Vortioxetine exerts anti-inflammatory and immunomodulatory effects on human monocytes/macrophages

Abstract

Background and purpose: A crosstalk between the immune system and depression has been postulated, with monocytes/macrophages and cytokines having a key role in this interaction. In this study, we examined whether vortioxetine, a multimodal anti-depressive drug, was endowed with anti-inflammatory and antioxidative activity, leading to immunomodulatory effects on human monocytes and macrophages.

Experimental approach: Human monocytes were isolated from buffy coats and used as such or differentiated into M1 and M2 macrophages. Cells were treated with vortioxetine before or after differentiation, and their responsiveness was evaluated. This included oxy-radical and TNFα production, TNFα and PPARγ gene expression and NF-κB translocation.

Key results: Vortioxetine significantly reduced the PMA-induced oxidative burst in monocytes and in macrophages (M1 and M2), causing a concomitant shift of macrophages from the M1 to the M2 phenotype, demonstrated by a significant decrease in the expression of the surface marker CD86 and an increase in CD206. Moreover, treatment of monocytes with vortioxetine rendered macrophages derived from this population less sensitive to PMA, as it reduced the oxidative burst, NF-kB translocation, TNFα release and expression while inducing PPARγ gene expression. FACS analysis showed a significant decrease in the CD14+ /CD16+ /CD86+ M1 population.

Conclusions and implications: These results demonstrate that in human monocytes/macrophages, vortioxetine has antioxidant activity and anti-inflammatory effects driving the polarization of macrophages towards their alternative phenotype. These findings suggest that vortioxetine, alongside its antidepressive effect, may have immunomodulatory properties.

© 2017 The British Pharmacological Society.

Figures

Figure 1

Effect of vortioxetine on superoxide anion production in monocytes. Human monocytes were pre‐incubated for 1 h with the indicated drugs (Ctrl, control untreated cell) and then stimulated with PMA 1 μM for 30 min. (A) Dose‐dependent decrease of the PMA‐oxidative burst by vortioxetine in stimulated cells. Data are means ± SEM of six independent experiments from distinct donors, analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison. (B) FACS analysis of O2 − positive cells in the presence or absence of the indicated drugs (vortioxetine, v; rosiglitazone, rosig). Data are means ± SEM of six independent experiments from distinct donors, analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison. Significance levels: °P < 0.05 versus Ctrl; *P < 0.05 versus PMA.

Figure 2

Effect of vortioxetine on superoxide anion production in M1 and M2 macrophages. Macrophages were pre‐incubated for 1 h with the indicated drugs (Ctrl, control untreated cell) and then stimulated with PMA 1 μM for 30 min. Vortioxetine decreased the level of reduced CytC on PMA stimulated M1 (A) and M2 (B) macrophages in a dose‐dependent manner. Significance levels: °P < 0.05 versus Ctrl; *P < 0.05 versus respective PMA. Data are means ± SEM of five independent experiments from distinct donors, analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison.

Figure 3

FACS analysis of phenotype/superoxide anion production in M1 and M2 macrophages. Macrophages differentiated from human monocytes were pre‐incubated with the indicated drugs [Ctrl, (control) untreated cells; 5‐HT, 1 μM; rosiglitazone (rosig), 1 μM; PGJ2, 10 μM] and then stimulated with PMA 1 μM for 30 min. Cells were then stained with antibodies anti‐CD86, anti‐CD206 and anti‐O2 −. (A) Mean fluorescence intensity (MFI) of cells stained with the O2 − detection probe. Significance level: °P < 0.05 versus Ctrl; *P < 0.05 versus PMA. (B) Percentage of M1 macrophages positive for the O2 − staining correlated to CD86 and CD206 expression. Significance level: *P < 0.05 versus PMA, referred to O2 −. #P < 0.05 versus PMA, referred to CD206. (C) Percentage of M2 macrophages positive for the O2 − staining correlated to the CD86 and CD206 expression. Significance level: *P < 0.05 versus PMA, referred to CD86 curve. All data are expressed as means ± SEM of five independent experiments from distinct donors analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison.

Figure 4

Real‐time analysis of PPARγ and TNFα expression. PPARγ and TNFα gene expression in (A) monocytes, (B) M1 and (C) M2 macrophages, challenged for 6 h with vortioxetine, 5‐HT or PGJ2, 10 μM. Data are means ± SEM of eight independent experiments from distinct donors analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison. Significance level: °P < 0.05 versus respective control; *P < 0.05 versus respective control.

Figure 5

Superoxide anion production in M1 and M2 macrophages differentiated from vortioxetine‐treated monocytes. M1 (A) and M2 (B) macrophages were differentiated for 6 days, in the presence of the specific cytokine cocktail, from monocytes treated for 6 h with vortioxetine, 5‐HT 1 μM, rosiglitazone 1 μM or PGJ2 10 μM. On the day of the experiment, macrophages were stimulated with PMA 1 μM for 30 min. Results are expressed as nmol of reduced cytochrome C (CytC) in response to PMA. Significance level: °P < 0.05 versus Ctrl; *P < 0.05 versus PMA alone. Data are means ± SEM of five independent experiments from distinct donors, analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison.

Figure 6

Real‐time analysis of PPARγ and TNFα expression. M1 (A) and M2 (B) macrophages were differentiated for 6 days, in the presence of the specific cytokine cocktail, from monocytes treated for 6 h with vortioxetine, 5‐HT or PGJ2 10 μM. Data are means ± SEM of six independent experiments from distinct donors, analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison. Significance levels: *P < 0.05 versus Ctrl.

Figure 7

Effect of vortioxetine on NF‐κB activity and TNFα secretion in PMA‐stimulated cells. ELISAs were performed in M1 (A) and M2 (B) macrophages differentiated for 6 days, in the presence of the specific cytokine cocktail, from monocytes treated for 6 h with vortioxetine (v), 5‐HT 1 μM, rosiglitazone (rosig) 1 μM and PGJ2 10 μM. On the day of the experiment, macrophages were stimulated with PMA 1 μM for 30 min. NF‐κB activity is expressed as optical density (O.D.) at 450 nm and secreted TNFα as pg·mL−1. Data are means ± SEM of six independent experiments from distinct donors analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison. Significance levels: °P < 0.05; *P < 0.05 versus respective control.

Figure 8

Effect of vortioxetine on surface marker expression. M1 and M2 macrophages were obtained from differentiating monocytes treated for 6 h with vortioxetine (vortio) 7.5 nM or 5‐HT 1 μM. Cells were stained with the indicated antibodies and the co‐expression of CD14/CD16/CD86 and CD14/CD163/CD206 in the two populations was analysed. Vortioxetine significantly reduced the M1 population (CD14+/CD16+/CD86+ cells), while the M2 population (CD14+/CD163+/CD206+) was not influenced by the drug treatment. Data are expressed as mean ± SEM of six independent experiments from distinct donors, analysed by one‐way ANOVA with Kruskal–Wallis test for multiple comparison. Significance level: *P < 0.05 versus macrophages differentiated from untreated monocytes (Ctrl).