Interleukin-17 (IL-17) expression is reduced during acute myocardial infarction: role on chemokine receptor expression in monocytes and their in vitro chemotaxis towards chemokines

Abstract

The roles of immune cells and their soluble products during myocardial infarction (MI) are not completely understood. Here, we observed that the percentages of IL-17, but not IL-22, producing cells are reduced in mice splenocytes after developing MI. To correlate this finding with the functional activity of IL-17, we sought to determine its effect on monocytes. In particular, we presumed that this cytokine might affect the chemotaxis of monocytes important for cardiac inflammation and remodeling. We observed that IL-17 tends to reduce the expression of two major chemokine receptors involved in monocyte chemotaxis, namely CCR2 and CXCR4. Further analysis showed that monocytes pretreated with IL-17 have reduced in vitro chemotaxis towards the ligand for CCR2, i.e., MCP-1/CCL2, and the ligand for CXCR4, i.e., SDF-1α/CXCL12. Our results support the possibility that IL-17 may be beneficial in MI, and this could be due to its ability to inhibit the migration of monocytes.

Figure 1

A and B. Percentages of positive splenocytes isolated from either sham-operated wild-type mice (WT) or WT with myocardial infarction induced through left coronary artery ligation three days earlier (WT + MI). Intracellular labeling with antibodies against IL-17 was performed on intact splenocytes (A) or splenocytes activated with PMA plus ionomycin for 4 h before labeling (B). C and D. Percentages of positive cells labeled intracellularly with anti-IL-22 antibodies. The cells were either unstimulated (C) or were activated with PMA plus ionomycin for 4 h before labeling (D). Mean ± SEM of 5 experiments are shown.

Figure 2

(A) Monocytes were isolated by monocyte enrichment kit and were labeled with anti-CD11b. About 88% of monocytes were labeled with this antibody. This population was then stained with either anti-IL-17R or anti-IL-22R; (B) T cells were isolated using a T cell enrichment kit, labeled with anti-CD3 and then with anti-IL-17R or anti-IL-22R. One representative experiment indicating the percentages of positive cells is shown.

Figure 3

IL-17 may reduce the expression of CCR2 and CXCR4 on the surface of monocytes. Monocytes (1 × 10⁶) were either left intact (Control) or were pretreated with 250 ng/mL of IL-17 overnight. They were washed and then examined for the expression of CCR2 (A) or CXCR4 (B). The geometric mean of fluorescence intensity (Geo MFI) is shown. Each dot represents one independent experiment.

Figure 4

IL-17 inhibits monocytes chemotaxis towards MCP-1/CCL2. Monocytes were either untreated (control cells) or pretreated with 250 ng/mL of murine recombinant IL-17 overnight. The cells were washed, labeled with calcein-AM and then examined for migration towards different concentrations of MCP-1/CCL2. Two hours later, calcein-AM-labeled cells were collected and counted in the flow cytometer. Mean ± SEM of two experiments are shown.

Figure 5

IL-17 inhibits the chemotaxis of monocytes towards SDF-1α/CXCL12. (A) and (B) Monocytes were either untreated (control cells) or pretreated with 250 ng/mL IL-17 overnight. The cells were washed, labeled with calcein-AM and then examined for migration towards various concentrations of SDF-1α/CXCL12; (C) and (D) Migration index was calculated by measuring the numbers of migrating cells into the lower filters of the chemotaxis chambers. Mean ± SEM of two experiments are shown.