bluuebunny/crossref_metadata_2025 huggingface dataset:
Strain dependency of B and T lymphoma development in immunoglobulin heavy chain enhancer (E mu)-myc transgenic mice.
Authors: Yukawa K , Kikutani H , Inomoto T , Uehira M , Bin S H , Akagi K , Yamamura K , Kishimoto T
Published: June 2004

The transgenic mice were produced by injecting eggs of B6 and C3H/HeJ mice with the human E mu-myc gene. Preferential development of B lymphomas was observed in the B6 transgenic mice, whereas the C3H/HeJ transgenic mice developed mostly T lymphomas. The phenotypic activation of B lineage cells but not of T lineage cells was detected in the prelymphomatous transgenic mice of both strains. The transgene was similarly expressed in B and T cells of the transgenic mice of both strains. These results suggest that a high incidence of T lymphomas in the C3H/HeJ transgenic mice may not be due to the preferential activation of or the preferential E mu-myc expression in T lymphocytes. When the bone marrow or fetal liver cells from the prelymphomatous transgenic mice of both strains were transferred into irradiated normal C3H/HeJ mice, most of the recipients developed T lymphomas. Moreover, even when irradiated B6 mice received the hematopoietic stem cells from the prelymphomatous B6 transgenic mice, the incidence of T lymphoma increased up to 50%. These findings suggest that B6 and C3H/HeJ mice might provide the environment that supports the development or growth of B and T lymphomas, respectively, and that such an environment could be modified by irradiation of the mice.
Macrophage production of transforming growth factor beta and fibroblast collagen synthesis in chronic pulmonary inflammation.
Authors: Khalil N , Bereznay O , Sporn M , Greenberg A H
Published: June 2004

A rat model of bleomycin-induced pulmonary inflammation and fibrosis was used to examine the relationship between collagen synthesis and transforming growth factor beta (TGF-beta) production, and cellular distribution. Total lung TGF-beta was elevated within 2 h of intratracheal bleomycin administration and peaked 7 d later at levels 30-fold higher than controls. This was followed by a gradual decline with lower but persistent levels of production in the late phase of the response between 21 and 28 d later. The peak TGF-beta levels preceded the maximum collagen and noncollagen protein synthesis measured by [3H]proline incorporation into lung fibroblast explants of bleomycin-treated rats. The pattern of immunohistochemical staining localized TGF-beta initially in the cytoplasm of bronchiolar epithelium cells and subepithelial extracellular matrix. The peak of lung TGF-beta levels at 7 d coincided with intense TGF-beta staining of macrophages dispersed in the alveolar interstitium and in organized clusters. Later in the course of the response. TGF-beta was primarily associated with extracellular matrix in regions of increased cellularity and tissue repair, and coincided with the maximum fibroblast collagen synthesis. This temporal and spatial relationship between collagen production and TGF-beta production by macrophages suggests an important if not primary role for TGF-beta in the pathogenesis of the pulmonary fibrosis.
The contribution of mutant amino acids to alloantigenicity.
Authors: Bill J , Ronchese F , Germain R N , Palmer E
Published: June 2004

The I-Abm12 mutation has been used extensively to study the relationship between structure and function of murine class II major histocompatibility molecules. I-Abm12 differs from I-Ab by three amino acid replacements in the A beta chain, and the proposed structural model of the I-Abm12 molecule places these three amino acid substitutions along one of the alpha-helices where they may affect both antigen and TCR binding. Two of the substitutions, Ile----Phe67 and Thr----Lys71, are thought to point into the binding site, whereas the third substitution, Arg----Gln70, is thought to point up and hence, be available for binding to the TCR. These predicted orientations are consistent with serologic analysis of the bm12 molecule, which demonstrates that residue 70 is uniquely accessible to mAbs distinguishing I-Ab from I-Abm12. In this study we have determined the influence of each of these amino acid substitutions on the ability of the resulting molecules to stimulate a panel of I-Abm12 (allo) reactive T cell hybridomas. Our experiments indicate that reversion of the amino acid at position 70 from Gln (I-Abm12) to Arg (I-Ab) interferes with allorecognition by 33 of 35 I-Abm12-reactive hybridomas. On the other hand, many hybrids can tolerate amino acid substitutions at positions 67 or 71. Single amino acid substitutions at position 67, 70, or 71 are recognized by only a minority of I-Abm12-specific hybrids and usually the reactivity is greatly diminished. These data are most consistent with the idea that the amino acid at position 70 directly interacts with the TCR during allorecognition. The additional effects of residues 67 and 71 are consistent with a contribution by bound peptide to the allorecognition process.
The specificity of rejection and the absence of susceptibility of pancreatic islet beta cells to nonspecific immune destruction in mixed strain islets grafted beneath the renal capsule in the rat.
Authors: Sutton R , Gray D W , McShane P , Dallman M J , Morris P J
Published: June 2004

The specificity of rejection of isolated pancreatic islets was examined in the rat using a quantitative model in which syngeneic (DA) or a mixture of syngeneic and allogeneic (DA and LEW or PVG) islets were implanted beneath the capsule of the kidney of nondiabetic normal rats (DA). 3 wk after transplantation total insulin extraction assays of the kidney with its islet implant together with immunohistological examination of the site of transplantation for evidence of syngeneic or allogeneic tissue demonstrated the total destruction of allogeneic islets without any evidence of damage to syngeneic islets either distant or in immediate proximity to allogeneic islets. Pancreatic islets, and especially beta cells, appear to be particularly vulnerable to the effector arm of both autoimmune and alloimmune responses, a vulnerability that has been attributed to the cytotoxic effects of lymphokines, notably IL-1, released in both autoimmune and alloimmune responses. The experiments reported here demonstrate not only the exquisite specificity of the allograft reaction but are not compatible with a hypothesis that B cells within an intact islet are nonspecifically susceptible to destruction by lymphokines.
A trans-acting major histocompatibility complex-linked gene whose alleles determine gain and loss changes in the antigenic structure of a classical class I molecule.
Authors: Livingstone A M , Powis S J , Diamond A G , Butcher G W , Howard J C
Published: June 2004

The RT1.A locus of the rat MHC encodes the H chain of the single classical class I molecule of this species. One of the alleles of this polymorphic locus, RT1.Aa, is present in several laboratory inbred, congenic, and MHC recombinant rat strains. Studies of the RT1.Aa class I molecule from a number of these strains as a target for CTL show that its antigenicity, both as an alloantigen and a restricting element, is subject to gain and loss alterations by the action of a gene mapping in the MHC to the right of RT1.A. This locus is apparently present in two allelic forms (one possibly a null allele) corresponding to the presence or absence of a dominant transacting modifier, and has been named class I modification, or cim. The antigenic change brought about by cim is scarcely detectable serologically but highly immunogenic for CTL. Biochemical investigations show that cim affects the post-translational modification of RT1.Aa.
Lysis of human melanoma cells by autologous cytolytic T cell clones. Identification of human histocompatibility leukocyte antigen A2 as a restriction element for three different antigens.
Authors: Wölfel T , Klehmann E , Müller C , Schütt K H , Meyer zum Büschenfelde K H , Knuth A
Published: June 2004

From the peripheral blood of the melanoma patient (AV), we derived cytolytic T lymphocyte (CTL) clones that lysed the autologous tumor line SK-MEL-29, but not autologous EBV-B cells, K562, and other tumor targets. By immunoselection experiments it was shown that the CTL clones recognized at least three different antigens on the autologous tumor cells. We demonstrate here that these melanoma antigens are presented to the CTL in association with HLA-A2. First, HLA-A2-reactive pregnancy sera as well as an mAb against HLA-A2 inhibited the CTL lysis. Second, immunoselected melanoma subclones that were resistant to lysis by CTL clones against the three antigens described were found to lack expression of HLA-A2. By sensitizing the patient's lymphocytes against an HLA-A2- melanoma clone, we established a new series of CTL clones recognizing autologous AV melanoma cells. However, efficient lysis was only seen when target cells were pretreated with IFN-gamma. The lytic activity of these CTL was selectively inhibited by an mAb against a common HLA-B determinant. These results indicate that in addition to HLA-A2, other class I antigens are involved in the recognition of AV melanoma cells by autologous CTL.
Isolation and characterization of a distinct immunoregulatory isoform of alpha-fetoprotein produced by the normal fetus.
Authors: van Oers N S , Cohen B L , Murgita R A
Published: June 2004

In this report, we examine the functional significance of the molecular microheterogeneity of alpha-fetoprotein (AFP). In doing so, we have taken the direct approach of purifying the naturally occurring isomeric forms of fetal-derived AFP using a preparative anion exchange column linked to an automated fast protein liquid chromatography (FPLC) system followed by parallel testing of each isolated molecular variant for in vitro immunoregulatory activity. The data obtained demonstrate the presence of seven distinct variants of AFP as defined by their retention volumes on FPLC elution profiles, by their pIs on analytical IEF gels, and by Western blot analysis. Molecular mass determination by SDS-PAGE showed each isomer to be equivalent in size to 69,000-dalton native unfractionated AFP molecules. All the immunosuppressive activity of AFP was localized to a single variant representing only 6% of the total composition of native AFP. The immunoregulating isomer termed AFP-1 was the least acidic of the seven isolated variants with a pI of 5.1 and displayed a sialic acid content of 1 mol/mol of protein. The inhibitory activity of AFP-1 could be readily measured on T cell-dependent antibody synthesis, Con A-induced stimulation of Lyt-1+23- thymocyte DNA synthesis, and lymphokine-activated NK cell activity. All other isomers were without effect in these test systems. The immunosuppressive AFP-1 isomer also displayed the strongest growth-promoting influence on cultured bone marrow lymphocytes. There was no correlation between functional activity and degree of expression of sialic acid residues on the AFP molecules. These findings demonstrate that the immunoregulating function of AFP is confined to a distinct and relatively small subpopulation of native AFP molecules and should therefore contribute to the resolution of outstanding questions regarding the structure/function relationship of this onco-fetal glycoprotein.
Identification and purification of natural killer cell stimulatory factor (NKSF), a cytokine with multiple biologic effects on human lymphocytes.
Authors: Kobayashi M , Fitz L , Ryan M , Hewick R M , Clark S C , Chan S , Loudon R , Sherman F , Perussia B , Trinchieri G
Published: June 2004

We have identified and purified a novel cytokine, NK cell stimulatory factor (NKSF), from the cell-free supernatant fluid of the phorbol diester-induced EBV-transformed human B lymphoblastoid cell line RPMI 8866. NKSF activity is mostly associated to a 70-kD anionic glycoprotein. The purified 70-kD protein, isolated from an SDS-PAGE gel, yields upon reduction two small species of molecular masses of 40 and 35 kD, suggesting that this cytokine is a heterodimer. When added to human PBL, purified NKSF preparations induce IFN-gamma production and synergize with rIL-2 in this activity, augment the NK cell-mediated cytotoxicity of PBL preparations against both NK-sensitive and NK-resistant target cell lines, and enhance the mitogenic response of T cells to mitogenic lectins and phorbol diesters. The three activities remain associated through different purification steps resulting in a 9,200-fold purification, and purified NKSF mediates the three biological activities at concentrations in the range of 0.1-10 pM. These data strongly suggest that the same molecule mediates these three activities, although the presence of traces of contaminant peptides even in the most purified NKSF preparations does not allow us to exclude the possibility that distinct biologically active molecules have been co-purified. The absence of other known cytokines in the purified NKSF preparations, the unusual molecular conformation of NKSF, the high specific activity of the purified protein, and the spectrum of biological activities distinguish NKSF from other previously described cytokines.
Cytokines in chronic inflammatory arthritis. IV. Granulocyte/macrophage colony-stimulating factor-mediated induction of class II MHC antigen on human monocytes: a possible role in rheumatoid arthritis.
Authors: Alvaro-Gracia J M , Zvaifler N J , Firestein G S
Published: June 2004

Granulocyte/macrophage CSF (GM-CSF) has recently been identified in rheumatoid arthritis (RA) synovial effusions. To study a potential role for GM-CSF and other cytokines on the induction of HLA-DR expression on monocytes and synovial macrophages, we analyzed the relative ability of recombinant human cytokines to induce the surface expression of class II MHC antigens on normal peripheral blood monocytes by FACS analysis. GM-CSF (800 U/ml) (mean fluorescence channel 2.54 +/- 0.33 times the control, p less than 0.001) and IFN-gamma (100 U/ml) (5.14 +/- 0.60, p less than 0.001) were the most potent inducers of HLA-DR. TNF-alpha and IL-4 also increased HLA-DR expression, although to a lesser degree [1.31 +/- 0.06 (p less than 0.02) and 1.20 +/- 0.03 (p less than 0.01), respectively]. IL-1 (40 U/ml), IL-2 (10 ng/ml), IL-3 (50 U/ml), IL-6 (100 U/ml), and CSF-1 (1,000 U/ml) did not affect surface HLA-DR density. GM-CSF also increased HLA-DR mRNA expression and surface HLA-DQ expression, but decreased CD14 (a monocyte/macrophage antigen) expression. The effect of GM-CSF on HLA-DR was not mediated by the generation of IFN-gamma in vitro because it was not blocked by anti-IFN-gamma mAb. GM-CSF was additive with IL-4 and low amounts (less than 3 U/ml) of IFN-gamma and synergistic with TNF-alpha. Because we have recently reported that supernatants of cultured RA synovial cells produce a non-IFN-gamma factor that induces HLA-DR on monocytes, we then attempted to neutralize this factor with specific anti-GM-CSF mAb. Four separate synovial tissue supernatants were studied, and the antibody neutralized the HLA-DR-inducing factor in each (p less than 0.01).
Ia-mediated signal transduction leads to proliferation of primed B lymphocytes.
Authors: Cambier J C , Lehmann K R
Published: June 2004

One of the most controversial questions in immunology is the molecular basis by which Th lymphocytes deliver activating signals to quiescent B lymphocytes during T cell-dependent immune responses. Recent studies suggest that T cell-dependent activation of quiescent B lymphocytes may involve signaling mediated by direct T helper cell-B cell contact. Since B cell membrane-associated MHC-encoded class II molecules (Ia) must be recognized by Th lymphocytes for generation of T cell-dependent humoral immune responses, they are obvious candidates for receptors of this signal. Here we report that stimulation of quiescent murine B cells with IL-4 and antibodies against the B cell antigen receptor for 12-16 h primes cells to proliferate in response to immobilized mIa binding ligands. In the presence of additional lymphokines, these B cells differentiate to secrete Ig of IgM and IgG classes. These results suggest that Ia molecules are receptors for direct, T helper cell-B cell contact mediated signaling that results in B cell proliferation.
Inhibition of MHC class II-restricted T cell response by Lyt-2 alloantigen.
Authors: Kanagawa O , Maki R
Published: June 2004

T cell hybridomas were established by fusing a CD8+ V beta 8.1+ CTL clone and a CD4+ V beta 8.1+ helper T lymphocyte (HTL) clone to the thymoma cell line BW5147. In contrast to the HTL x BW hybridomas, which retain the same antigen specificity as the original T cell clone, the CTL x BW hybridomas lost the class I MHC-restricted antigen response but acquired a new specificity to Mlsa antigen. Mlsa reactivity of CTL x BW hybridomas was shown to be mediated by the CTL TCR as assayed by inhibition using an anticlonotypic antibody to the CTL clone. Since hybridomas established with BW5147 lose CD8 expression, we have introduced the CD8 molecule into CTL x BW5147 hybridomas by gene transfection. The CD8+ V beta 8.1+ hybridoma was no longer capable of reacting to Mlsa antigen but exhibited the same antigen specificity as the parental CTL clone. Furthermore, the presence of the transfected CD8 molecule in the HTL x BW hybridomas was found to be inhibitory to class II MHC-restricted antigen reactivity. These results demonstrate that, besides its role in increasing the overall avidity of T cell-class I MHC/antigen interaction, the CD8 molecule inhibits T cell-class II MHC gene product/antigen interaction. This negative effect of the CD8 molecule on a class II MHC-restricted response may account for the failure of CD8+ T cells using either V beta 8.1 or V beta 6, which impart reactivity to the Mlsa antigen on CD4+ T cells, to respond to the Mlsa antigen.
Antagonistic control of tumor necrosis factor receptors by protein kinases A and C. Enhancement of TNF receptor synthesis by protein kinase A and transmodulation of receptors by protein kinase C.
Authors: Scheurich P , Köbrich G , Pfizenmaier K
Published: June 2004

We have investigated control mechanisms of TNF receptor expression (TNF-R) in various human tumor cells and normal peripheral blood monocytes. Activators of protein kinase A (PKA) signal transduction pathways were found to enhance TNF-R expression up to sevenfold, whereas in the same cells, IFN-alpha and -gamma receptors remained unaffected. Inhibitors of protein kinases downregulate both constitutive and cAMP-enhanced TNF-R expression. Binding studies revealed an increase in TNF-R numbers without a change in receptor affinity. Both, direct activators of PKA and inhibitors of phosphodiesterase, raising intracellular levels of cAMP, were found to be effective. As activation of PKA does not slow down the degradation rate of TNF-Rs, but rather enhances protein synthesis-dependent reexpression of TNF-Rs after transient PKC-mediated transmodulation and after tryptic digestion of TNF-Rs, it is concluded that PKA stimulates TNF-R synthesis. Maximum TNF-Rs enhancement is reached after 24 h of stimulation and is reversible, suggesting that receptor upregulation is not linked to irreversible steps of cellular differentiation. PKA-mediated enhancement of TNF-R expression was predominantly observed in normal peripheral blood monocytes and tumor cell lines of myeloid origin. As in these typical TNF producer cells, the production of TNF is also controlled by PKA and PKC, a regulatory circuit is proposed, by which these two independent signal pathways antagonistically regulate TNF production and, at the receptor level, TNF sensitivity.
Murine hepatic accessory cells support the proliferation of Th1 but not Th2 helper T lymphocyte clones.
Authors: Magilavy D B , Fitch F W , Gajewski T F
Published: June 2004

The liver is the major site of clearance and degradation of foreign antigens from the portal circulation. Despite the presence of hepatic accessory cells, antibody responses to orally administered antigens are uncommon. To ascertain if hepatic accessory cells are incapable of stimulating specific subsets of T lymphocytes, freshly isolated hepatic nonparenchymal and splenic cells were cultured with a panel of antigen-specific, H-2-restricted Th1 and Th2 HTL clones. Whereas spleen cells stimulated the proliferation of both Th1 and Th2 clones, hepatic nonparenchymal cells (NPC) stimulated the proliferation of only Th1 and not Th2 clones. Adding rIL-1, rIL-6, and rIL-7, alone or in combination, to the cultures did not result in proliferation of the Th2 clones. Despite the absence of Th2 proliferation, NPC were able to stimulate the secretion of IL-3 and IL-4 by Th2 clones in the presence of antigen. Moreover, adding hepatic NPC did not inhibit spleen cells from stimulating Th2 clones in the presence of antigen. Thus, the inability of liver cells to stimulate the proliferation of Th2 helper T lymphocytes appears to be secondary to an absence of either an unknown accessory cell cofactor or an accessory cell that preferentially presents antigen to Th2 cells. The selective activation of Th1 and not Th2 cells by liver accessory cells may result in suppression of antibody responses to orally administered antigens.
Lack of immunodominance in the T cell response to herpes simplex virus glycoprotein D after administration of infectious virus.
Authors: Yamashita K , Heber-Katz E
Published: June 2004

Glycoprotein D (gD) of HSV has been shown to be a potent immunogen. To analyze the T cell antigenic determinants on gD, a series of 28 overlapping 20-mer peptides that span the extracellular portion of gD-1 were examined for their ability to stimulate T cells from rgD-1 or infectious HSV-1-primed H-2d mice in vitro. rgD-1-primed cells responded exclusively to peptide 241-260, the immunodominant determinant of gD in H-2d mice. In contrast, infectious HSV-primed T cells were shown to respond to 17 (and up to 22) of 28 synthetic gD peptides. These results indicate an extensive diversity in the T cell repertoire to gD in H-2d mice with T cells directed to a broad array of peptide determinants being recruited during the acute phase of an HSV infection.
Species-restricted interactions between CD8 and the alpha 3 domain of class I influence the magnitude of the xenogeneic response.
Authors: Irwin M J , Heath W R , Sherman L A
Published: June 2004

As compared with the vigorous T cell response normally observed against allogeneic MHC molecules, T cells recognize xenogeneic MHC molecules poorly. To define structural features of the MHC molecule important for such species-specific recognition, HLA-A2(A2)-specific murine CTL were examined for their recognition of transfected cell lines expressing the class I molecules A2 or A2/H-2Kb(A2/Kb). A2/Kb is a chimeric molecule consisting of the alpha 1 and alpha 2 domains of A2 and the alpha 3, transmembrane, and cytoplasmic regions of Kb. The majority of CTL clones showed enhanced recognition of transfected cell lines expressing this chimeric molecule. Enhanced recognition was shown to correlate with sensitivity of the CTL clones to inhibition by anti-CD8 antibody. These results suggested that CD8 may interact with class I in a species-specific manner, and that suboptimal CD8 interaction with the alpha 3 domain of xenogeneic molecules may be an important contribution to poor xenoreactivity. This conclusion was supported by the capacity of A2/Kb, but not A2 human cell transfectants, to induce a primary in vitro CTL xenoresponse specific for A2.
Identification of the molecular target for the suppression of contact hypersensitivity by ultraviolet radiation.
Authors: Applegate L A , Ley R D , Alcalay J , Kripke M L
Published: June 2004

This study was conducted to explore the involvement of DNA damage in the suppression of contact hypersensitivity (CHS) by UV irradiation. The opossum, Monodelphis domestica, was used because cells of these marsupials have an enzyme that is activated by visible light (photoreactivating enzyme) and repairs ultraviolet radiation (UVR)-induced pyrimidine dimers in DNA. A single dose of 1,500 J/m2 of UVB (280-320 nm) radiation, representing 2 minimal erythema doses, was administered to the dorsal skin of opossums. This treatment prevented the opossums from developing a CHS response to dinitrofluorobenze (DNFB) applied either at the site of irradiation or an unirradiated site. In addition, this dose of UVR decreased the number of ATPase+ epidermal Langerhans cells in the dorsal epidermis to approximately 3% of that in unirradiated skin at the time of DNFB application. Treatment of the animals with wavelengths that activate the repair enzyme (320-500 nm, photoreactivating light, PRL) for 120 min immediately after UV irradiation inhibited the UVR-induced suppression of CHS almost completely. Exposure to PRL before UVR did not prevent UVR-induced suppression of CHS. PRL treatment after UV irradiation also prevented the decrease in the number of ATPase+ Langerhans cells. Measurements of lesions in DNA indicated that PRL treatment removed around 85% of the UVR-induced pyrimidine dimers. These data provide direct evidence that DNA, and most likely, the pyrimidine dimer, is the primary molecular target for the UVB-induced suppression of contact hypersensitivity to haptens applied to irradiated or unexposed skin.
Activation of CD4 cells by fibronectin and anti-CD3 antibody. A synergistic effect mediated by the VLA-5 fibronectin receptor complex.
Authors: Matsuyama T , Yamada A , Kay J , Yamada K M , Akiyama S K , Schlossman S F , Morimoto C
Published: June 2004

In this study, fibronectin synergized with anti-CD3 antibody to promote CD4 cell proliferation in a serum-free culture system. The cell-adhesive domain plus additional regions of the fibronectin molecule are involved in this synergy. Anti4B4(CDw29) antibody blocked the activation of CD4 cells in this system. Furthermore, it is the VLA-5 protein within the set of molecules recognized by anti-4B4 that serves as a fibronectin receptor on the CD4 lymphocytes. The VLA-5 fibronectin receptor was mainly expressed on CD4+ CD45R-CDw29+ cells and may in part contribute to the unique function of these cells.
Infection of monocyte-derived macrophages with human immunodeficiency virus type 1 (HIV-1). Monocyte-tropic and lymphocyte-tropic strains of HIV-1 show distinctive patterns of replication in a panel of cell types.
Authors: Collman R , Hassan N F , Walker R , Godfrey B , Cutilli J , Hastings J C , Friedman H , Douglas S D , Nathanson N
Published: June 2004

To characterize the host range of different strains of HIV-1, we have used four types of cells, primary monocyte-derived macrophages (MDM), primary PBL, a promonocyte cell line (U937), and a CD4+ T cell line (SUP-T1). These cells were infected with three prototype strains of HIV-1, a putative lymphocyte-tropic strain (IIIB), and two putative monocyte-tropic strains (SF162 and DV). Infections were monitored by assays for infectious virus, for cell-free and cell-associated viral antigen (p24), and for the proportion of cells infected by immunohistochemical staining. It was concluded that: (a) the use of four different cell types provides a useful biological matrix for distinguishing the tropism of different strains of HIV-1; this matrix yields more information than the infection of any single cell type. (b) A monocyte-tropic strain of HIV-1, such as strain SF162, shows a reciprocal host range when compared with a lymphocyte-tropic strain such as IIIB; strain SF162 replicates well in primary MDM but not in U937 or SUP-T1 cells, while strain IIIB replicates well in both U937 and SUP-T1 cells but not in MDM. (c) Both lymphocyte-tropic and monocyte-tropic strains of HIV-1 replicate well in PBL. (d) The promonocyte cell line, U937, and the T cell line, SUP-T1, differ markedly from primary cells, such as MDM and PBL, in their ability to support the replication of different strains of HIV-1; these cell lines cannot be used as surrogates for primary cells in host range studies of HIV-1 strains.
Generation of immunoglobulin heavy chain diversity subsequent to cell surface immunoglobulin expression in the avian bursa of Fabricius.
Authors: Ratcliffe M J
Published: June 2004

The avian bursa of Fabricius represents a site for the generation of antibody diversity. Transfer of neonatal bursal cells into cyclophosphamide-treated neonatal chickens results in reconstitution of recipient bursae with donor-derived bursal stem cells. These stem cells express cell surface IgM and, under conditions of limiting donor bursal stem cell numbers, each reconstituted bursal follicle is colonized by a single precursor cell. The expression of an Ig VH idiotype, CVH-1, was found to be heterogenous within such clonal follicles. The diversity generated within the bursa is subsequently found within the peripheral B cell compartment. Thus, the generation of functional Ig H chain diversity is shown to occur subsequent to Ig H chain rearrangement and expression.
The human mannose-binding protein gene. Exon structure reveals its evolutionary relationship to a human pulmonary surfactant gene and localization to chromosome 10.
Authors: Sastry K , Herman G A , Day L , Deignan E , Bruns G , Morton C C , Ezekowitz R A
Published: June 2004

The human mannose-binding protein (MBP) plays a role in first line host defense against certain pathogens. It is an acute phase protein that exists in serum as a multimer of a 32-kD subunit. The NH2 terminus is rich in cysteines that mediate interchain disulphide bonds and stabilize the second collagen-like region. This is followed by a short intervening region, and the carbohydrate recognition domain is found in the COOH-terminal region. Analysis of the human MBP gene reveals that the coding region is interrupted by three introns, and all four exons appear to encode a distinct domain of the protein. It appears that the human MBP gene has evolved by recombination of an ancestral nonfibrillar collagen gene with a gene that encodes carbohydrate recognition, and is therefore similar to the human surfactant SP-A gene and the rat MBP gene. The gene for MBP is located on the long arm of chromosome 10 at 10q11.2-q21, a region that is included in the assignment for the gene for multiple endocrine neoplasia type 2A.
Evolution of antibody structure during the immune response. The differentiative potential of a single B lymphocyte.
Authors: Manser T
Published: June 2004

Changes in the structure and function of antibodies occur during the course of an immune response due to variable (V) region gene somatic mutation and isotype switch recombination. While the end products of both these processes are now well documented, their mechanisms, timing, and regulation during clonal expansion remain unclear. Here I describe the characterization of antibodies expressed by a large number of hybridomas derived from single B cell clones at an intermediate stage of an immune response. These data provide new insights into the mechanism, relative timing, and potential of V gene mutation and isotype switching. The data suggest that somatic mutation and isotype switching are completely independent processes that may, but need not, occur simultaneously during clonal expansion. In addition, the results of this analysis demonstrate that individual B cell clones are far more efficient than previously imagined at generating and fixing particular V region somatic mutations that result in increased affinity for the eliciting epitope. Models to account for this high efficiency are discussed. Taken together with previous data, the results of this analysis also suggest that the "somatic evolution" of V region structure to a single epitope takes place in two stages; the first in which particular mutations are sustained and fixed by antigen selection in the CDR regions of the V region genes expressed in a clone over a short period of clonal expansion, and the second in which these selected CDR mutations are maintained in the growing clone, deleterious mutations are lost, and selectively neutral mutations accumulate throughout the length of V genes over long periods of clonal expansion.
Lipopolysaccharide (LPS) binding protein opsonizes LPS-bearing particles for recognition by a novel receptor on macrophages.
Authors: Wright S D , Tobias P S , Ulevitch R J , Ramos R A
Published: June 2004

Lipopolysaccharide binding protein (LBP) is an acute-phase reactant that binds bacterial LPS. We show that LBP binds to the surface of live Salmonella and to LPS coated erythrocytes (ELPS), and strongly enhances the attachment of these particles to macrophages. LBP bridges LPS-coated particles to macrophages (MO) by first binding to the LPS, then binding to MO. Pretreatment of ELPS with LBP enabled binding to MO, but pretreatment of MO had no effect. Moreover, MO did not recognize erythrocytes coated with LBP unless LPS was also added, thus suggesting that interaction of LBP with LPS results in a conformational change in LBP that allows recognition by MO. Binding of LBP-coated particles appears to be mediated by a receptor found on blood monocytes and MO but not on other leukocytes or umbilical vein endothelium. The receptor is mobile in the plane of the membrane since binding activity on MO was downmodulated upon spreading of cells on surfaces coated with LBP-LPS complexes. The receptor appears to be distinct from other opsonic receptors since downmodulation of CR1, CR3, Fc gamma RI, Fc gamma RII, and Fc gamma RIII with mAbs did not affect binding of LBP-coated particles, and leukocytes from CD18-deficient patients bound LBP-coated particles normally. Coating of erythrocytes with LBP-LPS complexes strongly enhanced phagocytosis observed in the presence of suboptimal amounts of anti-erythrocyte IgG. However, binding mediated by LBP-LPS complexes alone caused neither phagocytosis of the LBP-coated erythrocytes nor initiation of an oxidative burst. The results of our studies define LBP as an opsonin. During the acute phase, LBP can be expected to bind gram-negative bacteria and bacterial fragments and promote the interaction of coated bacteria with phagocytes.
Insertion/deletion-related polymorphisms in the human T cell receptor beta gene complex.
Authors: Seboun E , Robinson M A , Kindt T J , Hauser S L
Published: June 2004

Insertion/deletion related polymorphisms (IDRP) involving stretches of 15-30 kb within the human TCR-beta gene complex were revealed by pulse-field gel electrophoresis. Two independent IDRP systems were detected by analysis of Sfi I- and Sal I-digested human DNA samples using probes for TCR C and V region gene segments. The allelic nature of these systems was verified in family studies, and mapping data allowed localization of one area of insertion/deletion among the V gene segments and the other near the C region genes. All but one of 50 individuals tested could be typed for the two allelic systems, and gene frequencies for the two allelic forms were 0.37/0.61 and 0.46/0.54, indicating that these polymorphisms are widespread.
Chlamydial disease pathogenesis. The 57-kD chlamydial hypersensitivity antigen is a stress response protein.
Authors: Morrison R P , Belland R J , Lyng K , Caldwell H D
Published: June 2004

Chlamydia trachomatis infection of humans is commonly a localized inflammation that can result in infertility, blindness, and perhaps arthritis. The pathogenic process(es) that cause these sequelae are thought to be immunological. A 57-kD protein that is common among Chlamydia elicits ocular inflammation when introduced onto the conjunctivae of guinea pigs or nonhuman primates previously sensitized by chlamydial infection. This protein is thought to mediate the immunopathology that follows chlamydial infection. To more thoroughly characterize this chlamydial component, we cloned its gene from a C. psittaci strain and identified a particular recombinant that produced the 57-kD polypeptide. The recombinant gene product was immunoreactive with a monospecific anti-57-kD serum, and elicited an ocular inflammation similar to that produced by the 57-kD antigen isolated from chlamydiae. Sequencing identified two ORFs that encode polypeptides of 11.2 and 58.1 kD and are co-transcribed. These two polypeptides show homology with Escherichia coli groE and Coxiella burnetii htp heat-shock proteins. Striking homology (greater than 50%) was found between the 57-kD protein and the HtpB, GroEL, 65-k Mycobacterium tuberculosis and Hsp60 proteins. Thus, the 57-kD chlamydial protein, previously implicated as mediating a deleterious immunologic response to chlamydial infections, is a stress-induced protein similar to those that occur universally in both prokaryotic and eukaryotic organisms.
Characterization of sheep afferent lymph dendritic cells and their role in antigen carriage.
Authors: Bujdoso R , Hopkins J , Dutia B M , Young P , McConnell I
Published: June 2004

We have ablated peripheral lymph nodes in sheep and subsequently cannulated the pseudo-afferent lymphatic vessel that arises as a consequence of afferent lymphatic vessels reanastomosing with the former efferent duct. This technique allows the collection of lymph with a cellular composition that resembles true afferent fluid, and in particular, containing 1-10% dendritic cells. A 16-h collection of this lymph may contain between 10(6) and 10(7) dendritic cells. This dendritic cell population may be enriched to greater than 75% by a single-density gradient centrifugation step. We have generated a mAb that recognizes sheep CD1. This monoclonal not only reacts with afferent dendritic cells, but with dendritic cells in the skin and paracortical T cell areas of lymph nodes. The expression of CD1 suggests afferent dendritic cells are related to skin Langerhans' cells and other dendritic cells that act as accessory cells for T cell responses. Consistent with this is the high level of expression by dendritic cells of molecules involved in antigen recognition by T cells, including MHC class I and class II. Afferent dendritic cells express high levels of the cellular adhesion molecule LFA-3, and at the same time express a ligand for this molecule, namely CD2. The accessory functions of afferent dendritic cells resemble those displayed by mature Langerhans' cells and by lymph node interdigitating cells. These include clustering with resting T cells and stimulating their proliferation in a primary response to antigen. Afferent dendritic cells are capable of acquiring soluble protein antigen in vivo or in vitro and presenting the material directly to autologous T cells in an antigen-specific manner. We conclude that afferent dendritic cells represent a lymph-borne Langerhans' cell involved in antigen carriage to the lymph node.
In vivo modulation of CD1 and MHC class II expression by sheep afferent lymph dendritic cells. Comparison of primary and secondary immune responses.
Authors: Hopkins J , Dutia B M , Bujdoso R , McConnell I
Published: June 2004

The experiments described in this article characterize the phenotypic and functional changes in afferent lymph cell populations that occur as a result of in vivo immune stimulation. During the primary immune response (in antigen-naive sheep) there are very transient increases in level of CD1 expression by subpopulations of dendritic cells (DC) but no alterations in cell kinetics or MHC class II expression. In contrast, secondary antigenic challenge (in primed sheep) into the drainage area of an afferent lymphatic causes profound changes in the cell output, characterized by a greater than threefold drop in total cell output on days 1-3 followed by an approximate fivefold rise on day 5. There is also a substantial increase in both the proportion of MHC class II-positive T lymphocytes (from 28 to 54%) and in the quantitative expression of class II by both DC and lymphocytes. Class II expression by DC increases five- to sixfold by day 5, while the level of expression of class II on lymphocytes approximately doubles. The increase in CD1 expression during the secondary response is more prolonged than during the primary response, being detectable between days 2 and 6 after challenge. The rise in class II affects the whole DC population, in contrast to CD1 where the increase affects only a subpopulation of cells. In terms of functional properties, afferent lymph DC isolated during a primary response show no alteration of their activity, whereas DC taken 4-5 d after secondary challenge are up to fivefold more active in their ability to present soluble antigen to primed autologous T cells and to antigen-specific cell lines as well as to stimulate in the MLR. The relative expression of class II correlates temporally with an increased capacity of DC to present antigen. Monoclonal anti-class II antibodies totally inhibit the in vitro assays but anti-CD1 antibodies have no effect. The previous paper has demonstrated that afferent DC can associate with antigen in vivo and can present that antigen to antigen-specific T cells. This article extends our knowledge of DC biology and demonstrates that DC, activated during secondary in vivo immune responses, have an enhanced ability to present an antigen, unrelated to that used for challenge, to specific T cell lines. This enhancement correlates directly with quantitative variation of expressed class II and not CD1 and suggests that this variation in class II expression plays a physiological role in in vivo immune regulation.
Selective decreases in T cell receptor V beta expression. Decreased expression of specific V beta families is associated with expression of multiple MHC and non-MHC gene products.
Authors: Vacchio M S , Hodes R J
Published: June 2004

Previous reports of TCR V beta usage, studying either expression of a single V beta in a wide panel of strains (6, 7, 10, 12, 13), or expression of multiple V beta s in a very limited strain distribution (14, 15), have identified instances of clonal deletion of potentially autoreactive T cells specific for either self E alpha E beta or minor lymphocyte stimulatory (Mls) antigens. The present study has investigated the range of self antigens that can influence V beta usage by evaluating expression of 16 V beta families in 30 strains of mice. It was found that significant decreases in expression occur in at least 8 of the 16 V beta families and that dominant influences on the T cell V beta repertoire are exerted by expression of Mlsa, Mlsc, and MHC gene products. Decreased expressions of V beta 5, -11, -12, and -16 were influenced by MHC gene products. The patterns of decreased expression seen in intra-MHC recombinant strains and strains of different non-MHC background were distinct for V beta 11, -12, and -16, suggesting that different ligands are involved in the deletion of T cells expressing each of these V beta genes. Mice expressing Mlsa show decreased expression of V beta 9 as well as V beta 6. Mlsc mice lacked V beta 3 expression in those strains where the expressed MHC type was compatible with a strongly stimulatory Mlsc phenotype. V beta 7 was strongly influenced by both MHC and non-MHC products that are not yet identified. These results demonstrate that strain-specific decreases of mRNA expression occur in a major portion of the TCR repertoire. Self antigens including Mlsa, Mlsc, and E alpha E beta, as well as additional MHC and non-MHC products, appear to induce these decreases in expression in the process of eliminating self-reactive T cells from the mature T cell pool.
Autoantibodies encoded by the most Jh-proximal human immunoglobulin heavy chain variable region gene.
Authors: Logtenberg T , Young F M , Van Es J H , Gmelig-Meyling F H , Alt F W
Published: June 2004

Little is known about the utilization of human Ig heavy chain variable gene segments (VH segments) in different B-lineage cell populations or in antibodies of particular specificity and function. We now demonstrate that human antibodies with Ig VH regions encoded by the most JH-proximal human VH segment (VH6) have specificities resembling those of autoantibodies present in sera of patients with systemic lupus erythematosus (e.g., anti-DNA and anticardiolipin). These specificities appear to be encoded by the germline VH6 gene because the activity was found in multiple independent VH6 antibodies in which the light chain varied with respect to isotype and V kappa subgroup. Features of CDR3 length and somatic mutation patterns in several VH6 antibodies suggested that they were selected by the immune system.
Structure and expression of human IgG FcRII(CD32). Functional heterogeneity is encoded by the alternatively spliced products of multiple genes.
Authors: Brooks D G , Qiu W Q , Luster A D , Ravetch J V
Published: June 2004

The structural heterogeneity of the human low affinity receptor for IgG, FcRII(CD32), has been elucidated through the isolation, characterization, and expression of cDNA clones derived from myeloid and lymphoid RNA. These clones predict amino acid sequences consistent with integral membrane glycoproteins with single membrane spanning domains. The extracellular domains display sequence homology to other Fc gamma Rs and members of the Ig supergene family. A minimum of three genes (Fc gamma RIIa, IIa', and Fc gamma RIIb) encode these transcripts, which demonstrate highly related extracellular and membrane spanning domains. IIa/IIa' differ substantially in the intracytoplasmic domain from IIb. Alternative splicing of the IIb gene generates further heterogeneity in both NH2- and COOH-terminal domains of the predicted proteins. Comparison to the murine homologues of these molecules reveals a high degree of conservation between the products of one of these genes, Fc gamma RIIb, and the murine beta gene in primary sequence, splicing pattern, and tissue distribution. In contrast, the sequence of IIa' indicates its relationship to the beta-like genes, with mutation giving rise to a novel cytoplasmic domain, while IIa is a chimera of both alpha- and beta-like genes. Expression of these cDNA molecules by transfection results in the appearance of IgG binding molecules that bear the epitopes defined by the FcRII(CD32) mAbs previously described.
Endothelial receptor-mediated binding of glucose-modified albumin is associated with increased monolayer permeability and modulation of cell surface coagulant properties.
Authors: Esposito C , Gerlach H , Brett J , Stern D , Vlassara H
Published: June 2004

Advanced glycosylation end products (AGE) of proteins accumulate in the vasculature with diabetes and aging, and are thought to be associated with vascular complications. This led us to examine the interaction of AGE-BSA as a prototype of this class of nonenzymatically glycosylated proteins subjected to further processing, with endothelium. Incubation of 125I-AGE-BSA with cultured bovine endothelium resulted in time-dependent, saturable binding that was half-maximal at a concentration of approximately 100 nM. Although unlabeled normal BSA was not a competitor, unlabeled AGE-BSA was an effective competitor of 125I-AGE-BSA-endothelial cell interaction. In addition, AGE modification of two alternative proteins, hemoglobin and ribonuclease, rendered them inhibitors of 125I-AGE-BSA binding to endothelium, although the native, unmodified forms of these proteins were not. At 37 degrees C, binding of 125I-AGE-BSA or gold-labeled AGE-BSA was followed by internalization and subsequent segregation either to a lysosomal compartment or to the endothelial-derived matrix after transcytosis. Exposure of endothelium to AGE-BSA led to perturbation of two important endothelial cell homeostatic properties, coagulant and barrier function. AGE-BSA downregulated the anticoagulant endothelial cofactor thrombomodulin, and induced synthesis and cell surface expression of the procoagulant cofactor tissue factor over the same range of concentrations that resulted in occupancy of cell surface AGE-BSA binding sites. In addition, AGE-BSA increased endothelial permeability, resulting in accelerated passage of an inert macromolecular tracer, [3H]inulin, across the monolayer. These results indicate that AGE derivatives of proteins, potentially important constituents of pathologic vascular tissue, bind to specific sites on the endothelial cell surface and modulate central endothelial cell functions. The interaction of AGE-modified proteins with endothelium may play an important role in the early stages of increased vascular permeability, as well as vessel wall-related abnormalities of the coagulation system, characteristic of diabetes and aging.