ZK91587: a novel synthetic antimineralocorticoid displays high affinity for corticosterone (type I) receptors in the rat hippocampus

In vitro cytosol binding assays have shown the properties of binding of a novel steroid, ZK91587 (15 beta, 16 beta-methylene-mexrenone) in the brain of rats. Scatchard and Woolf analyses of the binding data reveal the binding of [3H] ZK91587 to the total hippocampal corticosteroid receptor sites with high affinity (Kd 1.9 nM), and low capacity (Bmax 17.3 fmol/mg protein). When 100-fold excess RU28362 was included simultaneously with [3H] ZK91587, the labelled steroid binds with the same affinity (Kd 1.8 nM) and capacity (Bmax 15.5 fmol/mg protein). Relative binding affinities (RBA) of various steroids for the Type I or Type II corticosteroid receptor in these animals are: Type I: ZK91587 = corticosterone (B) greater than cortisol (F); Type II: B greater than F much greater than ZK91587. In the binding kinetic study, ZK91587 has a high association rate of binding in the rat (20.0 x 10(7) M-1 min-1). The steroid dissociates following a one slope pattern (t 1/2 30 h), indicating, the present data demonstrate that in the rat hippocampus, ZK91587 binds specifically to the Type I (corticosterone-preferring/mineralocorticoid-like) receptor.     

The amino acid sequences of two alpha chains of hemoglobins from Komodo dragon Varanus komodoensis and phylogenetic relationships of amniotes

To elucidate phylogenetic relationships among amniotes and the evolution of alpha globins, hemoglobins were analyzed from the Komodo dragon (Komodo monitor lizard) Varanus komodoensis, the world's largest extant lizard, inhabiting Komodo Islands, Indonesia. Four unique globin chains (alpha A, alpha D, beta B, and beta C) were isolated in an equal molar ratio by high performance liquid chromatography from the hemolysate. The amino acid sequences of two alpha chains were determined. The alpha D chain has a glutamine at E7 as does an alpha chain of a snake, Liophis miliaris, but the alpha A chain has a histidine at E7 like the majority of hemoglobins. Phylogenetic analyses of 19 globins including two alpha chains of Komodo dragon and ones from representative amniotes showed the following results: (1) The a chains of squamates (snakes and lizards), which have a glutamine at E7, are clustered with the embryonic alpha globin family, which typically includes the alpha D chain from birds; (2) birds form a sister group with other reptiles but not with mammals; (3) the genes for embryonic and adult types of alpha globins were possibly produced by duplication of the ancestral alpha gene before ancestral amniotes diverged, indicating that each of the present amniotes might carry descendants of the two types of alpha globin genes; (4) squamates first split off from the ancestor of other reptiles and birds.      

The Effect of Three-Monthly Albendazole Treatment on Malarial Parasitemia and Allergy: A Household-Based Cluster-Randomized,Double-Blind,Placebo-Controlled Trial

Background

Helminth infections are proposed to have immunomodulatory activities affecting health outcomes either detrimentally or beneficially. We evaluated the effects of albendazole treatment, every three months for 21 months, on STH, malarial parasitemia and allergy.

Methods and Findings

A household-based cluster-randomized, double-blind, placebo-controlled trial was conducted in an area in Indonesia endemic for STH. Using computer-aided block randomization, 481 households (2022 subjects) and 473 households (1982 subjects) were assigned to receive placebo and albendazole, respectively, every three months. The treatment code was concealed from trial investigators and participants. Malarial parasitemia and malaria-like symptoms were assessed in participants older than four years of age while skin prick test (SPT) to allergens as well as reported symptoms of allergy in children aged 5–15 years. The general impact of treatment on STH prevalence and body mass index (BMI) was evaluated. Primary outcomes were prevalence of malarial parasitemia and SPT to any allergen. Analysis was by intention to treat. At 9 and 21 months post-treatment 80.8% and 80.1% of the study subjects were retained, respectively. The intensive treatment regiment resulted in a reduction in the prevalence of STH by 48% in albendazole and 9% in placebo group. Albendazole treatment led to a transient increase in malarial parasitemia at 6 months post treatment (OR 4.16(1.35–12.80)) and no statistically significant increase in SPT reactivity (OR 1.18(0.74–1.86) at 9 months or 1.37 (0.93–2.01) 21 months). No effect of anthelminthic treatment was found on BMI, reported malaria-like- and allergy symptoms. No adverse effects were reported.

Conclusions

The study indicates that intensive community treatment of 3 monthly albendazole administration for 21 months over two years leads to a reduction in STH. This degree of reduction appears safe without any increased risk of malaria or allergies.

Trial Registration

Controlled-Trials.com ISRCTN83830814     

Reduced transforming growth factor-beta signaling in cartilage of old mice: role in impaired repair capacity

Osteoarthritis (OA) is a common joint disease, mainly effecting the elderly population. The cause of OA seems to be an imbalance in catabolic and anabolic factors that develops with age. IL-1 is a catabolic factor known to induce cartilage damage, and transforming growth factor (TGF)-beta is an anabolic factor that can counteract many IL-1-induced effects. In old mice, we observed reduced responsiveness to TGF-beta-induced IL-1 counteraction. We investigated whether expression of TGF-beta and its signaling molecules altered with age. To mimic the TGF-beta deprived conditions in aged mice, we assessed the functional consequence of TGF-beta blocking. We isolated knee joints of mice aged 5 months or 2 years, half of which were exposed to IL-1 by intra-articular injection 24 h prior to knee joint isolation. Immunohistochemistry was performed, staining for TGF-beta1, -2 or -3, TGF-betaRI or -RII, Smad2, -3, -4, -6 and -7 and Smad-2P. The percentage of cells staining positive was determined in tibial cartilage. To mimic the lack of TGF-beta signaling in old mice, young mice were injected with IL-1 and after 2 days Ad-LAP (TGF-beta inhibitor) or a control virus were injected. Proteoglycan (PG) synthesis (³⁵S-sulfate incorporation) and PG content of the cartilage were determined. Our experiments revealed that TGF-beta2 and -3 expression decreased with age, as did the TGF-beta receptors. Although the number of cells positive for the Smad proteins was not altered, the number of cells expressing Smad2P strongly dropped in old mice. IL-1 did not alter the expression patterns. We mimicked the lack of TGF-beta signaling in old mice by TGF-beta inhibition with LAP. This resulted in a reduced level of PG synthesis and aggravation of PG depletion. The limited response of old mice to TGF-beta induced-IL-1 counteraction is not due to a diminished level of intracellular signaling molecules or an upregulation of intracellular inhibitors, but is likely due to an intrinsic absence of sufficient TGF-beta receptor expression. Blocking TGF-beta distorted the natural repair response after IL-1 injection. In conclusion, TGF-beta appears to play an important role in repair of cartilage and a lack of TGF-beta responsiveness in old mice might be at the root of OA development.     

Landscape-scale variation in the seed banks of floodplain wetlands with contrasting hydrology in China

1. At a local scale, the species composition, diversity and spatial variation of wetland plant communities are determined primarily by spatial and temporal heterogeneity in their environments. Less is known about variation at a landscape‐level. The floodplain of the Changjiang (Yangtze) River in China includes hydrologically connected, subtropical wetlands with different hydrological characteristics. 2. We examined seed‐bank species composition and richness in marshes of two contrasting hydrological types: permanent marshes, fed by local runoff, and lakeshore marshes more closely connected to the regulated river. Lakeshore marshes are flooded annually to depth of approximately 1 m and during flooding they support an alternate, aquatic vegetation type. The soil seed bank in March was a comparative estimator of species diversity. At the beginning of the growing season it included seeds from both phases of alternating vegetation types associated with the annual hydrological cycle. 3. A regional pool of 101 species was detected in the seed banks of six wetlands associated with the river and its tributaries: 56 occurred in permanent marshes and 59 in lakeshore marshes, with only 15 common to both. Species rarefaction curves indicated that more species occurred in permanent than lakeshore marshes at equal numbers of individuals sampled. However, the more heterogeneous lakeshore seed banks were estimated (Chao 2) to have greater total species richness (81) than permanent marsh (60). 4. Analysis using Sørensen's coefficient of similarity and DCA ordination revealed complex variation, with much greater differences between hydrological types than within them, irrespective of geographical distance. The types also differed significantly in the composition of four functional groups of species. 5. Despite the potential for dispersal of propagules via the annually pulsing river system (hydrochory), at a regional and landscape scale, diversity is maintained largely by large‐scale temporal hydrological heterogeneity and smaller scale spatial and topographic heterogeneity.     

Functional characterization of a three-component regulatory system involved in quorum sensing-based regulation of peptide antibiotic production in Carnobacterium maltaromaticum

Background  

Quorum sensing is a form of cell-to-cell communication that allows bacteria to control a wide range of physiological processes in a population density-dependent manner. Production of peptide antibiotics is one of the processes regulated by quorum sensing in several species of Gram-positive bacteria, including strains of Carnobacterium maltaromaticum. This bacterium and its peptide antibiotics are of interest due to their potential applications in food preservation. The molecular bases of the quorum sensing phenomenon controlling peptide antibiotic production in C. maltaromaticum remain poorly understood. The present study was aimed at gaining a deeper insight into the molecular mechanism involved in quorum sensing-mediated regulation of peptide antibiotic (bacteriocin) production by C. maltaromaticum. We report the functional analyses of the CS (autoinducer)-CbnK (histidine protein kinase)-CbnR (response regulator) three-component regulatory system and the three regulated promoters involved in peptide antibiotic production in C. maltaromaticum LV17B.     

Floral synorganization and its influence on mechanical isolation and autogamy in Marantaceae

The flowers of Marantaceae (～ 550 species) exhibit a highly derived pollination mechanism within Zingiberales, with a rapid and irreversible style movement based on a close synorganization of different floral parts. Given the complexity of the structure, we assume that little variation is possible if functionality is to be maintained. To test this, we investigated how much floral diversity exists in the clade and whether this diversity potentially influences the breeding system and placement of pollen on the pollinator. Flowers of 66 species covering the five major phylogenetic clades of the family were analysed. All species are similar in their basic flower construction: the fleshy staminode forms the tunnel‐shaped roof of the flower and narrows the tube with stiff swellings, and the hooded staminode holds the style under tension and narrows the flower entrance with its trigger appendage. Despite morphological diversity of the pollination apparatus, functionality is maintained by coordinated variation of the fleshy and hooded staminodes. Autogamy is usually avoided by herkogamy. However, in a few exceptions, subtle morphological changes alter the breeding system from allogamy to autogamy. Variable floral proportions allow for differential pollen deposition potentially causing mechanical isolation between sister taxa. This study clearly illustrates that structural variation is not only present in the highly synorganized flowers of Marantaceae, but that it also creates potentially new options for evolutionary diversification. © 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2012, 168 , 300–322.     

Immunolocalization of MMP-19 in the human intervertebral disc: implications for disc aging and degeneration

Matrix metalloproteinases (MMPs) degrade components of the extracellular matrix of the disc, but the presence of MMP-19 has not been explored. In other tissues, MMP-19 is known to act in proteolysis of the insulin-like growth factor (IGF) binding protein-3, thereby exposing this protein to make it available to influence cell behavior. MMP-19 also has been shown to inhibit capillary-like formation and thus play a role in the avascular nature of the disc. Using immunohistochemistry, normal discs from six subjects aged newborn through 10 years and 20 disc specimens from control donors or surgical patients aged 15-76 (mean age 40.2 years) were examined for immunolocalization of MMP-19; six Thompson grade I discs, five Thompson grade II, eight Thompson grade III, five Thompson grade IV, and one Thompson grade V discs were analyzed. The results indicate that in discs from young subjects, MMP-19 was uniformly localized in the outer annulus. In discs from adult donors and surgical patients, outer and inner annulus cells only occasionally showed MMP-19 localization. The greatest expression of MMP-19 was observed in young discs, and little expression was seen in older or degenerating discs. Because MMP-19 has been shown to regulate IGF-mediated proliferation in other tissues, its decline in the aging/degenerating disc may contribute to the age-related decrease in disc cell numbers.     

Brain corticosteroid receptor gene expression and neuroendocrine dynamics during aging

The present study examined the stress responsiveness of the hypothalamic-pituitary-adrenal axis in relation to the properties of corticosteroid receptors in the brain and pituitary in old (30 months) and young (3 months) male Brown Norway rats. The data demonstrate that circulating ACTH rather than the corticosteroid plasma level was elevated under basal conditions and following stress. Furthermore, a reduction of mineralocorticoid receptor (MR) number in the hippocampus and of glucocorticoid receptor (GR) number in the hypothalamus and the pituitary correspond to increased neuroendocrine responsiveness and negative feedback following stress. The changes in receptor binding do not parallel the changes in the amount of MR and GR mRNA measured with in situ hybridization. This suggests that the processing rather than the receptor gene expression is affected in senescence.     

Regulation of vasopressin messenger RNA levels in the small cell lung carcinoma cell line GLC-8: interactions between glucocorticoids and second messengers.

The role of glucocorticoids and second messenger systems in the regulation of the vasopressin (VP) gene was studied in the human small cell lung carcinoma cell line GLC-8. Small cell lung carcinoma GLC-8 cells express VP mRNA and contain both glucocorticoid and mineralocorticoid receptors. Treatment with the synthetic glucocorticoid dexamethasone when added alone at 10(-8) M had no effect on the VP mRNA level and decreased the level by 30% at 10(-6) M. However, the effect of dexamethasone changed to positive when cells were simultaneously treated with cAMP-enhancing agents. VP mRNA levels, which were elevated by 1.5- to 2-fold by the cAMP-enhancing agents alone, increased a further 1.5- to 3-fold by dexamethasone. Thus, the combined effect of dexamethasone and cAMP stimulation was a 3- to 7.5-fold increase in VP mRNA levels. Long term treatment with the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA) reduced the VP mRNA level by 75%. The TPA-suppressed VP mRNA levels could be up-regulated about 6-fold by simultaneous treatment with 8-bromo-cAMP. Dexamethasone did not alter the TPA-suppressed VP mRNA levels. These results indicate that both cAMP and protein kinase-C pathways as well as glucocorticoid receptors are involved in the regulation of VP mRNA levels and that these factors interact. This leads to a negative or positive response of VP gene expression to glucocorticoids in a state-dependent manner. The interactions may be of significance in a physiological context and relate to the different regulation of VP-expressing systems in the brain.