IL-17 and IL-22 are associated with protection against human kala azar caused by Leishmania donovani - PubMed (original) (raw)
. 2009 Aug;119(8):2379-87.
doi: 10.1172/JCI38813. Epub 2009 Jul 13.
Audrey Romano, Sandrine Cabantous, Sandrine Henri, Awad Hammad, Bouréma Kouriba, Laurent Argiro, Musa el Kheir, Bruno Bucheton, Charles Mary, Sayda Hassan El-Safi, Alain Dessein
Affiliations
- PMID: 19620772
- PMCID: PMC2719936
- DOI: 10.1172/JCI38813
IL-17 and IL-22 are associated with protection against human kala azar caused by Leishmania donovani
Maira G R Pitta et al. J Clin Invest. 2009 Aug.
Abstract
IL-17 and IL-22 have been shown to increase protection against certain bacteria and fungal pathogens in experimental models. However, no human studies have demonstrated a crucial role of IL-17 and IL-22 in protection against infections. We show here that Leishmania donovani, which can cause the lethal visceral disease Kala Azar (KA), stimulates the differentiation of Th17 cells, which produce IL-17, IL-22, and IFN-gamma. Analysis of Th1, Th2, and Th17 cytokine responses by cultured PBMCs from individuals in a cohort of subjects who developed KA or were protected against KA during a severe outbreak showed that IL-17 and IL-22 were strongly and independently associated with protection against KA. Our results suggest that, along with Th1 cytokines, IL-17 and IL-22 play complementary roles in human protection against KA, and that a defect in Th17 induction may increase the risk of KA.
Figures
Figure 1. Induction of IL-17 and IL-22 by L. donovani in cultures of PBMCs from healthy subjects.
(A) The production of IL-17 and IL-22 in cultures of PBMCs from healthy donors required priming with whole L. donovani parasites. Boosting was achieved using parasites or parasite extracts. Priming (on day 0; stimulation 1) and boost (on day 7; stimulation 2) were performed with opsonized (OpsLd) or non-opsonized (Ld) heat-killed L. donovani parasites or parasite extracts (Ld-ext). IL-17 and IL-22 were quantified by ELISA on days 7, 11, and 15. Two representative experiments (Exp 1 and Exp 2) from a series of 10 are shown. (B) IL-17 and IL-22 were produced by CD4+ T cells. CD4+ T cells were purified from the 12-day cultures of PBMCs primed and boosted with OpsLd. Purified CD4+ T cells were then stimulated for 48 hours with PMA plus ionomycin, concanavalin A (ConA), or phytohemagglutinin (PHA). IL-17 and IL-22 were quantified by ELISA. Three independent experiments with cells from different donors are shown. (C) Phenotypic characterization of IL-17+CD4+ T cells in cultures of normal PBMCs stimulated with OpsLd. Normal PBMCs were primed and boosted with OpsLd. Five days after the second stimulation, cells were stimulated with PMA plus ionomycin plus monensin for 5 hours. The x axis represents the fluorescence intensity after either CD4 labeling or IFN-γ labeling. The y axis corresponds to IL-17 labeling. Horizontal and vertical bars define the upper fluorescence intensity observed with labeled control cells. The numbers in each quadrant represent the percentage of cells in that quadrant relative to the total number of cells included in the analysis.
Figure 2. L. donovani induces the production of cytokines that are required for Th17 induction and maintenance.
IL-6, IL-23, and IL-1β production in IL-17– and IL-22–producing cultures of PBMCs (A) and monocytes (B) from healthy blood donors. PBMCs were primed (day 0) and boosted with OpsLd (day 7), as described in the legend of Figure 1. Monocytes were stimulated once with living L. donovani that were opsonized with either HIS or NHS. Cytokine levels were determined at the time points indicated. Results represent the arithmetic means of data from 4 independent experiments with PBMCs and 4 experiments with monocytes. The effects of the stimulation and culture duration were tested by linear regression (P < 0.05 was considered significant). Stimulations had a significant effect on IL-17 (P = 0.003), IL-22 (P = 0.05), IL-6 (P < 0.001), and IL-1β (P < 0.001) in cultures of PBMCs and on IL-23 (P = 0.05), IL-6 (P = 0.02), and IL-1β (P < 0.01) in cultures of monocytes. Culture duration had a significant effect (P < 0.01) in all cultures excepted for IL-1β (PBMCs and monocytes) and IL-23 (monocytes). Data are means ± SEM.
Figure 3. Resistance to KA is associated with enhanced IL-17 and IL-22 responses.
(A) IL-17, IL-22, and Th1 and Th2 cytokine production by PBMCs from gp1, gp2, and gp3 subjects. Cytokines were quantified by ELISA in supernatants of PBMCs stimulated with 10 μg/ml of L. donovani extracts, as indicated in the Methods. IL-4 was undetectable or present in small amounts in all cultures. The data presented are the arithmetic means ± SEM. Cytokine scales (y axes) were set to allow for a 5-fold increase in the mean levels above those of the group with the lowest levels to be represented. (B) Resistance to KA was associated with high levels of both IL-22 and IL-17, whereas KA was associated with low levels of these cytokines. IL-17 and IL-22 levels were each assigned to 3 classes of equal size. The 3 classes for IL-17 were ≤15, 16–120, and 121–1,250 pg/ml. The 3 classes for IL-22 were <190, 190–890, and 900-40,000 pg/ml. Dark bars represent subjects with the lowest IL-17 and IL-22 levels. Data analysis is presented in Table 3.
Figure 4. IL-23, IL-6, and IL-1β production in cultures of PBMCs from subjects from the 3 clinical groups.
(A) IL-6 and IL-1β levels were higher in cultures of PBMCs from gp1 subjects than in those of cells from gp2 subjects, whereas no difference was observed for IL-23. Cultures were stimulated as described in Figure 2A. (B) PBMCs from KA subjects produced small amounts of IL-17 and IL-22, even in the presence of normal to high levels of the regulatory cytokines IL-6, IL-1β, and IL-23. IL-23, IL-6, and IL-1β levels in cultures (from gp1 and gp2) were assigned to 3 classes of equal size, and IL-17 and IL-22 levels are presented for each cytokine class. A nonparametric Kruskal-Wallis test showed a significant increase of IL-17 (P < 0.03) in increasing IL-23, IL-6, and IL-1β classes in cultures of PBMCs from resistant subjects. No significant increase of IL-17 was observed with increasing concentrations of IL-6 and IL-1β in cultures of PBMCs from gp2 subjects, whereas IL-17 did increase (P < 0.001) with increasing IL-23 concentrations in the same cultures. IL-22 increased (P < 0.05) with increasing IL-23, IL-6, and IL-1β concentrations in cultures of PBMCs from gp1 and gp2 subjects. The data presented are the arithmetic means ± SEM.