Dendritic cell vaccines for cancer therapy.

Dendritic cells (DC) are professional antigen-presenting cells whose primary function is the initiation of immune response. Based on the finding that the immune system usually fails to identify and kill cancer cells, DC have been recently used as vaccines for stimulation of tumour-specific immunity. This review focuses on pitfalls related to DC-based vaccination against solid tumours and on improvement of this immunotherapeutic approach for routine treatment of cancer disease.

Complement-dependent control of viral dynamics in pathogenesis of human immunodeficiency virus and simian immunodeficiency virus infection.

Since the first contact with the host, human immunodeficiency virus (HIV) exploits the complement system to reach maximal spread of infection. HIV has adapted many strategies to avoid complement-mediated lysis and uses the opsonization with complement fragments for attachment to complement receptors (CR). From the pathogen's perspective, binding to CR-expressing cells is remarkably beneficial, bringing together virus and activated target cells that are highly susceptible to infection. Moreover, complement-mediated trapping on CR+ cells permits HIV to infect surrounding cells even in the presence of an excess of neutralizing antibodies. Thus, complement activation initiates the assumption of power over the host's immune system by HIV and thus augments viral spread and replication throughout the body. On the other hand, natural hosts of primate lentiviruses, such as sooty mangabeys, African green monkeys and chimpanzees, are generally considered to be resistant to the development of AIDS, despite persistent viral replication. This review focuses on the possible link between the resistance to disease and species-specific diversity in function of human and monkey complement system.

The supportive role of complement in HIV pathogenesis.

This review focuses on interactions of HIV with the first-line defence of native immunity, the complement system. In all body compartments tested so far, HIV meets complement. Activation of the complement system results in deposition of C3 fragments on the viral surface, but in contrast to other pathogens, most of HIV is not or is only poorly lysed by membrane attack complexes. To survive complement-mediated lysis, HIV has not only developed resistance mechanisms, but uses opsonisation with complement fragments for its own advantage. Opsonised virions interact with complement receptor-expressing cells, which are either subsequently infected with high efficiency or retain viral particles on their surface, which promotes transmission of virus to other permissive cells. Our knowledge of these mechanisms has increased enormously over the past few years. A complete understanding of these complex interactions of HIV with the complement system opens new perspectives for development of alternative therapeutic strategies.

C5a and C5a(desArg) enhance the susceptibility of monocyte-derived macrophages to HIV infection.

Mononuclear phagocytes, which include circulating blood monocytes and differentiated tissue macrophages, are believed to play a central role in the sexual transmission of HIV infection. The ability of HIV to productively infect these cells may be influenced by action of exogenous or host-derived substances at the site of viral entry. Given the potent capacities of inflammatory mediators to stimulate anaphylatoxic and immunomodulatory functions in mucosa, the effects of complement-derived anaphylatoxins on the susceptibility of monocytes and monocyte-derived macrophages (MDM) to HIV-1 infection were examined. In our in vitro system, the susceptibility to infection was up to 40 times increased in MDM that had been exposed to C5a or C5a(desArg), but not to C3a or C3a(desArg), for 2 days before adding of virus. By contrast, the treatment with complement anaphylatoxins did not affect HIV replication in fresh monocytes. Stimulatory effect of C5a and its desArg derivative on HIV infection correlated with the increase of TNF-alpha and IL-6 secretion from MDM. All these functional effects of C5a and C5a(desArg) were reversible by treatment of cells with the mAb that functionally blocks C5aR. Taken together, these results indicate that C5a and C5a(desArg) may increase the susceptibility of MDM to HIV infection through stimulation of TNF-alpha and IL-6 secretion from these cells.

B cell-mediated infection of stimulated and unstimulated autologous T lymphocytes with HIV-1: role of complement.

In vivo, human immunodeficiency virus type 1 (HIV-1) is opsonized with complement fragments and virus-specific antibodies (Ab). Thus, HIV is able to interact with complement receptor (CR) - and Fc receptor (FcR) - positive cells such as B cells, follicular dendritic cells or macrophages. In this study we demonstrate that the interaction between B cells and HIV has an impact on autologous primary T cell infection in vitro. We confirmed the presence of complement-fragments and virus-specific Ab on serum-treated HIV using a virus-capture assay. In experiments with CR2-specific Ab we showed that the virus/B cell interaction was mainly dependent on CR2. In infection experiments immobilisation of HIV on stimulated tonsil B cells greatly enhanced the infection of interleukin (IL)-2-activated autologous tonsil T cells. Surprisingly, enhancement of T cell infection by B cell-HIV complexes was observed even in the absence of mitogenic stimuli such as PMA and was independent of the addition of exogenous IL-2. Taken together, these results indicate that primary B cells are able to efficiently transmit opsonised HIV to autologous primary T cells and induce a massive enhancement of infection. These in vitro experiments mimic the in vivo situation in the lymphoid tissue and suggest an alternative mechanism for the infection of primary T cells.

Detachment of human immunodeficiency virus type 1 from germinal centers by blocking complement receptor type 2.

After the transition from the acute to the chronic phase of human immunodeficiency virus (HIV) infection, complement mediates long-term storage of virions in germinal centers (GC) of lymphoid tissue. The contribution of particular complement receptors (CRs) to virus trapping in GC was studied on tonsillar specimens from HIV-infected individuals. CR2 (CD21) was identified as the main binding site for HIV in GC. Monoclonal antibodies (MAb) blocking the CR2-C3d interaction were shown to detach 62 to 77% of HIV type 1 from tonsillar cells of an individual in the presymptomatic stage. Although they did so at a lower efficiency, these antibodies were able to remove HIV from tonsillar cells of patients under highly active antiretroviral therapy, suggesting that the C3d-CR2 interaction remains a primary entrapment mechanism in treated patients as well. In contrast, removal of HIV was not observed with MAb blocking CR1 or CR3. Thus, targeting CR2 may facilitate new approaches toward a reduction of residual virus in GC.

Interleukin-15 enhances HIV-1-driven polyclonal B-cell response in vitro.

Interleukin-15 (IL-15) is a recently described cytokine, produced by monocytes/macrophages, with biological activities similar to IL-2. Since IL-15 was shown to stimulate human B-cell proliferation and immunoglobulin secretion, we investigated its effect on human B-cells stimulated with heat-inactivated human immunodeficiency virus type 1 (iHIV-1) in vitro. We observed a dose-dependent elevation of [3H]-thymidine incorporation and immunoglobulin production by B-cells incubated in the presence of iHIV-1. Moreover, IL-15 stimulated HIV-1-driven B-cell proliferation similarly to IL-2. As to immunoglobulin secretion, IL-15 was able to potentiate the stimulatory effect of IL-10. The highest amounts of iHIV caused a decrease in B-cell proliferation and immunoglobulin secretion to baseline levels, even in the presence of cytokines. These findings indicate that during the late stages of AIDS, when monocytes/macrophages become the major site of viral production, IL-15, in concert with other monocyte-derived cytokines, may promote polyclonal B-cell activation and hypergammaglobulinaemia, which are frequently associated with HIV infection.

Neutralization of HIV type 1 by alloimmune sera derived from polytransfused patients.

Antibodies (Abs) against HLA and other cell surface molecules, which HIV-1 acquires during the budding process at the host cell surface, neutralize HIV-1 in vitro. Macaques were protected against infection by SIV grown in human cells after xenoimmunization with human MHC molecules. Besides the immune responses arising against xenogeneic antigens, the highly polymorphic character of the HLA antigens enables the induction of alloresponses after exposure to allogeneic HLA molecules. Since polytransfused (PT) patients develop alloresponses, including humoral anti-HLA responses, we assumed that sera derived from PT patients may neutralize HIV-1. In a model system two PT sera out of a panel of 12 PT and 6 normal control sera neutralized HIV IIIB in vitro. Neutralizing activity of the PT sera was comparable to the efficacy of anti-HIV sera. The neutralizing capacity coincided with strong IgG reactivity against (HIV-infected) cell lines, which were used for virus production, and recognition of cell-free viral particles. Active human complement enhanced HIV neutralization mediated by the sera. Our results suggest an IgG-mediated neutralization based on recognition of allogeneic HLA molecules expressed on the viral surface. A vaccination strategy based on alloimmunization appears conceivable and requires further investigation.

Involvement of adenylate cyclase and p70(S6)-kinase activation in IL-10 up-regulation in human monocytes by gp41 envelope protein of human immunodeficiency virus type 1.

Our previous results show that recombinant gp41 (aa565-647), the extracellular domain of HIV-1 transmembrane glycoprotein, stimulates interleukin-10 (IL-10) production in human monocytes. The signal cascade transducing this effect is not yet clear. In this study, we examined whether gp41-induced IL-10 up-regulation is mediated by the previously described synergistic activation of cAMP and NF-kappaB pathways. gp41 induced cAMP accumulation in monocytes in a time- and concentration-dependent manner and the adenylate cyclase inhibitor SQ 22536 suppressed gp41-induced IL-10 production in monocytes. In contrast, gp41 failed to stimulate NF-kappaB binding activity in as much as no NF-kappaB bound to the main NF-kappaB-binding site 2 of the IL-10 promoter after addition of gp41. We also examined the involvement of other signal transduction pathways. Specific inhibitors of p70(S6)-kinase (rapamycin), and Gi protein (pertussis toxin), prevented induction of IL-10 production by gp41 in monocytes, while inhibitors of the phosphatidylinositol 3-kinase (PI 3-kinase) (wortmannin) and mitogen-activated protein kinase (MAPK) pathway (PD 98059) did not. Thus HIV-1 gp41-induced IL-10 up-regulation in monocytes may not involve NF-kappaB, MAPK, or PI 3-kinase activation, but rather may operate through activation of adenylate cyclase and pertussis-toxin-sensitive Gi/Go protein to effect p70(S6)-kinase activation.

Human immunodeficiency virus type 1 derived from cocultures of immature dendritic cells with autologous T cells carries T-cell-specific molecules on its surface and is highly infectious.

During the budding process, human immunodeficiency virus type 1 (HIV-1) acquires cell surface molecules; thus, the viral surface of HIV-1 reflects the antigenic pattern of the host cell. To determine the source of HIV-1 released from cocultures of dendritic cells (DC) with T cells, immature DC (imDC), mature DC (mDC), T cells, and their cocultures were infected with different HIV-1 isolates. The macrophage-tropic HIV-1 isolate Ba-L allowed viral replication in both imDC and mDC, whereas the T-cell-line-tropic primary isolate PI21 replicated in mDC only. By a virus capture assay, HIV-1 was shown to carry a T-cell- or DC-specific cell surface pattern after production by T cells or DC, respectively. Upon cocultivation of HIV-1-pulsed DC with T cells, HIV-1 exclusively displayed a typical T-cell pattern. Additionally, functional analysis revealed that HIV-1 released from imDC-T-cell cocultures was more infectious than HIV-1 derived from mDC-T-cell cocultures and from cultures of DC, T cells, or peripheral blood mononuclear cells alone. Therefore, we conclude that the interaction of HIV-1-pulsed imDC with T cells in vivo might generate highly infectious virus which primarily originates from T cells.

Dendritic cells transmit human immunodeficiency virus type 1 to monocytes and monocyte-derived macrophages.

Previous studies have shown that human immunodeficiency virus type 1 (HIV-1) exploits dendritic cells (DC) to replicate and spread among CD4(+) T cells. To explain the predominance of non-syncytium-inducing (NSI) over syncytium-inducing (SI) strains during the initial viremia of HIV, we investigated the ability of blood monocyte (Mo)-derived DC to transmit HIV-1 to CD4(+) cells of the monocytoid lineage. First, we demonstrate that in our system, DC are able to transmit NSI strains, but not SI strains, of HIV-1 to fresh blood Mo and to Mo-derived macrophages (MDM). To establish a productive infection, a 10-fold-lower amount of virus was necessary for DC-mediated transmission of HIV-1 to Mo than in case of cell-free infection. Second, immature CD83(-) DC (imDC) transmit virus to Mo and MDM with higher efficacy compared to mature CD83(+) DC (maDC); this finding is in contrast to data previously obtained with CD4(+) T cells. Third, maturation from imDC to maDC efficiently silenced expression of beta2-integrins CD11b, CD11c, and CD18 by maDC. Moreover, monoclonal antibody against CD18 inhibited transmission of HIV-1 from imDC to Mo. We propose that the adhesion molecules of the CD11/CD18 family, involved in cell-cell interactions of DC with the microenvironment, may play a major role in imDC-mediated HIV-1 infection of Mo and MDM.

HIV-1 induces human monocyte-derived macrophages to produce C3 and to fix C3 on their surface.

Complement components, particularly C3, are known to be involved in the pathogenesis of AIDS and macrophages may serve as a source of C3 at sites of infection. We investigated whether the interaction between HIV-1 and monocytes has any effect on C3 production by the cells. Monocytes isolated from the blood of healthy volunteers were incubated with monocytotropic and T lymphocytotropic HIV-1 strains or with recombinant gp160 and cultured in serum-free medium up to 7 days. Supernatants were tested for secreted C3 by enzyme-linked immunosorbent assay. Our data show that monocytes cultured with either the monocytotropic or the T lymphocytotropic HIV-1 strains produce C3 in large amounts. The effect of both viruses is dose dependent and the amount of C3 induced by HIV was up to 20-fold higher than in the control samples. C3 production was also enhanced by gp160, the envelope protein of the virus. Secretion of IL-6 by the cells was also measured and found to be elevated up to threefold as a consequence of the interaction with the virus. HIV-1-activated monocyte-derived macrophages acquired the capacity to cleave exogenous C3 and to fix generated C3 fragments on their cell membrane.

gp41 envelope protein of human immunodeficiency virus induces interleukin (IL)-10 in monocytes, but not in B, T, or NK cells, leading to reduced IL-2 and interferon-gamma production.

The effect of extracellular domain of human immunodeficiency virus (HIV-1) transmembrane glycoprotein gp41 on interleukin (IL)-10, IL-2, interferon (IFN)-y, IL-4, and tumor necrosis factor-alpha production by human peripheral blood mononuclear cells (PBMC) was assessed by ELISA. Rapid gp41-induced increase of IL-10 production was detected in resting PBMC and isolated monocytes but not in B, T, or NK cells. Furthermore, gp41 also enhanced IL-10 production in staphylococcal enterotoxin B-stimulated PBMC, while synthesis of IL-2, IFN-gamma, and IL-4 in these cells was down-modulated. Kinetic studies revealed that increased IL-10 production preceded reduction of IL-2, indicating the possible IL-10 regulatory role in the gp41-induced down-modulation of this cytokine. Anti-IL-10 antibody reversed almost completely the gp41 inhibitory effect on IL-2 production. In this study, HIV-1 gp41 was a potent modulator of cytokine production by PBMC, in particular by increasing IL-10 secretion from normal monocytes/macrophages and consequently down-regulating IL-2 and IFN-gamma.

Pentacyclic oxindole alkaloids from Uncaria tomentosa induce human endothelial cells to release a lymphocyte-proliferation-regulating factor.

In the present study we show that pentacyclic but not tetracyclic oxindole alkaloids from Uncoria tomentosa (Willd.) DC. (Rubiaceae) induced EA.hy926 endothelial cells to release some yet to be determined factor(s) into the supernatant; this factor was shown to significantly enhance proliferation of normal human resting or weakly activated B and T lymphocytes. In contrast, proliferation of normal human lymphoblasts and of both the human lymphoblastoid B cell line Raji and the human lymphoblastoid T cell line Jurkat was inhibited significantly while cell viability was not affected. Tetracyclic oxindole alkaloids dose-dependently reduce the activity of pentacyclic oxindole alkaloids on human endothelial cells.

Role of IL-15 in HIV-1-associated hypergammaglobulinaemia.

IL-15 is a novel cytokine, produced by monocytes/macrophages, with biological activities similar to IL-2 but with no significant sequence homology. IL-15 also stimulates human B cells to proliferation and immunoglobulin secretion. We measured serum levels of IL-15 in 84 HIV-1-infected individuals at different stages of disease in reference to 41 healthy blood donors. Our results show a marked elevation of IL-15 serum levels during HIV-1 infection. Moreover, we found that this increase correlated with serum levels of IgG (r = 0.376, P < 0.0001), and partly with serum IgM (r = 0.265, P = 0.015). A significant increase of IL-15 production by cultured peripheral blood mononuclear cells (PBMC) and purified monocytes in the presence of HIV-1 virus suggests that monocytes/macrophages may be a source of higher IL-15 serum levels in HIV-1-infected individuals. These findings indicate a participation of IL-15 in the hypergammaglobulinaemia frequently associated with HIV-1 infection.

Complement receptors in HIV infection.

The complement system plays an important role in the antimicrobial defense of the organism. Its components recognize a large variety of pathogens and target them for destruction, either directly by formation of a membrane attack complex or indirectly by recruiting phagocytic cells. In addition, it has several functions in cell activation, clearance of immune complexes, control of inflammatory reactions, chemotaxis and autoimmunity. For mediation of all these tasks of the complement system, complement receptor molecules on the cell surface play a key role. Current knowledge on structure, function, signal transduction and associated molecules is briefly summarized here. The role of complement receptors for human immunodeficiency virus (HIV)-associated pathogenesis is ambiguous and varies depending on cell type. On the one hand, complement receptors support the infected host to manage HIV infection and to defend itself, at least partially, against viral spreading throughout the organism. Such complement receptor-mediated supporting mechanisms are activation of immune cells and lysis of viral particles and infected host cells. On the other hand, HIV employs complement receptors to intrude more easily into various cell types, to become localized into lymph follicles and to activate viral replication in latently infected cells. This review summarizes the complex interaction of virus and complement receptors in HIV infection for different cell types.

Immunomodulatory effect of some amphiphilic detergents on the human promyelocytic HL-60 cells.

The effect of two anionic (sodium dodecyl sulfate (SDS) and sodium laurate), two cationic (didodecyldimethylammonium bromide (DDAB) and [(1-methyldodecyl)-trimethylammonium bromide (2-ATDBr)] and a nonionic [(1-methyldodecyl)-dimethyl-amine N-oxide (2-ATDNO)] amphiphilic detergents on the metabolic activation of phagocytes derived from human promyelocytic leukemia cell line HL-60 was investigated. SDS in the concentrations of 10(-4) M significantly stimulated the INT-reduction and superoxide production by HL-60 cells which were preincubated with phorbol 12-myristate 13-acetate (PMA) for a macrophage-like differentiation. This effect was not significant in the case of cells differentiated along monocyte-like and granulocyte-like pathways. A similar influence was observed by sodium laurate, but this effect was significant (P < 0.01) by INT-reduction only. Concentrations higher than 5 x 10(-5) M DDAB, 2-ATDBr and 2-ATDNO inhibited INT-reduction and superoxide generation in HL-60 cells differentiated along all three ways. Our results indicated that differentiated HL-60 cells may be used for in vitro testing of immunotoxic effects on respiratory burst in phagocytes.

Anti-mutagenic and immuno-stimulatory properties of lactic acid bacteria.

Statistically significant antigenotoxic activity was exerted by six of nine strains of lactic acid bacteria tested (Lactobacillus delbrueckii subsp. bulgaricus, Staphylococcus carnosus, Streptococcus thermophilus, L. rhamnosus, Enterococcus faecium and En. faecalis) against nitrovin and 2-aminofluorene in Salmonella typhimurium TA100 and TA97. The mutagenic activity of both mutagens was substantially decreased by viable bacteria; cells heated to 100°C for 15 min were ineffective. In vitro, En. faecium stimulated the basic metabolic activities of human neutrophils which were essential for their antimicrobial and cytotoxic activity, whereas stimulation of guinea-pig macrophages was not so effective. Similar immuno-stimulatory effects were observed with both viable and heat-inactivated bacteria.

Immunomodulatory effect of sodium dodecyl sulfate on human neutrophils and the human promyelocytic HL-60 cell line.

The effect of sodium dodecyl sulfate (SDS), an anionic amphiphilic detergent, on the function of human neutrophils and of the human promyelocytic leukemia cell line HL-60 was investigated. SDS modulated the respiratory burst in human neutrophils and HL-60 cells which were stimulated with phorbol 12-myristate 13-acetate (PMA). In concentrations above 1 X 10(-6) M it also caused release of lysosomal enzymes (beta-D-glucuronidase, myeloperoxidase and lysozyme) from neutrophils. Our results demonstrate that SDS at concentrations 1 X 10(-6) M-1 X 10(-4) M strongly affect properties of human phagocytic cells.