Tumour-processed osteopontin and lactadherin drive the protumorigenic reprogramming of microglia and glioma progression

Abstract

Tumour tissue is infiltrated by myeloid cells that are reprogrammed into alternatively activated/regenerative (M2) macrophages. The contribution of major signalling pathways and their modulators/targets involved in the macrophage reprogramming is poorly known. Glioblastoma (malignant brain tumour) attracts and reprograms brain-resident microglia and peripheral macrophages into cells that increase invasion, angiogenesis and suppress antitumour immunity. Using a 'function-first' approach and glioma secretome proteomics we identified osteopontin and lactadherin as proteins that cooperatively activate amoeboid transformation, phagocytosis and motility of primary microglia cultures via integrins and FAK-Akt (focal adhesion kinase-Akt) signalling. A synthetic peptide interfering with integrin ligands blocks glioma-microglia communication, functional activation and M2 gene expression. We found that osteopontin/secreted phosphoprotein 1 (Spp1) produced by non-transformed cells acts as a proinflammatory factor inducing inflammatory signalling and M1 genes, and counteracts the action of lactadherin. Using constructs encoding functional mutants of osteopontin, we demonstrated sequential processing of Spp1 by thrombin and matrix metalloproteinase-3 and/or -7 (MMP-3 and/or -7) in glioma cells, which generates a microglia-activating form devoid of the inflammatory activity, while retaining the M2 reprogramming potential. A similar form of osteopontin is secreted by human glioma cells but not normal human astrocytes. Knockdown of osteopontin or lactadherin in glioma cells reduces intracranial glioma growth, blocks amoeboid transformation of myeloid cells and affects M2 reprogramming of microglia/macrophages. Our findings demonstrate how glioma cells misuse macrophage-activating signals and redesign primarily proinflammatory signals towards their advantage to induce M2 reprogramming of tumour-infiltrating brain macrophages.

Myeloid-derived suppressor cells in gliomas

“…These results are corroborated by our studies, which show a large number of Arg1 + but not Iba1 + cells (GAMs) infiltrating experimental C6 gliomas in rats [66]. Flow cytometry studies of double-positive cells – CD11b + Gr-1 + from experimental C6 gliomas (Fig.…”

“…The studies using pharmacological or genetic ablation of CD11b + cells in experimental glioma models demonstrated that tumours do not grow and invade the brain parenchyma if GAMs are ablated or functionally paralysed [59, 61–66]. In those studies the percentages of MDSCs and their functionality were not evaluated.…”

Dissecting functional phenotypes of microglia and macrophages in the rat brain after transient cerebral ischemia

“…Microglial cultures were prepared from 1‐day‐old Wistar rat pups as previously described (Ellert‐Miklaszewska et al, ; Ellert‐Miklaszewska et al, ). Briefly, cells were isolated from cerebral cortices by trypsinization, mechanically dissociated, and plated at a density of 3 × 10 5 cells/cm 2 on poly‐lysine‐coated flasks in Dulbecco's modified Eagle's medium (with Glutamax, 4.5 g/L glucose, 10% heat‐inactivated fetal bovine serum [Gibco], 100 U/mL penicillin, and 0.1 mg/mL streptomycin).…”

The Impact of the Tumor Microenvironment on the Properties of Glioma Stem-Like Cells

“…SPP1 is a secreted protein that shares similarities in features and mechanism of action with Periostin and contributes to the maintenance of an immune suppressive environment. It has been shown, in fact, to be highly expressed in M2 polarized macrophages and GBM-infiltrating CD14 + cells compared to matched blood monocytes and brain microglia (77). …”