) of patients with different diagnoses of gastrointestinal tract disease obtained by endoscopy and ERCP. RESULTS: We designed and optimized the real-time PCR conditions and tested various amount of PNA in PCR reaction to suppress amplification of the wild-type DNA. We determined the interassay variability of the melting temperatures and the results of mutation testing were confirmed by DNA sequencing with the 100 % accuracy. Incidence of searched mutations was 67.5 % in cohort of 40 patients; for KRASG12D it was in 44.4 %, KRASG12V in 22.2 %, KRASG12S in 14.8 %, KRASG12A in 14.8 % and KRASG12C in 3.8 %. The sensitivity of the assays is 1x10-5. CONCLUSIONS: Advantages of this technique are rapidity, accuracy and it is generally easy to perform. This method can be adapted for synchronic detection of multiple mutations and after readjustment by other type mutation of KRAS gene may serve as useful clinical tool for analyzing point mutations in various clinical samples (Tab. 3, Fig. 3, Ref. 42).

[Endoscopic therapy of pancreatic pseudocysts]. / Endoskopická liecba pseudocýst pankreasu.

BACKGROUND: Pancreatic pseudocysts are a complication of both acute and chronic pancreatitis. Incidence in patients with acute pancreatitis is 2-50%, in patients with chronic pancreatitis 20-40%. Pseudocysts are a cause of many symptoms, e.g. nausea, vomitus, pain, biliary obstruction, bleeding and perforation. Successful treatment of pseudocysts is not only surgical and percutaneous, but also endoscopic. OBJECTIVES: The aim of this study was to answer the following questions. First, what is the clinical success rate of endoscopic drainage of pancreatic pseudocysts? Second, what are the complications? Finally, how often is endoscopic drainage a definite treatment? METHODS: The records of all patients (11) with chronic pancreatitis and endoscopic drainage of symptomatic pseudocysts hospitalized between December 1993 and April 1999 at our clinic were retrospectively studied. RESULTS: Patients (5) were followed for a mean duration of 30 months. Endoscopic drainage was definitive treatment in 80%, after transgastric drainage in 50%, after transpapillary drainage in 100% and after the use of more than one drainage procedure in 0%. The prognostic factors for longterm success of endoscopic drainage could not be evaluated, because of the small number of treated patients. CONCLUSIONS: Endoscopic treatment of pancreatic pseudocysts (endoscopic cystogastrostomy, cystoduodenostomy and transpapillary drainage) is nowadays highly effective method, technically feasible in most patients, with a relative degree of safety when performed by experienced endoscopist. (Tab. 2, Ref. 16.)

[Hepatorenal syndrome]. / Hepatorenálny syndróm.

BACKGROUND: The association of renal failure with terminal liver insufficiency is referred to as hepatorenal syndrome (HRS). It is characterized by impaired renal function together with abnormalities in arterial circulation and activity of endogenous vasoactive system. In the pathogenesis of HRS, vasodilatation in the splanchnic circulation plays a major role. This leads to underfilling in arterial circulation in this area with subsequent vasoconstriction in renal arteries leading to lower renal circulation and functional renal failure. One of the most difficult tasks in clinical evaluation of patients with cirrhosis is to diagnose renal functions, because the standard methods used in evaluation of glomerular filtration are not reliable. The most important agents in the treatment of patients with cirrhosis, ascites and HRS are identification, prevention of precipitating factors, correction and treatment of prerenal failure and acute tubular necrosis. Up to now, the only effective treatment of HRS is ortothopic liver transplantation, which cannot be applied in all patients. CONCLUSIONS: HRS is a serious complication of liver diseases with a very bad prognosis qouad vitam and qouad sanationem, which is only rarely reversible. After diagnosing HRS, the renal insufficiency is already mostly progressive, and patients have only a minimal chance to improve their renal functions. The only currently proved treatment modality, which is successful in terms of improving renal functions and long-term survival rate is liver transplantation. (Tab. 2, Fig. 1, Ref. 28.)

Pulmonary complications in patients with inflammatory bowel disease.

BACKGROUND/AIMS: Between 1996 and 1998 we investigated the occurrence of lung disorders in 82 patients with inflammatory bowel disease (30 patients with ulcerative colitis and 52 patients with Crohn's disease) and a control group of 60 subjects. The aim of our study was to determine the occurrence of pulmonary complications in patients with inflammatory bowel disease, to investigate whether ulcerative colitis or Crohn's disease are connected with a typical lung function disorder, with the inflammatory activity of the disease or if they depend on the presence of other extraintestinal manifestations. METHODOLOGY: We investigated the occurrence of lung disorders in terms of the following parameters: clinical pulmonary symptoms, chest radiography and pulmonary function tests (body plethysmography, pneumotachography, lung transfer capacity for carbon monoxide, and blood gas analysis). RESULTS: Lung function abnormalities were significantly more frequent in patients with inflammatory bowel disease as compared to controls (p<0.001). There was no apparent correlation between these abnormalities and either bowel disease activity or drug administration (sulphasalazine, mesalazine). CONCLUSIONS: Despite the lack of radiological abnormalities, we identified a high incidence of pulmonary function abnormalities (suspicious of interstitial lung disorder) in patients with inflammatory bowel disease; 56.7% of patients with ulcerative colitis and 57.7% of patients with Crohn's disease had reduced lung transfer factor.