Urban-rural differences in the gene expression profiles of Ghanaian children.

Recent studies indicate that urbanization is having a pronounced effect on disease patterns in developing countries. To understand the immunological basis of this, we examined mRNA expression in whole blood of genes involved in immune activation and regulation in 151 children aged 5-13 years attending rural, urban low socioeconomic status (SES) and urban high-SES schools in Ghana. Samples were also collected to detect helminth and malaria infections. Marked differences in gene expression were observed between the rural and urban areas as well as within the urban area. The expression of both interleukin (IL)-10 and programmed cell death protein 1 increased significantly across the schools from urban high SES to urban low SES to rural (P-trend <0.001). Although IL-10 gene expression was significantly elevated in the rural compared with the urban schools (P<0.001), this was not associated with parasitic infection. Significant differences in the expression of toll-like receptors (TLRs) and their signaling genes were seen between the two urban schools. Genetic differences could not fully account for the gene expression profiles in the different groups as shown by analysis of IL-10, TLR-2 and TLR-4 gene polymorphisms. Immune gene expression patterns are strongly influenced by environmental determinants and may underlie the effects of urbanization seen on health outcomes.

Schistosome infection is negatively associated with mite atopy, but not wheeze and asthma in Ghanaian schoolchildren.

BACKGROUND: Epidemiological evidence suggests that helminth infection and rural living are inversely associated with allergic disorders. OBJECTIVE: The aim of the study was to investigate the effect of helminth infections and urban versus rural residence on allergy in schoolchildren from Ghana. METHODS: In a cross-sectional study of 1385 children from urban-high socio-economic status (SES), urban-low SES and rural schools, associations between body mass index (BMI), allergen-specific IgE (sIgE), parasitic infections and allergy outcomes were analysed. Allergy outcomes were skin prick test (SPT) reactivity, reported current wheeze and asthma. RESULTS: Helminth infections were found predominantly among rural subjects, and the most common were hookworm (9.9%) and Schistosoma spp (9.5%). Being overweight was highest among urban-high SES (14.6%) compared to urban-low SES (5.5%) and rural children (8.6%). The prevalence of SPT reactivity to any allergen was 18.3%, and this was highest among rural children (21.4%) followed by urban-high SES (20.2%) and urban-low SES (10.5%) children. Overall, SPT reactivity to mite (12%) was most common. Wheeze and asthma were reported by 7.9% and 8.3%, respectively. In multivariate analyses, factors associated with mite SPT were BMI (aOR 2.43, 95% CI 1.28-4.60, P = 0.007), schistosome infection (aOR 0.15, 95% CI 0.05-0.41) and mite sIgE (aOR 7.40, 95% CI 5.62-9.73, P < 0.001) but not area. However, the association between mite IgE and SPT differed by area and was strongest among urban-high SES children (aOR = 15.58, 95% CI 7.05-34.43, P < 0.001). Compared to rural, urban-low SES area was negatively associated with current wheeze (aOR 0.41, 95% CI 0.20-0.83, P = 0.013). Both mite sIgE and mite SPT were significantly associated with current wheeze and asthma. CONCLUSION AND CLINICAL RELEVANCE: Infection with schistosomes appeared to protect against mite SPT reactivity. This needs to be confirmed in future studies, preferably in a longitudinal design where schistosome infections are treated and allergic reactions reassessed.

Application of a circulating-cathodic-antigen (CCA) strip test and real-time PCR, in comparison with microscopy, for the detection of Schistosoma haematobium in urine samples from Ghana.

In the detection of parasitic infection, the traditional methods based on microscopy often have low sensitivity and/or specificity compared with the newer, molecular tests. An assay based on real-time PCR and a reagent strip test for detecting circulating cathodic antigen (CCA) have both now been compared with urine filtration and microscopy, in the detection of Schistosoma haematobium infections. Urine samples, obtained from 74 'cases' in areas of Ghana with endemic S. haematobium and 79 'controls' from non-endemic areas, were each checked using the three methods. With the results of the filtration and microscopy taken as the 'gold standard', real-time PCR was found to be 100% specific and 89% sensitive whereas the CCA strips were 91% specific and 41% sensitive. With the samples found to contain > or =50 eggs/10 ml (indicating relatively intense infections), the sensitivities of the PCR and CCA were higher, at 100% and 62%, respectively. As expected, egg counts were negatively correlated with the number of amplification cycles needed, in the PCR, to give a signal that exceeded the background (r=-0.38; P<0.01). Although the real-time PCR and CCA strip tests are very different, both show promise in the detection of S. haematobium infections. The PCR has optimal specificity and high sensitivity but the specificity of the CCA strips and the sensitivity of both tools could still be improved. A more thorough re-evaluation of the sensitivity and specificity of microscopy and these newer diagnostic methods, with an estimation of the cost-effectiveness of each technique, is recommended.

Cellular and humoral responses to tetanus vaccination in Gabonese children.

Protection to tetanus is often not optimal in developing countries due to incomplete vaccination schemes, or decreased efficacy of vaccination. In this study we investigated the immunological response to tetanus booster vaccination in school children living in a semi-urban or in a rural area of Gabon. Tetanus-specific total IgG as well as antibody subclasses of the IgG1, IgG2, IgG3 and IgG4 isotype and the avidity of the dominating IgG1 subclass were determined both before and 1 month after the booster vaccination. In addition, tetanus-specific cytokine responses were determined. We found a polarization towards a T helper 1 (Th1) profile in the semi-urban children, whereas the cytokine responses of the rural children showed a T helper 2 (Th2) skewed response. Furthermore, tetanus-specific antibodies of the different IgG subclasses were all increased upon a tetanus booster vaccination and levels of IgG1 and IgG3 were higher in the rural children. In conclusion, a tetanus booster vaccination induced a stronger Th2 over Th1 cytokine profile to tetanus toxoid (TT) in rural children who showed the highest levels of IgG1 and IgG3 anti-TT antibody responses.

Cellular and humoral responses to influenza in gabonese children living in rural and semi-urban areas.

BACKGROUND: With the current attention to the pandemic threat of avian influenza viruses, it is recognized that there is little information on influenza in Africa. In addition, the effects of influenza vaccination in African countries could be very different from the effects in regions with less exposure to microorganisms and parasites. METHODS: To monitor the presence of influenza viruses and investigate the immunological responses to influenza vaccination, schoolchildren in semi-urban and rural regions of Gabon were studied. Influenza-specific antibody responses to the 3 strains represented in the vaccine were determined in the serum. Furthermore, cytokine responses were measured after in vitro stimulation of whole blood by influenza antigens, before and after vaccination. RESULTS: Prevaccination titers of antibody against H3N2 were high. At vaccination, the titers of antibody against the 3 influenza strains increased significantly. The anti-H1N1 and anti-B responses after vaccination were weaker in rural schoolchildren than in semi-urban schoolchildren. Influenza-specific cytokine responses were induced within a week, showing significantly lower interferon- gamma and significantly higher interleukin-5 in the children from rural areas. CONCLUSIONS: Prevaccination antibody levels indicated that influenza viruses circulate in Gabon. Altogether, influenza vaccination induces weaker immune responses in a rural population than in a semi-urban population of Gabonese schoolchildren.

Co-infection of helminths and malaria: modulation of the immune responses to malaria.

Chronic helminth infections induce strong type 2 and regulatory immune responses and are known to influence immune activity to other antigens such as allergens and vaccines. Since malaria and helminth infections often coincide geographically in the same tropical regions, the question arises whether helminth infections modulate the immune responses towards the malaria parasite and affect its course of disease. Here, we will review studies on co-infections in both animal models and in human populations, and discuss the changes in the immune system seen. Furthermore, the implications of helminth infection for the efficacy of malaria vaccines will be discussed.

BLC (CXCL13) is expressed by different dendritic cell subsets in vitro and in vivo.

Dendritic cells (DC) attract both T and B lymphocytes to induce an efficient antigen-specific immune response. Recently, it was shown that naïve T cells are attracted to DC by dendritic cell chemokine 1 (DC-CK1, CCL18). The potent B lymphocyte chemoattractant BLC (CXCL13) was previously shown to be essential for homing of lymphocytes into secondary lymphoid organs and for the development of B cell follicles. As the cells that produce BLC are largely unknown and BLC could be a candidate chemokine for the recruitment of B cells to DC, we analyzed different DC subsets for expression of BLC. Here we demonstrate that monocyte-derived DC as well as activated blood DC indeed express and secrete BLC. Interestingly, ligation of the CD40 molecule down-regulated BLC expression in monocyte-derived DC. Staining of tonsilar sections indicated that BLC is expressed by follicular dendritic cells and germinal center dendritic cells (GCDC) in vivo. Real-time quantitative PCR confirmed the expression of BLC in isolated GCDC. Since both B cells and activated T cells express the receptor for BLC, our findings implicate an important role for BLC in establishing the interaction of DC with T cells and B cells. Furthermore, CD40/CD40 ligand interactions could modulate this process by down-regulating the expression of BLC.

Quantitative analysis of chemokine expression by dendritic cell subsets in vitro and in vivo.

Upon maturation, dendritic cells (DCs) have to adjust their chemokine expression to sequentially attract different leukocyte subsets. We used real-time quantitative polymerase chain reaction analysis to study in detail the expression of 12 chemokines involved in the recruitment of leukocytes into and inside secondary lymphoid organs, by DCs in distinct differentiation stages, both in vitro and in vivo. Monocyte-derived immature DCs expressed high levels of DC chemokine 1 (DC-CK1), EBI1-ligand chemokine (ELC), macrophage-derived chemokine (MDC), macrophage-inflammatory protein (MIP)-1alpha, and thymus and activation-regulated chemokine (TARC). Upon maturation, DCs up-regulated the expression of DC-CK1 (60-fold), ELC (7-fold), and TARC (10-fold). Activation of DCs by CD40 ligand further up-regulated the expression of ELC (25-fold). We found that freshly isolated blood DCs expressed only low levels of interleukin-8, lymphotactin, and MIP-1alpha. It is interesting that the chemokine profile expressed by activated CD11c(-) lymphoid-like as well as CD11c(+) myeloid blood DCs mimics that of monocyte-derived DCS: Additionally, purified Langerhans cells that had migrated out of the epidermis expressed a similar chemokine pattern. These data indicate that different DC subsets in vitro and in vivo can express the same chemokines to attract leukocytes.

The dendritic cell-specific CC-chemokine DC-CK1 is expressed by germinal center dendritic cells and attracts CD38-negative mantle zone B lymphocytes.

DC-CK1 (CCL18) is a dendritic cell (DC)-specific chemokine expressed in both T and B cell areas of secondary lymphoid organs that preferentially attracts CD45RA(+) T cells. In this study, we further explored the nature of DC-CK1 expressing cells in germinal centers (GCs) of secondary lymphoid organs using a newly developed anti-DC-CK1 mAb. Immunohistochemical analysis demonstrated a remarkable difference in the number of DC-CK1 expressing cells in adjacent GCs within one tonsil, implicating that the expression of DC-CK1 in GCs depends on the activation and/or progression stage of the GC reaction. Using immunohistology and RNA analysis, we demonstrated that GCDC are the source of DC-CK1 production in the GCs. Considering the recently described function of GCDC in (naive) B cell proliferation, isotype switching and Ab production, we investigated the ability of DC-CK1 to attract B lymphocytes. Here we demonstrate that DC-CK1 is a pertussis toxin-dependent chemoattractant for B lymphocytes with a preference in attracting mantle zone (CD38(-)) B cells. The findings that GCDC produce DC-CK1 and attract mantle zone B cells support a key role for GCDC in the development of GCs and memory B cell formation.

DC-STAMP, a novel multimembrane-spanning molecule preferentially expressed by dendritic cells.

Dendritic cells (DC) are unique in their ability to present antigen to naive T cells, and therefore play a central role in the initiation of immune responses. Characterization of DC-specific genes may help to unravel the mechanism underlying their potent antigen presenting capacity. Here we describe the identification of a novel transcript, isolated by random sequencing of a cDNA library prepared from monocyte-derived DC, which we termed DC-specific transmembrane protein (DC-STAMP). DC-STAMP is specifically expressed by DC, and not in a panel of other leukocytes or non-hematopoietic cells. Interestingly, DC-STAMP was also detected in activated but not resting blood DC. The DC-STAMP transcript encodes a 470-amino acid protein containing seven putative transmembrane domains. Expression of a DC-STAMP-GFP fusion protein in 293 cells indicates that DC-STAMP is expressed at the cell surface, and has an intracellular C terminus. Surprisingly, no sequence homology was found with any other protein or multimembrane-spanning receptor. Therefore, we propose that DC-STAMP is a novel DC-specific multimembrane-spanning protein, representing a new group of transmembrane proteins.

Analysis of dendritic cells at the genetic level.

To increase our understanding of dendritic cell (DC) function we have used two approaches to search at the genetic level for molecules which are specifically expressed by these cells. First, we have performed random sequencing of cDNA libraries prepared from DC. Second, we have employed differential display PCR (DD-PCR). DD-PCR is a powerful technique for the identification at the RNA level of molecules which are expressed in a cell type-specific manner. In our study, we have compared RNA from DC with RNA from a panel of leukocyte cell lines. Here we present a summary of our findings using these two approaches, and show that both methods are complementary and can be used to identify molecules that are specific to DC.