Regional differences in WT-1 and Tcf21 expression during ventricular development: implications for myocardial compaction

Background

Morphological and functional differences of the right and left ventricle are apparent in the adult human heart. A differential contribution of cardiac fibroblasts and smooth muscle cells (populations of epicardium-derived cells) to each ventricle may account for part of the morphological-functional disparity. Here we studied the relation between epicardial derivatives and the development of compact ventricular myocardium.

Results

Wildtype and Wt1^CreERT2/+ reporter mice were used to study WT-1 expressing cells, and Tcf21^lacZ/+ reporter mice and PDGFRα^-/-;Tcf21^LacZ/+ mice to study the formation of the cardiac fibroblast population. After covering the heart, intramyocardial WT-1+ cells were first observed at the inner curvature, the right ventricular postero-lateral wall and left ventricular apical wall. Later, WT-1+ cells were present in the walls of both ventricles, but significantly more pronounced in the left ventricle. Tcf21-^LacZ + cells followed the same distribution pattern as WT-1+ cells but at later stages, indicating a timing difference between these cell populations. Within the right ventricle, WT-1+ and Tcf21-lacZ+ cell distribution was more pronounced in the posterior inlet part. A gradual increase in myocardial wall thickness was observed early in the left ventricle and at later stages in the right ventricle. PDGFRα^-/-;Tcf21^LacZ/+ mice showed deficient epicardium, diminished number of Tcf21-^LacZ + cells and reduced ventricular compaction.

Conclusions

During normal heart development, spatio-temporal differences in contribution of WT-1 and Tcf21-^LacZ + cells to right versus left ventricular myocardium occur parallel to myocardial thickening. These findings may relate to lateralized differences in ventricular (patho)morphology in humans.     

Infection with Salmonella enterica Serovar Typhimurium Leads to Increased Proportions of F4/80+ Red Pulp Macrophages and Decreased Proportions of B and T Lymphocytes in the Spleen

Infection of mice with Salmonella enterica serovar Typhimurium (Salmonella) causes systemic inflammatory disease and enlargement of the spleen (splenomegaly). Splenomegaly has been attributed to a general increase in the numbers of phagocytes, lymphocytes, as well as to the expansion of immature CD71⁺Ter119⁺ reticulocytes. The spleen is important for recycling senescent red blood cells (RBCs) and for the capture and eradication of blood-borne pathogens. Conservation of splenic tissue architecture, comprised of the white pulp (WP), marginal zone (MZ), and red pulp (RP) is essential for initiation of adaptive immune responses to captured pathogens. Using flow cytometry and four color immunofluorescence microscopy (IFM), we show that Salmonella-induced splenomegaly is characterized by drastic alterations of the splenic tissue architecture and cell population proportions, as well as in situ cell distributions. A major cause of splenomegaly appears to be the significant increase in immature RBC precursors and F4/80⁺ macrophages that are important for recycling of heme-associated iron. In contrast, the proportions of B220⁺, CD4⁺ and CD8⁺ lymphocytes, as well as MZ MOMA⁺ macrophages decrease significantly as infection progresses. Spleen tissue sections show visible tears and significantly altered tissue architecture with F4/80⁺ macrophages and RBCs expanding beyond the RP and taking over most of the spleen tissue. Additionally, F4/80⁺ macrophages actively phagocytose not only RBCs, but also lymphocytes, indicating that they may contribute to declining lymphocyte proportions during Salmonella infection. Understanding how these alterations of spleen microarchitecture impact the generation of adaptive immune responses to Salmonella has implications for understanding Salmonella pathogenesis and for the design of more effective Salmonella-based vaccines.     

<Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> inhibits <Emphasis Type="Italic">in-vitro Candida</Emphasis> biofilm development

Background  

Elucidation of the communal behavior of microbes in mixed species biofilms may have a major impact on understanding infectious diseases and for the therapeutics. Although, the structure and the properties of monospecies biofilms and their role in disease have been extensively studied during the last decade, the interactions within mixed biofilms consisting of bacteria and fungi such as Candida spp. have not been illustrated in depth. Hence, the aim of this study was to evaluate the interspecies interactions of Pseudomonas aeruginosa and six different species of Candida comprising C. albicans, C. glabrata, C. krusei, C. tropicalis, C. parapsilosis, and C. dubliniensis in dual species biofilm development.     

Single-species microbial biofilm screening for industrial applications

While natural microbial biofilms often consist of multiple species, single-species biofilms are of great interest to biotechnology. The current study evaluates biofilm formation for common industrial and laboratory microorganisms. A total of 68 species of biosafety level one bacteria and yeasts from over 40 different genera and five phyla were screened by growing them in microtiter plates and estimating attached biomass by crystal violet staining. Most organisms showed biofilm formation on surfaces of polystyrene within 24 h. By changing a few simple conditions such as substratum characteristics, inoculum and nutrient availability, 66 strains (97%) demonstrated biofilm formation under at least one of the experimental conditions and over half of these strains were classified as strong biofilm formers, potentially suitable as catalysts in biofilm applications. Many non-motile bacteria were also strong biofilm formers. Biofilm morphologies were visualized for selected strains. A model organism, Zymomonas mobilis, easily established itself as a biofilm on various reactor packing materials, including stainless steel.     

Plasma proteome analysis in HTLV-1-associated myelopathy/tropical spastic paraparesis

Background

A new subgroup of HIV-1, designated Group P, was recently detected in two unrelated patients of Cameroonian origin. HIV-1 Group P phylogenetically clusters with SIVgor suggesting that it is the result of a cross-species transmission from gorillas. Until today, HIV-1 Group P has only been detected in two patients, and its degree of adaptation to the human host is largely unknown. Previous data have shown that pandemic HIV-1 Group M, but not non-pandemic Group O or rare Group N viruses, efficiently antagonize the human orthologue of the restriction factor tetherin (BST-2, HM1.24, CD317) suggesting that primate lentiviruses may have to gain anti-tetherin activity for efficient spread in the human population. Thus far, three SIV/HIV gene products (vpu, nef and env) are known to have the potential to counteract primate tetherin proteins, often in a species-specific manner. Here, we examined how long Group P may have been circulating in humans and determined its capability to antagonize human tetherin as an indicator of adaptation to humans.

Results

Our data suggest that HIV-1 Group P entered the human population between 1845 and 1989. Vpu, Env and Nef proteins from both Group P viruses failed to counteract human or gorilla tetherin to promote efficient release of HIV-1 virions, although both Group P Nef proteins moderately downmodulated gorilla tetherin from the cell surface. Notably, Vpu, Env and Nef alleles from the two HIV-1 P strains were all able to reduce CD4 cell surface expression.

Conclusions

Our analyses of the two reported HIV-1 Group P viruses suggest that zoonosis occurred in the last 170 years and further support that pandemic HIV-1 Group M strains are better adapted to humans than non-pandemic or rare Group O, N and P viruses. The inability to antagonize human tetherin may potentially explain the limited spread of HIV-1 Group P in the human population.     

A rapid and simple PCR analysis indicates there are two subgroups of Vibrio vulnificus which correlate with clinical or environmental isolation

Vibrio vulnificus is an estuarine bacterium which is the causative agent of both food-borne disease and wound infection. Although V. vulnificus is commonly found in molluscan shellfish at high numbers, the incidence of disease is relatively low, leading to the hypothesis that not all strains of V. vulnificus are equally virulent. Unfortunately, there is currently no easy test to identify virulent strains of this species. We have previously identified a 200 bp randomly amplified polymorphic DNA (RAPD) PCR amplicon associated with clinical isolates. DNA sequence data from this locus in six clinical and four environmental isolates showed that the strains could be divided into two groups, which we termed C-type (correlates with clinical origin) and E-type (correlates with environmental origin). We designed PCR primers that could distinguish between the two groups, and typed 55 randomly selected strains. We found that 90% of the C-type strains were clinical isolates, while 93% of environmental isolates were classified as E-type. The region directly downstream of this locus contained a heptanucleotide sequence repeated various times depending on the strain. Using a PCR-based assay to detect the repeat number present in a given strain, we found a statistically significant correlation with the C/E type classification and the number of repeats. The data reported here are consistent with the existence of two genotypes of V. vulnificus, with the C-type being a strong indicator of potential virulence.     

Development of a shuttle vector for Moraxella catarrhalis

Efforts to perform genetic analysis in Moraxella catarrhalis have been hampered by the lack of a cloning vector. M. catarrhalis strain E22 was previously shown to contain plasmid pLQ510 which lacked a selectable antibiotic resistance marker. Several methods were used to eliminate unnecessary DNA from pLQ510. Then, a 1.2 kb spectinomycin resistance cartridge, a multiple cloning site, and the origin of replication from pACYC184 were cloned into this plasmid backbone to obtain the 7.2 kb plasmid pWW102B. This new plasmid could replicate in M. catarrhalis as well as in both Escherichia coli and Haemophilus influenzae. This shuttle vector was used to clone and express two different M. catarrhalis genes, respectively, encoding an adhesin and a protein involved in serum resistance. When these two plasmids were introduced into appropriate M. catarrhalis mutants, they complemented the phenotypic deficiency of each mutant. This is the first report of functional complementation in trans in this pathogen.     

The accuracy of several multiple sequence alignment programs for proteins

Background  

There have been many algorithms and software programs implemented for the inference of multiple sequence alignments of protein and DNA sequences. The "true" alignment is usually unknown due to the incomplete knowledge of the evolutionary history of the sequences, making it difficult to gauge the relative accuracy of the programs.     

Compression or tension? The stress distribution in the proximal femur

Background  

Questions regarding the distribution of stress in the proximal human femur have never been adequately resolved. Traditionally, by considering the femur in isolation, it has been believed that the effect of body weight on the projecting neck and head places the superior aspect of the neck in tension. A minority view has proposed that this region is in compression because of muscular forces pulling the femur into the pelvis. Little has been done to study stress distributions in the proximal femur. We hypothesise that under physiological loading the majority of the proximal femur is in compression and that the internal trabecular structure functions as an arch, transferring compressive stresses to the femoral shaft.     

Ethylene Glycol Metabolism by Pseudomonas putida

In this study, we investigated the metabolism of ethylene glycol in the Pseudomonas putida strains KT2440 and JM37 by employing growth and bioconversion experiments, directed mutagenesis, and proteome analysis. We found that strain JM37 grew rapidly with ethylene glycol as a sole source of carbon and energy, while strain KT2440 did not grow within 2 days of incubation under the same conditions. However, bioconversion experiments revealed metabolism of ethylene glycol by both strains, with the temporal accumulation of glycolic acid and glyoxylic acid for strain KT2440. This accumulation was further increased by targeted mutagenesis. The key enzymes and specific differences between the two strains were identified by comparative proteomics. In P. putida JM37, tartronate semialdehyde synthase (Gcl), malate synthase (GlcB), and isocitrate lyase (AceA) were found to be induced in the presence of ethylene glycol or glyoxylic acid. Under the same conditions, strain KT2440 showed induction of AceA only. Despite this difference, the two strains were found to use similar periplasmic dehydrogenases for the initial oxidation step of ethylene glycol, namely, the two redundant pyrroloquinoline quinone (PQQ)-dependent enzymes PedE and PedH. From these results we constructed a new pathway for the metabolism of ethylene glycol in P. putida. Furthermore, we conclude that Pseudomonas putida might serve as a useful platform from which to establish a whole-cell biocatalyst for the production of glyoxylic acid from ethylene glycol.