Date Published: January 25, 2017
Publisher: Public Library of Science
Author(s): Nynke M. Kannegieter, Dennis A. Hesselink, Marjolein Dieterich, Rens Kraaijeveld, Ajda T. Rowshani, Pieter J. M. Leenen, Carla C. Baan, Kwan Man.
Monocytes and macrophages play key roles in many disease states, including cellular and humoral rejection after solid organ transplantation (SOT). To suppress alloimmunity after SOT, immunosuppressive drug therapy is necessary. However, little is known about the effects of the immunosuppressive drugs tacrolimus and mycophenolic acid (MPA) on monocyte activation and function. Here, the effect of these immunosuppressants on monocytes was investigated by measuring phosphorylation of three intracellular signaling proteins which all have a major role in monocyte function: p38MAPK, ERK and Akt. In addition, biological functions downstream of these signaling pathways were studied, including cytokine production, phagocytosis and differentiation into macrophages. To this end, blood samples from healthy volunteers were spiked with diverse concentrations of tacrolimus and MPA. Tacrolimus (200 ng/ml) inhibited phosphorylation of p38MAPK by 30% (mean) in CD14+ monocytes which was significantly less than in activated CD3+ T cells (max 60%; p < 0.05). This immunosuppressive agent also partly inhibited p-AKT (14%). MPA, at a therapeutic concentration showed the strongest effect on p-AKT (27% inhibition). p-ERK was inhibited with a maximum of 15% after spiking with either tacrolimus or MPA. The production of IL-1β and phagocytosis by monocytes were not affected by tacrolimus concentrations, whereas MPA did inhibit IL-1β production by 50%. Monocyte/macrophage polarization was shifted to an M2-like phenotype in the presence of tacrolimus, while MPA increased the expression of M2 surface markers, including CD163 and CD200R, on M1 macrophages. These results show that tacrolimus and MPA do not strongly affect monocyte function, apart from a change in macrophage polarization, to a clinically relevant degree.
Monocytes have numerous biologic functions that make them key players in solid organ transplantation (SOT)-related conditions, including ischemia-reperfusion injury and its repair, as well as regulation of allograft rejection [1–5]. After SOT, cells of the monocyte/macrophage lineage, process and present alloantigen to the recipients’ immune system, induce inflammation and contribute to allograft rejection, through the secretion of pro-inflammatory cytokines and by providing help to alloreactive T- and B-cells. For example, after ischemia and reperfusion injury, monocytes infiltrate the allograft where, after differentiation into macrophages, they produce inflammatory cytokines, can present donor antigen and also contribute to tissue injury and repair processes . Furthermore, immunohistochemistry of acute rejection kidney specimens demonstrated massive infiltration of the transplanted organ by CD68+ monocyte/macrophages [3, 7]. In antibody-mediated rejection, monocytes control and induce cell injury via the activation of their Fcγ-receptor by allo-antibodies [8, 9]
Cells from both the adaptive and innate immune system play an important role in the immune response after SOT but the effects of the immunosuppressants tacrolimus and MPA on human monocyte differentiation and functions have not been studied in-depth in previous studies. This study demonstrates that 1) these drugs partially inhibit phosphorylation of signaling molecules involved in CD14+ monocyte activation, i.e., p38MAPK, ERK and Akt kinases, but 2) have only limited effects on cytokine production, phagocytosis, phenotypic maturation, and polarization (an overview of de results is given in S1 Table).