Glioma Initiating Cells Form a Differentiation Niche Via the Induction of Extracellular Matrices and Integrin αV

Abstract

Glioma initiating cells (GICs) are considered responsible for the therapeutic resistance and recurrence of malignant glioma. To clarify the molecular mechanism of GIC maintenance/differentiation, we established GIC clones having the potential to differentiate into malignant gliomas, and subjected to DNA microarray/iTRAQ based integrated proteomics. 21,857 mRNAs and 8,471 proteins were identified and integrated into a gene/protein expression analysis chart. Gene Ontology analysis revealed that the expression of cell adhesion molecules, including integrin subfamilies, such as α2 and αV, and extracellular matrices (ECMs), such as collagen IV (COL4), laminin α2 (LAMA2), and fibronectin 1 (FN), was significantly upregulated during serum-induced GIC differentiation. This differentiation process, accompanied by the upregulation of MAPK as well as glioma specific proteins in GICs, was dramatically accelerated in these ECM (especially FN)-coated dishes. Integrin αV blocking antibody and RGD peptide significantly suppressed early events in GIC differentiation, suggesting that the coupling of ECMs to integrin αV is necessary for GIC differentiation. In addition, the expression of integrin αV and its strong ligand FN was prominently increased in glioblastomas developed from mouse intracranial GIC xenografts. Interestingly, during the initial phase of GIC differentiation, the RGD treatment significantly inhibited GIC proliferation and raised their sensitivity against anti-cancer drug temozolomide (TMZ). We also found that combination treatments of TMZ and RGD inhibit glioma progression and lead the longer survival of mouse intracranial GIC xenograft model. These results indicate that GICs induce/secrete ECMs to develop microenvironments with serum factors, namely differentiation niches that further stimulate GIC differentiation and proliferation via the integrin recognition motif RGD. A combination of RGD treatment with TMZ could have the higher inhibitory potential against the glioma recurrence that may be regulated by the GICs in the differentiation niche. This study provides a new perspective for developing therapeutic strategies against the early onset of GIC-associated glioma.

Integrative Network Analysis Combined with Quantitative Phosphoproteomics Reveals Transforming Growth Factor-beta Receptor type-2 (TGFBR2) as a Novel Regulator of Glioblastoma Stem Cell Properties

“…Our phosphoproteomic result indicated that the phosphorylation levels at Ser 21 and Tyr 420 were elevated in serum-free cultured GB2 cells, which reflected the previous evidence that the high phosphorylation level of Tyr 420 on Fyn was observed in clinical samples from glioblastoma patients (44). In serum cultured GB2 cells, the functional sites for the kinase activity of MAPK1 (Thr 185 / Tyr 187 ) and MAPK3 (Tyr 204 ) were also highly phosphorylated in agreement with the previous study regarding glioblastoma stem cells (14).…”

“…Fyn-mediated phosphorylation of this molecule are known to contribute to promote self-renewal and tumor formation of glioblastoma stem cells (32). The phosphorylated Thr 185 /Tyr 187 residues of MAPK1 and Tyr 204 of MAPK3 were also reported to be up-regulated in glioblastoma stem cells after serummediated loss of stem-like characteristics in the previous study (14).…”

“…Schulte et al compared the global gene expression profiles of glioblastoma stem cells with those of serum cultured monolayer cells and conventional glioblastoma cells, leading to identification of C-X-C chemokine receptor type 4 (CXCR4) as a target to block the growth of invasive glioblastoma stem cells (13). The integrated analysis of gene and protein expression in glioblastoma stem cells after serum-mediated alteration showed that integrin family members and extracellular matrixes, such as collagen type 4, laminin alpha 2, and fibronectin, were up-regulated in the serum cultured cells at both of the mRNA and protein levels (14). This report also revealed that interaction of integrin 5 and fibronectin functioned in the formation of the niches of glioblastoma stem cells.…”

“…Although the previous transcriptome and proteome analysis suggested some key molecules for maintenance of glioblastoma stem cell properties, the global changes of protein phosphorylation in serum-induced alteration remain unclear (13,14). Thus, we aimed to reveal the phosphoproteome dynamics in glioblastoma stem cells named GB2, which were established from the tumor tissues of the glioblastoma patient (15)(16)(17)(18).…”

Glia to glioma: A wrathful journey

“…GIC stays in a niche reported to be composed of stem cells, supportive cells, serum growth factors, immunological cells and eCM [76,77] . The αV integrin, one of the key factors of eCM, seems to have a pivotal role in the increased pathological outcome of the GIC.…”