**1. Introduction**

The idea that cellular survival and growth could be maintained outside the body was recog‐ nized as possible almost hundred years ago when the German zoologist Wilhelm Roux described a successful experiment where he cultured chick neural crest in warm saline for a few days [1]. Nobel Prize winner Alexis Carrel performed numerous experiments clearly showing that tissue explants, including connective tissue and heart tissue, could be cultured in vitro preserving their characteristics for prolonged periods of time [2] supporting the notion that entire organs could be cultured in vitro. A defined synthetic mixture of amino acids, salts, carbohydrates, vitamins and serum was shown to support cells in vitro[3], thus unifying a major variable in cell culturing experiments and providing a possibility for rapid development of this novel method. Since the establishment of the first cell line by Gey et al.[4] in 1951 cell culturing has become one of the most widely used methods with exceptional contribution to the advances in almost all fields of contemporary biology – cell biology, genetics, cell bio‐ chemistry, physiology etc. Significant progress in the field made possible numerous achieve‐ ments that were believed to be the foundations of personalized medicine. Among these is the isolation of the first line of murine stem cells [5, 6] in 1981, followed by establishment of the first human embryonic stem cell lines by Thompson [7].

Current knowledge of cellular behavior is mainly acquired by studies concerning homogenous populations of cells cultured as monolayers. This simplified approach towards understanding the essence of the mechanisms, underlying the processes determining life and death of a cell has undoubtedly provided scientists with enormous amount of knowledge. However, recent advances in the field of three-dimensional cell cultures have revealed a lot of imperfections in

© 2013 Evangelatov and Pankov; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2013 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

this limiting approach. Growing fibroblasts on flat, rigid surface of the Petri dish or flask results in major differences in adhesion formation and maturation, proliferation, cell signaling, migration and cytoskeletal function, compared to the three-dimensional environment [8-12]. Even changes in stiffness of the two-dimensional (2D) substrate have shown significant difference in fibroblasts response to the environment [13, 14]. Being cultured on 2D surfaces, these non-polar cells are forced to adapt to polar setting, thus significantly changing their response to the environment. One could argue that this culturing method would be ideal for epithelial cells, allowing them to form distinct apical and basal layers. Nevertheless growth in 3D environment allows normal polarization and differentiation of epithelial cells [15] and is a prerequisite for the formation of duct-like structures in vitro [16]. Thereby the emerging differences between conventional, monolayer cell cultures and in vivo cellular behavior led to the increasing number of scientists aiming to provide in vivo-relevant results.

The idea of cell culturing in 3D environment is not a novel one though [17], but until recently the importance of the cell environment – its dimensionality, stiffness, elasticity, composition and remodeling during tissue morphogenesis and disease, remained neglected. Number of studies have shown that knockdown of extracellular matrix (ECM) components like fibronec‐ tin, collagen, laminin, aggrecan, etc. lead to lethal phenotypes or severe pathologies during embryonic or post-natal development [18-24]. Furthermore, recent discoveries like the importance of Cdc42 for the appropriate acquisition of apical and basal polarity during morphogenesis or cell specification during tubulogenesis wouldn't have been possible without the use of 3D model systems [25, 26]. Thus the concept of the importance of the surrounding settings emerged and an entirely new scientific direction evolved, focusing on the role of the extracellular matrix and its importance in cell biology.

In the living organism cells are usually embedded in a complex three-dimensional extracellular matrix that is dynamic in its structure[27] and rarely do have the opportunity to attach to planar, rigid substrates. Reciprocal interactions between cells and the ECM facilitate signaling to and from cells and lead to continuous reorganization of the environment [10]. Investigation of the dynamics of the ECM, its structure in different tissues and cellular response to changes in the mechanical properties of the extracellular matrices have shown that cells are not only able to feel and respond to the environment, but also to cause changes to the environment's mechanical properties [14]. These interactions deliver further signals responsible for cell growth and differentiation, survival, migration and reorganization of the resident tissue [28]. In addition when one thinks of cells in the third dimension it has to be considered that cells are not just randomly incorporated in the ECM but form complex 3D structures, characteristic of the specific tissue or organ [29].

The recognition of the importance of dimensionality of cellular environment encouraged the creation of a variety of three-dimensional culturing systems. Essentially they could be divided into two groups – three-dimensional systems made of artificial materials, mimicking the natural components of the matrix and three-dimensional culturing systems based on natural ECM components, or cell-synthesized ECM. This article's focus will be mainly on the later 3D culturing models.
