Biomedical subjects

G Aboagye-Mathiesen

Publications and source records attributed to G Aboagye-Mathiesen.

31 records · Page 2Linked to original sources

PubMed1993

Differential replication of human immunodeficiency virus type 1 in CD8- and CD8+ subsets of natural killer cells: relationship to cytokine production pattern.

CD8+ and CD8- subsets of peripheral blood natural killer (NK) cells were examined for susceptibility to infection with human immunodeficiency virus type 1 (HIV-1) and for the ability to produce various types of interferon (IFN) and tumor necrosis factor (TNF). HIV-1 was preferentially grown in CD8+ NK cells. The ability of CD8- NK cells to suppress HIV-1 replication was related to their ability to produce alpha IFN (IFN-alpha) upon viral induction. Induction with interleukin-2 resulted in IFN-gamma production in both subsets of NK cells. In the CD8+ subset, IFN-gamma and HIV-1 mutually enhanced the production of TNF alpha, leading to hyperactivation of viral replication, whereas in CD8- NK cells IFN-gamma primed HIV-induced IFN-alpha production. The dichotomous effects of IFN-gamma on HIV-1 replication were dependent on the IFN-alpha-producing ability of the cellular targets. These findings can explain the selective depletion of the CD16+ CD8+ subset that begins early in the in vivo HIV-1 infection.

Antigens, CD↗

PubMed1992

Isolation, purification and biochemical characterization of human placental interferons by tandem high-performance affinity chromatography.

Human placental trophoblasts, fibroblasts and the trophoblast-derived malignant cell JAR are potent producers of interferons (IFNs) when stimulated with Sendai virus. The three cell lines produced different levels and compositions of IFN-alpha subtypes and IFN-beta. Anti-IFN globulins, Cibacron Blue F3GA and Concanavalin A were covalently immobilized on pressure-stable, macroporous polymeric matrices derivatized with vinyl sulphone (HEMA-BIO 1000 VS and HEMA 1000 VS). These supports were packed in biocompatible PEEK columns and were coupled with switching valves, to develop a tandem high-performance affinity chromatographic (HPAC) method for the isolation, purification and biochemical characterization of the IFNs produced in Sendai virus-stimulated human placental trophoblasts, fibroblasts and trophoblast-derived malignant cell, JAR, cultures. Silver-stained SDS-PAGE and gel densitometric analysis revealed the purity of the purified proteins to be between 94 and 98%. Specific activities of the purified IFNs ranged between 0.37-2.76 x 10(8) IU/mg of protein with cumulative recoveries between 90 and 92.2%. The purified IFN components exhibited quantitatively different antiviral activities in human and bovine cell lines. The utility of the tandem method for the purification and characterization of human type 1 IFNs produced from other cell lines are also discussed.

Chromatography, Affinity↗

PubMed1991

Enhanced chemiluminescence-based hybridization analysis for PCR-mediated HIV-1 DNA detection offers an alternative to 32P-labelled probes.

The efficiency of enhanced chemiluminescence based on a novel generation substrate for alkaline phosphatase, adamantyl-1,2-dioxetane phosphate, was compared with that of 32P-labelled probe for visualization of human immunodeficiency virus type 1 (HTV-1)-specific DNA-DNA hybrids. The probe used for nonisotopic detection was digoxigenin labelled and targeted by anti-digoxigenin antibody Fab-fragments conjugated to alkaline phosphatase. The dot-blot hybridization analysis performed on a dilution series of HIV-1 proviral DNA demonstrated a lower sensitivity limit of 0.5 pg with the nonisotopic method. However, one order of magnitude less DNA could still be detected by a random-primed 32P-labelled probe. The ability of nonradioactive and radioactive probes to detect 590-bp gag gene-specific target sequences generated by the polymerase chain reaction (PCR)-mediated amplification of HIV-1 DNA was also compared. Analysis of 20 samples from individuals at increased risk for HIV infection by using the two assayed systems produced virtually equivalent signal images on corresponding specimens. Furthermore, complete concordance in the performance was found when HIV-1 proviral DNA was investigated by PCR in additional 50 samples of human blood mononuclear cells.

Adamantane↗

PubMed1991

Purification of human placental trophoblast interferon by two-dimensional high performance liquid chromatography.

Human placental trophoblast challenged with Sendai virus induced IFNs mainly of the beta-type (75%) and relatively low levels of the alpha-type (25%). A two-step high performance liquid chromatographic procedure ("two-dimensional HPLC") has been developed for the complete purification of the placental trophoblast interferon beta (tro-IFN-beta) from serum-containing culture supernatant. The method involved a combination of high performance liquid affinity chromatography (HPLAC) on Cibacron Blue 3GA immobilized on an activated pressure stable macroporous synthetic polymer, 2-hydroxyethyl methacrylate vinyl sulphone (HEMA-BIO 1000 VS), as the first dimension and reversed-phase high performance liquid chromatography (RP-HPLC) on Separon SGX C-18 as the second. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot experiments showed that the tro-IFN-beta was present as a 24 kDa protein. Densitometric scanning analysis of Coomassie-stained gel revealed the purity of the final preparation to be greater than 99%. The purified tro-IFN-beta had a specific activity of 1.03 x 10(8) IU/mg of protein and the overall recovery was 81% of the total IFN-beta activity in the crude preparation and 61% of the total IFN activity.

Cells, Cultured↗

PubMed1991

Characterization of Sendai virus-induced human placental trophoblast interferons.

Human placental trophoblast cultures produce a mixture of interferons (IFNs) when challenged with Sendai virus. High-performance dye-ligand and immunoaffinity chromatography of a trophoblast IFN (tro-IFN) preparation enabled the isolation of three antigenically distinct IFNs, alpha I, alpha II 1 and beta, with Mrs of 16K, 22K and 24K respectively, by reducing and non-reducing SDS-PAGE. The major IFN, responsible for 75% of the total antiviral activity, was tro-IFN-beta, with the remaining activity being due to tro-IFN-alpha I and tro-IFN-alpha II 1, as determined by an antiviral neutralization test using specific anti-human IFN antibodies. The antiviral activities of the tro-IFNs were stable at pH 2.0 for 24 h and tro-IFN-alpha II 1 and -beta were shown to be glycoproteins. The three tro-IFNs showed different antiviral activities when assayed on human and bovine cell species; tro-IFN-alpha I and alpha II 1 protected both human and bovine (MDBK) cells from virus infection, whereas tro-IFN-beta showed a high degree of species specificity, protecting only the human cell types tested.

Animals↗

PubMed1990

Purification and initial characterization of human placental trophoblast interferon induced by polyriboinosinic.polyribocytidylic acid.

Human placental trophoblast interferon (tro-IFN), induced in trophoblast cultures by a superinduction procedure, was purified to a homogeneous product with retention of biological activity. The problems associated with isolation from serum-containing medium were overcome by a combination of Blue Sepharose affinity chromatography and reversed-phase HPLC (RP-HPLC) on Separon SGX C-18. This two-step purification procedure yielded tro-IFN with a specific activity of 3.4 x 10(7) international units/mg of protein. The overall recovery of interferon activity was 66.7%. The purified tro-IFN was shown to be a glycoprotein with an Mr of 24K on native and SDS-PAGE. Its antiviral activity was stable at pH 2.0 at 37 degrees C but was sensitive to heat at 56 degrees C for 1 h and was neutralized by antibodies to human IFN-beta.

Cell Line↗

PubMed1990

Interferon production by cultured human trophoblasts and choriocarcinoma cell lines induced by Sendai virus.

Human term-placental trophoblasts in primary culture were studied for an interferon (IFN) response when challenged with Sendai virus and compared to three choriocarcinoma cell lines, placental fibroblasts and placental macrophages. Normal trophoblasts were high producers and released both IFN-alpha and IFN-beta. In contrast, one choriocarcinoma cell line was a low producer and all malignant lines produced only IFN-beta. Circulating monocytes produce IFN-alpha but placental macrophages secreted IFN-beta and some IFN-alpha, suggesting that IFN production may be dependent on the stage of differentiation. A role for trophoblast IFNs in protection of the foetus against virus infections is proposed.

Antibodies↗

PubMedDate not supplied

Physiological oxygen tension is relevant to MHC-1 expression, spontaneous transformation, and interferon response of in vitro aging murine fibroblasts.

Working from the hypothesis that modest deviations from physiological oxygen tension will influence cell characteristics important for infections/immunity and tumor development, cells were studied at three oxygen tensions during in vitro aging. Primary mouse embryo fibroblasts were established and subsequently passaged at 3, 6, and 18 kPa oxygen tension (6 representing normal tissue tension and 18 being the conventionally tension at in vitro cultures). The growth rate was slightly higher at 6 than 3 and 18 kPa. All cultures eventually stopped growing and subsequently transformed to nonmalignant cells with unlimited growth capacity. Cells kept at 3 kPa reached the highest number of cell doublings before crisis. Stimulation with PolyI:C resulted in detectable interferon response only at the high oxygen tension, and after transformation none of the cultures responded with interferon production. Expression of the major histocompatibility complex H-2K was elevated above and below physiological oxygen tension, indicating regulatory processes optimizing MHC expression at about physiological oxygen tension.

Animals↗

PubMedDate not supplied

Differential HIV replication and HIV-induced interferon production in mononuclear phagocytes: relationship to cell maturation.

We have investigated the replication of human immunodeficiency virus (HIV) and HIV-induced interferon (IFN) production in human mononuclear phagocytes at 2 different stages of in vitro maturation. Blood monocytes and monocyte-derived macrophages from 6 healthy, HIV-seronegative donors were challenged with HIV1IIIB and HIV2ROD. Freshly separated monocytes produced IFN when inoculated with both HIV types. In these cultures, an inverse correlation was observed between the amount of IFN production and the rate of HIV replication. In contrast to the monocytes, 5-day-old monocyte-derived macrophages did not produce IFN when challenged with HIV, but a significant replication of HIV1IIIB and HIV2ROD was found in all cultures.

Acquired Immunodeficiency Syndrome↗

PubMedDate not supplied

Differential interferon production in human first and third trimester trophoblast cultures stimulated with viruses.

Stimulation of human placental first and third trimester trophoblast and syncytiotrophoblast cultures with viruses [Newcastle Disease Virus (NDV) and Sendai virus] led to a high interferon (IFN) production. The magnitude of the production was dependent on the gestational age of the trophoblast, type of inducer and the stage of differentiation of the trophoblast. The data obtained indicated that the first trimester trophoblast cultures produced five to sixfold more IFN than the third trimester trophoblast on per cell basis whereas syncytiotrophoblast at term produced twice as much IFN than the mononuclear term trophoblast when stimulated with the viruses. NDV and Sendai virus produced different levels and composition of IFN-alpha and -beta in both first and third trimester trophoblast and syncytiotrophoblast cultures. Purification of the virus-induced trophoblast interferons (tro-IFNs) by tandem high-performance affinity chromatography resulted in specific activities between 0.7 and 2.7 x 10(8) IU/mg of protein when assayed on human amniotic WISH cells. The tro-IFN-alpha protected both human and bovine MDBK cells from virus infection whereas the tro-IFN-beta protected only the human cell lines tested. The possible roles of the tro-IFNs are discussed in light of the observed differences in trophoblast IFN response.

Cells, Cultured↗

PubMedDate not supplied

Effects of human trophoblast-induced interferons on the expression of proto-oncogenes c-fms/CSF-1R, EGF-R and c-erbB2 in invasive and non-invasive trophoblast.

The human cytotrophoblast is the first fetal cell type to arise during embryogenesis and differentiate along two pathways to the invasive (extravillous) and non-invasive (villous) populations. The non-invasive villous trophoblast differentiate morphologically and biochemically to form terminally differentiated multinucleated syncytial trophoblast. First trimester invasive and non-invasive trophoblast were isolated from human placentae (5-12 weeks) and were cultured in vitro. The villous trophoblast cells differentiated in vitro to form aggregated syncytial cells which was associated with increased expression of epidermal growth factor receptor (EGF-R). The invasive trophoblast cells expressed colony-stimulating factor receptor (c-fms/CSF-1R) and c-erbB2 proteins but low levels of EGF-R. We studied the effects of human trophoblast-induced interferon (IFN)-alpha/beta on the expression of c-fms/CSF-1R, EGF-R and c-erbB2 whose ligands are reported to be involved in the regulation of growth and differentiation of normal invasive and non-invasive trophoblast cells. Human trophoblast-induced IFN-alpha/beta (100 IU/ml) reduced the expression of EGF-R in both invasive and non-invasive trophoblast cells as determined by quantitative enzyme-linked immunosorbant assay ('ELISA') and western immunoblot methods. The same amount of IFN activity reduced the expression of c-fms/CSF-1R and c-erbB2 proto-oncogene products in invasive trophoblast cells. These results may suggest a possible role of trophoblast-induced IFNs in the regulation of normal trophoblast growth, differentiation and function.

Blotting, Western↗

PubMedDate not supplied

In vitro infection of human placental trophoblast by wild-type vaccinia virus and recombinant virus expressing HIV envelope glycoprotein.

Short-time (< or = 7 days) cultures of trophoblast mononuclear cells isolated from term placentae were challenged with vaccinia virus. Cytopathic effects were induced in crude placental cell preparations as well as in cultures established after negative immunosorting of major histocompatibility complex class I epitope-expressing cells, i.e. cultures exclusively derived from villous cytotrophoblast according to our present state of knowledge. The trophoblast in vitro supported a full replicative cycle of both wild-type viruses and a recombinant clone serving as a vector for the human immunodeficiency virus type 1 envelope gene. Results may shed light on mechanisms involved in the rarely observed foetal damage caused by smallpox vaccination during pregnancy.

Animals↗

PubMedDate not supplied

Susceptibility of MHC class I expressing extravillous trophoblast cell lines to killing by natural killer cells.

Purified human first trimester extravillous trophoblast (EVT) cell lines HTR-8 and HT-116 were examined for susceptibility to natural killer (NK) cell-mediated lysis. Based upon nucleic acid sequencing of an amplified fragment of cDNA, Western blot analysis and immunostaining of fixed and live cells, it was shown that both EVT cell lines expressed HLA-G mRNA and protein within the cytoplasm when cultured on laminin-coated plates. Very weak HLA-G expression was detectable on the cell surface under these conditions. However, strong cell surface expression of a classical MHC class I molecule (most likely HLA-C) was exhibited by these EVT cell lines when grown on laminin, as indicated by W6/32 FACS analysis (Ab recognizing pan MHC class I), and Western immunoblotting with HC10 (Ab recognizing HLA-B/C). When these EVT cells, cultured on laminin, were used as targets for peripheral blood natural killer (NK) cells in a standard chromium release assay, both HTR-8 and HT-116 cells were lysed by NK cells in a dose-dependent manner. The respective percentage specific lysis at an effector to target (E/T) ratio of 100 was 28+/-7, and 48+/-14. The choriocarcinoma cell lines JAR and JEG-3 which were respectively MHC class I negative and HLA-G positive were resistant to NK cell lysis. Thus, there was no clear relationship between the MHC class I expression and NK cell resistance or susceptibility among the EVT cell lines and choriocarcinoma cells. These findings raise the possibility that NK cells may take part in the surveillance of the invasive EVT cells during normal placentation.

Cell Line↗