Cell culture techniques are ubiquitous in areas of developmental biology, drug discovery, regenerative medicine and protein production. Since the introduction of cell culture techniques, cells have been cultured in two-dimensions, attached to tissue culture plasticware or ECM attachment proteins. Cells in the physiological environment have constant interaction with the extracellular matrix, regulating complex biological functions like cellular migration, apoptosis, transcriptional regulation, and receptor expression1. In-vitro experimental data cannot be translated into clinical trials completely2 when cells are grown in 2D conditions since complicated cellular signals between cells and its matrix cannot be reproduced3. Three-dimensional cell cultures address this challenge and serve as a better model representing in vivo physiological conditions closely. Table 1 indicates the difference between 2D and 3D cell culture systems.
Moreover, several studies have reported a difference in gene and protein expression profiles of the cells grown in 3D environment when compared to its 2D counterpart. Also, expression profiles in 3D culture conditions are thought to be more physiologically relevant than 2D cell culture conditions.
Cellular events in 3D culture resemble physiological conditions closely and have the following distinct advantages over the 2D culture conditions
The choice of 3D cell culture technique should depend on several parameters, including the nature of the cells themselves (cell line, primary cell, tissue origin), or the final aim of the study. It’s crucial to evaluate these parameters before choosing the most relevant 3D cell culture technique.
Broadly, 3D cell culture techniques are classified as Scaffold-based or non-scaffold-based techniques.
In scaffold based techniques cells are grown in presence of a support. 2 major types of support can be used:
Suitability: +++ = High; ++ = Medium; + = Low; - = Unsuitable; +/- = varies with scaffold components
Scaffold-free techniques allow the cells to self-assemble to form non-adherent cell aggregates called spheroids. Spheroids mimic the solid tissues by secreting their own extracellular matrix and displaying differential nutrient availability. Spheroids grown via non-scaffold based techniques are consistent in size and shape and are better in-vitro cellular models for high-throughput screening. Different platforms, from specialized plate to more integrated systems, can be used to generate spheroids: attributes of these techniques are described in the following table.
Protocol Guide: Cancer Stem Cell Tumorsphere Formation Protocol.
Suitability : +++ = High; ++ = Medium; + = Low; - = Unsuitable
The evolution of 3D cell culture has the potential to bridge the gap between in vitro and in vivo experiments. The convenience of handling cells in vitro while obtaining results that reflect in-vivo condition and avoiding ethical concerns of animal usage is making 3D cell culture techniques increasingly popular among researchers, but choosing the right system to develop a 3D cell culture model is not a trivial question.
The future will see the emerging of some more complex and advanced technologies like 3D bioprinting, an offshoot of 3D printing, helpful to print both biomaterials and living cells. 3D bioprinting has a wide medical application like skin grafting, which avoids a second wound site, characteristic of the traditional grafting methods. The major components for 3D bioprinting, like bio-inks, scaffold material, and biomaterials, are relatively well known to the scientific world. By configuring the order and position of these components various tissue products can be developed while simulating the physiological environment19. At the moment, the technique is in the early stage but has the potential to evolve as an indispensable tool for drug discovery and toxicity studies.