Comparison of cytogenetic effects of 3,4-epoxy-1-butene and 1,2:3, 4-diepoxybutane in mouse, rat and human lymphocytes following in vitro G0 exposures

To understand better the species differences in carcinogenicity caused by 1,3-butadiene (BD), we exposed G0 lymphocytes (either splenic or peripheral blood) from rats, mice and humans to 3, 4-epoxy-1-butene (EB) (20 to 931 microM) or 1,2:3,4-diepoxybutane (DEB) (2.5 to 320 uM), two of the suspected active metabolites of BD. Short EB exposures induced little measurable cytogenetic damage in either rat, mouse, or human G0 lymphocytes as measured by either sister chromatid exchange (SCE) or chromosome aberration (CA) analyses. However, DEB was a potent inducer of both SCEs and CAs in G0 splenic and peripheral blood lymphocytes. A comparison of the responses among species showed that the rat and mouse were approximately equisensitive to the cytogenetic damaging effects of DEB, but the situation for the human subjects was more complex. The presence of the GSTT1-1 gene (expressed in the erythrocytes) reduced the relative sensitivity of the lymphocytes to the SCE-inducing effects of DEB. However, additional factors also appear to influence the genotoxic response of humans to DEB. This study is the first direct comparison of the genotoxicity of EB and DEB in the cells from all three species.     

Anti-HBV specific beta-L-2'-deoxynucleosides

A unique series of simple unnatural L-nucleosides that specifically inhibit hepatitis B virus (HBV) replication has been discovered. These molecules have in common a hydroxyl group in the 3'-position (3'-OH) of the beta-L-2'-deoxyribose sugar that confers antiviral activity specifically against hepadnaviruses. Replacement of the 3'-OH broadens activity to other viruses. Substitution in the base decreases antiviral potency and selectivity. Human DNA polymerases and mitochondrial function are not effected. Plasma viremia is reduced up to 8 logs in a woodchuck model of chronic HBV infection. These investigational drugs, used alone or in combination, are expected to offer new therapeutic options for patients with chronic HBV infection.     

Lymphosarcoma in the laboratory woodchuck (Marmota monax)

From 1979 to 1999,28 cases of lymphosarcoma were identified in the Cornell University woodchuck colony (prevalence rate: 152/100,000/yr). The prevalence of lymphosarcoma was similar in woodchucks not infected with the woodchuck hepatitis virus (WHV) and in chronic carriers of WHV. Males (13) and females (15) alike were affected (mean +/- SD age 4.7 +/- 2.92 years; range, 0.5 to 9 years). On the basis of the major organ system involved, woodchuck lymphosarcoma was classified as multicentric (12 cases, 43%), alimentary (5 cases, 18%), cranial mediastinal (5 cases, 18%), and miscellaneous (6 cases, 21%). A cutaneous form was not observed. Morphologic criteria similar to those of the Kiel classification were used for light microscopic classification. All Kiel categories-except the immunoblastic form-were found: 17 cases (61%) were centroblastic, and 6 were lymphocytic (21%). Other categories (centrocytic and plasmacytoid) were recognized less frequently. Immunophenotyping of 27 cases revealed 15 (56%) B cell (CD3-/CD79a+ or CD3-/BLA.36+), 7 (26%) T cell (CD3+/CD79a-/BLA.36-), and 5 (18%) non-T non-B cell (CD3-CD79a-/BLA.36-) lymphosarcomas. Lymphosarcoma in woodchucks develops at a higher rate than that observed in humans or companion animals, and WHV infection has no effect on prevalence. The anatomic and Kiel classification used in domestic species also can be used in woodchucks. Commercially available alpha-CD3, alpha-CD79a, and alpha-BLA.36 antibodies were useful for immunophenotyping woodchuck lymphosarcomas.     

Seasonal changes in serum leptin, food intake, and body weight in photoentrained woodchucks

Male woodchucks (Marmota monax) were maintained in northern vs. southern hemisphere photoperiods, provided feed and water ad libitum, and evaluated every 2 wk for 23 mo for body weight, absolute and relative food intake, body temperature, serum testosterone, and serum concentrations of leptin measured using an anti-mouse leptin enzyme-linked immunoassay. During late spring and summer, body weight increased 56 +/- 4% above winter nadirs, and during the autumn and early winter weights decreased 27 to 43% below midsummer maxima. Serum leptin initially increased during increases in body weight, in the late spring, reached peak values (490 +/- 32 pg/ml) in summer during the initial decline in body weight, and later decreased along with body weight to reach basal values (20 +/- 5 pg/ml) in late winter. Spontaneous declines in food intakes in summer began 2-6 wk before resulting declines in body weight and occurred during increases in leptin >100 pg/ml. The rate of decline in food intakes was greatest when serum leptin was at or near peak values. Food intake increased in late winter when leptin was low and 7-10 wk before resulting increases in body weight. Testis recrudescence occurred when leptin was declining to near basal levels. The results suggest that leptin is involved in the hormonal regulation of the circannual cycle in the drive for voluntary food intake in this species.     

Cytogenetic studies of three triazine herbicides. II. In vivo micronucleus studies in mouse bone marrow

Atrazine, simazine, and cyanazine are widely used preemergence and postemergence triazine herbicides that have made their way into the potable water supply of many agricultural communities. Although there are several contradictory genotoxicity studies in the literature, our previous in vitro studies with human lymphocytes showed that atrazine, simazine, and cyanazine did not induce sister chromatid exchanges (SCEs) or chromosome aberrations (CAs) up to the limits of solubility in aqueous medium using 0.5% dimethyl sulfoxide. To expand upon these results and to ensure that our in vitro findings could be replicated in an in vivo system, mice were treated with each triazine by two intraperitoneal injections, 24h apart. The animals were sacrificed and the bone marrow removed for micronucleus (MN) analysis, 24h after the last injection. Two to four independent trials were performed for MN analysis in polychromatic erythrocytes, and in some trials the spleen was removed, cultured, and analyzed for SCEs and CAs. None of the triazines investigated induced MN in the bone marrow, even at doses that caused significant bone marrow suppression and/or death. These results indicate that atrazine, simazine, and cyanazine are not genotoxic as measured by the bone marrow MN assay in mice following high dose exposures.     

Chemical shift as a probe of molecular interfaces: NMR studies of DNA binding by the three amino-terminal zinc finger domains from transcription factor IIIA

We report the NMR resonance assignments for a macromolecular protein/DNA complex containing the three amino-terminal zinc fingers (92 amino acid residues) of Xenopus laevis TFIIIA (termed zf1-3) bound to the physiological DNA target (15 base pairs), and for the free DNA. Comparisons are made of the chemical shifts of protein backbone¹ H^N, ¹⁵N,¹³ C and¹³ C and DNA base and sugar protons of the free and bound species. Chemical shift changes are analyzed in the context of the structures of the zf1-3/DNA complex to assess the utility of chemical shift change as a probe of molecular interfaces. Chemical shift perturbations that occur upon binding in the zf1-3/DNA complex do not correspond directly to the structural interface, but rather arise from a number of direct and indirect structural and dynamic effects.     

Invasive Salmonella Typhimurium ST313 with Naturally Attenuated Flagellin Elicits Reduced Inflammation and Replicates within Macrophages

Invasive non-typhoidal Salmonella (iNTS) are an important cause of septicemia in children under the age of five years in sub-Saharan Africa. A novel genotype of Salmonella enterica subsp. enterica serovar Typhimurium (multi-locus sequence type [ST] 313) circulating in this geographic region is genetically different to from S. Typhimurium ST19 strains that are common throughout the rest of the world. S. Typhimurium ST313 strains have acquired pseudogenes and genetic deletions and appear to be evolving to become more like the typhoidal serovars S. Typhi and S. Paratyphi A. Epidemiological and clinical data show that S. Typhimurium ST313 strains are clinically associated with invasive systemic disease (bacteremia, septicemia, meningitis) rather than with gastroenteritis. The current work summarizes investigations of the broad hypothesis that S. Typhimurium ST313 isolates from Mali, West Africa, will behave differently from ST19 isolates in various in vitro assays. Here, we show that strains of the ST313 genotype are phagocytosed more efficiently and are highly resistant to killing by macrophage cell lines and primary mouse and human macrophages compared to ST19 strains. S. Typhimurium ST313 strains survived and replicated within different macrophages. Infection of macrophages with S. Typhimurium ST19 strains resulted in increased apoptosis and higher production of proinflammatory cytokines, as measured by gene expression and protein production, compared to S. Typhimurium ST313 strains. This difference in proinflammatory cytokine production and cell death between S. Typhimurium ST19 and ST313 strains could be explained, in part, by an increased production of flagellin by ST19 strains. These observations provide further evidence that S. Typhimurium ST313 strains are phenotypically different to ST19 strains and instead share similar pathogenic characteristics with typhoidal Salmonella serovars.     

Transcriptional profiling of bovine intervertebral disc cells: implications for identification of normal and degenerate human intervertebral disc cell phenotypes

Introduction  

Nucleus pulposus (NP) cells have a phenotype similar to articular cartilage (AC) cells. However, the matrix of the NP is clearly different to that of AC suggesting that specific cell phenotypes exist. The aim of this study was to identify novel genes that could be used to distinguish bovine NP cells from AC and annulus fibrosus (AF) cells, and to further determine their expression in normal and degenerate human intervertebral disc (IVD) cells.