Dichotomy in duration and severity of acute inflammatory responses in humans arising from differentially expressed proresolution pathways

Abstract

Lipoxins (Lxs) and aspirin-triggered epi-Lxs (15-epi-LxA(4)) act through the ALX/FPRL1 receptor to block leukocyte trafficking, dampen cytokine/chemokine synthesis, and enhance phagocytic clearance of apoptotic leukocytes-key requisites for inflammatory resolution. Although studies using primarily inbred rodents have highlighted resolution as an active event, little is known about the role resolution pathways play in controlling the duration/profile of inflammatory responses in humans. To examine this, we found two types of responders to cantharidin-induced skin blisters in male healthy volunteers: those with immediate leukocyte accumulation and cytokine/chemokine synthesis followed by early resolution and a second group whose inflammation increased gradually over time followed by delayed resolution. In early resolvers, blister 15-epi-LxA(4) and leukocyte ALX were low, but increased as inflammation abated. In contrast, in delayed resolvers, 15-epi-LxA(4) and ALX were high early in the response but waned as inflammation progressed. Elevating 15-epi-LxA(4) in early resolvers using aspirin increased blister leukocyte ALX but reduced cytokines/chemokines as well as polymorphonuclear leukocyte and macrophage numbers. These findings show that two phenotypes exist in humans with respect to inflammation severity/longevity controlled by proresolution mediators, namely 15-epi-LxA(4). These data have implications for understanding the etiology of chronic inflammation and future directions in antiinflammatory therapy.

Fig. 1.

Aspirin reveals differential profiles of acute inflammatory responses in humans. Healthy male volunteers (n = 26) between the ages of 25 and 50 y had a cantharidin skin blister elicited on the ventral aspects of their forearm with baseline acute inflammatory responses established 24 h later. Each volunteer was then given ASA^low (75 mg) for 10 d, followed by creation of another blister. (A) The difference in blister total cell numbers before and after aspirin for each volunteer (y axis) against their corresponding baseline cell numbers on x axis (r = −0.81, P ≤ 0.001). Therefore, those above 0 on the y axis (n = 16) showed a reduction in cell after aspirin (i.e., responders) whereas those on or below 0 on the y axis showed no change or increased cell numbers, respectively (n = 10; nonresponders). Reduction in blister cell numbers after aspirin was highly significant in responders (B) but not in aspirin nonresponders (C), with differences in baseline inflammation severity between aspirin responders compared with nonresponders also significant (D). Data are presented as mean ± SEM; *P < 0.05 and ***P < 0.001.

Fig. 2.

Dichotomy in the profile of acute inflammatory responses in humans. The inflammatory profile of cantharidin-elicited skin blisters in aspirin responders was charted over time (in h) and compared with nonresponders showing that blister total cell numbers (A), CD16b-positive PMNs (Bi), and CD14-positive monocytes/macrophages (Bii) in aspirin responders (solid line) peaked in an immediate early manner and resolved quicker than in aspirin nonresponders (dotted line); with (Biii) a representative FACS dot plot charting the change from primarily CD16b-positive PMNs at 24 h to CD14-positive monocytes/macrophages at resolution in E^rvs. (C) Blister edema accumulation did not follow this trend. (D–L) Blister fluid exudate proinflammatory cytokines and chemokines with biologically relevant levels broadly mirrored inflammation in both groups. These profiles gave rise to us defining responses as early resolvers (E^rvs) or delayed resolvers (D^rvs). Data are presented as mean ± SEM; *P < 0.05 and ***P < 0.001 represent differences in inflammatory parameters between E^rvs and D^rvs at 24 h.

Fig. 3.

The 15-epi-LxA₄ and its ALX receptor dictate inflammation severity and longevity in humans with acute inflammation. Of the endogenous antiinflammatory and proresolution factors measured over time (in h), significant differences were found only in levels of blister fluid 15-epi-LxA₄ (A) as well as ALX expression on total blister leukocyte (B), CD16b-positive PMN (C), and CD14-positive monocytes/macrophages (D) between E^rvs (solid line) and D^rvs (dotted line). (C) Representative FACS histograms illustrate increased ALX expression on CD16b-positive PMNs in E^rvs compared with D^rvs with similar comparative histograms (D) illustrating increased ALX expression on CD14-positive monocytes/macrophages. Data are presented as mean ± SEM; *P < 0.05 and ***P < 0.001 represent differences in inflammatory parameters between E^rvs and D^rvs at 24 h. AU represents arbitrary units of ALX expression intensity determined by FACS.

Fig. 4.

Modulating 15-epi-LxA₄ with ASA^low dampens inflammation in E^rvs. ASA^low (75 mg/d for 10 d, data represented by red line) elevated 15-epi-LxA₄ in the blister fluids of 16 volunteers (A) as well as ALX expression on total blister leukocyte (B), PMN (C), and monocytes (D) compared with controls (black line). This elevation in endogenous 15-epi-LxA₄/ALX was associated with reduced blister fluid IL-β (E) and IL-8 (F) as well as blister total leukocyte (G), PMN (H), and monocyte numbers (I). Data are presented as mean ± SEM; *P < 0.05 represent differences after 24 h in cantharidin-induced skin blister inflammatory parameters in E^rvs before and after aspirin. AU represents arbitrary units of ALX expression intensity determined by FACS.

Fig. 5.

ASA^low is without effect on D^rvs. ASA^low (75 mg/d for 10 d; red dotted line) did not alter blister 15-epi-LxA₄ or ALX expression (A) on total blister leukocyte (B), PMN (C), and monocytes (D) compared with controls (black dotted line) in 10 human volunteers with a delayed resolving phenotype. Also, there was no effect of ASA^low on blister fluid IL-β (E) and IL-8 (F) as well as blister total leukocyte (G), PMN (H), and monocyte numbers (I). Data are presented as mean ± SEM; *P < 0.05. AU represents arbitrary units of ALX expression intensity determined by FACS.