Human anti-V3 HIV-1 monoclonal antibodies encoded by the VH5-51/VL lambda genes define a conserved antigenic structure

Preferential usage of immunoglobulin (Ig) genes that encode antibodies (Abs) against various pathogens is rarely observed and the nature of their dominance is unclear in the context of stochastic recombination of Ig genes. The hypothesis that restricted usage of Ig genes predetermines the antibody specificity was tested in this study of 18 human anti-V3 monoclonal Abs (mAbs) generated from unrelated individuals infected with various subtypes of HIV-1, all of which preferentially used pairing of the VH5-51 and VL lambda genes. Crystallographic analysis of five VH5-51/VL lambda-encoded Fabs complexed with various V3 peptides revealed a common three dimensional (3D) shape of the antigen-binding sites primarily determined by the four complementarity determining regions (CDR) for the heavy (H) and light (L) chains: specifically, the H1, H2, L1 and L2 domains. The CDR H3 domain did not contribute to the shape of the binding pocket, as it had different lengths, sequences and conformations for each mAb. The same shape of the binding site was further confirmed by the identical backbone conformation exhibited by V3 peptides in complex with Fabs which fully adapted to the binding pocket and the same key contact residues, mainly germline-encoded in the heavy and light chains of five Fabs. Finally, the VH5-51 anti-V3 mAbs recognized an epitope with an identical 3D structure which is mimicked by a single mimotope recognized by the majority of VH5-51-derived mAbs but not by other V3 mAbs. These data suggest that the identification of preferentially used Ig genes by neutralizing mAbs may define conserved epitopes in the diverse virus envelopes. This will be useful information for designing vaccine immunogen inducing cross-neutralizing Abs.     

SEEDS AND SYSTEMATICS IN HYDROPHYLLACEAE: TRIBE HYDROPHYLLEAE

Scanning electron microscopic investigation of seed surfaces in tribe Hydrophylleae (Hydrophyllaceae) and seed development in Nemophila indicates the presence of four closely related genera (Ellisia, Hydrophyllum, Nemophila, Pholistoma) and one aberrant genus (Eucrypta), which should probably be transferred elsewhere. Comparative evidence from embryology, morphology, cytology, and palynology is roughly corroborative.     

Evaluation of dietary practices of National Collegiate Athletic Association Division I football players

The objective of this study is to evaluate the dietary practices of 28 football athletes on a National Collegiate Athletic Association (NCAA) Division I team using 3-day diet records. Student athletes completed 3-day diet records at 2 individual points of time, when no training table was available. Diet records were evaluated and were compared with the Third National Health and Nutrition Survey (NHANES III) data for the same ages and gender group. No differences in dietary practices of collegiate football athletes were observed when compared with data for the same ages and gender group culled from NHANES III. Inadequacies in energy intake for activity level were significant (p < 0.05). Influences of fad dieting trends were noted when the diets were mapped onto the United States Department of Agriculture (USDA) food guide pyramid. Changes in diet would be necessary to sustain the activity level of these athletes.     

Determination of methadone, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine, 2-ethyl-5-methyl-3,3-diphenylpyraline and methadol in meconium by liquid chromatography atmospheric pressure chemical ionization tandem mass spectrometry

This paper details a validated liquid chromatography atmospheric pressure chemical ionization tandem mass spectrometry (LC-APCI-MS/MS) method for the quantification of methadone, and its metabolites 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), 2-ethyl-5-methyl-3,3-diphenylpyraline (EMDP) and methadol in human meconium. Limits of detection (LOD) were determined to be 1.0 ng/g for methadone, EDDP and EMDP and 2.5 ng/g for methadol. The limits of quantitation (LOQ) for methadone, EDDP, EMDP were 5 and 25 ng/g for methadol. Linearity ranged from 5.0 to 500 ng/g. Following solid-phase extraction, no matrix effect was observed. This method proved to be suitable for the quantification of methadone, EDDP and EMDP and the semi-quantitation of methadol in meconium. Literature review revealed no other published LC-APCI-MS/MS method for the detection of methadone and its three main metabolites in meconium specimens.     

Methionine adenosyltransferase:adrenergic-cAMP mechanism regulates a daily rhythm in pineal expression

(S)-adenosylmethionine (SAM) is a critical element of melatonin synthesis as the methyl donor in the last step of the pathway, the O-methylation of N-acetyl 5-hydroxytryptamine by hydroxyindole-O-methyltransferase. The activity of the enzyme that synthesizes SAM, methionine adenosyltransferase (MAT), increases 2.5-fold at night in the pineal gland. In this study, we found that pineal MAT2A mRNA and the protein it encodes, MAT II, also increase at night, suggesting that the increase in MAT activity is caused by an increase in MAT II gene products. The night levels of MAT2A mRNA in the pineal gland were severalfold higher than in other neural and non-neural tissues examined, consistent with the requirement for SAM in melatonin synthesis. Related studies indicate that the nocturnal increase in MAT2A mRNA is caused by activation of a well described neural pathway that mediates photoneural-circadian regulation of the pineal gland. MAT2A mRNA and MAT II protein were increased in organ culture by treatment with norepinephrine (NE), the sympathetic neurotransmitter that stimulates the pineal gland at night. NE is known to markedly elevate pineal cAMP, and here it was found that cAMP agonists elevate MAT2A mRNA levels by increasing MAT2A mRNA synthesis and that drugs that block cAMP activation of cAMP dependent protein kinase block effects of NE. Therefore, the NE-cAMP dependent increase in pineal MAT activity seems to reflect an increase in MAT II protein, which occurs in response to cAMP-->protein kinase-dependent increased MAT2A expression. The existence of this MAT regulatory system underscores the importance that MAT plays in melatonin biogenesis. These studies also point to the possibility that SAM production in other tissues might be regulated through cAMP.     

Gallium arsenide exposure impairs processing of particulate antigen by macrophages: modification of the antigen reverses the functional defect

Gallium arsenide (GaAs), a semiconductor used in the electronics industry, causes systemic immunosuppression in animals. The chemical's impact on macrophages to process the particulate antigen, sheep red blood cells (SRBC), for a T cell response in culture was examined after in vivo exposure of mice. GaAs-exposed splenic macrophages were defective in activating SRBC-primed lymph node T cells that could not be attributed to impaired phagocytosis. Modified forms of SRBC were generated to examine the compromised function of GaAs-exposed macrophages. SRBC were fixed to maintain their particulate nature and subsequently delipidated with detergent. Delipidation of intact SRBC was insufficient to restore normal antigen processing in GaAs-exposed macrophages. However, chemically exposed cells efficiently processed soluble sheep proteins. These findings suggest that the problem may lie in the release of sequestered sheep protein antigens, which then could be effectively cleaved to peptides. Furthermore, opsonization of SRBC with IgG compensated for the macrophage processing defect. The influence of signal transduction and phagocytosis via Fcgamma receptors on improved antigen processing could be dissociated. Immobilized anti-Fcgamma receptor antibody activated macrophages to secrete a chemokine, but did not enhance processing of unmodified SRBC by GaAs-exposed macrophages. Restoration of normal processing of particulate SRBC by chemically exposed macrophages involved phagocytosis through Fcgamma receptors. Hence, initial immune responses may be very sensitive to GaAs exposure, and the chemical's immunosuppression may be averted by opsonized particulate antigens.     

Changes in Oregon white oak (Quercus garryana Dougl. ex Hook.) following release from overtopping conifers

Oregon white oak or Garry oak (Quercus garryana Dougl. ex Hook.) is a shade-intolerant, deciduous species that has been overtopped by conifers during the past century in parts of its range due to an altered disturbance regime. We examined the response of suppressed Oregon white oak trees in western Washington, USA, to three levels of release from overtopping Douglas-fir (Pseudotsuga menziesii [Mirb.] Franco). We treated individual oak trees with either full release from competition, partial (“half”) release from competition, or a stand-level thinning of Douglas-fir not directed toward release (control). Five years after treatment, oak trees had suffered no mortality or windthrow. Stem diameter growth was 194% greater in the full-release treatment relative to the control. Acorn production varied widely by year, but in years of higher production, acorn production was significantly greater in both release treatments than in the control. Frequency of epicormic branch formation was significantly increased for years 1 and 2 by the full release; the greatest response occurred between 2 and 6 m above ground level. The greatest number of epicormic branches formed on trees on which the majority of original limbs had died back prior to treatment. Trees with relatively less crown dieback at the time of treatment generally had greater stem growth and acorn production responses to release treatments. Our findings indicate that these released Oregon white oak trees are beginning to recover after an extended period of suppression.     

Notch 1 inhibits photoreceptor production in the developing mammalian retina

The transmembrane receptor Notch1 plays a role in development and homeostasis in vertebrates and invertebrates. The mammalian retina is an excellent tissue in which to dissect the precise role of Notch signaling in regulating cell fate and proliferation. However, a systematic analysis has been limited by the early embryonic lethality of Notch1-null mice. Here, Notch1 was conditionally removed from the murine retina either early or late in development. Removal of Notch1 early led to a reduction in the size of the retina as well as aberrant morphology. A decrease in the number of progenitor cells and premature neurogenesis accounted for the reduction in size. Unexpectedly, ablation of Notch1 in early progenitor cells led to enhanced cone photoreceptor production, and ablation of Notch1 at later points led to an almost exclusive production of rod photoreceptor cells. These data suggest that Notch1 not only maintains the progenitor state, but is required to inhibit the photoreceptor fate. These cone enriched mutant mice should prove to be a valuable resource for the study of this relatively rare mammalian photoreceptor cell type.