Convective exchange between the nose and the atmosphere

It is generally accepted that there is little rebreathing of gas exhaled through the nose. A detailed physical model system has been used to quantify and identify the mechanisms responsible for this phenomenon. By the use of a cast of the upper respiratory tract and oscillating flows with a Reynolds number of 500 and nondimensional frequency of 1.6, corresponding to quiet tidal breathing through the nose, dye dilution measurements indicated an efficiency of tidal exchange of 0.95. Flow visualization studies performed to trace the expiratory flow, as well as the streamlines during steady inspiratory flow, support the hypothesis that the high efficiency of exchange is due to radical differences in the velocity fields between inspiratory and expiratory phases of this oscillatory flow. These findings confirm that convective gas exchange between the nose and the atmosphere is highly efficient; however, the underlying mechanism responsible for this exchange also maximizes the exposure of the respiratory system to aerosols contained in the ambient atmosphere.     

The mechanisms controlling ribosomal protein L1 pre-mRNA splicing are maintained in evolution and rely on conserved intron sequences.

Sequences corresponding to the third intron of the X.laevis L1 ribosomal protein gene were isolated from the second copy of the X.laevis gene and from the single copy of X.tropicalis. Sequence comparison revealed that the three introns share an unusual sequence conservation which spans a region of 110 nucleotides. In addition, they have the same suboptimal 5' splice sites. The three introns show similar features upon oocyte microinjection: they have very low splicing efficiency and undergo the same site specific cleavages which lead to the accumulation of truncated molecules. Computer analysis and RNAse digestions have allowed to assign to the conserved region a specific secondary structure. Mutational analysis has shown that this structure is important for conferring the cleavage phenotype to these three introns. Competition experiments show that the cleavage phenotype can be prevented by coinjection of excess amounts of homologous sequences.     

Cytokine gene polymorphisms in endemic pemphigus foliaceus: a possible role for IL6 variants

Endemic pemphigus foliaceus (EPF) is a complex autoimmune disease characterized by the presence of antibodies against desmoglein 1, which lead to the loss of adhesion among keratinocytes (acantholysis). Variants of HLA class II genes have been the only genetic factors found to modulate susceptibility to EPF. This study aims at investigating the influence of cytokine genetic variants in the pathogenesis of EPF, since they may affect the expression levels of these immunomodulatory molecules. The sample included 168 patients and 189 controls and was comprised of mostly Caucasoids and Mulattos. The approach consisted of a case-control association study and the alleles were identified by mismatched PCR-RFLP. No associations were found with variants of IL1A, IL1B, IL1RN, IL4R and IL10. There was a weak negative association with the haplotype -1082G -592C (OR=0.49) of the IL10 gene in Mulattos. In regard to polymorphism -590 of the IL4 gene, a positive association with the T/T genotype (OR=2.71) and a negative association with the C variant (OR=0.37) were found. Associations with IL6 -174 variants suggest that the C/C genotype has a protective effect (OR=0.13) while carriers of the G allele are more susceptible (OR=7.66) to EPF.     

Development of novel plasmid vectors and a promoter trap system in Francisella tularensis compatible with the pFLN10 based plasmids

Francisella tularensis is a category A bioterror pathogen which in some cases can cause a severe and fatal human infection. Very few virulence factors are known in this species due to the difficulty in working with it as well as the lack of tools for genetic manipulation. This work describes the construction of a shuttle vector that can replicate in Escherichia coli and F. tularensis as well as two distinct promoter trap constructs based on the shuttle vector backbone. Replication in F. tularensis is based on the promiscuous origin of replication from the Staphylococcus aureus plasmid pC194. We demonstrate the novel plasmids can coexist with established F. tularensis vectors based on the pFNL10 plasmid, the current workhorse of F. tularensis genetics. Our promoter trap can identify promoters that are activated during intracellular growth and survival. These new vectors provide additional tools for the genetic manipulation of F. tularensis.     

Type III Secretion System and Virulence Markers Highlight Similarities and Differences between Human- and Plant-Associated Pseudomonads Related to Pseudomonas fluorescens and P. putida

Pseudomonas fluorescens is commonly considered a saprophytic rhizobacterium devoid of pathogenic potential. Nevertheless, the recurrent isolation of strains from clinical human cases could indicate the emergence of novel strains originating from the rhizosphere reservoir, which could be particularly resistant to the immune system and clinical treatment. The importance of type three secretion systems (T3SSs) in the related Pseudomonas aeruginosa nosocomial species and the occurrence of this secretion system in plant-associated P. fluorescens raise the question of whether clinical isolates may also harbor T3SSs. In this study, isolates associated with clinical infections and identified in hospitals as belonging to P. fluorescens were compared with fluorescent pseudomonads harboring T3SSs isolated from plants. Bacterial isolates were tested for (i) their genetic relationships based on their 16S rRNA phylogeny, (ii) the presence of T3SS genes by PCR, and (iii) their infectious potential on animals and plants under environmental or physiological temperature conditions. Two groups of bacteria were delineated among the clinical isolates. The first group encompassed thermotolerant (41°C) isolates from patients suffering from blood infections; these isolates were finally found to not belong to P. fluorescens but were closely related and harbored highly conserved T3SS genes belonging to the Ysc-T3SS family, like the T3SSs from P. aeruginosa. The second group encompassed isolates from patients suffering from cystic fibrosis; these isolates belonged to P. fluorescens and harbored T3SS genes belonging to the Hrp1-T3SS family found commonly in plant-associated P. fluorescens.     

Structure-guided design of peptide-based tryptase inhibitors

Improved peptide-based inhibitors of human beta tryptase were discovered using information gleaned from tripeptide library screening and structure-guided design methods, including fragment screening. Our efforts sought to improve this class of inhibitors by replacing the traditional Lys or Arg P1 element. The optimized compounds display low nanomolar potency against the mast cell target and several hundred-fold selectivity with respect to serine protease off targets. Thus, replacement of Lys/Arg at P1 in a peptide-like scaffold does not need to be accompanied by a loss in target affinity.