RORγt modulates macrophage recruitment during a hydrocarbon oil-induced inflammation

Abstract

Hydrocarbon oils are often utilized as adjuvants in vaccines. In response to naturally occurring hydrocarbon oils, inflammation is initiated and persists with the continuous recruitment of immune cells such as macrophages and neutrophils. However, the mechanism underlying the chronic inflammation in response to hydrocarbon oils is not fully defined. In this study, we revealed an essential role of retinoid-related orphan receptor gamma t (RORγt) in sustaining the recruitment of macrophages following pristane treatment. RORγt absence resulted in the incompetent formation of mesenteric oil granulomas which may associate to a reduction in the migration of macrophages into the mesentery during pristane-induced inflammation. This is at least partially dependent on the expression of the monocyte chemoattractant protein-1 (MCP-1) in the mesentery and the decrease in the macrophage reservoir in the spleen. However, the absence of RORγt had no impact on the recruitment of neutrophils to the mesentery after pristane treatment. Our data uncovered an important role of RORγt in the recruitment of macrophages during hydrocarbon oil-induced chronic inflammation.

Figure 1. RORγt controls pristane-induced formation of oil granulomas on the mesentery.

(A–B) The mesenteric tissues associated with the gut were collected from naïve mice or pristane-treated C57BL/6 mice (3 weeks) and photographed. SG, serosal granulomas; MG, mesenteric granulomas. (C–D) Images shown represent the mesenteric tissues associated with the gut in either naïve or pristane-treated (3 weeks) RORγt^−/− mice. (E) SG was quantitated in wild type (WT) controls and RORγt^−/− mice after pristane treatment. (F), Analysis of cellularity of whole mesenteric tissue in WT and RORγt^−/− mice. (G-I), The number of B cells (B220⁺), macrophages (CD11b⁺Gr1^low) or neutrophils (CD11b⁺Gr1^hi) in whole mesenteric tissue was summarized. Data show the average ± SD of three mice, representing three independent experiments. *p<0.05, **p<0.01, compared with the WT group.

Figure 2. RORγt deficiency blocks macrophage recruitment to oil granulomas.

(A–B), Recruitment of macrophages and neutrophils to SG; data are shown as the absolute numbers of each cell type per individual SG. (C–D), Recruitment of macrophages and neutrophils to MG; data are shown as the absolute numbers of each cell type per mm² of MG. *p<0.05, **p<0.01, compared with the WT group.

Figure 3. RORγt deficiency alters the expression of chemokines by mesenteric cells during pristane-induced inflammation.

Levels of mRNA of indicated chemokines in mesenteric cells from naïve or pristane-injected WT and RORγt^−/− mice were measured by quantitative PCR. Data show the average ± SD of three samples, representing three independent experiments. *p<0.05, compared with the control group.

Figure 4. RORγt deficiency reduces splenic macrophage expansion during pristane-induced inflammation.

Total cells (A), macrophages (B) and neutrophils (C) in the spleens of naïve or pristane-treated WT or RORγt^−/− mice were numerated. Data show the average ± SD of three samples, representing three independent experiments. *p<0.05, compared with the control group.