gms | German Medical Science

Objective: Recent evidence indicates that myeloid-derived suppressor cells (MDSC) are markedly elevated in glioblastoma patients. Also, a subfraction of myeloid cells in blood and tumors has been reported to express variable immune receptors, i.e. T cell receptors. Here, we investigated circulating monocytes and tumor-associated macrophages (TAM) from glioblastoma patients for the expression of immunoglobulins (Ig) and their variability.

Method: Peripheral blood and tumor tissue of glioblastoma patients were used to isolate MDSCs, monocytes and TAM. MDSCs were collected and quantitated by FACS analyses. Monocytes and TAM were isolated using immunomagnetic microbeads. To evaluate the expression of Igs in the cell preparations, RNA isolation followed by PCRs was conducted using complement reverse and five different forward primers for constant and variable region Ig gene sequences. Positive bands were excised and the isolated DNA was used for TOPO-Cloning followed by Sanger sequencing. Analyses of the CDR3 region and the V-, D- and J-chain usage of the Igs were performed using the databases IgBLAST and VBASE2.

Results: First, our results showing elevated levels of MDSCs in the peripheral blood from glioblastoma patients are confirmed by independent recent reports. More importantly, we demonstrate the expression of variable Igs by circulating monocytes and TAM in glioblastoma patients. Purity of TAM/monocyte preparations was confirmed by the absence of T- and B-cell-specific gene expression indicating that all analyzed Igs are expressed by the purified monocytes and TAM. Furthermore, this work reveals that human monocytes and TAM express Ig repertoires in a highly restricted fashion compared to normal B cells. Specifically, analysis of the antigen-binding CDR3 regions showed remarkably smaller repertoires in TAM. This restriction in variability of TAM-Ig is even more pronounced with respect to V chain gene usage.

Conclusions: In this study, we demonstrate high levels of MDSCs in the peripheral blood of glioblastoma patients and the expression of variable Igs by circulating monocytes and TAM. The expressed Ig repertoires in glioblastoma patients vary significantly between TAM and circulating monocytes. Only a few V chain genes are used to build Igs according to a very limited number of CDR3 variants in the tumor environment. Further studies are needed to establish the relatedness of MDSCs and these new myeloid cells expressing variable immunoreceptors.