Binding of bile salts to pancreatic colipase and lipase.

The binding of conjugated bile salts to pancreatic colipase and lipase has been studied by equilibrium dialysis and gel filtration. The results indicate that at physiological ionic strength and pH, conjugated bile salts bind as micelles to colipase: 12-15 moles/mole of colipase for the dihydroxy conjugates and 2-4 for the trihydroxy conjugates. No binding of bile salt takes place from monomeric solutions. Under the same experimental conditions, only 1-2 moles of conjugated dihydroxy bile salts bind to pancreatic lipase.     

Accumulation of Natural Radionuclides in the Bottom Sediments and by Aquatic Organisms of Streams

The accumulation of natural radioisotopes by aquatic organisms and bottom sediments was studied in two small rivers, one uncontaminated and the other polluted by effluents of uranium ore mining and milling. Parabolic regressions between the water and uppermost sediment content of both uranium and total beta activity (corrected for ⁴⁰K content) is presented and demonstrates the water cumulative capacity of organically rich sediments. For ²²⁶Ra no significant differences between sediments with different organic content were found. The ²²⁶Ra content of bottom sediments is expressed as a power function of radium and calcium concentration in the water. In the given reaches, 53% and 85% of uranium and over 90% of ²²⁶Ra and total beta activity (corrected for ⁴⁰K) was accumulated in the upper two centimetres of sediments and biomass of aquatic vegetation. Filamentous algae, plankton, aquatic bryophyta and macrophyta from the present and other published data showed rather higher cumulation capacities as compared with bottom sediments.     

M234Glu is a component of the proton sponge in the reaction center from photosynthetic bacteria

Bacterial reaction centers use light energy to couple the uptake of protons to the successive semi-reduction of two quinones, namely Q_A and Q_B. These molecules are situated symmetrically in regard to a non-heme iron atom. Four histidines and one glutamic acid, M234Glu, constitute the five ligands of this atom. By flash-induced absorption spectroscopy and delayed fluorescence we have studied in the M234EH and M234EL variants the role played by this acidic residue on the energetic balance between the two quinones as well as in proton uptake. Delayed fluorescence from the P⁺Q_A^? state (P is the primary electron donor) and temperature dependence of the rate of P⁺Q_A^? charge recombination that are in good agreement show that in the two RC variants, both Q_A^? and Q_B^? are destabilized by about the same free energy amount: respectively ～ 100 ± 5 meV and 90 ± 5 meV for the M234EH and M234EL variants, as compared to the WT. Importantly, in the M234EH and M234EL variants we observe a collapse of the high pH band (present in the wild-type reaction center) of the proton uptake amplitudes associated with formation of Q_A^? and Q_B^?. This band has recently been shown to be a signature of a collective behaviour of an extended, multi-entry, proton uptake network. M234Glu seems to play a central role in the proton sponge-like system formed by the RC protein.     

Quantitative Properties of Random Amplified Polymorphic DNA (RAPD)

Random Amplified Polymorphic DNA analysis (RAPD) is a methodology that has been used as a tool for monitoring microbial communities. To be useful in this application RAPD, and any other methodology, must show properties that allows for the detection of quantitative changes in composition of the microbiota. Therefore, the objective of this study was to establish whether RAPD possesses such properties. The strategy was to use genomic DNA, extracted from a set of tertiary bacterial mixtures defined according to an experimental mixture design, and containing varying proportions of Escherichia coli, Bacillus subtilis, and Pseudomonas CF600. RAPD-PCR was performed on the mixed DNA extracts and the amplified DNA fragments were separated on sequencing gels to produce genomic fingerprints that were digitized and modeled by Partial Least Squares regression (PLS). Significant predictions were obtained using an external test set for validation, with Root Mean Square Error of Predictions (RMSEP) of 0.21, 0.19 and 0.20 for the proportion of E. coli, B. subtilis and Pseudomonas CF600 respectively. Taken together, the results showed that RAPD patterns quantitatively represented the initial mixture proportions. Therefore, the view that RAPD could be useful for whole microbial community monitoring was strengthened.     

Susceptibility of dopamine D5 receptor targeted mice to cysteamine

BACKGROUND: Recently we demonstrated that gastric mucosa of rats can synthesize, store and release dopamine. Out of five different subtypes, mRNA of D5 (=D1b) dopamine receptor is very abundant in the gastric epithelium. D1 receptor selective dopamine agonists have been shown to protect against experimental gastro-duodenal lesions. AIMS: To test the hypothesis that protective effects of dopamine involve D5 receptors, mucosal lesions were induced in D5 receptor deficient (KO) and wild-type (WT) mice using cysteamine. Morphology and gastric acid secretion of D5 KO mice were also studied. METHODS: Single doses of 600 mg/kg, 300 mg/kg cysteamine or vehicle were administered subcutaneously to fasted animals. After 24 h, number and severity of gastro-duodenal lesions were analyzed. Basal and histamine-induced maximal gastric acid output were measured by a stomach-sac wash-through method. RESULTS: All the KOs in the 600 mg/kg cysteamine group died within 4 h showing symptoms of toxicity while three out of four WTs survived (P<0.05). Mortality after 300 mg/kg cysteamine was significantly higher in KOs versus the WTs: 6/14 versus 2/11, P<0.05. Gastric lesion-index was also significantly higher in KOs (median, middle quartile): four (3-9) versus 0 (0-0), P<0.05. Duodenal lesions did not develop from this single dose of cysteamine in either genotype. Basal and histamine-induced maximal gastric acid output were comparable in the two genotypes. CONCLUSIONS: This study demonstrates that loss of D5 receptor causes mucosal vulnerability and increased toxicity of cysteamine in genetically manipulated mice. Thus, D5 receptor subtype is indeed likely to be involved in protective effects of dopamine in the stomach.     

Types of necroinflammation,the effect of cell death modalities on sterile inflammation

Distinct types of immune responses are activated by infections, which cause the development of type I, II, or III inflammation, regulated by Th1, Th2, Th17 helper T cells and ILC1, ILC2 and ILC3 cells, respectively. While the classification of immune responses to different groups of pathogens is widely accepted, subtypes of the immune response elicited by sterile inflammation have not yet been detailed. Necroinflammation is associated with the release of damage-associated molecular patterns (DAMP) from dying cells. In this review, we present that the distinct molecular mechanisms activated during apoptosis, necroptosis, pyroptosis, and ferroptosis lead to the release of different patterns of DAMPs and their suppressors, SAMPs. We summarize the currently available data on how regulated cell death pathways and released DAMPs and SAMPs direct the differentiation of T helper and ILC cells. Understanding the subtypes of necroinflammation can be crucial in developing strategies for the treatment of sterile inflammatory diseases caused by cell death processes.Subject terms: Necroptosis, Inflammasome     

Characterization of monoclonal antibodies for rapid identification of Actinomyces naeslundii in clinical samples

The purpose of this study was to generate highly specific serological reagents for the quantitative identification of Actinomyces naeslundii in clinical samples, in particular dental plaque. Balb/c mice were immunized with pasteurized human A. naeslundii strains representing different genospecies and serotypes. Ten hybrid cell lines secreting monoclonal antibodies reactive with A. naeslundii were isolated and characterized. Antibody specificity was determined by indirect immunofluorescence and enzyme-linked immunosorbent assay using strains from 59 species and by immunofluorescence analyses of supragingival plaque from 10 gingivitis patients. Nine monoclonal antibodies reacted selectively with A. naeslundii, whereas one additionally bound to Actinomyces israelii. They recognized at least nine different epitopes with characteristic expression patterns among the test strains. Six clusters of antigenically unique or closely related strains could be distinguished. Clusters 1, 4, and 5 represented by 12, 18, and 5 strains, respectively, comprised over 80% of the A. naeslundii strains tested. All reference strains for genospecies 1 grouped with cluster 1. Strains associated with genospecies 2 fell into clusters 4 and 5. Tests with mutant strains indicated that three monoclonal antibodies recognize type 2 and one type 1 fimbriae of genospecies 2. Only four isolates grouped with clusters 2 and 3 characterized by the expression of cluster-specific antigens. Interestingly, cluster 2 and 3 bacteria were markedly more abundant in vivo than indicated by their sparse representation in our strain collection. Overall, all but one of the new monoclonal antibodies should prove of value for the serological classification and rapid quantitative determination of A. naeslundii in clinical samples.     

Increased heart rate variability in mice overexpressing the Cu/Zn superoxide dismutase

Cu/Zn superoxide dismutase (SOD1) is implicated in various pathological conditions including Down's syndrome, neurodegenerative diseases, and afflictions of the autonomic nervous system (ANS). To assess the SOD1 contribution to ANS dysfunction, especially its influence on cardiac regulation, we studied the heart rate variability (HRV) and cardiac arrhythmias in conscious 12-month-old male and female transgenic mice for the human SOD1 gene (TghSOD1). TghSOD1 mice presented heart rate reduction as compared with control FVB/N individuals. All HRV parameters reflecting parasympathetic activity were increased in TghSOD1. Pharmacological studies confirmed that the parasympathetic tone was exacerbated and the sympathetic pathway was functional in TghSOD1 mice. A high frequency of atrioventricular block and premature ventricular contractions was observed in TghSOD1. By biochemical assays we found that SOD1 activities were multiplied by 9 and 4 respectively in the heart and brainstem of transgenic mice. A twofold decrease in cholinesterase activity was observed in the heart but not in the brainstem. We demonstrate that SOD1 overexpression induces an ANS dysfunction by an exacerbated vagal tone that may be related to impaired cardiac activity of the cholinesterases and may explain the high occurrence of arrhythmias.     

Postnatal gender-dependent maturation of cellular cysteine uptake

Background. In view of the functional capacity of glutathione synthesis in premature infants, and because the availability of cysteine is one the rate limiting steps in glutathione synthesis, we hypothesized that the low glutathione levels in premature infants may be due to immaturity of the active cellular uptake of cysteine.

Objective. To document in cells from newborn infants the effect of maturity and gender on cysteine uptake and consequently on glutathione levels.

Methods. Incorporation of L-[35S] cysteine was measured in leukocytes from cord blood and from tracheal aspirates (TAC) of newborn infants of varying (gestational as well as postnatal) ages and gender. Cysteine uptake was correlated with glutathione in TAC.

Results. The maturity of newborn girls positively influences cysteine uptake, which is responsible for 78% of the variation in their glutathione content. However, in newborn boys, gestational and postnatal ages did not influence the cysteine uptake.

Discussion. Cysteine uptake appears to be the limiting step explaining the reported gender-related differences in glutathione as well as the low levels of this central antioxidant found in premature infants. The immature cysteine uptake found in cells from premature infants raises questions about the bioavailability of this conditionally essential amino acid in regimens of parenteral nutrition for human neonates.