Platelet-activating factor, a molecular sensor for cellular damage, activates systemic immune suppression

Abstract

Ultraviolet (UV) radiation plays a critical role in the induction of nonmelanoma skin cancer. UV radiation is also immune suppressive, and the immune suppression induced by UV irradiation has been identified as a major risk factor for skin cancer induction. Previously, we showed that UV exposure activates a cytokine cascade involving prostaglandin (PG)E(2), interleukin (IL)-4, and IL-10 that induces immune suppression. However, the earliest molecular events that occur immediately after UV exposure, especially those upstream of PGE2, are not well defined. UV-irradiated keratinocytes secrete the inflammatory phospholipid mediator, platelet-activating factor (PAF). Because PAF upregulates the production of immunomodulatory compounds, including PGE2, we tested the hypothesis that UV-induced PAF activates cytokine production and initiates UV-induced immune suppression. Both UV and PAF activated cyclooxygenase (COX)-2 and IL-10 reporter gene construct transcription. PAF mimicked the effects of UV in vivo and suppressed delayed-type hypersensitivity (DTH). Furthermore, immune suppression was blocked when UV-irradiated mice were injected with PAF receptor antagonists. In addition to the well-known role of PAF as a proinflammatory lipid mediator, we propose that the PAF receptor senses cellular damage through the recognition of PAF and/or PAF-like molecules, such as oxidized phosphatidylcholine, which activates cytokine transcription and induces systemic immune suppression.

Figure 1.

Upregulation of cytokine gene transcription by PAF. Murine keratinocytes were transfected with a COX-2 reporter construct (A) or an IL-10 reporter construct (B) and exposed to 200 J/m² of UV radiation (○) or cultured with 10 nM cPAF (▴). The PAF receptor antagonist, PCA-4248, was added to the medium before UV radiation (□) or cPAF (♦). At the times indicated, the cells were lysed and the luciferase activity determined. The data is expressed as fold-induction by comparing the response of UV- or cPAF-treated cells to that of keratinocytes transfected with the reporter gene constructs and treated with tissue culture medium. Each point represents the mean ± SEM of three separate determinations.

Figure 2.

PAF suppresses the induction of DTH. Mice were injected with cPAF (50 to 5,000 pmol) or exposed to UV radiation (10 kJ/m²) 5 d before immunization with C. albicans. 9 d later, the mice were challenged with Candida antigen. DTH was measured 24 h after challenge. The background response (negative control) was measured in mice that were not immunized but were challenged. The positive control was measured in mice that were immunized and challenged. Results are expressed as means ± SEM. An asterisk (*) indicates a statistically significant difference (P < 0.01) from the positive control (two-tailed Student's t test, n = 5). A representative experiment is shown; this experiment was repeated three times with similar results.

Figure 3.

Injecting a PAF receptor antagonist blocks UV-induced immune suppression. Mice were injected with 5 to 500 nmol PCA-4248, a PAF receptor antagonist 1 h before UV-irradiation (10 kJ/m²). 5 d later, the mice were immunized with C. albicans, and 9 d later, challenged with Candida antigen. DTH was measured 24 h after challenge. The background response (negative control) was measured in mice that were not immunized but were challenged. The positive control was measured in mice that were immunized and challenged. Results are expressed as means ± SEM. An asterisk (*) indicates a statistically significant difference (P < 0.01) from the positive control (two-tailed Student's t test, n= 5). A representative experiment is shown; this experiment was repeated three times with similar results.

Figure 4.

Structurally diverse PAF receptor antagonists block UV-induced immune suppression. Mice were injected with 500 nmol PCA 4248, CV-3988 or dioxolane, and then exposed to 10 kJ/m² UV radiation. 5 d later, the mice were immunized with C. albicans, and 9 d later, challenged with Candida antigen. DTH was measured 24 h after challenge. The background response (negative control) was measured in mice that were not immunized but were challenged. The positive control was measured in mice that were immunized and challenged. Results are expressed as means ± SEM. * indicates a statistically significant difference (P < 0.01) from the positive control (two-tailed Student's t test, n = 5). # indicates a statistically significant difference (P < 0.01) from the response found in UV-irradiated mice. A representative experiment is shown; this experiment was repeated three times with similar results.

Figure 5.

UV-irradiated phosphatidylcholine induces immune suppression. Mice were injected with 500 pmol cPAF, 500 nmol normal phosphatidylcholine, (PC) or 5 to 500 nmol UV-PC, 5 d before immunization with C. albicans. 9 d later, the mice were challenged with Candida antigen. DTH was measured 24 h after challenge. The background response (negative control) was measured in mice that were not immunized but were challenged. The positive control was measured in mice that were immunized and challenged. Results are expressed as means ± SEM. An asterisk (*) indicates a statistically significant difference (P < 0.01) from the positive control (two-tailed Student's t test, n = 5). A representative experiment is shown; this experiment was repeated three times with similar results.

Figure 6.

Blocking COX-2 activity in vivo abrogates PAF receptor agonist-induced immune suppression. Mice were exposed to UV radiation (10 kJ/m²), or injected with cPAF (500 pmol) or UV-PC (500 nmol) with (black bars) or without (white bars) the selective COX-2 inhibitor, SC 236. 5 d later, the mice were immunized with C. albicans. 9 d later, the mice were challenged with Candida antigen. DTH was measured 24 h after challenge. The background response (negative control) was measured in mice that were not immunized but were challenged. The positive control was measured in mice that were immunized and challenged. Results are expressed as means ± SEM. An asterisk (*) indicates a statistically significant difference (P < 0.01) from the positive control (two-tailed Student's t test, n = 5). A representative experiment is shown; this experiment was repeated three times with similar results.