Primary Human Peripheral Blood Lymphocytes Research Articles

Ciprofloxacin induces an immunomodulatory stress response in human T lymphocytes.

Exposure of cells to adverse environmental conditions invokes a genetically programmed series of events resulting in the induction of specific genes. The fluoroquinolone antibiotic ciprofloxacin has recently been reported to upregulate interleukin-2 (IL-2) gene induction. In the present investigation, the effect of ciprofloxacin at supratherapeutic concentrations on immediate-early (<2 h) gene expression in primary human peripheral blood lymphocytes was studied with Northern blots. In addition, transcriptional activity of IL-2 and metallothionein enhancer and promoter regions and transcription factors AP-1, NF-kappaB, and NF-AT were analyzed by chloramphenicol acetyltransferase (CAT) and electrophoretic mobility shift assays, respectively. The concentration of c-fos, c-jun, c-myc, junB, and fra-1 mRNAs was increased in activated peripheral blood lymphocytes incubated with ciprofloxacin compared to that in untreated controls. Ciprofloxacin increased CAT activity in stimulated lymphocytes transfected with plasmids containing either the IL-2 or metallothionein enhancer. Furthermore, among the transcription factors tested, AP-1 activity was increased in stimulated purified T helper lymphocytes incubated with ciprofloxacin compared to drug-free controls. Taken together, ciprofloxacin increased the levels of immediate-early transcripts, enhanced IL-2 and metallothionein promoter induction, and upregulated AP-1 concentrations in primary lymphocytes, reflecting a program commonly observed in mammalian stress responses.

Neutralization titres of HIV-1-specific monoclonal antibodies vary according to the batch of primary human peripheral blood lymphocytes, but do not vary coordinately

Human peripheral blood lymphocytes (PBL) were collected from five healthy adults under standard conditions and on a number of different occasions, and used in neutralization assays of human immunodeficiency virus type 1 (HIV-1) strain IIIB with three monoclonal antibodies (mAbs). Variations in neutralization titre were observed with different batches of PBLs with, for example, titres of ICR39.3b ranging from 1/10 to over 1/40000. However titres were as high, or higher, in PBLs than in C8166 cells (a human CD4 + T lymphoblastoid cell line) in 82% (28/34) of tests made. Most surprising was that neutralization by the three mAbs did not vary coordinately. In one batch of PBLs the neutralization titre of one of the mAbs might be increased while that of another mAb did not increase, or decrease. Thus PBLs could not be described as giving high or low levels of neutralization without reference to a specific mAb. This was not an assay problem as infectivity titres were relatively constant (varying by 1 to 1.4 fold with respect to C8166 cells), and neutralization titres were reproducible with the same batch of frozen PBLs over a three month period. Only one donor gave consistently low neutralization titres (defined here as 1/200; 2/2 batches tested) with all three mAbs, but all other donors gave similarly low titres with one of their batches of PBLs. The non-coordinate variation in neutralization titre indicates the advisability of using antibodies of several different specificities in any kind of preventive or therapeutic immunity.

Expression of the human RAD51 gene during the cell cycle in primary human peripheral blood lymphocytes.

The S. cerevisiae RAD51 gene product exerts important functions in meiotic and mitotic recombination, as well as in the repair of DNA double-strand breaks. We have studied the expression of the human RAD51 (HsRAD51) gene in primary human peripheral blood lymphocytes (PBLs). The HsRAD51 mRNA level increased three fold in mitogen stimulated PBLs, with a peak in the late S phase. A five fold increase of HsRAD51 protein levels was observed in late G2. Specific inhibition of DNA synthesis with aphidicolin did not block the induction of the HsRAD51 protein, indicating that HsRAD51 expression is independent of DNA replication. In contrast, after inhibition of RNA synthesis with actinomycin D and protein synthesis with cycloheximide, the HsRAD51 protein level decreased rapidly. Taken together, these results indicate that the HsRAD51 gene is transcriptionally regulated in human PBLs, and exerts its function during the late S and G2 phases of the cell cycle.

Anti-HIV Viral Interference Induced by Retroviral Vectors Expressing a Nonproducer HIV-1 Variant

A Hut-78 cell clone (F12) harboring a nonproducer human immunodeficiency virus (HIV-1) variant shows complete resistance to HIV-1 or HIV-2 superinfection. The F12-HIV provirus produces an altered HIV-1 protein pattern and cannot generate even immature viral particles. We demonstrated that HeLa CD4+ cells transfected with the F12-HIV genome resist HIV superinfection through a CD4-independent mechanism. As F12-HIV appears to be a useful system to induce anti-HIV intracellular immunization, we constructed various retroviral vectors containing the F12-HIV genome, modified by elimination of the F12 3'LTR and part of its nef gene, inserted 'antisense' with respect to the Moloney murine leukemia virus 5' LTR. Here we show that recombinant retroviral particles carrying the N2/F12-HIV nef- (as) construct can stably transduce both CEMss human cells and primary human peripheral blood lymphocytes, inducing the expression of the F12-HIV genome. These results could open the way to an anti-AIDS gene therapy strategy based on F12-HIV-induced intracellular immunization.

Further evaluation of soluble CD4 as an anti-HIV type 1 gene therapy: demonstration of protection of primary human peripheral blood lymphocytes from infection by HIV type 1.

We previously reported on the construction of retroviral vectors that produce a secreted form of the HIV-1 receptor, T cell antigen CD4 (Morgan et al., AIDS Res Hum Retroviruses 1990;6:183-191). In this article we test the ability of these sCD4-expressing retroviral vectors to protect human T-cell lines or primary T cells from HIV-1 infection. To demonstrate that protection from HIV-1 infection is mediated by the soluble nature of this protein, two coculture protection experiments were conducted. In these experiments, sCD4-expressing retroviral vectors were used to engineer mouse NIH 3T3 cells. In one coculture experiment the human SupT1 cell line was added directly to the culture of sCD4-producing NIH 3T3 cells, and in another experiment the two cell types were separated physically by a semipermeable membrane. In both coculture configurations, the T cell line was protected from HIV-1 challenge as measured by syncytium formation and indirect immunofluorescent assays. In addition, the SupT1 line was directly engineered with sCD4-expressing retroviral vectors and shown to be protected from HIV-1 challenge. As a prelude to further preclinical studies, we tested the ability of retroviral vectors to transduce primary human peripheral blood lymphocytes (PBLs). Conditions used to stimulate T cell growth resulted in significant shifts in the CD4/CD8 cell in favor of CD8 cells. Retroviral-mediated gene transfer under these conditions resulted in low levels of gene transfer (< 5%).(ABSTRACT TRUNCATED AT 250 WORDS)

Dilemma of Neutralization Resistance of HIV-1 Field Isolates and Vaccine Development

Dilemma of Neutralization Resistance of HIV-1 Field Isolates and Vaccine Development

CGP 53437, an orally bioavailable inhibitor of human immunodeficiency virus type 1 protease with potent antiviral activity

CGP 53437 is a peptidomimetic inhibitor of human immunodeficiency virus type 1 (HIV-1) protease containing a hydroxyethylene isostere. The compound inhibited recombinant HIV-1 protease with a Ki of 0.2 nM. The inhibition constant versus human cathepsin D and human cathepsin E was 4 nM. Human pepsin and gastricsin were inhibited with Kis of 8 and 500 nM, respectively, and human renin was inhibited with a Ki of 190 microM. The replication of HIV-1/LAV, HIV-1/Z-84, and HIV-1/pLAI was inhibited with a 90% effective dose of 0.1 microM in acutely infected MT-2 cells. The 50% cytotoxic dose was 100 microM. Similar antiviral activity was observed when the compound was added up to 10 h after infection. At the effective concentration, processing of Gag precursor protein p55 was greatly reduced, confirming an action on the late stage of the virus life cycle, as expected. The efficacy of the inhibitor was also demonstrated by using primary human peripheral blood lymphocytes infected with the HIV-1/LAV strain, low-passage clinical isolates obtained from HIV-1-seropositive individuals (including a zidovudine-resistant strain), and HIV-2/ROD. In these cells, CGP 53437 delayed the onset of HIV replication in a dose-dependent fashion (substantial effects with concentrations of > or = 0.1 microM) as long as the inhibitor was maintained in the culture. CGP 53437 was orally bioavailable in mice. Concentrations in plasma 10-fold in excess of the in vitro antiviral 90% effective dose could be sustained for several hours after oral application of 120 mg/kg. Therefore, CGP 53437 has the potential to be a therapeutically useful anti-HIV agent for the treatment of AIDS.

Intracellular immunization: trans-dominant mutants of HIV gene products as tools for the study and interruption of viral replication.

AIDS Research and Human RetrovirusesVol. 8, No. 6 Intracellular Immunization: Trans-Dominant Mutants of HIV Gene Products as Tools for the Study and Interruption of Viral ReplicationMARK B. FEINBERG and DIDIER TRONOMARK B. FEINBERGSearch for more papers by this author and DIDIER TRONOSearch for more papers by this authorPublished Online:16 Mar 2009https://doi.org/10.1089/aid.1992.8.1013AboutSectionsPDF/EPUB Permissions & CitationsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail FiguresReferencesRelatedDetailsCited byReferences30 April 2014Cellular Cofactors of Lentiviral Integrase: From Target Validation to Drug DiscoveryMolecular Biology International, Vol. 2012Approaches to Gene Therapy for Human Immunodeficiency Virus Infection Gary L. Buchschacher and Flossie Wong-Staal6 July 2004 | Human Gene Therapy, Vol. 12, No. 9Inhibition of Human Immunodeficiency Virus Type 1 by Tat/Rev-Regulated Expression of Cytosine Deaminase, Interferon alpha2, or Diphtheria Toxin Compared with Inhibition by Transdominant Rev Jack A. Ragheb, Larry Couture, Craig Mullen, Anthony Ridgway, and Richard A. Morgan6 July 2004 | Human Gene Therapy, Vol. 10, No. 1Inhibitors of Human Immunodeficiency Virus IntegraseInhibition of HIV Type 1 Infectivity by Coexpression of a Wild-Type and a Defective Glycoprotein 120 OLE SØGAARD LUND, BRITT LOSMAN, KRISTIAN SCHØNNING, ANDERS BOLMSTEDT, SIGVARD OLOFSSON, and JOHN-ERIK STIG HANSEN15 March 2009 | AIDS Research and Human Retroviruses, Vol. 14, No. 16Mutations in the Leucine Zipper-Like Heptad Repeat Sequence of Human Immunodeficiency Virus Type 1 gp41 Dominantly Interfere with Wild-Type Virus InfectivityJournal of Virology, Vol. 72, No. 6Engineering Catalytically Defective Forms of HIV Protease to Modulate Its ActivityGene Therapy for Infectious DiseasesClinical Microbiology Reviews, Vol. 11, No. 1Molecular biology of human immunodeficiency virus Type-1Transfusion Science, Vol. 17, No. 3Gene TherapyResearch at the Interface between Chemistry and Virology: Development of a Molecular Flashlight1 February 1996 | Chemical Reviews, Vol. 96, No. 1Antiviral therapy of hepatitis B virus infection: Blocking viral gene expressionAdvanced Drug Delivery Reviews, Vol. 17, No. 3Effects of second-site mutations on dominant interference by a human immunodeficiency virus type 1 envelope glycoprotein mutantJournal of Virology, Vol. 69, No. 2Further Evaluation of Soluble CD4 as an Anti-HIV Type 1 Gene Therapy: Demonstration of Protection of Primary Human Peripheral Blood Lymphocytes from Infection by HIV Type 1 RICHARD A. MORGAN, GABRIELE BALER-BITTERLICH, JACK A. RAGHEB, FLOSSIE WONG-STAAL, ROBERT C. GALLO, and W. FRENCH ANDERSON16 March 2009 | AIDS Research and Human Retroviruses, Vol. 10, No. 11Gene Therapy for Human Immunodeficiency Virus Infection: Genetic Antiviral Strategies and Targets for Intervention Boro Dropulić and Kuan-Teh Jeang19 March 2008 | Human Gene Therapy, Vol. 5, No. 8Therapy Other Than Reverse Transcriptase Inhibitors for HIV InfectionClinics in Laboratory Medicine, Vol. 14, No. 2The Molecular Biology of HIVClinics in Perinatology, Vol. 21, No. 1HIV life cycle and genetic approachesPerspectives in Drug Discovery and Design, Vol. 1, No. 1Gene Transfer for Therapy and Prophylaxis of HIV-1 InfectionAnnals of the New York Academy of Sciences, Vol. 685, No. 1 ImmunomodulatPathogenesis of human immunodeficiency virus infectionMicrobiological Reviews, Vol. 57, No. 1Can Diphtheria Toxin be Used for Gene Therapy of Human Immunodeficiency Virus Infection? Manuel Caruso, Geneviève Tsikas, Michèle Roussel, Marc Alizon, and David Klatzmann16 March 2009 | AIDS Research and Human Retroviruses, Vol. 8, No. 12 Volume 8Issue 6Jun 1992 To cite this article:MARK B. FEINBERG and DIDIER TRONO.Intracellular Immunization: Trans-Dominant Mutants of HIV Gene Products as Tools for the Study and Interruption of Viral Replication.AIDS Research and Human Retroviruses.Jun 1992.1013-1022.http://doi.org/10.1089/aid.1992.8.1013Published in Volume: 8 Issue 6: March 16, 2009PDF download

Multiple Effects of Interferon on HIV-1 Replication

Journal of Interferon ResearchVol. 11, No. 6 Multiple Effects of Interferon on HIV-1 ReplicationPAULA M. PITHAPAULA M. PITHASearch for more papers by this authorPublished Online:5 May 2009https://doi.org/10.1089/jir.1991.11.313AboutSectionsPDF/EPUB ToolsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail FiguresReferencesRelatedDetailsCited ByThe Role of Cytokines in Viral Infections15 March 2010Interferon-induced exonuclease ISG20 exhibits an antiviral activity against human immunodeficiency virus type 1Journal of General Virology, Vol. 86, No. 8Current status of gene therapy strategies to treat HIV/AIDSMolecular Therapy, Vol. 11, No. 6Structure of the mitochondrial ATP synthase by electron cryomicroscopyThe EMBO Journal, Vol. 22, No. 23Trans-Activated Interferon-α2 Delivered to T Cells by SV40 Inhibits Early Stages in the HIV-1 Replicative Cycle9 July 2004 | Journal of Hematotherapy & Stem Cell Research, Vol. 11, No. 5Immunologic Profile of Highly Exposed Yet HIV Type 1-Seronegative Men5 July 2004 | AIDS Research and Human Retroviruses, Vol. 18, No. 14Low-Dose IFN-α Monotherapy in Treatment-Naive Individuals with HIV-1 Infection: Evidence of Potent Suppression of Viral Replication7 July 2004 | Journal of Interferon & Cytokine Research, Vol. 21, No. 10AIDS-Associated MalignanciesRegulation of chemokine/cytokine network during in vitro differentiation and HIV‐1 infection of human monocytes: possible importance in the pathogenesis of AIDS1 September 2000 | Journal of Leukocyte Biology, Vol. 68, No. 3A Randomized Trial of Interferon alpha Therapy for HIV Type 1 Infection5 July 2004 | AIDS Research and Human Retroviruses, Vol. 16, No. 3Phase II, Randomized, Open-Label, Community-Based Trial to Compare the Safety and Activity of Combination Therapy With Recombinant Interferon-α2b and Zidovudine Versus Zidovudine Alone in Patients With Asymptomatic to Mildly Symptomatic HIV InfectionJournal of Acquired Immune Deficiency Syndromes and Human Retrovirology, Vol. 20, No. 3Molecular Mechanisms of Interferon Resistance Mediated by Viral-Directed Inhibition of PKR, the Interferon-Induced Protein KinasePharmacology & Therapeutics, Vol. 78, No. 1Regulation of Human Immunodeficiency Virus Replication by 2′,5′-Oligoadenylate-Dependent RNase LJournal of Virology, Vol. 72, No. 2Interferon-α and -γ Differentially Reduce Rapid Immature T-Cell Death by Contact with HIV-1 Carrier Cell Clones In Vitro14 November 2013 | Microbiology and Immunology, Vol. 41, No. 9Obstruction of HIV-1 Particle Release by Interferon-α Occurs Before Viral Protease Processing and Is Independent of Envelope Glycoprotein28 March 2009 | Journal of Interferon & Cytokine Research, Vol. 17, No. 5What Happens Inside Lentivirus or Influenza Virus Infected Cells: Insights into Regulation of Cellular and Viral Protein SynthesisMethods, Vol. 11, No. 4Interferon-betaHIV-permissive cells from solid tissues: Cytokine induction and effectsPerspectives in Drug Discovery and Design, Vol. 5, No. 1Anti-HIV Potential of a New Interferon, Interferon-τ (Trophoblastin)Journal of Acquired Immune Deficiency Syndromes and Human Retrovirology, Vol. 11, No. 3IMMUNE-BASED THERAPY FOR HIVNursing Clinics of North America, Vol. 31, No. 1Regulation of human immunodeficiency virus type 1 and cytokine gene expression in myeloid cells by NF-kappa B/Rel transcription factorsMicrobiological Reviews, Vol. 59, No. 3Selective decrease in human immunodeficiency virus type 1 (HIV-1)-induced alpha interferon production by peripheral blood mononuclear cells during HIV-1 infectionClinical Diagnostic Laboratory Immunology, Vol. 2, No. 2Further Evaluation of Soluble CD4 as an Anti-HIV Type 1 Gene Therapy: Demonstration of Protection of Primary Human Peripheral Blood Lymphocytes from Infection by HIV Type 116 March 2009 | AIDS Research and Human Retroviruses, Vol. 10, No. 11Specific inhibition of viral protein synthesis in HIV-infected cells in response to interferon treatment.Journal of Biological Chemistry, Vol. 269, No. 37Regulation of HIV Replication in Monocytes by InterferonIncreased Efficacy of Human Natural Interferon α (IFN-αn3) Versus Human Recombinant IFN-α2 for Inhibition of HIV-1 Replication in Primary Human Monocytes16 March 2009 | AIDS Research and Human Retroviruses, Vol. 9, No. 11HIV-1 reverse transcriptase: Inhibition by 2',5'-oligoadenylates1 May 2002 | Biochemistry, Vol. 32, No. 45Preclinical evaluation of antiviral activity and toxicity of Abbott A77003, an inhibitor of the human immunodeficiency virus type 1 proteaseAntimicrobial Agents and Chemotherapy, Vol. 37, No. 1The War Against the Interferon-induced dsRNA-Activated Protein Kinase: Can Viruses Win?5 May 2009 | Journal of Interferon Research, Vol. 12, No. 4 Volume 11Issue 6Dec 1991 To cite this article:PAULA M. PITHA.Multiple Effects of Interferon on HIV-1 Replication.Journal of Interferon Research.Dec 1991.313-318.http://doi.org/10.1089/jir.1991.11.313Published in Volume: 11 Issue 6: May 5, 2009PDF download

The ADA Human Gene Therapy Clinical Protocol Original Covering Memo: February 23, 1990

Severe combined immunodeficiency (SCID) due to deficiency of the purine metabolic enzyme adenosine deaminase (ADA) is a fatal childhood immunodeficiency disease. Immune reconstitution by transplantation with HLA-identical bone marrow is the treatment of choice. For patients not candidates for bone marrow transplantation, we propose to attempt immune reconstitution by using infusions of autologous T lymphocytes expanded in tissue culture and genetically corrected by insertion of a normal ADA gene using retroviral-mediated gene transfer. The vector is LASN, in which the human ADA gene is promoted by the LTR while the NeoR gene is driven by the SV40 early gene promoter. The packaging line is PA317. The protocol is designed to have two parts. In Part 1, autologous gene-corrected T lymphocytes would be infused repeatedly in low numbers in order to build an immune repertoire of T cells and also to obtain information as to how long gene corrected T cells survive in vivo. In Part 2A, the gene-corrected T cells would be selected in G418 and/or 2'deoxyadenosine and reinfused into the patient at monthly intervals for approximately 6 months. The goals would be essentially the same as in Part 1. In Part 2B, the number of gene-corrected T cells would be escalated in half-log increments to the predicted therapeutic level (probably around 1 x 10(9)/kg). 1-3 x 10(9)/kg gene-corrected cells would be infused several times and the patients would be monitored in order to determine if significant clinical improvement has occurred.