Figure 6.

Effects of ADAM10 inhibitors on Hodgkin lymphoma cell growth in 3D scaffolds repopulated with lymph node-mesenchymal stromal and Hodgkin lymphoma cells. (A) A representative Avitene^TM scaffold repopulated by LN-MSC16412 (2x10⁵) and L428 cells (4x10⁵). Sections (5 mm) from paraffin-embedded repopulated scaffolds were stained with DAPI for nuclei (blue), anti-CD30 monoclonal antibody (mAb) followed by anti-mouse Alexa Fluor594 (red) for Hodgkin lymphoma (HL) cells and anti-Ki67 polyclonal antibody followed by anti-rabbit Alexa Fluor488 (green) to identify cycling cells. Images were taken with the Aperio VERSA Digital Pathology Scanner (Leica Biosystems) with a 10x objective. Subpanel b: enlargement of the blue rectangle in subpanel a. Scale bars as indicated. (B and C) Avitene^TM scaffolds repopulated with 2x10⁵ MSC16412 and 4x10⁵ L428 (B) or L540 cells (C) and exposed to 10 mM LT4 or MN8 ADAM10 inhibitors for 72 hours (h) or 96 h as indicated. Nil: solvent (dimethyl sulfoxide 1:1,000) were subjected to immunofluorescence (IF) as in (A). At least three sections/scaffold, cut at 15 μm distance, were acquired. Image data were analyzed with the Aperio Cellular IF Algorithm (Leica Biosystems) and the percentage of CD30⁺/Ki67⁺ cells was calculated as described in the Online Supplementary Figure S1. Results are the mean ± standard error of the mean (SEM) from three independent experiments performed in duplicate (two scaffolds). *P<0.05 vs. nil; **P<0.005 vs. nil.