Blood clotting supports glioma invasion and colonization

J. Pilch, A. Burkardt, C. Wolter, S. Urbschat, R. Ketter, D. Lessig, I. Müller, H. Eichler, L. M. Knowles (Homburg, Germany)

Acquired problems and alterations of coagulation
Date: 17.02.2017,
Time: 17:15 - 18:15

Objective: High-grade gliomas are highly invasive brain tumors that are extremely difficult to treat. High concentrations of pro-angiogenic factors cause glioma cells to induce vascular leakage, which in turn leads to the generation of a fibrin-rich edema. We hypothesize that the blood clot surrounding the glioma cells provides a provisional extracellular matrix that promotes tumor cell infiltration and growth. The objective of this study is to probe high-grade glioma cells for their capacity to infiltrate clotted plasma in comparison to benign meningioma and assess the underlying mechanism.

Methods: Tumor tissue from patients with high-grade glioma (WHO grade III-IV) or benign meningioma (grade I-II) was cultured in DMEM + 10% FBS to allow tumor cells to outgrow. Primary tumor cells were then embedded in clotted plasma, fibrin or Matrigel™ and scored for invadopodia formation as well as proliferation using phase contrast microscopy. Fibronectin mRNA expression was analyzed from tumor cell and tissue extracts using quantitative PCR. In addition, we assessed fibronectin and Slug/Snail2 protein expression by western blot.

Results: Primary tumor cells isolated from patients with high-grade glioma invaded and colonized extensively in clotted plasma and fibrin while lagging behind in Matrigel™. This clot-invasive tumor cell phenotype was specific for malignant brain tumors as tumor cells derived from benign meningioma were considerably less invasive in clotted plasma. Clot invasion correlated positively with fibronectin mRNA expression, which was strong in high-grade glioma, comparably weak in low-grade glioma (WHO grade II) as well as meningioma, and barely detectable in normal brain. High-grade glioma invasion in fibrin was accompanied by the generation of an elaborate fibronectin meshwork that serves to stabilize the adhesive interactions in the 3D environment. This in turn is relevant for the expression of the EMT transcription factor and tumor stem cell marker Slug/Snail 2, which we found to be upregulated in high grade glioma.

Conclusion: Our data show that clotted plasma, which is present in the fibrin-rich edema of the tumor extracellular matrix, strongly and specifically promotes invasion as well as colonization of high-grade glioma. Further research about the role of fibronectin and Slug/Snail2 in glioma invasion and colonization is warranted.