Caveolin-1 implicated as a pro-invasive gene in high-grade glioma cell models: implementation of a 3d spheroid matrix invasion assay

INTRODUCTION: The poor prognosis associated with Glioblastoma multiforme (GBM) is multifactorial but includes the capacity of residual tumour cells not removed by surgery and resistant to radio-/chemo-therapy undergoing diffuse invasion into the surrounding brain tissue. Caveolin-1 (Cav-1) is the major structural and functional component of caveolae. In a number of tumour types Cav-1 is recognised to participate in cytoskeletal rearrangement, integrin-mediated adhesion and/or matrix remodelling. We proposed Cav-1 serves to promote invasion of GBM cells. To investigate this we have employed in an in-vitro 3D cellular invasion assay. METHOD: The human GBM cell lines, UP007 and UP029 established from primary cultures of biopsy-derived brain tumours (University of Portsmouth), U-87 MG (ECACC) and U-373 MG (ECACC) were genetically modified to stably knock-out Cav-1 using a Lentiviral Cav-1 shRNA approach; corresponding stably transfected non-target (NT) shRNA cell lines were generated as controls. Neuropheres were formed and embedded within an extracellular matrix (Matrigel™). Over a two-/four-day period (depending on cell line) the migration of cells away from the neurophere core (CORE) was quantified by image capture and processing (Image J) using a custom-developed MatLab script for pixel density analysis indicative of the density of migrating cellular material. RESULTS: Cav-1 knockout resulted in significant (P0.05) towards reduced invasion. Depending upon the cell line the Cav-1 knockdown also resulted in reduced size and cellular density of the neurosphere core (UP007 and UP029) indicative of reduced proliferation and/or cell survival capacity. CONCLUSION: Using an in-vitro 3D cellular invasion assay we have found Cav-1 expression in a series of three GBM cell lines to promote cellular invasion capacity. Ongoing studies are addressing signalling mechanisms and the influence of the microenvironment

INSIDIA:a FIJI macro delivering high-throughput and high-content spheroid invasion analysis

Time-series image capture of in vitro 3D spheroidal cancer models embedded within an extracellular matrix affords examination of spheroid growth and cancer cell invasion. However, a customizable, comprehensive and open source solution for the quantitative analysis of such spheroid images is lacking. Here, the authors describe INSIDIA (INvasion SpheroID ImageJ Analysis), an open-source macro implemented as a customizable software algorithm running on the FIJI platform, that enables high-throughput high-content quantitative analysis of spheroid images (both bright-field gray and fluorescent images) with the output of a range of parameters defining the spheroid “tumor” core and its invasive characteristics

Pandemics, vulnerability and prevention: time to fundamentally reassess how we value and communicate risk?

Pandemics have long been recognised on the UK’s National Risk Register as both the likeliest and most severe of threats. Prior to the COVID-19 pandemic, the burden of non-communicable diseases (NCDs) such as obesity, heart failure and mental ill-health was already crippling our healthcare services and our economy. The quality of the places in which we live and to which we have access, most especially in deprived populations, compound that vulnerability significantly. When vulnerable, as we are now, we are forced to acknowledge what has weakened us, and to fundamentally reassess our actions and priorities

Induction of membrane-type-1 matrix metalloproteinase by epidermal growth factor-mediated signaling in gliomas

This study showed for the first time that one mechanism of EGFR-mediated invasiveness in gliomas may involve the induction of membrane-type metalloproteinase (MT1-MMP)