Hybridization potential of oligonucleotides comprising 3'-O-methylated altritol nucleosides.

A series of 3'-O-methylated-d-altrohexitol nucleoside analogs (MANA) was synthesized comprising all four base moieties, adenine, cytosine, uracil, and guanine. These monomers were incorporated into oligonucleotides (ONs) by automated solid phase synthesis and the thermal and thermodynamic stability of all new modified constructs were evaluated. Data were compared with results obtained for both anhydrohexitol (HNAs) and 3'-O-altrohexitol-modified ONs (ANAs). We hereby demonstrate that ONs modified with MANA monomers have an improved thermal and thermodynamic stability compared to RNA, ANA, or HNA containing ONs of which the extent depends on the number of incorporated moieties and their position in the sequence. Thermodynamic analysis afforded comparable or even improved results in comparison with the incorporation of locked nucleic acids. While the specificity of these new synthons is slightly lower compared to mismatches within RNA double strands, it is similar to the discrimination potential of other hexitol modifications (HNA and ANA) which already proved their biologic interest, highlighting the potential of MANA constructs in antisense and in siRNA applications.

Biomarkers of exposure and potential harm in adult smokers of 3-7 mg tar yield (Federal Trade Commission) cigarettes and in adult non-smokers.

The paper reports levels of 24-h urine nicotine and five of its major metabolites (expressed as nicotine-equivalents) and blood carboxyhaemoglobin as biomarkers of exposure to particulate- and gas-phase cigarette smoke, respectively, from an exploratory pilot study of adult smokers of 3.0-6.9 mg tar delivery (Federal Trade Commission (FTC) method) cigarettes. On multiple occasions over 6 weeks, blood high-sensitivity C-reactive protein (hs-CRP), fibrinogen, HDL- and LDL-cholesterol, and 24-h urine 8-epi-prostaglandin F2alpha (8-epi-PGF2alpha) and 11-dehydro-thromboxane B2 (11-dehydro-TxB2) were also evaluated as biomarkers of potential harm. All the biomarkers examined, except for LDL-cholesterol, discriminated with high sensitivity and specificity between adult smokers and non-smokers overall. Except for HDL-cholesterol, all biomarker medians were greater in adult smokers than in non-smokers: urine nicotine-equivalents 64.514 versus < 0.034 nmol mg-1 creatinine (p<0.001), carboxyhaemoglobin 4.0 versus 0.4% saturation (p<0.001), hs-CRP 0.27 versus 0.12 mg dl-1 (p=0.05), fibrinogen 292 versus 248 mg dl-1 (p<0.001), HDL-cholesterol 46 versus 53 mg dl-1 (p=0.003), LDL-cholesterol 119 versus 109 mg dl-1 (p=0.18), urine 8-epi-PGF2alpha 1935 versus 1034 pg mg-1 creatinine (p<0.001) and urine 11-dehydro-TxB2 973 versus 710 pg mg-1 creatinine (p<0.001). All the biomarkers of exposure and most of the biomarkers of potential harm showed no time of sampling (by visit week) effect.

First measurement of the velocity of slow antihydrogen atoms.

The speed of antihydrogen atoms is deduced from the fraction that passes through an oscillating electric field without ionizing. The weakly bound atoms used for this first demonstration travel about 20 times more rapidly than the average thermal speed of the antiprotons from which they form, if these are in thermal equilibrium with their 4.2 K container. The method should be applicable to much more deeply bound states, which may well be moving more slowly, and should aid the quest to lower the speed of the atoms as required if they are to be trapped for precise spectroscopy.

First laser-controlled antihydrogen production.

Lasers are used for the first time to control the production of antihydrogen (H ). Sequential, resonant charge exchange collisions are involved in a method that is very different than the only other method used so far-producing slow H during positron cooling of antiprotons in a nested Penning trap. Two attractive features are that the laser frequencies determine the H binding energy, and that the production of extremely cold H should be possible in principle-likely close to what is needed for confinement in a trap, as needed for precise laser spectroscopy.

Synthesis and antiviral evaluation of ribavirin congeners containing a hexitol moiety.

Several ribavirin congeners containing a hexitol moiety were prepared via ring opening of two different epoxides with the methylcarboxylate ester of triazole and further elaboration. Unfortunately, none of the newly synthesized compounds displayed appreciable antiviral activity.

Oligonucleotides containing disaccharide nucleosides: synthesis, physicochemical, and substrate properties.

The efficient synthesis of oligonucleotides containing 2'-O-beta-D-ribofuranosyl (and beta-D-ribopyranosyl)nucleosides, 2'-O-alpha-D-arabinofuranosyl (and alpha-L-arabinofuranosyl)nucleosides. 2'-O-beta-D-erythrofuranosylnucleosides, and 2'-O-(5'-amino-5-deoxy-beta-D-ribofuranosyl)nucleosides have been developed.

Methylated hexitol nucleic acids, towards congeners with improved antisense potential.

In an effort to further improve the hybridisation potential of anhydro-hexitol nucleoside analogues, the 1'-methoxyl and 3'-methoxyl substituents were introduced and evaluated for their antisense potential. In view of the selectivity of pairing with RNA, especially the introduction of a 3'-O-alkyl moiety deserves further study.

Toxicological evaluation of an electrically heated cigarette. Part 4: Subchronic inhalation toxicology.

The biological activity of mainstream smoke from an electrically heated cigarette (EHC) with controlled combustion and from the University of Kentucky Reference Cigarette 1R4F was determined in Sprague Dawley rats exposed nose-only for 90 days, 6 h a day, 7 days per week. For an equivalent response comparison between the two cigarette types, two doses were chosen for the EHC where the anticipated results were in the dynamic range of the 1R4F dose-response curve (four concentrations) for most end points. The number of cigarettes smoked per m(3) of diluted smoke resulted in total particulate matter concentrations of 40 and 90 microg l (-1) for the EHC and 40-170 microg l (-1) for the 1R4F. Biomonitoring indicated achievement of target doses. Mainstream smoke yields were lower for the EHC, with the exception of formaldehyde. No smoke-related mortality, remarkable in-life observations or abnormal gross pathological findings were observed. Smoke- and dose-related clinical pathology and organ weight changes included: increases in segmented neutrophils, some liver parameters and lung and adrenal weight relative to body weight; and decreases in lymphocytes, glucose concentration and spleen weight. Smoke-related histopathological findings in the respiratory tract included epithelial cell hyperplasia, squamous metaplasia, atrophy and accumulation of pigmented alveolar macrophages; they were mostly dose-dependent, more pronounced in the upper than lower respiratory tract and completely or partially reversed by 6 weeks post-inhalation. Qualitatively, the biological effects seen for the EHC and the 1R4F were comparable and similar to those observed in other mainstream smoke inhalation studies. Quantitatively, the biological activity of the EHC mainstream smoke was, on average, 65% lower than that of the 1R4F mainstream smoke on an equal cigarette basis and equivalent activity on an equal TPM basis.

Driven production of cold antihydrogen and the first measured distribution of antihydrogen states.

Cold antihydrogen is produced when antiprotons are repeatedly driven into collisions with cold positrons within a nested Penning trap. Efficient antihydrogen production takes place during many cycles of positron cooling of antiprotons. A first measurement of a distribution of antihydrogen states is made using a preionizing electric field between separated production and detection regions. Surviving antihydrogen is stripped in an ionization well that captures and stores the freed antiproton for background-free detection.

Background-free observation of cold antihydrogen with field-ionization analysis of its states.

A background-free observation of cold antihydrogen atoms is made using field ionization followed by antiproton storage, a detection method that provides the first experimental information about antihydrogen atomic states. More antihydrogen atoms can be field ionized in an hour than all the antimatter atoms that have been previously reported, and the production rate per incident high energy antiproton is higher than ever observed. The high rate and the high Rydberg states suggest that the antihydrogen is formed via three-body recombination.

Improved hybridisation potential of oligonucleotides comprising O-methylated anhydrohexitol nucleoside congeners.

The hybridising potential of anhydrohexitol nucleoside analogues (HNAs) is well documented, but tedious synthesis of the monomers hampers their development. In a search for better analogues, the synthesis of two new methylated anhydrohexitol congeners 1 and 2 was accomplished and the physico-chemical properties of their respective oligomers were evaluated. Generally, oligonucleotides (ONs) containing the 3'-O-methyl derivative 1 showed a small increase in thermal stability towards complementary sequences as compared to HNA. Compared to the altritol modification, 3'-O-methylation seems to cause a small decrease in thermal stability of duplexes, especially when targeting RNA. These results suggest the possibility of derivatisation of the 3'-hydroxyl group of altritol-containing congeners without significantly affecting the thermal stability of the duplexes. The methyl glycosidic analogues 2 likewise increased the affinity for RNA in comparison with well-known HNA, while at the same time being economically more favorable monomers. However, homopolymers of 2 displayed self-pairing, but not so homopolymers of 1. Upon incorporation of the hexitols within RNA sequences in an effort to induce a beneficial pre-organised structure, the positive effect of the 3'-O-methyl derivative 1 proved larger than that of 2.

Increased RNA affinity of HNA analogues by introducing alkoxy substituents at the C-1 or C-3 position.

1,5-Anhydrohexitol nucleoside congeners with alkoxy substituents, were prepared, resulting in a further improvement of their RNA affinity and antisense potential.

Mouse Sox8 is located between, not within,the t-complex deletions t(w18) and t(h20) on chromosome 17.

The SOX family of developmental transcription factors is known to play critical roles in cell lineage specification, fate determination and differentiation during development in diverse phyla. Their importance is underscored by their involvement in a number of human diseases and mouse mutants, and by targeted mutation in mice. SOX8 is broadly expressed during development and is located on human chromosome 16p and within the t-complex on mouse chromosome 17, in the vicinity of two mutations t(w18) and t(h20). Here we analyse mutant genomic DNA to show that the Sox8 gene locus lies outside the deletion regions of both t(w18) and t(h20) and between these deletions. These data exclude Sox8 from contributing to the t(w18) and t(h20) phenotypes, and provide an additional marker for structural characterization of this complex genomic region.

Hemoglobin adducts in rats chronically exposed to room-aged cigarette sidestream smoke and diesel engine exhaust.

Under controlled conditions with exaggerated concentrations of environmental aerosols, the biologically effective dose markers suggested in the literature as being specific for ETS (i.e., HPB Hb adducts for TSNA exposure) and DEE (i.e., l-aminopyrene Hb adducts for l-nitropyrene exposure) did not respond. A slight but dose-dependent increase in 4-ABP Hb adduct levels was seen in RASS-exposed rats.

Hybridization between "six-membered" nucleic acids: RNA as a universal information system.

Within the polyA:polyT recognition system, cross-pairing between several nucleic acids with a phosphorylated six-membered carbohydrate (mimic) as repeating unit in the backbone structure has been observed. All investigated nucleic acids (except for beta-homo-DNA) hybridize with RNA, leaving RNA as a versatile biopolymer for informational transfer. [reaction: see text]

Phylogeny of the SOX family of developmental transcription factors based on sequence and structural indicators.

Members of the SOX family of transcription factors are found throughout the animal kingdom, are characterized by the presence of a DNA-binding HMG domain, and are involved in a diverse range of developmental processes. Previous attempts to group SOX genes and deduce their structural, functional, and evolutionary relationships have relied largely on complete or partial HMG box sequence of a limited number of genes. In this study, we have used complete HMG domain sequence, full-length protein structure, and gene organization data to study the pattern of evolution within the family. For the first time, a substantial number of invertebrate SOX sequences have been included in the analysis. We find support for subdivision of the family into groups A-H, as has been suggested in some previous studies, and for the assignment of two new groups, I and J. For vertebrate genes, it appears that relatedness as suggested by HMG domain sequence is congruent with relatedness as indicated by overall structure of the full-length protein and intron-exon structure of the genes. Most of the SOX groups identified in vertebrates were represented by a single SOX sequence in each invertebrate species studied. We have named anonymous sequences and, where appropriate, have suggested systematic names for some previously identified sequences. In addition, we identify an HMG domain signature motif which may be considered representative of the SOX family. Based on our data, we propose a robust phylogeny of SOX genes that reflects their evolutionary history in metazoans.

Cloning and characterisation of the Sry-related transcription factor gene Sox8.

SOX proteins form a large family of transcription factors related by a DNA-binding domain known as the HMG box. Some 30 Sox genes have been identified in mammals and orthologues have been found in a wide range of other metazoans. Sox genes are highly conserved and are known to play important roles in embryonic development, including roles in gonadal, central nervous system, neural crest and skeletal development. Several SOX genes have been implicated in human congenital diseases. We report here the isolation of Sox8 and its characterisation in mice and humans. This gene has a remarkably similar primary structure and genomic organisation to the campomelic dysplasia gene SOX9 and the Waardenburg-Shah syndrome gene SOX10. SOX8 protein is able to bind to canonical SOX target DNA sequences and activate transcription in vitro through two separate trans -activation regions. Further, Sox8 is expressed in the central nervous system, limbs, kidneys, gonads and craniofacial structures during mouse embryo development. Sox8 maps to the t complex on mouse chromosome 17 and to human chromosome 16p13.3, a region associated with the microphthalmia-cataract syndrome CATM and the alpha-thalassemia/mental retardation syndrome ATR-16.

A long-term-care setting pilot study evaluating predictors of success in medication self-administration.

OBJECTIVE: To determine whether any specific patient variables or a short battery of neuropsychological tests of cognition and memory predict success in medication self-administration. DESIGN: A prospective, single-blinded design. SETTING: An extended care center of a university-affiliated VA Medical Center. PATIENTS: Thirty predominately male, older veteran patients, mean age 70 +/- 4 years with a range of 63 to 79 years. INTERVENTIONS: Neuropsychological testing [Mini-Mental Status Examination (MMSE), Delayed Word Recall Test (DWRT), Shipley Institute of Living Scale (SILS), Neurobehavioral Cognitive Status Examination (NCSE), Hopkins Verbal Learning Test (HVLT)], twice a week unannounced bedside medication counts and medication administration record inspections, and educational instruction, if needed, by nurses and pharmacists. MAIN OUTCOME MEASURES: Patient characteristics such as age, number of medications, presence of a disorder that can alter cognitive or memory function, years of education, and results from the above listed neuropsychological tests. The dependent variable was successful or not successful as defined by whether the patient required a re-education intervention. RESULTS: Fifteen patients required one or more re-education intervention(s) as a result of meeting the criteria for not being successful. The absence of major depression, stroke, or anxiety disorder did tend to predict success (P = 0.0716) in medication self-administration. The other patient specific characteristics did not predict success. Among the neuropsychological tests administered, only the Judgment Subtest of the NCSE tended to predict success (P = 0.098). The MMSE, DWRT, SILS,HVLT tests did not predict successful performance in the self-medication program. CONCLUSIONS: Although the presence of a diagnosis that could potentially alter cognitive and memory function tended to predict success, no patient characteristics were found that predicted success independently. Among the neuropsychological tests, only the Judgment subtest of the NCSE tended to predict success in medication self-administration. We conclude that the NCSE and characterization of patient-specific factors, including diseases that may affect cognitive and memory function, seem to be the best predictors of success in medication self-administration in a long-term-care setting.

5'-hydroxycotinine-N-oxide, a new nicotine metabolite isolated from rat urine.

1. 5'-Hydroxycotinine-N-oxide, 5-(3-pyridyl-N-oxide)-5-hydroxy-1-methyl-pyrrolidone-2, was identified as a new in vivo metabolite of nicotine. 2. The new metabolite was isolated from the urine of rats treated with S-nicotine and characterized using chemical and spectrometric methods. 3. 5'-Hydroxycotinine-N-oxide was synthesized and characterized by MS and by infrared as well as 1H- and 13C-NMR spectroscopy. 4. Identity of the new metabolite with synthetic 5'-hydroxycotinine-N-oxide was demonstrated by comparing the MS and 1H-NMR spectroscopy data as well as by co-chromatography of a spiked urine sample.