The response of L5178Y lymphoma sublines to oxidative stress: antioxidant defence, iron content and nuclear translocation of the p65 subunit of NF-kappaB

We examined the response to hydrogen peroxide of two L5178Y (LY) sublines which are inversely cross-sensitive to hydrogen peroxide and X-rays: LY-R cells are radio-resistant and hydrogen peroxide-sensitive, whereas LY-S cells are radiosensitive and hydrogen peroxide-resistant. Higher initial DNA breaks and higher iron content (potentially active in the Fenton reaction) were found in the hydrogen peroxide sensitive LY-R cells than in the hydrogen peroxide resistant LY-S cells, whereas the antioxidant defence of LY-R cells was weaker. In particular, catalase activity is twofold higher in LY-S than in LY-R cells. The content of monobromobimane-reactive thiols is 54% higher in LY-S than in LY-R cells. In contrast, the activity of glutathione peroxidase (GPx) is about two times higher in LY-R than in LY-S cells; however, upon induction with selenium the activity increases 15.6-fold in LY-R cells and 50.3-fold in LY-S cells. Altogether, the sensitivity difference is related to the iron content, the amount of the initial DNA damage, as well as to the efficiency of the antioxidant defence system. Differential nuclear translocation of p65-NF-kappaB in LY sublines is due to the more efficient antioxidant defence in LY-S than in LY-R cells.     

The reactions of hypochlorous acid, the reactive oxygen species produced by myeloperoxidase, with lipids

Myeloperoxidase (MPO), an abundant enzyme in phagocytes, has been implicated in the pathogenesis of various inflammatory diseases including atherosclerosis. The major oxidant produced by MPO, hypochlorous acid (HOCl), is able to modify a great variety of biomolecules by chlorination and/or oxidation. In this paper the reactions of lipids (preferentially unsaturated fatty acids and cholesterol) with either reagent HOCl or HOCl generated by the MPO-hydrogen peroxide-chloride system are reviewed. One of the major issues has been whether the reaction of HOCl with lipids of low density lipoprotein (LDL) yields predominantly chlorohydrins or lipid hydroperoxides. Electrospray mass spectrometry provided direct evidence that chlorohydrins rather than peroxides are the major products of HOCl- or MPO-treated LDL phosphatidylcholines. Nevertheless lipid peroxidation is a possible alternative reaction of HOCl with polyunsaturated fatty acids if an additional radical source such as pre-formed lipid hydroperoxides is available. In phospholipids carrying a primary amino group such as phosphatidylethanolamine chloramines are the preferred products compared to chlorohydrins. Cholesterol can be converted by HOCl to great variety of oxysterols besides three isomers of chlorohydrins. For the situation in vivo it appears that the type of reaction occurring between HOCl and lipids would very much depend on the circumstances, e.g. the pH and the presence of radical initiators. The biological effects of lipid chlorohydrins are not yet well understood. It has been shown that chlorohydrins of both unsaturated fatty acids as well as of cholesterol may cause lysis of target cells, possibly by disruption of membrane structures.     

Evaluation of the usefulness of various PCR method variations and nucleic acid hybridization for CMV infection in immunosuppressed patients

In diagnosis of CMV infection various laboratory methods are used. The methods based on detection of viral nucleic acids have been introduced routinely in many laboratories. The aim of this study was to compare nucleic acid hybridisation method and various variants of PCR methods with respect to their ability to detect CMV DNA. The studied material comprised 60 blood samples from 19 patients including 13 renal transplant recipients and 6 with acute leukaemia. The samples were subjected to hybridisation (Murex Hybrid Capture System CMV DNA) and PCR carried out in 3 variants: with one pair of primers (single PCR), nested PCR and Digene SHARP System with detection of PCR product using a genetic probe in ELISA system. The sensitivity of the variants ranged from 10(0) particles of viral DNA in nested PCR to 10(2) in single PCR. The producer claimed the sensitivity of the hybridisation test to be 3 x 10(5) and it seems to be sufficient for detection of CMV infection. The obtained results show that sensitivity of hybridisation was comparable to that of single PCR and the possibility of obtaining quantitative results makes it superior, on efficacy of antiviral therapy, especially in monitoring CMV infection in immunossuppressed patients and in following the efficacy of antiviral treatment.     

Determination of specific immunoglobin G avidity in serodiagnosis of toxoplasmosis

A total of 37 serum samples from pregnant women were examined for the avidity of toxoplasma IgG antibodies using commercially available enzyme immunoassay (Vidas Toxo IgG Avidity, bio Merieux). Low values of the avidity index were found for 4 samples and borderline ones were found in further 3 cases. Out of 19 serum samples showing the presence of IgG and IgM toxoplasma antibodies, 13 had high avidity index, including 9 samples from women in the first trimester of pregnancy. The results of examinations in these 9 cases allowed to exclude the possibility that toxoplasma infection was acquired during gestation. It is to be concluded that determinations of antitoxoplasma IgG avidity should become routine in every pregnant woman positive in the test for specific IgM antibody.     

Bactericidal properties of neutrophils from peripheral blood (PMN)of patients with Lyme borreliosis

In recent years in Poland, the interest has increased in studies about tick borne diseases, mainly Lyme borreliosis. Immune response and genotype of pathogen play an important role in the course of this disease. Phagocytic cells, especially PMN are dominant in defence mechanisms against bacterial infections. The main feature of PMN is their ability to destroy pathogenic microorganisms by phagocytosis. The aim of this study was to estimate the phagocytic activity of PMN connected with intracellular respiratory burst in patients with Lyme borreliosis. The PMN activity tests completed were: phagocytosis, spontaneous and reduced of nitrotetralizate blue test (NBT). Decreased phagocytic activity and oxygen metabolism of PMN from patients with borreliosis in comparison with values of controls were found. Normalization of these parameters after treatment was observed. Changed phagocytic activity connected with intracellular oxygen metabolism during the course of therapy was the main observation. Depression of phagocytic activity of PMN connected with oxygen metabolism can influence defence reactions in patients with Lyme borreliosis. It is suggested that changes observed are acquired and associated with Borrelia burgdorferi presence.     

The effect of selected factors on the course of wound healing after surgery for laryngeal neoplasms

Finding of the a etiologic factors and participation of bacteria flora in wound healing in laryngeal cancer treatment was the purpose of our study. Investigations were performed in 27 patients. Swabs were taken from the postoperative wounds. Detailed identifications of the bacteria flora and antibiotic susceptibility of bacteria were performed. Wound healing was estimated according to extension of the carcinoma, applied antibiotics, state of the oral cavity, the kind of bacteriological flora isolated after surgical treatment from postoperative wounds. It was found that wound healing depended on the extension of carcinoma, as well as, type of isolated bacteria and antibiotic therapy used. The proper healing of postoperative wounds was not dependent on the state of the oral cavity and the dentition. The main cause of postoperative complication of wounds was methicillin-resistant Staphylococcus aureus (MRSA).     

Sesquiterpenoids from roots of Taraxacum laevigatum and Taraxacum disseminatum

Chromatographic separation of ethanolic root extracts of Taraxacum laevigatum and Taraxacum disseminatum afforded a total of eight germacrane- and eudesmane-type sesquiterpenoids. including new compounds, 1beta,3beta,6alpha-trihydroxy-4alpha( 15)-dihydrocostic acid methyl ester and its 1-O-beta-glucopyranoside. Their structures were established by spectroscopic analyses. In addition, the structure of 4alpha(15), 11beta(13)-tetrahydroridentin B-1-O-beta-glucopyranoside was elucidated by extensive NMR studies.     

Sesquiterpenoids and phenolics from Crepis mollis

From the roots of Crepis mollis, one new and two known guaianolides were isolated together with eight known guaianolide glycosides, one known germacranolide glycoside and two known phenylpropanoids. The structure and relative configuration of the new compound were established as 9alpha-hydroxy-4beta, 15, 11beta, 13-tetrahydro-dehydrozaluzanin C by spectral methods.     

Molecular modelling of the interaction of carbocyclic analogues of netropsin and distamycin with d(CGCGAATTCGCG)2

A molecular mechanics and molecular dynamics approach was used to examine the structure of complexes formed between the d(CGCGAATTCGCG)2 duplex and netropsin, distamycin, and four carbocyclic analogues of netropsin and distamycin (1-4). The resulting structures of the ligand-DNA model complexes and their energetics were examined. It is predicted that the compounds 1-4 should have a decreased affinity for the minor groove of AT-rich regions in comparison to netropsin and distamycin. From the energetic analysis it appears that van der Waals and electrostatic interactions are more important than specific hydrogen bonds in stabilizing the ligand-duplex complexes. We predict that compounds 1 and 2 are effectively isohelical with the DNA minor groove. The superior DNA-binding afforded by 1 and 2 in comparison to 3 and 4 results from their more effective penetration into the minor groove and smaller perturbation of molecular structure upon complex formation.     

Connexin43 in porcine myocardium and non-pregnant myometrium

It is generally accepted that connexin43 (Cx43) is a major constituent of heart and myometrial gap junctions. However, the presence of Cx43 gap junctions in non-pregnant myometrium is still poorly documented. Tissue sections of porcine heart and non-pregnant uterus and myometrial smooth muscle cell cultures were immunostained with monoclonal antibody against Cx43. In the heart, intensive immunostaining was confined to the intercalated discs as previously reported. In the non-pregnant uterus, punctuate immunostaining of Cx43 was seen throughout the myometrium along cell interfaces between myocytes. The expression of Cx43 was sustained in cultured smooth muscle cells isolated from non-pregnant myometrium. Western blotting has detected single isoform of Cx43 in both, cardiac and myometrial tissues. The electrophoretic mobility of porcine heart Cx43 was similar to that of myometrial isoform but different from the pattern of mobility of Cx43 of the rat heart. Hence, porcine myometrium may provide attractive model for studying cellular mechanisms triggering expression of gap junction protein in normal (non-pregnant) uterus.