Tenascin in the human intervertebral disc: alterations with aging and disc degeneration

Our objective for this study was to determine the presence and distribution of tenascin in the human intervertebral disc. The tenascins are a family of extracellular matrix proteins with repeated structural domains homologous to epidermal growth factor, fibronectin type III and the fibrinogens. Little is known about the presence of this protein in the disc. Ten normal human discs donated from subjects newborn to 15 years old, 10 control discs from adult donors aged 24-41 years, and 11 surgical disc specimens from patients aged 26-76 years were examined for immunolocalization of tenascin. In young discs, tenascin was localized throughout the annulus; in the nucleus, localization was confined to pericellular matrix. In adult control and degenerating disc specimens, tenascin in the annulus was localized primarily in pericellular matrix regions encircling either single cells or clusters of disc cells; in rare instances localization was more diffuse in the intraterritorial matrix. In young, healthy disc, tenascin was abundant throughout the annulus. In contrast, degenerating discs in adults showed a localization restricted to the pericellular, and rarely, more restricted intraterritorial matrix. These observations indicate that changes in the amount and distribution of tenascin may have a role in disc aging and degeneration, possibly by modulating fibronectin-disc-cell interactions, and causing alterations in the shape of disc cells.     

A re-evaluation of the Macrothrix rosea-triserialis group, with the description of two new species (Crustacea Anomopoda: Macrothricidae)

We redescribe Macrothrix rosea (females and males) based on material collected in Belgium. We also compare seven populations of Macrothrix `triserialis' from different parts of the world, including a topotypical population of M. triserialis s. str. from Sri Lanka, and males from South India (here first described), relying heavily on the structure of the trunk limbs, beside classical features of morphology. M. rosea and M. triserialis are extremely closely related: males are easily separated, but the identification of females requires micro-characters such as the relative length of the apical segment of the setae natatoriae and the adornment of the first antenna and of the longest swimming seta of the antenna. M. rosea and triserialis together constitute a sub-group of the rosea-group. Macrothrix triserialis-like animals occur in the tropical–subtropical belts of four continents. We compare populations from Asia, South America and Africa, and find differences in microcharacters of the trunk limbs, but cannot decide whether these represent random variation or sound taxonomical differences. One of the basic characters of the Macrothrix rosea-triserialis subgroup is that the setae natatoriae of the postabdomen are implanted on a prominence, the `heel'. Other characters include the fact that the Fryer' forks are adorned with one or two big teeth only, and that the scrapers of trunk limb two form a row of eight without any doublings. Possibly, scraper five, and scraper four to a lesser degree has an enlarged subapical tooth. The exopodite of trunk limb three has four plumose setae, the back and front row of the endopodite six setae and/or receptors, the exopodite of trunk limb four has two setae, and the back row of the endopodite six setae, plus one on the gnathobase. The pre-epipodite of trunk limb five consists of three lobes, the `endopodite' is small, and the `exopodite' is reduced to a single seta. The male postabdomen has a tubular ending, without true end-claws, although a rudiment of an end-claw is seen in M. triserialis. Two new species are described: M. tabrizensis and M. agsensis. A comparison, including the males of Macrothrix triserialis, M. rosea, M. smirnovi and M. tabrizensis confirms the relationship of all these taxa, but also reveals a morphological series in the shape of the postabdomen, from a complete absence of end-claws, over rudiments of a pair of end-claws, to complete endclaws. Absence of end-claws is here considered to represent an evolved character state. Macrothrix smirnovi Ciros & Elías (1987) is less closely related to the rosea-triserialis group, and is considered to form a sub-group in its own right. It shows a short `heel' on the postabdomen, but carries a supplementary seta behind scraper 4 of the endopodite of trunk limb two, and has a male with a postabdomen that closely resembles that of the female. These are primitive characters, which are also found in Wlassicsia, Bunops and Onchobunops and provide a possible phylogenetic link between Macrothrix and these three genera, although the genetic distance between them is considered to be quite large.     

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.

     

Host range of Gonatocerus sp. near tuberculifemur ‘Clade 1’ in Argentina, an egg parasitoid newly associated to the glassy-winged sharpshooter, Homalodisca vitripennis (Hemiptera: Cicadellidae), and candidate for its biological control in California, USA

The South American egg parasitoid Gonatocerus sp. near tuberculifemur “Clade 1” (G. sp. “Clade 1”) (Hymenoptera: Mymaridae) is a new association of the glassy-winged sharpshooter (GWSS), Homalodisca vitripennis (Germar) (Cicadellidae) and a candidate for its biological control in California, USA. In Argentina, G. sp. “Clade 1” was screened in the laboratory (no-choice tests) and in the field (multiple choice tests) against eggs of 32 Auchenorrhyncha host species and other four potential hosts unrelated to sharpshooters. In no-choice assays, it parasitized only eggs within the leafhopper tribe Proconiini. In contrast, in the long term field tests, it emerged not only from eggs of the Proconiini but also from two species of Cicadellini at low numbers (five wasps out of 698 exposed eggs). Two interpretations arise from the results: (1) Host associations of G. sp. “Clade 1” are restricted to the Proconiini whereas field parasitization of the Cicadellini species were false positive, or (2) G. sp. “Clade 1” parasitizes also some Cicadellini species and its rejection in the laboratory was a false negative. Both interpretations are discussed. Insect motivation could be the explanation for the negative results in the no-choice tests. On the other hand, in the more natural field situations, the host selection process and oviposition behavior should not have been affected and host range would be more realistic. The parasitism of the Cicadellini species would be indicative of a potential non-target effect on the sharpshooters in the USA.     

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.