DC modulation via PRR signaling
The capacity to recognize and distinguish different classes of pathogens is of critical importance for the initiation of appropriate immune responses. For this purpose, DC express PRR, which recognize and differentiate between different pathogen-derived molecules, the so-called PAMP. One of the most thoroughly studied classes of PRR are the TLR, which have primarily been implicated in the priming of pro-inflammatory/Th1 responses by DC in response to PAMP from bacterial or viral origin 6. Nonetheless, some helminth products have also been shown to prime Th2 or regulatory responses through ligation of TLR. For instance, excretory/secretory-62 (ES-62), a phosphorylcholine-containing glycoprotein of the nematode Acanthocheilonema viteae, conditions DC to induce Th2 responses through TLR4 7, 8. Moreover, the glycoconjugate LNFPIII, carrying a Lewis-X (Lex) carbohydrate epitope found in schistosoma soluble egg antigens (SEA), has been implicated in TLR4-dependent priming of Th2 responses via DC 9. However, SEA is also known to be capable of modulating DC for Th2 priming in the absence of TLR signaling 10. Furthermore, phosphatidylserine (PS) lipids derived from schistosomes and ascaris worms, which carry TLR2- activating molecules, have been shown to promote Th2 responses via DC, but this is not TLR2 dependent 11, whereas mono-acetylated PS lipids from schistosomes were found to specifically instruct DC to preferentially induce IL-10-producing Treg in a TLR2-dependent fashion12. Although TLR2 does not seem to be essential for Th2 polarization by schistosomes in vivo10, 13, there is evidence that TLR2 plays an important role in the induction of Treg responses during natural infection 14. Finally, double-stranded RNA from schistosome eggs has been implicated in the activation of DC via TLR3, resulting in a Th1-polarized response 13, 15.
Apart from TLR, a group of carbohydrate-recognizing PRR, the C-type lectins (CLR) have been shown to play an important role in the sensing of helminth glycans by DC. For instance, SEA, which contains glycoproteins, is recognized and internalized by human DC in a DC-specific ICAM-3-grabbing nonintegrin (DC-SIGN)-, mannose receptor- and macrophage galactose-type lectin-dependent manner 16, 17. Antigen preparations of other stages of the schistosome life cycle have also been shown to interact with DC-SIGN 18. Binding of SEA to DC-SIGN is dependent on the sugar motifs Lex and LDN-F 17, while chemical modification of the glycans present in SEA abolishes the Th2-driving capacity of SEA 19. This, together with the observation that Lex-containing LNFPIII favors Th2-biased responses 20, suggests that CLR play an important role in conditioning DC for induction of Th2 responses by schistosomal antigens. Moreover, antigens from Toxocara canis were found to be recognized by DC-SIGN expressed on DC 21, and the induction of a Th2 response in vivo by antigens of the parasitic nematode Brugia malayi, as well as the free-living nematode Caenorhabditis elegans, was found to be dependent on intact glycans 22. These findings together suggest that certain helminth glycans may serve as PAMP that instruct DC via CLR to drive Th2-polarized responses.
Finally, there is evidence that the class A scavenger receptor, which is a member of a family of receptors that bind chemically modified low-density lipoproteins, can function as PRR (reviewed in 23), and mediate recognition of helminth components that result in the induction of Th2-polarized responses. It was found that calreticulin, a secreted protein expressed by tissue-invasive larvae of the gastrointestinal helminth Heligmosomoides polygyrus, binds class A scavenger receptor on DC and has the capacity in the absence of adjuvants to predominantly induce IL-4 production in vivo24.
In contrast to the prevailing view of DC activation by microbial ligands via PRR, DC primed by helminth products often fail to show signs of classical maturation 25. The absence of DC maturation is in-line with several studies in which stimulation of p38 MAPK, a signaling molecule crucial for PRR-mediated DC activation 26, was not observed in DC exposed to helminth products such as SEA, ES-62 or LNFPIII 9, 27–29. Instead, these helminth-derived components preferentially induce the activation of ERK MAPK, in the case of SEA and ES-62, or NF-κB1 and ERK, in the case of LNFPIII. Signaling through ERK in DC has been shown to result in suppression of IL-12 and induction of IL-10 expression, in line with the observation that ERK−/− mice are prone to develop autoimmunity. As a result, ERK has been implicated in conditioning DC for Th2 priming 11, 30, 31. Likewise, signaling through NF-κB1 appears to be important for priming of DC for Th2 polarization, since both SEA- 32 and LNFPIII-pulsed NF-κB1-deficient DC 29 are incapable of inducing a Th2 response (Fig. 1).
Figure 1. Molecular mechanisms through which DC become conditioned by helminth products via signaling-dependent and signaling-independent pathways for priming of Th2 responses. Helminth-derived molecules condition DC for induction of Th2 polarization through interactions with PRR, which in signaling-dependent fashion induce the expression of Th2-promoting molecules while suppressing the expression of Th1-polarizing factors. In addition, helminth-derived molecules may favor induction of Th2 responses by DC by suppressing antigen presentation, costimulation and/or expression of Th1-promoting cytokines by directly interfering with these pathways in a signaling-independent fashion.
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A feature shared by many helminths is their capacity to suppress TLR-mediated DC activation by microbial PAMP. Numerous studies have reported the inhibitory effects of helminth-derived components on TLR-induced activation as determined by pro-inflammatory cytokine production and expression of MHC class II/costimulatory molecules 28, 33–39. The pathways underlying this suppression are, however, still poorly understood. Interestingly, the suppression of TLR-mediated responses by helminth antigens has striking similarities with the effects induced by several microbial pathogens that target DC-SIGN 40–42. Recently, significant advances have been made in the identification of pathways downstream of DC-SIGN that result in the modulation of TLR signaling. Lex has been found to modulate, via leukocyte specific protein-1, LPS-induced signaling, by elevating IL10 and reducing IL-12 43. As SEA also alters LPS responses by enhancing IL-10 and reducing IL-12 secretion, it is possible that these signaling pathways play a role in DC conditioning by schistosomes; however, since not all helminth-derived modulatory molecules contain or are glycans, it is reasonable to assume that exploiting CLR to modulate DC function is just one of the ways through which helminths exert their effects on DC activation.
DC modulation through PRR-signaling independent mechanisms
Evidence is emerging that helminth manipulation of DC may also be mediated by mechanisms other than receptor-mediated signaling events, for example, by the enzymatic activities of helminth-derived products. Helminth parasites are known to release a wide variety of enzymatically active products that are thought to play an important role in establishing and maintaining infection by contributing to the degradation of soluble anti-parasitic molecules or the impairment of innate immune cells (reviewed in 44).
With regard to the effects of such molecules on DC, omega-1, a glycosylated RNase secreted by schistosome eggs and present in SEA, was recently found to drive Th2 responses via functional modulation of DC 37, 45. Interestingly, omega-1 could modulate DC function in vitro with characteristics similar to those of SEA, while depletion of omega-1 from SEA abrogated to a large extent its potential to modulate DC function as determined by its capacity to suppress DC LPS-induced IL-12 secretion, expression of maturation markers, and its ability to drive Th2 polarization. In addition, chemical inactivation of omega-1's RNase activity through DEPC treatment led to a significant attenuation of Th2 polarization by omega-1. This suggests that although, as discussed earlier (see DC modulation via PRR signaling), sugars present in SEA may modulate DC function for Th2 priming, the RNase activity present in SEA may also be very important for conditioning DC to drive Th2 responses. Although omega-1 is the first RNase from helminths to be described to induce Th2, other RNases have been linked to Th2 responses. For instance, the allergen Aspf-1, which is an RNase 46, as well as eosinophil-derived neutrotoxin, an RNase from eosinophils, have been found to drive Th2 responses via DC 47. Interference with translation would be in line with reports that several plant-derived enzymes that inactivate ribosomes, so-called ribotoxins, have been identified as allergens, thereby linking Th2 responses with the inhibition of protein synthesis 48, 49.
Furthermore, a number of studies have documented the potent suppressory effects of cystatins, a class of molecules expressed by filarial nematodes, on host immune responses 50–52. This is thought to reflect their capacity to interfere with antigen presentation by DC 53, 54, through blocking the host cysteine protease activity required for the removal of the invariant chain that is necessary for peptide loading onto MHC class II. Thus, rather than promoting Th2 or Treg by DC, cystatins seem to suppress the capacity of DC to prime T-cell responses in general.
Finally, helminth pathogens express cysteine proteases, termed cathepsins, that lead to immune deviation by suppressing Th1 immunity 55. Whether helminth proteases can mediate these effects by targeting DC is at the moment unclear; however, based on the observations that numerous allergens are known to be cysteine proteases 56 and that the cysteine protease Der p 1, one of the major allergens of the house dust mite Dermatophagoides pteronyssinus, has been found to prime monocyte-derived DC for Th2 polarization in a protease dependent manner 57, it is conceivable that such helminth-derived proteases have the potential to favor Th2 polarization through functional modulation of DC.
Taken together, these studies illustrate that DC can become conditioned via PRR signaling dependent (see DC modulation via PRR signaling) and independent mechanisms (this section) by helminth products that include carbohydrates, lipids and proteins or a combination of these, with or without enzymatic activities. A schematic overview of the events leading to DC modulation is shown in Fig. 1, with a summary of helminth-derived components that modulate DC in Table 1. Nonetheless, this wide spectrum of helminth products shares important features, in that the products generally fail to induce conventional DC maturation and suppress activation by pro-inflammatory PAMP, which results in the impairment of Th1 development and a bias of the immune response toward Th2 or Treg. This observation, together with the fact that helminth products are recognized by DC via receptors that also recognize Th1- or Th17-inducing microbial products, suggests that it may not be engagement of the type of receptor itself, but the anti-inflammatory signaling resulting in a muted DC activation profile that sets helminth-derived molecules apart from their microbial counterparts and enables DC to induce Th2- or Treg-polarized immune responses.
Table 1. Helminth-derived antigens/factors that modulate DC function
|Group||Species||Component||PRR||Signaling or mode of action||Resulting DC phenotype||T-cell polarization||Refs.|
|Trematodes||Fasciola hepatica||Tegumental antigen||TLR independent|| NF-κB p65|| IL-12,||Th1||36|
| || || || || || IL-10|| || |
| ||Schistosoma mansoni||LNFPIII||TLR4|| NF-κB1||Low IL-12||Th2||9, 20, 29|
| || || || || ERK|| || || |
| || ||SEA||DC-SIGN, MR, MGL|| ERK,|| IL-12,||Th2||16, 17|
| || || || ||c-fos|| IL-10|| || |
| || || || || || OX40-L|| || |
| || ||Egg ES||CLR?|| ERK|| IL-12,||Th2||37, 45 unpublished|
| || || || || || IL-10|| || |
| || ||omega-1||CLR?||Interference with translation?|| IL-12,||Th2||37, 45|
| || || || || || Ag presentation|| || |
| || ||PS lipids||TLR2|| p38|| IL-12||Th2||11, 12|
| || ||Lyso-PS lipids||TLR2||ND|| IL-12||Treg (Tr1)||12|
| || ||double stranded RNA||TLR3||STAT-1, ISG||IFNα||Th1||13, 15|
| || ||0–3 h cercarial ES||DC-SIGN||ND||Low IL-12||Th2||18, 127|
|Cestodes||Echinococcus granulosus||AgB||TLR?|| IRAK1|| IL-12||Th2||39|
| || || || || NF-κB1|| CD40|| || |
|Nematodes||Acanthocheilonema viteae||ES-62||TLR4|| ERK||Low IL-12||Th2||8, 27, 128|
| ||Ascaris lumbricoides||PS lipids||TLR2|| p38|| IL-12||Th2||11|
| || || || || || IL-10|| || |
| ||Heligmosomoides polygyrus||calreticulin||SR-A signaling?||ND||ND||Th2||24|
| || ||ES||ND||ND|| IL-12||Treg||34, 61|
| || || || || || IL-10|| || |
| ||Nippostrongylus brasiliensis||NES||ND||ND|| IL-12|| ||33, 61|
| || || || || || OX40-L|| || |
| || || || || || CD86|| || |
|All||Multiple helminth species||Cathepsins||Cleavage of specific proteins?||ND||ND||Th1||55, 129|
| || ||Cystatins||ND|| peptide loading on MHC||Ag presentation|| T cell activation||50–52, 53, 54|