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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Comparative Analysis of 3D Bladder Tumor Spheroids Obtained by Forced Floating and Hanging Drop Methods for Drug Screening

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Author(s):
Amaral, Robson L. F. ; Miranda, Mariza ; Marcato, Priscyla D. ; Swiech, Kamilla
Total Authors: 4
Document type: Journal article
Source: FRONTIERS IN PHYSIOLOGY; v. 8, AUG 22 2017.
Web of Science Citations: 12
Abstract

Introduction: Cell-based assays using three-dimensional (3D) cell cultures may reflect the antitumor activity of compounds more accurately, since these models reproduce the tumor microenvironment better. Methods: Here, we report a comparative analysis of cell behavior in the two most widely employed methods for 3D spheroid culture, forced floating (Ultra-low Attachment, ULA, plates), and hanging drop (HD) methods, using the RT4 human bladder cancer cell line as a model. The morphology parameters and growth/metabolism of the spheroids generated were first characterized, using four different cell-seeding concentrations (0.5, 1.25, 2.5, and 3.75 x 10(4) cells/mL), and then, subjected to drug resistance evaluation. Results: Both methods generated spheroids with a smooth surface and round shape in a spheroidization time of about 48 h, regardless of the cell-seeding concentration used. Reduced cell growth and metabolism was observed in 3D cultures compared to two-dimensional (2D) cultures. The optimal range of spheroid diameter (300-500 mu m) was obtained using cultures initiated with 0.5 and 1.25 x 10(4) cells/mL for the ULA method and 2.5 and 3.75 x 10(4) cells/mL for the HD method. RT4 cells cultured under 3D conditions also exhibited a higher resistance to doxorubicin (IC50 of 1.00 and 0.83 mu g/mL for the ULA and HD methods, respectively) compared to 2D cultures (IC50 ranging from 0.39 to 0.43). Conclusions: Comparing the results, we concluded that the forced floating method using ULA plates was considered more suitable and straightforward to generate RT4 spheroids for drug screening/cytotoxicity assays. The results presented here also contribute to the improvement in the standardization of the 3D cultures required for widespread application. (AU)

FAPESP's process: 15/05102-1 - Comparative analysis of different three-dimensional culture systems to establish an in vitro bladder cancer model for toxicological assays
Grantee:Robson Luis Ferraz do Amaral
Support type: Scholarships in Brazil - Doctorate (Direct)