bluuebunny/crossref_metadata_2025 huggingface dataset:
The basal phylogeny of Scarabaeoidea (Insecta : Coleoptera) inferred from larval morphology
Authors: Grebennikov Vasily V. , Scholtz Clarke H.
Published: June 2004

Larvae of 60 genera representing the following families and subfamilies of Scarabaeoidea were studied and analysed phylogenetically: Lucanidae (Aesalinae, Nicaginae, Syndesinae, Lampriminae, Lucaninae), Passalidae (Passalinae, Aulacocyclinae), Trogidae, Pleocomidae, Geotrupidae (Taurocerastinae, Lethrinae, Geotrupinae), Bolboceratidae, Ceratocanthidae, Hybosoridae, Glaphyridae, Scarabaeidae (Aphodiinae, Scarabaeinae, Melolonthinae, Dynastinae, Cetoniinae). Seventy-eight larval morphological characters were employed in the analysis. Our data confirm that Dascillidae are not closely related to Scarabaeoidea. The monophyly of the superfamily is supported by 20 apomorphic character states, 18 of them unique. Monophyly of the following scarabaeoid clades is supported (with the number of larval synapomorphies followed by the bootstrap value in parentheses): Scarabaeoidea without Passalidae (6/67), Passalidae (9/100), Pleocomidae (11/93), Trogidae (8/93), Glaphyridae (10/96), Lucanidae (9/95), Ceratocanthidae + Hybosoridae (5/74), Scarabaeinae (9/98). The family Ceratocanthidae was found to be paraphyletic with respect to Hybosoridae. Monophyly of the family Scarabaeidae is not supported. The resolution of the basal parts of the strict consensus tree is higher when using Dascillidae + Eulichadidae v. Agyrtidae + Helophoridae as an outgroup, but the differences in topology become insignificant after bootstrapping. It is suggested that larval morphology alone is not an adequate tool to address basal relationships of Scarabaeoidea and a total evidence analysis should be performed.
The pattern of growth in the early lifecycle of individual Sepia pharaonis
Authors: Minton Jonathan W.
Published: June 2004

The pattern of growth in the early lifecycle of the pharaoh cuttlefish, Sepia pharaonis, was investigated by rearing hatchlings at 26°C in two separate trials. In each trial, the mean weight and mantle length (ML) was recorded in 5-day intervals. In addition, in each trial the growth of 20 group-reared cuttlefish was measured as a control to compare against the individual data. After 60 days of growth, the mean size for individuals in trial 1 was 2.75 g (maximum size 3.32 g) and in trial 2 was 12.76 g (maximum size 14.99 g) at 90 days. Each individual went through distinct growth phases during the first 90 days after hatching. The first growth phase matched exponential curves with an R-value of 0.98 or better, and the second growth phase corresponded with linear and power growth curves at an R-value of 0.98 or better. In trial 1, the mean growth rate for individuals during the first phase was 5.91% BW day–1 and the control group growth rate was 6.36% BW day–1. In trial 2, the mean growth rate for individuals during the first phase was 6.06% BW day–1 and the control group growth rate was 6.70% BW day–1.
Performance of grass and rainforest riparian buffers in the wet tropics, Far North Queensland. 2. Water quality
Authors: McKergow Lucy A. , Prosser Ian P. , Grayson Rodger B. , Heiner Dale
Published: June 2004

Riparian lands have the potential to buffer streams from hillslope sediment and nutrient transport. Most research on buffers has been conducted under laboratory or manipulated field experimental conditions. Few quantitative data exist on buffer performance under natural field conditions. This study reports measured soil loss and evaluates the effectiveness of riparian buffers on planar and convergent slopes under field hydrological conditions in Far North Queensland. The conditions are extreme for testing the effectiveness of riparian buffers as the land is steep, intensely cropped and receives high intensity rainfall. Hillslopes cropped with bananas were monitored using paired flumes. Runoff, bedload, and suspended loads were measured leaving the crop (upper sites) and leaving the riparian buffers (lower sites). Highly variable hillslope soil losses of <1 to >70 t/ha per wet season were recorded. High rates of hillslope soil loss were from areas of steep gradient with little ground cover experiencing high rainfall intensity. On planar slopes, even with high soil loss, grass buffer strips were able to trap >80% of the incoming bedload. Total N (TN), total P (TP) and suspended sediment (SS) loads were reduced between 25 and 65% by the planar slope grass buffer and within the first 15 m of the moderately convergent grass buffer. Loads leaving the moderately convergent buffer were often higher than those delivered from the crop, due to seepage after prolonged or high frequency rainfall. Under these conditions the buffer's main function is to prevent erosion rather than trap sediment and nutrients. Results from a highly convergent 5-ha hillslope, suggest that for buffers to be more effective in such topography, they should also be placed at the end of the crop rows, where contributing areas are smaller. Flow was able to concentrate within the crop and on at least one occasion was able to scour a 30-cm-wide channel through the entire width of the buffer releasing previously trapped material and making the buffer ineffective. A remnant rainforest buffer, receiving runoff from a planar slope, acted as a temporary store of sediment and nutrients that were reworked during subsequent events. This study demonstrates both a need for managed buffer strips on sloping tropical cropped land and identifies limitations on their potential effectiveness.
Spatial variability studies of soil hydro-physical properties using GIS for sustainable crop planning of a watershed of eastern India and its testing in a rainfed rice area
Authors: Kar Gouranga , Singh Ravender , Verma Harsh Nath
Published: June 2004

In this study soil hydro-physical properties such as soil texture, available water capacity, and organic carbon, along with existing land use and topography of a representative watershed of eastern India (Kadalipal watershed, 20.80–20.86°N and 85.54–86.50°E, Dhenkanal district, Orissa), were spatially mapped after collecting data from different sources (ground-truthing, profile survey, remote sensing satellite imagery). Since availability of water is confined to the south-west monsoon period (rainy season, June–September), eastern India is mainly mono-cropped, dominated by rice but second crops can be planned based on the soil hydro-physical properties, mainly water retention and available water of the soil profile utilising residual soil moisture. Keeping the existing land use as a base and considering the potential and prospects of soil hydro-physical properties in different topographies, in this investigation, an alternative land-use plan was developed in different parts of the watershed and tested in its rainfed rice area. Since direct measurement of available water capacity or soil water constants over a large area is time-consuming, pedo-transfer functions were also developed to predict soil water constants using easily measured parameters (e.g. cation exchange capacity, organic carbon, oven-dry bulk density, soil texture, calcium carbonate, etc.) of arable land of the watershed. Results of implementation revealed that the new cropping system was practical for increasing production, productivity, profitability, and sustainability of the rainfed rice area of the eastern India.
Evidence for a significant urbanization effect on climate in China
Authors: Zhou Liming , Dickinson Robert E. , Tian Yuhong , Fang Jingyun , Li Qingxiang , Kaufmann Robert K. , Tucker Compton J. , Myneni Ranga B.
Published: June 2004

China has experienced rapid urbanization and dramatic economic growth since its reform process started in late 1978. In this article, we present evidence for a significant urbanization effect on climate based on analysis of impacts of land-use changes on surface temperature in southeast China, where rapid urbanization has occurred. Our estimated warming of mean surface temperature of 0.05°C per decade attributable to urbanization is much larger than previous estimates for other periods and locations. The spatial pattern and magnitude of our estimate are consistent with those of urbanization characterized by changes in the percentage of urban population and in satellite-measured greenness.
An enantioselective, modular, and general route to the cytochalasins: Synthesis of L-696,474 and cytochalasin B
Authors: Haidle Andrew M. , Myers Andrew G.
Published: June 2004

The cytochalasins are structurally complex natural products with a broad range of apparently unrelated effects in different biological systems. Different members of the family have variously demonstrated inhibitory activity toward the formation of actin filaments, toward the functioning of HIV protease, and toward the process of angiogenesis. The structural series is defined by a largely conserved, rigid bicyclic isoindolone core that is fused to a macrocyclic appendage. The latter structural component varies widely within the cytochalasins and seems to play an important role in the determination of biological activity. In this work, we describe the development of a convergent and enantioselective synthetic route to the cytochalasins that allows for the late-stage introduction of macrocyclic appendages of different sizes and constitutions. We illustrate the route with the synthesis of the 14-membered macrolactone cytochalasin B (1, an inhibitor of the formation of actin filaments) and the 11-membered macrocarbocyclic cytochalasin L-696,474 (2, an inhibitor of HIV protease) by using common precursors.
A one-headed class V myosin molecule develops multiple large (≈32-nm) steps successively
Authors: Watanabe Tomonobu M. , Tanaka Hiroto , Iwane Atsuko Hikikoshi , Maki-Yonekura Saori , Homma Kazuaki , Inoue Akira , Ikebe Reiko , Yanagida Toshio , Ikebe Mitsuo
Published: June 2004

Class V myosin (myosin-V) was first found as a processive motor that moves along an actin filament with large (≈36-nm) successive steps and plays an important role in cargo transport in cells. Subsequently, several other myosins have also been found to move processively. Because myosin-V has two heads with ATP- and actin-binding sites, the mechanism of successive movement has been generally explained based on the two-headed structure. However, the fundamental problem of whether the two-headed structure is essential for the successive movement has not been solved. Here, we measure motility of engineered myosin-V having only one head by optical trapping nanometry. The results show that a single one-headed myosin-V undergoes multiple successive large (≈32-nm) steps, suggesting that a novel mechanism is operating for successive myosin movement.
An
Arabidopsis
homeodomain transcription factor gene,
HOS9
, mediates cold tolerance through a CBF-independent pathway
Authors: Zhu Jianhua , Shi Huazhong , Lee Byeong-ha , Damsz Barbara , Cheng Shie , Stirm Vicki , Zhu Jian-Kang , Hasegawa Paul M. , Bressan Ray A.
Published: June 2004

To investigate essential components mediating stress signaling in plants, we initiated a large-scale stress response screen using
Arabidopsis
plants carrying the firefly luciferase reporter gene under the control of the stress-responsive
RD29A
promoter. Here we report the identification and characterization of a mutant,
hos9-1
(for high expression of o
s
motically responsive genes), in which the reporter construct was hyperactivated by low temperature, but not by abscisic acid or salinity stress. The mutants grow more slowly, and flower later, than do wild-type plants and are more sensitive to freezing, both before and after acclimation, than the wild-type plants. The
HOS9
gene encodes a putative homeodomain transcription factor that is localized to the nucleus.
HOS9
is constitutively expressed and not further induced by cold stress. Cold treatment increased the level of transcripts of the endogenous
RD29A
, and some other stress-responsive genes, to a higher level in
hos9-1
than in wild-type plants. However, the C repeat/dehydration responsive element-binding factor (CBF) transcription factor genes that mediate a part of cold acclimation in
Arabidopsis
did not have their response to cold altered by the
hos9-1
mutation. Correspondingly, microarray analysis showed that none of the genes affected by the
hos9-1
mutation are controlled by the CBF family. Together, these results suggest that HOS9 is important for plant growth and development, and for a part of freezing tolerance, by affecting the activity of genes independent of the CBF pathway.
Human and yeast Rad52 proteins promote DNA strand exchange
Authors: Bi Baoyuan , Rybalchenko Nataliya , Golub Efim I. , Radding Charles M.
Published: June 2004

Studies of
rad52
mutants in
Saccharomyces cerevisiae
have revealed a critical role of Rad52 protein in double-strand break repair and meiosis, and roles in both
RAD51
-dependent and -independent pathways of recombination.
In vitro
, both yeast and human Rad52 proteins play auxiliary roles with RPA in the action of Rad51. Rad52 also has annealing activity and promotes the formation of D-loops in superhelical DNA. The experiments described here show that
Homo sapiens
(Hs)Rad52 and yeast Rad52 proteins promote strand exchange as well. Strand exchange was promoted by the N-terminal domain of HsRad52 that contains residues 1–237, which includes the residues required to form rings of Rad52, whereas other truncated domains, both N-terminal and C-terminal, were inactive. For both yeast Rad52 and HsRad52, the yield of strand-exchange reactions was proportional to the fractional A·T content of the DNA substrates, but both enzymes catalyzed exchange with substrates that contained up to at least 50% G·C. Observations made on
S. cerevisiae
Rad52 protein from mutants with severe recombination deficiencies indicate that the strand-exchange activity measured
in vitro
reflects a biologically significant property of Rad52 protein.
The worldwide costs of marine protected areas
Authors: Balmford Andrew , Gravestock Pippa , Hockley Neal , McClean Colin J. , Roberts Callum M.
Published: June 2004

Declines in marine harvests, wildlife, and habitats have prompted calls at both the 2002 World Summit on Sustainable Development and the 2003 World Parks Congress for the establishment of a global system of marine protected areas (MPAs). MPAs that restrict fishing and other human activities conserve habitats and populations and, by exporting biomass, may sustain or increase yields of nearby fisheries. Here we provide an estimate of the costs of a global MPA network, based on a survey of the running costs of 83 MPAs worldwide. Annual running costs per unit area spanned six orders of magnitude, and were higher in MPAs that were smaller, closer to coasts, and in high-cost, developed countries. Models extrapolating these findings suggest that a global MPA network meeting the World Parks Congress target of conserving 20–30% of the world's seas might cost between $5 billion and $19 billion annually to run and would probably create around one million jobs. Although substantial, gross network costs are less than current government expenditures on harmful subsidies to industrial fisheries. They also ignore potential private gains from improved fisheries and tourism and are dwarfed by likely social gains from increasing the sustainability of fisheries and securing vital ecosystem services.
Evidence for sequential signals in the induction of the arachidonic acid cascade in macrophages.
Authors: Aderem A A , Scott W A , Cohn Z A
Published: June 2004

We have examined the requirement for Na+, Ca2+, and protein synthesis in the induction of the arachidonic acid (20:4) cascade in cultured murine peritoneal macrophages. Replacement of extracellular Na+ with choline or with K+ inhibited receptor-mediated 20:4 release by 60-90%, but did not inhibit release stimulated by the soluble triggers PMA and A23187. Cells that had preingested zymosan particles in a K+ medium could be induced to secrete 20:4 metabolites merely by changing the medium to one containing Na+. The Ca2+ ionophore A23187 caused cells in Na+-free medium to release and metabolize 20:4 to prostacyclin, PGE2, leukotriene C, and hydroxyeicosatetraenoic acids, suggesting that the phospholipase(s), cyclooxygenase, and lipoxygenase enzymes do not have a requirement for extracellular Na+. These data suggest that receptor-mediated 20:4 secretion has a requirement for extracellular Na+, while 20:4 release triggered by soluble stimuli do not. Immune complex- and A23187-induced 20:4 release was absolutely dependent on extracellular Ca2+. PMA-triggered 20:4 secretion was inhibited 50% in Ca2+-free medium, but could be inhibited completely by preloading the cells with the Ca2+ antagonist quinine. Protein and RNA synthesis was required for 20:4 release induced by zymosan, immune complex, and PMA, but not by A23187. Cycloheximide and emetine were effective within 15 min of addition, while actinomycin D was an effective inhibitor within 45 min. We suggest that receptor-mediated signal response coupling in the 20:4 cascade in macrophages comprises a sequential series of signals that includes an Na+ influx, synthesis of a rapid turnover-protein, and finally an increase in intracellular Ca2+.
Intrathymic presentation of circulating non-MHC antigens by medullary dendritic cells. An antigen-dependent microenvironment for T cell differentiation.
Authors: Kyewski B A , Fathman C G , Rouse R V
Published: June 2004

We present evidence for intrathymic presentation of soluble circulating antigens in vivo. Our results show that proteins of different molecular weight enter the mouse thymus rapidly after i.v. injection. The intrathymic presence of antigen was assayed by proliferation of cloned antigen-specific T helper cells, which were cocultured with purified thymic stromal cells; stromal cells were isolated and purified as lymphostromal cell complexes, which preexist in vivo. Antigen presentation copurified with non-adherent medullary dendritic cells (DC) (interdigitating cells). I-A- cortical macrophages forming thymocyte rosettes in vivo and I-A+ cortical epithelial cells forming thymic nurse cells (TNC) in vivo did not act as antigen presenting cells (APC) after antigen pulsing in vivo or in vitro. Thymic APC turn over physiologically and are rapidly replaced (within 2-5 wk) after lethal irradiation by donor bone marrow-derived cells. The frequency of thymocyte-DC interactions in vivo strictly correlates with thymic T cell differentiation, and is independent of the immune status of the animal. Fetal thymic APC seem to be secluded from antigen in the maternal circulation. Thymic DC-ROS probably represent the microenvironment where maturing T cells first encounter non-MHC antigens in the context of self-MHC antigens.
Accessory cell-T lymphocyte interactions. Antigen-dependent and -independent clustering.
Authors: Inaba K , Steinman R M
Published: June 2004

Previous work documented the capacity of dendritic cells (DC) to stimulate primary immune responses and to physically cluster with the responding lymphocytes. Rapid cell-cell aggregation assays were used here to study the interaction of DC and other types of APC with T lymphocytes. Graded doses of APC were sedimented with T cells that had been primed to alloantigens, soluble proteins, or lectin, and then labeled with carboxyfluorescein diacetate. The number of clustered T cells was measured after 10 min at 4 or 37 degrees C. At 4 degrees, binding was antigen-dependent and included greater than 50% of the added T cells. Clustering was mediated by all types of APC tested, including DC, macrophages, B lymphocytes, and fresh Langerhans cells, although DC were the most effective. Specificity was evident in the findings that alloreactive T lymphoblasts bound to allogeneic but not syngeneic APC; KLH- and OVA-reactive T cells bound to syngeneic APC in the presence of specific protein: and Con A blasts needed lectin to cluster. A 30 min pretreatment with chloroquine, a drug known to inhibit APC activity, markedly blocked the specific binding of alloreactive and protein-specific T blasts at 4 degrees C. Since Lyt-2- alloreactive blasts should specifically recognize Ia, presentation of Ia seems to be altered by chloroquine. Binding assays at 37 degrees C gave similar results to those performed at 4 degrees C, with one exception. When DC were used as APC, striking antigen-independent clustering occurred. DC could efficiently cluster primed T cells in the absence of alloantigen, soluble protein, or lectin. We suggest that antigen-independent binding contributes to the distinctive capacity of DC to prime T cells in the afferent limb of the immune response, whereas antigen-dependent binding between other APC and sensitized lymphocytes is critical in the efferent limb.
Functional expression of the murine Thy-1.2 gene in transfected human T cells.
Authors: Gunter K C , Kroczek R A , Shevach E M , Germain R N
Published: June 2004

The interaction of certain mAbs with the Thy-1 molecules of murine T lymphocytes leads to cell activation and proliferation. To examine the signal transduction mechanism underlying this process and to determine what, if any, relationship exists between Thy-1-dependent triggering and T cell activation mediated through the T3-antigen receptor (T3-Ti) complex, a genomic clone of murine Thy-1.2 was isolated and transfected into the human T cell tumor, Jurkat. The transfected gene was actively transcribed in these human cells and high levels of Thy-1.2 glycoprotein were found on the cell membrane. Although certain mAbs to Thy-1.2 failed to bind to the Thy-1 transfected Jurkat cells, several known mitogenic anti-Thy-1 mAbs did react, and in the presence of phorbol ester, induced IL-2 secretion. One Thy-1+ transfectant out of five failed to produce IL-2 in response to anti-T3/Ti antibodies even though it retained the ability to increase intracytoplasmic calcium concentration [( Ca2+]i) in response to these ligands. A Thy-1 negative revertant of this cell regained anti-T3/Ti reactivity, suggesting a regulatory defect in signal transmission via T3/Ti in the original transfectant. These data confirm the ability of Thy-1 to act as an activation receptor for T cells. They reveal a potential role for changes in [Ca2+]i in this process, in common with other pathways of T cell activation, but also indicate a more complex series of events is involved.
Use of a novel colony assay to evaluate the cytotoxicity of an immunotoxin containing pokeweed antiviral protein against blast progenitor cells freshly obtained from patients with common B-lineage acute lymphoblastic leukemia.
Authors: Uckun F M , Gajl-Peczalska K J , Kersey J H , Houston L L , Vallera D A
Published: June 2004

We report a novel colony assay for B-lineage progenitor cells in acute lymphoblastic leukemia (ALL). The primary plating efficiency of blast progenitors freshly obtained from common B-lineage ALL patients varied between 0.09 and 2.63%. Morphological, cytochemical, and immunological analyses of cells from day 7 colonies provided the evidence that they are B-lineage lymphoblasts. Immunological marker analyses of cultured blasts using BA-2 (anti-CD9), BA-3 (anti-CD10), BA-1 (anti-CD24), and B43 mAb have allowed us to define two distinct immunological groups. The first group had BA-2+, BA-3+, BA-1+, B43+ marker profiles, consistent with the phenotype of uncultured bone marrow blasts. The second group differed in that the cells in the blast colonies were BA-3 (anti-CD10)-negative, although many of the cells in the bulk population were BA-3+ before culture. Blasts from both groups were able to proliferate and form secondary colonies when recultured. A pan-B immunotoxin was synthesized by linking B43, a human B cell-specific mAb, to pokeweed antiviral protein (PAP). This study showed that B43-PAP can effectively eradicate leukemic progenitor cells freshly obtained from patients with common B-lineage ALL. B43-PAP eliminated greater than 99.96% of blast progenitors under conditions in which only minimal inhibition of normal bone marrow progenitor cells (CFU-GM, CFU-E, CFU-MK, CFU-GEMM) was observed. Our results establish that the surface determinant recognized by B43 is expressed on B-lineage progenitor cells in ALL, and that these cells are sensitive to PAP at the ribosomal level. To our knowledge, B43-PAP is the first IT to prove effective against common B-lineage ALL cells.
Distribution of blood monocytes between a marginating and a circulating pool.
Authors: van Furth R , Sluiter W
Published: June 2004

Studies on the blood volume of SPF Swiss mice in the steady state led to reevaluation of the distribution of monocytes in the blood compartment. It was concluded from the results of our calculations that monocytes entering the circulation are distributed over a circulating and a marginating pool. The circulating pool accounts for approximately 40%, and the marginating pool or approximately 60% of the population of peripheral blood monocytes.
The major neutral proteinase of Entamoeba histolytica.
Authors: Keene W E , Petitt M G , Allen S , McKerrow J H
Published: June 2004

FPLC anion-exchange and chromatofocusing chromatography were used to purify the major neutral proteinase from secretions of axenically cultured Entamoeba histolytica trophozoites. HM-1 strain trophozoites, which were more proteolytically active than the less virulent HK-9 strain, were used for purification of the enzyme. It is a thiol proteinase with a subunit Mr of approximately 56,000, a neutral pH optimum, and a pI of 6. The importance of this enzyme in extraintestinal amoebiasis is suggested by its ability to degrade a model of connective tissue extracellular matrix as well as purified fibronectin, laminin, and type I collagen. The enzyme caused a loss of adhesion of mammalian cells in culture, probably because of its ability to degrade anchoring proteins. Experiments with a peptide substrate and inhibitors indicated that the proteinase preferentially binds peptides with arginine at P-1. It is also a plasminogen activator, and could thus potentiate host proteinase systems.
Delineation of a defect in T cell receptor beta genes of NZW mice predisposed to autoimmunity.
Authors: Noonan D J , Kofler R , Singer P A , Cardenas G , Dixon F J , Theofilopoulos A N
Published: June 2004

In an attempt to determine whether genes involved in T cell antigen recognition are structurally abnormal and thereby promote murine systemic lupus, we analyzed the structural integrity of the D, J, and C region elements of the T cell receptor alpha and beta chain genes in all major lupus strains and several normal strains. Within the limits of restriction fragment length polymorphism analysis, all strains had an identical genomic organization, except the NZW mice, in which a deletion of the C beta 1-D beta 2-J beta 2 elements was found. Sequence analysis of NZW genomic elements containing this deletion placed its probable origin within the first exon of C beta 1, and extending to a complementary region within the first exon of C beta 2. The significance of this abnormality in the pathogenesis of systemic autoimmune disease remains to be determined.
I-A-restricted T cell antigen recognition. Analysis of the roles of A alpha and A beta using DNA-mediated gene transfer.
Authors: Lechler R I , Ronchese F , Braunstein N S , Germain R N
Published: June 2004

The contributions of A alpha and A beta chains, and of subregions of A beta, to Ia-restricted recognition of antigen by Th lymphocytes were analyzed using a panel of L cells transfected with various pairs of A alpha b,d, or k genes and recombinant or wild-type A beta b,d, or k genes. The A beta genes included all possible exchanges of the whole NH2-terminal (beta 1) domain or halves of the beta 1 domain among these three allelic A beta genes. The Ia+ L cells derived from such transfections were used as antigen-presenting cells with a 21 member panel of responding Ia-restricted T hybridoma cells of differing nominal antigen specificity and Ia-restriction. Special care was taken to account for quantitative variation in levels of Ia expression throughout the experiments. The results of this analysis reveal that (a) only 2 of the 21 Th cells recognized Ia molecules involving either a nonparental A alpha or a nonparental A beta chain, and in both cases the degeneracy extended to only one of the two other alleles tested. This suggests that allele specific contributions from both A alpha and A beta chains are important in restricted recognition for most, if not all I-A-restricted Th cells. (b) In no case did substitution of the A beta 2 domain from either of the alternative haplotypes lead to any functionally detectable effects, demonstrating that polymorphisms in the A beta 1 domain can entirely account for the restriction imposed on Th cell responses by the entire A beta chain. (c) For 90% of the cells tested, replacement of the NH2-terminal portion of the beta 1 domain with an allogeneic segment led to Ia molecules unable to elicit Th responses. Furthermore, of all the cells permissive of the substitution of one or other half of the beta 1 domain, only two permitted the substitution of sequence from both alternative haplotypes. Taken together, these data strongly suggest that antigen recognition by most, if not all, I-A-restricted Th cells involves contributions from both halves of the A beta 1 domain. These data suggest that the role of I-A molecules in restricted Th cell recognition of antigen depends on conformational determinants unique to a particular combination of polymorphic alpha and beta chains, and that multiple such sites exist on a single Ia molecule.(ABSTRACT TRUNCATED AT 400 WORDS)
Compartmentalized regulation of macrophage arachidonic acid metabolism.
Authors: Fels A O , Pawlowski N A , Abraham E L , Cohn Z A
Published: June 2004

We show that downregulation of arachidonic acid (20:4) metabolism which occurs following i.p. injection of C. parvum can occur in a single, localized macrophage population, and is therefore unlikely to be mediated solely by a systemic factor.
Differentiation of dendritic cells in cultures of rat bone marrow cells.
Authors: Bowers W E , Berkowitz M R
Published: June 2004

Although dendritic cells (DC) originate from bone marrow, they were not observed in fresh preparations of bone marrow cells (BMC). Likewise, accessory activity was barely measurable in a sensitive assay for this potent function of DC. However, both DC and accessory activity developed when BMC were cultured for 5 d. Based on fractionation before culture, nearly all of the accessory activity could be attributed to only 5% of the total BMC recovered in a low-density (LD) fraction. The LD-DC precursors differed from mature DC in a number of important respects. Removal of Ia+ cells from the LD fraction by panning did not decrease the production of DC when the nonadherent cells were cultured. Thus, the cell from which the DC is derived does not express or minimally expresses Ia antigens, in contrast to the strongly Ia+ DC that is produced in bone marrow cultures. Irradiation of LD cells before culture prevented the development of DC. When irradiation was delayed by daily intervals, progressive increases in the number of DC resulted, up to the fifth day. These findings, together with preliminary autoradiographic data, indicate that cell division has occurred, in contrast to the DC, which does not divide. We conclude that bone marrow-derived DC arise in culture from the division of LD, Ia- precursors.
Differences in antigen presentation to MHC class I-and class II-restricted influenza virus-specific cytolytic T lymphocyte clones.
Authors: Morrison L A , Lukacher A E , Braciale V L , Fan D P , Braciale T J
Published: June 2004

We have examined requirements for antigen presentation to a panel of MHC class I-and class II-restricted, influenza virus-specific CTL clones by controlling the form of virus presented on the target cell surface. Both H-2K/D- and I region-restricted CTL recognize target cells exposed to infectious virus, but only the I region-restricted clones efficiently lysed histocompatible target cells pulsed with inactivated virus preparations. The isolated influenza hemagglutinin (HA) polypeptide also could sensitize target cells for recognition by class II-restricted, HA-specific CTL, but not by class I-restricted, HA-specific CTL. Inhibition of nascent viral protein synthesis abrogated the ability of target cells to present viral antigen relevant for class I-restricted CTL recognition. Significantly, presentation for class II-restricted recognition was unaffected in target cells exposed to preparations of either inactivated or infectious virus. This differential sensitivity suggested that these H-2I region-restricted CTL recognized viral polypeptides derived from the exogenously introduced virions, rather than viral polypeptides newly synthesized in the infected cell. In support of this contention, treatment of the target cells with the lysosomotropic agent chloroquine abolished recognition of infected target cells by class II-restricted CTL without diminishing class I-restricted recognition of infected target cells. Furthermore, when the influenza HA gene was introduced into target cells without exogenous HA polypeptide, the target cells that expressed the newly synthesized protein product of the HA gene were recognized only by H-2K/D-restricted CTL. These observations suggest that important differences may exist in requirements for antigen presentation between H-2K/D and H-2I region-restricted CTL. These differences may reflect the nature of the antigenic epitopes recognized by these two CTL subsets.
Transport of macrophage Fc receptors and Fc receptor-bound ligands to lysosomes.
Authors: Ukkonen P , Lewis V , Marsh M , Helenius A , Mellman I
Published: June 2004

Mouse macrophage Fc receptors specific for IgG1/IgG2b mediate the binding and pinocytic uptake of soluble IgG-containing antibody-antigen complexes. Internalization of these multivalent IgG complexes is accompanied not only by the intracellular degradation of the ligand, but also by a net decrease in the number of plasma membrane Fc receptors and an accelerated rate of receptor turnover. In contrast, internalized receptors bound to a monovalent ligand, the high affinity Fab fragment of the antireceptor mAb 2.4G2, escape degradation by rapidly recycling to the cell surface. In this paper, we have characterized the intracellular pathway involved in the endocytosis and transport of Fc receptors in the J774 macrophage cell line. The results show that the uptake of multivalent ligands follows the normal pathway of receptor-mediated endocytosis: internalization in clathrin-coated pits and coated vesicles, delivery to endosomes, and finally to acid hydrolase-rich lysosomes. Immunoprecipitation of radiolabeled receptor from Percoll density gradients showed that endocytosis of the IgG complexes also results in the concomitant transport of the receptor to lysosomes. Although uptake of the monovalent Fab fragment had no detectable effect on intracellular receptor distribution, preparations of 2.4G2 Fab rendered multivalent by adsorption to colloidal gold were as effective as the IgG complexes at causing lysosomal accumulation of internalized receptors. Thus, it is likely that the down-regulation and degradation of Fc receptors which occurs during the endocytosis of antibody-antigen complexes is due to the transport of internalized receptors to lysosomes. Moreover, the ability of certain Fc receptor-bound ligands to interfere with receptor recycling and trigger lysosomal transport seems to depend on ligand valency rather than on the presence or absence of Fc domains on intact IgG molecules.
Antibodies to basement membrane heparan sulfate proteoglycans bind to the laminae rarae of the glomerular basement membrane (GBM) and induce subepithelial GBM thickening.
Authors: Miettinen A , Stow J L , Mentone S , Farquhar M G
Published: June 2004

Antibodies specific for the core protein of basement membrane HSPG (Mr = 130,000) were administered to rats by intravenous injection, and the pathologic consequences on the kidney were determined at 3 min to 2 mo postinjection. Controls were given antibodies against gp330 (the pathogenic antigen of Heymann nephritis) or normal rabbit IgG. The injected anti-HSPG(GBM) IgG disappeared rapidly (by 1 d) from the circulation. The urinary excretion of albumin increased in a dose-dependent manner during the first 4 d, was increased 10-fold at 1-2 mo, but remained moderate (mean = 12 mg/24 h). By immunofluorescence the anti-HSPG(GBM) was seen to bind rapidly (by 3 min) to all glomerular capillaries, and by immunoperoxidase staining the anti-HSPG was seen to bind exclusively to the laminae rarae of the GBM where it remained during the entire 2-mo observation period. C3 was detected in glomeruli immediately after the injection (3 min), where it bound exclusively to the lamina rara interna; the amount of C3 bound increased up to 2 h but decreased rapidly thereafter, and was not detectable after 4 d. Mononuclear and PMN leukocytes accumulated in glomerular capillaries, adhered to the capillary wall, and extended pseudopodia through the endothelial fenestrae to contact in the LRI of the GBM where the immune deposits and C3 were located. At 1 wk postinjection, staining for C3 reappeared in the glomeruli of some of the rats, and by this time most of the rats, including controls injected with normal rabbit IgG, had circulating anti-rabbit IgG (by ELISA) and linear deposits of rat IgG along the GBM (by immunofluorescence). Beginning at 9 d, there was progressive subepithelial thickening of the GBM which in some places was two to three times its normal width. This thickening was due to the laying down of a new layer of basement membrane-like material on the epithelial side of the GBM, which gradually displaced the old basement membrane layers toward the endothelium. The results show that the core proteins of this population of basement membrane HSPG (Mr = 130,000), which are ubiquitous components of basement membranes, are exposed to the circulation and can bind anti-HSPG(GBM) IgG in the laminae rarae of the GBM. Binding of these antibodies to the GBM leads to changes (C3 deposition, leukocyte adherence, moderate proteinuria, GBM thickening) considered typical of the acute phase of anti-GBM glomerulonephritis. Antibody binding interferes with the normal turnover of the GBM, presumably by affecting the biosynthesis and/or degradation of basement membrane components.
Interferon-gamma depresses binding of ligand by C3b and C3bi receptors on cultured human monocytes, an effect reversed by fibronectin.
Authors: Wright S D , Detmers P A , Jong M T , Meyer B C
Published: June 2004

Cultivation of human monocytes with recombinant IFN-gamma causes a 5-10-fold depression in their binding of EC3b or EC3bi. This effect is observed within 18 h and is expressed for 5 d in the presence of 100 U/ml IFN-gamma. The capacity of IFN-gamma-treated phagocytes to bind EC3b and EC3bi is fully restored if the phagocytes are allowed to spread for 45 min on surfaces coated with Fn. IFN-gamma-treated cells express normal levels of cell surface C3b and C3bi receptors as measured with monoclonal anti-receptor antibodies, and spreading on Fn does not alter receptor number. We conclude that cultivation with IFN-gamma causes a change in the nature of these receptors that prevents them from interacting with ligand. Immunoelectron microscopy shows that C3bi receptors are expressed on the apical surface of the IFN-gamma-treated MO and that these receptors exhibit normal capacity to migrate in the plane of the membrane. Thus, the nature of the change caused by IFN-gamma is not related to changes in receptor number, location, or mobility. While spreading of IFN-gamma-treated cells on Fn enables C3 receptors to bind ligand, it does not enable them to promote phagocytosis. Treatment of cells with PMA alone does not affect binding or phagocytosis, but treatment of cells with both Fn and PMA enables cells to phagocytose EC3b and EC3bi. These data indicate that the binding and signaling activities of C3 receptors are separately regulated. Fn enables receptors to bind ligand and PMA enables them to signal phagocytosis.
Interleukin 1 and lipopolysaccharide induce an inhibitor of tissue-type plasminogen activator in vivo and in cultured endothelial cells.
Authors: Emeis J J , Kooistra T
Published: June 2004

Human IL-1, recombinant murine IL-1 and E. coli LPS were found to be potent inducers of plasminogen activator (PA)-inhibitor activity, both in vivo, in rats, as well as in cultured human endothelial cells. In vivo, LPS rapidly and dose-dependently (0.01-1,000 micrograms/kg) increased plasma PA-inhibitor activity. Infusion of IL-1 into rats resulted in a small but significant increase in PA-inhibitor activity in rat plasma. Likewise, in cultured human umbilical vein endothelial cells, LPS and IL-1 induced increased synthesis of PA-inhibitor. We suggest that the induced rat plasma inhibitor might be of endothelial origin.
Different classes of T lymphocytes have different mRNAs for the leukocyte-common antigen, T200.
Authors: Lefrancois L , Thomas M L , Bevan M J , Trowbridge I S
Published: June 2004

The leukocyte common antigen, T200, is expressed on all white blood cells but not on other differentiated cells. Within the hematopoietic lineage, specific cell types display characteristic structural forms of the molecule on their surface. We show that murine cytotoxic T lymphocyte clones and helper T cell clones contain different size mRNA for this molecule, and that the early precursors of T200 glycoprotein made in the helper and cytotoxic T cells differ in Mr. Thus, in addition to differences in posttranslational modifications, it is highly likely that a difference in protein structure contributes to the distinct forms of T200 glycoprotein found on these functional T cell subsets.
Involvement of the T cell antigen receptor and of Lyt-2 in the cytotoxic function of aged killer (AK) T cells.
Authors: Jankovic D L , Woodland D , Melchers I , Weltzien H U , Kubo R T , Eichmann K
Published: June 2004

Aged killer (AK) T cells are antigen-independent, IL-2-requiring variants of antigen-dependent CTL clones that have lost their original antigen specificity and have acquired, instead, specific cytotoxicity for P815 target cells. In this report we study whether AK cells use a similar or a different target cell recognition system than that of bona fide CTL. To this end, we selected from a cloned AK line variants that are partially or completely deficient in specific target recognition and/or in cytotoxic function, and analyzed these variants for expression of the T cell antigen receptor and of Lyt-2. Variants were selected from the prototype AK line (Cl 96) with specific, as well as lectin-facilitated, cytotoxicity for P815 tumor cells. Variants could be grouped into four types with increasing degrees of functional deficiency, which correlated with loss of T cell receptor and/or loss of Lyt-2. In short, loss of Lyt-2 was reflected in loss of specific target recognition, and loss of the T cell antigen receptor was reflected in loss of all cytotoxic activity. We conclude from these results that both Lyt-2 and the T cell antigen receptor are required for specific target cell recognition and the T cell antigen receptor is, in addition, required for cytotoxic function. Moreover, since AK cells express a somatically acquired specificity that differs from that of their clonal precursors, it appears that cytotoxic T cells may change their antigen receptor from one specificity to another during tissue culture.
Mapping of a second recombination hot spot within the I-E region of the mouse H-2 gene complex.
Authors: Lafuse W P , Berg N , Savarirayan S , David C S
Published: June 2004

The crossover points of nine intra-I region recombinant mouse strains were determined by restriction fragment analysis. The recombinants were examined for the presence of k and p haplotype specific DNA restriction endonuclease sites. These restriction sites were a Sac I site between the E beta and E beta 2 genes, a Hpa I site within the E beta 2 gene, and a Rsa I site approximately 1 kb to the right of the E alpha gene. Seven of the recombinants were found to have crossovers between the Hpa I and the Rsa I site. This analysis suggests that a recombination hot spot exists within this segment. This segment is approximately 12-14 kb, and contains the E alpha gene and the intervening sequence between the E beta 2 and E alpha genes.
Experimental systemic amyloidosis induced by immunization with syngeneic organ extracts in mice.
Authors: Mori Y , Akikusa B , Mori T , Ueda S , Iesato K , Yoshida H , Ogawa M , Kato I , Wakashin Y , Wakashin M
Published: June 2004

Systemic amyloidosis was induced consistently in mice by intramuscular injection of syngeneic organ (liver and kidney) extracts mixed with CFA six times at weekly intervals. Syngeneic organ extract with CFA also induced amyloidosis of a lesser degree. All three strains of mice (C57BL/6, C3H/He, and BALB/c) injected with a syngeneic liver extract mixed with CFA developed systemic amyloidosis; the most prominent amyloid deposition occurred in C57BL/6 (B6) mice, followed by C3H/He and BALB/c. The amyloid substance deposited in these animals was identified as mouse amyloid A protein (AA). Furthermore, an organ specificity of the immunogen in inducing amyloidosis was suggested with liver and kidney extracts. Primed spleen cells of the immunized B6 mice were fractionated by a nylon-wool column and injected to normal recipient mice via the tail vein. Organs of the recipient mice developed systemic amyloidosis 8 wk after the transfer, and the most prominent histological changes occurred in the recipient mice given nylon-wool column adherent spleen cells. Using anti-Thy-1,2; Ly-1; Ly-2, antibody and complement, it was suggested that T cells, especially Ly-1,2,3+ T cell populations in the primed nylon-wool adherent cells, play an important role in the induction of systemic amyloidosis. It was shown further that the amyloidosis-inducing substance in liver extract was composed of unstable proteins or protein-bound substance.
A lymphokine that activates the cytolytic program of both cytotoxic T lymphocyte and natural killer clones.
Authors: Milanese C , Siliciano R F , Schmidt R E , Ritz J , Richardson N E , Reinherz E L
Published: June 2004

A 10-12 kD lymphokine, herein termed TCAF, was recently shown to be secreted from Th after crosslinking of their antigen/MHC (T3-Ti) receptors. TCAF stimulates resting T lymphocyte proliferation via binding to surface components of the T11 pathway. To determine whether TCAF could induce antigen-independent activation of the lytic machinery of cytotoxic cells, the present studies were conducted. In the presence of TCAF, both T8+ class I MHC-specific and T4+ class II MHC-specific cytotoxic T cell clones were induced to kill targets, including those lacking the appropriate MHC molecules. This effect was unique to TCAF, since IL-1, IL-2, IFN-gamma could not stimulate lytic activity. Furthermore, both T3+T11+ and T3-T11+ NK clones were triggered to lyse NK-resistant target cells. These findings suggest that TCAF can function in an antigen-independent fashion to amplify cytotoxic effector responses.
Administration of recombinant interleukin 2 in vivo induces a polyclonal IgM response.
Authors: Weyand C M , Goronzy J , Dallman M J , Fathman C G
Published: June 2004

We studied the potential immunoenhancing effects of high doses of rIL-2 on murine T and B cell functions in vivo. Injection of rIL-2 caused a threefold or more increase in the frequencies of antigen-specific proliferative T cells, suggesting that rIL-2 initiated a polyclonal T cell response. In primary and secondary humoral immune responses, administration of rIL-2 in vivo selectively enhanced the production of IgM antibodies, whereas the IgG response was unaffected. Coadministration of rIL-2 with antigen failed to induce an isotype switch from IgM to IgG in genetically low-responding mice. Interestingly, in mice treated with rIL-2 alone (in the absence of exogenous antigen), polyclonal IgM production was induced. Polyclonal IgM production of lesser magnitude was found when mice were immunized with specific antigen in the absence of exogenous rIL-2, suggesting that local IL-2 concentrations in a primary immune response might be sufficient to elicit a polyclonal IgM response.