P2Y14 receptor activation decreases interleukin-6 production and glioma GL261 cell proliferation in microglial transwell cultures

Gliomas are rich in extracellular nucleotides that modulate glioma cell production of multiple cytokines including interleukin (IL)-6, which strongly contributes to glioma cell proliferation. However, little is known about how nucleotide signaling modulates microglial/macrophage (MG/MP) cytokine production in the context of gliomas, nor how MG/MP purinergic P2 receptor expression changes in the tumor micro-environment. We hypothesized that: (1) expression of key P2Y receptors will be augmented in glioma-derived MG/MP, and (2) selective activation of these receptors in vitro will regulate microglial production of IL-6 and glioma cell proliferation. We tested these hypotheses using the murine GL261 glioma model. Compared to MG/MP isolated from the normal brain tissue, CD11b⁺ cells isolated from GL261 tumors expressed higher levels of several P2 receptors, including P2Y14 receptors. To evaluate microglial P2Y14 receptor function in the context of tumor cells, we first cultured N9 microglia in transwells with GL261 cells and found that microglial P2Y14 mRNA levels were similarly increased in transwell cultures. GL261 cells did not express detectable P2Y14 levels either when they were cultured alone or in transwell cultures with N9 cells. Selective P2Y14 receptor activation with UDP-glucose (UDPG) did not affect IL-6 levels in either cell type cultured alone, but in transwell cultures, UDPG decreased IL-6 protein levels in the medium. Application of conditioned medium from UDPG-treated microglia reduced GL261 cell proliferation. Together, these data suggest that P2Y14 receptors may be a key a receptor involved in glioma cell-MG/MP communication in the tumor environment.