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Biomedical subjects

C Larcher

Publications and source records attributed to C Larcher.

At least 55 records · Page 3Linked to original sources
PubMed1990

Characterization of monoclonal antibodies to human immunodeficiency virus type 1 gp41 by HIV-1 polypeptides expressed in Escherichia coli.

Two monoclonal antibodies (MAbs) were produced in Balb/c mice by immunization with recombinant gp41 derived from expression of lambda-BH10 cDNA of the human immunodeficiency virus-1 (HIV-1) in the prokaryotic expression vector pEX-41. Characterization of the epitopes recognized by these MAbs was done with HIV-1 envelope (env) fusion proteins expressed in Escherichia coli encoding ten distinct segments of the env proteins. In comparison, another mouse MAb, M25, a human MAb directed against gp41, which was produced by the xeno hybridoma line 3D6 and a pool of human patient sera containing antibodies to HIV-1 were tested. We were able to demonstrate that the epitopes recognized by our MAbs are located between arg732 and ser759 of the HIV-1 env glycoprotein gp160 of HTLV-III strain B. M25 reacted with epitopes between ser647 and pro731, which includes the hydrophobic transmembrane region of gp41. The human MAb against gp41, 3D6 reacts with epitopes between ile474 and trp646, a polypeptide stretch consisting of gp120 and gp41 specific amino acids. The human serum pool, positive for HIV-1 antibodies, reacted predominantly with antigenic determinants located between ile474 and leu863. The recombinant env fusion proteins were initially produced to test the immunoreactivity with patient sera and to characterize epitopes which are relevant for immunodiagnostic purposes. In this study, we showed that the set of recombinant env proteins is also a simple and accurate tool for the characterization of MAbs directed to the HIV envelope proteins.

Animals↗
PubMed1990

Expression of the C3d/EBV receptor and of other cell membrane surface markers is altered upon HIV-1 infection of myeloid, T, and B cells.

Human cell lines (the T-cell lines H9, Jurkat, and HUT102, the myeloid lines U937 and HL60, and the Raji B cell line) were infected with HIV-1. HIV-1 antigen could be detected by immunofluorescence analysis in more than 50% of T cells and myeloid cells 15 days after infection. Infection of Raji cells took more than 2-3 months. Studies of cell surface marker expression revealed remarkable changes after HIV-1 infection of Raji cells: expression of CR2 (C3d/EBV receptor, CD19, CD20, CD22, CD23, CD10, and surface IgM) were highly reduced, in the case of CR2 and membrane-IgM from 100 to 0%, whereas levels of CD37 and CD38 remained unaltered by HIV-1 infection. U937 cells showed a reduction of CD4 expression from 14 to 5% after HIV-1 infection; the CR3 expression slightly increased from 25 to 30%. In contrast, HLA-DR was only expressed (21%) after HIV-1 infection but not in uninfected U937 cells. Expression of HLA-DR could be detected also in HL60 cells (33%) after HIV-1 infection. In H9 cells, CD4 was reduced from 60 to 30% after HIV-1 infection, whereas HLA-DR and CD25/IL-2 receptor expression increased from 16 to 90% and from 0 to 50%, respectively. CD4 was reduced from 70 to 0% from Jurkat cells after HIV-1 infection, whereas expression of CR2 was only slightly diminished from 8 to 4%. Expression of CR1 and HLA-DR was slightly increased in these cells (1 to 3%).(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Differentiation, B-Lymphocyte↗
PubMed1990

[Human immunodeficiency virus (HIV): a review].

Since its discovery in 1983, a lot of knowledge about the human immunodeficiency virus (HIV) has been accumulated. Our paper gives a brief survey on what is known at present about the structure, molecular biology, and the cell tropism of this virus. We discuss its relationship to other lentiviruses as well as its possible origin; in addition, we refer to the immune response to HIV and its interactions with the infected host. We also briefly summarize the difficulties encountered in the attempts to produce a vaccine against HIV and highlight some promising approaches in the development of such a vaccine.

DNA Replication↗
PubMed1989

[Human herpesvirus 6 (HHV-6): incidence in healthy blood donors and in patients with acute infections caused by other herpes viruses].

Antibodies to HHV-6 were detected by immunofluorescence in 8.04% of 460 healthy blood donors in West Austria. Testing sera from patients with acute or reactivated infections with other herpesviruses, such as cytomegalovirus (CMV), Epstein-Barr virus (EBV), herpes simplex virus (HSV) and varicella zoster virus (VZV) we observed a remarkably higher prevalence of antibodies to HHV-6 in patients with CMV infections (75%) and also in patients with EBV infections (50%). Patients with HSV and VZV infections were positive for HHV 6 in 12.5% and 6.6% of cases, respectively. Many of the patients with CMV infection were transplant recipients. The high incidence of positive HH 6 serology in these patients could be due to new infection by HHV 6 or to the reactivation of a previous infection with HHV 6 by means of allogenic cell stimulation. Furthermore, preliminary results from our laboratory point to a serological cross-reaction between HHV 6 and CMV, which may also contribute to this result.

Antibodies, Viral↗
PubMed1989

Human complement factor H: an additional gene product of 43 kDa isolated from human plasma shows cofactor activity for the cleavage of the third component of complement.

In addition to the 150-kDa factor H protein, we have previously described a 43-kDa factor H molecule in human plasma, which probably represents a translational product of the additional 1.8-kb mRNA for factor H. This factor H was isolated from human plasma by means of immunoaffinity chromatography and high-performance liquid chromatography. When tested for its functional activity, this purified 43-kDa H protein was shown to act as cofactor for factor I- mediated cleavage of fluid-phase C3b to iC3b.

Antibodies, Monoclonal↗
PubMed1989

HIV and HIV-infected cells differentially activate the human complement system independent of antibody.

The human retroviruses HTLV-I and HIV-I have previously been shown not to be lysed by human serum. An interaction between HIV and the complement system, however, has not been investigated in any detail. In this report we show that purified HIV as well as HIV-infected cells activate the complement system. In the case of virus-infected cells this activation is mediated by the alternative pathway of complement, whereas the classical pathway seems to be in operation for the triggering of the complement system by purified virus and recombinant envelope glycoprotein (gp 160). We demonstrate that this leads to the deposition of C3b and/or C3bi on the surface of infected cells. But the HIV-infected cells are not lysed by human complement. C3 fragments deposited on the surface of HIV-infected cells are capable of mediating immune adherence to complement receptor-bearing cells, such as human erythrocytes and phagocytes. Whether this might have an influence on infectivity of HIV for certain cells bearing complement receptors has yet to be shown.

Cell Line↗
PubMed1989

Recombinant peptides derived from the env-gene of HIV-2 in the serodiagnosis of HIV-2 infections.

We produced recombinant envelope-derived peptides of HIV-2 for use in a diagnostic enzyme-linked immunosorbent assay (ELISA) or Western blot by expressing several restriction enzyme fragments from the env gene of HIV-2 in the bacterial fusion vector pEX-3. On Western blots, 17 out of 18 anti-HIV-2-positive sera available to us reacted strongly with those recombinant peptides which were derived from, or extended into, the transmembrane protein of HIV-2. In contrast, recombinant peptides derived from the external envelope glycoprotein were only weakly recognized by two sera. We observed a cross-reactivity of some human sera containing antibodies to HIV-1 with the HIV-2 peptides derived from the transmembrane protein of HIV-2. In spite of this cross-reactivity, a serological distinction between anti-sera to HIV-1 and HIV-2 can be attempted by simultaneous testing in ELISA on recombinant peptides derived from the transmembrane protein of HIV-1 and HIV-2.

AIDS Serodiagnosis↗
PubMed1989

The central segment of herpes simplex virus type 1 glycoprotein C (gC) is not involved in C3b binding: demonstration by using monoclonal antibodies and recombinant gC expressed in Escherichia coli.

Three monoclonal antibodies (MAbs) have been raised against cell membrane-derived herpes simplex virus type 1 glycoprotein C (gC-1). By using different DNA constructs of gC-1 expressed in Escherichia coli the sites recognized by these antibodies could be assigned to a peptide in the more hydrophobic and probably non-glycosylated middle third of the gC-1 molecule. This peptide segment corresponds to a 571 bp segment on the gC-1 gene located between the NcoI and the NruI restriction sites. None of the three MAbs interfered with the binding of the human serum complement component C3b to gC, which has been shown to be rather glycosylation-dependent. Since the data of other groups have suggested that antibodies directed against all regions of gC-1 inhibit C3b binding to gC-1, whereas our results suggest that the central part of gC-1 is not actually involved in C3b-binding activity, it can be inferred that the N- and C-terminal segments are involved in C3b binding by gC-1.

Animals↗
PubMed1989

C3bi-binding protein on Candida albicans: temperature-dependent expression and relationship to human complement receptor type 3.

We investigated in detail the previously described capacity of pseudohyphae of Candida albicans to bind C3-coated particles. We show that the expression of the C3bi receptor of C. albicans was dependent upon the growth temperature of the fungi. C. albicans grown at 30 degrees C bound strongly to EAC1423bi, whereas those cells grown at 38.5 degrees C were completely devoid of this capacity. The molecule responsible for the attachment of EAC1423bi was heat labile and trypsin sensitive. Several, but not all, monoclonal antibodies to the alpha-chain of human complement receptor type 3 (CR3) stained C. albicans, and this reactivity was expressed in parallel with the capacity of C. albicans to bind EAC1423bi, i.e., both were dependent on the growth temperature of the fungi and were trypsin sensitive. In contrast to CR3, the binding of EAC1423bi to C. albicans did not require the presence of divalent cations. Rabbit immunoglobulin G antibodies directed against C. albicans inhibited the binding of EAC1423bi to C. albicans but not to human CR3. These inhibiting IgG antibodies recognized antigens expressed on the surface of pseudohyphae but not those of yeast cells. OKM-1, a monoclonal antibody to human CR3 inhibited the attachment of EAC1423bi to CR3 and also to C. albicans. OKM-1 precipitated a 130-kilodalton band from solubilized 125I-labeled C. albicans. We conclude that the complement receptors on C. albicans and human CR3 were antigenically related but not identical and that they differed in their functional characteristics.

Animals↗
PubMed1989

Effect of D-penicillamine on the expression and propagation of the human immunodeficiency virus by H9 T-lymphoblastoid cells.

The effect of D-penicillamine on the replication of the Human Immunodeficiency Virus (HIV) in H9 T-lymphoblastoid cells was evaluated. HIV-infected and uninfected H9 cells were incubated for a period of 6 weeks with various concentrations of D-penicillamine. During the first 8 days, D-penicillamine (at concentrations of 0-150 micrograms/ml) did not influence growth and viability of H9/H9-IIIB cells. However, long term cultures and higher doses of the drug caused a decay in cell number and viability of the cultured cells. Up to the 8th day of culture, D-penicillamine did not exert an inhibiting effect on the number of HIV-infected cells but did suppress HIV-antigen production by up to 29%. After 6 weeks, in addition to a decrease of the relative viral antigen concentration from 390-500 ng/ml in the medium control to 210-320 ng/ml in cultures containing 150 micrograms D-penicillamine/ml, D-penicillamine also reduced the percentage of HIV-positive cells. Thus our results demonstrate an antiviral effect of D-penicillamine in H9 cultures.

Cells, Cultured↗
PubMed1988

[Etiology--characteristics of HIV infection].

In this study the structures of the AIDS-virus are presented, furthermore the cross-reactivity of HIV-1 and HIV-2 is described as well as the known genes of both viruses. The mechanism of cell infection, its kinetics and the special role of activated T-cells and macrophages in this process are outlined. This interrelation characterized phenomenologically based on the neopterin data available is now supported by experiments at a molecular level. This should lead to therapeutical and diagnostical consequences.

Acquired Immunodeficiency Syndrome↗
PubMed1988

Selective induction of mononuclear phagocytes to produce neopterin by interferons.

Interferon-gamma (IFN-gamma) has been shown to be a potent inducer of neopterin secretion by human peripheral blood monocytes/macrophages (1). In this paper, it is shown that other known stimuli of monocytes (e.g., to secrete proteases or to migrate) such as zymosan-activated human serum, lipopolysaccharide, human C3/iC3 and zymosan coated with complement were unable to trigger monocytes/macrophages to release neopterin. Monocytes/macrophages could be stimulated solely by IFN-gamma (25 U/ml) and IFN-alpha at very high concentrations (10,000 U/ml). In the case of human peripheral blood mononuclear cells (PBMNC), basically the same pattern was observed. If however, in the buffer controls PBMNC showed some neopterin release, all stimuli triggered an increase of neopterin secretion: 10,000 U/ml IFN-alpha induced the same amount of secreted neopterin as did 25 U/ml of IFN-gamma. Both caused higher levels of neopterin secretion than ZAS, LPS and C3/iC3. Amongst the supernatants from PBMNC, only those which were obtained from cells activated with IFN-gamma or -alpha stimulated monocytes/macrophages to produce neopterin. Supernatants from lymphocytes activated with zymosan, lipopolysaccharide and interferon did not contain neopterin, nor did the latter induce monocytes/macrophages to generate and secrete neopterin. Antibodies against IFN-gamma inhibited the triggering effect of the supernatants except when generated by IFN-alpha at 10,000 U/ml. These results demonstrate that both interferons, IFN-gamma and IFN-alpha, the latter only at a 400-fold higher concentration, can trigger monocytes/macrophages directly to secrete neopterin. ZAS, LPS and C3/iC3 are weakly effective only on a mixture of lymphocytes and monocytes/macrophages, provided this cell mixture shows already a basic spontaneous neopterin release.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1988

Comparison of Western blot (immunoblot) based on recombinant-derived p41 with conventional tests for serodiagnosis of human immunodeficiency virus infections.

To evaluate the performance of a serological test for human immunodeficiency virus type 1 (HIV-1) infections based on the use of a recombinant envelope gene-derived protein as the antigen, we caused expression of a 1.4-kilobase fragment of HIV.DNA that codes for the complete gp41 transmembrane protein in an Escherichia coli expression vector and used Western blots (WB; immunoblots) prepared with recombinant material (pEX-41) to detect antibodies to HIV-1. This test detected all 339 sera which were positive by a combination of conventional serodiagnostic assays and produced no false-positive results with 311 negative samples. Also no false-positive results were obtained with 20 sera from systemic lupus erythematosus patients which had high titers of cross-reactive autoantibodies. In six cases, the pEX-41 WB proved to be more sensitive than individual assays applied on their own, and in five cases it was even more sensitive than a combination of conventional assays. We tested 221 sera in both our pEX-41 WB and a commercially available recombinant enzyme immunoassay (EIA [Abbott]). The results were identical in 188 cases. A total of 27 sera containing antibodies to gp41 as demonstrated in the pEX-41 WB, as well as the Abbott recombinant EIA, had no antibodies to the recombinant core antigen as measured in the Abbott EIA. However, 25 of these sera did stain the 24-kilodalton band on a WB with purified virus. Six sera that were positive in all of the conventional confirmatory assays and reacted strongly with the pEX-41 WB did not recognize the surface antigen used in the Abbott recombinant EIA. We conclude that the use of WB prepared with recombinant-derived p41 offers a very sensitive and specific method to detect antibodies to HIV.

Acquired Immunodeficiency Syndrome↗
PubMed1987

[Detection of antibodies to HTLV-III: a comparison of various ELISA methods as screening tests and the Western Blot with immunofluorescence as confirmatory procedures].

386 sera were examined with three commercially available ELISAs for antibodies to HTLV-III. Western Blot and an indirect immunofluorescence assay were performed on sera showing a positive reaction in one or more ELISAs as confirmatory assay. 299 sera reacted negatively in all ELISAs, 48 were positive in all ELISAs and the confirmatory assays, whilst 33 ELISA positive sera reacted negatively in the confirmatory assays. In the case of 5 sera both Western Blot as well as the immunofluorescence assay had to be undertaken to obtain conclusive results.

Acquired Immunodeficiency Syndrome↗