Utilizing QCM-D to characterize the adhesive mucilage secreted by two marine diatom species in-situ and in real-time

The quartz crystal microbalance with dissipation monitoring (QCM-D) was used to monitor the deposition of adhesive extracellular polymeric substances (EPS) employed by the marine biofouling diatoms Craspedostauros australis Cox and Amphora coffeaeformis Cleve during initial adhesion and subsequent motility. Upon injection into the QCM chamber, initial negative frequency (f) shifts and positive dissipation (D) shifts were measured that correlated to cells impacting and adhering to the QCM sensor surface. Following this "initial adhesion" response, f continued to decrease while D increased logarithmically. Rather than the result of any cell morphological alterations at the substrate surface, the shifts were correlated to the time-dependent deposition of EPS upon the substrate surface as a result of cellular motility, or gliding. Experiments utilizing comparable cell concentrations of the diatom species C. australis and A. coffeaeformis revealed significant differences between the parameter responses recorded, where A. coffeaeformis produced Deltaf and DeltaD values of -32 Hz and 6.6, and C. australis produced values of -82 Hz and 42, respectively, after 20 h post-inoculation. The viscoelastic properties of the adhered EPS adlayer from both species were examined via a Deltaf/DeltaD plot, providing reproducible signature "ratio" values for each species that likely correlate to differences in EPS interactions with the substrate that may be associated directly to differences in the fouling potential of the two species. There is a distinct lack of knowledge regarding the chemical nature of the adhesive polymers engaged, and few quantitative techniques are applicable to the study of diatom EPS. We propose that QCM-D may be a useful tool in identifying differences in the EPS employed by diatoms of different fouling potential.     

Sexual dimorphism in age-related changes in UCP2 and leptin gene expression in subcutaneous adipose tissue in humans

The influence of age and gender on uncoupling protein 2 (UCP2) expression and its relationship with leptin expression in subcutaneous adipose tissue has been studied in humans. Samples of subcutaneous adipose tissue were obtained from 41 adult subjects (20 women and 21 men), with an age range of 28 to 84 years, and body mass index (BMI) of 19 to 36 Kgm(minus sign2). UCP2 and leptin mRNA expression was determined by northern blot. In women, both leptin and UCP2 expression in the subcutaneous adipose tissue increased significantly with age (r = 0.490 p < 0.05 and r = 0.475 p < 0.05, respectively). In men, in contrast, a negative correlation was found between leptin expression and age (r = minus sign0.678 p < 0.001), while no significant correlation was apparent between UCP2 expression and age (r = minus sign0.077). In addition, there was a positive correlation between UCP2 and leptin expression in women (r = 0.656 p < 0.01). These data show important gender dependent differences in the age-related changes in leptin and UCP2 expression in subcutaneous adipose tissue in humans.     

Structure-functional analysis of the Dictyoglomus cell envelope

Several closely related strains of the thermophilic bacterium Dictyoglomus have been isolated from various hot springs on the Philippine archipelago. These strains as well as Dictyoglomus thermophilum H-6-12 were analyzed in view of the structure-functional relationships of the cell envelopes. All envelopes of Dictyoglomus strains show several peculiar features that are apparently either unique for the genus or common for other phylogenetically related Thermotogales. The filamentous cells develop pili at the cell poles, guided by large columnar protein assemblies that traverse the periplasm. Filamentous protein complexes span the periplasmic space at the longitudinal sides of the cell. By the end of the exponential growth phase, Dictyoglomus strains form multicellular aggregates ("rotund bodies") inside a compartment surrounded by a single, continuous outer envelope. The formation of these rotund bodies which are also found in some other deeply branching thermophilic phyla (Thermotoga, Thermus) was studied in detail. The transition between unicellular and multicellular lifestyle can be explained by the partial detachment of a protoplast from the outer envelope during cell division. When the outer envelope is partially detached from the protoplast, mechanical forces generated by protoplast elongation may drive cell rearrangement of daughter cells inside the compartment. During the following rounds of cell division, the overall shape of the compartment changes from spindle-like to globular geometry. Analysis of subcellular fractions of Dictyoglomus cells shows that glucan hydrolases are associated with the compartment. This feature is discussed in view of the multicellular life style of Dictyoglomus.     

Phylogeography and population dynamics of dengue viruses in the Americas

Changes in Dengue virus (DENV) disease patterns in the Americas over recent decades have been attributed, at least in part, to repeated introduction of DENV strains from other regions, resulting in a shift from hypoendemicity to hyperendemicity. Using newly sequenced DENV-1 and DENV-3 envelope (E) gene isolates from 11 Caribbean countries, along with sequences available on GenBank, we sought to document the population genetic and spatiotemporal transmission histories of the four main invading DENV genotypes within the Americas and investigate factors that influence the rate and intensity of DENV transmission. For all genotypes, there was an initial invasion phase characterized by rapid increases in genetic diversity, which coincided with the first confirmed cases of each genotype in the region. Rapid geographic dispersal occurred upon each genotype's introduction, after which individual lineages were locally maintained, and gene flow was primarily observed among neighboring and nearby countries. There were, however, centers of viral diversity (Barbados, Puerto Rico, Colombia, Suriname, Venezuela, and Brazil) that were repeatedly involved in gene flow with more distant locations. For DENV-1 and DENV-2, we found that a "distance-informed" model, which posits that the intensity of virus movement between locations is inversely proportional to the distance between them, provided a better fit than a model assuming equal rates of movement between all pairs of countries. However, for DENV-3 and DENV-4, the more stochastic "equal rates" model was preferred.     

Chemical structure requirements and cellular targeting of microRNA-122 by peptide nucleic acids anti-miRs

Anti-miRs are oligonucleotide inhibitors complementary to miRNAs that have been used extensively as tools to gain understanding of specific miRNA functions and as potential therapeutics. We showed previously that peptide nucleic acid (PNA) anti-miRs containing a few attached Lys residues were potent miRNA inhibitors. Using miR-122 as an example, we report here the PNA sequence and attached amino acid requirements for efficient miRNA targeting and show that anti-miR activity is enhanced substantially by the presence of a terminal-free thiol group, such as a Cys residue, primarily due to better cellular uptake. We show that anti-miR activity of a Cys-containing PNA is achieved by cell uptake through both clathrin-dependent and independent routes. With the aid of two PNA analogues having intrinsic fluorescence, thiazole orange (TO)-PNA and [bis-o-(aminoethoxy)phenyl]pyrrolocytosine (BoPhpC)-PNA, we explored the subcellular localization of PNA anti-miRs and our data suggest that anti-miR targeting of miR-122 may take place in or associated with endosomal compartments. Our findings are valuable for further design of PNAs and other oligonucleotides as potent anti-miR agents.     

Central representation of spatial and temporal surface wave parameters in the African clawed frog

Xenopus laevis employs mechano-sensory lateral lines to, for instance, capture arthropods on the surface of turbid waters with poor visibility based on incoming wave signals. To characterise central representations of surface waves emitted from different locations, responses to several wave parameters were extracellularly recorded across brainstem, midbrain and thalamic areas. Overall, 339 of 411 statistically analysed responses showed significantly altered spike rates during the presentation of surface waves. Of these units, 45.1?% were obtained in the torus semicircularis including its laminar subnucleus (23.3?%) that is known to process auditory cues. Wave parameters contributing to central object representations were indicated by response rates that systematically varied with amplitude (76.3?% of 160 tested units), frequency (74.4?% of 270 tested units), source angle (93.7?% of 79 tested units), or source distance (63.8?% of 218 tested units). Map-like parameter representations were rather diffuse, yet an increased fraction of units tuned to frontal source angles was observed at deeper tissue layers (>180?μm), and an increased fraction of best neuronal responses to low wave frequencies (≤25?Hz) at rostral midbrain sections. Responses to wave frequencies remained largely robust across tested unit samples independent of source angles, and distances (N?=?62). In comparison, spatial response characteristics seemed fragile across different wave frequencies in 68.3?% of 41 recordings.     

Renal IL-17 expression in human ANCA-associated glomerulonephritis

Interleukin-17A (IL-17) promotes inflammatory renal tissue damage in mouse models of crescentic glomerulonephritis, including murine experimental autoimmune anti-myeloperoxidase glomerulonephritis, which most likely depends on IL-17-producing Th17 cells. In human anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, however, the cellular sources of IL-17 remain to be elucidated. Therefore, we analyzed human kidney biopsies of active necrotizing and crescentic ANCA-associated glomerulonephritis by immunohistochemistry using an IL-17-specific antibody and by immunofluorescent colocalization with cell type markers. We detected numerous IL-17-expressing (IL-17(+)) cells in the glomeruli and in the tubulointerstitium. Unexpectedly, most of these IL-17(+) cells were polymorphonuclear neutrophilic granulocytes, while IL-17(+) T cells and IL-17(+) mast cells were present at significantly lower frequencies. IL-17 was not detected in other infiltrating or resident kidney cells. In those patients who had not received immunosuppressive treatment before biopsy, serum creatinine levels were positively correlated with tubulointerstitial IL-17(+) neutrophils as well as IL-17(+) T cells. Furthermore, we could demonstrate that purified human blood neutrophils expressed IL-17 protein and released it upon stimulation in vitro. In conclusion, these results support a pathogenic role for IL-17 in human ANCA-associated glomerulonephritis. Our data suggest that in the acute stage of the disease neutrophils may act as an important immediate-early innate source of IL-17 and may thereby initiate and promote ongoing renal inflammation. IL-17 may thus be a target for treating acute ANCA-associated glomerulonephritis.     

Protective role for CCR5 in murine lupus nephritis

Leukocyte infiltration is a characteristic feature of human and experimental lupus nephritis and is closely correlated with loss of renal function. The chemokine receptor CCR5 is expressed on monocyte and T cell subsets and is thought to play an important role in recruiting these cells into inflamed organs. To investigate the functional role of CCR5 in lupus nephritis, CCR5-deficient mice were backcrossed onto the lupus-prone MRL-Fas(lpr) (MRL/lpr) genetic background. Unexpectedly, CCR5(-/-) MRL/lpr mice developed an aggravated course of lupus nephritis in terms of glomerular tissue injury and albuminuria. Deterioration of the nephritis was associated with an overall increase in mononuclear cell infiltration into the kidney, whereas renal leukocyte subtype balance, systemic T cell response, and autoantibody formation were unaffected by CCR5 deficiency. Renal and systemic protein levels of the CCR5 ligand CCL3, which can also attract leukocytes via its alternate receptor CCR1, were significantly increased in nephritic CCR5(-/-) MRL/lpr mice. Further studies revealed that the systemic increase in the CCR5/CCR1 ligand is also observed in nonimmune CCR5(-/-) C57BL/6 mice and that this increase was due to a reduced clearance, rather than an overproduction, of CCL3. Taken together, our data support the hypothesis that CCR5-dependent consumption of its own ligands may act as a negative feedback loop to restrain local chemokine levels within inflamed tissues, thereby limiting inflammatory cell influx.