Comparative studies of cellular viability levels on 2D and 3D in vitro culture matrices
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In this study, the cellular viability and function of immortalized human cervical and dermal cells are monitored and compared in conventional 2D and two commercial 3D membranes, Collagen and Geltrex, of varying working concentration and volume. Viability was monitored with the aid of the Alamar Blue assay, cellular morphology was monitored with confocal microscopy, and cell cycle studies and cell death mechanism studies were performed with flow cytometry. The viability studies showed apparent differences between the 2D and 3D culture systems, the differences attributed in part to the physical transition from 2D to 3D environment causing alterations to effective resazurin concentration, uptake and conversion rates, which was dependent on exposure time, but also due to the effect of the membrane itself on cellular function. These effects were verified by flow cytometry, in which no significant differences in viable cell numbers between 2D and 3D systems were observed after 24 h culture. The results showed the observed effect was different after shorter exposure periods, was also dependent on working concentration of the 3D system and could be mediated by altering the culture vessel size. Cell cycle analysis revealed cellular function could be altered by growth on the 3D substrates and the alterations were noted to be dependent on 3D membrane concentration. The use of 3D culture matrices has been widely interpreted to result in “improved viability levels” or “reduced” toxicity or cellular “resistance” compared to cells cultured on traditional 2D systems. The results of this study show that cellular health and viability levels are not altered by culture in 3D environments, but their normal cycle can be altered as indicated in the cell cycle studies performed and such variations must be accounted for in studies employing 3D membranes for in vitro cellular screening.
KeywordsCollagen I Geltrex® 3D matrices Confocal microscopy In vitro screening
This study were funded by the Government of Libya for M. Gargotti and Consejo Nacional de Cienciay Tecnología, Mexico for U. Lopez-Gonzalez and this work has been enabled by Science Foundation Ireland Principle Investigator Award 11/PI/1108.
- Cody D, Casey A, Naydenova I, Mihaylova E (2013) A comparative cytotoxic evaluation of acrylamide and diacetone acrylamide to investigate their suitability for holographic photopolymer formulations. Int J Polym Sci 2013. doi: 10.1155/2013/564319
- European Union—Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010Google Scholar
- Gilbert TW, Sellaro TL, Badylak SF (2006) Decellularization of tissues and organs. Biomaterials 27:3675–3683Google Scholar
- Riss T (2014) Overview of 3D Cell culture model systems validating cell-based assays for use with 3D cultures [PowerPoint slides]. Retrieved from https://worldwide.promega.com/-/media/files/promega-worldwide/north-america/promega-us/webinars-and-events/2014/3d-cell-culture-webinar-march-2014.pdf?la=en
- Stacey G, Bar P, Granville R (2009) Primary cell cultures and immortal cell lines. Encycl Life Sci, pp. 1–6. http://orca.cf.ac.uk/24620/
- Vega-Avila E, Pugsley MK (2011) An overview of colorimetric assay methods used to assess survival or proliferation of mammalian cells. Proc West Pharmacol Soc 54:10–14Google Scholar
- Worthington P, Pochan DJ, Langhans SA (2015) Peptide hydrogels—versatile matrices for 3D cell culture in cancer medicine. Front Oncol 2015:92Google Scholar