bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of the p53 oncogene in acute myeloblastic leukemia.
Authors: Smith L J , McCulloch E A , Benchimol S
Published: June 2004

We have investigated whether the p53 oncogene is expressed in the blast cells of patients with acute myeloblastic leukemia. p53 protein was detected in the blast cells of 19 out of 34 patients, but not in normal myelopoietic cells. We find a highly significant correlation between p53 protein synthesis in leukemic blast cells and the secondary plating efficiency of these cells (p = 0.0001). The latter provides an estimate for the self renewal capacity of progenitor cells in the blast population. These data indicate that p53 may be involved in leukemic stem cell renewal.
Induction of antibodies to hyaluronic acid by immunization of rabbits with encapsulated streptococci.
Authors: Fillit H M , McCarty M , Blake M
Published: June 2004

The immunogenicity of hyaluronic acid was investigated. Rabbits were immunized with encapsulated group A and C streptococci. Intact long-chain hyaluronate was conjugated to BSA for use as antigen in an ELISA. Antibodies to the hyaluronate-BSA conjugate were detected in peak immune sera. The specificity of the antibodies for both mammalian and streptococcal hyaluronate was shown by inhibition studies. To further confirm the presence of antihyaluronate antibodies, hyaluronidase-digested streptococcal hyaluronate was conjugated to biotin and used as an antigen in the ELISA. A clear immunization effect was shown for each rabbit by the study of preimmune and postimmunization bleedings. Titers for each rabbit increased by greater than 32 - 256 - fold. Inhibition studies using hyaluronidase-digested hyaluronate and periodate-treated hyaluronate showed that the immunodominant site of antibody reactivity was a terminal glucuronic acid residue. Further studies showed that the carboxyl group of the terminal glucuronide was the major immunoreactive site. Both mammalian and streptococcal hyaluronate inhibited the immune rabbit sera reaction to streptococcal hyaluronate, demonstrating crossreactivity of these molecules. Thus, hyaluronate was shown to be immunogenic in rabbits.
Human neutrophil plasminogen activator is localized in specific granules and is translocated to the cell surface by exocytosis.
Authors: Heiple J M , Ossowski L
Published: June 2004

The subcellular localization of plasminogen activator (PA) in human neutrophils was studied. The cells were disrupted by nitrogen cavitation and fractionated on Percoll density gradients into three major components containing the plasma membranes, the specific granules, and the azurophilic granules. The biochemical markers we used to identify these organelles were alkaline phosphatase, vitamin B12-binding protein, and beta-glucuronidase, respectively. Using the radioactive fibrin plate method, PA activity and plasminogen-independent fibrinolytic activity were measured. In resting neutrophils, PA was associated mainly with the membranes of the specific granules. In five individual experiments the activity of this fraction varied from 79 to 100% of the total; the remaining activity was found to be associated with the plasma membrane, and no activity was present in the azurophilic granules. In neutrophils that were activated by exposure to PMA (20 ng/ml for 15 min at 37 degrees C), the total recoverable PA activity remained unchanged; however, the main peak of activity (85% of total) shifted from the specific granules to the plasma membranes. The magnitude of the reduction of the enzyme in the specific granules paralleled that of vitamin B12-binding protein. PMA-activated, intact neutrophils had approximately 12-fold more surface-bound PA activity than resting cells. Recovery of PA activity from neutrophils was critically dependent on pretreatment of the intact cells with DFP before cavitation; 100-fold more PA activity was detected in DFP-pretreated cells. At the same time, this pretreatment reduced the plasminogen-independent fibrinolytic activity by approximately sevenfold. We determined that PA present in the neutrophils is of the urokinase (UK) type and that the enzyme is produced and stored as a pro-UK, a form insensitive to DFP inhibition. The reduction in the level of proteases (measured as fibrinolytic activity) and the resistance of pro-UK to DFP are most likely the two major reasons for the greatly improved recovery of PA from the DFP-pretreated cells. These findings show that in resting neutrophils PA is stored in the specific granules, and that during activation, it translocates to the outer surface of the plasma membranes, thus equipping the cell with an ecto-proteolytic potential.
LFA-1 immunodeficiency disease. Definition of the genetic defect and chromosomal mapping of alpha and beta subunits of the lymphocyte function-associated antigen 1 (LFA-1) by complementation in hybrid cells.
Authors: Marlin S D , Morton C C , Anderson D C , Springer T A
Published: June 2004

Lymphocyte function associated antigen 1 (LFA-1) is a leukocyte cell adhesion protein. We have studied a novel human immunodeficiency disease in which LFA-1 and two other proteins which share the same beta subunit are lacking from the surface of leukocytes. The basis of the inherited defect in cell surface expression of both the alpha and beta subunits of LFA-1 was determined by somatic cell fusion of patient or normal human cells with an LFA-1+ mouse T cell line. Human LFA-1 alpha and beta subunits from normal cells could associate with mouse LFA-1 subunits to form interspecies hybrid alpha beta complexes. Surface expression of the alpha but not the beta subunit of patient cells was rescued by the formation of interspecies complexes. The findings show that the LFA-1 alpha subunit in genetically deficient cells is competent for surface expression in the presence of an appropriate beta subunit, and suggest that the genetic lesion affects the beta subunit. The human LFA-1 alpha and beta subunits were mapped to chromosomes 16 and 21, respectively. The genetic defect is inferred to be on chromosome 21.
HLA-DO polymorphism associated with resistance to type I diabetes detected with monoclonal antibodies, isoelectric point differences, and restriction fragment length polymorphism.
Authors: Schreuder G M , Tilanus M G , Bontrop R E , Bruining G J , Giphart M J , van Rood J J , de Vries R R
Published: June 2004

A new HLA-DQ-related genetic system with two alleles, 2B3 and TA10, defined serologically by MAbs and alloantisera, showed an almost perfect correlation with charge differences on DQ beta molecules, as well as with two polymorphic DNA fragments hybridizing with a DQ beta probe and various restriction enzymes on a panel of 14 DR4+ homozygous typing cells. It was therefore concluded that the serologically defined alleles 2B3 and TA10 are coded by the DQ beta gene and situated on the HLA-DQ beta chain. This 2B3/TA10 polymorphism is independent of HLA-D and segregates with HLA in families. The TA10 allele appears to be a new marker for resistance to type I diabetes, which is independent from the known resistance marker DR2, whereas no association was observed between this DQ beta polymorphism and rheumatoid arthritis.
Induction of the c-myc protooncogene after antigen binding to hapten-specific B cells.
Authors: Snow E C , Fetherston J D , Zimmer S
Published: June 2004

The stimulation of hapten-specific B cell populations with the thymus-dependent antigen, TNP-KLH, was found to induce elevated levels of c-myc mRNA by 2 h. A similar treatment with carrier protein alone did not elevate c-myc mRNA above the level seen in the nonstimulated, resting B cells. These results indicate that antigen binding to the sIg receptor, in the absence of Th cell involvement, directly signals the antigen-binding cell and implicates the active participation of sIg during the process of antigen-mediated B cell activation.
Regulation of colony-stimulating factor 1 during pregnancy.
Authors: Bartocci A , Pollard J W , Stanley E R
Published: June 2004

Pregnancy results in an elevation in serum and tissue concentrations of the mononuclear phagocytic growth factor, CSF-1 (colony-stimulating factor 1). These increases are associated with an increase in the number of monocytes in the circulation, and with increases in the number of splenic macrophage precursors. In contrast to the approximately 2-fold elevation of the CSF-1 concentrations in serum and most tissues, pregnancy results in a 1,000-fold increase in the concentration of uterine CSF-1. The roughly fivefold elevation in uterine CSF-1 concentration observed at day 5 of pregnancy could be mimicked by administration of chorionic gonadotrophin in intact but not ovariectomized mice. These dramatic changes in uterine CSF-1 concentrations may indicate a role for CSF-1 in the regulation of nonmononuclear phagocytic cell types.
Expression of HLA-DR molecules by keratinocytes, and presence of Langerhans cells in the dermal infiltrate of active psoriatic plaques.
Authors: Gottlieb A B , Lifshitz B , Fu S M , Staiano-Coico L , Wang C Y , Carter D M
Published: June 2004

Immunoperoxidase staining of skin sections and immunofluorescence analysis of keratinocyte suspensions obtained from suction blisters of psoriatic plaques were performed using an mAb, Josh 524.4.1, and Fab'2 fragments of a rabbit antiserum, both of which are directed against nonpolymorphic determinants of HLA-DR molecules. HLA-DR+ keratinocytes were present in plaques, but not normal-appearing skin, from a significant portion of patients with active psoriasis. Double-labelling immunofluorescence experiments with either the monoclonal or polyclonal anti-HLA-DR antibody, in conjunction with the mAb OKT6, which identifies DR+ Langerhans cells, demonstrated that HLA-DR molecules were present on OKT6- keratinocytes. The dermal infiltrate of psoriatic plaques contained T cells expressing the activation antigens, IL-2 receptor (Tac) and HLA-DR, as well as macrophages and OKT6+ cells. There was little difference in the characteristics of the dermal infiltrate between the lesions with or without HLA-DR+ keratinocytes. OKT6+ presumptive Langerhans cells were also found in the dermal infiltrates of patients with lichen planus, contact dermatitis, spongiotic dermatitis, erythema multiforme, basal and squamous cell carcinoma. Studies of keratinocyte suspensions showed that 7-84% of keratinocytes were HLA-DR+. Flow cytometry experiments showed that keratinocytes at all stages of differentiation were HLA-DR+. However, the stem cell-enriched population contained the highest proportion of HLA-DR+ cells. HLA-DR expression by keratinocytes correlated with disease activity. The expression was reversible with successful medical therapy. HLA-DR+ keratinocytes may activate T cells directly or may present an as yet unknown antigen to T cells. These studies provide further support for the hypothesis that immunological mechanisms play an important role in the pathogenesis of psoriasis.
Thymic B lymphocyte clones from patients with myasthenia gravis secrete monoclonal striational autoantibodies reacting with myosin, alpha actinin, or actin.
Authors: Williams C L , Lennon V A
Published: June 2004

Striational autoantibodies (StrAb), which react with elements of skeletal muscle cross-striations, occur frequently in patients with thymoma associated with myasthenia gravis (MG). Dissociated thymic lymphocytes from 22 of 72 MG patients secreted StrAb when cultured with PWM. A high yield of EBV-transformed B cell lines was established from thymus, thymoma, and peripheral blood of seven patients with MG, but clones secreting StrAb arose only from the three patients who had StrAb in their sera. The monoclonal StrAb bound to A bands or I bands in skeletal muscle of human, rat, and frog. One bound to mitochondria in addition to myofibrillar I bands. None bound to nuclei, smooth muscle, or gastric mucosal cells. In immunoblot analyses and ELISAs the monoclonal StrAb bound to muscle and nonmuscle isotypes of myosin, alpha actinin, and/or actin. All bound to contractile proteins common to thymus and muscle, and one selectively immunostained epithelial cells of the thymic medulla. From these antigenic specificities we suggest that StrAb might arise as an immune response directed against the cytoskeletal anchoring proteins associated with nicotinic acetylcholine receptors in thymic epithelial cells undergoing neoplastic transformation to thymoma.
Binding and internalization of interleukin 1 by T cells. Direct evidence for high- and low-affinity classes of interleukin 1 receptor.
Authors: Lowenthal J W , MacDonald H R
Published: June 2004

In this report we describe, on the basis of direct IL-1 binding assays and IL-1 internalization studies, the existence of two classes of IL-1-R on a variety of T cell types. Cells of the EL4-6.1 thymoma express large numbers (approximately 20,000 per cell) of IL-1-R that have a Kd of approximately 300 pM for IL-1. Even though these receptors make up 98-99% of the total IL-1-R per cell, they appear to be nonfunctional, based on their inability to endocytose IL-1. A minor class of IL-1-R (200-400/cell) has an approximately 100-fold higher affinity for IL-1 (Kd, approximately 5 pM) and can rapidly internalize the ligand upon binding. All of the biological activity of IL-1 can be shown to occur via binding to high-affinity IL-1-R since the IL-1 concentration giving half-maximum biological activity in EL4-6.1 cells corresponds precisely to the Kd of this class of receptor. Other cell types, including normal T cells, also express both high- and low-affinity IL-1-R, but the absolute number of receptors per cell is considerably less.
Recombinant gamma interferon induces hypertriglyceridemia and inhibits post-heparin lipase activity in cancer patients.
Authors: Kurzrock R , Rohde M F , Quesada J R , Gianturco S H , Bradley W A , Sherwin S A , Gutterman J U
Published: June 2004

Animals suffering from malignancy or chronic infection develop characteristic metabolic abnormalities, including a well-defined hypertriglyceridemic state. These abnormalities have been attributed to release of one or more mediators from activated macrophages. We report that cancer patients receiving RIFN-gamma, a potent macrophage activator, at doses of greater than or equal to 0.25 mg/m2/d i.m. show marked increases in triglyceride but not in cholesterol levels (pretreatment triglyceride level of 180 +/- 190 mg/dl [mean +/- SD] vs. a day-14 level of 370 +/- 242 mg/dl, n = 23, p less than 0.001 by the paired t test). This hypertriglyceridemia was characterized by an increase in very low-density lipoproteins and a decrease in plasma post-heparin lipase activity, consistent with defective triglyceride clearance (mean pretreatment lipase level of 2.1 mumol/ml/h vs. a day-14 level of 1.2 mumol/ml/h, n = 6, p = 0.02 by the paired t test). rIFN-gamma did not directly inhibit lipoprotein lipase enzymatic activity in vitro. Other possible mechanisms of action, such as suppression of lipase by an rIFN-gamma-induced mediator released from activated macrophages, or a direct effect of interferon on lipase biosynthesis, require further investigation. Our observations provide evidence that factors produced by the immune system can regulate lipid metabolism in man.
Defective lymphopoiesis in bone marrow of motheaten (me/me) and viable motheaten (mev/mev) mutant mice. I. Analysis of development of prothymocytes, early B lineage cells, and terminal deoxynucleotidyl transferase-positive cells.
Authors: Greiner D L , Goldschneider I , Komschlies K L , Medlock E S , Bollum F J , Schultz L
Published: June 2004

This study identifies defects in the early stages of lymphopoiesis that may contribute to the abnormalities in the development and/or function of peripheral T and B lymphocytes in mice homozygous for the motheaten (me/me) and viable motheaten (mev/mev) mutations. The results indicate that in me/me and mev/mev mice prothymocytes in bone marrow are present in essentially normal numbers, as determined by intrathymic injection, but apparently lack the ability to home effectively to the thymus, as determined by intravenous transfer; early B lineage cells in bone marrow, identified by the B220 antigen, are markedly depleted, including immature B cells (sIg+), pre-B cells (cIg+, sIg-), and pro-B cells (B220+, cIg-, sIg-); TdT+ bone marrow cells, especially a subset that expresses the B220 B lineage antigen, are markedly depleted by two weeks of age; normal numbers of TdT+ thymocytes are present during the first 3 wk of postnatal life, but rapidly decrease thereafter. The results further indicate that neither the defective thymus homing capacity of prothymocytes nor the deficiency of TdT+ bone marrow cells is due to autoantibodies. The possible relationship of the defective development of lymphoid precursor cells to the premature onset of thymic involution and to the abnormalities of peripheral T and B lymphocytes in me/me and mev/mev mice is discussed; as are the results of in vitro studies (presented in a companion paper), which suggest that a primary defect in the stromal microenvironment of the bone marrow is responsible for the abnormal development of the lymphoid precursor cells.
Isolation of a Treponema pallidum gene encoding immunodominant outer envelope protein P6, which reacts with sera from patients at different stages of syphilis.
Authors: Peterson K M , Baseman J B , Alderete J F
Published: June 2004

A phage directing the synthesis of an abundant 45-kD Treponema pallidum surface protein was isolated from an EMBL-4 bacteriophage lambda library of T. pallidum DNA. The recombinant phage was identified using an mAb that was directed toward an immunodominant, outer envelope T. pallidum protein designated P6. The recombinant P6 protein possessed the same mol mass as the native treponemal antigen detected from total T. pallidum protein preparations, confirming the cloning of the structural gene for this molecule. Furthermore, E. coli was transformed by a 4.5-kb Eco RI lambda insert fragment subcloned into the plasmid vector pUC19. These transformed cells expressed and translocated the 45-kD protein to their outer membranes. Finally, all sera from patients with different stages of syphilis (primary, secondary, and latent) contained antibody reactive to this protein.
Aging and arteriosclerosis. I. Development of myointimal hyperplasia after endothelial injury.
Authors: Hariri R J , Alonso D R , Hajjar D P , Coletti D , Weksler M E
Published: June 2004

Old Fischer 344 rats are more susceptible to vascular lesions after arterial endothelial injury than are young animals. Thus, 20-26-mo-old Fischer 344 rats developed greater and more persistent intimal proliferative lesions than did 2-5-mo-old rats after aortic endothelial denudation. 3 d after deendothelialization, intimal thickness was increased two-fold in both old and young animals. However, 14 d after endothelial injury, intimal thickness had increased nearly five times in old animals, but had regressed to normal in young animals. Intimal thickness of young aortic grafts transplanted into young recipients did not differ significantly from adjacent host aorta or autotransplanted aortic segments 6 wk after surgery. In contrast, intimal thickness of old grafts transplanted into young recipients was eight times greater than adjacent young host aorta 6 wk after surgery. The density of cell nuclei in the intima of old grafts was also much greater than that in young grafts. Thus, in two experimental models of vascular injury, old rats have consistently had greater myointimal hyperplasia than young rats. The increased proliferative response of aortic smooth muscle cells after vascular injury of old animals may contribute to the increased prevalence of vascular disease with age.
T cell suppressors of antitumor immunity. The production of Ly-1-,2+ suppressors of delayed sensitivity precedes the production of suppressors of protective immunity.
Authors: DiGiacomo A , North R J
Published: June 2004

The results of this study show that during growth of the immunogenic Meth A fibrosarcoma, two different types of suppressor T lymphocytes are generated in sequence. One type is generated during early tumor growth, reaches peak number around day 6, and is progressively lost thereafter. It is defined by its ability, upon passive transfer, to suppress the expression of a DTH reaction to tumor antigens in tumor-immunized recipients. It bears the Ly1-,2+ membrane phenotype and is sensitive to relatively low doses of cyclophosphamide. In contrast, the second type of suppressor cell is not detected until after day 9 of tumor growth, and is defined by its ability to inhibit, upon passive transfer, the expression of adoptive immunity against an established tumor in T cell-deficient recipients. According to previous studies it bears the Ly1+,2-, L3T4a+ membrane phenotype and is less sensitive to cyclophosphamide than the T cell suppressor of DTH. It is argued that this second type of suppressor T cell seems likely to be responsible for the escape of immunogenic tumors from antitumor immunity, because it can suppress the expression of a powerful mechanism of antitumor immunity in recipient mice, and is generated progressively as the tumor-bearing host loses concomitant immunity. In contrast, although the Ly-1-,2+ T cell suppressors of DTH can efficiently suppress a DTH reaction to an implant of living tumor cells, they fail to suppress the expression of immunity to the same implant.
Lymphokine-activated killer cells. Analysis of progenitors and effectors.
Authors: Ortaldo J R , Mason A , Overton R
Published: June 2004

IL-2 has been examined for its ability to regulate lymphokine-activated killer (LAK) activity. IL-2 is a potent activator of cytolytic activity against a wide array of tumor cells, including those from fresh autologous and allogeneic tumors. Using subpopulations of lymphoid cells that were separated on Percoll density gradients, and subsequently purified by immunoadsorbance, studies were performed to examine the phenotypes of progenitor and effector cells of human LAK cells and to compare them with the phenotype of activated NK cells. From these studies, it was evident that several lymphoid subsets, including CD3+, CDw16- and CD3-, CDw16+ cells could mediate LAK lysis of fresh tumor cells. Our examination of the kinetics of activation revealed that CDw16+, NKH1+ (NK-active) cells were maximally activated by 1-2 d. In contrast, CD3+ cells appeared not to achieve maximal cytolytic activity against fresh and cultured tumor cells until days 2-3. Using limiting-dilution frequency analysis, we showed that a large percentage of cytolytically active progenitors was present among the CDw16+, NKH1+ cells. The progenitor and effector cell frequencies appear to be 10-50 times higher in these populations compared to CD3+ cells. In addition, the selective blockage by mAb to the CD3 determinant of the T cell receptor complex indicated that these two effector cell phenotypes relied on different receptors to mediate their cytotoxic activity against tumor cells. Therefore, the accumulated data suggest that there is not a single unique progenitor of LAK activity, but rather that multiple subsets of lymphocytes become cytotoxic in response to IL-2. However, the NK cell population forms the largest single component of LAK cell activity in human peripheral blood.
Immunochemical localization and amino acid sequences of crossreactive epitopes within the group A streptococcal M6 protein.
Authors: Jones K F , Khan S A , Erickson B W , Hollingshead S K , Scott J R , Fischetti V A
Published: June 2004

mAbs 10A11, 10B6, and 10F5, raised against the native group A streptococcal M6 protein, were examined for their crossreactivity with non-laboratory passaged clinical isolates, representing 58 M serotypes, by bacterial dot blot immunoassay. mAb 10A11 crossreacted with 9, mAb 10B6 with 30, and mAb 10F5 with 30 different non-M6 serotypes. To identify the epitopes for these antibodies, the native M6 protein was cleaved with pepsin or staphylococcal V8 protease. Resultant peptides were purified by HPLC, examined for binding to crossreactive mAbs in ELISA, and reactive peptides were subjected to amino acid sequence analysis. Peptides were aligned with the amino acid sequence of the entire M6 protein predicted by the DNA sequence of the M6 gene. Competitive inhibition studies using peptides synthesized on the basis of peptide and DNA sequences, in concert with selective blocking of amino acid residues, allowed for the further identification and placement of these crossreactive epitopes within the M6 molecule. The 10A11 epitope was located within the six amino acid residues at position 134-139, which repeat at positions 159-164 and 184-189 within the variable amino terminal half of the native molecule. The conserved 10B6 and 10F5 epitopes were positioned within a 15-amino-acid span at position 275-289, with the possibility that either epitope could have been repeated at residues 239-247. Chemical modification of amino acids within this sequence aided in the differentiation of these two epitopes. Such studies should aid in the recognition of a sequence(s) common to a greater number of M serotypes, which may be useful for future vaccine development or group A streptococcal identification.
Dendritic cell-lymphoid cell aggregation and major histocompatibility antigen expression during rat cardiac allograft rejection.
Authors: Forbes R D , Parfrey N A , Gomersall M , Darden A G , Guttmann R D
Published: June 2004

To determine the pattern of cellular expression of donor MHC class I and class II antigens during the course of rat cardiac allograft rejection, ACI cardiac allografts transplanted to BN recipients were examined from day 2 to day 6 using immunohistologic and immunoelectron microscopic methods. We used both monomorphic and donor-specific mouse anti-rat MHC class I and class II mAbs in this study. In normal ACI hearts, MHC class I reactivity was confined to the vascular endothelium and to interstitial cells. Ongoing rejection was characterized by an increased donor MHC class I staining intensity of microvascular endothelium and induction of donor class I surface reactivity on cardiac myofibers. Donor MHC class II reactivity was exclusively confined to interstitial dendritic cells (IDC) in both normal ACI hearts and in rejecting allografts, although rejection was associated with marked fluctuations in class II IDC frequency. An early numerical depression in class II IDC present in both allografts and syngeneic heart grafts was attributed to a direct effect of the transplantation procedure. By days 3-4, allografts showed an absolute overall increase in donor class II IDC frequency, which was associated with the presence of multiple localized high-density IDC-lymphocyte aggregates. The lymphocytes present in the focal areas were predominantly of the class II-reactive Th cell subpopulation. These aggregates may thus represent the in vivo homologue of dendritic cell-lymphocyte clustering, which has been shown to be required for primary class II allosensitization in the rat and mouse in vitro. During the late phase of rejection, there was a marked numerical fall in donor class II IDC, which correlated with extensive overall graft destruction. This study has shown that acute rat cardiac allograft rejection can occur in the absence of donor MHC class II expression by allograft vascular endothelium and cardiac myofibers. The IDC, which are believed to represent the principal class II alloantigen presenting cells in the rat heart, remain the sole class II-expressing cellular constituents of the graft throughout the course of rejection.
Immune regulation of the L5178Y murine tumor-dormant state. I. In vivo and in vitro effects of prostaglandin E2 and indomethacin on tumor cell growth.
Authors: Liu C M , Okayasu T , Goldman P , Suzuki Y , Suzuki K , Wheelock E F
Published: June 2004

Immunization and intraperitoneal challenge of DBA/2 mice with L5178Y lymphoma cells results in the suppression and maintenance of the L5178Y cells in a tumor-dormant state in the peritoneal cavity for many months. Cell-mediated immune responses involving lymphocytes and macrophages are involved in maintenance of the tumor-dormant state. Macrophages that have increased immunosuppressive activity and that produce increased amounts of PGE2 appear in the peritoneal cavity of tumor-dormant mice before the breakdown of the tumor-dormant state and formation of ascitic tumors. We report here that the tumor-dormant state can be terminated with formation of ascitic tumors by treatment of tumor-dormant mice with PGE2. Treatment with indomethacin results in inhibition of tumor cell growth and elimination of all recoverable tumor cells. Cultures of peritoneal cells (PC) from mice harboring L5178Y cells in a tumor-dormant state were used to analyze the PGE2 and indomethacin effects. Tumor cells did not grow out in the high-cell density PC cultures prepared from many tumor-dormant mice, but addition of PGE2 to these cultures resulted in tumor cell growth. The tumor cell growth that did occur in the PC cultures from some tumor-dormant mice was associated with PGE2 production by the associated host cells, and the addition of indomethacin to these cultures inhibited both PGE2 synthesis and tumor cell growth. Removal of plastic-adherent cells from the PC cultures eliminated the restraint on tumor cell growth. These experiments suggest that L5178Y tumor cells are maintained in a tumor-dormant state by host peritoneal cells, which are under PGE2 regulation.
Novel macrophage receptor for glucose-modified proteins is distinct from previously described scavenger receptors.
Authors: Vlassara H , Brownlee M , Cerami A
Published: June 2004

A high-affinity macrophage receptor has been identified that recognizes proteins modified by a common in vivo process, long-term nonenzymatic reaction of glucose with proteins (AGE proteins). This receptor for glucose-modified proteins is now shown to be distinct from previously described scavenger receptors, using competition and crosscompetition experiments between AGE-modified protein and a variety of in vitro-modified scavenger receptor ligands, including unmodified BSA, unmodified low-density lipoproteins (LDL), acetyl-LDL, maleyl-BSA, and formaldehyde-treated BSA. Furthermore, the specific pattern of AGE-protein receptor inhibition by the polyanionic compounds polyinosinic acid, polyadenylic acid, polyglutamic acid, polycytidylic acid, fucoidin, and heparin was distinctly different from that of acetyl-LDL. By thus selectively recognizing a time-dependent in vivo protein modification, macrophages may preferentially degrade senescent macromolecules, thereby having an important role in the regulation of extracellular protein turnover.
Idiotypic network connectivity and a possible cause of myasthenia gravis.
Authors: Dwyer D S , Vakil M , Kearney J F
Published: June 2004

Extensive idiotypic connectivity has been discovered between the antibodies composing the immune responses against the acetylcholine receptor (AChR) and alpha-1,3-dextran. The idiotypic connections form an elaborate network linking these disparate antigen systems, and there is an hierarchical organization of the antibodies in this network. The key anti-Ids that interconnect these two responses are more crossreactive, lower-affinity antibodies. Interestingly, 15% of patients with MG, which is caused by autoantibodies against the AChR, have serum antibodies against DEX. Control sera are negative for anti-DEX antibodies. Certain anti-DEX antibodies also bind to anti-AChR antibodies via idiotypic interactions. These findings suggest a model for the initiation of autoimmunity in MG. Antibodies made in response to DEX epitopes on the surface of certain bacteria would elicit the production of anti-Ids. However, some of these anti-Ids would also be autoantibodies against the AChR. Thus, is some circumstances, autoimmunity may develop as a consequence of the normal operation of regulatory idiotypic networks.
Macrophage deactivation. Altered kinetic properties of superoxide-producing enzyme after exposure to tumor cell-conditioned medium.
Authors: Tsunawaki S , Nathan C F
Published: June 2004

Incubation of activated mouse peritoneal macrophages with tumor cell-conditioned medium (TCM) results in their deactivation, as measured by ability to release reactive oxygen intermediates and kill protozoal pathogens. The mechanism of suppression by macrophage deactivation factor (MDF) was studied. Inhibition of H2O2 release could not be overcome by increasing the concentration of phorbol diesters used to trigger the respiratory burst. Deactivated macrophages consumed H2O2 at the same rate as activated cells (t1/2, 35-40 min for 25 nmol H2O2 per 10(6) peritoneal cells). They transported glucose with the same kinetics (Km, 1 mM; Vmax, approximately 100 nmol per 6 min per milligram cell protein), and maintained similar intracellular concentrations of NADPH and NADP (approximately 0.62 mM and approximately 0.11 mM, respectively), as measured by enzymatic cycling methods and determinations of the volume of cell water (3.6 microliter/mg cell protein). To study the kinetics of the PMA-triggered NADPH oxidase in cell lysates, mixed detergents were used (deoxycholate and Tween 20). These stabilized the oxidase for approximately 3.3-fold longer than deoxycholate alone, which was used in previous studies. Incubation of activated macrophages in MDF resulted in a marked increase in the Km of the oxidase for NADPH, from 0.06 mM to 0.67 mM. The Vmax fell approximately 1.7-fold. These kinetic changes, together with the measured intracellular concentration of NADPH, account quantitatively for the suppression of H2O2 release by deactivated macrophages, and are nearly the mirror image of the kinetic changes observed during macrophage activation.
Macrophage type 3 complement receptors mediate serum-independent binding of Leishmania donovani. Detection of macrophage-derived complement on the parasite surface by immunoelectron microscopy.
Authors: Wozencraft A O , Sayers G , Blackwell J M
Published: June 2004

In this study, direct visual evidence for local opsonization of L. donovani by macrophage (M phi)-derived complement components was obtained using immunoelectron microscopy. C3 deposition was detected on the surface of both promastigotes and amastigotes after 20 min serum-free incubation with murine resident peritoneal M phi (RPM), followed by fixation and incubation first with specific antibody directed against C3 and then with gold-labelled protein A. Gold deposition was not observed around either form of the parasite if the anti-C3 antibody was omitted. For promastigotes, the degree of C3 deposition under serum-free conditions was comparable with that observed in the presence of an exogenous (serum) source of C3, but did not result in the same severe damage to the parasite as did the latter. Addition of sodium salicyl hydroxamate, which prevents covalent binding of C3 to activator surfaces, abrogated promastigote binding. Hence, although the anti-C3 antibody did not distinguish between native C3 and its breakdown product iC3b, these data support our earlier conclusion that promastigote binding to the CR3 of murine RPM is complement dependent. For amastigotes, gold deposition and binding to murine RPM were not eliminated by sodium salicyl hydroxamate. The presence of normal mouse serum resulted in increased gold deposition, but did not mediate either enhanced binding to M phi or damage to the amastigote. These data suggest that a proportion of C3 binding to the amastigote surface may be via noncovalent linkages, and that the C3 bound may not be in the correct form to mediate binding to CR3.
Release of the rat T cell alloantigen RT-6.2 from cell membranes by phosphatidylinositol-specific phospholipase C.
Authors: Koch F , Thiele H G , Low M G
Published: June 2004

The mechanism by which the rat T cell alloantigen, RT-6.2, is attached to the membrane was investigated. Treatment of rat lymph node and T-hybridoma cells with phosphatidylinositol-specific phospholipase C (PI-PLC) caused a substantial reduction in the amount of RT-6.2 on the cell surface. No significant release of a rat T helper marker (visualized by the mAb W3/25) was observed in response to PI-PLC treatment. This is in sharp contrast to the effects of trypsin, which removes most of the T helper marker but had little effect on RT-6.2. SDS-PAGE analysis of the RT-6.2 released by PI-PLC indicated that the Mr was not significantly changed by this treatment. Phase separation of the released RT-6.2 in Triton X-114 showed that the PI-PLC had converted it from an amphiphilic membrane form to a water-soluble form, apparently by removing its hydrophobic membrane anchoring domain. These results strongly suggest that RT-6.2, in common with Thy-1 and several other cell surface proteins, is anchored in the membrane by the 1,2-diacylglycerol moiety of a covalently attached phosphatidylinositol molecule.
Effects of activation on lipoprotein lipase secretion by macrophages. Evidence for autoregulation.
Authors: Behr S R , Kraemer F B
Published: June 2004

Lipoprotein lipase (LPL) activity was measured in the media of cultured mouse peritoneal macrophages that were isolated after the intraperitoneal injection of inflammatory agents in order to yield a variety of states of activation. Fully activated macrophages obtained from Corynebacterium parvum-injected mice secreted very low levels of LPL when compared to unstimulated macrophages, while inflammatory and primed macrophages had increased LPL secretion. When inflammatory macrophages were incubated with conditioned medium obtained from fully activated macrophages, LPL secretion decreased in a time- and dose-dependent fashion. The factor(s) secreted by fully activated macrophages that inhibited LPL secretion was shown to be thermolabile and distinct from tumor necrosis factor. These results demonstrate that activation dramatically alters macrophage LPL secretion.
Streptococcal M protein size mutants occur at high frequency within a single strain.
Authors: Fischetti V A , Jarymowycz M , Jones K F , Scott J R
Published: June 2004

Streptococcal M protein, the antiphagocytic molecule on the surface of the organism, was previously found to exhibit extensive size heterogeneity between as well as within M serotypes. In this study, methods were devised to isolate M protein size mutants within a laboratory-grown culture. We were able to isolate three independent M protein deletion mutants and one additional mutant, which was derived from the first deletion mutant. We found that these deletion mutants occur at a frequency of approximately 1 in 2 X 10(3) CFUs in culture. Functional studies revealed that the deletion mutants were able to survive as well as the parental strain in human blood. They also had the determinants necessary to absorb opsonic antibodies as well as the parent. Pepsin digestion experiments localized the deletions within the N-terminal half of the M molecule, which is distal to the cell wall surface. This is the region of the molecule in which extensive sequence repeats are found. This is consistent with the suggestion that the size changes may be the result of homologous recombination between the repeat regions in the gene. These results support the idea that strains showing M protein size variation within successive clinical isolates from single patients may be derived from the initial infecting organisms, and are not the result of separate unrelated acquisitions of the same serotype. This size change may be important in the survival of the streptococcus in vivo.
Membrane-bound C4b interacts endogenously with complement receptor CR1 of human red cells.
Authors: Kinoshita T , Medof M E , Hong K , Nussenzweig V
Published: June 2004

Activation of the classical complement pathway on the membrane of autologous cells results in the deposition of C4b on their surface and in the assembly of the C3 convertase C4b2a, one of the amplifying enzymes of the cascade. Here we study the sequence of events leading to irreversible inactivation of the potentially harmful C4b bound to human red cells. We show that deposited C4b interacts endogenously with complement receptor type 1 (CR1) present on the membrane of the same red cell. Complexes containing CR1 and C4b are found in extracts of membranes of C4b-bearing red cells after treatment of the intact cells with a bifunctional crosslinking reagent. The amount of complexed CR1 increases with the number of deposited C4b molecules. Only small amounts of free CR1 are observed on red cells bearing as few as 1,900 molecules of C4b, suggesting that the binding avidity between C4b and endogenous CR1 is high. In agreement with this observation, we find that the deposited C4b inhibits the exogenous cofactor activity of the red cell CR1 for the factor I-mediated cleavage of target-bound clustered C3b. The C4b bound to the human red cells is cleaved by the serum enzyme C3b/C4b inactivator (factor I) and a large fragment (C4c) is released in the incubation medium. The cleavage is totally inhibited by mAbs against CR1, showing that the complement receptor is an essential cofactor for the activity of I. When the number of bound C4b per red cell is relatively small (less than 1,000 molecules) the substrate for the enzymatic activity of factor I is mostly or exclusively the C4b bound endogenously to CR1. Indeed, the kinetics or the extent of cleavage of C4b are not affected by greatly augmenting the concentration of exogenous CR1 or of C4b-bearing red cells in the incubation mixture, thereby increasing the frequency of collisions between CR1 on the surface of one cell with C4b deposited on the membrane of a different cell. On the basis of the present and prior observations, we speculate that both DAF and CR1 act endogenously to inactivate the function of autologous red cell-bound C4b and prevent the progression of the cascade. DAF binding prevents the formation of the C3 convertase, C4b2a. The cleavage and irreversible inactivation of C4b only occurs after the concerted activities of endogenous CR1 and serum factor I.(ABSTRACT TRUNCATED AT 400 WORDS)
HLA B37 determines an influenza A virus nucleoprotein epitope recognized by cytotoxic T lymphocytes.
Authors: McMichael A J , Gotch F M , Rothbard J
Published: June 2004

Human influenza A virus-specific, cytotoxic T cells have been shown previously to recognize the virus nucleoprotein on infected cells. CTL preparations from four HLA B37-positive donors were shown to recognize a synthetic peptide that corresponded to amino acids 335-349 of the nucleoprotein sequence. Influenza-specific CTL from 10 donors of other HLA types failed to recognize this epitope. CD8+ CTL lines were derived from lymphocytes of two HLA B37-positive donors and used to show that the peptide was represented on virus-infected cells and to determine the probable boundaries of the epitope.
The human invariant chain is the core protein of the human class II-associated proteoglycan.
Authors: Giacoletto K S , Sant A J , Bono C , Gorka J , O'Sullivan D M , Quaranta V , Schwartz B D
Published: June 2004

The human class II-associated chondroitin sulfate proteoglycan (CSPG) was analyzed biochemically and immunologically to determine a possible relationship with the human invariant chain (gamma 1) and its related components. The CSPG was purified by a three-step procedure involving associative ion-exchange chromatography, immunoprecipitation, and dissociative ion-exchange chromatography. Treatment of the CSPG with chondroitinase revealed core proteins of Mr approximately 46,000, 38,000, and 28,000, with the 38,000 species most highly represented. Tryptic peptide analysis revealed identity of the peptides of the 38,000 Mr core protein and gamma 1, and of the 28,000 Mr species and p25. The CSPG and its core proteins were shown to react directly with the mouse anti-human invariant chain monoclonal antibody VIC-Y1 and a rabbit antiserum produced against a synthetic peptide corresponding to the C-terminal end of invariant chain. These results demonstrate that the invariant chain is the core protein of the class II-associated CSPG. In addition, virtually all the CSPG was shown to be present on the cell surface.
Immunologically relevant peptide antigen exists on the presenting cell in a manner accessible to macromolecules in solution.
Authors: Cease K B , Buckenmeyer G , Berkower I , York-Jolley J , Berzofsky J A
Published: June 2004

Although studies of the association of antigen with APC have been complicated by antigen-processing requirements, recent studies have suggested that immunologically relevant antigen should be present on the APC surface. Nevertheless, blocking of antigen presentation with antibody to the antigen has not been demonstrable in most systems. To study this problem we developed a system using avidin to block presentation of amino-terminal biotinylated synthetic peptide 132-146 of sperm whale myoglobin (B132) to a murine T cell clone specific for this site in association with I-Ed. greater than 95% specific inhibition was observed with doses of B132 equipotent to unmodified peptide. Specific blocking could be observed: (a) after pulsing APC with antigen, washing, and incubating for a chase period of 8-16 h before addition of avidin and T cells to assure adequate time for intracellular trafficking and maximal display of antigen on the cell surface, or (b) when monensin is present during the antigen pulse to inhibit such traffic. Therefore, the inhibition appeared to be occurring at the cell surface unless dissociation and reassociation were constantly occurring. To distinguish these, B10.GD APC (I-Ed-negative) were pulsed with antigen and cocultured with B10.D2 APC (I-Ed-positive). No detectable antigen presentation resulted. Thus, minimal dissociation and reassociation between antigen and APC occurs and, consequently, blocking by extracellular solution-phase binding of avidin to antigen is unlikely. Taken together, these data suggest that the blocking is occurring at the cell surface. Thus, under physiologic conditions, immunologically relevant antigen necessary for T cell activation appears to be present on the APC surface and is freely accessible to macromolecules the size of avidin. These findings hold specific implications for models of antigen presentation for T cell recognition.
Fc epsilon receptor, a specific differentiation marker transiently expressed on mature B cells before isotype switching.
Authors: Kikutani H , Suemura M , Owaki H , Nakamura H , Sato R , Yamasaki K , Barsumian E L , Hardy R R , Kishimoto T
Published: June 2004

The expression of Fc epsilon R on human lymphocytes was studied with the anti-Fc epsilon R mAbs. Fc epsilon R was expressed on most mu+,delta+ circulating B cells, whereas T cells did not express Fc epsilon R even in patients with hyper-IgE syndrome. B cells with gamma, alpha, or epsilon phenotype did not express Fc epsilon R, moreover its expression could not be induced, suggesting that the Fc epsilon R expression was correlated with isotype switching. mu+delta+ B cells in bone marrow did not express Fc epsilon R, but PHA-sup (supernatant from PHA-stimulated cell cultures) could induce its expression, and the addition of IgE augmented this induction. Recombinant IL-2, IL-1, IFN-gamma or -beta, or purified B cell differentiation factor (BSF-2 B cell-stimulatory factor 2) could not induce Fc epsilon R expression in bone marrow B cells. IFN-gamma inhibited the Fc epsilon R expression induced by PHA-sup, suggesting that the human counterpart of BSF-1 may be responsible for Fc epsilon R expression in bone marrow B cells. B cells from patients with common variable immunodeficiency and ataxia telangiectasia did not express Fc epsilon R, but PHA-sup could induce its expression, indicating that circulating B cells of these patients are at a differentiation stage similar to B cells in bone marrow. The study showed that Fc epsilon R is a B cell-specific differentiation marker, the expression of which is restricted to a defined stage of B cell differentiation.