Properties of Immature Myeloid Progenitors with Nitric-Oxide-Dependent Immunosuppressive Activity Isolated from Bone Marrow of Tumor-Free Mice

Abstract

Myeloid derived suppressor cells (MDSCs) from tumor-bearing mice are important negative regulators of anti-cancer immune responses, but the role for immature myeloid cells (IMCs) in non-tumor-bearing mice in the regulation of immune responses are poorly described. We studied the immune-suppressive activity of IMCs from the bone marrow (BM) of C57Bl/6 mice and the mechanism(s) by which they inhibit T–cell activation and proliferation. IMCs, isolated from BM by high-speed FACS, inhibited mitogen-induced proliferation of CD4+ and CD8+ T-cells in vitro. Cell-to-cell contact of T-cells with viable IMCs was required for suppression. Neither neutralizing antibodies to TGFβ1, nor genetic disruption of indolamine 2,3-dioxygenase, abrogated IMC-mediated suppressive activity. In contrast, suppression of T-cell proliferation was absent in cultures containing IMCs from interferon-γ (IFN-γ) receptor KO mice or T-cells from IFN-γ KO mice (on the C57Bl/6 background). The addition of NO inhibitors to co-cultures of T-cells and IMC significantly reduced the suppressive activity of IMCs. IFN-γ signaling between T-cells and IMCs induced paracrine Nitric Oxide (NO) release in culture, and the degree of inhibition of T-cell proliferation was proportional to NO levels. The suppressive activity of IMCs from the bone marrow of tumor-free mice was comparable with MDSCs from BALB/c bearing mice 4T1 mammary tumors. These results indicate that IMCs have a role in regulating T-cell activation and proliferation in the BM microenvironment.

Granulocytic myeloid-derived suppressor cells promote angiogenesis in the context of multiple myeloma

“…However, no difference was seen in the suppressive potential of MDSCs isolated from MM-bearing mice or healthy controls. In contrast to studies reporting that immature myeloid cells from naive mice are not immunosuppressive [14, 22], our results are in accordance with a report from Forghani et al who observed a comparable suppressive capacity for MDSCs isolated from the BM of tumor-free or mammary tumor-bearing mice, suggesting a physiological role of MDSCs in the control of T-cell proliferation and activation in the BM microenvironment [23]. Similarly, no significant difference was reported between the suppressive potential of MDSCs isolated from MM patients or healthy donors [16].…”

PD-1 and CTLA-4 up regulation on donor T cells is insufficient to prevent GvHD in allo-HSCT recipients

“…Our data indicate that the expression levels of immune checkpoint blockade molecules and Tregs varies dynamically over time and across different hematolymphoid tissues and organs in transplant recipients. Among transplant recipients with established GvHD, the expression levels and numbers of PD-1 and CTLA-4 positive donor T cells were higher in the BM than the spleen, consistent with a recent publication showing that the BM microenvironment is relatively immune-suppressive [ 39 ]. Host APCs surviving in the spleen and BM after lethal irradiation have been shown to play a pivotal role in activating allo-reactive donor T cells that cause GvHD [ 40 – 42 ].…”

Poly (I: C) modulates the immunosuppressive activity of myeloid-derived suppressor cells in a murine model of breast cancer

“…Spleen and tumor suspension were stained with allophycocyanin (APC)-cyanine7 (Cy7)-conjugated anti-CD11b, fluorescein isothiocyanate (FITC)-conjugated Gr-1, and PE-conjugated lineage markers for 20 min at 4°C and FACS was performed as we have previously described [18].…”

“…CD80, CD86, and MHC class II (I-A d ) expression levels were used to evaluate the maturation status of MDSCs [18,23]. Splenocyte-derived MDSCs from 4T1 tumor-bearing mice were co-cultured with Poly (I: C) (20 lg/ml) overnight.…”

“…Of note, even when MDSC from Poly (I: C)-treated mice were re-cultured without additional exposure to Poly (I: C), ROS levels remained lower than in PBS-treated mice. We also analyzed the effect of Poly (I: C) on the immunosuppressive activity of MDSCs using proliferation of splenocytes stimulated with anti-CD3/CD28 beads as an in vitro readout, as described [18]. Briefly, (5 9 10 5 ) sorted purified splenocyte-derived MDSCs were co-cultured in FBS-supplemented RPMI 1640 in the presence of Poly (I: C) in 24-well flat-bottom plates.…”