Search

IL-2 is currently used in HIV-infected patients to treat CD4+ T lymphopenia. In order to document a mechanism accounting for its capacity to restore immune function, we studied the effects of IL-2 administration in mice. IL-2 treatment of C57BL/6 mice for 4 days leads to a transient accumulation of CD4+ T lymphocytes. Whereas memory and activated CD4+ T lymphocytes accumulate after IL-2 treatment in both lymphoid and nonlymphoid organs, naive CD4+ T cells only accumulate in the former. IL-2 transiently increases CD4+ T lymphocyte numbers in lymphopenic IL-7(-/-) mice. Studies in T-cell-reconstituted Rag(-/-) gamma c(-/-) mice and in thymectomized mice demonstrated that IL-2 acts directly on peripheral CD4+ T lymphocytes. In vivo labeling of thymocytes showed that IL-2 also stimulates the release of CD4+ thymocytes from the thymus. Therefore, IL-2 treatment acts centrally and peripherally to increase the size of the naive CD4+ T lymphocyte compartment. This dual activity of IL-2 treatment may influence the quality of restoration of this compartment, especially regarding the ability to reconstitute a normal T lymphocyte repertoire.

Human immunodeficiency virus infection leads to a deregulated production of a number of cytokines. Some of them (IL-1, IL-6, TNF-alpha, interferon-gamma) are produced in increased amounts in vivo, whereas the production of IL-2 is decreased. This latter abnormality plays a pivotal role in the establishment of the immunodeficiency. Some cytokines (IL-1, IL-6, TNF-alpha) stimulate the in vitro replication of HIV, whereas others (mainly the interferons) inhibit it. The effect of cytokines in vivo in the spreading of HIV remains, however, largely unknown. Cytokines may also be involved in the development of many clinical manifestations associated with HIV infection. IL-1, IL-6 and TNF-alpha may play a role in tissue damages associated with opportunistic infections, in HIV-related encephalopathy and in cachexia. Cytokines, mainly IL-6, IL-10 and IL-13, may stimulate the growth of malignant cells during Kaposi sarcoma or lymphomas. Better knowledge of the role of cytokines during HIV infection should allow new therapeutic approaches based on the use of either recombinant cytokines or specific antagonists, with the aim of limiting both HIV spreading and the clinical manifestations of this infection.

Interferon-gamma (IFN-gamma) has been shown to inhibit human immunodeficiency virus (HIV) replication in macrophages. However, the site of its effect on the HIV infectious cycle is unknown. We show here that IFN-gamma inhibits the transactivation of HIV long terminal repeat (LTR) during viral infection and that it antagonizes tat effect in HT4LacZ-1 cells. HT4LacZ-1 is an indicator CD4+ HeLa cell line for HIV infectivity, because it harbors a HIV LTR-LacZ gene susceptible to transactivation by tat. It was used in combination with a computer-assisted image analyzer to quantify: (i) the number of transactivated foci following HIV infection, (ii) their individual level of transactivation, and (iii) the fusion potency of infected cells. IFN-gamma induced a 75% decrease of the number of transactivated foci following infection of HT4LacZ-1 cells by HIV. The remaining 25% foci still susceptible to transactivation were transactivated at a lower level than in control cultures, and the fusion potency of infected cells was strongly decreased. IFN-gamma acted after HIV entry into the cell and independently of reverse transcription. IFN-gamma antagonized tat-induced LTR transactivation: it inhibited transactivation of HT4LacZ-1 cells when tat was provided either from a SV40-based expression vector of tat or by polyethylene glycol-induced cell fusion with HeLa-tat-III cells. These results suggest that IFN-gamma affects the expression or the activity of cellular factors interacting with tat and that the high level of IFN-gamma production associated with HIV infection plays a role in the establishment of HIV latency.

Serine esterase B (SE B) is a protein contained in cytoplasmic granules of cytotoxic T lymphocytes and natural killer cells; SE B gene is transcribed upon activation of these cytotoxic cells. In order to show the in vivo interactions between HIV-infected cells and anti-HIV cytotoxic cells we analysed, by in situ hybridization, the expression of the SE B gene in eight hyperplastic lymph nodes from HIV-1-infected patients presenting with persistent generalized lymphadenopathy. We detected numerous cells expressing the SE B gene. The mean number of positive cells was 3.2 times higher in HIV lymph nodes than in six non-HIV hyperplastic lymph nodes studied in parallel (P less than 0.05). In control lymph nodes, the SE B gene was expressed only in interfollicular areas; virtually no cells expressed the SE B gene within follicles. In contrast, in HIV lymph nodes cells expressing the SE B gene were distributed either in interfollicular areas or within follicles. Expression of the SE B gene inside follicles was thus a specific feature of HIV lymph nodes (P less than 0.001) and was associated with the presence of HIV antigens and RNA at the same site. These results suggest that cytotoxic cells are activated in follicles of HIV lymph nodes and may be involved in the lysis of HIV-infected cells. Such a phenomenon may explain the development of follicle lysis, a specific feature of HIV lymph nodes. It may also inhibit the spreading of HIV infection.

We used a quantitative bioassay (the beta-gal assay) to visualize and quantify syncytium induction by fresh HIV isolates. This bioassay is based on the transactivation by tat of a chimeric gene comprising an HIV-1 long terminal repeat (LTR) fused to a modified lacZ gene of Escherichia coli. The chimeric gene encodes a beta-galactosidase which is translocated to the nucleus. It allows the enzymatic staining of all nuclei from HIV-induced syncytia. Using this unequivocal assay (the beta-gal assay), we could assess the syncytium-inducing properties of fresh HIV isolates after only 4 days of coculture of patient lymphocytes with activated normal lymphocytes. Syncytium-inducing HIV isolates were detected in 11 out of 40 seropositive patients studied. They were isolated mainly from AIDS patients: eight out of 17 grade IV (according to Centers for Disease Control criteria) patients were infected with syncytium-inducing strains. However, of 23 grade II and III patients tested, syncytium-inducing HIV strains were isolated from three cases. These three patients displayed no detectable p24 antigenaemia and had a CD4+ cell count of greater than 300 cells/microliter. The in vitro replication rate of HIV grown from 36 patient blood samples was then examined by sequential p24 antigen measurements in coculture supernatants. The 10 samples leading to syncytium formation also exhibited the highest replication rate. The possibility of unequivocally detecting syncytium-inducing strains after only a few days of coculture will make this detection routine and rapid. In addition, the limited period of amplification required is a significant advantage as it minimizes the emergence of HIV variants selected during long-term in vitro cultures.

Interleukin (IL)-10 and glucocorticoids (GCs) inhibit the ability of antigen-presenting dendritic cells (DCs) to stimulate T lymphocytes. We show that induction of GILZ (GC-induced leucine zipper) is involved in this phenomenon. IL-10, dexamethasone (DEX), and transforming growth factor (TGF)β stimulate GILZ production in human immature DCs derived from monocytes and from CD34+ cells. GILZ is necessary and sufficient for DEX, IL-10, and TGFβ modulation of CD80, CD83, CD86, immunoglobulin-like transcript (ILT)-3, and B7-H1 expression by DCs, and alteration of DC functions. GILZ stimulates the production of IL-10 by immature DCs and prevents the production of inflammatory chemokines by CD40L-activated DCs. In contrast, GILZ does not prevent CD40 ligand-mediated inhibition of phagocytosis, indicating that it affects some but not all aspects of DC maturation. GILZ prevents DCs from activating antigen-specific T lymphocyte responses. Administration of GCs to patients stimulates GILZ expression in their circulating antigen-presenting cells, and this contributes to the weak lymphocyte responses of GC-treated patients. Thus, regulation of GILZ expression is an important factor determining the decision of DCs whether or not to stimulate T lymphocytes, and IL-10, GCs, and TGFβ share this mechanism for influencing DC functions and the balance between immune response and tolerance.

To document the in vivo interactions occurring between the immune system and HIV replicating cells, we analyzed using in situ hybridization the production of IL-1 beta, IL-6, IL-2, and INF-gamma in eight hyperplastic lymph nodes from HIV-1 infected patients. Numerous IL-1 beta- and IL-6-producing cells associated in clusters were detected in sinuses. Few individual IL-1 beta- and IL-6-producing cells were present in interfollicular and follicular areas. IL-2- and INF-gamma-producing cells were observed in all lymph node compartments, with a selective enrichment in germinal centers. The amount and distribution of IL-1 beta, IL-6-, and IL-2-producing cells in HIV lymph nodes were not different from those found in six HIV unrelated hyperplastic lymph nodes. In contrast, a higher level of INF-gamma production was observed in HIV-1 lymph nodes. The CD8+ cells that accumulate in germinal centers of HIV lymph nodes (and not in non-HIV germinal centers) were actively involved in this INF-gamma production. INF-gamma synthesizing cells were in direct contact with cells containing HIV core antigens and HIV RNA. Thus a high INF-gamma production may characterize anti-HIV T cell immune response, potentially contributing to control of viral spreading as well as to the development of follicle lysis.

We analysed the cellular and molecular signals involved in the induction of non-specific T suppressor (Ts) cells generated in mouse spleen cell suspensions after 3-4 days' culture in the absence of conventional antigen. Using an in vitro model of primary antibody response as a suppressor assay, we showed that purified T cells were unable to generate suppressive activity. This suppression was fully restored when T cells were cultured with macrophages. In this case, Ts-cell generation was markedly increased by the addition of indomethacin suggesting a negative effect of prostaglandins in this phenomenon. Macrophages could be replaced by semi-purified IL1 as well as by recombinant IL2. Thus, in conditions of severe macrophage depletion, IL1 and/or IL2 are fully capable of inducing Ts-cell generation.

The role of macrophages in the in vitro response of mouse spleen cells to the insolubilized, T-independent antigen trinitrophenylated polyacrylamide (TNP-PAA) is demonstrated by the following points. The response is abolished by filtration on Sephadex G-10 and can be restored by the addition of splenic adherent cells, deficient in either B or T cells, or by 2-mercaptoethanol (2-ME). It is suppressed upon elimination of phagocytic cells by silica, and restored by 2-ME. 2-ME can restore a normal response from zero, in cultures depleted of both adherent and phagocytic cells, and is efficient in the absence of mature T cells. Experiments in microcultures show that large numbers of macrophages can stimulate a supra-optimal response from B cells. This response is only obtained in the presence of the antigen, and is specific for TNP. These results show that macrophages, probably by their polyclonal B-cell activator (PBA) property, play a role in the specific response to TNP-PAA. This prompts us to discuss the respective roles in the B-cell response of this PBA activity and of the interaction of the antigen with the specific B-cell receptors.

We have studied the effect of cyclophosphamide (CY) administration on the subsequent in vitro antibody response in the mouse. Treatment with a low dose (20 mg/kg) of CY four days before culture results in an increased IgM response to the T-independent antigen trinitrophenylated polyacrylamide (TNP-PAA), without affecting the background response of unstimulated cultures. This suggests that CY treatment eliminates a short-lived suppressor cell, involved in the regulation of the in vitro B cell response. In contrast, the same regimen decreases the ability of nude mouse spleen cells to respond to TNP-PAA, showing that the target of CY-enhancing effect is a mature T cell. The increased response observed in conventional mice should be the result of a balance between the direct suppressive effect of CY on B cells and the elimination of a suppresor T cell, the latter phenomenon being of predominant significance in our conditions. The target of CY-enhancing effect is nonadherent to plastic, but adherent to Sephadex G-10 columns.