Fluorescent virions: dynamic tracking of the pathway of adenoviral gene transfer vectors in living cells

T cells specific for nucleosomal autoepitopes are selectively expanded in lupus mice and these Th cells drive autoimmune B cells to produce pathogenic antinuclear antibodies. We transfected the TCR-alpha and -beta chain genes of a representative, pathogenic autoantibody-inducing Th clone specific for the nucleosomal core histone peptide H471-94 into TCR-negative recipient cells. Although the autoimmune TCRs were originally derived from SNF1 (I-Ad/q) mice, the transfectants could recognize the nucleosomal autoepitope presented by APC-bearing I-A molecules of all haplotypes tested, as well as human DR molecules. Competition assays indicated that the autoepitopes bound to the MHC class II groove. Most remarkably, MHC-unrestricted recognition of the nucleosomal peptide epitope was conferred by the lupus TCR-alpha chain even when it paired with a TCR-beta chain of irrelevant specificity. Several other disease-relevant Th clones and splenic T cells of lupus mice had similar properties. The TCR-alpha chains of these murine lupus Th clones shared related motifs and charged residues in their CDRs, and similar motifs were apparent even in TCR-alpha chains of human lupus Th clones. The lupus TCR-alpha chains probably contact the nucleosomal peptide complexed with MHC with relatively high affinity/avidity to sustain TCR signaling, because CD4 coreceptor was not required for promiscuous recognition. Indeed, pathogenic autoantibody-inducing, CD4-negative, TCR-alphabeta+ Th cells are expanded in systemic lupus erythematosus. These results have implications regarding thymic selection and peripheral expansion of nucleosome-specific T cells in lupus. They also suggest that universally tolerogenic epitopes could be designed for therapy of lupus patients with diverse HLA alleles. We propose to designate nucleosomes and other antigens bearing universal epitopes "Pantigens" (for promiscuous antigens).     

Punching shear behavior of recycled aggregate concrete slabs with and without steel fibres

A study on the punching shear behavior of 8 slabs with recycled aggregate concrete (RAC) was carried out. The two main factors considered were the recycled coarse aggregate (RCA) replacement percentage and the steel fibre volumetric ratio. The failure pattern, load-displacement curves, energy consumption, and the punching shear capacity of the slabs were intensively investigated. It was concluded that the punching shear capacity, ductility and energy consumption decreased with the increase of RCA replacement percentage. Research findings indicated that the incorporation of steel fibres could not only improve the energy dissipation capacity and the punching shear capacity of the slab, but also effectively improve the integrity of the slab tension surface and thereby changing the trend from typical punching failure pattern to bending-punching failure pattern. On the basis of the test, the punching shear capacity formula of RAC slabs with and without steel fibres was proposed and discussed.     

A new RTD-FET logic family

We describe a new family of clocked logic gates based on the resonant-tunneling diode (RTD). Pairs of RTDs form storage latches, and these are connected by networks consisting of field-effect transistors (FETs), saturated resistors, and RTDs. The design, operation, and expected performance of both a shift register and a matched filter using this logic are discussed. Simulations show that the RTD circuits can achieve higher performance in terms of speed and power in many signal processing applications. Compared to circuits using III-V FETs alone, the RTD circuits are expected to run nearly twice as fast at the same power or at the same speed with reduced power. Compared to circuits using Lincoln Laboratory's fully depleted silicon-on-insulator CMOS, implementation using state-of-the-art RTDs should operate five times faster when both technologies follow the CMOS design rules     

A role for perforin in activation-induced T cell death in vivo: increased expansion of allogeneic perforin-deficient T cells in SCID mice

Despite defective granule exocytosis, T cells from mice whose perforin gene was ablated by homologous recombination (pko mice) caused a similar degree of graft-vs-host disease as normal T cells after injection into sublethally irradiated C.B-17 SCID mice. Moreover host spleens contained significantly greater numbers of T cells from pko mice than from wild-type mice following their i.v. injection. This increase could not be explained by persistence of host APCs that were not cleared by defective donor cytotoxic effector cells. The absence of functional perforin-dependent suppressor cells or an altered cytokine profile of donor T cells could also not account for the behavior of pko cells. Spontaneous and Fas-mediated apoptosis of in vivo activated donor T cells were independent of donor origin. However, pko T blasts exhibited less growth inhibition and cell death after reactivation in vitro. The results are compatible with a model of a defective activation-induced cell death (AICD) pathway, controlled by perforin, accounting for the increased expansion of alloreactive pko T cells.     

Independence of evolutionary and mutational rates after transmission of avian influenza viruses to swine

In 1979, an H1N1 avian influenza virus crossed the species barrier, establishing a new lineage in European swine. Because there is no direct or serologic evidence of previous H1N1 strains in these pigs, these isolates provide a model for studying early evolution of influenza viruses. The evolutionary rates of both the coding and noncoding changes of the H1N1 swine strains are higher than those of human and classic swine influenza A viruses. In addition, early H1N1 swine isolates show a marked plaque heterogeneity that consistently appears after a few passages. The presence of a mutator mutation was postulated (C. Scholtissek, S. Ludwig, and W. M. Fitch, Arch. Virol. 131:237-250, 1993) to account for these observations and the successful establishment of an avian H1N1 strain in swine. To address this question, we calculated the mutation rates of A/Mallard/New York/6750/78 (H2N2) and A/Swine/Germany/2/81 (H1N1) by using the frequency of amantadine-resistant mutants. To account for the inherent variability of estimated mutation rates, we used a probabilistic model for the statistical analysis. The resulting estimated mutation rates of the two strains were not significantly different. Therefore, an increased mutation rate due to the presence of a mutator mutation is unlikely to have led to the successful introduction of avian H1N1 viruses in European swine.     

Molecular cloning of a full-length cDNA for human type 3 adenylyl cyclase and its expression in human islets

The GK (Goto-Kakizaki) rat is a lean model of type 2 diabetes in which the diabetic state was spontaneously induced. We recently demonstrated the presence in GK rats of two functional point mutations in the promoter region of the type 3 adenylyl cyclase (AC3) gene that resulted in overexpression of AC3 mRNA associated with increased cAMP generation. The AC3 gene promoter mutations are the first molecular changes to be described in any specific gene in the GK rat. Here we report cloning of a full-length cDNA encoding human AC3 from a human fetal brain cDNA library using a PCR-based screening method. This 4142-bp cDNA predicts an open reading frame encoding 1144 amino acids containing putative 12 transmembrane-spanning domains which are typically found in other mammalian AC isoforms. Comparison of the translated amino acid sequence of the AC3 gene between human and rat shows 95% homology. Using RT-PCR, clear AC3 expression was detected in isolated human islets as well as a cDNA panel containing templates from eight different tissues (brain, heart, kidney, liver, lung, pancreas, placenta, and skeletal muscle). This wide distribution of AC3 expression may involve a number of physiological and pathophysiological metabolic processes.     

Experimental demonstration that offspring sex ratio varies with maternal condition

Sex ratio theory predicts that, if prevailing ecological or social circumstances differentially influence the fitness benefits of offspring of each sex, parents should adjust their production accordingly to maximize fitness. For species in which sex is chromosomally determined, such as birds and mammals, a differential effect of maternal condition on the fitness of male and female young is one important route whereby selection is expected to favor a bias in the offspring sex ratio at birth or egg laying. However, despite its central place in sex allocation theory, this hypothesis has rarely been tested in wild populations. We manipulated maternal condition upward and downward in a sexually dimorphic wild bird and examined the effect on offspring survival and on offspring sex ratio. The survival to fledging of male, but not female, young was substantially reduced if they came from less well provisioned eggs produced by females in relatively poor condition. As female condition, and thereby her capacity to produce high quality eggs, declined, she progressively skewed the sex ratio of her eggs toward females; i.e., she produced more of the sex with the higher survival prospects. The decline in the survival of male offspring, and the sex ratio bias, was removed when maternal condition was enhanced. These results provide experimental evidence of an adaptive, facultative adjustment of sex ratio in response to changes in maternal condition in wild birds.