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PubMed2005

Pre-incubation of cell-free HIV-1 group M isolates with non-nucleoside reverse transcriptase inhibitors blocks subsequent viral replication in co-cultures of dendritic cells and T cells.

In order to study the inhibitory effect of various reverse transcriptase inhibitors (RTIs) on cell-free HIV, we adapted a recently described in vitro system, based on co-cultures of dendritic cells and resting CD4 T cells, modelling early target cells during sexual transmission. The compounds tested included the second-generation non-nucleoside RTI (NNRTI) TMC-120 (R147681, dapivirine) and TMC-125 (R165335, travertine), as well as the reference nucleoside RTI AZT (zidovudine), the nucleotide RTI PMPA (tenofovir) and the NNRTI UC-781. The virus strains included the reference strain HIV-1Ba-L and six primary isolates, representative of the HIV-1 group M pandemic. They all display the non-syncytium-inducing and CCR5 receptor-using (NSI/R5) phenotype, important in transmission. Cell-free virus was immobilized on a poly-L-lysine (PLL)-treated microwell plate and incubated with compound for 1 h. Afterwards, the compound was thoroughly washed away; target cells were added and cultured for 2 weeks, followed by an extended culture with highly susceptible mitogen-activated T cells. Viral production in the cultures was measured on supernatant with HIV antigen ELISA. Negative results were confirmed by showing absence of proviral DNA in the cells. TMC-120 and TMC-125 inhibited replication of HIV-1Ba-L with average EC50 values of 38 nM and 117 nM, respectively, whereas the EC50 of UC-781 was 517 nM. Complete suppression of virus and provirus was observed at compound concentrations of 100, 300 and 1000 nM, respectively. Inhibition of all primary isolates followed the same pattern as HIV-1Ba-L. In contrast, pre-treating the virus with the nucleotide RTI PMPA and AZT failed to inhibit infection even at a concentration of 100000 nM. These data clearly suggest that NNRTIs inactivate RT enzymatic activity of different viral clades (predominant in the epidemic) and might be proposed for further testing as a sterilizing microbicide worldwide.

CD4-Positive T-Lymphocytes↗

PubMed1990

Reactivation of viral replication in anti-HBe positive chronic HBsAg carriers.

Reactivation of hepatitis B virus replication was investigated in an unselected group of 44 HBV DNA negative, anti-HBe positive chronic HBsAg carriers. Twenty-five patients (54%) were intravenous drug addicts and 7 (16%) were male homosexuals. Sixteen patients had evidence of delta infection and five of the seven male homosexuals had human immunodeficiency virus infection. The patients were followed for 1 to 180 months (median, 24 months) while HBV DNA negative, anti-HBe positive. Reactivation, defined as reappearance of HBV DNA or HBeAg, or both, was detected in six patients corresponding to an annual reactivation rate of 5%. Reactivation in four patients was detected by reversion to HBV DNA positivity only, whereas HBeAg/anti-HBe status remained unchanged. Two patients became both HBV DNA and HBeAg positive. None of the patients developed hepatitis-like symptoms and transaminase elevation was only observed in two patients. Reactivation in two patients was ascribed to human immunodeficiency virus infection and in one patient to chronic lymphatic leukaemia. It is concluded that HBV DNA seems to be superior to HBeAg in the detection of reactivation of HBV replication and that reactivation associated with clinical symptoms leading to progression in chronic liver disease is a rare event in the population studied.

Adolescent↗

PubMed1994

Presence of hepatitis C virus (HCV) genomic RNA and viral replicative intermediates in bone marrow and peripheral blood mononuclear cells from HCV-infected patients.

The cellular tropism of hepatitis C virus (HCV) was studied in vivo in samples from patients with persistent HCV infection. Plasma, liver, peripheral blood mononuclear cell (PBMC), and bone marrow cell (BMC) samples from 15 subjects positive for anti-HCV antibodies were tested for the presence of HCV RNA sequences by reverse transcription PCR. Virus-specific RNA sequences were found to be present in liver samples from all subjects (100%), in plasma samples from 13 of 15 patients (86.7%), in PBMC samples from 3 patients (20%), and in BMC samples from 9 (60%) of the 15 anti-HCV-positive patients enrolled in this study. The presence of the molecular intermediate of HCV replication (the negative-stranded HCV RNA) was evident in the two of the three PBMC and in five of the nine BMC HCV RNA-positive samples. Finally, we studied the nucleotide sequence of a large portion (-270 to -59) of the 5'untranslated region of HCV amplified from plasma samples of 12 of the 15 patients with and without HCV in BMCs; the degree of heterogeneity compared with the prototype HCV sequence was similar in both groups. The data principally indicate that HCV infection of PBMCs and BMCs is frequent in persistently infected patients, as shown by the occurrence of positive- and negative-stranded HCV RNA, thus suggesting the possibility of extrahepatic replication of HCV.

Adult↗

PubMed2004

Suppression of HIV-1 viral replication and cellular pathogenesis by a novel p38/JNK kinase inhibitor.

OBJECTIVE: To analyze a novel compound, which inhibits serine-threonine protein kinase p38, for its possible bioactivity against HIV-1 infection. METHODS: Proteins involved in cellular signal transduction pathways represent a novel class of host therapeutic targets for infectious diseases. In this regard the serine/threonine kinase p38 MAPK, a member of the mitogen-activated protein (MAP) kinase superfamily of signal transduction molecules may play an important role in HIV-1 infection. We analyzed the ability of this compound (RWJ67657) to inhibit HIV replication in primary T cells and monocytes. Cellular expression of phospho-p38MAPK was studied by Western blot analysis. Blockade of HIV infection induced apoptosis was measured by Annexin V staining. RESULTS: p38 inhibitor RWJ67657 was effective in inhibiting HIV-1 replication in both T-cell and monocyte cell lines, irrespective of the coreceptor used by the virus for entry into the cell. Importantly, both reverse transcriptase and protease resistant escape mutant viruses were effectively suppressed by RWJ67657. In addition, the tested compounds block HIV-induced T-cell apoptosis, a critical means of T-cell depletion linked to AIDS progression. CONCLUSION: Several steps in the HIV-1 virus life cycle appear to depend on cellular activation, including activation of the p38 pathway. Without activation virus replication is thought to be blocked due to incomplete reverse transcription and a lack of proviral DNA integration. The data collectively illustrate that inhibition of the p38 pathway can affect HIV-1 replication. Interruption of HIV infection by p38 inhibitors underscores the value of exploring antiviral drugs that target host cellular proteins.

Analysis of Variance↗

PubMed2002

Residual viral replication during antiretroviral therapy boosts human immunodeficiency virus type 1-specific CD8+ T-cell responses in subjects treated early after infection.

Human immunodeficiency virus type 1 (HIV-1)-infected subjects treated early after infection have preserved HIV-1-specific CD4+ T-cell function. We studied the effect of highly active antiretroviral therapy (HAART) on the frequency of HIV-1-specific CD8+ T cells in patients treated during early (n = 31) or chronic (n = 23) infection. The degree of viral suppression and time of initiation of treatment influenced the magnitude of the CD8+ T-cell response. HIV-1-specific CD8+ T cells can increase in number after HAART in subjects treated early after infection who have episodes of transient viremia.

Acute Disease↗

PubMed1997

Loss of inducible virus in CD45RA naive cells after human immunodeficiency virus-1 entry accounts for preferential viral replication in CD45RO memory cells.

Controversy exists concerning the preferential infection and replication of human immunodeficiency virus-1 (HIV-1) within naive (CD45RA+) and memory (CD45RO+) subsets of CD4+ lymphocytes. To explore the susceptibility of these subsets to HIV-1 infection, we purified CD45RA+/CD4+ (RA) and CD45RO+/CD4+ (RO) cells from normal donors and subjected them to a novel monokine activation culture scheme. Following HIV-1 infection and interleukin-2 (IL-2) induction, viral production measured on day 13 was 19-fold greater in RO cultures compared with RA cultures. IL-2-stimulated proliferation in uninfected control cultures was equivalent. To explore the mechanisms by which RA cells were reduced in viral production capacity, RA and RO cells were exposed to HIV-1 followed by treatment with trypsin, and then phytohemagglutinin antigen (PHA)-stimulated at days 4, 7, and 10 postinfection. HIV-1 production in day 4 postinfection RA and RO cultures was analogous, indicating that viral fusion and entry had occurred in both cell types. However, whereas similarly treated day 7 and 10 postinfection RO cultures produced virus, HIV-1 was markedly reduced or lost in the corresponding RA cultures. These results suggest that a temporally labile postfusion HIV-1 complex exists in unstimulated RA cells that requires cellular activation signals beyond that provided by IL-2 alone for productive infection.

CD4-Positive T-Lymphocytes↗

PubMed2005

Point mutations in the C-terminus of HIV-1 gp160 reduce apoptosis and calmodulin binding without affecting viral replication.

One hallmark of AIDS progression is a decline in CD4+ T lymphocytes, though the mechanism is poorly defined. There is ample evidence that increased apoptosis is responsible for some, if not all, of the decline. Prior studies have shown that binding of cellular calmodulin to the envelope glycoprotein (Env) of HIV-1 increases sensitivity to fas-mediated apoptosis and that calmodulin antagonists can block this effect. We show that individual mutation of five residues in the C-terminal calmodulin-binding domain of Env is sufficient to significantly reduce fas-mediated apoptosis in transfected cells. The A835W mutation in the cytoplasmic domain of gp41 eliminated co-immunoprecipitation of Env with calmodulin in studies with stably transfected cells. Four point mutations (A835W, A838W, A838I, and I842R) and the corresponding region of HIV-1 HXB2 were cloned into the HIV-1 proviral vector pNL4-3 with no significant effect on viral production or envelope expression, although co-immunoprecipitation of calmodulin and Env was decreased in three of these mutant viruses. Only wild-type envelope-containing virus induced significantly elevated levels of spontaneous apoptosis by day 5 post-infection. Fas-mediated apoptosis levels positively correlated with the degree of calmodulin co-immunoprecipitation, with the lowest apoptosis levels occurring in cells infected with the A835W envelope mutation. While spontaneous apoptosis appears to be at least partially calmodulin-independent, the effects of HIV-1 Env on fas-mediated apoptosis are directly related to calmodulin binding.

Apoptosis↗

PubMed1993

Human immunodeficiency virus type 1 DNA synthesis, integration, and efficient viral replication in growth-arrested T cells.

Human immunodeficiency virus type 1 (HIV-1) replicates efficiently in nonproliferating monocytes and macrophages but not in resting primary T lymphocytes. To determine the contribution of cell division to the HIV-1 replicative cycle in T cells, we evaluated HIV-1 expression, integration of proviral DNA, and production of infectious progeny virus in C8166 T-lymphoid cells blocked in cell division by treatment with either mitomycin, a DNA cross-linker, or aphidicolin, a DNA polymerase alpha inhibitor. The arrest of cell division was confirmed by assay of [3H]thymidine uptake; the nondividing cells remained viable for at least 3 days after treatment. HIV-1 was expressed and replicated equally well in nondividing and dividing C8166 cells, as judged by the comparison of the levels of p24 core antigens in culture supernatants, the proportion of cells expressing HIV-1 specific antigens, the pattern and quantity of HIV-1 DNA present in the extrachromosomal and total cellular DNA fractions, and the biological activity of progeny viruses. A polymerase chain reaction-based viral DNA integration assay indicated that HIV-1 provirus was integrated in C8166 cells treated with either of the two inhibitors of cell division. Similar results were obtained by using growth-arrested Jurkat T-lymphoid cells. We conclude that cell division and cellular DNA synthesis are not required for efficient HIV-1 expression in T cells.

Aphidicolin↗

PubMed1991

Plasma viraemia as a marker of viral replication in HIV-infected individuals.

Two hundred and twenty-eight blood samples from 154 HIV-seropositive subjects were investigated for the presence of HIV in peripheral blood mononuclear cells (PBMC) and plasma (without prior ultracentrifugation or filtration), using normal PBMC as the target for replication. HIV was recovered from PBMC and plasma in 80.5 and 19.5% of the patients, respectively. Plasma viraemia was significantly associated with clinical manifestations of HIV infection, indicating that HIV replication increased as disease progressed. This was confirmed by the statistically significant correlations between plasma viraemia and low CD4 cell counts (P less than 0.01) and low anti-p24 antibody titers (P less than 0.01). On patient follow-up, detection of HIV in plasma was transient. p24 antigenaemia was only detected in 13.6% of cases and was also associated with advanced clinical stages of the disease. When HIV RNA detection by polymerase chain reaction (PCR) was compared with plasma viraemia, HIV RNA was detected in plasma in all symptomatic cases and in 53.8% (seven out of 13) of asymptomatic patients [Centers for Disease Control (CDC) stages II and III], confirming that PCR was far more sensitive than plasma culture. These results indicate that cell-free virus production is associated with the clinical stage of HIV infection and may serve as a marker for disease progression. Detection of HIV RNA by PCR appears to be the most sensitive method to detect viraemia.

AIDS-Related Complex↗

PubMed1992

Relationship between frequency of infectious human immunodeficiency virus type 1-harboring cells and kinetics of viral replication: a simple procedure for quantitation of infectious virus-carrying cells in blood samples.

Statistical analysis of a limiting dilution assay (LDA) showed that the occurrence of infectious human immunodeficiency virus type 1 (HIV-1)-harboring cells in serially diluted samples of peripheral blood mononuclear cells (PBMCs) of HIV-1-seropositive patients fits the model describing a single-hit Poisson distribution. This observation led to the discovery that there is a direct correlation (r = 0.957) between the number of HIV-1-positive cells and the time when viral culture produces 1 ng of the HIV-1 p24 gag protein per ml. Frequency estimates based on this relationship were highly accurate (P less than 0.01) within the first 15 days of viral culture, which consisted of coculture of 10(6) normal PBMCs with the equivalent number of test PBMCs. This approach was less cumbersome than LDA and was sensitive enough to detect a single infectious HIV-1-harboring cell among as many as 320,000 cells. The values obtained for 57 patients agreed well with the data in the literature and showed that the frequencies of infectious cells in PBMCs reflect the advancement in the clinical stage, being 1/38,000, 1/11,000, and 1/7,000 for asymptomatic patients (Centers for Disease Control [CDC] group II/III), patients with AIDS-related complex (CDC group IVa), and patients with AIDS (CDC group IVb/c), respectively. A nearly 10-fold disparity in mean frequencies was observed when these values were correlated with the numbers of CD4-positive cells (1/9,000, 1/1,500, and 1/300, respectively, for asymptomatic patients, patients with AIDS-related complex, and patients with AIDS). The described method provides a simple means of determining infectious HIV-1-positive cells in blood samples.

HIV Infections↗

PubMed1994

Basal and Tat-transactivated expression from the human immunodeficiency virus type 1 long terminal repeat in human placental trophoblast rules out promoter-enhancer activation as the partial block to viral replication.

We have analysed the capacity of the trophoblast-derived malignant cell lines BeWo, JAR and JEG-3, and primary cultures of highly purified trophoblast cells to support the basal and Tat-mediated trans-activation-enhanced transcriptional activity of two distinct human immunodeficiency virus type 1 (HIV-1) isolates. Kinetic studies based on expression of long terminal repeat (LTR)-chloramphenicol acetyltransferase (CAT) constructs revealed that LTRs of both the prototype strain 3B and the highly cytopathic Zairean variant NDK were activated significantly in all target cells. Overall, the strongest activation was observed in primary trophoblasts. A novel modification of quantitative PCR was used to normalize LTR expression for transfection efficiency, enabling the calculation of specific expression rates in terms of muU CAT enzyme per fmol of transfected DNA. Using the latter criterion we determined that LTRs of both viruses were activated in decreasing order from trophoblasts to JAR, JEG-3 and BeWo cells; furthermore, the expression of HIV-1 3B LTR always significantly surpassed that of HIV-1 NDK. The effects of trans-activation on either of the LTRs, when assayed in cotransfection assays with various amounts of HIV-1 NDK-Tat expression vector, increased in a dose-dependent fashion and were comparable in a particular neoplastic cell line. Furthermore, the cell-specific LTR activity patterns did not correspond to the abundance of transcription factors binding specifically to the viral NF kappa B and SP1 motifs. Unlike SP1-binding proteins which were relatively abundant, substantially smaller amounts of proteins with NF kappa B specificity were found in all cells. Despite this apparent deficit in NF kappa B activity, trophoblasts supported a high basal activity of both LTRs. These data indicate that an insufficiency of basal or Tat-trans-activated LTR activity cannot account for the low level of HIV-1 replication in this important cell type.

Base Sequence↗

PubMed2005

Receptor modulation in viral replication: HIV, HSV, HHV-8 and HPV: same goal, different techniques to interfere with MHC-I antigen presentation.

Evasion of host immunity is a common objective of viruses that cause chronic infections. Viruses involved in sexually transmitted infections constitute no exception to this phenomenon. HIV, HPV, HSV, and HHV-8 subvert the class I major histocompatibility complex (MHC-I) antigen presentation pathway, thereby evading the cellular immune response. Although the goal of these viruses is the same and efficient MHC-I downregulation in infected cells is achieved, their techniques vary considerably. Whether viral inhibition occurs at the transcriptional level, during assembly of MHC-I complexes in the endoplasmic reticulum, during its journey to the cell surface, or after reaching the cell surface, each one of these viruses ingeniously achieves MHC-I downregulation and avoids the cellular immune response. Unraveling the mechanisms of interference with MHC-I antigen presentation employed by these viruses is not only crucial to understand their pathogenesis, but also reveals novel mechanisms of regulation of cellular receptors. When employed as modulators of cellular trafficking pathways, viruses become tools to dissect fundamental cell processes. In return, the precise dissection of these processes may offer new weapons against the ruses viruses employ to propagate and establish chronic infections.

Antigen Presentation↗

PubMed1998

Antisense RNA complementary to hepatitis B virus specifically inhibits viral replication.

BACKGROUND & AIMS: Chronic infection with the hepatitis B virus (HBV) is a major public health problem, and currently available therapies have limited efficacy. Gene therapy strategies for HBV infection are under active investigation. We evaluated the potential of antisense RNA transcribed from antisense genes to interfere with HBV replication. METHODS: Subgenomic fragments of the HBV genome were studied with respect to the property of inhibiting HBV replication when intracellularly expressed in the antisense orientation. RESULTS: Antisense RNAs derived from the HBV genome specifically inhibited HBV replication and antigen expression in human hepatocellular carcinoma cells by 60%-75%. DNA sequences corresponding to the identified RNAs had no effect on HBV replication, indicating that inhibitory effects are mediated by RNA. Transcripts corresponding to the inhibitory subgenomic fragments were present at high levels. One antisense RNA was found to reduce the amount of pregenomic RNA encapsidated into core particles as a molecular mechanism of antiviral effects. CONCLUSIONS: Certain antisense RNA molecules will have substantial antiviral effects against HBV. Antisense RNAs derived from the HBV genome are promising candidates as antiviral agents and may serve as novel tools to identify functionally important regions of HBV transcripts.

Base Sequence↗

PubMed2000

Persistence of viral replication after anti-HBe seroconversion during antiviral therapy for chronic hepatitis B.

BACKGROUND/AIMS: Hepatitis B virus genome mutants may be selected during the immune-mediated clearance of infection or during long-term nucleoside analog administration and may escape both antiviral pressures. The pattern of anti-HBe seroconversion was analyzed in patients receiving new nucleoside analogs, lamivudine or famciclovir, in comparison with patients treated with interferon alpha. METHODS: Eighteen consecutive patients who seroconverted to anti-HBe were included in the study. Serial serum samples were studied with the quantitative determination of HBV DNA by the branched DNA assay (Chiron) and by a quantitative PCR assay (Roche diagnostics), determination of pre-S1 Ag, the genetic analysis of the viral genome with the determination of pre-core promoter or pre-core region mutations with a line probe assay (Innogenetics) and, in selected samples of polymerase gene mutations. RESULTS: The quantitative PCR assay was found to be more sensitive than the bDNA assay, allowing a 25-log decrease in viral DNA levels to be demonstrated after anti-HBe seroconversion. Viral persistence after anti-HBe seroconversion induced by interferon, lamivudine or famciclovir, was often associated with circulating HBV genomes harboring mutations in the precore promoter. The clinical significance of these findings was demonstrated by the observation of reversion to HBeAg in two patients treated with interferon and one with lamivudine. CONCLUSION: Persistence of significant levels of viremia that are not detected by the branched DNA assay may be observed after anti-HBe seroconversion. A precise monitoring of viremia levels with more sensitive assays and HBV mutant strains is warranted in patients undergoing antiviral therapy.

2-Aminopurine↗

PubMed1989

Postpartum subsidence of hepatitis B viral replication in HBeAg-positive carrier mothers.

To elucidate the effects of pregnancy and delivery on hepatitis B e antigen (HBeAg)-positive carrier mothers, 31 HBeAg-positive carrier mothers were followed-up postpartum 1 year, with 30 HBeAg-positive nonpregnant female carriers as controls. Serum hepatitis B surface antigen (HBsAg), HBeAg titer, and hepatitis B virus (HBV)-DNA concentration were studied at defined intervals. The results revealed that in the control group HBeAg titers and HBV-DNA concentrations fluctuated, whereas the HBsAg titers showed little change, but HBeAg clearance or seroconversion to anti-HBe were not noted on follow-up. In contrast, one carrier mother seroconverted to anti-HBe during pregnancy and the antibody persisted thereafter. Five of the remaining 30 carrier mothers cleared HBeAg postpartum, and among these five cases, one also seroconverted to anti-HBe. In addition, in another five of the 30 cases, the HBV-DNA fell to undetectable level (less than 0.04 ng/ml). All these ten cases had a common tendency of showing a decrease in HBeAg titers and/or HBV-DNA concentrations 1-2 months after delivery. The HBeAg titers and HBV-DNA concentrations in the other 11 cases remained unchanged, whereas the remaining nine cases had increased levels. It is concluded that subsidence of HBV replication is precipitated by delivery in one-third of HBeAg-positive carrier mothers in Taiwan, and this occurs most frequently 1-2 months postpartum.

Adult↗

PubMed2000

Intrinsic susceptibility of rhesus macaque peripheral CD4(+) T cells to simian immunodeficiency virus in vitro is predictive of in vivo viral replication.

Previous studies with simian immunodeficiency virus (SIV) infection of rhesus macaques suggested that the intrinsic susceptibility of peripheral blood mononuclear cells (PBMC) to infection with SIV in vitro was predictive of relative viremia after SIV challenge. The present study was conducted to evaluate this parameter in a well-characterized cohort of six rhesus macaques selected for marked differences in susceptibility to SIV infection in vitro. Rank order relative susceptibility of PBMC to SIVsmE543-3-infection in vitro was maintained over a 1-year period of evaluation. Differential susceptibility of different donors was maintained in CD8(+) T-cell-depleted PBMC, macrophages, and CD4(+) T-cell lines derived by transformation of PBMC with herpesvirus saimiri, suggesting that this phenomenon is an intrinsic property of CD4(+) target cells. Following intravenous infection of these macaques with SIVsmE543-3, we observed a wide range in plasma viremia which followed the same rank order as the relative susceptibility established by in vitro studies. A significant correlation was observed between plasma viremia at 2 and 8 weeks postinoculation and in vitro susceptibility (P < 0.05). The observation that the two most susceptible macaques were seropositive for simian T-lymphotropic virus type 1 may suggests a role for this viral infection in enhancing susceptibility to SIV infection in vitro and in vivo. In summary, intrinsic susceptibility of CD4(+) target cells appears to be an important factor influencing early virus replication patterns in vivo that should be considered in the design and interpretation of vaccine studies using the SIV/macaque model.

Animals↗

PubMed2001

Hepatitis B virus reactivation during cytotoxic chemotherapy-enhanced viral replication precedes overt hepatitis.

The diagnosis of HBV reactivation during cytotoxic chemotherapy is based on an abrupt rise in levels of serum HBV DNA in conjunction with a hepatitic picture and in the absence of other causes of hepatitis. However, several cases of "hepatitis" have been noted during chemotherapy in HBsAg-positive patients, for which no cause could be found and in which HBV DNA levels were negative. One possible explanation is that HBV reactivation may, indeed, have been the cause but that HBV DNA levels became negative by the time ALT levels peaked and the clinical diagnosis of hepatitis was made. During the course of a longitudinal study of HBsAg-seropositive cancer patients who were monitored during standard chemotherapy, the opportunity was available to test this hypothesis.

Adult↗

PubMed1993

Cytokines from vaccine-induced HIV-1 specific cytotoxic T lymphocytes: effects on viral replication.

Cytolytic T lymphocytes (CTLs) specific for the human immunodeficiency virus (HIV-1) envelope glycoproteins have been cloned from HIV-1-seronegative human volunteers immunized with HIV-1 gp160-based candidate vaccines. Although vaccine-induced CTLs can potentially contribute to the antiviral response by direct lysis of infected cells, these CTLs may also produce cytokines that alter HIV-1 gene expression in other infected cells present in the microenvironment where CTL-target cell interactions occur. Vaccine-induced CTL clones were therefore examined for production of cytokines that affect HIV-1 gene expression in chronically infected T lymphocytic and promonocytic cell lines. Enhancement of HIV-1 gene expression was observed with supernatants from CD4+ CTL clones and with supernatants from a subset of CD8+ CTL clones. For each clone studied, upregulation of HIV-1 gene expression in chronically infected T cell lines resulted from the antigen-specific release by CTLs of tumor necrosis factor alpha (TNF-alpha). CD4+ and CD8+ CTLs that released TNF-alpha on antigen stimulation were also shown to express a biologically active 26-kDa transmembrane form of TNF-alpha, which was sufficient to induce upregulation of HIV-1 gene expression in chronically infected T cells placed in direct contact with the CTLs. Supernatants from antigen-activated, vaccine-induced CD4+ and CD8+ CTLs also caused upregulation of HIV-1 gene expression in chronically infected promonocytic cells. A subset of CD8+ CTL clones also produced a soluble factor(s) that inhibited HIV-1 replication in acutely infected autologous CD4+ blasts. Supernatants from CD4+ CTLs had no effect on HIV-1 replication in acutely infected CD4+ blasts. These results suggest that cytokine production as well as cytolytic activity should be evaluated in the analysis of the potential antiviral effects of vaccine-induced CTLs.

AIDS Vaccines↗