Abstract
Tumor-associated macrophages (TAM) are abundant in glioblastoma (GBM), composing up to 30% of the total tumor mass. However, their genotype/phenotype and exact role in tumor progression and immune suppression are not fully understood. Macrophages are believed to be polarized along a spectrum spanning from M0, M1, and M2 states. M1 TAMs are proinflammatory while M2 TAMs are associated with anti-inflammatory, pro-tumor responses. For example, using gelatin zymography we saw an increase in MMP 2 and 9 activities in co-culture of GBM cells and M2 polarized macrophages. In search for factors determining M2 type TAMs, we found Aldehyde Dehydrogenase 1 family A2 (ALDH1A2) to be highly over-expressed in M2 polarized THP1 cells using a gene expression array. We also performed single cell sequencing on CD45+ cells isolated from GBM tumors using antibody-conjugated magnetic beads. Monocytic/macrophage cells were found in 2 clusters and ALDH1A2 expression was increased in 2.1% of CD68+/CD163+ cells in Cluster 1 and 2.7% of CD68+/CD163+ cells in Cluster 2. Notably, ALDH1A2 was not detected in peripheral blood mononuclear cells. Analysis of the single cell sequencing data has led to identification of several genes whose expression is increased in ALDH1A2+ cells compared to ALDH1A2- cells. These genes are associated with lipid metabolism, regulation of neoplastic transformation, and insulin growth factor signaling. To further validate these results, we co-stained GBM tumor sections for CD163 and ALDH1A2 and observed that ALDH1A2 is co-localized in M2 macrophages. We have noticed a propensity of ALDH1A2+ cells to be associated with tumor neovasculature. Being that ALDH1A2 is the main enzyme in retinoic acid (RA) synthesis, it is plausible that this could represent another possible function of these enzyme-positive TAMs. Taken together these data suggest a role for ALDH1A2 as a novel putative marker of a subset of M2 TAM phenotype and function in GBM.