Duration of alpha 1-antichymotrypsin gene activation by interleukin-1 is determined by efficiency of inhibitor of nuclear factor kappa B alpha resynthesis in primary human astrocytes

Expression of alpha1antichymotrypsin (ACT) is significantly activated by interleukin-1 (IL-1) in human astrocytes; however, it is barely affected by IL-1 in hepatocytes. This tissue-specific regulation depends upon an enhancer that contains both nuclear factor kappaB (NF-kappaB) and activating protein 1 (AP-1) elements, and is also observed for an NF-kappaB reporter but not for an AP-1 reporter. We found efficient activation of NF-kappaB binding in both cell types; however, this binding was persistent in glial cells and only transient in hepatocytes. IL-1-activated NF-kappaB complexes consisted of p65 and p50, with p65 transiently phosphorylated on serine 536 in glial cells whereas more persistently in hepatic cells. Overexpression of p65 or constitutively active IKKbeta (inhibitor of NF-kappaB kinase beta) resulted in an efficient activation of the ACT reporter in hepatic cells, indicating that a specific mechanism exists in these cells terminating IL-1 signaling. IL-1 effectively induced the degradation of inhibitor of NF-kappaBalpha (IkBalpha) and IkBepsilon in both cell types but IkBbeta was not affected. However, IkBalpha was resynthesized much more rapidly in hepatic cells in comparison to glial cells. In addition, the initial levels of IkBalpha were much lower in glial cells. We propose that the tissue-specific regulation of the ACT gene expression by IL-1 is determined by different efficiencies of IkBalpha resynthesis in glial and hepatic cells.     

Astrocyte- and hepatocyte-specific expression of genes from the distal serpin subcluster at 14q32.1 associates with tissue-specific chromatin structures

The distal serpin subcluster contains genes encoding alpha1-antichymotrypsin (ACT), protein C inhibitor (PCI), kallistatin (KAL) and the KAL-like protein, which are expressed in hepatocytes, but only the act gene is expressed in astrocytes. We show here that the tissue-specific expression of these genes associates with astrocyte- and hepatocyte-specific chromatin structures. In hepatocytes, we identified 12 Dnase I-hypersensitive sites (DHSs) that were distributed throughout the entire subcluster, with the promoters of expressed genes accessible to restriction enzyme digestion. In astrocytes, only six DHSs were located exclusively in the 5' flanking region of the act gene, with its promoter also accessible to restriction enzyme digestion. The acetylation of histone H3 and H4 was found throughout the subcluster in both cell types but this acetylation did not correlate with the expression pattern of these serpin genes. Analysis of histone modifications at the promoters of the act and pci genes revealed that methylation of histone H3 on lysine 4 correlated with their expression pattern in both cell types. In addition, inhibition of methyltransferase activity resulted in suppression of ACT and PCI mRNA expression. We propose that lysine 4 methylation of histone H3 correlates with the tissue-specific expression pattern of these serpin genes.     

Expression of estrogen receptor {alpha} in the anteroventral periventricular nucleus of hypogonadal mice

Gonadotropin-releasing hormone-1 (GnRH-1) neurons play critical roles in the development and maintenance of reproductive function in all vertebrates. Due to a truncation in the GnRH-1 gene, hypogonadal (hpg) mice are unable to synthesize GnRH-1 and are infertile. These animals develop in the complete absence of exposure to gonadal steroid hormones, making them an interesting model for understanding brain sexual differentiation and dimorphism. We studied expression of the estrogen receptors (ERs) alpha and beta in the medial anteroventral periventricular nucleus (mAVPV), an important reproductive neuroendocrine brain region, in wild-type and hpg mice of both sexes. Adult wild-type and hpg mice of the same genetic background were used to quantify numbers of ERalpha and ERbeta immunoreactive cells in the mAVPV using a stereologic approach. Quantitative analyses showed that ERalpha cell numbers were significantly higher in hpg than wild-type mice, irrespective of sex. Qualitatively, ERalpha-immunoreactive cells were concentrated more densely along the ventricular zone of the AVPV of wild-type female mice compared with wild-type male mice or hpg male and female mice. No ERbeta-immunoreactive cells were detected in the mAVPV of any mice, a result that was surprising because ERbeta expression is abundant in the mAVPV of rats. These results on ERalpha provide additional evidence that the female brain is not the "default" organizational pattern, because neither ERalpha cell number nor its distribution in hpg mice, which develops with a deficiency of reproductive hormones, resembles that of the wild-type female mouse. Differences in ERalpha expression may be due in part to the absence of gonadal steroid hormones, although they more likely to also involve other factors, potentially GnRH itself.     

Cutting edge: human CD4+CD25+ T cells restrain the maturation and antigen-presenting function of dendritic cells

The characteristics and functions of CD4(+)CD25(+) regulatory cells have been well defined in murine and human systems. However, the interaction between CD4(+)CD25(+) T cells and dendritic cells (DC) remains unclear. In this study, we examined the effect of human CD4(+)CD25(+) T cells on maturation and function of monocyte-derived DC. We show that regulatory T cells render the DC inefficient as APCs despite prestimulation with CD40 ligand. This effect was marginally reverted by neutralizing Abs to TGF-beta. There was an increased IL-10 secretion and reduced expression of costimulatory molecules in DC. Thus, in addition to direct suppressor effect on CD4(+) T cells, regulatory T cells may modulate the immune response through DC.     

Involvement of a glycerol-3-phosphate dehydrogenase in modulating the NADH/NAD+ ratio provides evidence of a mitochondrial glycerol-3-phosphate shuttle in Arabidopsis

A mitochondrial glycerol-3-phosphate (G-3-P) shuttle that channels cytosolic reducing equivalent to mitochondria for respiration through oxidoreduction of G-3-P has been extensively studied in yeast and animal systems. Here, we report evidence for the operation of such a shuttle in Arabidopsis thaliana. We studied Arabidopsis mutants defective in a cytosolic G-3-P dehydrogenase, GPDHc1, which, based on models described for other systems, functions as the cytosolic component of a G-3-P shuttle. We found that the gpdhc1 T-DNA insertional mutants exhibited increased NADH/NAD+ ratios compared with wild-type plants under standard growth conditions, as well as impaired adjustment of NADH/NAD+ ratios under stress simulated by abscisic acid treatment. The altered redox state of the NAD(H) pool was correlated with shifts in the profiles of metabolites concerning intracellular redox exchange. The impairment in maintaining cellular redox homeostasis was manifest by a higher steady state level of reactive oxygen species under standard growth conditions and by a significantly augmented hydrogen peroxide production under stress. Loss of GPDHc1 affected mitochondrial respiration, particularly through a diminished capacity of the alternative oxidase respiration pathway. We propose a model that outlines potential involvements of a mitochondrial G-3-P shuttle in plant cells for redox homeostasis.     

Evaluation of the stress-inducible production of choline oxidase in transgenic rice as a strategy for producing the stress-protectant glycine betaine

Glycine betaine (GB) is a compatible solute that is also capable of stabilizing the structure and function of macromolecules. Several GB-producing transgenic rice lines were generated in which the Arthrobacter pascens choline oxidase (COX) gene, fused to a chloroplast targeting sequence (TP) was expressed under the control of an ABA-inducible promoter (SIP; stress-inducible promoter) or a ubiquitin (UBI) gene promoter that is considered to be constitutive. This comparison led to interesting observations that suggest complex regulation with respect to GB synthesis and plant growth response under stress. In spite of the use of the well-studied stress-inducible promoter, the highest level of GB accumulation (up to 2.60 micromol g(-1) DW) in the SIP lines grown under saline conditions was not as high as in the UBI lines (up to 3.12 micromol g(-1) DW). Therefore, the use of an ABA-inducible promoter was not more beneficial for de novo production of GB. Interestingly, saline growth conditions enhanced GB accumulation by up to 89% in the SIP lines, whereas up to 44% increase was seen in a UBI line. In all these cases the GB levels were many-fold below the range reported for plant species that produce GB naturally. In spite of lower GB concentrations, statistically greater levels of stress tolerance were found in SIP lines than in UBI lines, suggesting that the stress protection observed in SIP plants cannot be totally explained by the increase in the GB content.     

Nexplorer: phylogeny-based exploration of sequence family data

SUMMARY: Nexplorer is a web-based program for interactive browsing and manipulation of character data in NEXUS format, well suited for use with alignments and trees representing families of homologous genes or proteins. Users may upload a sequence family dataset, or choose from one of several thousand already available. Nexplorer provides a flexible means to develop customized views that combine a tree and a data matrix or alignment, to create subsets of data, and to output data files or publication-quality graphics. AVAILABILITY: Web access is from http://www.molevol.org/nexplorer     

Inhibition of calmodulin-dependent cyclic AMP phosphodiesterase by phenoxazines

Phenoxazine derivatives were examined for their ability to inhibit the calmodulin-mediated activation of phosphodiesterase, which is based on the hydrolysis of cAMP to AMP by phosphodiesterase in the presence or absence of inhibitor, followed by quantitative analysis by HPLC method. Anticalmodulin activity of phenoxazines with respect to substitution at C-2 position follows the order: 2-trifluoromethyl>2-chloro>unsubstituted phenoxazines. The interaction of phenoxazines with calmodulin using fluorescence spectroscopy has been performed. Binding study showed that calmodulin has two types of binding sites for phenoxazines. One is high affinity binding site (Kd value 0.07-0.46 microM) and the other, a low affinity binding site (Kd value 0.7-34.5 microM). The change in secondary structure of calmodulin upon binding to phenoxazines was studied by circular dichroism (CD) method, which showed that the percentage of helicity decreased with an extensive change in tertiary structure of calmodulin. Kinetic analysis of the phenoxazine-calmodulin interaction showed that phenoxazines competitively inhibited the activation of phosphodiesterase without affecting Vmax. Thus, these studies showed a good correlation between the ability of phenoxazines to block the activation of phosphodiesterase and their ability to bind to the activator.     

Addressing Women's Non-Maternal Healthcare Financing in Developing Countries: What Can We Learn from the Experiences of Rural Indian Women?

Background and Objectives

This paper focuses on the inadequate attention on women''s non-maternal healthcare in low- and middle-income countries. The study assessed the purchase of and financial access to non-maternal healthcare. It also scoped for mainstreaming household financial resources in this regard to suggest for alternatives.

Methods

A household survey through multi-stage stratified sampling in the state of Orissa interviewed rural women above 15 years who were neither pregnant nor had any pregnancy-related outcome six weeks preceding the survey. The questions explored on the processes, determinants and outcomes of health seeking for non-maternal ailments. The outcome measures were healthcare access, cost of care and financial access. The independent variables for bivariate and multivariate analyses were contextual factors, health seeking and financing pattern.

Results

The survey obtained a response rate of 98.64% and among 800 women, 43.8% had no schooling and 51% were above 60 years. Each woman reported at least one episode of non-maternal ailment; financial constraints prevented 68% from receiving timely and complete care. Distress coping measures (e.g. borrowings) dominated the financing source (67.9%) followed by community–based measures (32.1%). Only 6% had financial risk-protection; financial risk of not obtaining care doubled for women aged over 60 years (OR 2.00, 95% CI 0.84–4.80), seeking outpatient consultation (OR 2.01, 95% CI 0.89–4.81), facing unfavourable household response (OR 2.04, 95% CI 1.09–3.83), and lacking other financial alternatives (OR 2.13, 95% CI 1.11–4.07). When it comes to timely mobilization of funds and healthcare seeking, 90% (714) of the households preferred maternal care to non-maternal healthcare.

Conclusion

The existing financing options enable sub-optimal purchase of women''s non-maternal healthcare. Though dominant, household economy extends inadequate attention in this regard owing to its unfavourable approach towards non-maternal healthcare and limited financial capacity and support from other financial resources.