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Date of download: 4/7/2017 Copyright © ASME. All rights reserved. From: Engineering a Three-Dimensional In Vitro Drug Testing Platform for Glioblastoma J. Nanotechnol. Eng. Med. 2016;6(4):041002-041002-6. doi:10.1115/1.4032903 Figure Legend: Schematic representation of microwell platform to generate 3D GBM tumors and drugs testing. Step 1: thin layer of hydrogel was fabricated on cover glass treated with TMSPMA. Step 2: second hydrogel layer with microwells was prepared by the photolithography technique. Step 3: cell seeding and spheroid formations. Step 4: anticancer drug sensitivity testing on tumor spheroids. Date of download: 4/7/2017 Copyright © ASME. All rights reserved. From: Engineering a Three-Dimensional In Vitro Drug Testing Platform for Glioblastoma J. Nanotechnol. Eng. Med. 2016;6(4):041002-041002-6. doi:10.1115/1.4032903 Figure Legend: (a) Treatment of Irinotecan or/and Pitavastatin on 2D monolayer cells. U87 cells were seeded into six-well plate to final concentration 0.6 × 106 cells/well. On the next day, they were treated with Pitavastatin or Irinotecan at working concentrations 0, 1, 10, 50, and 100 μM. At day 4 of treatment, U87 cells were observed to lose their integrity and monolayer structure at dose 50 μM or higher with Irinotecan, 10 μM or higher with Pitavastatin, and Pita 1 μM + Iri 10 μM with combinatorial drugs. (b) U87 cells appeared to lose monolayer structure in 2D monolayer cell culture and cell—cell interaction in 3D cell culture and undergo cell lysis. Autophagic cell death was shown with arrows in both 2D monolayer and 3D cell culture spheroids. Scale bars represent 200 μm. Date of download: 4/7/2017 Copyright © ASME. All rights reserved. From: Engineering a Three-Dimensional In Vitro Drug Testing Platform for Glioblastoma J. Nanotechnol. Eng. Med. 2016;6(4):041002-041002-6. doi:10.1115/1.4032903 Figure Legend: (a) Time-lapse image of control untreated U87 spheroids and spheroids treated with 10, 50, 100, 150, and 200 μM Pitavastatin on days 1, 4, and 7 after treatment. The spheroids continued to growth for 7 days after treatment of concentrations 10 μM and control group. At drug concentration 50 μM, the spheroids lost shape after 7 days while spheroids treated by 100 μM and higher drug concentrations started from day 1. Scale bars represent 200 μm. (b) Cell viability of 3D spheroids. Pitavastatin-induced cell lysis was quantified on days 1, 4, and 7 post-treatment, using trypan blue. Error bars represent standard deviation (n = 3). Date of download: 4/7/2017 Copyright © ASME. All rights reserved. From: Engineering a Three-Dimensional In Vitro Drug Testing Platform for Glioblastoma J. Nanotechnol. Eng. Med. 2016;6(4):041002-041002-6. doi:10.1115/1.4032903 Figure Legend: (a) Time-lapse image of control untreated U87 spheroids and spheroids treated with 10, 50, 100, 150, and 200 μM Irinotecan on days 1, 4, and 7 post-treatment. The spheroids continued to grow for 7 days after treatment with concentrations 10, 50 μM, and control group. At drug concentration 100 μM, the spheroids lost their shape after 7 days while 150 and 200 μM Irinotecan treated spheroid shape starting from day 1. Scale bars represent 200 μm. (b) Cell viability of 3D spheroids. Irinotecan-induced cell lysis was quantified on days 1, 4, and 7 post-treatment, using trypan blue. Error bars represent standard deviation (n = 3). Date of download: 4/7/2017 Copyright © ASME. All rights reserved. From: Engineering a Three-Dimensional In Vitro Drug Testing Platform for Glioblastoma J. Nanotechnol. Eng. Med. 2016;6(4):041002-041002-6. doi:10.1115/1.4032903 Figure Legend: (a) Time-lapse image of U87 spheroids treated with Pita 1 μM + Iri 50 μM, Pita 5 μM + Iri 50 μM, Pita 10 μM + Iri 50 μM, Pita 1 μM + Iri 100 μM, Pita 5 μM + Iri 100 μM, and Pita 10 μM + Iri 100 μM on days 1, 4, and 7 after treatment. Lysis of 3D spheroids treated by combination drug treatments Pita 1 μM + Iri 50 μM and Pita 1 μM + Iri 100 μM on day 7 while at higher concentrations treated spheroid shape changed starting from day 1. Scale bars represent 200 μm. (b) Comparison of cell viability between single drugs (Irinotecan 50 μM and Pitavastatin 10 μM) and combination drug treatment (Pita 5 μM + Iri 50 μM and Pita 10 μM + Iri 50 μM) on 3D spheroids on days 1, 4, and 7 post-treatment, using trypan blue. (c) Comparison of cell viability between single drugs (Irinotecan 100 μM and Pitavastatin 10 μM) and combination drug treatments (Pita 5 μM + Iri 100 μM and Pita 10 μM + Iri 100 μM) on 3D spheroids on days 1, 4, and 7 post-treatment, using trypan blue. Error bars represent standard deviation (n = 3).