Rapamycin induces apoptosis in monocyte- and CD34-derived dendritic cells but not in monocytes and macrophages

Abstract

Rapamycin (Rapa), a recently introduced immunosuppressive drug, seems to be effective in preventing acute allograft rejection. Although its antiproliferative effect on T lymphocytes has been investigated extensively, its effect on the initiators of the immune response, the dendritic cells (DCs), is not known. Therefore, the effect of Rapa on monocyte-(mo-DCs) and CD34 ؉ -derived DCs in vitro but also on other myeloid cell types, including monocytes and macrophages, was examined. The present study shows that Rapa does not affect phenotypic differentiation and CD40L-induced maturation of mo-DCs. However, Rapa dramatically reduced cell recovery (40%-50%). Relatively low concentrations of Rapa (10 ؊9 M) induced apoptosis in both moDCs and CD34 ؉ -derived DCs, as visualized by phosphatidylserine exposure, nuclear condensation and fragmentation, and DNA degradation. In contrast, Rapa did not affect freshly isolated monocytes, macrophages, or myeloid cell lines. The sensitivity to Rapa-induced apoptosis was acquired from day 2 onward of mo-DC differentiation. Rapa exerts its apoptotic effect via a reversible binding to the cytosolic receptor protein FKBP-12, as demonstrated in competition experiments with FK506, which is structurally related to Rapa. Partial inhibition of Rapa-induced apoptosis was obtained by addition of ZVAD-fmk, which implies caspase-dependent and caspase-independent processes. The fact that Rapa exerts a specific effect on DCs but not on monocytes and macrophages might contribute to the unique actions of IntroductionDendritic cells (DCs) are professional antigen-presenting cells that are specialized in the uptake of antigens and their transport from peripheral tissues to the lymphoid organs. Because of their capacity to stimulate naive T cells, DCs have a central role in the initiation of primary immune responses. 1 Human DCs can be grown in vitro from CD34 ϩ progenitors in medium supplemented with granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor-␣ 2,3 or from monocytes in the presence of GM-CSF and interleukin (IL)-4 or IL-13. 4 It has been demonstrated also that in vivo monocytes can differentiate into DCs upon transmigration through endothelial layers. 5,6 DCs are thought to play a critical role in diverse facets of immune regulation, ranging from tolerance induction and the prevention of autoimmunity to the induction of antitumor immunity and the protection against infectious agents. 1,7-9 A similar dichotomy of DC functions has been proposed to play a role in allograft transplantation. Various experimental models have demonstrated that DCs are critical for the induction of transplant tolerance. [10][11][12][13] As a consequence DCs have been genetically modified to promote their tolerogenic potential. 14 On the other hand, it is clear that the immunostimulatory potential of DCs may also contribute to disease pathogenesis of allograft rejection. 15 Donor DCs transplanted en bloc with the allograft migrate to the recipient spleen, 16 where they can activate na...

Rapamycin specifically interferes with GM-CSF signaling in human dendritic cells, leading to apoptosis via increased p27KIP1 expression

“…Inhibition of PI3K, ERK, or p38 in monocytes slightly induced apoptosis ( Figure 2A). As shown previously, 15 monocytes were completely insensitive for the proapoptotic activity of Rapa, demonstrating that the longevity of DCs and their precursors, ie, the monocytes, are regulated by different survival mechanisms.…”

“…Like treatment of DCs with Rapa to inhibit mTOR, 15 blockade of PI3K in DCs by addition of LY294002 time-and dose-dependently induced apoptosis that was characterized by nuclear condensation and fragmentation ( Figure 2B) and DNA fragmentation (data not shown). In addition to the data obtained from GM-CSF deprivation experiments, both LY294002 and Rapa demonstrated loss of ⌬⌿ m associated with the percentage of apoptosis induced ( Figure 2C).…”

“…15 Rapa, which is an immunosuppressive drug introduced to prevent allograft rejection, 16 is extensively studied for its effect on T lymphocytes and is known mainly for its antiproliferative effect. 17 The drug is structurally related to FK506 that also inhibits T-cell proliferation.…”

Rapamycin inhibits macropinocytosis and mannose receptor–mediated endocytosis by bone marrow–derived dendritic cells

“…21 Having established that rapamycin inhibited DC endocytosis, we analyzed whether this effect was due to increased apoptotic cell death. In contrast to a recent report, 22 the incidence of apoptosis at day 7 of culture was consistently low (Ͻ 10%) and was not affected significantly by rapamycin, even at a suprapharmacological dose of 100 ng/mL, as determined independently by annexin-V/7-AAD and TUNEL staining ( Figure 2D, E). Similar results were obtained with GM-CSFϩIL-4-expanded DCs and at day 4 of culture (data not shown).…”

Cutting Edge: Sanglifehrin A, a Novel Cyclophilin-Binding Immunosuppressant Blocks Bioactive IL-12 Production by Human Dendritic Cells

“…We reported recently that Rapa impairs Ag uptake of murine dendritic cells (DC) (8), a finding that was subsequently confirmed in human DC (9). Moreover, it was suggested that Rapa promotes in vitro apoptosis of human DC by interfering with GM-CSF signaling (10) and suppresses in vivo DC development and function (11). DC are professional APC with a unique ability to induce and control adaptive and innate immune responses (12)(13)(14)(15).…”