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Background: Strongyloides stercoralis is an intestinal nematode unique in its ability to replicate in the human host, allowing ongoing cycles of autoinfection, persisting for decades within the same host. Although usually asymptomatic, overwhelming infections can occur in Strongyloides and HTLV-1 co-infected individuals (SS/HTLV-1). Regulatory T cells (Tregs) are able to blunt specific Th2 responses necessary to control the parasite. We previously reported that peripheral blood Tregs are increased in SS/HTLV-1 and correlate with low Th2 responses. We hypothesized that Tregs are also increased at the site of infection in duodenal mucosa. Methods: Paraffin embedded duodenal biopsies were obtained from 10 SS/HTLV-1 patients, 3 controls with non-parasitic chronic duodenitis, and 2 healthy controls. Immunohistochemistry was performed using monoclonal antibodies against human CD3, CD8, IgE and FoxP3. The number of cells were counted using a conventional light microscope. The number of CD3+, CD8+, FP3+ and IgE positive cells per 0.35 mm2 was measured using ImagePro Plus software comparing areas adjacent or distant from parasite material. Results: In patients with SS/HTLV-1, T lymphocyte counts and CD8+ cells were lower in areas adjacent to the parasite compared to non-adjacent areas (CD3+: adjacent: 6.5 [Interquartile range (IQR: 2.8–12.3)]; non-adjacent: 24.5 [IQR: 20.9–34.4]; Mann-Whitney p = 0.0003; CD8+: adjacent: 4.5 [IQR: 2.3–11.8]; non-adjacent: 21 [IQR: 15.3–42.9]; Mann-Whitney p = 0.0011). Tregs cells in the intestines (FoxP3+ expressing cells) were increased in patients with SS/HTLV-1 compared with patients with chronic duodenitis (SS/HTLV-1: 1.5 [IQR: 0.7–2.3]; duodenitis controls: 0 [range 0–0.7]; healthy controls: 0; Mann-Whitney p = 0.034). There was also a trend towards fewer eosinophils adjacent to the parasites. Among SS/HTLV-1 patients the number of IgE expressing cells was increased for in areas not adjacent to the parasite compared to non-adjacent areas (ANOVA, p = 0.001). Conclusions: Our data shows increased Treg cell numbers localized adjacent to the parasites in the duodenum SS/HTLV-1 patients. In addition, other T lymphocytes and IgE expressing cells were decreased adjacent to the parasites, suggesting an important role for Tregs in down-regulating local parasite effector responses. [ABSTRACT FROM AUTHOR]

Regulatory T cells (Tregs) blunt uncontrolled immune responses. In advanced human immunodeficiency virus (HIV) infection, the total number of Tregs is decreased, but the proportion of T cells with a regulatory phenotype is highly variable. We studied CD4+CD25+FoxP3+ T cells from patients successfully treated with combination antiretroviral therapy (ART). The proportion of CD4+CD25+FoxP3+ cells transiently increased and then decreased from a median of 13% at baseline to 5.1% at 48 weeks, similar to values in normal subjects. These data suggest that with effective therapy, the regulatory cell numbers normalize, and that the inflammatory signals driving their production may also abate. [ABSTRACT FROM AUTHOR]

Background: Human strongyloidiasis varies from a chronic but limited infection in normal hosts to hyperinfection in patients treated with corticosteroids or with HTLV-1 co-infection. Regulatory T cells dampen immune responses to infections. How human strongyloidiasis is controlled and how HTLV-1 infection affects this control are not clear. We hypothesize that HTLV-1 leads to dissemination of Strongyloides stercoralis infection by augmenting regulatory T cell numbers, which in turn down regulate the immune response to the parasite. Objective: To measure peripheral blood T regulatory cells and Strongyloides stercoralis larval antigen-specific cytokine responses in strongyloidiasis patients with or without HTLV-1 co-infection. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from newly diagnosed strongyloidiasis patients with or without HTLV-1 co-infection. Regulatory T cells were characterized by flow cytometry using intracellular staining for CD4, CD25 and FoxP3. PBMCs were also cultured with and without Strongyloides larval antigens. Supernatants were analyzed for IL-5 production. Results: Patients with HTLV-1 and Strongyloides co-infection had higher parasite burdens. Eosinophil counts were decreased in the HTLV-1 and Strongyloides co-infected subjects compared to strongyloidiasis-only patients (70.0 vs. 502.5 cells/mm3, p = 0.09, Mann-Whitney test). The proportion of regulatory T cells was increased in HTLV-1 positive subjects co-infected with strongyloidiasis compared to patients with only strongyloidiasis or asymptomatic HTLV-1 carriers (median = 17.9% vs. 4.3% vs. 5.9 p,0.05, One-way ANOVA). Strongyloides antigen-specific IL-5 responses were reduced in strongyloidiasis/HTLV-1 coinfected patients (5.0 vs. 187.5 pg/ml, p = 0.03, Mann-Whitney test). Reduced IL-5 responses and eosinophil counts were inversely correlated to the number of CD4+CD25+FoxP3+ cells. Conclusions: Regulatory T cell counts are increased in patients with HTLV-1 and Strongyloides stercoralis co-infection and correlate with both low circulating eosinophil counts and reduced antigen-driven IL-5 production. These findings suggest a role for regulatory T cells in susceptibility to Strongyloides hyperinfection. [ABSTRACT FROM AUTHOR]

Abstract: Flow cytometry provides accurate relative cellular quantitation (percent abundance) of cells from diverse samples, but technical limitations of most flow cytometers preclude accurate absolute quantitation. Several quantitation standards are now commercially available which, when added to samples, permit absolute quantitation of CD4+ T cells. However, these reagents are limited by their cost, technical complexity, requirement for additional software and/or limited applicability. Moreover, few studies have validated the use of such reagents in complex biological samples, especially for quantitation of non-T cells. Here we show that addition to samples of known quantities of polystyrene fluorescence standardization beads permits accurate quantitation of CD4+ T cells from complex cell samples. This procedure, here termed single bead-enhanced cytofluorimetry (SBEC), was equally capable of enumerating eosinophils as well as subcellular fragments of apoptotic cells, moieties with very different optical and fluorescent characteristics. Relative to other proprietary products, SBEC is simple, inexpensive and requires no special software, suggesting that the method is suitable for the routine quantitation of most cells and other particles by flow cytometry. [Copyright &y& Elsevier]

Regulatory T-cells (Tregs) are increased in patients with HTLV-1/ Strongyloides stercoralis co-infection, and they may modify otherwise protective antigen-specific cytokine production. We hypothesized that effective anti-helminthic treatment would decrease Tregs and restore antigen-specific cytokine responses. We enrolled 19 patients with Strongyloides larvae in their stool by Baerman's test. Six were positive and 13 negative for antibody to HTLV-1 by ELISA, with positive tests confirmed by immunoblot. Before treatment, co-infected subjects had higher Tregs percentages and lower antigen-stimulated IL-5 levels compared to subjects with Strongyloides without HTLV-1. All patients were treated with ivermectin. After effective treatment, Tregs percentages decreased in patients with HTLV-1; however, antigen-specific IL-5 production remained blunted in co-infected subjects. These results suggest that treating strongyloidiasis infection decreases circulating Tregs, but antigen-specific cytokine remains altered. This may reflect blunting of sensitization by Tregs. [ABSTRACT FROM AUTHOR]