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Biomedical subjects

S C Peiper

Publications and source records attributed to S C Peiper.

At least 55 records · Page 3Linked to original sources
PubMed1996

Identification of a major co-receptor for primary isolates of HIV-1.

Entry of HIV-1 into target cells requires cell-surface CD4 and additional host cell cofactors. A cofactor required for infection with virus adapted for growth in transformed T-cell lines was recently identified and named fusin. However, fusin does not promote entry of macrophage-tropic viruses, which are believed to be the key pathogenic strains in vivo. The principal cofactor for entry mediated by the envelope glycoproteins of primary macrophage-tropic strains of HIV-1 is CC-CKR-5, a receptor for the beta-chemokines RANTES, MIP-1alpha and MIP-1beta.

3T3 Cells↗
PubMed1996

Lymphoblastoid transmembrane potential in bipolar patients, their siblings, and unrelated healthy comparison subjects.

Bipolar illness appears to be characterized by alterations in ionic homeostasis that are related to mood state. This is reflected by reports of altered intracellular sodium and calcium concentrations, altered sodium pump (Na+,K+-adenosine triphosphatase), and calcium pump (Ca2+-adenosine triphosphatase) activity. Recent ex vivo studies with fresh lymphocytes and in vitro studies with cultured lymphoblasts suggest that there may be an enduring trait-related abnormality in Na+,K+-adenosine triphosphatase as well. We have previously found that the lymphocyte transmembrane potential (TMP) varies with mood state. To examine the question of trait-related changes in TMP, we studied TMP of immortalized lymphoblasts from 14 patients with bipolar illness, nine unaffected siblings, and eight normal subjects. TMP was the same in all three groups. These preliminary data suggest that TMP is a state-- rather than a trait-related marker.

Adult↗
PubMed1996

The Duffy antigen receptor for chemokines: structural analysis and expression in the brain.

The Duffy antigen receptor for chemokines (DARC) is expressed in human erythrocytes and on endothelial cells lining postcapillary venules in kidney and spleen. DARC is a promiscuous chemokine receptor and a binding protein for the malarial parasite Plasmodium vivax. The expression of DARC by subsets of endothelial cells and neurons in discrete anatomic sites in the brain suggests that this enigmatic receptor may have multiple roles in normal and pathological physiology. Conservation of this promiscuous chemokine binding function is evident from the similarity in nucleotide sequence of DARC homologues from multiple species, as well as the high-affinity binding of human chemokines to murine and avian erythrocytes. Analysis of the functional domains of DARC using chimeric receptors and and monoclonal antibodies to multiple extracellular domains localized chemokine binding to structures in the amino terminal extracellular domain (E1). Scatchard analysis demonstrated that a chimeric DARC receptor, composed of the E1 domain of DARC and the predicted hydrophobic helices and loops of interleukin-8RB (IL-8RB), bound IL-8, and MGSA with KD values almost identical to the wild type receptors and bound a repertoire of C-X-C and C-C chemokines characteristic of DARC. Although numerous reports have demonstrated that chemokines such as IL-8 are expressed in the brain, presumably by glial cells, little insight into the nature of their role in normal or pathological physiology in the nervous system has developed because the target cells that express the corresponding receptors have not yet been identified. Northern blotting experiments suggest that mRNA encoding DARC are expressed in the central nervous system, however, interpretation of this is unclear because of the ubiquitous expression of DARC lining postcapillary venules. This study provides direct evidence to localize expression of DARC in the central nervous system. Immunohistochemical examination of human archival sections of the brain with monoclonal antibodies specific for DARC localize expression of DARC to cell bodies and processes of Purkinjie cells in the cerebellum. The immunohistochemical findings were supported by analysis of chemokine binding and radioligand crosslinking with membranes made from various brain fractions. The hierarchical expression of DARC in neurons in the cerebellum suggest that chemokines may play an important role in the modulation of neuronal activity by glial cells.

Animals↗
PubMed1996

Identification of the Fy6 epitope recognized by two monoclonal antibodies in the N-terminal extracellular portion of the Duffy antigen receptor for chemokines.

The epitope Fy6 recognized by two monoclonal antibodies (i3A and BG6), which inhibit binding of chemokines to the Duffy antigen, was characterized by means of peptides synthesized on pins (Epitope Scanning Kit) and deletion mutagenesis. Both antibodies showed very similar specificities. They recognized a linear epitope, the essential portion of which was the heptapeptide Gln-Leu-Asp-Phe-Glu-Asp-Val comprising amino acid residues 21-27, located between two glycosylation sites of the Duffy protein. All the amino acid residues of the epitope, except Glu, were essential for antibody binding, since they could not be replaced by any other amino acid residues or by only one or two. The Glu residue could be replaced by most other amino acid residues, and its replacement by 10 amino acid residues gave a distinct increase in the antibody binding. The results were in full agreement with the finding that the mutant of the Duffy antigen, lacking amino acid residues 23-25 (-Asp-Phe-Glu-), did not bind the i3A antibody, but bound the anti-Fy3 monoclonal antibody similarly to the wild type of the Duffy antigen. The apparent affinity constants of both anti-Fy6 antibodies were determined by surface plasmon resonance, using immunopurified Duffy protein as a ligand.

Amino Acid Sequence↗
PubMed1995

The promiscuous chemokine binding profile of the Duffy antigen/receptor for chemokines is primarily localized to sequences in the amino-terminal domain.

The Duffy antigen (DARC) is a promiscuous chemokine receptor that also binds Plasmodium vivax. DARC belongs to a family of heptahelical chemokine receptors that includes specific (IL-8RA) and shared (IL-8RB) IL-8 receptors. Ligand binding specificity of IL-8 receptors was localized to the amino-terminal extracellular (E1) domain. To determine the basis for promiscuous chemokine binding by DARC, a chimeric receptor composed of the E1 domain of DARC and hydrophobic helices and loops from IL-8RB (DARCe1/IL-8RB) was constructed. Scatchard analysis of stable transfectants demonstrated that the DARCe1/IL-8RB chimeric receptor bound IL-8 and melanoma growth stimulating activity (MGSA) with KD values almost identical to the native receptors. The hybrid receptor also bound RANTES, MCP-1, and MGSA-E6A (which binds DARC, but not IL-8RB), but not MIP-1 alpha, similarly to DARC. Ligand binding to DARC transfectants was unaltered by anti-Fy3, but inhibited by Fy6, which binds an epitope in the E1 domain. The epitope recognized by Fy3 was localized to the third extracellular loop by analysis of insect cells expressing chimeric receptors composed of complementary portions of DARC and IL-8RB. These findings implicate the E1 domain of DARC in multispecific chemokine binding.

Amino Acid Sequence↗
PubMed1995

Development and characterization of three recombinant single chain antibody fragments (scFvs) directed against the CD19 antigen.

Antibodies that recognize antigens restricted to leukemia, lymphoma, and normal hematopoietic cells represent a unique opportunity to develop therapeutics, because they have the potential for relatively selective treatment of these diseases. Antibodies that recognize the CD19 antigen found on normal and malignant B cells, but not on stem cells, have been used to develop immunoconjugates. However, these conjugates are large and might be suboptimal in tumor penetration when compared to molecules using smaller single chain Fv (scFv) antibody fragments. scFv has the advantage of being a molecularly engineered homogeneous molecule. In this report, we demonstrate the cloning, expression, and binding of three anti-CD19 antibodies as scFvs. All three scFvs were successfully cloned and expressed. FVS191, derived from cell line B43, and FVS192, derived from SJ25C1, were properly refolded and bound CD19 antigen in FACS competition assays. These anti-CD19 scFv should be useful in the further development of diagnostic and therapeutic molecules.

Antibody Specificity↗
PubMed1995

The Duffy antigen/receptor for chemokines (DARC) is expressed in endothelial cells of Duffy negative individuals who lack the erythrocyte receptor.

The Duffy antigen/receptor for chemokines (DARC), first identified on erythrocytes, functions not only as a promiscuous chemokine receptor but also as a receptor for the malarial parasite, Plasmodium vivax. The recent finding that DARC is ubiquitously expressed by endothelial cells lining postcapillary venules provides a possible insight into the function of this receptor because this anatomic site is an active interface for leukocyte trafficking. However, the biological significance of DARC is questionable since it has not yet been determined whether individuals lacking the expression of this protein on their erythrocytes (Duffy negative individuals), who are apparently immunologically normal, express the receptor on endothelial cells. However, we report here that DARC is indeed expressed in endothelial cells lining postcapillary venules and splenic sinusoids in individuals who lack the erythrocyte receptor. These findings are based on immunohistochemical, biochemical, and molecular biological analysis of tissues from Duffy negative individuals. We also present data showing that, in contrast to erythrocyte DARC, cells transfected with DARC internalize radiolabeled ligand. We conclude that the DARC may play a critical role in mediating the effects of proinflammatory chemokines on the interactions between leukocyte and endothelial cells since the molecular pathology of the Duffy negative genotype maintains expression on the latter cell type.

Amino Acid Sequence↗
PubMed1995

Murine monoclonal antibodies (MCS-1 and MCS-2) reactive with human myeloid leukemia cells.

In an attempt to identify the antigens expressed on human myeloid leukemia cells, two murine monoclonal antibodies (mAb) designated as MCS-1 (isotype; IgG3) and MCS-2 (IgG1) were raised. MCS-1 reacted with peripheral blood granulocytes, but not with monocytes, whereas MCS-2 reacted with both granulocytes and monocytes. The incidence of MCS-1 and MCS-2 reactivity with the cells from a total of 121 patients with various type of leukemias was as follows: 25/46 (54%) and 39/47 (83%) in acute myeloblastic leukemia and acute monoblastic leukemia, 8/16 and 16/16 in chronic myeloid leukemia in blastic crisis (CML-BC) of myeloid type, 0/7 and 2/7 in CML-BC of lymphoid type, 0/26 and 2/32 in acute lymphoblastic leukemia (ALL), 7/7 and 7/7 in chronic phase of CML, respectively. Two cases of MCS-2 positive ALL had a Philadelphia chromosome marker (Ph1). In contrast, neither MCS-1 nor MCS-2 reacted with lymphocytes or any of leukemic and non-leukemic lymphoid cell lines tested. These results indicate that MCS-1 and MCS-2 mAb recognized two different differentiation antigens expressed on granulocytes and monocytes. These mAb would be useful reagents to determine differentiation antigens on the cells in granulocyte and monocyte lineage.

Animals↗
PubMed1995

Distributions of melanoma growth stimulatory activity of growth-regulated gene and the interleukin-8 receptor B in human wound repair.

The alpha-chemokines have been implicated as regulators of proliferation and differentiation of normal keratinocytes and as mediators of keratinocyte maturation and migration in inflammatory processes that involve the skin. Using the cutaneous wound repair model, we examined the sites and temporal sequence of the appearance of melanoma growth stimulatory activity or growth-regulated gene (MGSA/GRO;ligand) and the type B interleukin (IL)-8 receptor (IL-8RB) to which MGSA/GRO binds. Human burn tissues (n = 44) representing days 2 to 12 after injury were obtained during surgical debridement, fixed in 4% paraformaldehyde, and embedded in paraffin. Immunolocalizations were performed with polyclonal antisera for both ligand and receptor, as well as a monoclonal antibody for the IL-8 RB. Western blot analysis confirmed the presence of the IL-8 RB in immunoprecipitates of epidermal keratinocyte lysates. In normal skin, MGSA/GRO protein was restricted to sites populated by differentiated keratinocytes (suprabasal compartments, inner root sheath cells, and dermal sweat ducts). MGSA/GRO protein was barely detectable within epithelial margins and islands of burn wounds where the migrating/proliferating keratinocyte populations reside, but staining intensities increased as cells matured into the outer layers. Weak diffuse staining was detected in areas of neutrophilic infiltration (granulation tissue and overlying exudates). By contrast, in normal skin the IL-8 RB was detected in specific locations within epidermal and dermal compartments of healing wounds. In the dermis, polyvalent antibodies detected receptor immunoreactivity most prominently in dermal sweat ducts and endothelium of capillaries, whereas this immunoreactivity was inconspicuous in sections stained with the monoclonal antibody. Receptor immunostaining was noted in migrating/proliferating keratinocytes in epithelial margins and islands but was in the outer layers or in hypertrophic epidermis adjacent to wounds. This same pattern was observed in epidermal appendages such as hair follicles and eccrine sweat ducts. In granulation tissues, IL-8 RB was noted in numerous fibroblasts and in subpopulations of macrophages and smooth muscle. The presence of both MGSA/GRO and its receptor in human burn wounds implicate this cytokine as an autocrine or paracrine mediator of epidermal regeneration in both the inflammatory and proliferative phases of cutaneous wound repair.

Animals↗
PubMed1994

Posttransplantation plasma cell dyscrasias.

BACKGROUND: Lymphoproliferative disorders that occur in patients receiving cyclosporine for immunosuppression after solid organ transplantation typically are B-cell neoplasms associated with Epstein-Barr virus (EBV), which may be polyclonal or monoclonal in origin. Although these tumors may have partial B-cell differentiation, (manifested as plasmacytoid features), terminal differentiation to plasma cells that secrete a monoclonal immunoglobulin is rare. The case of a patient who developed a posttransplantation lymphoproliferative disorder that was composed of multiple plasmacytomas located in the abdomen and urinary bladder after liver transplantation is presented. The patient also had high levels of an immunoglobulin-G kappa monoclonal paraprotein. METHODS: The plasmacytoma was examined for the presence of EBV by both polymerase chain reaction and in situ hybridization, and the possibility of a codon-12 mutation in the ras gene was investigated by digestion of DNA amplification products with the HpaII-restriction endonuclease. RESULTS: Epstein-Barr virus genomes were demonstrated by DNA amplification of sequences in the long, internal, direct repeat region, and in situ hybridization showed expression of EBV RNA transcripts that annealed to an EBER-1 probe. Immunohistochemistry showed clonally restricted expression of kappa light chains but failed to reveal evidence of expression of the latent membrane protein 1 encoded by EBV. Mutations of codon-12 in the H-ras gene were not detected. CONCLUSIONS: Resolution of the tumor and the paraprotein after radiation and reduction of immunosuppression indicates that terminal plasmacytic differentiation does not necessarily portend an unfavorable prognosis, even in a clonal lesion.

Abdominal Neoplasms↗
PubMed1994

Identification and characterization of a promiscuous chemokine-binding protein in a human erythroleukemic cell line.

The erythrocyte chemokine receptor is a cell surface protein that binds a wide array of chemokines including interleukin-8 (IL-8), melanoma growth stimulating activity (MGSA), monocyte chemotactic protein-1 (MCP-1), and RANTES (Regulated on Activation, Normal T Expressed and Secreted). This protein has also been identified as the Duffy blood group antigen, a cell surface receptor for the malarial parasite Plasmodium vivax. In the present study, we have identified a chemokine receptor-like binding protein in a human erythroleukemic cell line (HEL), which, based on its molecular properties, may be related to the erythrocyte chemokine receptor. Saturation binding studies with 125I-IL-8 revealed a single class of IL-8 binding sites in HEL cells with a KD of 7.4 +/- 1.9 nM and a receptor density of 12,818 +/- 965 binding sites/cell. In competition studies unlabeled IL-8 MGSA, MCP-1, and RANTES were fully able to inhibit the binding of 125I-IL-8 to HEL cells. Chemical cross-linking with radiolabeled IL-8 resulted in a cross-linked species of 60 kDa in membranes from HEL cells. The labeling was specific since it was inhibited by pre-incubation with 1 microM unlabeled IL-8 or MGSA. A monoclonal antibody (Fy6) to the human erythrocyte Duffy blood group antigen/chemokine receptor blocked the binding of IL-8 and other chemokines to the HEL cell chemokine receptor-like binding protein. Cell membranes from HEL cells and from erythrocyte ghosts were subjected to SDS-PAGE and analyzed by Western blotting with anti-Fy6. The antibody bound to a molecule with a molecular mass of 50 kDa in HEL cell membranes and 40 kDa in erythrocyte ghosts. Northern blot analysis of mRNA revealed that the HEL chemokine-binding protein hybridized to a cDNA probe to the Duffy antigen/chemokine receptor.

Binding Sites↗
PubMed1994

Cytometric detection of DNA amplified with fluorescent primers: applications to analysis of clonal bcl-2 and IgH gene rearrangements in malignant lymphomas.

Follicular lymphomas comprise almost two thirds of the US adult non-Hodgkin's lymphomas (NHL) and are the most common malignancy of B-lineage lymphocytes. Polymerase chain reaction (PCR) protocols have been developed to detect the t(14;18) translocation, which juxtaposes the bcl-2 proto-oncogene to the Ig heavy-chain (IgH) gene in 85% of follicular lymphomas and monoclonal rearrangements of the IgH gene in B-cell NHL that lack bcl-2 rearrangements. We used PCR to amplify bcl-2 and IgH rearrangements in DNA from patients with lymphoproliferative disorders and analyzed the products in parallel by gel electrophoresis and flow cytometry, which detected PCR products incorporating fluoresceinated oligonucleotide primers by sequence-specific capture to oligonucleotide-coated magnetic beads. Overall, flow cytometry was superior to electrophoresis of ethidium-bromide-stained agarose gels for detection of products of nested PCR to detect intergenic rearrangements involving bcl-2 and single primer-pair amplification of clonal rearrangement of IgH. Flow cytometric analysis detected bcl-2 translocations in 12 of 13 CD10+ B-cell lymphomas and clonal IgH rearrangements in 14 of 17 monoclonal B-cell populations. In contrast, analysis by gel electrophoresis detected bcl-2 translocations in only 10 of 13 CD10+ and clonal IgH gene rearrangements in only 9 of 17 monoclonal B-cell populations. Flow cytometric analysis was more sensitive than gel electrophoresis and could detect a 16-fold greater dilution of a bcl-2-amplified product than gel electrophoresis. Similarly, flow cytometry could detect an amplification product when template DNA was diluted 10,000-fold, whereas gel electrophoresis only detected amplification products when template was subjected to dilution between 100- and 1,000-fold. This shows the utility of flow cytometry for the analysis of DNA amplification products incorporating fluorochrome-labeled primers as a rapid, objective alternative to conventional strategies. Because current-generation clinical laboratories emphasize automation, flow cytometric analysis of PCR-amplified products shows increased analytic sensitivity and offers a vehicle for automation of DNA amplification tests.

Base Sequence↗
PubMed1994

Postcapillary venule endothelial cells in kidney express a multispecific chemokine receptor that is structurally and functionally identical to the erythroid isoform, which is the Duffy blood group antigen.

The human erythrocyte chemokine receptor has recently been shown to be identical to the Duffy blood group antigen and is expressed in multiple organs, including kidney. Here we have examined the molecular properties of the renal isoform. Immunoblot analysis of erythrocyte and kidney detergent lysates, with a monoclonal antibody (Fy6) to the Duffy antigen, revealed that the renal isoform had a molecular mass of 43-45 kD, which could be distinguished from that observed in erythroid cells (38-47 kD). Chemical cross-linking of kidney membranes to 125I-melanoma growth stimulatory activity (MGSA) indicated that the renal chemokine receptor had a molecular mass of 38-45 kD. Binding of 125I-labeled MGSA to kidney membranes was competitively inhibited by the addition of unlabeled MGSA, IL-8, regulated on activation, normal T expressed and secrted, and monocyte chemotactic protein-1. Scatchard analysis of MGSA binding showed that the chemokine receptor from renal tissues had a binding affinity of 3.5 nM similar to that observed for the erythroid isoform (5-10 nM). The primary structure of the renal chemokine receptor predicted from the nucleotide sequence of cDNA from renal tissues is identical to that reported for the erythroid isoform. Immunocytochemical staining of kidney with Fy6 localized expression to endothelial cells present in postcapillary venules. These studies implicate the Duffy antigen/chemokine receptor in the complex interactions between postcapillary endothelial cells and granulocytes, which are modulated by pro-inflammatory chemokines.

Antibodies, Monoclonal↗
PubMed1994

An investigation of the viral pathogenesis of Kikuchi-Fujimoto disease. Lack of evidence for Epstein-Barr virus or human herpesvirus type 6 as the causative agents.

Histiocytic necrotizing lymphadenitis of Kikuchi and Fujimoto is a well-defined clinicopathologic entity of unknown cause. Both the Epstein-Barr virus (EBV) and human herpesvirus type 6 (HHV-6) have been suggested as potential etiologic agents. Twenty cases of Kikuchi-Fujimoto disease were studied for the presence of EBV DNA and HHV-6 DNA by the polymerase chain reaction (PCR), and in situ hybridization in the case of EBV. Cellular DNA from sections of formalin-fixed, paraffin-embedded lymph node tissue was amplified using the PCR technique and oligonucleotide primers to the EBV BamH1 W, lymphocyte-determined membrane antigen, or the EBNA-1 region. These studies were performed in three separate laboratories. In addition, 12 cases were examined by in situ hybridization, eight of which had shown at least one positive PCR signal for EBV. The presence of HHV-6 was assessed by PCR using primers to part of the pZVH14 sequence. Biopsy specimens from eight patients (40%) showed a strong positive signal for EBV in at least one laboratory, while an additional three specimens (15%) showed a weaker positive signal. Five cases studied showed rare positive cells by in situ hybridization, and one case had scattered positive cells. All samples lacked HHV-6 genomic templates. These findings indicate that HHV-6 does not play a role in the pathogenesis of Kikuchi-Fujimoto disease and do not implicate EBV as a causal agent for Kikuchi-Fujimoto disease, since EBV was detected in only a fraction of cases with a low number of positive cells detected by in situ hybridization. Further, some discrepancies were identified in the positive results for EBV in samples studied by multiple laboratories. These results indicate that inconsistent results by PCR may occur with very low levels of viral genomes and that different laboratories perform DNA amplification at different efficiencies. Alternatively, laboratory contamination may give rise to false-positive results. Therefore, a positive result for EBV should be interpreted with caution and should be confirmed by repeated study (PCR) or by independent methodology (in situ hybridization).

Adolescent↗