NF-kappaB protects lung epithelium against hyperoxia-induced nonapoptotic cell death-oncosis

Prolonged exposure to hyperoxia induces pulmonary epithelial cell death and acute lung injury. Although both apoptotic and nonapoptotic morphologies are observed in hyperoxic animal lungs, nonapoptotic cell death had only been recorded in transformed lung epithelium cultured in hyperoxia. To test whether the nonapoptotic characteristics in hyperoxic animal lungs are direct effects of hyperoxia, the mode of cell death was determined both morphologically and biochemically in human primary lung epithelium exposed to 95% O(2). In contrast to characteristics observed in apoptotic cells, hyperoxia induced swelling of nuclei and an increase in cell size, with no evidence for any augmentation in the levels of either caspase-3 activity or annexin V incorporation. These data suggest that hyperoxia can directly induce nonapoptotic cell death in primary lung epithelium. Although hyperoxia-induced nonapoptotic cell death was associated with NF-kappaB activation, it is unknown whether NF-kappaB activation plays any causal role in nonapoptotic cell death. This study shows that inhibition of NF-kappaB activation can accelerate hyperoxia-induced epithelial cell death in both primary and transformed lung epithelium. Corresponding to the reduced cell survival in hyperoxia, the levels of MnSOD were also low in NF-kappaB-deficient cells. These results demonstrate that NF-kappaB protects lung epithelial cells from hyperoxia-induced nonapoptotic cell death.     

Anoxia tolerance in rice seedlings: exogenous glucose improves growth of an anoxia-'intolerant', but not of a 'tolerant' genotype

This study demonstrated that, in rice seedlings, genotypic differencein tolerance to anoxia only occurred when anoxia was imposedat imbibition, but not at 3 d after imbibition. When seeds wereimbibed and grown in anoxia, IR22 (anoxia-‘intolerant’)grew much slower and had lower soluble sugar concentrationsin coleoptiles and seeds than Amaroo (anoxia-‘tolerant’),while Calrose was intermediate. After 3 d in anoxia, the sugarconcentrations in embryos and endosperms of anoxic seedlingswere nearly 4-fold lower in IR22 than in Amaroo. Sugar deficitin the embryo of IR22 is presumably due to the limitation ofsugar mobilization rather than the capacity of transport asshown by similar sugar accumulation ratios of 1.8 between embryoand endosperm in IR22 and Amaroo at 3 d in anoxia. With 20 molm^–3 exogenous glucose, coleoptile extension and freshweight increments in anoxic seedlings of IR22 were much closerto those in the two other genotypes, nevertheless protein concentrationremained lowest on a fresh weight basis in the coleoptiles ofIR22; indicating that protein synthesis has a lower priorityfor energy apportionment during anoxia than processes crucialto coleoptile extension. In contrast to these responses to anoxiaimposed at imbibition, IR22 had nearly the same high toleranceto anoxia as Calrose and Amaroo, when anoxia was imposed onseedlings subsequent to 48 h aeration followed by 16 h hypoxicpretreatment. In fact, coleoptiles of anoxic IR22 had highersugar concentrations and grew faster than Calrose, and exogenousglucose had no effect on the coleoptile extension of IR22. Excisedcoleoptile tips of IR22 and Amaroo with exogenous glucose hadsimilar rates of ethanol production and were equally tolerantto anoxia. In conclusion, much of the anoxia ‘intolerance’of IR22 when germinated in anoxia could be attributed to limitedsubstrate availability to the embryo and coleoptile, presumablydue to slow starch hydrolysis in the endosperm. Key words: Anoxia, coleoptile, embryo, endosperm, ethanol production, germination, growth, Oryza sativa L., solute net uptake or loss, sugar availability.     

Comparison of the linkage results of two phenotypic constructs from longitudinal data in the Framingham Heart Study: analyses on data measured at three time points and on the average of three measurements

Background

Family studies are often conducted in a cross-sectional manner without long-term follow-up data. The relative contribution of a gene to a specific trait could change over the lifetime. The Framingham Heart Study offers a unique opportunity to investigate potential gene × time interaction. We performed linkage analysis on the body mass index (BMI) measured in 1970, 1978, and 1986 for this project.

Results

We analyzed the data in two different ways: three genome-wide linkage analyses on each exam, and one genome-wide linkage analysis on the mean of the three measurements. Variance-component linkage analyses were performed by the SOLAR program. Genome-wide scans show consistent evidence of linkage of quantitative trait loci (QTLs) on chromosomes 3, 6, 9, and 16 in three measurements with a maximum multipoint LOD score > 2.2. However, only chromosome 9 has a LOD score = 2.14 when the mean values were analyzed. More interestingly, we found potential gene × environment interactions: increasing LOD scores with age on chromosomes 3, 9, and 16 and decreasing LOD scores on chromosome 6 in the three exams.

Conclusion

The results indicate two points: 1) it is possible that a gene (or genes) influencing BMI is (are) up- or down-regulated as people aged due to aging process or changes in lifestyle, environments, or genetic epistasis; 2) using mean values from longitudinal data may reduce the power to detect linkage and may have no power to detect gene × time, and/or gene × gene interactions.

     

A transgenomic cytogenetic sorghum (Sorghum propinquum) bacterial artificial chromosome fluorescence in situ hybridization map of maize (Zea mays L.) pachytene chromosome 9, evidence for regions of genome hyperexpansion

A cytogenetic FISH map of maize pachytene-stage chromosome 9 was produced with 32 maize marker-selected sorghum BACs as probes. The genetically mapped markers used are distributed along the linkage maps at an average spacing of 5 cM. Each locus was mapped by means of multicolor direct FISH with a fluorescently labeled probe mix containing a whole-chromosome paint, a single sorghum BAC clone, and the centromeric sequence, CentC. A maize-chromosome-addition line of oat was used for bright unambiguous identification of the maize 9 fiber within pachytene chromosome spreads. The locations of the sorghum BAC-FISH signals were determined, and each new cytogenetic locus was assigned a centiMcClintock position on the short (9S) or long (9L) arm. Nearly all of the markers appeared in the same order on linkage and cytogenetic maps but at different relative positions on the two. The CentC FISH signal was localized between cdo17 (at 9L.03) and tda66 (at 9S.03). Several regions of genome hyperexpansion on maize chromosome 9 were found by comparative analysis of relative marker spacing in maize and sorghum. This transgenomic cytogenetic FISH map creates anchors between various maps of maize and sorghum and creates additional tools and information for understanding the structure and evolution of the maize genome.     

Radioiodinated aza-diphenylacetylenes as potential SPECT imaging agents for beta-amyloid plaque detection

Two new iodinated fluoro- and hydroxy-pegylated aza-diphenylacetylene derivatives, 1 and 2, targeting beta-amyloid (Abeta) plaques have been successfully prepared. In vitro binding carried out in tissue homogenates prepared from postmortem AD brains with [(125)I]IMPY (6-iodo-2-(4'-dimethylamino)phenyl-imidazo[1,2-a]pyridine) as the radioligand indicated good binding affinities (K(i)=9.2 and 16.8 nM for 1 and 2, respectively). Brain penetrations of the corresponding radioiodinated ligands, evaluated in the normal mice, showed good initial brain penetrations (3.55% and 5.67% ID/g for [(125)I]1 and [(125)I]2 at 2 min post-injection). The washout from normal mice brain was relatively fast (0.33% and 0.91% ID/g at 2h post-injection). The specific binding of these radioiodinated ligands to beta-amyloid plaques was clearly demonstrated using film autoradiography of AD brain sections. Taken together, these preliminary results strongly suggest that these novel iodinated aza-diphenylacetylenes may be potentially useful for imaging Abeta plaques in the living human brain.     

Clinical and immunological effects of human recombinant interleukin-2 given by repetitive weekly infusion to normal dogs

Four normal adult dogs received two consecutive weekly cycles of human recombinant interleukin-2 (IL-2) by continuous infusion for 4 days/week. The dose of IL-2 given to each dog was 3×10⁶ units m^–2 day^–1. Toxicities consisted of mild vomiting, diarrhea, and lethargy to varying degrees in all the dogs. These side-effects were reversed when the treatment was discontinued. Fever, tachypnea, and weight gain were not seen. A marked lymphocytosis and eosinophilia developed in all dogs after completion of each course of IL-2 (resulting in a more than sevenfold increase in each cell type) and persisted for more than 1 month in some. Fresh peripheral blood lymphocytes (PBL) obtained during this lymphocytosis mediated enhanced in vitro lysis of a natural-killer-cell-sensitive canine tumor cell line (CTAC). The in vitro proliferative responses of these same PBL to IL-2 could be detected earlier, progressed faster, and involved more cells than PBL tested prior to IL-2 infusion. Thus, a relatively well-tolerated regime of IL-2 in dogs can induce dramatic increases in lymphocyte numbers and activation, which is associated with augmentation of their in vitro antitumor reactivity. The clinical effectiveness of this immunotherapeutic approach remains to be tested in tumor-bearing dogs where it could serve as a relevant large-animal model for immunotherapy of cancer with IL-2.This work was supported by grants from the University of Wisconsin Graduate School. University of Wisconsin School of Veterinary Medicine Companion Animal Fund, NIH CA-32685, CM-87290, and American Cancer Society CH-237     

Synthesis and biological evaluation of 3-alkyl-dihydrotetrabenazine derivatives as vesicular monoamine transporter-2 (VMAT2) ligands

In the search of new probes for in vivo brain imaging of vesicular monoamine transporter type 2 (VMAT2), we have developed an efficient synthesis of a novel series of 3-alkyl-dihydrotetrabenazine (DTBZ) derivatives. The affinity of VMAT2 was evaluated by an in vitro inhibitory binding assay using [¹²⁵I]-iodovinyl-TBZ or [¹⁸F](+)-FP-DTBZ as radioligands in rat striatal tissue homogenates. New DTBZ derivatives exhibited moderate to good binding affinity to VMAT2. Among these new ligands, compound 4b showed the best affinity for VMAT2 (K_i = 5.98 nM) and may be a useful lead compound for future structure-activity studies.     

Association of ORAI1 haplotypes with the risk of HLA-B27 positive ankylosing spondylitis

Ankylosing spondylitis (AS) is a chronic inflammation of the sacroiliac joints, spine and peripheral joints. The aetiology of ankylosing spondylitis is still unclear. Previous studies have indicated that genetics factors such as human leukocyte antigen HLA-B27 associates to AS susceptibility. We carried out a case-control study to determine whether the genetic polymorphisms of ORAI1 gene, a major component of store-operated calcium channels that involved the regulation of immune system, is a susceptibility factor to AS in a Taiwanese population. We enrolled 361 AS patients fulfilled the modified New York criteria and 379 controls from community. Five tagging single nucleotides polymorphisms (tSNPs) at ORAI1 were selected from the data of Han Chinese population in HapMap project. Clinical statuses of AS were assessed by the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Bath Ankylosing Spondylitis Functional Index (BASFI), and Bath Ankylosing Spondylitis Global Index (BAS-G). Our results indicated that subjects carrying the minor allele homozygote (CC) of the promoter SNP rs12313273 or TT homozygote of the SNP rs7135617 had an increased risk of HLA-B27 positive AS. The minor allele C of 3′UTR SNP rs712853 exerted a protective effect to HLA-B27 positive AS. Furthermore, the rs12313273/rs7135617 pairwise allele analysis found that C-G (OR 1.69, 95% CI 1.27, 2.25; p = 0.0003) and T-T (OR 1.75, 95% CI 1.36, 2.27; p<0.0001) haplotypes had a significantly association with the risk of HLA-B27-positive AS in comparison with the T-G carriers. This is the first study that indicate haplotypes of ORAI1 (rs12313273 and rs7135617) are associated with the risk of HLA-B27 positive AS.     

Study the effect of a pseudo-carrier on pharmacokinetics of 9-fluoropropyl-(+)-dihydrotetrabenazine in rat plasma by ultra-performance liquid chromatography-tandem mass spectrometry

To evaluate the effect of a pseudo-carrier (9-hydroxypropyl-(+)-dihydrotetrabenazine, AV-149) on pharmacokinetics of 9-fluoropropyl-(+)-dihydrotetrabenazine (AV-133), an ultra-performance liquid chromatography–tandem mass spectrometric (UPLC–MS/MS) method was developed and validated for the determination of AV-133 and AV-149 in rat plasma. AV-133 and AV-149 were extracted from plasma following protein precipitation. The chromatographic analysis was performed on an ACQUITY UPLC BEH? C₁₈ column (50 mm × 2.1 mm × 1.7 μm) by a gradient elution. The mass spectrometer was operated in positive mode using electrospray ionization. The analytes were measured using the multiple reaction-monitoring mode (MRM). An external calibration was used, and the calibration curves were linear in the range of 1.00–800 ng/mL for AV-133 and AV-149. The accuracy ranged from 90.8% to 113.2% and the precision ranged from 2.7% to 9.9% for each analyte. The effect of a pseudo-carrier on pharmacokinetics of AV-133 was studied using the presented method.