Involvement of interleukin 18 in Crohn's disease: evidence from in vitro analysis of human gut inflammatory cells and from experimental colitis models

An imbalance of immunoregulatory factors and/or cells contributes to uncontrolled mucosal T cell activation and inflammation in Crohn's disease (CD). Bioactive interleukin (IL)-18 has been shown to be produced by macrophages in CD lesions. We report here that T cells freshly isolated from inflamed tissue of CD patients (and not T cells from control intestinal tissue) were responsive to IL-18. In the presence of IL-18, these T cells produced more interferon (IFN)-gamma and less IL-10. To analyse further the role of IL-18 in this disease, an acute and a chronic model of murine colitis were used. IL-18 mRNA was significantly enhanced in trinitrobenzene sulphonic acid (TNBS) induced colitis, and treatment with IL-18 binding protein (IL-18BPa), which neutralizes IL-18 bioactivity, significantly reduced the severity of colitis. However, IL-18BPa did not affect the course of chronic colitis in CD45RBhighCD4+ T cell reconstituted SCID mice. Production of IFN-gamma in lamina propria mononuclear cell cultures from IL-18BPa-treated SCID mice was decreased, but at the same time fewer lamina propria CD4+ T cells harvested from IL-18BPa-treated mice compared to non-treated mice were in apoptosis. We conclude that IL-18 clearly has a modulatory role in the inflammatory cascade of CD and experimental colitis by affecting IFN-gamma and IL-10 production, and apoptosis. In view of the divergent effects of IL-18 neutralization in the two different murine colitis models, it is unlikely that IL-18 is at the top of this cascade.     

Cytokine levels during mild and cerebral falciparum malaria in children living in a mesoendemic area

Summary Cell-mediated immunity and cytokines are probably involved in the pathogenesis of malaria. To investigate the role and the activity of different immune cells, we measured levels of tumour necrosis factor-(TNF-α), gamma interferon (IFN-γ) and several interleukins (IL-2, IL-4, IL-6 and IL-10) in children with mild (MM) and cerebral (CM) Plasmodium falciparum malaria and compared them with those of healthy children from Guadalupe - Lobata District, St. Tomé Island, where malaria is mesoendemic. Both groups of patients had significantly higher levels of IL-6, IL-10 and TNF-α than controls. For IL-2, IL-4 and IFN-γ we found no difference between the groups. However, 24 h after admission the levels of IL-10 and IL-6 were significantly higher in CM than in MM patients, although 7 days after treatment they returned to normal levels, similar to those found in control children. Therefore, TNF-α IL-6 and IL-10 increase during Plasmodium falciparum attacks in all children, not only in those with cerebral malaria. This finding suggests the activation of the monocyte/macrophage system during the early stage of clinical malaria.     

Ex vivo models of HIV sexual transmission and microbicide development

Here, we review the armamentarium on in vitro/ex vivo models of HIV sexual transmission and discuss how these models can be applied to study candidate microbicides.     

Survey of the temporal changes in HIV-1 replicative fitness in the Amsterdam Cohort

Changes in virulence and fitness during an epidemic are common among pathogens. Several studies have shown that HIV fitness increases within a patient during disease progression, while bottlenecks, such as sexual transmission, immune pressure and drug treatment can reduce fitness. In this study, we analyzed how these opposing forces have shaped HIV-1 fitness over time. Therefore, we compared the replicative fitness of HIV-1 isolates from newly infected untreated individuals, diagnosed for HIV-1 infection early in the AIDS epidemic in Amsterdam, the Netherlands, with more recent isolates. Twenty-five early and late HIV-1 isolates, carefully matched for seroconversion time, were competed head-to-head in a dual infection/competition assay, employing peripheral blood mononuclear cells. In contrast with previous studies, we observed a trend of increasing fitness over time in the HIV epidemic of Amsterdam. Apparently, the bottleneck, occurring with each transmission event, does not completely reset the fitness increase acquired during disease progression.     

The intraspleen huPBL NOD/SCID model to study the human HIV-specific antibody response selected in the course of natural infection

The intrasplenic injection of human peripheral blood mononuclear cells (PBMCs) into severely immune deficient NOD/SCID mice, causes a massive and transient dominant expansion of human B cells in the spleen. This permits the easy isolation of human monoclonal antibodies specific for different antigens by a Kohler and Milstein-based method. Here we studied the human HIV-specific antibody response in the circulation of mice after intrasplenic transfer of PBMC from untreated HIV-infected patients with detectable to high viral load as well as from HAART-treated and from untreated patients, who kept an undetectable viral load (the latter referred to as “natural suppressors”). Excellent B cell expansion was obtained for all PBMC. High level replication of virus was observed after transfer of PBMC of untreated viremic patients only. A strong and multispecific HIV-specific antibody response was observed after transfer of PBMC of untreated viremic patients and natural suppressors. In contrast, only a weak and pauci-specific antibody response was detected in mice reconstituted with PBMC from successfully treated patients. Based on these observations we conclude that the use of the intraspleen mouse model confirmed a) the presence of HIV-specific circulating memory B cells in untreated patients and natural suppressors; b) the nearly complete absence of circulating memory B cells in patients receiving highly active antiretroviral therapy. Using the intraspleen model we generated large numbers of immortalized B cells and isolated two anti-p24 human monoclonal antibodies. We further conclude that the intraspleen huPBL NOD/SCID model is a small animal model useful for the analysis of the antibody response against HIV found in patients.     

Replicative fitness of historical and recent HIV-1 isolates suggests HIV-1 attenuation over time

BACKGROUND: Changes in virulence during an epidemic are common among pathogens, but still unexplored in the case of HIV-1. Here we used primary human cells to study the replicative fitness of primary HIV-1 isolates from untreated patients, comparing historical (1986-1989) and recent samples (2002-2003). METHODS: Head-to-head dual virus infection/competition assays were performed in both peripheral blood mononuclear cells and human dendritic cell/T-cell co-cultures with pairs of 12 carefully matched historical and recent HIV-1 isolates from untreated patients. Sensitivity to inhibition by lamivudine (3TC) and TAK-779 of historical and recent R5 HIV-1 isolates was measured in a subset of samples. RESULTS: Overall, the historical HIV-1 out-competed the recent HIV-1 isolates in 176 of 238 competitions and in 9 of 12 competitions carefully matched for CD4 cell count. The mean relative replicative fitness (W) of all historical HIV-1 strains was significantly greater than that of recent HIV-1 isolates (W(1986-1989) = 1.395 and W(2002-2003) = 0.545, P < 0.001 (t test)). The more fit viruses (mean W > 1) from 1986-1989 appeared less sensitive to TAK-779 and 3TC than did the less fit (mean W < 1) 2002-2003 viruses. CONCLUSIONS: These findings suggest that HIV-1 replicative fitness may have decreased in the human population since the start of the pandemic. This 'attenuation' could be the consequence of serial bottlenecks during transmission and result in adaptation of HIV-1 to the human host.     

Neutralization of primary HIV-1 SF13 can be detected in extended incubation phase assays with sera from monkeys immunized with recombinant HIV-1 SF2 gp120

Phase III efficacy trials of recombinant human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins were postponed. In Phase I and II trials these candidate vaccines had failed to induce neutralizing antibodies against virus which had been isolated by co-culture with human peripheral blood mononuclear cells (PBMC). The aim of the present study was to determine assay conditions for detecting neutralization of primary HIV-1 isolates with sera from immunized individuals. We show that in two immunogenicity trials in rhesus macaques, recombinant HIV-1 SF2 gp120 induced antibodies which neutralized the primary HIV-1 SF13 isolate. Statistically significant in vitro neutralization required assays in which the incubation phase was extended. Sterile immunity was only seen with the highest level of neutralization, induced by a recombinant prime, peptide boost strategy. We recommend that neutralization assays with extended incubation phases should be used to monitor Phase III efficacy trials.