Pseudotype Formation of Moloney Murine Leukemia Virus with Sendai Virus Glycoprotein F

Mixed infection of cells with both Moloney murine leukemia virus (MoMLV) and related or heterologous viruses produces progeny pseudotype virions bearing the MoMLV genome encapsulated by the envelope of the other virus. In this study, pseudotype formation between MoMLV and the prototype parainfluenza virus Sendai virus (SV) was investigated. We report for the first time that SV infection of MoMLV producer cells results in the formation of MoMLV(SV) pseudotypes, which display a largely extended host range compared to that of MoMLV particles. This could be associated with SV hemagglutinin-neuraminidase (SV-HN) glycoprotein incorporation into MoMLV envelopes. In contrast, solitary incorporation of the other SV glycoprotein, SV fusion protein (SV-F), resulted in a distinct and narrow extension of the MoMLV host range to asialoglycoprotein receptor (ASGP-R)-positive cells (e.g., cultured human hepatoma cells). Since stably ASGP-R cDNA-transfected MDCK cells, but not parental ASGP-R-negative MDCK cells, were found to be transduced by MoMLV(SV-F) pseudotypes and transduction of ASGP-R-expressing cells was found to be inhibited by ASGP-R antiserum, a direct proof for the ASGP-R-restricted tropism of MoMLV(SV-F) pseudotypes was provided. Cultivation of ASGP-R-positive HepG2 hepatoma cells on Transwell-COL membranes led to a significant enhancement of MoMLV(SV-F) titers in subsequent flowthrough transduction experiments, thereby suggesting the importance of ASGP-R accessibility at the basolateral domain for MoMLV(SV-F) pseudotype transduction. The availability of such ASGP-R-restricted MoMLV(SV-F)-pseudotyped vectors opens up new perspectives for future liver-restricted therapeutic gene transfer applications.     

Metformin Induces Apoptosis and Cell Cycle Arrest Mediated by Oxidative Stress,AMPK and FOXO3a in MCF-7 Breast Cancer Cells

Recent studies have demonstrated that the anti-diabetic drug, metformin, can exhibit direct antitumoral effects, or can indirectly decrease tumor proliferation by improving insulin sensitivity. Despite these recent advances, the underlying molecular mechanisms involved in decreasing tumor formation are not well understood. In this study, we examined the antiproliferative role and mechanism of action of metformin in MCF-7 cancer cells treated with 10 mM of metformin for 24, 48, and 72 hours. Using BrdU and the MTT assay, it was found that metformin demonstrated an antiproliferative effect in MCF-7 cells that occurred in a time- and concentration- dependent manner. Flow cytometry was used to analyze markers of cell cycle, apoptosis, necrosis and oxidative stress. Exposure to metformin induced cell cycle arrest in G₀-G₁ phase and increased cell apoptosis and necrosis, which were associated with increased oxidative stress. Gene and protein expression were determined in MCF-7 cells by real time RT-PCR and western blotting, respectively. In MCF-7 cells metformin decreased the activation of IRβ, Akt and ERK1/2, increased p-AMPK, FOXO3a, p27, Bax and cleaved caspase-3, and decreased phosphorylation of p70S6K and Bcl-2 protein expression. Co-treatment with metformin and H₂O₂ increased oxidative stress which was associated with reduced cell number. In the presence of metformin, treating with SOD and catalase improved cell viability. Treatment with metformin resulted in an increase in p-p38 MAPK, catalase, MnSOD and Cu/Zn SOD protein expression. These results show that metformin has an antiproliferative effect associated with cell cycle arrest and apoptosis, which is mediated by oxidative stress, as well as AMPK and FOXO3a activation. Our study further reinforces the potential benefit of metformin in cancer treatment and provides novel mechanistic insight into its antiproliferative role.     

Synthesis and biological evaluation of Esaprazole analogues showing σ1 binding and neuroprotective properties in vitro

Esaprazole, a molecule previously acknowledged to protect against stomach and intestinal ulcers was surprisingly discovered to have neuroprotective activities and σ₁ binding in vitro. A highly diverse set of Esaprazole analogues 2–5 was prepared in order to increase blood–brain barrier penetration. The analogues showed a structure–activity relationship at the σ₁ receptor closely matching already published pharmacophores. Many of the analogues were shown to have neuroprotective properties in two assays using primary cultures of cortical neurons exposed to glutamate and hydrogen peroxide. However, no apparent SAR for these two assays could be developed. Metabolic stability of the analogues were also investigated and the structure of R¹ had a significant bearing on the ADME properties of the compound resulting in two series of compounds. Compounds in which R¹ was a H or acyl group had good metabolic stability in RLM but poor BBB penetration, whereas compounds where R¹ was a cyclo- or bicyclo-alkyl group had poor metabolic stability but good BBB penetration.     

Bosnia Hotel: Kenyan Warriors in Bosnia

Bosnia Hotel: Kenyan Warriors in Bosnia. 1996. 52 minutes, color. video by Thomas Balmès. For more information contact Filmmakers Library, 124 East 40th Street, New York, NY 10016(212/808-4980).     

Damage and Recovery of Cycas micronesica after Typhoon Paka1

Typhoon Paka passed over Guam on 16 December 1997, and caused extensive damage to the Cycas micronesica population. Defoliation reduced subsequent stem extension and leaf size, and increased synchrony of flushes. Decapitated plants developed numerous adventitious buds on damaged stems during recovery. Toppled plants were selectively browsed by feral deer and pigs. Epiphytes increased susceptibility to decapitation or toppling. Growth reduction was minimal and ephemeral, indicating that this species is highly resilient to tropical cyclone damage.     

A novel flat-embedding method to prepare ultrathin cryosections from cultured cells in their in situ orientation.

Immunogold labeling of ultrathin cryosections provides a sensitive and quantitative method to localize proteins at the ultrastructural level. An obligatory step in the routine preparation of cryosections from cultured cells is the detachment of cells from their substrate and subsequent pelleting. This procedure precludes visualization of cells in their in situ orientation and hampers the study of polarized cells. Here we describe a method to sample cultured cells from a petri dish or coverslip by embedding them in a 12% gelatin slab. Subsequently, sections can be prepared in parallel or perpendicular to the plane of growth. Our method extends the cryosectioning technique to applications in studying polarized cells and correlative light-electron microscopy.     

GENETIC AND MATERNAL EFFECTS ON SEEDLING CHARACTERS OF SOLIDAGO ALTISSIMA (COMPOSITAE)

The relative contributions of genetic and maternal factors to phenotypic variation were studied in seedlings of the clonal herb Solidago altissima L. Pairwise crosses were made to establish 19 full-sibships. Because crosses were reciprocal, each parent served as seed parent to half its offspring and as pollen parent to the other half. For 18 characters, genetic variation was estimated from the between sibship variance and maternal variation from the variance between maternal groups within sibships. Early in the growing season, maternal effects exerted strong influence over variation in leaf and stem dimensions, while genetic influences were weak or absent. At the end of the season genetic influences on leaf and stem dimensions, relative biomass allocation between stem and leaves, and the propensity to flower were significant but maternal effects were not. However, both genetic and maternal influences were found on the number and lengths of new rhizomes. Phenotypic, genetic, and maternal correlation matrices were examined through principal component analysis for trade-offs among vegetative growth, clonal expansion and floral reproduction which could constrain the evolution of life history patterns. At the phenotypic level, seedlings with strong vegetative growth produced more and longer rhizomes, and were more likely to flower. Genetically, correlations among these functions were weak or absent. Strong maternal contributions to seedling vegetative growth correlated positively with the number and length of rhizomes. Thus, no trade-offs acting during the seedling stage were found. However, there were indications of a genetically influenced trade-off between the number of rhizomes produced in the seedling year with the size of the ramets produced in the second year.     

Research on dead human bodies: African perspectives on moral status

A useful concept that can be invoked to resolve complex bioethical issues is that of moral status (or, human dignity). In this article, we apply this concept to dead human bodies in order to support our view that research on such bodies is permissible. Instead of drawing from salient Western theories of human dignity that account for it by appeals to autonomy or rationality, we will base our investigation on emerging conceptions in African theories of moral status as articulated by Thaddeus Metz and Munamato Chemhuru. Our conclusion is that neither of these theories of moral status, one secular, and the other religious, can accommodate the intuition that we have direct duties of respect towards such bodies because, in relation to Metz's conception, they lack the capacity to enter into social relationships while on Chemhuru's conception, dead bodies appear not to have any meaningful purpose within the hierarchy that captures the ontological systems that determine such status. Consequently, we argue that our analysis provides support for the use of unclaimed cadavers in training and research in medical institutions.     

Diversity of Insulin Resistance in Monkeys with Normal Glucose Tolerance

Insulin resistance has been proposed as a critical factor in the development of Type II diabetes, hypertension, dyslipidemia, and coronary artery disease. However, even in normal healthy individuals, a wide range of in vivo insulin action has been found. In the present study we sought to examine this heterogeneity in Insulin action in both normal and spontaneously obese nonhuman primates. Maximal insulin responsiveness as measured by a hyperinsulinemic euglycemic clamp, fasting plasma glucose, and insulin levels, β-cell insulin response to glucose, glucose tolerance, and adiposity were measured in 22 male rhesus monkeys. Results showed that lean animals (body fat ≤ 22%) had higher insulin-stimulated glucose uptake (M rate: 14.42±1.8 mg/kg FFM/min) compared to obese (8.08±0.8). The obese monkeys, with 23–49% body fat, had a wide range of M values (5.32-14.29 mg/kg FFM/min) which showed no relationship to degree of adiposity. In all monkeys, M values had a strong inverse correlation with fasting plasma insulin levels (r=-0.76; p<0.001), but not with fasting glucose or glucose disappearance rate. We conclude that neither degree of obesity above a critical threshold nor range of glucose tolerance is related to insulin resistance; however, in individuals with normal glucose tolerance an early reliable indicator of defective insulin action appears to be fasting insulin concentration. Longitudinal determination of basal insulin levels obtained under standardized conditions so as to minimize extraneous variability is likely to strengthen the ability to predict insulin resistance and possible later development of overt Type II diabetes.     

EXTREME KARYOTYPIC VARIATION IN A MUS MUSCULUS DOMESTICUS HYBRID ZONE: THE TOBACCO MOUSE STORY REVISITED

The Robertsonian fusion is a common chromosomal mutation among mammal species and is especially prevalent in the West European house mouse, Mus musculus domesticus. More than 40 races of the house mouse exist in Europe, including the famous “tobacco mouse” (Poschiavo race) of Val Poschiavo, Switzerland. Documented here is the discovery of an extreme case of karyotypic variation in the neighboring Upper Valtellina, Italy. In a 20-km stretch of the valley, 32 karyotypes were observed, including five chromosomal races and 27 hybrid types. One previously unknown race is reported, the “Mid Valtellina” race, with a diploid number of 2n = 24 and the Robertsonian fusions Rb(1.3), Rb(4.6), Rb(5.15), Rb(7.18), Rb(8.12), Rb(9.14), Rb(11.13), and Rb(16.17). The Poschiavo race (2n = 26), Upper Valtellina race (2n = 24), Lower Valtellina race (2n = 22) and all-acrocentric race (2n = 40) were also present. The races form a patchy distribution, which we term a “mottled hybrid zone.” Geographical position, isolation, extinction, recolonization, and selection against hybrids are all believed to be instrumental in the origin and evolution of this complex system. Previous studies of house mice from Upper Valtellina indicated that two of the races in the valley (the Upper Valtellina and Poschiavo races) may have speciated in the village of Migiondo. We discuss the possibility that there may have been a reinforcement event in this village.