**1. Introduction**

The growth of cells in an environment outside of an organism's body is referred to as a cell culture [1, 2]. Cells obtained from human tissues are often used for cell cultures to mimic the physiological and metabolic functions of humans for *in vitro* studies [3]. Cell cultures are

© 2016 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. © 2018 The Author(s). Licensee IntechOpen. 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.

maintained in controlled environments and under controlled temperatures for cells to maintain their ability to survive and to grow [3]. For this reason, a medium for the cells to obtain the required nutrients for their growth is essential [2].

medium in 2D cell cultures initially consisted of blood plasma; however, over the years, synthetic medium with the right amount of nutrients and even antifungal and antibiotic ingredients have been developed [11]. Cell culture medium is expected to contain amino acids, a buffering system, vitamins, trace elements and more, to maintain cell viability [12]. Eagle's minimum essential medium (EMEM) and Dulbecco's modified Eagle's medium (DMEM) are two examples of cell culture media that have been developed [13]. Different cell types may

2D vs. 3D Cell Culture Models for *In Vitro* Topical (Dermatological) Medication Testing

http://dx.doi.org/10.5772/intechopen.79868

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The very first cell cultures developed were derived from human skin and were of primarily keratinocytes [5, 8, 9]. As the largest organ in the human body, [14] the skin consists of the epidermal and dermal layers [14, 15]. In the epidermal layer, the main cell types are keratinocytes, while fibroblasts are the main cell types in the dermal layer of the skin [10]. Initially, 2D skin cell cultures consisted primarily of keratinocytes [8, 9]. It was however determined that fibroblasts were essential for the growth of keratinocyte cultures [16] as fibroblasts are the cells responsible for secreting substances of the extracellular matrix such as collagen [17]. The primary fibroblast cell line, cultured by George Todardo and Howard Green, is known as the 3T3 cell line and was named based on its culturing method of transferring cultures every 3 days [16, 18]. The 3T3 cell line is typically used as a feeder layer in keratinocyte cultures [18]. HaCaT cells are a cell line

Prior to animal testing and clinical trials, traditionally, when testing pharmaceutical ingredients, the first step has been to test *in vitro* using 2D cell cultures [7]. This has however provided limitations with respect to testing as the results obtained from *in vitro* 2D cell culture studies have not translated to *in vivo* studies [7]. For this reason, *in vitro* 3D models have become increasingly popular as they are considered to more closely resemble *in vivo* processes [11, 20].

Cell cultures that are 3D involve cells that are combined and shaped into a 3D form using a surrounding medium or specialized condition to help maintain the shape [7]. The equipment used for 2D cell cultures such as a biosafety hood, cell incubator with a temperature of 37°C

used for 3D cell cultures. Supplies used for 3D cultures which differ from those in 2D cell cultures include the matrices, scaffolds, and proprietary plasticware for aggregate formation (e.g., AlgiMatrix, microplates, and multidishes) [21]. Other supplies used in 3D cell cultures may include cell reagents for determining cell health and viability, such as the live/dead Viability/Cytotoxicity Kit and CellTracker Deep Red Dye by ThermoFisher Scientific [21].

The techniques used in creating 3D cell cultures are typically divided into methods that use a scaffold and methods that do not (**Figure 1**) [22]. Methods that do not use a scaffold are typically considered to be a better representation of *in vivo* activity as the cells in this method aggregate on their own [22]. When cells come together and aggregate, they are referred to as

, refrigerator for culture medium storage and other supplies [2, 3] may also be

require different culture media, and thus, the nutrient requirements may vary [13].

**2.1. 2D skin cell cultures**

of keratinocytes [19].

**3. 3D cell cultures**

and 95% O2

Cells obtained directly from living tissues and then cultured are referred to as primary cell cultures [4]. These cells have a short time frame of survival and scientific use [4]. Cells that have been modified to survive indefinitely with continuous cell division are referred to as either continuous cell lines [1] or immortalized cell lines [2]. Cancer cells are an example of the type of cells that may be used to create cell lines due to their genetic mutations and ability to divide continuously [2].

The very first cell cultures developed were of human skin which allowed for a better understanding of the physiological functions of the skin [5]. Cell cultures are available today in twodimensional (2D) and three-dimensional (3D) forms. Cell cultures that grow in controlled flat environments, such as a Petri dish, are 2D cell cultures [6]. Cell cultures that are 3D, however, combine and shape cells into a 3D form using a surrounding medium or specialized conditions to help maintain a 3D figure [7]. *In vitro* skin cell cultures developed initially were primarily 2D with keratinocytes as the primary cell types [8, 9]. In the epidermal layer, the main cell types are keratinocytes, while fibroblasts are the main cell types in the dermal layer of the skin [10]. It is thus evident that an *in vitro* model consisting of both keratinocytes and fibroblasts is required to better mimic the physiological functions of the human skin, especially with relation to the wound healing properties of the skin [5]. There are therefore ongoing efforts to develop various *in vitro* skin models that more closely resemble the skin's physiological functions and that will allow for testing of dermatological products [8]. To have a better understanding of the type of cell cultures used for testing dermatological products, this chapter aims to outline the differences between 2D and 3D skin cell cultures while considering the advantages and disadvantages of each culture. Different types of cell cultures used for wound healing and for inflammatory skin conditions such as psoriasis will also be discussed.
