Is It Time to Start Transitioning From 2D to 3D Cell Culture?

Cell culture is an important and necessary process in drug discovery, cancer research, as well as stem cell study. Most cells are currently cultured using two-dimensional (2D) methods but new and improved methods that implement three-dimensional (3D) cell culturing techniques suggest compelling evidence that much more advanced experiments can be performed yielding valuable insights. When performing 3D cell culture experiments, the cell environment can be manipulated to mimic that of a cell in vivo and provide more accurate data about cell-to-cell interactions, tumor characteristics, drug discovery, metabolic profiling, stem cell research, and other types of diseases. Scaffold based techniques such as hydrogel-based support, polymeric hard material-based support, hydrophilic glass fiber, and organoids are employed, and each provide their own advantages and applications. Likewise, there are also scaffold free techniques used such as hanging drop microplates, magnetic levitation, and spheroid microplates with ultra-low attachment coating. 3D cell culture has the potential to provide alternative ways to study organ behavior via the use of organoids and is expected to eventually bridge the gap between 2D cell culture and animal models. The present review compares 2D cell culture to 3D cell culture, provides the details surrounding the different 3D culture techniques, as well as focuses on the present and future applications of 3D cell culture.

[1]  Benjamin S. Freedman,et al.  Organoid cystogenesis reveals a critical role of microenvironment in human polycystic kidney disease , 2017, Nature materials.

[2]  Anandika Dhaliwal,et al.  Three Dimensional Cell Culture : A Review , 2012 .

[3]  Kin Fong Lei,et al.  Real-Time Monitoring of Ascorbic Acid-Mediated Reduction of Cytotoxic Effects of Analgesics and NSAIDs on Tenocytes Proliferation , 2019, Dose-response : a publication of International Hormesis Society.

[4]  Anh-Vu Do,et al.  3D Printing of Scaffolds for Tissue Regeneration Applications , 2015, Advanced healthcare materials.

[5]  Georges Noel,et al.  Three-Dimensional Cell Culture: A Breakthrough in Vivo , 2015, International journal of molecular sciences.

[6]  Magdi H. Yacoub,et al.  Hydrogel scaffolds for tissue engineering: Progress and challenges , 2013, Global cardiology science & practice.

[7]  Apurva R. Patel,et al.  AlgiMatrix™ Based 3D Cell Culture System as an In-Vitro Tumor Model for Anticancer Studies , 2013, PloS one.

[8]  Yong Teng,et al.  Simultaneously inactivating Src and AKT by saracatinib/capivasertib co-delivery nanoparticles to improve the efficacy of anti-Src therapy in head and neck squamous cell carcinoma , 2019, Journal of Hematology & Oncology.

[9]  Bei Peng,et al.  New advances in microfluidic flow cytometry , 2018, Electrophoresis.

[10]  Chun-Ting Lee,et al.  3D brain Organoids derived from pluripotent stem cells: promising experimental models for brain development and neurodegenerative disorders , 2017, Journal of Biomedical Science.

[11]  Yong Teng,et al.  Nck-associated protein 1 associates with HSP90 to drive metastasis in human non-small-cell lung cancer , 2019, Journal of Experimental & Clinical Cancer Research.

[12]  Thomas Geiser,et al.  A lung-on-a-chip array with an integrated bio-inspired respiration mechanism. , 2015, Lab on a chip.

[13]  Subhas C. Kundu,et al.  A Non‐Mulberry Silk Fibroin Protein Based 3D In Vitro Tumor Model for Evaluation of Anticancer Drug Activity , 2012 .

[14]  Margaret Nowicki,et al.  Fabrication of a Highly Aligned Neural Scaffold via a Table Top Stereolithography 3D Printing and Electrospinning. , 2016, Tissue engineering. Part A.

[15]  S. L. Forsberg,et al.  Epigenetics and cerebral organoids: promising directions in autism spectrum disorders , 2018, Translational Psychiatry.

[16]  Zoe Cesarz,et al.  Spheroid Culture of Mesenchymal Stem Cells , 2015, Stem cells international.

[17]  S. Richon,et al.  A three-dimensional tumor cell defect in activating autologous CTLs is associated with inefficient antigen presentation correlated with heat shock protein-70 down-regulation. , 2003, Cancer research.

[18]  Ümit Hakan Yildiz,et al.  Biomimetic hybrid scaffold consisting of co-electrospun collagen and PLLCL for 3D cell culture. , 2019, International journal of biological macromolecules.

[19]  James A Bankson,et al.  Three-dimensional tissue culture based on magnetic cell levitation. , 2010, Nature nanotechnology.

[20]  Z. Werb,et al.  Remodelling the extracellular matrix in development and disease , 2014, Nature Reviews Molecular Cell Biology.

[21]  Sharanya Sankar,et al.  Enhanced osteodifferentiation of MSC spheroids on patterned electrospun fiber mats - An advanced 3D double strategy for bone tissue regeneration. , 2019, Materials science & engineering. C, Materials for biological applications.

[22]  N. Peppas,et al.  Hydrogels in Pharmaceutical Formulations , 1999 .

[23]  L. O’Driscoll,et al.  Three-dimensional cell culture: the missing link in drug discovery. , 2013, Drug discovery today.

[24]  Roger D. Kamm,et al.  A 3D neurovascular microfluidic model consisting of neurons, astrocytes and cerebral endothelial cells as a blood-brain barrier. , 2017, Lab on a chip.

[25]  Donald E Ingber,et al.  Modeling radiation injury-induced cell death and countermeasure drug responses in a human Gut-on-a-Chip , 2018, Cell Death & Disease.

[26]  E. Danen,et al.  3D Cell-Based Assays for Drug Screens: Challenges in Imaging, Image Analysis, and High-Content Analysis , 2019, SLAS discovery : advancing life sciences R & D.

[27]  Hiroaki Onoe,et al.  ECM-based Stretchable Microfluidic System for in vitro 3D Tissue Culture , 2019, 2019 20th International Conference on Solid-State Sensors, Actuators and Microsystems & Eurosensors XXXIII (TRANSDUCERS & EUROSENSORS XXXIII).

[28]  Liesbet Geris,et al.  Towards Self-Regulated Bioprocessing: A Compact Benchtop Bioreactor System for Monitored and Controlled 3D Cell and Tissue Culture. , 2019, Biotechnology journal.

[29]  Donald E Ingber,et al.  Organ‐on‐Chip Recapitulates Thrombosis Induced by an anti‐CD154 Monoclonal Antibody: Translational Potential of Advanced Microengineered Systems , 2018, Clinical pharmacology and therapeutics.

[30]  Sigrid A. Langhans Three-Dimensional in Vitro Cell Culture Models in Drug Discovery and Drug Repositioning , 2018, Front. Pharmacol..

[31]  P. Ma,et al.  Polymeric Scaffolds for Bone Tissue Engineering , 2004, Annals of Biomedical Engineering.

[32]  Peter X. Ma,et al.  Scaffolds for tissue fabrication , 2004 .

[33]  Jian-wen Liu,et al.  A collagen-based multicellular tumor spheroid model for evaluation of the efficiency of nanoparticle drug delivery , 2016, Artificial cells, nanomedicine, and biotechnology.

[34]  David R. K. Harding,et al.  Synthesis, characterisation and evaluation of poly[lactose acrylate-N-vinyl-2-pyrrolidinone] hydrogels for drug delivery , 2003 .

[35]  Manoel Luis Costa,et al.  2D and 3D-Organized Cardiac Cells Shows Differences in Cellular Morphology, Adhesion Junctions, Presence of Myofibrils and Protein Expression , 2012, PloS one.

[36]  Jacqueline Alblas,et al.  Cellular immunotherapy on primary multiple myeloma expanded in a 3D bone marrow niche model , 2018, Oncoimmunology.

[37]  Lee L. Rubin,et al.  Large-Scale Production of Mature Neurons from Human Pluripotent Stem Cells in a Three-Dimensional Suspension Culture System , 2016, Stem cell reports.

[38]  Chia Chia Ong,et al.  Effect of Modified Solvent Casting/Particulate Leaching (SCPL) Technique on the Properties of Bioactive Glass Reinforced Polyurethane Scaffold for Biomedical Applications , 2019, Journal of Physical Science.

[39]  David A Ferrick,et al.  Advances in measuring cellular bioenergetics using extracellular flux. , 2008, Drug discovery today.

[40]  G. Souza,et al.  Engineering innervated secretory epithelial organoids by magnetic three-dimensional bioprinting for stimulating epithelial growth in salivary glands. , 2018, Biomaterials.

[41]  Jeffrey M Karp,et al.  Engineering Stem Cell Organoids. , 2016, Cell stem cell.

[42]  George A. Truskey,et al.  Modeling the mitochondrial cardiomyopathy of Barth syndrome with iPSC and heart-on-chip technologies , 2014, Nature Medicine.

[43]  Daniel R. Berger,et al.  Cell diversity and network dynamics in photosensitive human brain organoids , 2017, Nature.

[44]  Kelly Johanson,et al.  Diamagnetic levitation changes growth, cell cycle, and gene expression of Saccharomyces cerevisiae , 2007, Biotechnology and bioengineering.

[45]  Lin Lu,et al.  Three-dimensional cell culture: A powerful tool in tumor research and drug discovery. , 2017, Oncology letters.

[46]  Felicia Fält,et al.  Unique animal friendly 3D culturing of human cancer and normal cells. , 2019, Toxicology in vitro : an international journal published in association with BIBRA.

[47]  John W Haycock,et al.  3D cell culture: a review of current approaches and techniques. , 2011, Methods in molecular biology.

[48]  Alan Wells,et al.  Liver ‘organ on a chip’ , 2017, Experimental cell research.

[49]  Cheng Zhang,et al.  Development of a primary human Small Intestine-on-a-Chip using biopsy-derived organoids , 2018, Scientific Reports.

[50]  Julien Picot,et al.  Flow cytometry: retrospective, fundamentals and recent instrumentation , 2012, Cytotechnology.

[51]  Hwan-You Chang,et al.  Recent advances in three‐dimensional multicellular spheroid culture for biomedical research , 2008, Biotechnology journal.

[52]  Juergen A. Knoblich,et al.  Organogenesis in a dish: Modeling development and disease using organoid technologies , 2014, Science.

[53]  William L. Haisler,et al.  Three-dimensional cell culturing by magnetic levitation , 2013, Nature Protocols.

[54]  M. Bissell,et al.  Interaction with basement membrane serves to rapidly distinguish growth and differentiation pattern of normal and malignant human breast epithelial cells. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[55]  K. Anseth,et al.  Hydrogel Cell Cultures , 2007, Science.

[56]  Deepak Choudhury,et al.  A pump‐free tricellular blood–brain barrier on‐a‐chip model to understand barrier property and evaluate drug response , 2019, Biotechnology and bioengineering.

[57]  A. E. Rossi,et al.  A Novel Three-Dimensional Immune Oncology Model for High-Throughput Testing of Tumoricidal Activity , 2018, Front. Immunol..

[58]  Michael R Rosen,et al.  Mesenchymal Stem Cells Support Migration, Extracellular Matrix Invasion, Proliferation, and Survival of Endothelial Cells In Vitro , 2007, Stem cells.

[59]  Massimo Messori,et al.  Development of solvent-casting particulate leaching (SCPL) polymer scaffolds as improved three-dimensional supports to mimic the bone marrow niche. , 2019, Materials science & engineering. C, Materials for biological applications.

[60]  Sidra Waheed,et al.  3D printed microfluidic devices: enablers and barriers. , 2016, Lab on a chip.

[61]  Rémi Courson,et al.  Fabrication of 3D scaffolds reproducing intestinal epithelium topography by high-resolution 3D stereolithography. , 2019, Biomaterials.

[62]  Drosophila M Brumby,et al.  Modelling Cancer in , 2008 .

[63]  Chung-Ho Sun,et al.  Modeling aberrant wound healing using tissue-engineered skin constructs and multiphoton microscopy. , 2004, Archives of facial plastic surgery.

[64]  Dana M Spence,et al.  PolyJet 3D-Printed Enclosed Microfluidic Channels without Photocurable Supports. , 2019, Analytical chemistry.

[65]  M J Bissell,et al.  Influence of a reconstituted basement membrane and its components on casein gene expression and secretion in mouse mammary epithelial cells. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[66]  Robert J Gillies,et al.  Metabolic Profiling of healthy and cancerous tissues in 2D and 3D , 2017, Scientific Reports.

[67]  Kin Fong Lei,et al.  Real-time and non-invasive impedimetric monitoring of cell proliferation and chemosensitivity in a perfusion 3D cell culture microfluidic chip. , 2014, Biosensors & bioelectronics.

[68]  Yuejun Kang,et al.  Freeze-drying prepared ready-to-use gelatin @polypropylene nonwoven hybrid sheet for stacking 3D cell culture , 2019, Cellulose.

[69]  Kang Zhang,et al.  3D printing of functional biomaterials for tissue engineering. , 2016, Current opinion in biotechnology.

[70]  Allan S Hoffman,et al.  Hydrogels for biomedical applications. , 2002, Advanced drug delivery reviews.

[71]  Andrea Pavesi,et al.  Characterizing the Role of Monocytes in T Cell Cancer Immunotherapy Using a 3D Microfluidic Model , 2018, Front. Immunol..

[72]  L P Ferreira,et al.  Design of spherically structured 3D in vitro tumor models -Advances and prospects. , 2018, Acta biomaterialia.

[73]  Xin Zhao,et al.  Metastasis-on-a-chip mimicking the progression of kidney cancer in the liver for predicting treatment efficacy , 2020, Theranostics.

[74]  D. Harding,et al.  Preparation and in-vitro evaluation of poly[N-vinyl-2-pyrrolidone-polyethylene glycol diacrylate]-chitosan interpolymeric pH-responsive hydrogels for oral drug delivery. , 2000, International journal of pharmaceutics.

[75]  LeeSe-Jun,et al.  Fabrication of a Highly Aligned Neural Scaffold via a Table Top Stereolithography 3D Printing and Electrospinning . , 2016 .

[76]  Donald E. Ingber,et al.  Modelling cancer in microfluidic human organs-on-chips , 2019, Nature Reviews Cancer.

[77]  O. Okay General Properties of Hydrogels , 2009 .

[78]  Richard Novak,et al.  Matched-Comparative Modeling of Normal and Diseased Human Airway Responses Using a Microengineered Breathing Lung Chip. , 2016, Cell systems.

[79]  T. Maekawa,et al.  POLYMERIC SCAFFOLDS IN TISSUE ENGINEERING APPLICATION: A REVIEW , 2011 .

[80]  J. Collins,et al.  Contributions of microbiome and mechanical deformation to intestinal bacterial overgrowth and inflammation in a human gut-on-a-chip , 2015, Proceedings of the National Academy of Sciences.

[81]  M J Bissell,et al.  Functional differentiation and alveolar morphogenesis of primary mammary cultures on reconstituted basement membrane. , 1989, Development.

[82]  Ernesto Reverchon,et al.  Biodegradable synthetic scaffolds for tendon regeneration. , 2012, Muscles, ligaments and tendons journal.

[83]  T. Mukohara,et al.  Comparison of 2D- and 3D-culture models as drug-testing platforms in breast cancer. , 2015, Oncology reports.

[84]  Donald E Ingber,et al.  Human Organ Chip Models Recapitulate Orthotopic Lung Cancer Growth, Therapeutic Responses, and Tumor Dormancy In Vitro. , 2017, Cell reports.

[85]  Dagmar Kulms,et al.  A 3D Organotypic Melanoma Spheroid Skin Model , 2018, Journal of visualized experiments : JoVE.

[86]  Wenxin Wang,et al.  Application of a microfluidic chip-based 3D co-culture to test drug sensitivity for individualized treatment of lung cancer. , 2013, Biomaterials.

[87]  Younan Xia,et al.  Electrospinning and Electrospun Nanofibers: Methods, Materials, and Applications. , 2019, Chemical reviews.

[88]  Rudolf Hausmann,et al.  Beyond bread and beer: whole cell protein extracts from baker’s yeast as a bulk source for 3D cell culture matrices , 2017, Applied Microbiology and Biotechnology.

[89]  Tim D Holmes,et al.  A Human NK Cell Activation/Inhibition Threshold Allows Small Changes in the Target Cell Surface Phenotype To Dramatically Alter Susceptibility to NK Cells , 2011, The Journal of Immunology.

[90]  Maddaly Ravi,et al.  3D Cell Culture Systems: Advantages and Applications , 2015, Journal of cellular physiology.

[91]  Vernella Vickerman,et al.  Design, fabrication and implementation of a novel multi-parameter control microfluidic platform for three-dimensional cell culture and real-time imaging. , 2008, Lab on a chip.

[92]  John M. Walker,et al.  Metabolic Profiling , 2011, Methods in Molecular Biology.

[93]  F. Sonntag,et al.  A four-organ-chip for interconnected long-term co-culture of human intestine, liver, skin and kidney equivalents. , 2015, Lab on a chip.

[94]  Tetsuya Nakamura,et al.  Transplantation of Expanded Fetal Intestinal Progenitors Contributes to Colon Regeneration after Injury , 2013, Cell stem cell.

[95]  Ali Khademhosseini,et al.  Controlling the porosity and microarchitecture of hydrogels for tissue engineering. , 2010, Tissue engineering. Part B, Reviews.

[96]  Dana M. Spence,et al.  Review of 3D Cell Culture with Analysis in Microfluidic Systems. , 2019, Analytical methods : advancing methods and applications.

[97]  K. Lamperska,et al.  2D and 3D cell cultures – a comparison of different types of cancer cell cultures , 2016, Archives of medical science : AMS.

[98]  Liju Yang,et al.  Three-dimensional cell culture systems and their applications in drug discovery and cell-based biosensors. , 2014, Assay and drug development technologies.

[99]  Gaoyan Zhong,et al.  Scaffold Techniques and Designs in Tissue Engineering Functions and Purposes: A Review , 2019, Advances in Materials Science and Engineering.

[100]  Uwe Marx,et al.  Bone marrow-on-a-chip: Long-term culture of human hematopoietic stem cells in a 3D microfluidic environment , 2017 .

[101]  Shuichi Takayama,et al.  High-throughput 3D spheroid culture and drug testing using a 384 hanging drop array. , 2011, The Analyst.

[102]  Vicky M. Avery,et al.  Advanced Cell Culture Techniques for Cancer Drug Discovery , 2014, Biology.

[103]  Qixu Zhang,et al.  Human decellularized adipose tissue scaffold as a model for breast cancer cell growth and drug treatments. , 2014, Biomaterials.

[104]  Vítor M Gaspar,et al.  3D tumor spheroids: an overview on the tools and techniques used for their analysis. , 2016, Biotechnology advances.