A novel colonic repressor element regulates intestinal gene expression by interacting with Cux/CDP

Intestinal gene regulation involves mechanisms that direct temporal expression along the vertical and horizontal axes of the alimentary tract. Sucrase-isomaltase (SI), the product of an enterocyte-specific gene, exhibits a complex pattern of expression. Generation of transgenic mice with a mutated SI transgene showed involvement of an overlapping CDP (CCAAT displacement protein)-GATA element in colonic repression of SI throughout postnatal intestinal development. We define this element as CRESIP (colon-repressive element of the SI promoter). Cux/CDP interacts with SI and represses SI promoter activity in a CRESIP-dependent manner. Cux/CDP homozygous mutant mice displayed increased expression of SI mRNA during early postnatal development. Our results demonstrate that an intestinal gene can be repressed in the distal gut and identify Cux/CDP as a regulator of this repression during development.     

Reactivity of new adhesion molecules on lymphocytes from patients with chronic graft versus host disease

Reaction patterns of the 7th Human Leukocyte Differentiation Antigen Workshop blind panel adhesion molecules were studied on CD3/CD4, CD3/CD8, CD3/TCR gamma delta double positive T cells from peripheral blood of patients with chronic graft versus host disease (n = 8) and healthy controls (n = 4). Reactivity of 14 adhesion antibodies was tested by three-colour immunophenotyping. The mean proportion of CD3+ T cells (69 +/- 19%). CD3/CD8++ (31 +/- 13%) and CD3/TCR gamma delta++ (4 +/- 2%) T sub-populations of patients were comparable with the healthy controls. However, the mean percentage of CD3/CD4++ T cell subset in patients (14 +/- 12%) proved to be significantly decreased in comparison with the normal control value (34 +/- 16%) presumably due to secondary immunodeficiency. The workshop antibodies proved to be reactive with three T cell subsets expressing the examined antigens. Based on the results of the adhesion molecule workshop new CD categories have been introduced: CD156b as a transmembrane protein, CD167a as an epithelial tyrosin kinase receptor, CD168 as a receptor for hyaluronan mediated motility (RHAMM) and CD171 as a co-stimulatory adhesion molecule. There were significant differences in the expression of the CD167a and CD156b antigens on the CD3/CD4++ subset between the samples of patients compared with the controls characterizing the CD4+ T lymphocyte subpopulation in chronic graft versus host disease.     

Removal and destruction of high concentrations of gaseous toluene in a two-phase partitioning bioreactor by Alcaligenes xylosoxidans

A two-phase bioreactor consisting of hexadecane dispersed in an aqueous, cell-containing medium (organic fraction = 0.33) was used to trap toluene vapours from an air stream. The affinity for toluene by the solvent resulted in high efficiency of removal and transfer to the aqueous phase based on equilibrium transfer. The system was readily able to handle a loading capacity of 748 mg l^–1 h^–1 at a toluene degradation efficiency of greater than 98%.     

Increased nitric oxide levels as an early sign of premature aging in diabetes

The levels of different reactive species, especially those of nitric oxide and peroxynitrite, were determined in streptozotocin-induced diabetic rat tissues, before the development of histopathological damages. Significantly higher steady state free radical concentrations were found in the liver 3 weeks after the onset of diabetes compared to age-matched control groups. Increased nitric oxide levels in diabetic vasculature and kidney decreased the production of detectable reactive oxygen species. High peroxynitrite generation suggested the onset of processes characteristic to premature aging of the endothelium. According to the histopathological results, there were no signs of late complications in the tissues up to 7 weeks after induction of diabetes. These results support the idea that oxidative stress is increased at a very early stage of diabetes and, in particular, that high levels of nitric oxide and peroxynitrite could play a decisive role in the development of late complications in the diabetic vasculature and kidney.     

The Nac2 gene of Chlamydomonas encodes a chloroplast TPR-like protein involved in psbD mRNA stability

The psbD mRNA, which encodes the D2 reaction center polypeptide of photosystem II, is one of the most abundant chloroplast mRNAs. We have used genomic complementation to isolate the nuclear Nac2 gene, which is required for the stable accumulation of the psbD mRNA in Chlamydomonas reinhardtii. Nac2 encodes a hydrophilic polypeptide of 1385 amino acids with nine tetratricopeptide-like repeats (TPRs) in its C-terminal half. Cell fractionation studies indicate that the Nac2 protein is localized in the stromal compartment of the chloroplast. It is part of a high molecular weight complex that is associated with non-polysomal RNA. Change of a conserved alanine residue of the fourth TPR motif by site-directed mutagenesis leads to aggregation of Nac2 protein and completely abrogates its function, indicating that this TPR is important for proper folding of the protein and for psbD mRNA stability, processing and/or translation.     

Sonic Hedgehog Controls the Phenotypic Fate and Therapeutic Efficacy of Grafted Neural Precursor Cells in a Model of Nigrostriatal Neurodegeneration

The expression of soluble growth and survival promoting factors by neural precursor cells (NPCs) is suggested to be a prominent mechanism underlying the protective and regenerative effects of these cells after transplantation. Nevertheless, how and to what extent specific NPC-expressed factors contribute to therapeutic effects is not well understood. Using RNA silencing, the current study investigated the roles of two donor NPC molecules, namely glial cell-line derived neurotrophic factor (GDNF) and sonic hedgehog (SHH), in the protection of substantia nigra dopamine neurons in rats treated with 6-hydroxydopamine (6-OHDA). Analyses indicate that as opposed to the knock-down of GDNF, SHH inhibition caused a profound decline in nigrostriatal neuroprotection. Further, SHH silencing also curbed endogenous neurogenesis and the migration of host brdU⁺/dcx⁺ neural precursors into the striatum, which was present in the animals receiving control or GDNF silenced NPCs. A change in graft phenotype, mainly reflected by a reduced proportion of undifferentiated nestin⁺ cells, as well as a significantly greater host microglial activity, suggested an important role for these processes in the attenuation of neuroprotection and neurogenesis upon SHH silencing. Overall these studies reveal core mechanisms fundamental to grafted NPC-based therapeutic effects, and delineate the particular contributions of two graft-expressed molecules, SHH and GDNF, in mediating midbrain dopamine neuron protection, and host plasticity after NPC transplantation.     

Segregation distortion and linkage analysis in eggplant (Solanum melongena L.)

An anther-derived doubled haploid (DH) population and an F2 mapping population were developed from an intraspecific hybrid between the eggplant breeding lines 305E40 and 67/3. The former incorporates an introgressed segment from Solanum aethiopicum Gilo Group carrying the gene Rfo-sa1, which confers resistance to Fusarium oxysporum; the latter is a selection from an intraspecific cross involving two conventional eggplant varieties and lacks Rfo-sa1. Initially, 28 AFLP primer combinations (PCs) were applied to a sample of 93 F2 individuals and 93 DH individuals, from which 170 polymorphic AFLP fragments were identified. In the DH population, the segregation of 117 of these AFLPs as well as markers closely linked to Rfo-sa1 was substantially distorted, while in the F2 population, segregation distortion was restricted to just 10 markers, and thus the latter was chosen for map development. A set of 141 F2 individuals was genotyped with 73 AFLP PCs (generating 406 informative markers), 32 SSRs, 4 tomato RFLPs, and 3 CAPS markers linked to Rfo-sa1. This resulted in the assignment of 348 markers to 12 major linkage groups. The framework map covered 718.7?cM, comprising 238 markers (212 AFLPs, 22 SSRs, 1 RFLP, and the Rfo-sa1 CAPS). Marker order and inter-marker distances in this eggplant map were largely consistent with those reported in a recently published SSR-based map. From an eggplant breeding perspective, DH populations produced by anther culture appear to be subject to massive segregation distortion and thus may not be very efficient in capturing the full range of genetic variation present in the parental lines.     

Population dynamics of Empetrum hermaphroditum (Ericaceae) on a subarctic sand dune: Evidence of rapid colonization through efficient sexual reproduction

The importance of sexual reproduction for clonal plant species has long been underestimated, perhaps as a consequence of the difficulty in identifying individuals, preventing the study of their population dynamics. Such is the case for Empetrum hermaphroditum, an ericaceous species, which dominates the ground vegetation of subarctic ecosystems. Despite abundant seed production, seedlings are rarely observed. Therefore, prevalent seedling recruitment on a subarctic dune system provided an opportunity to study the population dynamics and spatial pattern of the colonization phase of this species. We established a 6-ha grid on the dune systems that extended from the shoreline to the fixed dunes and mapped and measured all E. hermaphroditum individuals in the grid. Moreover, we sampled 112 individuals just outside the grid to identify any allometric relationship between the size and age of the individuals, which allowed us to reconstruct population expansion. The overall size structure suggests that the population is still expanding. In the last 50 yr, E. hermaphroditum advanced more than 200 m in the dune system. Expansion started in the 1960s simultaneously at different distances from the shoreline. Colonization did not proceed gradually from the fixed dune toward the shoreline but instead individuals established earlier in the troughs between the dunes, with an increasingly clumped spatial pattern as the population filled in with time.     

Developing oligodendrocytes express functional GABA(B) receptors that stimulate cell proliferation and migration

GABA(B) receptors (GABA(B)Rs) are involved in early events during neuronal development. The presence of GABA(B)Rs in developing oligodendrocytes has not been established. Using immunofluorescent co-localization, we have identified GABA(B)R proteins in O4 marker-positive oligodendrocyte precursor cells (OPCs) in 4-day-old mouse brain periventricular white matter. In culture, OPCs, differentiated oligodendrocytes (DOs) and type 2 astrocytes (ASTs) express both the GABA(B1abcdf) and GABA(B2) subunits of the GABA(B)R. Using semiquantitative PCR analysis with GABA(B)R isoform-selective primers we found that the expression level of GABA(B1abd) was substantially higher in OPCs or ASTs than in DOs. In contrast, the GABA(B2) isoform showed a similar level of expression in OPCs and DOs, and a significantly higher level in ASTs. This indicates that the expression of GABA(B1) and GABA(B2) subunits are under independent control during oligodendroglial development. Activation of GABA(B)Rs using the selective agonist baclofen demonstrated that these receptors are functionally active and negatively coupled to adenylyl cyclase. Manipulation of GABA(B)R activity had no effect on OPC migration in a conventional agarose drop assay, whereas baclofen significantly increased OPC migration in a more sensitive transwell microchamber-based assay. Exposure of cultured OPCs to baclofen increased their proliferation, providing evidence for a functional role of GABA(B)Rs in oligodendrocyte development. The presence of GABA(B)Rs in developing oligodendrocytes provides a new mechanism for neuronal-glial interactions during development and may offer a novel target for promoting remyelination following white matter injury.     

Population screening for coeliac disease in primary care by district nurses using a rapid antibody test: diagnostic accuracy and feasibility study

Objective To evaluate the feasibility and diagnostic accuracy of screening for coeliac disease by rapid detection of IgA antibodies to tissue transglutaminase performed in primary care.Design District nurses screened 6 year old children using rapid antibody testing of finger prick blood. They also collected capillary blood samples for laboratory determination of IgA and IgG antibodies to endomysium and IgA antibodies to tissue transglutaminase. Children with positive rapid test results were directly sent for biopsy of the small intestine.Setting Primary care in Jász-Nagykun-Szolnok county, Hungary.Participants 2690 children (77% of 6 year olds living in the county) and 120 nurses.Main outcome measures Positivity for antibodies to endomysium or transglutaminase in the laboratory and coeliac disease confirmed at biopsy.Results 37 children (1.4%, 95% confidence interval 0.9% to 1.8%) had biopsy confirmed coeliac disease. Only five of these children had been diagnosed clinically before screening. Rapid testing had a 78.1% sensitivity (70.0% to 89.3%) and 100% specificity (88.4% to 100%) for a final diagnosis of coeliac disease by biopsy. Sensitivity was 65.1% (50.2% to 77.6%) and specificity was 100% (99.8% to 100%) compared with combined results of IgA and IgG laboratory tests. Trained laboratory workers detected 30 of the 31 newly diagnosed IgA competent patients with the rapid test kit used blindly. Median time to biopsy after a positive rapid test result was significantly shorter (20 days, range 4-148) than after a positive laboratory result (142 days, 70-256; P<0.001). Children with coeliac disease detected at screening were smaller and had worse health status than their peers but they improved on a gluten-free diet.Conclusions A simple rapid antibody test enabled primary care nurses to detect patients with coeliac disease in the community who were not picked up in clinical care. Extra training is needed to improve sensitivity.