Journal: PLoS ONE

Article Title: Glioblastoma: A Pathogenic Crosstalk between Tumor Cells and Pericytes

doi: 10.1371/journal.pone.0101402

Figure Lengend Snippet: GFP-actin-GBM cells (green) contacting an NG2-DsRed + -pericyte ( A , red iso-surface in magnified box) and a DLP ( B , blue; confocal section) through flectopodia (arrowheads indicate moniliform actin). Note the presence of GFP-actin within the DLP (merged channels, inset in B ). v, vessels (DiD-blue in A ; Ink-filled-grey in B ). C , Scheme showing pericyte (colored cells) in-vivo (top; BV, blood vessel), in-vitro (middle) and on silicone-substrate (bottom). Wrinkling is associated with high αSMA-expression (red-color). D and E ( boxed area in G) , DIC-optic video-frames of the same field before ( D ) and after ( E ) GBM cell addition to pre-plated pericytes. Pericytes alone produce stable drifting wrinkles (red arrows) that are de-stabilized by GBM cells. White and yellow arrowheads indicate the appearance and disappearance of wrinkles, respectively. Dashed line marks the upper-limit of GBM cell population, transposed from F and G , which show the low magnification of FR Dextran-labeled GBM cells (white false-color in F and magenta in G ), plated on cultured pericytes. Time in minutes. H , Traces of two wrinkles, produced before (i) and after (ii) U87-GBM cell-addition, revealing the spatial evolution and colored to indicate lengthening (violet to green) or shortening (green to red) for each time-frame (numbers). I , 3D-plot summarizing the wrinkling behavior of pericytes, either alone (red points, n = 40) or with U87-GBM cells (green points, n = 23). Note the lack of green points in clusters 1 and 2. E1, E2 and C: track-straightness of the ends (E) and center (C) of each wrinkle. Scale bars: 10 µm ( A , B ), 30 µm ( D ), 100 µm ( G ).

Article Snippet: For immunocytochemistry and immunohistochemistry, we used mouse monoclonal antibodies against human-CD44 (1∶100, BD Pharmingen), mouse/human-Cdc42 (1∶50, BD-Transduction Lab), human-Nestin (1∶200, Chemicon) and human-alpha3beta1 Integrin (1∶50, Chemicon); chicken polyclonal against mouse/human-Cdc42 (1∶50, GeneTex); mouse monoclonal against mouse/human-Vimentin (1∶60; clone V-9, Sigma) and alpha Smooth Muscle Actin (αSMA) (1∶70, in vitro, and 1∶50 in thick vibratome slices, Abcam); rabbit polyclonal against Laminin (1∶100; Chemicon), NG2 chondroitin sulphate proteoglycan (1∶80, Chemicon), Nitrotyrosine (1∶50, Millipore) and RFP (1∶400 and 1∶250, in vitro and thick vibratome slices, respectively; MBL); rat anti mouse-CD44 (1∶80, BD Pharmingen), mouse-CD68 (1∶60, Abd-Serotec) and clone rat-401 against mouse-Nestin (1∶150, Millipore).

Techniques: In Vivo, In Vitro, Expressing, Labeling, Cell Culture, Produced