Application of chimeric glucanase comprising mutanase and dextranase for prevention of dental biofilm formation

Water‐insoluble glucan (WIG) produced by mutans streptococci, an important cariogenic pathogen, plays an important role in the formation of dental biofilm and adhesion of biofilm to tooth surfaces. Glucanohydrolases, such as mutanase (α‐1,3‐glucanase) and dextranase (α‐1,6‐glucanase), are able to hydrolyze WIG. The purposes of this study were to construct bi‐functional chimeric glucanase, composed of mutanase and dextranase, and to examine the effects of this chimeric glucanase on the formation and decomposition of biofilm. The mutanase gene from Paenibacillus humicus NA1123 and the dextranase gene from Streptococcus mutans ATCC 25175 were cloned and ligated into a pE‐SUMOstar Amp plasmid vector. The resultant his‐tagged fusion chimeric glucanase was expressed in Escherichia coli BL21 (DE3) and partially purified. The effects of chimeric glucanase on the formation and decomposition of biofilm formed on a glass surface by Streptococcus sobrinus 6715 glucosyltransferases were then examined. This biofilm was fractionated into firmly adherent, loosely adherent, and non‐adherent WIG fractions. Amounts of WIG in each fraction were determined by a phenol‐sulfuric acid method, and reducing sugars were quantified by the Somogyi–Nelson method. Chimeric glucanase reduced the formation of the total amount of WIG in a dose‐dependent manner, and significant reductions of WIG in the adherent fraction were observed. Moreover, the chimeric glucanase was able to decompose biofilm, being 4.1 times more effective at glucan inhibition of biofilm formation than a mixture of dextranase and mutanase. These results suggest that the chimeric glucanase is useful for prevention of dental biofilm formation.     

Serine protease inhibitors modulate chemotactic cytokine production by human lung fibroblasts in vitro

Chemotactic chemokines can be released from lung fibroblasts in response to interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha. An imbalance between proteases and antiproteases has been observed at inflammatory sites, and, therefore, protease inhibitors might modulate fibroblast release of chemotactic cytokines. To test this hypothesis, serine protease inhibitors (FK-706, alpha(1)-antitrypsin, or N(alpha)-p-tosyl-L-lysine chloromethyl ketone) were evaluated for their capacity to attenuate the release of neutrophil chemotactic activity (NCA) or monocyte chemotactic activity (MCA) from human fetal lung fibroblasts (HFL-1). Similarly, the release of the chemoattractants IL-8, granulocyte colony-stimulating factor, monocyte chemoattractant protein-1, macrophage colony-stimulating factor, and granulocyte/macrophage colony-stimulating factor, from HFL-1, were evaluated in response to IL-1beta and TNF-alpha. NCA, MCA, and chemotactic cytokines were attenuated by FK-706. However, matrix metalloproteinase inhibitors were without effect, and cysteine protease inhibitors only slightly attenuated chemotactic or cytokine release. These data suggest that IL-1beta and TNF-alpha may stimulate lung fibroblasts to release NCA and MCA by a protease-dependent mechanism and that serine protease inhibitors may attenuate the release.     

Phytochrome- and blue-light-absorbing pigment-mediated directional movement of chloroplasts in dark-adapted prothallial cells of fernAdiantum as analyzed by microbeam irradiation

When prothalli ofAdiantum capillus-veneris L. were kept for 2 d in the dark, chloroplasts gathered along the anticlinal walls (Kagawa and Wada, 1994, J Plant Res 107: 389–398). In these dark-adapted prothallial cells, irradiation with a microbeam (10 gm in diameter) of red (R) or blue light (B) for 60 s moved the chloroplasts towards the irradiated locus during a subsequent dark period. Chloroplasts located less than 20 gm from the center of the R microbeam (18 J·m^–2) moved towards the irradiated locus. The higher the fluence of the light, the greater the distance from which chloroplasts could be attracted. The B microbeam was less effective than the R microbeam. Chloroplasts started to move anytime up to 20 min after the R stimulus, but with the B microbeam the effect of the stimulus was usually apparent within 10 min after irradiation. The velocity of chloroplast migration was independent of light-fluence in both R and B and was about - 0.3 m·min^–1 between 15 min and 30 min after irradiation. Whole-cell irradiation with far-red light immediately after R- and B-microbeam irradiations demonstrated that these responses were mediated by phytochrome and a blue-light-absorbing pigment, respectively. Sequential treatment with R and B microbeams, whose fluence rates were less than the threshold values when applied separately, resulted in an additive effect and induced chloroplast movement, strongly suggesting that signals from phytochrome and the blue-light-absorbing pigment could interact at some point before the induction of chloroplast movement.Abbreviations B blue light - FR far-red light - IR infrared light - R red light     

Chemical Synthesis and Characterization of α-N-Octanoyl and Other α-N-Acyl Colistin Nonapeptide Derivatives

Eight α-N-acyl colistin nonapeptide derivatives including three aliphatic, four aromatic and one alicyclic derivatives were synthesized by the reaction of colistin nonapeptide with corresponding acid chlorides. This acylation reaction was carried out under the condition kept restrictedly at pH 5,0 in order to introduce an acyl group only to α-amino group but not to γ-amino group existing in a colistin nonapeptide molecule. Synthetic method and several physico-chemical natures of these acyl colistin nonapeptide derivatives are given in this paper.

All of the acylated derivatives thus synthesized exhibited characteristic antimicrobial activities. Antimicrobial spectra were substantially based upon a gram-negative type and not so much altered by chemical structures of acyl groups which were considerably differentiated from each other such as cyclic or chain form. Thus, more possible response of carbon size than its structure to the antimicrobial effectiveness was inferred. In spite of almost no toxicity and feeble antimicrobial activity of colistin nonapeptide itself, these acylated colistin nonapeptide derivatives showed a toxicity against mice at a dose of 16.9~70 mg/kg in LD₅₀, which, however, was inferior to the toxicity of colistin sulfate, possibly correspondent to their much weaker antimicrobial activities, as a whole. Hence, it seems likely that acyl part of these acylated colistin nonapeptide derivatives including that of colistin is seriously responsible for the biological activities.     

Formation of Threonine Aldolase by Bacteria and Yeasts

Threonine aldolase was found to be formed in various strains of bacteria and yeasts when they were grown in media containing l-threonine as a sole source of carbon. As the other sources of carbon, d, l-allothreonine, l-serine and glycine were effective but glucose and sucrose were inert for the formation of the enzyme.

The maximal formation of the enzyme was observed in the initial of stationary phase of growth and, thereafter, the enzyme disappeared with the consumption of l-threonine. It seems that the enzyme is adaptive in nature and that it is responsible for the growth in threonine as the carbon source.     

Chemical forms of selenium for cancer prevention.

Cancer is becoming an increasingly significant disease worldwide. Currently, more than 7 million people die each year from cancer. With the existing knowledge, at least one-third of worldwide cancer cases could be prevented. Searching for naturally occurring agents in routinely consumed foods that may inhibit cancer development, although challenging, constitutes a valuable and plausible approach to the control and prevention of cancer. To date, the use of the micronutrient selenium (Se) in human clinical trials is limited, but the outcome indicates that Se is among the most promising agents. Although it is convenient to describe the effects of Se in terms of the element, it must always be kept in mind that the chemical form of Se and the dose are determinants of its biological activities. Hyphenated techniques based on coupling chromatographic separation with inductively coupled plasma mass spectrometric (ICP-MS) detection are now established as the most realistic and potent analytical tools available for real-life speciation analysis. These speciation investigations provide evidence that the Se compounds, which can generate monomethylated Se (e.g., Se-methylselenocysteine and methylseleninic acid), are more efficacious than other Se compounds because of their chemoprevention activity.     

Risk Score to Predict 1-Year Mortality after Haemodialysis Initiation in Patients with Stage 5 Chronic Kidney Disease under Predialysis Nephrology Care

Background

Few risk scores are available for predicting mortality in chronic kidney disease (CKD) patients undergoing predialysis nephrology care. Here, we developed a risk score using predialysis nephrology practice data to predict 1-year mortality following the initiation of haemodialysis (HD) for CKD patients.

Methods

This was a multicenter cohort study involving CKD patients who started HD between April 2006 and March 2011 at 21 institutions with nephrology care services. Patients who had not received predialysis nephrology care at an estimated glomerular filtration rate (eGFR) of approximately 10 mL/min per 1.73 m² were excluded. Twenty-nine candidate predictors were selected, and the final model for 1-year mortality was developed via multivariate logistic regression and was internally validated by a bootstrapping technique.

Results

A total of 688 patients were enrolled, and 62 (9.0%) patients died within one year of HD initiation. The following variables were retained in the final model: eGFR, serum albumin, calcium, Charlson Comorbidity Index excluding diabetes and renal disease (modified CCI), performance status (PS), and usage of erythropoiesis-stimulating agent (ESA). Their β-coefficients were transformed into integer scores: three points were assigned to modified CCI≥3 and PS 3–4; two to calcium>8.5 mg/dL, modified CCI 1–2, and no use of ESA; and one to albumin<3.5 g/dL, eGFR>7 mL/min per 1.73 m², and PS 1–2. Predicted 1-year mortality risk was 2.5% (score 0–4), 5.5% (score 5–6), 15.2% (score 7–8), and 28.9% (score 9–12). The area under the receiver operating characteristic curve was 0.83 (95% confidence interval, 0.79–0.89).

Conclusions

We developed a simple 6-item risk score predicting 1-year mortality after the initiation of HD that might help nephrologists make a shared decision with patients and families regarding the initiation of HD.