J Biol Chem, 2003 Jun 13, 278(24), 21517 - 25 Epub 2003 Apr 03. Inhibition of proteasome activity induces concerted expression of proteasome genes and de novo formation of Mammalian proteasomes; Meiners S et al.; The 26 S proteasome is a high molecular mass proteinase complex that is built by at least 32 different protein subunits . Such protease complexes in bacteria and yeast are systems that undergo a highly sophisticated network of gene expression regulation . However, regulation of mammalian proteasome gene expression has been neglected so far as a possible control mechanism for the amount of proteasomes in the cell . Here, we show that treatment of cells with proteasome inhibitors and the concomitant impairment of proteasomal enzyme activity induce a transient and concerted up-regulation of all mammalian 26 S proteasome subunit mRNAs . Proteasome inhibition in combination with inhibition of transcription revealed that the observed up-regulation is mediated by coordinated transcriptional activation of the proteasome genes and not by post-transcriptional events . Our experiments also demonstrate that inhibitor-induced proteasome gene activation results in enhanced de novo protein synthesis of all subunits and in increased de novo formation of proteasomes . This phenomenon is accompanied by enhanced expression of the proteasome maturation factor POMP . Thus, our experiments present the first evidence that the amount of proteasomes in mammalia is regulated at the transcriptional level and that there exists an autoregulatory feedback mechanism that allows the compensation of reduced proteasome activity. Appl Environ Microbiol, 2003 Apr, 69(4), 2201 - 8 Interstrain inhibition in the sweet potato pathogen Streptomyces ipomoeae: purification and characterization of a highly specific bacteriocin and cloning of its structural gene; Zhang X et al.; Strains of the sweet potato soil rot pathogen Streptomyces ipomoeae had previously been divided into three groups based on their ability to inhibit one another during pairwise cocultivation . While group I strains are not antagonistic to members of the other groups, group II and group III strains produce separate substances that are inhibitory to strains outside their respective cognate groups . Here, we purified the group III inhibitory substance from the culture supernatant of a representative strain and found that it consists of a single 10-kDa cationic protein which is bacteriolytic for S . ipomoeae group I and II strains but which showed no inhibitory function against other streptomycetes or other bacterial genera tested . The structural gene for the inhibitor was cloned from a chromosomal library of the producing strain, and while the gene sequence revealed that the inhibitor is initially made in a larger precursor form, the deduced mature protein showed no significant homology to other known proteins . Our results demonstrate that S . ipomoeae group III inhibitory activity is manifested in the form of a highly specific, potentially novel bacteriocin, which we have designated ipomicin. Appl Environ Microbiol, 2003 Apr, 69(4), 1980 - 9 Effect of signal compounds and incubation conditions on the culturability of freshwater bacterioplankton; Bruns A et al.; The effect of signal compounds and of different incubation conditions on the culturability (i.e., the fraction of all cells capable of growth) of natural bacterioplankton from the eutrophic lake Zwischenahner Meer was investigated over a period of 20 months . Numbers of growing cells were determined by the most-probable-number technique in liquid media containing low concentrations (10 micro M) of the signal compounds N-(oxohexanoyl)-DL-homoserine lactone, N-(butyryl)-DL-homoserine lactone, cyclic AMP (cAMP), or ATP . cAMP was the most effective signal compound, leading to significantly increased cultivation efficiencies of up to 10% of the total bacterial counts . Microautoradiography with {2,8-(3)H}cAMP, combined with fluorescence in situ hybridization, demonstrated that cAMP was taken up by 18% of all cells . The bacterial cAMP uptake systems had a very low K(m) value of </=1 nM . Analysis of the cultured bacteria by 16S rRNA gene fingerprinting showed that different bacterial phylotypes were recovered in the presence and in the absence of cAMP . Consequently, the addition of cAMP caused a stimulation of otherwise nonculturable bacteria . Phylogenetically different bacteria were also recovered at different temperatures and oxygen partial pressures . Throughout the study period, mainly members of the beta-subclass of the Proteobacteria were cultivated . In addition, some members of the Actinomycetales were enriched . Quantification by culture-independent fluorescence in situ hybridization demonstrated that beta-Proteobacteria and Actinomycetales also dominated the natural bacterioplankton assemblage . Sequence comparison revealed that two members of the Actinomycetales which reached high numbers in the natural bacterioplankton assemblage could actually be enriched by our cultivation approach. Appl Environ Microbiol, 2003 Apr, 69(4), 1959 - 66 5-keto-D-gluconate production is catalyzed by a quinoprotein glycerol dehydrogenase, major polyol dehydrogenase, in gluconobacter species; Matsushita K et al.; Acetic acid bacteria, especially Gluconobacter species, have been known to catalyze the extensive oxidation of sugar alcohols (polyols) such as D-mannitol, glycerol, D-sorbitol, and so on . Gluconobacter species also oxidize sugars and sugar acids and uniquely accumulate two different keto-D-gluconates, 2-keto-D-gluconate and 5-keto-D-gluconate, in the culture medium by the oxidation of D-gluconate . However, there are still many controversies regarding their enzyme systems, especially on D-sorbitol and also D-gluconate oxidations . Recently, pyrroloquinoline quinone-dependent quinoprotein D-arabitol dehydrogenase and D-sorbitol dehydrogenase have been purified from G . suboxydans, both of which have similar and broad substrate specificity towards several different polyols . In this study, both quinoproteins were shown to be identical based on their immuno-cross-reactivity and also on gene disruption and were suggested to be the same as the previously isolated glycerol dehydrogenase (EC 1.1.99.22) . Thus, glycerol dehydrogenase is the major polyol dehydrogenase involved in the oxidation of almost all sugar alcohols in Gluconobacter sp . In addition, the so-called quinoprotein glycerol dehydrogenase was also uniquely shown to oxidize D-gluconate, which was completely different from flavoprotein D-gluconate dehydrogenase (EC 1.1.99.3), which is involved in the production of 2-keto-D-gluconate . The gene disruption experiment and the reconstitution system of the purified enzyme in this study clearly showed that the production of 5-keto-D-gluconate in G . suboxydans is solely dependent on the quinoprotein glycerol dehydrogenase. Appl Environ Microbiol, 2003 Apr, 69(4), 1884 - 9 Identification of Streptomyces coelicolor proteins that are differentially expressed in the presence of plant material; Langlois P et al.; Streptomyces coelicolor and Lemna minor were used as a model to study the modulation of bacterial gene expression during plant-streptomycete interactions . S . coelicolor was grown in minimal medium with and without L . minor fronds . Bacterial proteomes were analyzed by two-dimensional gel electrophoresis, and a comparison of the two culture conditions resulted in identification of 31 proteins that were induced or repressed by the presence of plant material . One-half of these proteins were identified by peptide mass fingerprinting by using matrix-assisted laser desorption ionization-time of flight mass spectrometry . The induced proteins were involved in energetic metabolism (glycolysis, pentose phosphate pathway, oxidative phosphorylation), protein synthesis, degradation of amino acids, alkenes, or cellulose, tellurite resistance, and growth under general physiological or oxidative stress conditions . The repressed proteins were proteins synthesized under starvation stress conditions . These results suggest that root exudates provide additional carbon sources to the bacteria and that physiological adaptations are required for efficient bacterial growth in the presence of plants. Genomics, 2003 Apr, 81(4), 356 - 60 Reversible gene inactivation in the mouse; Mallo M et al.; Gene-inactivation techniques in the mouse have become an essential tool for modern biomedical research . Both ubiquitous and tissue-specific inactivation are possible with current approaches, and recent developments facilitate a temporal control of the inactivation process . However, one of the limitations of current procedures is that inactivation is irreversible . We have produced complete and reversible inactivation of the Hoxa2 gene in the mouse using the control elements of the tetracycline-resistance operon . We show that a Hoxa2 allele containing tetracycline operator (tetO) sequences is susceptible to controlled regulation by tTS, a chimeric molecule containing the tetracycline repressor and a transcriptional repressing domain . This inhibition was specific to the tetO-modified allele, did not affect neighboring genes, and was reversible by administration of doxycycline to the pregnant female . This procedure allows the production of gene inactivation that is complete, is reversible, and can be controlled at the spatial and temporal levels. Bioresour Technol, 2003 Aug, 89(1), 57 - 62 The effect of temperature fluctuations on psychrophilic anaerobic sequencing batch reactors treating swine manure; Masse DI et al.; Under northern climatic conditions, a temporary decrease in the temperature of anaerobic reactors treating swine manure is likely to happen at the farm . The objective of this study was to evaluate the impact of temperature fluctuations, between 10 and 20 degrees C, on the stability and performance of psychrophilic anaerobic sequencing batch reactors (ASBRs) treating swine manure . Methane yield decreased from 0.266+/-0.014 l/g of total chemical oxygen demand (TCOD) fed to the ASBRs at 20 degrees C to 0.218+/-0.022 and 0.080+/-0.002 l/g TCOD (fed) at 15 and 10 degrees C, respectively . Soluble chemical oxygen demand (SCOD) reduction decreased from 94.2+/-1.1% at 20 degrees C to 78.8+/-3.0% at 15 degrees C and 60.4+/-6.4% at 10 degrees C . Total COD removal also tended to decrease as temperature was lowered, but difference between operating temperatures was not as pronounced . A lower methanogenic activity in the ASBRs operated at 10 degrees C probably favoured quiescent conditions during the settling period, thereby increasing physical removal of the TCOD through sedimentation of the solids with the biomass . When the operating temperature was increased back to 15 and 20 degrees C, methane yield and SCOD reduction improved, but reactor performance remained significantly (P<0.05) lower than that achieved before the cycles at 10 degrees C . Results from this experiment nevertheless suggested that fluctuation in the operating temperature of psychrophilic ASBRs should only have temporary effects on the performance and stability of the process. Toxicol Lett, 2003 Apr 11, 140-141, 391 - 401 Long-term effects of heavy metals and microelements on nematode assemblage; Bakonyi G et al.; Effects of Cd, Cr, Se and Zn at a maximum rate of 270 mg kg(-1) were studied on a nematode assemblage after 6-10 years of application . Winter wheat, sunflower, sorrel, barley and rape were grown on the experimental field . Cd had a moderate effect on nematodes in spite of the fact that this element significantly decreased plant biomass . Cr was harmful to plants only in the first year of the study . However, Cr decreased Aporcelaimellus density and maturity index, increased Pratylenchus density and bacterial-fungal ratio, changed the c-p structure and feeding type composition . Se proved to be very toxic at a concentration of 11 mg kg(-1) (NH(4)-acetate+EDTA soluble form) . Some advantageous effects of Zn were found in the first year . These disappeared later on . Remarkable between-year fluctuations of the nematode assemblage composition were observed. Environ Int, 2003 Jun, 29(2-3), 213 - 39 Ecological effects of particulate matter; Grantz DA et al.; Atmospheric particulate matter (PM) is a heterogeneous material . Though regulated as un-speciated mass, it exerts most effects on vegetation and ecosystems by virtue of the mass loading of its chemical constituents . As this varies temporally and spatially, prediction of regional impacts remains difficult . Deposition of PM to vegetated surfaces depends on the size distribution of the particles and, to a lesser extent, on the chemistry . However, chemical loading of an ecosystem may be determined by the size distribution as different constituents dominate different size fractions . Coating with dust may cause abrasion and radiative heating, and may reduce the photosynthetically active photon flux reaching the photosynthetic tissues . Acidic and alkaline materials may cause leaf surface injury while other materials may be taken up across the cuticle . A more likely route for metabolic uptake and impact on vegetation and ecosystems is through the rhizosphere . PM deposited directly to the soil can influence nutrient cycling, especially that of nitrogen, through its effects on the rhizosphere bacteria and fungi . Alkaline cation and aluminum availability are dependent upon the pH of the soil that may be altered dramatically by deposition of various classes of PM . A regional effect of PM on ecosystems is linked to climate change . Increased PM may reduce radiation interception by plant canopies and may reduce precipitation through a variety of physical effects . At the present time, evidence does not support large regional threats due to un-speciated PM, though site-specific and constituent-specific effects can be readily identified . Interactions of PM with other pollutants and with components of climate change remain important areas of research in assessment of challenges to ecosystem stability. J Comput Biol, 2003, 10(1), 83 - 93 Temporal changes in phosphoglycerate kinase coding sequences: a quantitative measure; Chattopadhyay S et al.; The ratio of the average of the square of the number of the nucleotides to that of the random sequence of the same strand bias is proposed as a quantitative measure of evolution in some coding DNA sequences . Applying this measure to the phosphoglycerate kinase gene we observe a monotonic rise of the ratio with evolution . We present an interpretation of this data on some bacteria. J Vet Med B Infect Dis Vet Public Health, 2003 Mar, 50(2), 90 - 4 Differentiation of twelve Actinobacillus pleuropneumoniae serotypes by outer membrane lipoprotein gene-based restriction fragment length polymorphism; Cho WS et al.; Twelve Actinobacillus pleuropneumoniae serotypes were differentiated by restriction fragment length polymorphism (RFLP) of polymerase chain reaction (PCR)-amplified fragments from the outer membrane lipoprotein (omlA) gene . All 12 reference serotypes and 80 field isolates produced the expected 950-base pair (bp) fragment of the omlA gene by PCR . Combining the RFLP patterns obtained with SfaNI, Bst71I, AluI, NciI, nine distinct patterns were observed in the 12 serotype reference strains . The PCR-based RFLP analysis of omlA genes allows differentiation among the 12 serotypes, with the exception of group 1 (serotypes 1, 9 and 11), and group 2 (serotypes 2 and 8) . When the PCR products from the 70 field isolates were subjected to RFLP analysis, 68 showed the same RFLP patterns as their respective serotype reference strain . Two isolates that could not be typed had the same RFLP patterns as those of serotype 5 . These results suggest that PCR-based RFLP analysis of the omlA genes may be of value in differentiating among 12 A . pleuropneumoniae serotypes. Russ J Immunol, 2002 Oct, 7(3), 219 - 28 New approaches to immunoprevention and immunotherapy of neoplasms; Glushkov AN; The problems in the creation of vaccines for the prevention and treatment of neoplasms and the perspectives of their application are discussed . The positive and negative properties of the conjugates of the chemical carcinogens with proteins, DNA and enterotoxins, as well as the properties of the monoclonal anti-idiotype antibodies to the carcinogens are being analyzed . We are describing here a new technology of the anti-carcinogen vaccine production that we have developed . We suggest using transgenic plants and bacteria carrying the Fv-genes of antibodies for the passive immunoprevention of cancer diseases . We assume that lymphoproliferative neoplasms mainly arise from the clones carrying membrane receptors for the carcinogens . Therefor, here it is proposed to use the xenogenic anticarcinogens antibodies for the active immunotherapy of leukemia and other lymphoproliferative neoplasms. Indian J Med Res, 2002 Oct, 116, 140 - 4 Effect of rifampicin & isoniazid on cytochrome P-450 in mycobacteria; Ramachandran G et al.; BACKGROUND & OBJECTIVES: Rifampicin and isoniazid are the most important first line drugs used in the treatment of tuberculosis . These drugs are also used in combination with other medications to treat co-infections . It, therefore, becomes important to study the effect of these drugs on the drug metabolizing system, namely, cytochrome P-450, not only in the host but also in the bacteria . We report the effect of rifampicin and isoniazid on the cytochrome P-450 activity in Mycobacterium smegmatis and M . tuberculosis H37Rv . METHODS: Subinhibitory concentrations of rifampicin and isoniazid were added to the organisms after they had attained the growth phase and cytochrome P-450 activity was estimated in the membranous fractions of the bacteria at different time points . RESULTS: Rifampicin was able to significantly enhance cytochrome P-450 in both M . smegmatis and M . tuberculosis H37Rv . Isoniazid was found to inhibit cytochrome P-450 in M . tuberculosis H37Rv, while there seemed to be no effect in M . smegmatis . INTERPRETATION & CONCLUSION: We report here the effect of rifampicin and isoniazid on mycobacterial cytochrome P-450 . These findings are similar to those found in eukaryotic organisms . The role of mycobacterial cytochrome P-450 in the metabolism of drugs within the bacteria needs to be elucidated. Sheng Wu Gong Cheng Xue Bao, 2002 Nov, 18(6), 703 - 8 {Catalysis of lyase-isomerase PecE/PecF for several apophycobiliproteins}; Zhu JP et al.; Phycoerythrocyanin(PEC) lyase-isomerase PecE/PecF from Mastigocladus laminosus is the specific enzyme for biosynthesis of PEC alpha-subunit(alpha-PEC) . In this work, the specificity of PecE/PecF on substrate apoproteins was reported . PecE/PecF could catalyse the reconstitution of phycocyanobilin(PCB) with apoproteins of alpha-PEC from two different subspecies of Mastigocladus laminosus, as well the site-directed mutated apoprotein of alpha-PEC with Trp at 128 to Phe in vitro, but could not catalyse the reconstitution of PCB with apoprotein of phycocyanin alpha-subunit(alpha-CPC) from Mastigocladus laminosus . The surfactant Triton X-100 had no effect for the reconstitution of alpha-PEC, while it could improve the reconstitution of PCB with apoprotein of alpha-CPC. J Eukaryot Microbiol, 2003 Jan-Feb, 50(1), 43 - 8 Trichomonas vaginalis: in vitro attachment and internalization of HIV-1 and HIV-1-infected lymphocytes; Rendon-Maldonado J et al.; Sexually transmitted diseases (STDs) caused by bacteria and protozoa play an important role in the epidemiology of human immunodeficiency virus (HIV-1) infection . Human trichomoniasis, produced by the protozoan parasite Trichomonas vaginalis, is one of the most common STDs, and is a cause of mucosal lesions in the urogenital tract, which may increase the risk for HIV infection . However, there are no reports concerning the outcome of in vitro interactions between HIV particles and trichomonads . Therefore, we incubated T . vaginalis with three subtypes of HIV-1 (A, B, and D), as well as with HIV-1-infected lymphocytes, and analyzed the interactions with immunofluorescence microscopy and transmission electron microscopy . Our results demonstrated that HIV-1 particles attach and are incorporated into T . vaginalis through endocytic vesicles and are degraded within cytoplasmic vacuoles in approximately 48 h . There was no ultrastructural evidence of HIV-1 replication in trichomonads . These results demonstrated that trichomonads may internalize and harbor HIV-1 particles for short periods of time . In addition, under in vitro conditions, T . vaginalis ingests and digests HIV-1-infected lymphocytes. Adv Biochem Eng Biotechnol, 2003, 78, 75 - 96 The effect of plants on the degradation and toxicity of petroleum contaminants in soil: a field assessment; Banks MK et al.; A field project located at the US Naval Base at Port Hueneme, California was designed to evaluate changes in contaminant concentrations and toxicity during phytoremediation . Vegetated plots were established in petroleum (diesel and heavy oil) contaminated soil and were evaluated over a two-year period . Plant species were chosen based on initial germination studies and included native California grasses . The toxicity of the impacted soil in vegetated and unvegetated plots was evaluated using Microtox, earthworm, and seed germination assays . The reduction of toxicity was affected more by contaminant aging than the establishment of plants . However, total petroleum hydrocarbon concentrations were lower by the end of the study in the vegetated plots when compared to the unvegetated soil . Although phytoremediation is an effective approach for cleaning-up of petroleum contaminated soil, a long-term management plan is required for significant reductions in contaminant concentrations. Quintessence Int, 2003 Jan, 34(1), 61 - 70 Identification of hierarchical factors to guide clinical decision making for successful long-term pulp capping; Murray PE et al.; OBJECTIVE: Clinicians have few quantitative studies that rank the in vivo pulp capping effects of commonly used restorative materials . These factors were investigated to provide guidance to clinicians . METHOD AND MATERIALS: One hundred sixty-one standardized pulp-exposed cavities were prepared in nonhuman primate teeth . Exposed pulps were capped with calcium hydroxide, resin-modified glass-ionomer cements, and resin composites . Teeth were collected from 7 to 720 days to observe a full range of responses . Pulpal reactions were categorized according to the standards set by the International Standards Organization . Bacteria were detected with McKay's stain . RESULTS: The incidence of bacterial microleakage was 19.7% with resin composite, 21.1% with resin-modified glass-ionomer cement, and 47.0% with calcium hydroxide . The severity of pulpal inflammation increased with the presence of bacteria or tunnel defects . The severity of pulpal inflammation prevented dentinal bridge formation at varying levels: slight for resin composite and resin-modified glass-ionomer cement, and severe with calcium hydroxide . The incidence of severe inflammation or pulpal necrosis was 7.9% with resin composite, 10.6% with calcium hydroxide, and 10.5% with resin-modified glass-ionomer cement . Other variables, such as pulpal exposure width and tertiary dentin formation, were not highly correlated to pulpal inflammation . CONCLUSION: Pulp capping with resin composite provided the lowest incidence of bacterial microleakage, the lowest levels of pulpal inflammation, and the lowest incidence of necrosis. J Physiol Pharmacol, 2003 Mar, 54(1), 33 - 51 Triple eradication therapy counteracts functional impairment associated with Helicobacter pylori infection in Mongolian gerbils; Brzozowski T et al.; Gastric Helicobacter pylori (Hp) infection in Mongolian gerbils is an established experimental model of gastric carcinogenesis resulting from the long-term Hp infection but functional aspects accompanying this Hp-induced progression from gastritis to the cancer, especially changes in gastric acid secretion, gastric blood flow (GBF) and gastrin-somatostatin link have been little studied . It is unclear whether Hp eradication therapy alters the functional and the histopathological changes in this animal model of Hp-infection . We examined the effects of intragastric (i.g.) inoculation of Mongolian gerbils with Hp strain (cagA+ vacA+, 5 x 10(6) CFU/ml) that had been isolated from a patient with gastric ulcer as compared to those induced by vehicle (saline) in gerbils with or without gastric fistula (GF) at 1.2, 4, 6, 9, 12 and 30 wks upon gastric inoculation with this bacteria . An attempt was made to evaluate the influence of anti-Hp triple therapy with omeprazole, amoxicillin and tinidazol on gastric Hp-infection and Hp-induced functional impairment of the gastric mucosa . Gastric mucosal biopsy specimens were taken for the assessment of the morphological changes and the presence of Hp infection using rapid urease test (CLO-test) and the density of Hp-colonization were assessed by counting of the number of bacterial colonies per plate . Gastric blood flow (GBF) was measured by H2-gas clearance technique and the venous blood and the gastric content were collected for the measurement of plasma gastrin levels and the gastric luminal somatostatin level by radioimmunoassay (RIA) . The Hp in gastric mucosa was detected in all animals by culture and rapid urease test at various periods upon Hp inoculation . Basal gastric acid in non-infected conscious gerbils with GF reached the level of about 28 +/- 4 micromol/h and this was reduced by over 50% immediately upon the Hp-inoculation and persisted for time intervals tested up to 30 wk . Early lesions were seen 4 wks after the Hp-inoculation and consisted of chronic gastritis with thickened gastric mucosal foldings and elongated interfoveolar ridges . Edema and congestion as well as significant mucosal inflammatory infiltration with lymphoid infiltrate in lamina propria of the mucosa occurred in all infected gerbils . Adenomatous hyperplasia with cellular atypia was observed at 12 wk upon Hp-inoculation together with increased mitotic activity and numerous apoptotic bodies formation, while lamina propria was reduced leaving dilated atypical gastric gland situated "back-to-back" . This glandular atypia failed to show lamina propria or submucosa infiltration corresponding to gastric intraepithelial neoplasia . The GBF in Hp-infected gerbils was significantly lower, and a 6-7 fold increase in plasma gastrin levels combined with a significant fall in gastric luminal somatostatin contents observed at all tested periods as compared to vehicle-controls and these effects were counteracted by anti-Hp triple therapy . We conclude that: 1) . Hp-infection in Mongolian gerbils in early stages before adenocarcinoma formation results in the development of typical functional and pathological changes such as suppression of gastric secretion and impairment of both, gastric mucosal microcirculation and gastrin-somatostatin link, and 2) . this deleterious influence of Hp on gastric morphology and gastric functions is greatly attenuated in gerbils treated with Hp-eradication therapy. Gastric Cancer, 2003, 6(1), 1 - 7 Helicobacter pylori infection and gastric carcinogenesis in animal models; Tatematsu M et al.; The effects of helicobacter pylori infection on gastric disorders have been proven by many epidemiological and experimental studies . To explore the relationships between h . Pylori infection and gastric carcinogenesis, many factors, including host responses, environmental status, and the virulence factors of the bacteria should be taken into account . Mongolian gerbils ( meriones unguiculatus) can be easily infected with h . Pylori, and provide an excellent in-vivo experimental model to clarify the role of h . Pylori in active gastritis, peptic ulcers, intestinal metaplasia, and gastric carcinoma . Studies have revealed that h . Pylori infection markedly enhances all histological types of gastric cancers in gerbils treated with a chemical carcinogen . Eradication reduced the enhancing effect of h . Pylori on gastric carcinogenesis, whereas a high-salt diet synergistically enhanced the effect of h . Pylori . Various factors involving inflammation, cell proliferation, and cell differentiation could be examined with this experimental model to help elucidate this mechanisms of gastric carcinogenesis. Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 2003 Apr, 35(4), 311 - 6 Targeting strategies in cancer gene therapy; Wang JH et al.; Targeting to the tumor tissues can improve the therapeutic effect of gene transfer by preventing damage of healthy tissues and decreasing the risk of germ line transduction . Although targeting seems not important for intratumoral gene delivery, it becomes crucial when systemic gene transfer is performed . Targeted gene therapy of malignancies can be achieved through targeted gene delivery or targeted gene transcription . Recent advances in targeted delivery include the successful use of bifunctional crosslinkers to target adenoviral and retroviral vectors, inserting short targeting peptides and larger polypeptide-binding domains into the coat proteins of a number of different viral vectors, and replication-competent vectors which have been shown to be promise as anti-cancer agents . Some other non-viral therapeutic agents, including receptor-mediated DNA or liposome-DNA complex, and bacteria vehicles have also been developed . Some of these delivery systems are currently in clinical trials . For targeted and regulable gene transcription, tissue or tumor specific promoters and some manual regulatory systems are used to regulate therapeutic gene expression . Antisense oligonucleotides, some ribozyme and DNAzyme molecules are developed to inactivate genes that are essential to the development of many tumors. Plant Cell, 2003 Apr, 15(4), 845 - 53 Increased lysine synthesis coupled with a knockout of its catabolism synergistically boosts lysine content and also transregulates the metabolism of other amino acids in Arabidopsis seeds; Zhu X et al.; To elucidate the relative significance of Lys synthesis and catabolism in determining Lys level in plant seeds, we expressed a bacterial feedback-insensitive dihydrodipicolinate synthase (DHPS) in a seed-specific manner in wild-type Arabidopsis as well as in an Arabidopsis knockout mutant in the Lys catabolism pathway . Transgenic plants expressing the bacterial DHPS, or the knockout mutant, contained approximately 12-fold or approximately 5-fold higher levels, respectively, of seed free Lys than wild-type plants . However, the combination of these two traits caused a synergistic approximately 80-fold increase in seed free Lys level . The dramatic increase in free Lys in the knockout mutant expressing the bacterial DHPS was associated with a significant reduction in the levels of Glu and Asp but also with an unexpected increase in the levels of Gln and Asn . This finding suggested a special regulatory interaction between Lys metabolism and amide amino acid metabolism in seeds . Notably, the level of free Met, which competes with Lys for Asp and Glu as precursors, was increased unexpectedly by up to approximately 38-fold in the various transgenic and knockout plants . Together, our results show that Lys catabolism plays a major regulatory role in Lys accumulation in Arabidopsis seeds and reveal novel regulatory networks of seed amino acid metabolism. J Bacteriol, 2003 Apr, 185(8), 2582 - 91 Signal transduction protein P(II) is required for NtcA-regulated gene expression during nitrogen deprivation in the cyanobacterium Synechococcus elongatus strain PCC 7942; Fadi Aldehni M et al.; The transcription factor of the cyclic AMP receptor protein/FNR family, NtcA, and the P(II) signaling protein play central roles in global nitrogen control in cyanobacteria . A dependence on P(II) for NtcA-regulated transcription, however, has not been observed . In the present investigation, we examined alterations in gene expression following nitrogen deprivation in Synechococcus elongatus strain PCC 7942 and specifically the roles of NtcA and P(II) . Global changes in de novo protein synthesis following combined-nitrogen deprivation were visualized by in vivo {(35)S}methionine labeling and two-dimensional polyacrylamide gel electrophoresis analysis . Nearly all proteins whose synthesis responded specifically to combined-nitrogen deprivation in wild-type cells of S . elongatus failed to respond in P(II)- and NtcA-deficient mutants . One of the proteins whose synthesis was down-regulated in a P(II)- and NtcA-dependent manner was RbcS, the small subunit of RubisCO . Quantification of its mRNA revealed that the abundance of the rbcLS transcript following combined-nitrogen deprivation rapidly declined in wild-type cells but not in P(II) and NtcA mutant cells . To investigate further the relationship between P(II) and NtcA, fusions of the promotorless luxAB reporter genes to the NtcA-regulated glnB gene were constructed and these constructs were used to transform wild-type cells and P(II)(-) and NtcA(-) mutants . Determination of bioluminescence under different growth conditions showed that NtcA represses gene expression in the presence of ammonium in a P(II)-independent manner . By contrast, NtcA-dependent activation of glnB expression following combined-nitrogen deprivation was impaired in the absence of P(II) . Together, these results suggest that under conditions of combined-nitrogen deprivation, the regulation of NtcA-dependent gene expression requires the P(II) signal transduction protein. Sci Total Environ, 2003 Apr 15, 305(1-3), 169 - 76 Bioaerosol characteristics in hospital clean rooms; Li CS et al.; Bioaerosol characteristics were evaluated in hospital clean rooms with different class levels . For total particles, an airborne particle counter was used to determine the particle size ranges (0.1, 0.2, 0.3, 0.5, 1 and 5 microm) for air inlets and patient beds . An Andersen 1-STG sampler was used for bacterial and fungal collection . For aerosol characteristics, it was found that some air inlet particle levels were higher than 100000/foot(3) in class 100000 clean rooms . In addition, it was clearly demonstrated that particle concentrations in patient beds were much higher than those for air inlets . Human activity might play a role in these particle concentration differences . Moreover, it was demonstrated that bacterial and fungal concentrations ranged from 1 to 423 and from 0 to 319 CFU/m(3), respectively . For class 100 clean rooms, no particles were ever found . In addition, bacterial concentrations were found to be in the range of 0-32 CFU/m(3) and there were no fungal aerosols . For operating rooms of class 10000, some of the particle levels observed were higher than 10000/foot(3) . Furthermore, the average level of bacterial aerosols was 88 with a range of 13-336 CFU/m(3) . In addition, fungal levels ranged from 0 to 51 with a mean value of 4 CFU/m(3) . It was indicated that bacterial levels were higher than fungal ones, which might be related to human sources . Moreover, there were weak relationships among class level, particle concentration and bioaerosol levels. FEMS Microbiol Lett, 2003 Mar 28, 220(2), 271 - 5 Activation of the complement cascade by Bordetella pertussis; Barnes MG et al.; Bordetella pertussis must survive the defenses of the human respiratory tract including the complement system . The BrkA (Bordetella resistance to killing) protein prevents killing by the antibody-dependent classical pathway . In this study, the ability of B . pertussis to activate the human complement cascade by other pathways was examined . B . pertussis was not killed in serum depleted of C2, however serum depleted for factor B killed B . pertussis as efficiently as intact serum, suggesting complement activation occurred exclusively by the classical pathway . B . pertussis was not killed by serum depleted of antibody, suggesting the bacteria fail to activate the antibody-independent branches of the classical pathway, including the mannose binding lectin pathway . Mutants lacking the terminal trisaccharide of lipopolysaccharide retained the complement-resistant phenotype, suggesting this structure does not influence activation of complement. Int J Parasitol, 2003 Mar, 33(3), 257 - 67 Programmed cell death in trypanosomatids and other unicellular organisms; Debrabant A et al.; In multicellular organisms, cellular growth and development can be controlled by programmed cell death (PCD), which is defined by a sequence of regulated events . However, PCD is thought to have evolved not only to regulate growth and development in multicellular organisms but also to have a functional role in the biology of unicellular organisms . In protozoan parasites and in other unicellular organisms, features of PCD similar to those in multicellular organisms have been reported, suggesting some commonality in the PCD pathway between unicellular and multicellular organisms . However, more extensive studies are needed to fully characterise the PCD pathway and to define the factors that control PCD in the unicellular organisms . The understanding of the PCD pathway in unicellular organisms could delineate the evolutionary origin of this pathway . Further characterisation of the PCD pathway in the unicellular parasites could provide information regarding their pathogenesis, which could be exploited to target new drugs to limit their growth and treat the disease they cause. J Am Chem Soc, 2003 Apr 9, 125(14), 4012 - 3 Hybridization-based unquenching of DNA hairpins on au surfaces: prototypical "molecular beacon" biosensors; Du H et al.; There is a keen interest in developing techniques for rapid genetic analysis that do not require labeling of an analyte . Here we demonstrate that fluorophore-tagged DNA hairpins attached to gold films can function as immobilized "molecular beacons" . Two DNA hairpins incorporating portions of the Staphlococcus aureus FemA and mecR methicillin-resistance genes were attached to a gold substrate . Upon exposure to the complement, a approximately 26-fold increase in fluorescence intensity was measured corresponding to a 96 +/- 5% quenching efficiency . Studies with nonspecific DNA indicate that DNA hairpins immobilized on a gold surface retain their ability to bind complementary DNA sequences selectively. J Int Acad Periodontol, 2002 Jul, 4(3), 110 - 4 The future of periodontology: new treatments for a new era; Tonetti MS; Scientific evidence that has been emerging over the last decade has provided the basis for a radical change in the understanding, diagnosis and treatment of periodontal diseases . A new era in periodontal research and practice is being defined along the following lines: Better understanding of disease . Emerging needs . Novel treatment strategies and therapeutics . Choice between multiple therapeutic modalities . The purpose of this manuscript is to critically discuss changes in understanding and to provide a rationale to change daily practice. J Int Acad Periodontol, 2002 Jul, 4(3), 101 - 9 The periodontal infection-systemic disease link: a review of the truth or myth; Paquette DW; Observational studies indicate periodontal infections as a risk factor for systemic conditions like cardiovascular disease and preterm low birth weight . This paper reviews and argues the biological plausibility for a periodontal infection-systemic disease link and reviews the available experimental data from animal models and human intervention trials . Five principal lines of evidence can be used to explain the biological plausibility of a link . First, infection in general has been implicated in the pathogenesis of both atherosclerosis and preterm delivery . Periodontal infection secondly causes transient and low-grade bacteraemias and endotoxaemias in patients . Thirdly, periodontal infection promotes systemic inflammatory and immune responses that may play roles in disease . Periodontal pathogens express specific virulence factors that can affect atherogenic or parturition events . Lastly, periodontal pathogens have also been isolated from non-oral tissues like atheromatous plaques . Experimental data derived from rodent and pig models indicate that infection or bacteraemias with the periodontal pathogen, Porphyromonas gingivalis, can increase atheroma size or reduce litter weights as compared to controls . While human intervention data are lacking for patients at risk for cardiovascular disease, early data indicate that periodontal therapy administered to pregnant mothers with periodontitis can reduce the incidence of preterm low birth weight deliveries . Nevertheless, more and larger intervention trials are needed before we can fully accept periodontal infection as a true risk factor in the causal pathways of cardiovascular disease and preterm low birth weight. J Antibiot (Tokyo), 2003 Jan, 56(1), 24 - 9 Core structure in roselipins essential for eliciting inhibitory activity against diacylglycerol acyltransferase; Tomoda H et al.; Fungal roselipins, discovered as inhibitors of diacylglycerol acyltransferase (DGAT), consist of three parts; highly methylated C20 fatty acid, mannose and arabinitol . Demannosyl and/or dearabinitoyl roselipins were prepared chemically or enzymatically . Demannnosyl roselipins conserved the DGAT inhibitory activity, but the others lost the activity, indicating that the arabinitoyl fatty acid core is essential for eliciting the activity. Genetika, 2003 Feb, 39(2), 244 - 9 {Expression analysis of proteins encoded by genes of the tag7/tagL (PGRP-S,L) family in human peripheral blood cells}; Kibardin AV et al.; Various types of human blood cells were tested for expression of the Tag7/PGRP-SA and TagL/PGRP-L proteins, which belong to the family of proteins possessing the lysozyme-like peptidoglycan recognition protein (PGRP) domain . Expression regulation by several factors was demonstrated. Nat Rev Immunol, 2003 Apr, 3(4), 331 - 41 Anatomical basis of tolerance and immunity to intestinal antigens; Mowat AM; The intestinal immune system has to discriminate between harmful and beneficial antigens . Although strong protective immunity is essential to prevent invasion by pathogens, equivalent responses against dietary proteins or commensal bacteria can lead to chronic disease . These responses are normally prevented by a complex interplay of regulatory mechanisms . This article reviews the unique aspects of the local microenvironment of the intestinal immune system and discuss how these promote the development of regulatory responses that ensure the maintenance of homeostasis in the gut. J Exp Med, 2003 Apr 7, 197(7), 845 - 60 Epub 2003 Mar 31. B cell receptor-independent stimuli trigger immunoglobulin (Ig) class switch recombination and production of IgG autoantibodies by anergic self-reactive B cells; Phan TG et al.; In both humans and animals, immunoglobulin (Ig)G autoantibodies are less frequent but more pathogenic than IgM autoantibodies, suggesting that controls over Ig isotype switching are required to reinforce B cell self-tolerance . We have used gene targeting to produce mice in which hen egg lysozyme (HEL)-specific B cells can switch to all Ig isotypes (SWHEL mice) . When crossed with soluble HEL transgenic (Tg) mice, self-reactive SWHEL B cells became anergic . However, in contrast to anergic B cells from the original nonswitching anti-HEL x soluble HEL double Tg model, self-reactive SWHEL B cells also displayed an immature phenotype, reduced lifespan, and exclusion from the splenic follicle . These differences were not related to their ability to Ig class switch, but instead to competition with non-HEL-binding B cells generated by VH gene replacement in SWHEL mice . When activated in vitro with B cell receptor (BCR)-independent stimuli such as anti-CD40 monoclonal antibody plus interleukin 4 or lipopolysaccharide (LPS), anergic SWHEL double Tg B cells proliferated and produced IgG anti-HEL antibodies as efficiently as naive HEL-binding B cells from SWHEL Ig Tg mice . These results demonstrate that no intrinsic constraints to isotype switching exist in anergic self-reactive B cells . Instead, production of IgG autoantibodies is prevented by separate controls that reduce the likelihood of anergic B cells encountering BCR-independent stimuli . That bacteria-derived LPS could circumvent these controls may explain the well-known association between autoantibody-mediated diseases and episodes of systemic infection. Development, 2003 May, 130(10), 2139 - 48 Isolation of COV1, a gene involved in the regulation of vascular patterning in the stem of Arabidopsis; Parker G et al.; The molecular mechanisms that control the ordered patterning of vascular tissue development in plants are not well understood . Several models propose a two-component system for vascular differentiation . These components include an inducer of vascular tissue development and an inhibitor that prevents the formation of vascular bundles near pre-existing bundles . We have identified two recessive allelic mutants in Arabidopsis, designated continuous vascular ring (cov1), that display a dramatic increase in vascular tissue development in the stem in place of the interfascicular region that normally separates the vascular bundles . The mutant plants exhibited relatively normal vascular patterning in leaves and cotyledons . Analysis of the interaction of cov1 with a known auxin signalling mutant and direct analysis of auxin concentrations suggests that cov1 affects vascular pattering by some mechanism that is independent of auxin . The COV1 protein is predicted to be an integral membrane protein of unknown function, highly conserved between plants and bacteria . In plants, COV1 is likely to be involved in a mechanism that negatively regulates the differentiation of vascular tissue in the stem. Microb Pathog, 2003 Apr, 34(4), 187 - 94 The Legionella pneumophila GacA homolog (LetA) is involved in the regulation of icm virulence genes and is required for intracellular multiplication in Acanthamoeba castellanii; Gal-Mor O et al.; Legionella pneumophila, the causative agent of legionnaires' disease, is a broad-host-range facultative intracellular pathogen . Thus far, 24 genes (icm/dot genes) required for L . pneumophila intracellular growth, have been discovered . In this study, a deletion substitution was constructed in the L . pneumophila homolog of the gacA response regulator (letA) and its involvement in L . pneumophila pathogenicity and icm/dot gene expression was characterized . The letA mutant constructed had no intracellular growth defect when analyzed in HL-60 derived human macrophages, but it was found to be severely attenuated for intracellular growth in the protozoan host Acanthamoeba castellanii . The growth defect in amoebae was fully complemented by introducing the L . pneumophila letA gene on a plasmid . In addition, the LetA regulator was found to be involved in the expression of three icm genes (icmT, icmP and icmR) . The level of expression of the icmT::lacZ and icmR::lacZ fusions was found to be higher, while the level of expression of the icmP::lacZ fusion was found to be lower when analyzed in the letA mutant strain, in comparison to the wild-type strain . We concluded that LetA has a moderate effect on icm/dot gene expression, but it probably plays a major role in the expression of L . pneumophila genes required for intracellular growth in protozoan hosts . A similar host specific phenotype was previously described for the RpoS sigma factor and the type II general secretion system of L . pneumophila . Microb Pathog, 2003 Apr, 34(4), 179 - 86 Differential expression of the invasion-associated locus B (ialB) gene of Bartonella bacilliformis in response to environmental cues; Coleman SA et al.; Bartonella bacilliformis is the causative agent of the biphasic human disease, Oroya fever . During the primary disease phase, up to 100% of the circulating erythrocytes can be parasitized and 80% lysed . During the secondary phase of this disease, bacterial invasion shifts to endothelial cells lining the vasculature . B . bacilliformis is transferred between human hosts by the sandfly, Lutzomyia verrucarum . To investigate the regulation of ialB by environmental cues signaling vector-to-host transmission; nuclease protection assays were performed to compare the amount of ialB mRNA in bacteria subjected to temperature shift, pH change, oxidative stress, or hemin limitation . The amount of ialB mRNA increased by 223-310% in acid-treated samples and decreased by 28-39% in base-treated samples as compared to bacteria kept at pH 7.2 . B . bacilliformis samples showed a 56-63% and 74-80% decrease in ialB mRNA when shifted to 37 degrees C from growth temperatures of 20 and 30 degrees C, respectively . Oxidative stress (1 mM H(2)O(2)) and hemin limitation had no significant effect on mRNA levels . Determination of IalB protein amounts using SDS-PAGE and immunoblotting showed the greatest amounts of IalB under acidic conditions or at 20 degrees C . The least amount of IalB was synthesized under basic conditions or at 37 degrees C . The viability of wild-type B . bacilliformis under the various experimental culture conditions was determined and found not to affect ialB mRNA amounts in these experiments . Finally, we compared the survival of wild-type and ialB mutant B . bacilliformis and found no difference in the viability of these two strains, demonstrating that IalB does not aid bacterial survival under these conditions . Virology, 2003 Mar 15, 307(2), 341 - 57 Expression and characterization of a recombinant novel reverse transcriptase of a porcine endogenous retrovirus; Avidan O et al.; The study of porcine endogenous retroviruses (PERVs) becomes increasingly important due to the potential use of pig cells, tissues, and organs as a source for xenogenic cell therapy and xenotransplantation into humans . Consequently, we have constructed a plasmid that induces in bacteria the synthesis of a soluble and highly active reverse transcriptase (RT) of PERV-B . The purified PERV RT was studied biochemically in comparison with the RT of murine leukemia virus (MLV), because of the high-sequence homology between these two RTs . The data show that in several properties the two enzymes are similar, particularly regarding the monomeric subunit composition of the proteins in solution, the high resistance to deoxynucleoside analogues, and the pattern of RNA cleavage by the ribonuclease H activity (RNase H) of the RTs . However, in several cases there are apparent differences between the two RTs, most notable the divalent cation preference (Mn(+2) versus Mg(+2)) in the DNA polymerase reactions . As already shown for viral PERV RT, the novel recombinant PERV RT exhibits a relatively high resistance to several deoxynucleoside analogue inhibitors, suggesting that they might not be very efficient in inhibiting the replication of PERV virions . Therefore, the availability of large amounts of the recombinant RT can be useful for a wide screening of novel drugs against infectious PERV. Arch Biochem Biophys, 2003 Apr 15, 412(2), 231 - 41 Evolutionary conservation of physical and functional interactions between phospholipase D and actin; Kusner DJ et al.; Phospholipase D (PLD) enzymes from bacteria to mammals exhibit a highly conserved core structure and catalytic mechanism, but whether protein-protein interactions exhibit similar commonality is unknown . Our objective was to determine whether the physical and functional interactions of mammalian PLDs with actin are evolutionarily conserved among bacterial and plant PLDs . Highly purified bacterial and plant PLDs cosedimented with mammalian skeletal muscle alpha-actin, indicating direct interaction with F-actin . The binding of bacterial PLD to G-actin exhibited two affinity states, with dissociation constants of 1.13 pM and 0.58 microM . The effects of actin on the activities of bacterial and plant PLDs were polymerization dependent; monomeric G-actin inhibited PLD activity, whereas polymerized F-actin augmented PLD activity . Actin modulation of bacterial and plant PLDs demonstrated kinetic characteristics, efficacies, and potencies similar to those of human PLD1 . Thus, physical and functional interactions between PLD and actin in PLD family members from bacteria to mammals are highly conserved throughout evolution. Ann Intern Med, 2003 Apr 1, 138(7), 550 - 9 Gene vaccines; Srivastava IK et al.; Gene vaccines are a new approach to immunization and immunotherapy in which, rather than a live or inactivated organism (or a subunit thereof), one or more genes that encode proteins of the pathogen are delivered . The goal of this approach is to generate immunity against diseases for which traditional vaccines and treatments have not worked, to improve vaccines, and to treat chronic diseases . Gene vaccines make use of advances in immunology and molecular biology to more specifically tailor immune responses (cellular or humoral, or both) against selected antigens . They are still under development in research and clinical trials . The mechanisms for inducing cellular (as opposed to humoral) responses against a particular antigen have been elucidated . Gene vaccines provide a means to generate specific cellular responses while still generating antibodies, if desired . In addition, by delivering only the genes that encode the particular proteins against which a protective or therapeutic immune response is desired, the potential limitations and risks of certain other approaches can be avoided . This article describes the rationale for, immunologic mechanisms involved in, and design of gene vaccines under development . Preclinical and clinical studies of these vaccines are discussed for various clinical applications, focusing on infectious diseases. J Int Acad Periodontol, 2001 Apr, 3(2), 31 - 7 A transmission electron microscopy study of supragingival calculus; Galgut PN et al.; In this study the ultrastructure of human dental calculus was investigated using transmission electron microscopy (TEM) . The aim of the study was to observe and clarify the calcification process and the part played by dental plaque in this process . Supragingival calculus from 10 patients was harvested and processed . It was shown that both calculus, and dental plaque within non-mineralised lacunae in calculus, are heterogeneous . Furthermore it was shown that the calculus/dental plaque interface was distinct, although also variable . As calcification occurred apparently independently of dental plaque in some places and when calcification of dental plaque bacteria occurred, it was also varied, it was concluded that supragingival calculus is not simply 'mineralised plaque' but a complex heterogeneous material consisting of numerous phases of mineralisation and non-mineralised areas . The role of dental plaque in the mineralisation process is unclear . However, calculus was not observed arising out of mineralising dental plaque . Mineralising dental calculus and maturing dental plaque are two distinct and different phases occurring simultaneously, albeit with great variability. J Int Acad Periodontol, 2000 Oct, 2(4), 115 - 9 Removal of supragingival plaque in an intraoral model by use of the Sonicare toothbrush; Stanford CM et al.; This study was performed to evaluate plaque removal efficacy of a mechanical toothbrush in a controlled in vivo trial . The study used enamel sections, obtained from extracted human teeth, to evaluate the efficacy of supragingival plaque removal by a mechanical powered toothbrush in various modes of use . Enamel sections were positioned in milled depressions on metal extensions of a maxillary partial denture that bilaterally extended along the buccal corridors . Four sections were used per side, positioned zero, 2 or 3mm from the surface of the metal extension . The prosthesis was worn for 16 hours, whereupon four enamel sections (two per side) were removed . Intra-oral brushing was then performed for 5 or 15 seconds on the remaining four sections . The total of adherent bacteria was then assayed from all enamel sections . Ten to twenty trials were performed (n=4 pairs/trial) for each set of variables . When the electric toothbrush was used as a manual brush ('off'), it was equivalent in plaque removal to a conventional toothbrush (Oral-B 35; P = 0.49) . However, when the powered brush was in its active mode ('on'), it removed more bacteria (P < 0.0001); this efficacy was observed even when the bristle tips were at a distance from the plaque surface. J Int Acad Periodontol, 1999 Oct, 1(4), 117 - 20 Effect of amine fluoride-stannous fluoride containing toothpaste (Meridol) on plaque and gingivitis in adults: a six-month clinical study; Shapira L et al.; Treatments that aim to reduce the accumulation of bacteria on dental surfaces are considered today as the most efficient way of treating gingivitis and the resultant destruction of periodontal tissues . Mechanical plaque removal was found to be very effective in the control of gingival inflammation for most patients, but the supplement of toothpastes and mouthwashes with anti-plaque properties may benefit individuals with unperfected plaque control . The aim of the present study was to investigate the long-term effect of amine fluoride/stannous fluoride (AmF/SnF2) toothpaste on plaque levels and gingivitis in adults, as an additional measure to standard oral hygiene . One hundred and three adult subjects with gingivitis were divided into two randomised groups . One group used NaF toothpaste (control) and the second group AmF/SnF2 (Meridol) toothpaste . The subjects were examined every two months under double-blind conditions . The recorded parameters were plaque index (PlI), gingival index (GI) and bleeding on probing . The differences in PlI between baseline and the end of the study were statistically significant for both treatments, and no significant differences were found between the two treatment groups . There was a significant drop in GI between baseline and the concluding examination only for the AmF/SnF2 group, but the differences between the two treatment groups were not significant . A significant reduction in the percentage of bleeding sites during the six-month study period was found in the AmF/SnF2 group, but not in the control group . The results of the present study suggest that the long-term use of AmF/SnF2 toothpaste in adults may be of benefit to gingival health. J Int Acad Periodontol, 1999 Oct, 1(4), 110 - 6 Epidemiology of periodontal diseases: an update; Papapanou PN; This article provides a brief review of findings from epidemiological studies of periodontal disease that have been generated over the past five years . In line with the conclusions of the 1996 World Workshop in Periodontics, the recent data support the concept that early onset periodontitis appears to be rather infrequent, while advanced adult periodontitis, leading to severe loss of supporting periodontal tissues and tooth loss, does not exceed a prevalence of 10-15% in most populations . However, a number of important issues remain unresolved . It is still not clear whether the prevalence of these diseases has shown an overall decline in recent years . In fact, retention of teeth in older age may contribute to an increase . Analytical epidemiological studies have been increasingly successful in identifying a handful of risk factors for disease onset and progression . These include colonisation at high levels by certain subgingival bacteria, environmental exposures such as cigarette smoking, and systemic conditions such as diabetes mellitus . Importantly, the molecular basis of host susceptibility has recently begun to be unraveled . Research efforts are now focused on creating multi-factorial models to assess the risk for disease, prior to the development of irreversible damage . Importantly, the role of periodontal infections as a modifier of systemic health is being increasingly explored. J Periodontol, 2003 Feb, 74(2), 219 - 24 Constitutionally hyperreactive neutrophils in periodontitis; Fredriksson MI et al.; BACKGROUND: Neutrophils constitute the first line of host defense, but their extracellular release of oxygen radicals and proteases may cause tissue destruction . Conflicting results have been reported on the neutrophilic production of oxygen radicals in periodontal patients . These may vary with different forms of periodontitis, the pathway of neutrophil activation studied, and the influence of smoking . METHODS: Venous blood was taken from 15 patients with severe chronic periodontitis, treated with scaling, with or without surgery . They were matched by age and gender to periodontally healthy controls; all participants were non-smokers . Peripheral neutrophils from the participants were activated with IgG-opsonized bacteria, complement-opsonized zymosan, or phorbol myristate acetate (PMA) . The total release of oxygen radicals was measured with luminol-enhanced chemiluminescence (CL) and the extracellular release of oxygen radicals with isoluminol-enhanced CL . Antibodies to endotoxin and the concentrations of interleukin (IL)-8 and acute phase proteins in plasma were analyzed to see if these parameters were related to hyperreactivity of the neutrophils . RESULTS: Neutrophils from periodontal patients had significantly higher CL when activated via the FcgammaR pathway, measured as both total generation of oxygen radicals and extracellular generation . The neutrophil generation of oxygen radicals was also higher in patients when complement-opsonized zymosan or PMA was used, but these differences were less pronounced . No significant differences in plasma proteins were found . CONCLUSION: Our findings suggest that the increased CL from peripheral neutrophils in periodontitis may be related to a constitutionally greater responsiveness of the Fcgamma-receptor. Dtsch Tierarztl Wochenschr, 2003 Feb, 110(2), 73 - 5 The effect of outdoor production on the seroprevalence of Lawsonia intracellularis in growing-finishing pigs in a large pig production unit infected with endemic porcine proliferative enteropathy; Bona B et al.; The study was performed in a large Croatian production unit from May 2000 till June 2002 . Blood samples form late-pregnant gilts were tested by indirect immunofluorescence antibody (IFA) serum assay for Lawsonia intracellularis . The offspring of 301 positively tested gilts were dislocated after the nursery phase either to indoor or outdoor growing-finishing facilities . Ten percent of these animals (142 indoor, 143 outdoor raised pigs) were tested at 2, 6, 10, 14, 18, 22 and 26 weeks of age for seroprevalence of Lawsonia intracellularis . All offspring of IFA positive gilts were seronegative at 2 and 6 weeks of age . At 10 weeks of age 71.1% (101 animals) of indoor and 32.8% (47 animals) of outdoor pigs were tested positive (P < 0.05) . While at 14 weeks of age 71.1% of indoor raised pigs showed seropositivity, the seropositivity declined in outdoor units to 7.6% (P < 0.01) . At weeks 18 (52.1%), 22 (47.8%) and 26 (21.7%) indoor raised pigs still showed marked seropositivity and but their outdoor raised counterparts returned to seronegativity. J Comput Chem, 2003 Apr 30, 24(6), 741 - 53 Mechanistic insights into oxidosqualene cyclizations through homology modeling; Schulz-Gasch T et al.; 2,3-Oxidosqualene cyclases (OSC) are key enzymes in sterol biosynthesis . They catalyze the stereoselective cyclization and skeletal rearrangement of (3S)-2,3-oxidosqualene to lanosterol in mammals and fungi and to cycloartenol in algae and higher plants . Sequence information and proposed mechanism of 2,3-oxidosqualene cyclases are closely related to those of squalene-hopene cyclases (SHC), which represent functional analogs of OSCs in bacteria . SHCs catalyze the cationic cyclization cascade converting the linear triterpene squalene to fused ring compounds called hopanoids . High stereoselectivity and precision of the skeletal rearrangements has aroused the interest of researchers for nearly half a century, and valuable data on studying mechanistic details in the complex enzyme-catalyzed cyclization cascade has been collected . Today, interest in cyclases is still unbroken, because OSCs became targets for the development of antifungal and hypocholesterolemic drugs . However, due to the large size and membrane-bound nature of OSCs, three-dimensional structural information is still not available, thus preventing a complete understanding of the atomic details of the catalytic mechanism . In this work, we discuss results gained from homology modeling of human OSC based on structural information of SHC from Alicyclobacillus acidocaldarius and propose a structural model of human OSC . The model is in accordance with previously performed experimental studies with mechanism-based suicide inhibitors and mutagenesis experiments with altered activity and product specificity . Structural insight should strongly stimulate structure-based design of antifungal or cholesterol-lowering drugs . FASEB J, 2003 Apr, 17(6), 644 - 51 A new approach to treat tissue destruction in periodontitis with chemically modified dextran polymers; Escartin Q et al.; Periodontitis are diseases of the supportive tissues of the teeth provoked by bacteria and characterized by gingival inflammation and bone destruction . We have developed a new strategy to repair tissues by administrating agents (RGTA) that mimic heparan sulfates by protecting selectively some of the growth factors naturally present within the injured tissue and interfering with inflammation . After periodontitis induction in hamsters, the animals were left untreated or received weekly i.m . injections of RGTA1507 at a dose of 100 microg/kg, 400 microg/kg, 1.5 mg/kg, or 15 mg/kg for 4 wk . RGTA treatment significantly reduced gingival tissue inflammation, thickened the pocket epithelium by increasing cell proliferation, and enhanced collagen accumulation in the gingiva . A marked reduction in bone loss was observed, resulting from depression of osteoclasia and robust stimulation of bone formation at the dose of 1.5 mg/kg . RGTA treatment for 8 wk at this dose reversed macroscopic bone loss, sharply contrasting with the extensive bone destruction in the untreated animals . RGTA treatment decreased gelatinase A (MMP-2) and B (MMP-9) pro-forms in gingival tissues . Our data indicate that a 4 wk treatment dose-dependently attenuated gingival and bone manifestations of the disease, whereas a longer treatment restored alveolar bone close to controls . By modulating and coordinating host responses, RGTA has unique therapeutic properties and is a promising candidate for the treatment of human periodontitis. Photochem Photobiol Sci, 2003 Feb, 2(2), 118 - 23 Replicated mesocosm study on the role of natural ultraviolet radiation in high CDOM, shallow lakes; Perez AP et al.; The role of ultraviolet radiation on shallow, high CDOM (colored dissolved organic matter) lakes was investigated during two consecutive summers (1999 and 2000) in replicated mesocosms (rectangular fiberglass tanks) . Each tank (volume: 300 L; depth: 40 cm) was covered with a layer (approximately 3 cm) of sediment from lake El Toro (40 degrees 14' S; 70 degrees 22' W) and filled with filtered water . The experimental design consisted of two treatments: full natural radiation (UV-exposed) and natural radiation without ultraviolet radiation (UV-shielded) . UV-exposed and UV-shielded treatments differed in most studied variables as revealed by repeated measures ANOVA . UV-exposed tanks displayed lower CDOM levels (dissolved absorbance) of lower average molecular size (absorbance ratio between 250 and 365 nm), higher bacterial biomass, and lower chlorophyll a concentration . The effect on consumers (rotifers and crustaceans) was less noticeable . The results are consistent with UV stimulation of bacteria production mediated by higher rates of CDOM photobleaching, and the photoinhibition of planktonic algae . Thus, a major effect of UVR in shallow, high CDOM ecosystems appears to be the stimulation of heterotrophic pathways and a simultaneous inhibition of photoautotrophs. Eksp Klin Gastroenterol . 2003;(1):57, 183. {Role of dysbiotic disruption in etiology and pathogenesis of irritable bowel syndrome}; Ruchkina IN et al.; Based on the obtained data, it was established that the development of dysbiotic changes in the large and small intestines after enteric infections is a cause for exacerbation of the irritable bowel syndrome. Extremophiles, 2003 Apr, 7(2), 159 - 68 Epub 2003 Jan 10. Purification and characterization of 5'-methylthioadenosine phosphorylase from the hyperthermophilic archaeon Pyrococcus furiosus: substrate specificity and primary structure analysis; Cacciapuoti G et al.; 5'-Methylthioadenosine phosphorylase (MTAP) was purified to homogeneity from the hyperthermophilic archaeon Pyrococcus furiosus . The protein is a homoexamer of 180 kDa . The enzyme is highly thermoactive, with an optimum temperature of 125 degrees C, and extremely thermostable, retaining 98% residual activity after 5 h at 100 degrees C and showing a half-life of 43 min at 130 degrees C . In the presence of 100 mM phosphate, the apparent T(m) (137 degrees C) increases to 139 degrees C . The enzyme is extremely stable to proteolytic cleavage and after incubation with protein denaturants, detergents, organic solvents, and salts even at high temperature . Thiol groups are not involved in the catalytic process, whereas disulfide bond(s) are present, since incubation with 0.8 M dithiothreitol significantly reduces the thermostability of the enzyme . N-Terminal sequence analysis of the purified enzyme is 100% identical to the predicted amino acid sequence of the gene PF0016 from the partially sequenced P . furiosus genome . The deduced amino acid sequence of the gene revealed a high degree of identity (52%) with human MTAP . Nevertheless, unlike human MTAP, MTAP from P . furiosus is not specific for 5'-methylthioadenosine, since it phosphorolytically cleaves adenosine, inosine, and guanosine . The calculated k(cat)/ K(m) values for 5'-methylthioadenosine and adenosine, about 20-fold higher than for inosine and guanosine, indicate that 6-amino purine nucleosides are preferred substrates of MTAP from P . furiosus . The structural features and the substrate specificity of MTAP from P . furiosus document that it represents a 5'-methylthioadenosine-metabolizing enzyme different from those previously characterized among Archaea, Bacteria, and Eukarya . The functional and structural relationships among MTAP from P . furiosus, human MTAP, and two putative MTAPs from P . furiosus and Sulfolobus solfataricus are discussed here for the first time. Extremophiles, 2003 Apr, 7(2), 123 - 30 Epub 2003 Jan 16. Arsenic resistance in the archaeon "Ferroplasma acidarmanus": new insights into the structure and evolution of the ars genes; Gihring TM et al.; Arsenic resistance in the acidophilic iron-oxidizing archaeon " Ferroplasma acidarmanus" was investigated . F . acidarmanus is native to arsenic-rich environments, and culturing experiments confirm a high level of resistance to both arsenite and arsenate . Analyses of the complete genome revealed protein-encoding regions related to known arsenic-resistance genes . Genes encoding for ArsR (arsenite-sensitive regulator) and ArsB (arsenite-efflux pump) homologues were found located on a single operon . A gene encoding for an ArsA relative (anion-translocating ATPase) located apart from the arsRB operon was also identified . Arsenate-resistance genes encoding for proteins homologous to the arsenate reductase ArsC and the phosphate-specific transporter Pst were not found, indicating that additional unknown arsenic-resistance genes exist for arsenate tolerance . Phylogenetic analyses of ArsA-related proteins suggest separate evolutionary lines for these proteins and offer new insights into the formation of the arsA gene . The ArsB-homologous protein of F . acidarmanus had a high degree of similarity to known ArsB proteins . An evolutionary analysis of ArsB homologues across a number of species indicated a clear relationship in close agreement with 16S rRNA evolutionary lines . These results support a hypothesis of arsenic resistance developing early in the evolution of life. Int Immunol, 2003 Apr, 15(4), 525 - 34 Antigen-specific, CD4+CD25+ regulatory T cell clones induced in Peyer's patches; Tsuji NM et al.; Since intestine is exposed to numerous exogenous antigens such as food and commensal bacteria, the organ bears efficient mechanisms for establishment of tolerance and induction of regulatory T cells (T(reg)) . Intestinal and inducible T(reg) include T(r)1-like and T(h)3 cells whose major effector molecules are IL-10 and transforming growth factor (TGF)-beta . These antigen-specific T(reg) are expected to become clinical targets to modify the inflammatory immune response associated with allergy, autoimmune diseases and transplantation . In the present study, we characterized the antigen-specific T(reg) induced in the intestine by orally administering high-dose beta-lactoglobulin (BLG) to BALB/c mice . Seven days after feeding, only Peyer's patch (PP) cells among different organs exerted significant suppressive effect on antibody production upon in vitro BLG stimulation . This suppressive effect was also prominent in six BLG-specific CD4(+) T cell clones (OPP1-6) established from PP from mice orally administered with high doses of BLG and was partially reversed by antibodies to TGF-beta . Intravenous transfer of OPP2 efficiently suppressed BLG-specific IgG1 production in serum following immunization, indicating the role of such T(reg) in the systemic tolerance after oral administration of antigen (oral tolerance) . OPP clones secrete TGF-beta, IFN-gamma and low levels of IL-10, a cytokine pattern similar to that secreted by anergic T cells . OPP clones bear a CD4(+)CD25(+) phenotype and show significantly lower proliferative response compared to T(h)0 clones . This lower response is recovered by the addition of IL-2 . Thus, antigen-specific CD4(+)CD25(+) T(reg), which have characteristics of anergic cells and actively suppress antibody production are induced in PP upon oral administration of protein antigen. J Pineal Res, 2003 May, 34(4), 233 - 41 Non-vertebrate melatonin; Hardeland R et al.; Melatonin has been detected in bacteria, eukaryotic unicells, macroalgae, plants, fungi and various taxa of invertebrates . Although precise determinations are missing in many of these organisms and the roles of melatonin are still unknown, investigations in some species allow more detailed conclusions . Non-vertebrate melatonin is not necessarily circadian, and if so, not always peaking at night, although nocturnal maxima are frequently found . In the cases under study, the major biosynthetic pathway is identical with that of vertebrates . Mimicking of photoperiodic responses and concentration changes upon temperature decreases have been studied in more detail only in dinoflagellates . In plants, an involvement in photoperiodism seems conceivable but requires further support . No stimulation of flowering has been demonstrated to date . A participation in antioxidative protection might be possible in many aerobic non-vertebrates, although evidence for a contribution at physiological levels is mostly missing . Protection from stress by oxidotoxins or/and extensions of lifespan have been shown in very different organisms, such as the dinoflagellate Lingulodinium, the ciliate Paramecium, the rotifer Philodina and Drosophila . Melatonin can be taken up from the food, findings with possible implications in ecophysiology as well as for human nutrition and, with regard to high levels in medicinal plants, also in pharmacology. Biochem J, 2003 May 15, 372(Pt 1), 33 - 40 Monitoring conformational changes of proteins in cells by fluorescence lifetime imaging microscopy; Calleja V et al.; To be able to detect in situ changes in protein conformation without perturbing the physiological environment would be a major step forward in understanding the precise mechanism occurring in protein interaction . We have developed a novel approach to monitoring conformational changes of proteins in intact cells . A double-labelled fluorescent green fluorescent protein-yellow fluorescent protein (GFP-YFP) fusion protein has been constructed, allowing the exploitation of enhanced-acceptor-fluorescence (EAF)-induced fluorescence resonance energy transfer (FRET) . Additionally, a novel fusion partner, YFP(dark), has been designed to act as a sterically hindered control for EAF-FRET . Any conformational changes will cause a variation in FRET, which, in turn, is detected by fluorescence lifetime imaging microscopy ("FLIM") . Protein kinase B (PKB)/Akt, a key component of phosphoinositide 3-kinase-mediated signalling, was selected for this purpose . Although conformational changes in PKB/Akt consequent to lipid binding and phosphorylation have been proposed in models, its behaviour in intact cells has not been tractable . We report here that platelet-derived-growth-factor ("PDGF") stimulation of NIH3T3 cells expressing the GFP-Akt-YFP construct resulted in a loss of FRET at the plasma membrane and hence a change in PKB/Akt conformation . We also show that the GFP-Akt-YFP construct conserves fully its functional integrity . This novel approach of monitoring the in situ conformational changes has broad application for other members of the AGC kinase superfamily and other proteins. J Org Chem, 2003 Apr 4, 68(7), 2529 - 39 Discovering nature's diverse pathways to vitamin B12: a 35-year odyssey; Scott AI; The chronology of the discoveries along the pathway of vitamin B(12) biosynthesis is reviewed from a personal perspective, including discussion of the most recent finding that two pathways to B(12) exist-one aerobic and one anaerobic-which differ mainly in the ring contraction mechanisms that convert porphyrin to corrin. Photochem Photobiol Sci, 2002 Apr, 1(4), 225 - 36 UV-induced DNA damage and repair: a review; Sinha RP et al.; Increases in ultraviolet radiation at the Earth's surface due to the depletion of the stratospheric ozone layer have recently fuelled interest in the mechanisms of various effects it might have on organisms . DNA is certainly one of the key targets for UV-induced damage in a variety of organisms ranging from bacteria to humans . UV radiation induces two of the most abundant mutagenic and cytotoxic DNA lesions such as cyclobutane-pyrimidine dimers (CPDs) and 6-4 photoproducts (6-4PPs) and their Dewar valence Isomers . However, cells have developed a number of repair or tolerance mechanism to counteract the DNA damage caused by UV or any other stressors . Photoreactivation with the help of the enzyme photolyase is one of the most important and frequently occurring repair mechanisms in a variety of organisms . Excision repair, which can be distinguished into base excision repair (BER) and nucleotide excision repair (NER), also plays an important role in DNA repair in several organisms with the help of a number of glycosylases and polymerases, respectively . In addition, mechanisms such as mutagenic repair or dimer bypass, recombinational repair, cell-cycle checkpoints, apoptosis and certain alternative repair pathways are also operative in various organisms . This review deals with UV-induced DNA damage and the associated repair mechanisms as well as methods of detecting DNA damage and its future perspectives. Kansenshogaku Zasshi, 2003 Feb, 77(2), 110 - 5 {Evaluation of rapid identification method for Mycobacterium tuberculosis complex using the immunochromatographic slide test kit}; Hasegawa M et al.; Capilia TB, a lateral flow immunochromatographic slide test kit for directly identifying Mycobacterium tuberculosis complex (MTC), was evaluated by using culture-positive specimens from Mycobacteria Growth Indicator Tubes (MGIT) . Sputum specimens from patients suspected of having tuberculosis were treated with NALC-NaOH and cultivated in MGIT960 . Liquid specimens were collected from the positive tubes and directly inoculated with Capilia TB . Liquid specimens were also directly tested with AccuProbe . Of the organisms isolated from the 100 MGIT positive tubes, M . tuberculosis complex was identified in 49 (49%) tubes with Capilia TB and not identified in 51 (51%) with Capilia TB . Mycobacterium avium-intracellulare complex (MAC) was identified in 46 (46%) with AccuProbe MAC and other acid-fast bacteria were identified in 5 (5%) by DNA-DNA hybridization method . There were one tube in which M . tuberculosis complex was detected with Capilia TB and M . tuberculosis complex was not detected with AccuProbe MTC, but no tubes in which M . tuberculosis complex was detected with AccuProbe MTC and M . tuberculosis complex was not detected with Capilia TB . Capilia TB is excellent in sensitivity and specificity and very suitable for rapid diagnosis of tuberculosis and is considered to contribute to public health intervention measures taken for the tuberculosis control in Japan. J Infect Dis, 2003 Apr 1, 187(7), 1126 - 36 Epub 2003 Mar 19. Type I interferon induction pathway, but not released interferon, participates in the maturation of dendritic cells induced by negative-strand RNA viruses; Lopez CB et al.; Signaling through toll-like receptors (TLRs) is essential for dendritic cell (DC) maturation induced by bacteria and other pathogens . The mechanism for virus-induced DC maturation is not known . By use of pairs of live viruses with different abilities to induce the interferon (IFN) pathway, a strong correlation between DC maturation and the ability of the virus to induce type I IFN synthesis was demonstrated . The secreted IFN was not necessary, nor was it sufficient to induce full DC maturation . Intracellular viral replication is necessary for this process, and the transcription factor nuclear factor-kappaB was crucial for cytokine induction . The double-stranded RNA-dependent protein kinase was not essential for DC maturation . Similar to TLR-induced DC maturation, after virus infection, separate pathways for the induction of cytokine secretion and the up-regulation of major histocompatibility complex and costimulatory molecules were activated . It was demonstrated that these pathways have different sensitivities to the presence of viral stimulus. Minerva Stomatol, 2002 Dec, 51(11-12), 495 - 9 {Risk factors in periodontal disease}; Sartori S et al.; Some life styles and systemic pathologies represent risk factors for the periodontal disease . Cigarette smoking in particular increases the speed of progression of periodontitis . Other conditions such as diabetes mellitus, presence of genetic markers and some bacteria are significantly correlated with the periodontal disease. J Leukoc Biol, 2003 Apr, 73(4), 493 - 501 Fever-like thermal conditions regulate the activation of maturing dendritic cells; Tournier JN et al.; Fever is one of the most frequent clinical signs encountered in pathology, especially with respect to infectious diseases . It is currently thought that the role of fever on immunity is limited to activation of innate immunity; however, its relevance to activation of adaptive immunity remains unclear . Dendritic cells (DCs) that behave as sentinels of the immune system provide an important bridge between innate and adaptive immunity . To highlight the role of fever on adaptive immunity, we exposed murine bone marrow-derived lipopolysaccharide (LPS)- or live bacteria-maturing DCs over a 3-h period to 37 degrees C or to fever-like thermal conditions (39 degrees C or 40 degrees C) . At these three temperatures, we measured the kinetics of cytokine production and the ability of DCs to induce an allogeneic mixed lymphocyte reaction . Our results show that short exposure of DCs to temperatures of 39 degrees C or 40 degrees C differentially increased the secretion of interleukin (IL)-12p70 and decreased the secretion of IL-10 and tumor necrosis factor alpha by maturing DCs . These fever-like conditions induced a regulation of cytokine production at the single-cell level . In addition, short-term exposed LPS-maturing DCs to 39 degrees C induced a stronger reaction with allogeneic CD4(+) T cells than maturing DCs incubated at 37 degrees C . These results provide evidence that temperature regulates cytokine secretion and DC functions, both of which are of particular importance in bacterial diseases. Biochem Biophys Res Commun, 2003 Apr 4, 303(2), 640 - 4 Binding and internalization of Helicobacter pylori VacA via cellular lipid rafts in epithelial cells; Kuo CH et al.; In this study we investigated the roles of lipid rafts and glycosylphosphatidylinositol-anchored proteins (GPI-APs) in the process of VacA binding and internalization into epithelial cells . Vacuolating activity analysis in AGS, CHO cells, and a CHO-derived line that highly expresses GPI-linked fasI proteins indicated the significance of cholesterol and GPI-APs for VacA activity . Flow cytometric analysis along with VacA-cholesterol co-extraction experiments showed a cholesterol-dependent manner for VacA cell-binding activity, while GPI-APs were not related to it . Differential detergent extraction and fractionation in sucrose density gradient showed co-association of VacA and fasI with rafts on cell membranes . Subcellular distribution of fasI visualized by confocal microscope suggested that fasI trafficked via a newly defined endocytic pathway for GPI-APs in the derived line . Upon VacA intoxication, VacA was visualized to co-migrate along with fasI and finally induced vacuolation coupled with dramatic redistribution of fasI molecules . These results suggest that VacA exploits rafts for docking and entering the cell via the endocytic pathway of GPI-APs. Biochem Biophys Res Commun, 2003 Apr 4, 303(2), 475 - 82 One-step on-column affinity refolding purification and functional analysis of recombinant human VDAC1; Shi Y et al.; The outer mitochondrial membrane porin, voltage-dependent anion-selective channel (VDAC), is believed to play an important role in mediating mitochondria-dependent apoptosis . However, detailed structure-function studies of VDAC have been hindered by the difficulties to obtain a soluble, correctly folded, and fully active form of the recombinant VDAC and its mutant variants due to its transmembrane nature . Here we report a high-throughput one-step chromatographic procedure in purification of recombinant human VDAC1 (rhVDAC1) protein overexpressed in bacteria . The improved methodology could generate a large quantity of rhVDAC1 with correct folding in terms of the secondary structure, with full biological activities in mediating cytochrome c release and in interaction with Bcl-X(L) . The method will significantly benefit genetic, biochemical, and structural studies of this critical channel protein. Alcohol Clin Exp Res, 2003 Mar, 27(3), 503 - 8 Phagocytosis and production of reactive oxygen species by peripheral blood phagocytes in patients with different stages of alcohol-induced liver disease: effect of acute exposure to low ethanol concentrations; Parlesak A et al.; BACKGROUND: In rodents, the development of alcoholic liver disease (ALD) after chronic alcohol feeding was shown to depend on the activity of enzymes that are necessary for production of reactive oxygen species (ROS) in phagocytes . The aim of this study was to determine the formation of ROS by resting and challenged phagocytes of patients with different stages of ALD in the presence of ethanol concentrations commonly found in the blood of alcohol abusers . PATIENTS AND METHODS: The release of ROS and the phagocytosis of bacteria by neutrophils and monocytes obtained from 60 patients, who were categorized in three groups due to the severity of ALD, were compared to that of 28 healthy controls . ROS release by these phagocytes was measured after challenging with endotoxin and the addition of ethanol (22 and 44 mM) . RESULTS: Resting neutrophils but not monocytes from patients with severe stages of ALD produced significantly more ROS than those of healthy controls . Basal values of ROS production from neutrophils correlated closely to markers of the severity of ALD . ROS formation was depressed dose-dependently by ethanol in the healthy controls but not in alcohol abusers . CONCLUSIONS: Changes in the ROS metabolism of phagocytes found in this study might contribute to both the development of ALD and the impaired immune response occurring in patients with severe ALD. Pancreas, 2003 Apr, 26(3), 274 - 8 Adenovirus-mediated gene transfer in the treatment of pancreatic cancer; Pan X et al.; INTRODUCTION: Suicide gene therapy is a new experimental form of cancer chemotherapy that is currently being evaluated in human trials.AIM: To evaluate the killing effects of the cytosine deaminase (CD) gene mediated by an adenovirus vector on human pancreatic carcinoma in vitro and in vivo . METHODOLOGY: The CD gene was cloned into pAdTrack-CMV-CD, pAdTrack-CMV-CD, and pAdEasy-1, which underwent recombination in bacteria BJ5183 . The newly recombinant Ad-CD containing green fluorescent protein was propagated in 293 cells and purified by cesium chloride gradient centrifugation . The human pancreatic carcinoma cell line PaTu8988/SW1990 was infected with this virus, and then 5-fluorocytosine (5-FC) was added; XTT assay was used to estimate relative numbers of viable cells . An in vivo model of pancreatic cancer was established by injecting 1.0 x 10(7) PaTu8988/SW1990 cells subcutaneously in Balb/c nude mice . When tumors were palpable, Ad-CD was injected into each tumor, and 5-FC was administered . The positive clones were selected by endonuclease digestion of the combinations, and the concentration of viral liquids containing the CD gene was 2 x 10(11) pfu/mL . RESULTS: It was found that significant cytotoxic activities were possessed by 5-FC for CD gene-transduced PaTu8988/SW1990 cells, but there was little effect on the nontransduced pancreatic carcinoma cells . The antitumor effect was observed in PaTu8988/SW1990 xenografts from nude mice with in situ CD gene transduction . CONCLUSION: These results indicate that the CD gene mediated by adenovirus has a high level of infectivity and is efficient for gene therapy for pancreatic carcinoma. J Neurosci, 2003 Mar 15, 23(6), 2170 - 81 Synchronized formation and remodeling of postsynaptic densities: long-term visualization of hippocampal neurons expressing postsynaptic density proteins tagged with green fluorescent protein; Ebihara T et al.; To explore mechanisms governing the formation and remodeling of postsynaptic density (PSD), we used dissociated cultures of hippocampal neurons isolated from transgenic embryos expressing green fluorescent protein (GFP)-tagged PSD proteins PSD-Zip45 (Homer 1c) and PSD-95 . Expression of GFP-tagged PSD molecules was stable, and the remodeling process of PSDs could be followed for >1 week . A higher expression level of GFP-PSD-Zip45 enabled us to quantitatively analyze the amount of PSD-Zip45 clusters during development . Repetitive imaging of the same cell populations between 11 and 17 d in culture revealed an increase of the average PSD-Zip45 cluster density from 0.32 to 0.73/microm . Newly generated dendrites rapidly acquired GFP-PSD-Zip45 clusters, and their density reached the level of parental dendrites within a few days . Temporal profiles of GFP-PSD-Zip45 cluster density showed a variety of patterns . Some dendrites showed a monotonous increase of clusters, whereas others showed complex patterns, including short decremental stages . Analysis of long-term remodeling of PSD-95-GFP clusters confirmed that the decremental stages were not specific to the PSD-Zip45 clusters . Comparison of the temporal profiles of the cluster density among neurons indicated synchronization of both GFP-PSD-Zip45 and PSD-95 clustering within individual cells . Furthermore, activation of cAMP-dependent protein kinase suppressed the decremental stages of cluster remodeling . These observations suggest the presence of signaling mechanisms that can induce synchronized addition or elimination of PSD proteins throughout dendritic arborization of a single neuron. Gene, 2003 Mar 13, 306, 37 - 44 Molecular cloning and characterization of a novel gene family of four ancient conserved domain proteins (ACDP); Wang CY et al.; We have recently cloned four novel human genes that encode the ancient conserved domain proteins (ACDP) . The full-length cDNA sequence of ACDP1 consists of 5898 bp and encodes a predicted protein of 951 amino acids (AA) . The transcript for ACDP2 has 4058 bp of cDNA sequence, encoding a protein of 875 AA . ACDP3 contains 3113 bp of cDNA sequence and encodes a putative protein of 707 AA . ACDP4 contains 4765 bp of cDNA sequence and encodes a protein of 775 AA . The ACDP genes belong to a highly conserved new gene family . The conserved region showed 62.8% of nucleotide sequence identity, and 65.5% of AA identity with 92% of AA homologies among ACDP members . The conserved domain is also found in genes from evolutionarily divergent species from bacteria, yeast, Caenorhabditis elegans, and Drosophila melanogaster to mammals . All ACDP genes except ACDP1 have a ubiquitous expression pattern while ACDP1 expression is restricted to the brain and testis . Immunofluorescence staining of premeablized HeLa cells showed that ACDP proteins are predominantly localized in the nucleus . Sequence homology analyses revealed AA property and structural homologies between the ACD domain and cyclin molecules. Int Endod J, 2003 Feb, 36(2), 106 - 16 Histomorphometric analysis of odontoblast-like cell numbers and dentine bridge secretory activity following pulp exposure; Murray PE et al.; AIM: The purpose of this study was to collect quantitative information about the numbers and dentine bridge secretory activity of odontoblast-like cells following dental pulp exposure . METHODOLOGY: The numbers and secretory activity of odontoblast-like cells were measured histomorphometrically between 7 days and 2 years in 161 pulp-exposed nonhuman primate teeth . The area of dentine bridges and the dimensions of cavity preparations were measured . The density of odontoblast-like cells and subjacent reorganizing tissue cells were measured beneath dentine bridge formation . The presence of operative dentine debris and tunnel defects in bridges was noted . Pulp inflammation was categorized according to ISO standards . Bacteria were detected using McKay's stain . RESULTS: The area of dentine bridges was mediated by the density and secretory activity of odontoblast-like cells over time . The cell density of subjacent reorganizing tissue was found to be strongly associated with that of odontoblast-like cells . Bacterial microleakage was found to impede dentine bridge secretion by odontoblast-like cells . CONCLUSIONS: Pulp reparative activity occurs naturally beneath capping materials in the absence of bacterial microleakage . The outcome of pulp-capping treatments could be beneficially influenced by concentrating attention on limiting the width of pulp exposure, minimizing pulp injury by limiting the creation of operative debris and placing materials which prevent bacterial microleakage. Aliment Pharmacol Ther, 2003 Apr 1, 17(7), 895 - 904 A randomized controlled trial of a probiotic, VSL#3, on gut transit and symptoms in diarrhoea-predominant irritable bowel syndrome; Kim HJ et al.; AIM: To investigate the effects of a probiotic formulation, VSL#3, on gastrointestinal transit and symptoms of patients with Rome II irritable bowel syndrome with predominant diarrhoea . METHODS: Twenty-five patients with diarrhoea-predominant irritable bowel syndrome were randomly assigned to receive VSL#3 powder (450 billion lyophilized bacteria/day) or matching placebo twice daily for 8 weeks after a 2-week run-in period . Pre- and post-treatment gastrointestinal transit measurements were performed in all patients . Patients recorded their bowel function and symptoms daily in a diary during the 10-week study, which was powered to detect a 50% change in the primary colonic transit end-point . RESULTS: There were no significant differences in mean gastrointestinal transit measurements, bowel function scores or satisfactory global symptom relief between the two treatment groups, pre- or post-therapy . Differences in abdominal bloating scores between treatments were borderline significant (P = 0.09, analysis of covariance) . Further analysis revealed that abdominal bloating was reduced (P = 0.046) with VSL#3 {mean post- minus pre-treatment score, - 13.7; 95% confidence interval (CI), - 2.5 to - 24.9}, but not with placebo (P = 0.54) (mean post- minus pre-treatment score, - 1.7; 95% CI, 7.1 to - 10.4) . With the exception of changes in abdominal bloating, VSL#3 had no effect on other individual symptoms: abdominal pain, gas and urgency . All patients tolerated VSL#3 well . CONCLUSION: VSL#3 appears to be promising in the relief of abdominal bloating in patients with diarrhoea-predominant irritable bowel syndrome . This is unrelated to an alteration in gastrointestinal or colonic transit. Drugs, 2003, 63(6), 535 - 53 Fixed-dose combination drugs for tuberculosis: application in standardised treatment regimens; Blomberg B et al.; Short-course chemotherapy is highly efficacious in treating tuberculosis (TB) . However, the length (>/=6 months) and complexity (three or four different drugs) of the treatment makes adherence difficult . Erratic treatment not only fails to cure patients but also creates chronically contagious cases, who may excrete drug-resistant TB bacteria . The Directly Observed Treatment Short-course (DOTS) strategy recommended by WHO provides a comprehensive organisational and infrastructural framework for the rational use of diagnosis, drug supply, as well as case and programme management services, in TB control . WHO and other organisations recommend fixed-dose combination formulations (FDCs) as a further step to facilitate the optimal drug treatment of TB . Using FDCs in TB control will simplify the doctor's prescription and patient's drug intake, as well as the drug supply management of the programme . By preventing monotherapy and facilitating the ingestion of adequate doses of the constituent anti-TB drugs, FDCs are expected to help prevent the emergence of drug resistance.This article presents the international recommendations for the use of FDCs in TB programmes . The fundamental issue is to obtain drug supplies of good quality . A laboratory network for quality testing, including bioavailability testing of FDCs exists, and the recently established Global TB Drug Facility (GDF) supplies quality TB drugs, including 4-drug FDCs, to countries requesting assistance . This articles deals with the requirements for a successful transition to FDC-based treatment . It emphasises the need for appropriately revised programme documentation (programme manual, training modules, treatment guidelines and forms), training of staff at all levels, carefully calculated drug needs, and a plan for the exhaustion of existing stocks of loose tablets and the phasing-in of FDCs at all levels of the programme at the same time . Loose drugs for individualised treatment of patients with adverse effects should be kept at district or central health institutions. Chemosphere, 2003 Jan, 50(1), 165 - 9 The inhibitory effects of lindane in batch and upflow anaerobic sludge blanket reactors; Erguder TH et al.; In this study, the inhibitory effects of lindane (LIN) on originally unacclimated mixed anaerobic cultures were investigated by anaerobic toxicity assay (ATA) experiments . ATA experiments revealed that 10 mg/l LIN exerted inhibitory effects on anaerobic cultures, which was recoverable . Continuous reactor experiments conducted to determine the inhibitory effects of LIN and the maximum LIN loading rate achievable in two-stage upflow anaerobic sludge blanket (UASB) reactors indicated that anaerobic granular cultures were successfully acclimated to 30 mg/l LIN . The maximum LIN loading rate and minimum hydraulic retention time (HRT) possible for the UASB system were 10 mg/l day and 18 h, respectively, which resulted in the overall chemical oxygen demand (COD) removal efficiency of 89%. J Biol Chem, 2003 Jun 6, 278(23), 20882 - 9 Epub 2003 Mar 24. Lectins homologous to those of monocotyledonous plants in the skin mucus and intestine of pufferfish, Fugu rubripes; Tsutsui S et al.; We have characterized pufflectin, a novel mannose-specific lectin, from the skin mucus of the pufferfish, Fugu rubripes . Molecular mass estimations by gel filtration and matrix-assisted laser desorption ionization time-of-flight mass spectrometry and the SDS-PAGE pattern suggest that pufflectin is a homodimer composed of non-covalently associated subunits of 13 kDa . The full-length pufflectin cDNA consists of 527 bp, with 116 amino acid residues deduced from the open reading frame . The amino acid sequence of pufflectin shows no homology with any known animal lectin . Surprisingly, pufflectin shares sequence homology with mannose-binding lectins of monocotyledonous plants and has conserved two of three carbohydrate recognition domains of these plant lectins . The pufflectin gene is expressed in gills, oral cavity wall, esophagus, and skin . In addition, an isoform occurs exclusively in the intestine . Pufflectin differs from mannose-binding lectins purified from the blood plasma of Fugu . Whereas pufflectin did not agglutinate five bacterial species tested, it was demonstrated to bind to the parasitic trematode, Heterobothrium okamotoi . This finding suggests that pufflectin contributes to the parasite-defense system in Fugu. Infect Immun, 2003 Apr, 71(4), 1995 - 2001 Anaplasma phagocytophilum reduces neutrophil apoptosis in vivo; Scaife H et al.; Ovine neutrophils spontaneously underwent apoptosis during culture in vitro, as assessed by morphological changes and exposure of annexin V binding sites on their cell surfaces . The addition of conditioned medium from concanavalin A-treated ovine peripheral blood mononuclear cells (PBMC) could partially protect against this progression into apoptosis, but dexamethasone and sodium butyrate could not . Actinomycin D accelerated the rate at which ovine neutrophils underwent apoptosis . Neutrophils isolated from sheep experimentally infected with Anaplasma phagocytophilum showed significantly delayed apoptosis during culture ex vivo, and the addition of conditioned medium from PBMC to these cells could not delay apoptosis above the protective effects observed after in vivo infection . The ability of neutrophils from A . phagocytophilum-infected sheep to activate a respiratory burst was increased compared to the activity measured in neutrophils from uninfected sheep, but chemotaxis was decreased in neutrophils from infected sheep . These data are the first demonstration that in vivo infection with A . phagocytophilum results in changes in rates of apoptosis of infected immune cells . This may help explain how these bacteria replicate in these normally short-lived cells. Antimicrob Agents Chemother, 2003 Apr, 47(4), 1430 - 2 Novel tetracycline resistance determinant from the oral metagenome; Diaz-Torres ML et al.; A major drawback of most studies on how bacteria become resistant to antibiotics is that they concentrate mainly on bacteria that can be cultivated in the laboratory . In the present study, we cloned part of the oral metagenome and isolated a novel tetracycline resistance gene, tet(37), which inactivates tetracycline. Drug Discov Today, 2003 Apr 1, 8(7), 297 - 306 Targeting RNA: new opportunities to address drugless targets; Zaman GJ et al.; Historically, pharmaceutical industries have focussed on the discovery of compounds that target the protein products of genes . The intermediary product between gene and protein, consisting of RNA, has remained largely unexplored . Several drugs targeting the rRNA of bacteria have been, however, in clinical use for over half a century . One of these drug classes, the aminoglycoside antibiotics, also targets human rRNA, and have been developed as therapeutics for genetic disorders . Targeting at the RNA level is an economical approach to address non-drugable proteins and targets that have failed to give leads by hits in HTS, as it can build on biological knowledge gathered over years . RNA also offers entirely new opportunities for drug development, such as targeting of non-coding RNA sequences. J Mol Biol, 2003 Apr 4, 327(4), 867 - 84 Hammond behavior versus ground state effects in protein folding: evidence for narrow free energy barriers and residual structure in unfolded states; Sanchez IE et al.; Apparent transition state movement upon mutation or changes in solvent conditions is frequently observed in protein folding and is often interpreted in terms of Hammond behavior . This led to the conclusion that barrier regions in protein folding are broad maxima on the free energy landscape . Here, we use the concept of self-interaction and cross-interaction parameters to test experimental data of 21 well-characterized proteins for Hammond behavior . This allows us to characterize the origin of transition state movements along different reaction coordinates . Only one of the 21 proteins shows a small but coherent transition state movement in agreement with the Hammond postulate . In most proteins the structure of the transition state is insensitive to changes in protein stability . The apparent change in the position of the transition state upon mutation, which is frequently observed in phi-value analysis, is in most cases due to ground-state effects caused by structural changes in the unfolded state . This argues for significant residual structure in unfolded polypeptide chains of many proteins . Disruption of these residual interactions by mutation often leads to decreased folding rates, which implies that these interactions are still present in the transition state . The failure to detect Hammond behavior shows that the free energy barriers encountered by a folding polypeptide chain are generally rather narrow and robust maxima for all experimentally explorable reaction coordinates . Pediatr Int, 2003 Feb, 45(1), 31 - 4 Therapeutic effect of spiramycin in brucellosis; Geyik MF et al.; OBJECTIVE: This study was undertaken to investigate the usefulness of spiramycin in treatment for brucellosis in an animal model . METHODS: Eighty-four Sprague-Dawley rats were infected by intraperitoneal injection of Brucella melitensis suspension . Seven days after inoculation, four rats were selected randomly, killed and spleen cultures and Brucella standard tube agglutination test were carried out . All four rats were found to be infected . Eighty adult rats were randomly divided into four groups of 20 rats each . Tap water was given to the first group . Rifampicin 50 mg/kg per day and doxycycline 40 mg/kg per day were given to the second group, spiramycin 50 mg/kg per day orally was given to the third group, and a combination of spiramycin and rifampicin at the same dose and period was given to the fourth group . Duration of therapy regimens in all groups was 21 days . The spleens of all 80 rats were removed aseptically, homogenized, and placed onto Brucella agar plates to determine if viable bacteria were present . RESULTS: Bacterial growth occurred in all of the rats' spleens in the first group and in two rats' spleens in the spiramycin group . Mean colony forming unit (c.f.u.) values were at the highest in the first group . The effectivities of spiramycin and rifampicin-spiramycin were similar to rifampicin-doxycycline . There were no differences in the treatment results between the three groups that received combined rifampicin-doxycycline, rifampicin-spiramycin and only spiramycin (P>0.05) . CONCLUSIONS: The results show that spiramycin cures experimental rat brucellosis and may be an effective alternative in the therapy of human brucellosis. J Oral Pathol Med, 2003 Apr, 32(4), 219 - 23 Helicobacter pylori DNA in recurrent aphthous stomatitis; Victoria JM et al.; BACKGROUND: Considering not only the fact that recurrent aphthous stomatitis (RAS) and stomach ulcers are immunologically mediated ulcers associated with Helicobacter pylori, but also the recent evidence that anaemia can be associated with both diseases, and the discovery of H . pylori in the oral mucosa led us to hypothesize that this bacteria may be related to RAS pathogenesis . METHODS: Thirty-six consecutive subjects affected by minor and major forms of RAS and 48 healthy volunteers were included in the present study . The nested polymerase chain reaction (PCR) technique was used to detect the presence of H . pylori in the oral lesion, the normal contralateral mucosa of patients affected by RAS and the oral mucosa of control subjects . The chi2- and Fisher's tests were used for statistical analysis . RESULTS: No association between RAS lesions and H . pylori was observed . However, 14 out of 36 (38.9%) of the patients with RAS were found to show the presence of H . pylori DNA in the lesion and/or contralateral mucosa . Sixteen out of 48 (33.3%) of the patients without RAS (control subjects) were positive (P > 0.05) . CONCLUSION: The present study does not give support to the assumption that H . pylori could be involved in RAS development. Arch Pathol Lab Med, 2003 Mar, 127(3), 289 - 96 Reactivity of resident immunocytes in normal and prepsoriatic skin using an ex vivo skin-explant model system; Curry JL et al.; CONTEXT: While it is well known that both exogenous and endogenous stimuli can trigger appearance of psoriatic lesions, the initial cellular and molecular events mediated by immunocompetent cells normally resident in prepsoriatic (PN) skin are not well understood . Moreover, it is unclear whether there are any fundamentally important differences in the innate immune response of normal healthy skin (NN skin) versus PN skin . Since acute tissue responses to stimuli involve both resident cells and immunocytes recruited rapidly from circulation, it is difficult to discern the contribution of endogenous cells normally present in skin . OBJECTIVE: To solely characterize the reactivity of resident immunocytes using an experimental system . DESIGN: To probe the activation potential of resident immunocytes in NN (n = 18) and PN skin (n = 10), a short-term ex vivo organ culture system containing interleukin (IL)-2 was established and characterized . To mimic exogenous or environmental trigger factors, bacteria-derived superantigens and lipopolysaccharide were added to the skin-explant assays, whereas endogenous trigger factors were investigated using heat shock proteins . RESULTS: Using this skin-explant assay, both NN and PN skin gave rise to an expansion of various T-cell subsets, which could differentially produce various cytokines and a growth factor (keratinocyte growth factor), depending on the stimulus and source of skin . Bacterial superantigens were relatively potent inducers of interferon-gamma, and natural killer-T cells were observed proliferating from PN skin . CONCLUSIONS: Despite relatively few T cells normally residing in either NN or PN skin, initiation of skin explants from both sets of individuals in the presence of IL-2 triggered vigorous T-cell proliferation and cytokine/growth factor release . These results demonstrate the utility of this skin-explant assay system to further investigate quantitative and qualitative immune responses of NN and PN skin. Biochemistry, 2003 Apr 1, 42(12), 3446 - 56 Structural studies on ADP activation of mammalian glutamate dehydrogenase and the evolution of regulation; Banerjee S et al.; Glutamate dehydrogenase (GDH) is found in all organisms and catalyzes the reversible oxidative deamination of L-glutamate to 2-oxoglutarate . Unlike GDH from bacteria, mammalian GDH exhibits negative cooperativity with respect to coenzyme, activation by ADP, and inhibition by GTP . Presented here are the structures of apo bovine GDH, bovine GDH complexed with ADP, and the R463A mutant form of human GDH (huGDH) that is insensitive to ADP activation . In the absence of active site ligands, the catalytic cleft is in the open conformation, and the hexamers form long polymers in the crystal cell with more interactions than found in the abortive complex crystals . This is consistent with the fact that ADP promotes aggregation in solution . ADP is shown to bind to the second, inhibitory, NADH site yet causes activation . The beta-phosphates of the bound ADP interact with R459 (R463 in huGDH) on the pivot helix . The structure of the ADP-resistant, R463A mutant of human GDH is identical to native GDH with the exception of the truncated side chain on the pivot helix . Together, these results strongly suggest that ADP activates by facilitating the opening of the catalytic cleft . From alignment of GDH from various sources, it is likely that the antenna evolved in the protista prior to the formation of purine regulatory sites . This suggests that there was some selective advantage of the antenna itself and that animals evolved new functions for GDH through the addition of allosteric regulation. Biomed Pharmacother, 2002, 56 Suppl 2, 319s - 326s Geomagnetics and society interact in weekly and broader multiseptans underlying health and environmental integrity; Cornelissen G et al.; Evidence for the ubiquity and partial endogenicity of about-weekly (circaseptan) components and multiples and/or submultiples thereof (the multiseptans) accumulates as longer and denser records become available . Often attributed to a mere response to the soc