Study of adhesion and survival of lactobacilli and bifidobacteria on table olives with the aim of formulating a new probiotic food

With the aim of developing new functional foods, a traditional product, the table olive, was used as a vehicle for incorporating probiotic bacterial species. Survival on table olives of Lactobacillus rhamnosus (three strains), Lactobacillus paracasei (two strains), Bifidobacterium bifidum (one strain), and Bifidobacterium longum (one strain) at room temperature was investigated. The results obtained using a selected olive sample demonstrated that bifidobacteria and one strain of L. rhamnosus (Lactobacillus GG) showed a good survival rate, with a recovery of about 10(6) CFU g(-1) after 30 days. The Lactobacillus GG population remained unvaried until the end of the experiment, while a slight decline (to about 10(5) CFU g(-1)) was observed for bifidobacteria. High viability, with more than 10(7) CFU g(-1), was observed throughout the 3-month experiment for L. paracasei IMPC2.1. This strain, selected for its potential probiotic characteristics and for its lengthy survival on olives, was used to validate table olives as a carrier for transporting bacterial cells into the human gastrointestinal tract. L. paracasei IMPC2.1 was recovered from fecal samples in four out of five volunteers fed 10 to 15 olives per day carrying about 10(9) to 10(10) viable cells for 10 days.     

Exporting calcium from cells

All eukaryotic cells import Ca2+ through a number of variously gated plasma membrane channels. Once inside cells, Ca2+ transmits information to a large number of (enzyme) targets. Eventually, it must be exported again, to prevent the overloading of the cytosol with Ca2+. Two systems export Ca2+ from cells: a high affinity, low capacity Ca2+-ATPase, and a lower affinity, but much larger capacity, Na+/Ca2+ exchanger. The ATPase (commonly called the Ca2+ pump) is the fine-tuner of cell Ca2+, as it functions well even if the concentration of the ion drops below the microM level. It is a large enzyme, with 10 transmembrane domains and a C-terminal cytosolic tail that contains regulatory sites, including a calmodulin-binding domain. Four distinct gene products plus a large number of splice variants have been described. Some are tissue specific, the isoform 2 being specifically expressed in the sensorial cells of the Corti organ in the inner-ear. Its genetic absence causes deafness in mice. Two different families of the Na+/Ca2+ exchanger exist, one of which, originally described in photoreceptors, transports K+ and Ca2+ in exchange for Na+. The exchanger is particularly active in excitable cells, e.g., heart, where the necessity cyclically arises to rapidly eject large amounts of Ca2+. In addition to heart, the exchanger is particularly important to neurons: the cleavage of the most important neuronal isoform (NCX3) by calpains activated by excitotoxic treatments generates Ca2+ overload and eventually cell death.     

Specific targeting of hepatitis C virus NS3 RNA helicase. Discovery of the potent and selective competitive nucleotide-mimicking inhibitor QU663

Hepatitis C virus (HCV) infection is an emerging global epidemic, and no effective cure is yet available. Interferon-alpha (INFalpha) and pegylated INFs, in combination or otherwise with ribavirin, have proven to be effective in no more than 50% of chronically infected patients. New and better therapeutic strategies are therefore needed. HCV nonstructural protein 3 (NS3) RNA helicase (h) is a promising target for developing new therapeutics. QU663 was discovered as a potent new selective inhibitor of the helicase reaction of HCV NS3 (K(i) = 0.75 microM), competing with the nucleic acid substrate without affecting ATPase function, even at high concentrations. QU663 is one of a new generation of small-molecule nucleotide-mimicking inhibitors which are potential anti-HCV agents. A thorough molecular modeling study was carried out to explain the molecular basis of NS3h inhibition by QU663. The resulting three-dimensional interaction model is discussed.     

Synthesis, biological evaluation and molecular modelling studies on benzothiadiazine derivatives as PDE4 selective inhibitors

A series of 2,1,3- and 1,2,4-benzothiadiazine derivatives (BTDs) were synthesized and evaluated for their inhibitory activity versus enzymatic isoforms PDE3, PDE4 and PDE7. The compounds characterized by the 3,5-di-tert-butyl-4-hydroxybenzyl moiety at N1 position of 2,1,3-benzothiadiazine core (8, 13, 18), were found active and selective at micromolar level versus PDE4 and could be studied as new leads for the treatment of asthma and COPD (Chronic Obstructive Pulmonary Disease). The antioxidant activity evaluation on the same compounds highlighted 13 as the most significative. Molecular modelling studies gave further support to biological results and suggested targeted modifications so as to improve their potency.     

Vitamin C inhibits platelet expression of CD40 ligand

Upon stimulation with agonists, platelets express CD40 ligand (CD40L), a transmembrane protein implicated in the initiation and progression of atherosclerotic disease. We have recently discovered that oxidative stress plays a major role in platelet CD40L expression. In this study, we sought to determine whether vitamin C, a known antioxidant, is able to influence platelet CD40L expression. In vitro experiments were done by stimulating platelets with collagen in the presence or absence of vitamin C (50–100 μM) or vehicle as control. An in vivo study was done in 10 healthy subjects who were randomized to intravenous infusion of placebo or 1 g vitamin C for 45 min in a crossover design. At the end of infusion platelet CD40L and O_2- were measured. The in vitro study demonstrated that vitamin C dose dependently inhibited platelet CD40L expression without affecting agonist-induced platelet aggregation. In subjects treated with placebo no changes of platelet CD40L and O_2- were observed; conversely, vitamin C infusion caused a significant and parallel decrease of platelet O_2- (−70%, P < 0.001) and CD40L (−68%, P < 0.001). Platelet aggregation was not modified by either treatment. This study suggests that water-soluble antioxidants, which scavenge superoxide radicals, may reduce platelet CD40L expression.     

Hypoglycosylation with increased fucosylation and branching of serum transferrin N-glycans in untreated galactosemia

Untreated classic galactosemia (galactose-1-phosphate uridyltransferase [GALT] deficiency) is known as a secondary congenital disorders of glycosylation (CDG) characterized by galactose deficiency of glycoproteins and glycolipids (processing defect or CDG-II). The mechanism of this undergalactosylation has not been established. Here we show that in untreated galactosemia, there is also a partial deficiency of whole glycans of serum transferrin associated with increased fucosylation and branching as seen in genetic glycosylation assembly defects (CDG-I). Thus galactosemia seems to be a secondary "dual" CDG causing a processing as well as an assembly N-glycosylation defect. We also demonstrated that in galactosemia patients, transferrin N-glycan biosynthesis is restored upon dietary treatment.     

Meiothermus timidus sp. nov., a new slightly thermophilic yellow-pigmented species

Several yellow-pigmented isolates, with optimum growth temperatures between 55 and 60 degrees C, were recovered from hot springs in Central Portugal and the Azores. Phylogenetic analysis of the 16S rDNA showed that these organisms represented a new species of the genus Meiothermus. The new isolates could be distinguished from other strains of the species of the genus Meiothermus by biochemical characteristics and the fatty acid composition because they had very high levels of iso C15:0 and iso C17:0 and very low levels of anteiso C17:0 and iso C16:0. On the basis of the results presented here we propose the name Meiothermus timidus for the new species represented by strains SPS-243(T) (=LMG 22897(T)=CIP 108604(T)), RQ-10 and RQ-12.     

Worldwide genomic diversity of the high-risk human papillomavirus types 31, 35, 52, and 58, four close relatives of human papillomavirus type 16

Among the more than one hundred formally described human papillomavirus (HPV) types, 18 are referred to as high-risk HPV types due to their association with anogenital cancer. Despite pathogenic similarities, these types form three remotely related taxonomic groups. One of these groups is called HPV species 9 and is formed by HPV-16, the most common and best-studied type, together with HPV-31, -33, -35, -52, -58, and -67. Previous worldwide comparisons of HPV-16 samples showed about 2% nucleotide diversity between isolates, which were subsequently termed variants. The distribution of divergent variants has been found to correlate frequently with the geographic origin and the ethnicity of the infected patients and led to the concept of unique African, European, Asian, and Native American HPV-16 variants. In the current study, we address the question of whether geography and ethnicity also correlate with sequence variations found for HPV-31, -35, -52, and -58. This was done by sequencing the long control region in samples derived from Europe, Asia, and Africa, and from immigrant populations in North and South America. We observed maximal divergence between any two variants within each of these four HPV types ranging from 1.8 to 3.6% based on nucleotide exchanges and, occasionally, on insertions and deletions. Similar to the case with HPV-16, these mutations are not random but indicate a relationship between the variants in form of phylogenetic trees. An interesting example is presented by a 16-bp insert in select variants of HPV-35, which appears to have given rise to additional variants by nucleotide exchanges within the insert. All trees showed distinct phylogenetic topologies, ranging from dichotomic branching in the case of HPV-31 to star phylogenies of the other three types. No clear similarities between these types or between these types and HPV-16 exist. While variant branches in some types were specific for Europe, Africa, or East Asia, none of the four trees reflected human evolution and spread to the extent illustrated by HPV-16. One possible explanation is that the rare HPV types that we studied spread and thereby diversified more slowly than the more abundant HPV-16 and may have established much of today's variant diversity already before the worldwide spread of humans 100,000 years ago. Most variants had prototypic amino acid sequences within the E6 oncoprotein and a segment of the L1 capsid protein. Some had one, two, or three amino acid substitutions in these regions, which might indicate biological and pathogenic diversity between the variants of each HPV type.