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Biomedical subjects

L M Pinchuk

Publications and source records attributed to L M Pinchuk.

At least 19 recordsLinked to original sources
PubMed1999

Isolation and characterization of macaque dendritic cells from CD34(+) bone marrow progenitors.

We have developed a method for isolating and characterizing pigtailed macaque dendritic cells (DCs) generated from CD34(+) bone marrow (BM) progenitors based on methods previously developed for isolating human DCs. Macaque DCs displayed a characteristic morphology and were potent stimulators of allogeneic T cell proliferation. They expressed a set of DC-associated markers, such as MHC class II, CD1a, CD4, CD11a, CD40, CD58, CD80, CD83, CD86, and CXCR4. Macaque DCs, as well as peripheral blood CD4(+) T cells, were highly susceptible to HIV-2 infection, as detected by DNA-PCR. The expression of HIV-2 in macaque DCs was downregulated by treatment with the beta-chemokine RANTES. Macaque DCs will be useful for defining the in vivo role of DCs in HIV pathogenesis and for optimizing and testing peptide-DC vaccines or tolerizing regimens.

Animals↗
PubMed1997

Nuclear import of HIV-1 DNA in resting CD4+ T cells requires a cyclosporin A-sensitive pathway.

Using PCR to monitor early (LTR/LTR (long terminal repeat)) and late (LTR/gag) products of reverse transcription and the formation of HIV-1 LTR circles (indicating nuclear import), we explored the relationship between T cell activation signals and early events in the life cycle of HIV-1 infection. The combination of TCR ligation with either CD28 cross-linking or exogenous IL-2 was required for HIV-1 LTR circle formation in resting CD4+ T cells. Ligation of the TCR or CD28 receptors or addition of IL-2 alone did not induce this process. However, cross-linking the TCR, or IL-2 alone, unlike CD28 ligation, could induce the completion of viral reverse transcription. In contrast, the initiation of HIV-1 reverse transcription could occur in resting CD4+ T cells without any stimulation. Cyclosporin A (CsA), an inhibitor of T cell activation, completely blocked HIV-1 DNA nuclear import in activated CD4+ T cells. The completion of HIV-1 reverse transcription was blocked by CsA in infected CD4+ T cells activated by TCR ligation and IL-2, but not in cells stimulated by TCR and CD28 ligation. The costimulation of CD3 and CD28 mAbs in the presence of IL-2 could not overcome the CsA inhibitory effect on nuclear import of viral DNA. Therefore, the factor(s) involved in a CsA-sensitive pathway plays a critical role in HIV-1 DNA transport from the cytoplasm into the nucleus. Production and movement of HIV-1 DNA in resting CD4+ T cells require two signals: one signal from the TCR, which normally regulates the G0 to G1 transition, induces completion of viral reverse transcription; the other signal through CD28 or an IL-2R-dependent process is sensitive to CsA treatment and regulates viral DNA entry into the nucleus.

Antibodies, Monoclonal↗
PubMed1996

Functional CD40 ligand expressed by human blood dendritic cells is up-regulated by CD40 ligation.

We examined whether human blood dendritic cells (DCs) express a functional ligand for CD40 (CD40L). Human blood DCs expressed significant amounts of cell surface CD40L identical to that expressed on activated T cells, as detected by mAb to CD40L or a chimeric CD40.Ig fusion protein (CD40.Ig). Stimulation through CD40 up-regulated protein and mRNA CD40L expression in DCs, B cells, and B cell lines. CD40-mediated CD40L expression was inhibited by a protein tyrosine kinase inhibitor, herbimycin, in a dose-dependent manner, suggesting that the induction of CD40L expression via CD40 requires protein tyrosine kinase activity. CD40L surface expression correlated with constitutive or inducible levels of CD40L-specific mRNA, as determined by reverse transcribed PCR analysis (RT-PCR) using CD40L-homologous primers. Furthermore, CD40L on DCs was functional, since CD40L+ DCs, unlike CD40L- DCs, induced B cell IgG and IgA production, and this induction could be inhibited by blocking CD40L-CD40 interactions with mAb to CD40L. Thus, CD40L on DCs and CD40L induced by crosslinking CD40 may regulate B cell activation and maturation. The importance of DC CD40L expression on B cell function is discussed.

B-Lymphocytes↗
PubMed1996

Immunodeficiency virus cDNA synthesis in resting T lymphocytes is regulated by T cell activation signals and dendritic cells.

We explored the relationship between T cell activation signals and dendritic cells (DC) in the replication cycle of immunodeficiency viruses. First we analyzed the effect of two cell cycle inhibitors (mimosine and aphidicolin) on SIV reverse transcription, circularization, and integration in macaque resting T cells stimulated with anti-CD3 mAb at the time of infection. The formation of SIV LTR circles was blocked by the G1 inhibitor mimosine. The G1/S inhibitor aphidicolin neither affected circularization nor integration of SIV DNA. Therefore, the induction of SIV LTR circle production is likely to be mediated by signaling events normally regulating the G1 to S transition. We further characterized DC-dependent HIV-expression in human T cells. We examined the effect of ligating two novel receptors, IPO-3 and Bgp95, on DC-dependent HIV-1 expression. Activation of DCs through IPO-3 receptors, and to a lesser extent Bgp95 ligation, upregulated HIV spread in these cells. The mechanisms by which IPO-3 vs. Bgp95 increase HIV-1 levels appear to be different. In particular, IPO-3 ligation alone on T cells also increased HIV-1 levels. Activation of T cells via defined surface receptors or with DCs is required for establishing HIV/SIV cDNA synthesis in T cells.

Animals↗
PubMed1995

Cell-cell interactions regulate dendritic cell-dependent HIV-1 production in CD4+ T lymphocytes.

We investigated the role of blood dendritic cells (DC) in transmission of HIV-1 from infected to uninfected CD4+ T cells, and the accessory molecules involved. DC promoted transmission from infected to uninfected CD4+ cells, but blood DC themselves were not infectable. DC-mediated transmission was blocked by mAb to CD4 and MHC class II, but strongly increased by mAb to CD40 on DC or CD28 on T cells. The DC-dependent infection was inhibitable by anti-CD80 and a soluble fusion protein of the CD80 ligand, CTLA4; soluble CTLA4Ig also blocked infection augmented by crosslinking CD40. We also demonstrated that mAb to CD40 up-regulate the expression of CTLA4 ligands CD80 and B70/B7-2 (CD86) on DC. These data suggest that the dialog between CD40-CD40 ligand (CD40L) and CD28-CD80 counter-receptors on DC and T cells may be linked to HIV infection in vivo.

Abatacept↗
PubMed1994

Costimulation through CD28 enhances T cell-dependent B cell activation via CD40-CD40L interaction.

Changes in T cell helper function were analyzed when anti-CD3-activated T cells were costimulated with mAbs to the CD28 receptor (anti-CD28). T cell-dependent B cell growth and differentiation were consistently augmented if anti-CD3 stimulated-T cells were simultaneously activated with anti-CD28. Although anti-CD28 enhanced IL-2 and IL-4 production, it did not increase B cell responses solely by augmenting production of soluble lymphokines. Anti-CD28 costimulation induced increases on T cells of CD40 ligand (CD40L), known to promote B cell proliferation and Ig secretion. Because anti-CD28 promoted T cell helper functions and expression of CD40L, we examined the dependence for CD40L during T cell-dependent B cell responses. Although soluble CD40 fusion proteins only partially inhibited T cell-dependent B cell activation, we found a strict requirement for CD40L expression at initiating B cell responses. Both CD40L expression and T cell help were blocked by cyclosporin A after TCR cross-linking, and, unlike T cell proliferation, both remained cyclosporin A sensitive during CD28 costimulation. In addition, anti-CD28 could not compensate for the T cell helper deficiency of hyper IgM syndrome patients who lack functional CD40L. Thus, anti-CD28-induced T cell help is delivered via a CD40L-dependent process. The fact that cross-linking CD40 on B cells promotes expression of the B7/BB-1 ligand for CD28 suggest T and B interactions may have a reciprocal amplification mechanism.

Antigens, CD↗
PubMed1994

The role of CD40 and CD80 accessory cell molecules in dendritic cell-dependent HIV-1 infection.

We investigated the role of blood dendritic cells (DCs) in transmission of HIV-1 from infected to uninfected CD4+ T cells, and the accessory molecules involved. DCs promoted transmission from infected to uninfected CD4+ cells, but DCs themselves were not infectable. DC-mediated transmission was blocked by MAb to CD4 and MHC class II, but strongly increased by MAb to CD40 on DCs or CD28 on T cells. The DC-dependent infection was inhibitable by anti-CD80 and a soluble fusion protein of the CD80 ligand, CTLA4; soluble CTLA4 immunoglobulin also blocked infection augmented by cross-linking CD40. These data suggest a linkage between CD40-CD40L and CD28-CD80 counterreceptors on DCs and T cells, and spread of HIV infection in vivo.

Antibodies, Monoclonal↗
PubMed1994

[Identification of cultures of Mycobacteria cultivated using sodium salicylate medium].

Sodium salicylate test is used in bacteriological practice to distinguish M. tuberculosis and M. bovis from other mycobacterial species. The species of the strains growing in sodium salicylate media are not as a rule identified. This was done using gas chromatography with cultures which had grown in Lowenstein-Jensen media with sodium salicylate and isolated from the patients admitted to the Tuberculous hospital in 1991-1993. Only 20% of the strains belonged to human mycobacteria, the others were opportunistic or saprophyte mycobacteria (M. avium, M. fortuitum, M. phlei, M. flavescens, M. vaccae, M. smegmatis), some of the cultures were not of the Mycobacterium genus, but belonged to other Actinomycetales. It is evident that with sodium salicylate test one cannot be absolutely sure of Mycobacteria isolation. The strains growing in sodium salicylate medium call for further investigations.

Chromatography, Gas↗
PubMed1990

[Identification of Mycobacteria species by liquid-gas chromatography].

Principles and characteristic features of gas chromatographic indentification of Mycobacterium from the fatty acid composition were considered. The results of their differentiation by gas chromatography agree with Myco. indentification according to the complex of morphological, cultural and biochemical, and other characteristics. Copyrolysis has a number of advantages compared to the routine identification techniques, i.e. it reduces the time needed for the analysis down to 2-3 hours; allows one to use small portions of microbial mass; and provides for a complete automation of the procedure. The advisability of putting the gas chromatographic identification of Myco. species into practice of the antituberculosis medical and veterinary institutions is pointed out.

Carbohydrate Metabolism↗
PubMed1983

[Characteristics of the composition of higher fatty acids in methicillin-resistant and methicillin-sensitive Staphylococci].

The composition of fatty acids of methicillin-resistant (MR) and methicillin-sensitive (MS) strains of Staph. aureus and Staph. epidermidis was determined with the method of reactive gas liquid chromatography. The MS staphylococci of the above species differed by the content of acids with branched chains of iso- and anteisostructures and straight chains. Anteisoacids in the cells of Staph. epidermidis amounted almost to 80 per cent of the total number of the acids, while in the cells of Staph. aureus, their total number amounted only to a half of the fatty acid pool. Comparison of the composition of the fatty acids of the MS and MR strains of Staph. aureus revealed differences in the proportions of the anteiso- and isoacids. The total number of the long-chain C20.0 acid in the cells of Staph. epidermidis resistant to methicillin was lower as compared to that in the sensitive cells.

Chromatography, Gas↗
PubMed1982

[Higher fatty acid composition of enterococci].

The content of unsaturated fatty acids in enterococcal cells has been found to have no essential relation to the composition of the culture medium. When cultivated in the same media, S. faecium had the degree of lipid unsaturation 1.5-2 times higher than S. faecalis. Mobile enterococci are sharply differentiated from immobile species by the content of cyclopropanic acid with 19 carbon atoms, constitute a heterogenous group and consist of at least 2 taxons, differing in the content of acids with 18 carbon atoms and the degree of lipid unsaturation.

Chromatography, Gas↗