Good news is on the way for the unlucky ones who've suffered from an itchy rash after an encounter with poison ivy. Researchers have made significant progress in discovering the underlying mechanisms that mediate the reaction to irritant agents such as poison ivy. In fact, poison ivy is one of many agents that produce a response that falls under the classification of allergic contact dermatitis (ACD), or contact hypersensitivity (CHS). However, to truly appreciate this relevant research it's important to understand a bit about CHS.
CHS is essentially a hypersensitive immune response to reactive molecules that bind to your skin (1). All immune hypersensitivities (not just CHS) occur in two phases. In the primary phase, the sensitization stage, a normal primary immune response is mounted to a pathogen. The effector stage consists of a hypersensitive or excessive response to that same pathogen upon subsequent exposure (1). These pathogens can be therefore be referred to as allergens. This type of allergic reaction is mediated by T cells (a type of white blood cell that helps defend against pathogens) and can be caused by a variety of molecules. However, it's important to note that dendritic cell (DC) activation must occur before a T cell response can be mounted. DCs are the immune cells that absorb invading pathogens and subsequently present proteins derived from that pathogen to other immune cells, such as T cells, so that an immune response can occur. Ultimately, this reaction results in excessive inflammation of the skin, as demonstrated by the aforementioned rash. Current research examines exactly how these DCs are activated in the first place.
Research has discovered that the purinergic receptor, P2X7, a ligand-gated ion channel expressed on DCs that has been shown to play an important role in activating T cells. Specifically, it's been shown that ATP released from skin cells is capable of activating this receptor (2)and causing the release of IL-1β (3), a molecule important for the sensitization process mentioned earlier. This process also involves the NLRP3 inflammasome (a structure within the DC) though the exact mechanism is unclear. In Weber et al. 2010, the role of this ATP receptor, P2X7, in CHS and its specific mechanism of action is illuminated.
Initially, researchers wanted to reinforce the role of P2X7 in CHS. Therefore, CHS was induced in mice by applying contact allergens, such as TNCB and oxazalone, to the ear skin. The intensity of the TNCB-induced CHS was analyzed by measuring ear thickness caused by the swelling. Mice deficient for the P2X7 receptor (P2X7-/-) were resistant to CHS, whereas wild-type (WT) controls were not. Additionally, P2X7 receptor antagonists (KN-62 and suramin) were injected into WT mice before TNCB treatment. These mice also failed to produce CHS. Lastly, when WT mice were injected with apyrase, an ATP degrading molecule, CHS was also prevented. Taken together, these experiments reinforce the role of the P2X7 receptor in the CHS response.
However, researchers wanted to see if ATP was in fact released in the skin after TNCB treatment. This is important because ATP is physiologically relevant to us. ATP- P2X7 receptor binding is therefore the initiation signal. Application of TNCB did in fact trigger the release of ATP, as seen by bioluminescence (imaging technique). This data showed that P2X7 is activated by the endogenous danger signal ATP released from cells after TNCB application.
Researchers then wanted to identify which part of the immune response (sensitization or effector) the receptor was mediating. DCs taken from P2X7-/- mice, and modified with TNBS, were not able to induce sensitization in WT mice. However, these same P2X7-/- DCs, when also pretreated with alum, were able to induce sensitization. Alum works by directly activating the NLRP3 inflammasome without the need to activate the P2X7 receptor first. As previously discussed, NLRP3 and its production of IL-1β are important in the sensitization process. In contrast, WT DC’s (also modified) were able to induce sensitization in WT mice without the need of alum. This experiment identified that the P2X7 receptor specifically mediates the sensitization/induction of CHS, and that it likely does so via the inflammasome-dependent processing and release of IL-1β.
Researchers, having just implicated the NLRP3 inflammasome as a participating agent in this mechanism, wanted to further prove that this structure within the DC mediates the sensitization response. They therefore then took bone marrow-derived DCs (BMDCs) from NLRP3 -/- mice and adoptively transferred to WT mice. CSH was not elicited. This lack of response was seen even with the alum pretreatment, reinforcing the notion that alum worked via the NLRP3 inflammasome. This proved that a functional NLP3 is required for sensitization. Furthermore, when the BMDCs taken from NLRP3 -/- mice were stimulated in vitro (in well plates) with LPS and ATP (inflammatory agents), no IL-1β was produced. Taken together, this suggests that P2X7 receptor activation results in NLRP3 inflammasome-mediated IL-1β release.
To further examine the effects of P2X7 deficiency, researchers examined the effects on secretion of IL-1β, again, a cytokine vital for the sensitization process. BMDCs from P2X7-/- mice were not able to secrete mature IL-1β when stimulated with ATP and LTP. This suggests that IL-1β release is dependent on the signaling through the P2X7 receptor. These results likely explained the failed sensitization experiments: defective IL-1β release failed to induce CHS.
To further address the role of IL-1β in the sensitization process, this cytokine was neutralized by injection of IL-1Ra anakinra before TNCB treatment. IL-1β mediates its effects through IL-1R.This injection failed to produce a CHS response. This effect was also seen when TNCB was applied to IL-1R -/- mice. Finally, an injection of IL-1β restored this sensitization to P2X7-/- mice.
This study sheds light on a particular receptor that could make a viable target for therapeutic strategies aimed at ACD/CHS. Researchers showed that blocking the activation of the P2X7 receptor on DCs and its effector mechanisms (knocking out P2X7, NLRP3 or IL-1β) alleviated CHS severity. Treatment may therefore aim to look at P2X7 receptor antagonists. Though the antagonists (KN-62 and suramin) used in this study worked effectively, there is still a need for antagonists that are highly specific for P2X7. Additionally, future studies hope to look at whether the effects of P2X7 activation extends beyond the sensitization phase into the effector phase. Though more studies must be done before clinical trials are even considered, this study provides evidence that significant progress is being made on this front!
Weber, F. C., Esser, P. R., Muller, T., Ganesan, J., Pellegatti, P., Simon, M. M., . . . Martin, S. F. (2010). Lack of the purinergic receptor P2X(7) results in resistance to contact hypersensitivity. The Journal of Experimental Medicine, 207(12), 2609-2619. doi:10.1084/jem.20092489
(1) Mak, Tak & Saunders, Mary. Primer to the Immune Response. California. Elsevier, 2011. Print.
(2) Mizumoto, N., Kumamoto, T., Robson, S. C., Sevigny, J., Matsue, H., Enjyoji, K., & Takashima, A. (2002). CD39 is the dominant langerhans cell-associated ecto-NTPDase: Modulatory roles in inflammation and immune responsiveness. Nature Medicine, 8(4), 358-365. doi:10.1038/nm0402-358
(3) Granstein, R. D., Ding, W., Huang, J., Holzer, A., Gallo, R. L., Di Nardo, A., & Wagner, J. A. (2005). Augmentation of cutaneous immune responses by ATP gamma S: Purinergic agonists define a novel class of immunologic adjuvants. Journal of Immunology (Baltimore, Md.: 1950), 174(12), 7725-7731.