Both hydrostatic and osmotic pressures are altered in the tumour microenvironment. Glioblastoma (GBM) is a brain tumour with high invasiveness and poor prognosis. We hypothesized that physical and osmotic forces regulate glioblastoma (GBM) invasiveness. the osmotic pressure of GBM cell culture medium was adjusted using sodium chloride or water. Alternatively, cells were subjected to increased hydrostatic force. The proteolytic profile and epithelial-mesenchymal transition (EMT) were investigated using zymography and real-time qPCR. The EMT markers assessed were Snail-1, Snail-2, N-cadherin, Twist and vimentin. Invasion was investigated in vitro using extracellular matrixcoated Transwell inserts. In response to osmotic and mechanical pressure, GBM cell lines U87 and U251 and patient-derived neural oncospheres upregulated the expression of urokinase-type plasminogen activator (upA) and/or matrix metalloproteinases (MMps) as well as some of the eMt markers tested. the adherent cell lines invaded more when placed in media of increased osmolality. therefore, GBM respond to osmotic or mechanical pressure by increasing matrix degrading enzyme production, and adopting a phenotype reminiscent of eMt. Better understanding the molecular and cellular mechanisms by which increased pressure promotes GBM invasiveness may help to develop innovative therapeutic approaches. Physical solid and fluid forces play a key role when solid tumours grow, progress and also respond to therapy 1. Compressive stresses affect cancer cells by promoting invasiveness and metastasis 2. Tumours are generally hypoperfused, and interstitial fluid pressure is increased compared to normal tissue 1,3,4 with both increased hydrostatic pressure 4 and oncotic pressure 5,6. Increased interstitial fluid pressure results from abnormal blood and lymphatic vessels, fibrosis and contraction of the matrix by stromal cells 7. In addition to these stresses common to most solid tumours, brain tumours experience pressure when the tumour grows within a space limited by the skull 8. Glioblastoma is the most common primary brain cancer. The average survival time is approximately one year after diagnosis. A major feature of GBM that contributes to its poor prognosis is its high invasiveness. The urokinase-type plasminogen activator (uPA) derives its name from its ability to activate plasminogen into plasmin. While tissue-type plasminogen activator (tPA) plays a role in the fibrinolytic process, uPA is involved in cell migration and tissue remodelling, thereby playing a major role in cancer development and spreading. This role is especially crucial in glioblastoma 9-11. Equally important and complementary to the uPA system, MMPs play a key role in the control of the tumour microenvironment and ECM, thereby modulating tumor growth, angiogenesis, invasion and metastasis. Recently reports showed that the MMPs play pivotal roles in the invasiveness of GBM by degrading surrounding tissue, activating signal transduction, releasing ECM-bound growth factors, activating