Expansion of myeloid-derived suppressor cells (MDSCs) is associated with cancer progression and suppression of T cell function. However, further defining MDSCs and their role is hindered by the fact that no specific cell marker is available to distinguish MDSCs from non-suppressive leukocytes. Here we report a separation method that clearly separate granulocytic MDSC (G-MDSC) from non-suppressive granulocytes. In this study, mouse bone marrow cells from healthy and tumor bearing mice were collected, followed by Percoll density gradients centrifugation. Naive CD11b+Ly6G+Ly6Clow granulocytes from the fraction between 50-60% Percoll effectively inhibited the proliferation of T cells via production of ROS and arginase. These suppressive cells were expanded during tumor formation and the inhibitory function was enhanced in tumor bearing mice, indicating that G-MDSC was enriched at the 50-60% Percoll interface. Unlike G-MDSC, high density granulocyte (~60-70% Percoll fraction) failed to suppress T cells proliferation but had a high phagocytic activity and chemotatic capacity toward CXCL1, suggesting that they were mature neutrophils. Further study found that CXCR2 in G-MDSC from tumor bearing mice was significantly upregulated, leading to a massive migration of G-MDSC into spleen and tumor site. Collectively, our findings provide a new way to discriminate G-MDSC from mature granulocytes and reveal a critical role of CXCR2 in G-MDSC mobilization during tumor formation.

2014