Chimeric yellow fever/dengue virus as a candidate dengue vaccine: quantitation of the dengue virus-specific CD8 T-cell response

We have constructed a chimeric yellow fever/dengue (YF/DEN) virus, which expresses the premembrane (prM) and envelope (E) genes from DEN type 2 (DEN-2) virus in a YF virus (YFV-17D) genetic background. Immunization of BALB/c mice with this chimeric virus induced a CD8 T-cell response specific for the DEN-2 virus prM and E proteins. This response protected YF/DEN virus-immunized mice against lethal dengue encephalitis. Control mice immunized with the parental YFV-17D were not protected against DEN-2 virus challenge, indicating that protection was mediated by the DEN-2 virus prM- and E-specific immune responses. YF/DEN vaccine-primed CD8 T cells expanded and were efficiently recruited into the central nervous systems of DEN-2 virus challenged mice. At 5 days after challenge, 3 to 4% of CD8 T cells in the spleen were specific for the prM and E proteins, and 34% of CD8 T cells in the central nervous system recognized these proteins. Depletion of either CD4 or CD8 T cells, or both, strongly reduced the protective efficacy of the YF/DEN virus, stressing the key role of the antiviral T-cell response.     

Inter‐Fullerene Electronic Coupling Controls the Efficiency of Photoinduced Charge Generation in Organic Bulk Heterojunctions

Photoinduced charge generation (PCG) dynamics are notoriously difficult to correlate with specific molecular properties in device relevant polymer:fullerene organic photovoltaic blend films due to the highly complex nature of the solid state blend morphology. Here, this study uses six judiciously selected trifluoromethylfullerenes blended with the prototypical polymer poly(3‐hexylthiophene) and measure the PCG dynamics in 50 fs–500 ns time scales with time‐resolved microwave conductivity and femtosecond transient absorption spectroscopy. The isomeric purity and thorough chemical characterization of the fullerenes used in this study allow for a detailed correlation between molecular properties, driving force, local intermolecular electronic coupling and, ultimately, the efficiency of PCG yield. The findings show that the molecular design of the fullerene not only determines inter‐fullerene electronic coupling, but also influences the decay dynamics of free holes in the donor phase even when the polymer microstructure remains unchanged.     

The Computerized Laboratory Notebook concept for genetic toxicology experimentation and testing

We describe a microcomputer system utilizing the Computerized Laboratory Notebook (CLN) concept developed in our laboratory for the purpose of automating the Battery of Leukocyte Tests (BLT). The BLT was designed to evaluate blood specimens for toxic, immunotoxic, and genotoxic effects after in vivo exposure to putative mutagens. A system was developed with the advantages of low cost, limited spatial requirements, ease of use for personnel inexperienced with computers, and applicability to specific testing yet flexibility for experimentation. This system eliminates cumbersome record keeping and repetitive analysis inherent in genetic toxicology bioassays. Statistical analysis of the vast quantity of data produced by the BLT would not be feasible without a central database. Our central database is maintained by an integrated package which we have adapted to develop the CLN. The clonal assay of lymphocyte mutagenesis (CALM) section of the CLN is demonstrated. PC-Slaves expand the microcomputer to multiple workstations so that our computerized notebook can be used next to a hood while other work is done in an office and instrument room simultaneously. Communication with peripheral instruments is an indispensable part of many laboratory operations, and we present a representative program, written to acquire and analyze CALM data, for communicating with both a liquid scintillation counter and an ELISA plate reader. In conclusion we discuss how our computer system could easily be adapted to the needs of other laboratories.     

Optimizing healthy gestational weight gain in women at high risk of gestational diabetes: A randomized controlled trial

Objective:

Optimizing gestational weight gain (GWG) in early pregnancy is of clinical and public health importance, especially in higher risk pregnancies.

Design and Methods:

In a robustly designed, randomized controlled trial, 228 pregnant women at risk of developing gestational diabetes mellitus (GDM) were allocated to either control (written health information only) or intervention (four‐session lifestyle program). All women received standard maternal care. Measures were completed at 12‐15 and 26‐28 weeks gestation. Measures included anthropometrics (weight and height), physical activity (pedometer and International Physical Activity Questionnaire), questionnaires (risk perception), and GDM screening.

Results:

The mean (SD) age [31.7 (4.5) and 32.4 (4.7) years] and body mass index [BMI; 30.3 (5.9) and 30.4 (5.6) kg/m²] were similar between control and intervention groups, respectively. By 28 weeks, GWG was significantly different between control and intervention groups [6.9 (3.3) vs. 6.0 (2.8) kg, P < 0.05]. When stratified according to baseline BMI, overweight women in the control group gained significantly more weight compared to overweight women in the intervention group [7.8 (3.4) vs. 6.0 (2.2) kg, P < 0.05], yet in obese women, GWG was similar in both groups. Physical activity levels declined by 28 weeks gestation overall (P < 0.01); however, the intervention group retained a 20% higher step count compared to controls [5,203 (3,368) vs. 4,140 (2,420) steps/day, P < 0.05]. Overall, GDM prevalence was 22%, with a trend toward less cases in the intervention group (P = 0.1).

Conclusions:

Results indicate that a low‐intensity lifestyle intervention, integrated with antenatal care, optimizes healthy GWG and attenuates physical activity decline in early pregnancy. Efficacy in limiting weight gain was greatest in overweight women and in high‐risk ethnically diverse women.     

Characterization of a cDNA encoding rat sterol carrier protein2

Sterol carrier protein2 (SCP2) is a 13.2-kD protein that is thought to be involved in the intracellular transport of cholesterol. Using synthetic oligonucleotides based on the protein sequence of SCP2, a clone (SP43) was isolated from a rat liver cDNA library. The DNA sequence revealed that the cDNA could encode a polypeptide of 273 amino acids (28.9 kD) or 143 amino acids (15.3 kD) in which the carboxy-terminal 123 amino acids are identical to the SCP2 protein. RNA blot hybridization revealed that a variety of rat tissues contain a homologous RNA of a size similar to SP43 (approximately 1.5 kb). Levels of SCP2 mRNA increased in parallel with cytochrome P450scc mRNA in the immature gonadotropin-primed rat ovary. The isolation of a cDNA clone encoding SCP2 will facilitate studies on its role in cholesterol metabolism.     

3-hydroxy fatty acids found in capsules of Cryptococcus neoformans

Using immunofluorescence confocal laser scanning microscopy, immunogold transmission electron microscopy and gas chromatography--mass spectrometry, we demonstrated the presence of 3-hydroxy fatty acids in Cryptococcus neoformans. Our results suggest that these oxylipins accumulate in capsules where they are released as hydrophobic droplets through tubular protuberances into the surrounding medium.     

A spin-labeled abasic DNA substrate for AP endonuclease.

We report the first observation of a spin-labeled ds 23-mer oligonucleotide by high-field electron spin resonance (ESR) and demonstrate that it interacts with AP endonuclease, the key enzyme in DNA abasic site repair. The spin labeled 23-mer with a U at position 12 of the upper strand is processed by uracil DNA glycosylase to provide the abasic substrate. With a spin-label two nucleotides away from the abasic site, AP endo binds and cleaves when the label is 3' but not 5' to the abasic site. These results confirm that the disposition of the bases immediately upstream of the abasic site is particularly critical for cleavage by AP endo, and establish that DNA-protein interactions in this important enzyme can be examined using spin-labeled substrates.     

Activation of Cl− Currents in Cultured Rat Retinal Pigment Epithelial Cells by Intracellular Applications of Inositol-1,4,5-triphosphate: Differences Between Rats with Retinal Dystrophy (RCS) and Normal Rats

Using the whole-cell configuration of the patch-clamp technique, we studied the conditions necessary for the activation of Cl⁻-currents in retinal pigment epithelial (RPE) cells from rats with retinal dystrophy (RCS) and nondystrophic control rats. In RPE cells from both rat strains, intracellular application of 10 μm inositol-1,4,5-triphosphate (IP3) via the patch pipette led to a sustained activation of voltage-dependent Cl⁻ currents, blockable by 1 mm 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS). IP3 activated Cl⁻ currents in the presence of a high concentration of the calcium chelator BAPTA (10 mm) in the pipette solution, but failed to do so when extracellular calcium was removed. Intracellular application of 10⁻⁵ m Ca²⁺ via the patch pipette also led to a transient activation of Cl⁻ currents. When the cells were preincubated in a bath solution containing thapsigargin (1 μm) for 5 min before breaking into the whole-cell configuration, IP3 failed to activate voltage-dependent currents. Thus, IP3 led to release of Ca²⁺ from cytosolic calcium stores. This in turn activated an influx of extracellular calcium into the submembranal space by a mechanism as yet unknown, leading to an activation of calcium-dependent chloride currents. In RPE cells from RCS rats, which show an increased membrane conductance for calcium compared to normal rats, we observed an accelerated speed of Cl⁻-current activation induced by IP3 which could be reduced by nifedipine (1 μm). Thus, the increased membrane conductance to calcium in RPE cells from RCS rats changes the response of the cell to the second messenger IP3. Received: 17 July 1995/Revised: 31 January 1996     

Bone Growth Dynamics of the Facial Skeleton and Mandible in <Emphasis Type="Italic">Gorilla gorilla</Emphasis> and <Emphasis Type="Italic">Pan troglodytes</Emphasis>

Adult craniofacial morphology results from complex processes that involve growth by bone modelling and interactions of skeletal components to keep a functional and structural balance. Previous analyses of growth dynamics in humans revealed critical changes during late ontogeny explaining particular morphological features in our species. Data on bone modelling patterns from other primate species could help us to determine whether postnatal changes in the growth dynamics of the craniofacial complex are human specific or are shared with other primates. However, characterizations of bone modelling patterns through ontogeny in non-human hominids are scarce and restricted to isolated data on facial and mandibular regions. In the present study, we analyse the bone modelling patterns in an ontogenetic series of Pan and Gorilla to infer the growth dynamics of their craniofacial complex during postnatal development. Our results show that both Pan troglodytes and Gorilla gorilla are characterized by species-specific bone modelling patterns indicative of a mainly forward growth direction during postnatal development. Both species show minor but consistent ontogenetic changes in the distribution of bone modelling fields in specific regions of the face and mandible, in contrast to other regions which show more constant bone modelling patterns. In addition, we carry out a preliminary integrative study merging histological and geometric morphometric data. Both approaches yield highly complementary data, each analysis providing details on specific growth dynamics unavailable to the other. Moreover, geometric morphometric data show that ontogenetic variation in the modelling pattern of the mandibular ramus may be linked to sexual dimorphism.