The chemokine,CXCL16, and its receptor,CXCR6, are constitutively expressed in human annulus fibrosus and expression of CXCL16 is up-regulated by exposure to IL-1ß in vitro

Chemokines are an important group of soluble molecules with specialized functions in inflammation. The roles of many specialized chemokines and their receptors remain poorly understood in the human intervertebral disc. We investigated CXCL16 and its receptor, CXCR6, to determine their immunolocalization in disc tissue and their presence following exposure of cultured human annulus fibrosus cells to proinflammatory cytokines. CXCL16 is a marker for inflammation; it also can induce hypoxia-inducible factor 1α (HIF-1α), which is a phenotypic marker of heathy nucleus pulposus tissue. We found CXCL16 and CXCR6 immunostaining in many cells of the annulus portion of the disc. Molecular studies showed that annulus fibrosus cells exposed to IL-1ß, but not TNF-α, exhibited significant up-regulation of CXCL16 expression vs. control cells. There was no significant difference in the percentage of annulus cells that exhibited immunolocalization of CXCL16 in grade I/II, grade III or grade IV/V specimens. The presence of CXCL16 and its receptor, CXCR6, in the annulus in vivo suggests the need for future research concerning the role of this chemokine in proinflammatory functions, HIF-1α expression and disc vascularization.     

Geographical patterns of genetic variation in the world collections of wild annual Cicer characterized by amplified fragment length polymorphisms

Cicer reticulatum, C. echinospermum, C. bijugum, C. judaicum, C. pinnatifidum, C. cuneatum and C. yamashitae are wild annual Cicer species and potential donors of valuable traits to improve chickpea (C. arietinum). As part of a large project to characterize and evaluate wild annual Cicer collections held in the world gene banks, AFLP markers were used to study genetic variation in these species. The main aim of this study was to characterize geographical patterns of genetic variation in wild annual Cicer germplasm. Phylogenetic analysis of 146 wild annual Cicer accessions (including two accessions in the perennial C. anatolicum and six cultivars of chickpea) revealed four distinct groups corresponding well to primary, secondary and tertiary gene pools of chickpea. Some possible misidentified or mislabelled accessions were identified, and ILWC 242 is proposed as a hybrid between C. reticulatum and C. echinospermum. The extent of genetic diversity varied considerably and was unbalanced between species with greatest genetic diversity found in C. judaicum. For the first time geographic patterns of genetic variation in C. reticulatum, C. echinospermum, C. bijugum, C. judaicum and C. pinnatifidum were established using AFLP markers. Based on the current collections the maximum genetic diversity of C. reticulatum, C. echinospermum, C. bijugum and C. pinnatifidum was found in southeastern Turkey, while Palestine was the centre of maximum genetic variation for C. judaicum. This information provides a solid basis for the design of future collections and in situ conservation programs for wild annual Cicer.     

Paleoenvironmental history of the Popayán area since 27 000 yr BP at Timbio, Southern Colombia

A pollen record from Timbio, located at an elevation of 1750m on the high plain of Popayán (2 degrees 24'N, 76 degrees 36'W) is presented. This forms the basis for reconstructing the vegetation and climate history for the periods from 27000 to 9200 radiocarbon years before the present (14Cyr BP) and 2100 14Cyr BP to sub-recent. The 5m sediment core has time control based on seven AMS radiocarbon dates. Four pollen assemblage zones (TIM-1 to TIM-4) are recognized. During the period of 27200 to 26000 14Cyr BP, an Andean forest was near the site. The vegetation consisted of forest and open herb-rich vegetation, climatic conditions were moist and temperatures some 6 degrees C lower than compared to those of today. During the period of 26000 to 16000 14Cyr BP forest was less open. The observed succession from a Podocarpus-Weinmannia dominated forest to a Hedyosmum dominated forest, and finally to a forest with Ilex, Myrica and ferns indicates a progressive decrease of temperature during this period, with a maximum temperature depression of ca. 5-7.5 degrees C compared to present-day conditions. During the period of 16000 to 9200 14Cyr BP, temperature decrease is estimated at ca. 7.5 degrees C and the climate was the driest. During the period of 2100 to 600 14C2600m altitude (ca. 8 degrees C) and those at sea-level (2.5-6 degrees C) and supports the observation that glacial lapse rates were higher than in modern times.     

The vocal expression of feeding motivation and frustration in the domestic laying hen, Gallus gallus domesticus

Thwarting of feeding behaviour in the laying hen results in an increase in stereotyped pacing, displacement preening, and the gakel-call. These behaviours therefore reflect the frustration arousal caused by the thwarting of feeding behaviour. This raises the question whether the level of frustration also varies with the intensity of the motivation to perform the thwarted behaviour. This study investigated the relationship between the intensity of the motivation and level of frustration on the one hand and the gakel-call on the other hand. In Experiment 1, the strength of the motivation to feed was varied by thwarting hens in their feeding behaviour in an operant procedure after different durations of food deprivation (0, 8, 23 and 47 h). Trend analysis showed that with increasing hunger state, an increasing number of gakel-calls was given. No effect of treatments on temporal characteristics of the gakel-call was found. In Experiment 2, the level of frustration was varied by reducing or increasing the duration of access to food for food-deprived hens compared to the duration of access during training. It was assumed that a shorter duration of access to food compared to training would elicit frustration, which in turn would affect the performance of behaviours indicative of thwarting. However, we found neither a relation between the number of gakel-calls nor the temporal features of the gakel-call and the duration of access to food. Possibly, the differences between treatments were not large enough to induce differences in frustration level. Also, other factors that might have influenced the motivation are discussed.     

De novo production of the flavonoid naringenin in engineered Saccharomyces cerevisiae

Background

Specific strains of Lactobacillus plantarum are marketed as health-promoting probiotics. The role and interplay of cell-wall compounds like wall- and lipo-teichoic acids (WTA and LTA) in bacterial physiology and probiotic-host interactions remain obscure. L. plantarum WCFS1 harbors the genetic potential to switch WTA backbone alditol, providing an opportunity to study the impact of WTA backbone modifications in an isogenic background.

Results

Through genome mining and mutagenesis we constructed derivatives that synthesize alternative WTA variants. The mutants were shown to completely lack WTA, or produce WTA and LTA that lack D-Ala substitution, or ribitol-backbone WTA instead of the wild-type glycerol-containing backbone. DNA micro-array experiments established that the tarIJKL gene cluster is required for the biosynthesis of this alternative WTA backbone, and suggest ribose and arabinose are precursors thereof. Increased tarIJKL expression was not observed in any of our previously performed DNA microarray experiments, nor in qRT-PCR analyses of L. plantarum grown on various carbon sources, leaving the natural conditions leading to WTA backbone alditol switching, if any, to be identified. Human embryonic kidney NF-κB reporter cells expressing Toll like receptor (TLR)-2/6 were exposed to purified WTAs and/or the TA mutants, indicating that WTA is not directly involved in TLR-2/6 signaling, but attenuates this signaling in a backbone independent manner, likely by affecting the release and exposure of immunomodulatory compounds such as LTA. Moreover, human dendritic cells did not secrete any cytokines when purified WTAs were applied, whereas they secreted drastically decreased levels of the pro-inflammatory cytokines IL-12p70 and TNF-α after stimulation with the WTA mutants as compared to the wild-type.

Conclusions

The study presented here correlates structural differences in WTA to their functional characteristics, thereby providing important information aiding to improve our understanding of molecular host-microbe interactions and probiotic functionality.

     

X-linked adrenoleukodystrophy (X-ALD): clinical presentation and guidelines for diagnosis, follow-up and management

Background

Pompe disease (Glycogen storage disease type II, GSD II, acid alpha-glucosidase deficiency, acid maltase deficiency, OMIM # 232300) is an autosomal-recessive lysosomal storage disorder due to a deficiency of acid alpha-glucosidase (GAA, acid maltase, EC 3.2.1.20, Swiss-Prot P10253). Clinical manifestations are dominated by progressive weakness of skeletal muscle throughout the clinical spectrum. In addition, the classic infantile form is characterised by hypertrophic cardiomyopathy.

Methods

In a cross-sectional single-centre study we clinically assessed 3 patients with classic infantile Pompe disease and 39 patients with non-classic presentations, measured their acid alpha-glucosidase activities and analysed their GAA genes.

Results

Classic infantile patients had nearly absent residual enzyme activities and a typical clinical course with hypertrophic cardiomyopathy until the beginning of therapy. The disease manifestations in non-classic patients were heterogeneous. There was a broad variability in the decline of locomotive and respiratory function. The age of onset ranged from birth to late adulthood and correlated with enzyme activities. Molecular analysis revealed as many as 33 different mutations, 14 of which are novel. All classic infantile patients had two severe mutations. The most common mutation in the non-classic group was c.-32-13?T?>?G. It was associated with a milder course in this subgroup.

Conclusions

Disease manifestation strongly correlates with the nature of the GAA mutations, while the variable progression in non-classic Pompe disease is likely to be explained by yet unknown modifying factors. This study provides the first comprehensive dataset on the clinical course and the mutational spectrum of Pompe disease in Germany.     

Cell cycle population effects in perturbation studies

Growth condition perturbation or gene function disruption are commonly used strategies to study cellular systems. Although it is widely appreciated that such experiments may involve indirect effects, these frequently remain uncharacterized. Here, analysis of functionally unrelated Saccharyomyces cerevisiae deletion strains reveals a common gene expression signature. One property shared by these strains is slower growth, with increased presence of the signature in more slowly growing strains. The slow growth signature is highly similar to the environmental stress response (ESR), an expression response common to diverse environmental perturbations. Both environmental and genetic perturbations result in growth rate changes. These are accompanied by a change in the distribution of cells over different cell cycle phases. Rather than representing a direct expression response in single cells, both the slow growth signature and ESR mainly reflect a redistribution of cells over different cell cycle phases, primarily characterized by an increase in the G1 population. The findings have implications for any study of perturbation that is accompanied by growth rate changes. Strategies to counter these effects are presented and discussed.     

Biologically relevant effects of mRNA amplification on gene expression profiles

Background  

Gene expression microarray technology permits the analysis of global gene expression profiles. The amount of sample needed limits the use of small excision biopsies and/or needle biopsies from human or animal tissues. Linear amplification techniques have been developed to increase the amount of sample derived cDNA. These amplified samples can be hybridised on microarrays. However, little information is available whether microarrays based on amplified and unamplified material yield comparable results.     

High-level production of animal-free recombinant transferrin from Saccharomyces cerevisiae

Background

Microorganisms that are exposed to pollutants in the environment, such as metals/metalloids, have a remarkable ability to fight the metal stress by various mechanisms. These metal-microbe interactions have already found an important role in biotechnological applications. It is only recently that microorganisms have been explored as potential biofactories for synthesis of metal/metalloid nanoparticles. Biosynthesis of selenium (Se⁰) nanospheres in aerobic conditions by a bacterial strain isolated from the coalmine soil is reported in the present study.

Results

The strain CM100B, identified as Bacillus cereus by morphological, biochemical and 16S rRNA gene sequencing [GenBank:GU551935.1] was studied for its ability to generate selenium nanoparticles (SNs) by transformation of toxic selenite (SeO₃ ^2-) anions into red elemental selenium (Se⁰) under aerobic conditions. Also, the ability of the strain to tolerate high levels of toxic selenite ions was studied by challenging the microbe with different concentrations of sodium selenite (0.5 mM-10 mM). ESEM, AFM and SEM studies revealed the spherical Se⁰ nanospheres adhering to bacterial biomass as well as present as free particles. The TEM microscopy showed the accumulation of spherical nanostructures as intracellular and extracellular deposits. The deposits were identified as element selenium by EDX analysis. This is also indicated by the red coloration of the culture broth that starts within 2-3 h of exposure to selenite oxyions. Selenium nanoparticles (SNs) were further characterized by UV-Visible spectroscopy, TEM and zeta potential measurement. The size of nanospheres was in the range of 150-200 nm with high negative charge of -46.86 mV.

Conclusions

This bacterial isolate has the potential to be used as a bionanofactory for the synthesis of stable, nearly monodisperse Se⁰ nanoparticles as well as for detoxification of the toxic selenite anions in the environment. A hypothetical mechanism for the biogenesis of selenium nanoparticles (SNs) involving membrane associated reductase enzyme(s) that reduces selenite (SeO₃ ^2-) to Se⁰ through electron shuttle enzymatic metal reduction process has been proposed.