**5. Skin cell cultures for psoriasis**

platelets are activated and migrated to the site of injury [57]. The second phase, inflammation, begins about 1 day postinjury and inflammatory mediators such as histamine are released, providing the typical traits of inflammation such as heat and swelling [57]. Proliferation is the phase in which granulation tissue forms at the site of injury and after which re-epithelization occurs [58]. In the last phase, remodeling of the tissue occurs to improve the strength of the skin tissue, which is only ever within 80% of the original tissue's strength [58]. The normal wound healing process could be affected, however, leading to chronic ulcers or excessive wound healing resulting in hypertrophic scars [59]. For this reason, topical agents to improve wound healing or reduce scarring may be of interest and as such *in vitro* testing models for wound healing will

Both 2D and 3D skin cell cultures are available as wound healing models [60]. 2D wound healing models involve the creation of a site of injury in a monolayer of skin cells, either through mechanical or chemical means, in which cells then migrate to the site of injury [60, 61]. Cells in 2D monolayer cultures are thought to adhere to the flat environments, such as a Petri dish, in which they are cultured and will therefore migrate to areas of free space within the dish, an activity thought to mimic *in vivo* migration involved in cell differentiation [62]. One method of mechanical introduction of a wound to a 2D cell culture is through the scratch assay that utilizes materials such as pipette tips or needles to introduce a wound or scratch into the monolayer cell culture [63, 64]. Images of the wound are taken within set time frames to assess the migration of cells [63]. Typically, however, it is difficult to ensure wounds that are equal in size using this method [63], and thus for this reason, testing of pharmaceutical products on these types of cultures are not ideal. 2D skin cell cultures also lack essential functions that could mimic *in vivo* processes, such as immune functionality and blood perfusion [65]. 3D wound healing models have thus attempted to more closely mimic *in vivo* wound healing processes and are available

Histocultures are cultures of intact tissues, consisting of more than one type of skin cell, such as neutrophils and other cells involved in wound healing, and are thus able to better mimic *in vivo* skin responses of wound healing [42, 43]. HSEs, on the other hand, are 3D cell culture models created from various human skin cells and materials that mimic the extracellular matrix [45] and are created as either epidermal equivalents, dermal equivalents or skin equivalents consisting of both layers [8, 45]. Some examples of 3D wound healing skin cell

A 3D wound healing skin equivalent developed by Herman et al. [66] included capillary endothelial cells which were capable of creating a similar formation to *in vivo* capillaries in the skin. This wound healing model involved several different cells, such as keratinocytes and epithelial cells, and used 3D matrices composed of Matrigel™ and collagen [66]. As angiogenesis is involved in the process of wound healing [66], this model is an excellent example of improved 3D wound healing cell culturing protocols to more closely mimic *in vivo* wound

Another example of a 3D skin cell culture model that could be used for wound healing was reported by Sidgwick et al. [67]. This model allows for the immersion of biopsies in Williams E culture media with the epidermal layer of the skin uncovered and uses whole tissue biopsies

be discussed.

10 Cell Culture

as histocultures or HSEs.

healing processes.

cultures and HSEs are described below.

As an inflammatory skin condition that involves the immune system, psoriasis has the characteristic appearance of silver scales that arise from increased proliferation of the keratinocytes in the epidermal layer [68, 69]. Psoriasis is typically treated with topical steroid medications or with vitamin D analogue medications such as calcipotriol as well as with moisturizing agents [68]. The treatment of psoriasis also includes systemic medications that suppress the immune system such as methotrexate and cyclosporine as well as biologic drugs [68, 70].

Psoriasis has been linked to various mental health illnesses such as anxiety and depression which are thought to result from having a chronic visible skin condition [71], and therefore the need for developing new pharmaceutical products and testing of these products is evident. As psoriasis is primarily treated with topical medications [70], having *in vitro* cell cultures or models for testing the safety and efficacy of these topical medications is essential.

Skin cell cultures that are 2D for dermatological conditions such as psoriasis and other autoimmune disorders exist [39]. 2D cell cultures of psoriasis initially consisted of psoriatic keratinocytes; however, this proved to be an ineffective model for psoriasis as the cells were not able to grow and lost their psoriatic genes with time [72]. As a result, alternative methods such as adding cytokines to normal human keratinocytes to induce psoriatic features were developed [73]. As stated previously, however, testing of medications on 2D cell culture models does not always translate to *in vivo* responses [7], and thus, 3D models to better mimic *in vivo* responses for testing of topical medications for psoriasis have been developed.

Barker et al. [74] introduce an *in vitro* model of psoriatic human skin, whereby a monolayer cell culture of psoriatic keratinocytes on top of collagen and fibroblasts from the dermis was created, onto which an epidermal layer was formed approximately 3 weeks later. Similarly, a 3D skin substitute using psoriatic skin cells was developed by Jean et al. [75] with either both psoriatic keratinocytes and fibroblasts or only one type of psoriatic cells (either keratinocytes or fibroblasts were psoriatic). Skin biopsies were obtained from patients with plaque psoriasis, and keratinocytes were extracted and seeded on to mouse fibroblasts and incubated at a temperature of 37°C [75]. For skin substitutes, ascorbic acid was used to culture the fibroblasts which formed dermal sheets that were altered to create a dermal layer onto which keratinocytes were then seeded to form an epidermal layer [75]. This method is the self-assembly method as the fibroblasts release their own extracellular matrix to maintain their growth [75]. A serum-free 3D psoriatic skin cell culture was similarly developed by Duque-Fernandez et al. [66]. This method is similar to the one presented by Jean et al. [75] and may be due to the fact that some authors are the same in both articles. The model by Duque-Fernandez et al. [76] was even used for testing percutaneous permeation of benzoic acid, caffeine, and hydrocortisone, using a Franz-diffusion cell method. This was compared to healthy skin substitutes and revealed that the psoriatic skin substitute had a greater permeability response to the three compounds [76].

cultures, [8] or the lack of interactions in 2D cell cultures of cell-to-cell or cell-to-extracellular matrix may be possible causes [5, 80]. However, advantages to using a 2D cell culture include

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

3D cell cultures, on the other hand, provide a better imitation of *in vivo* processes as they allow for better understanding of the interactions that take place between cell-to-cell and cell-to-extracellular matrix [7]. Cell cultures that are 3D have also revealed to possess gene expression abilities, thus mimicking *in vivo* processes [83]. However, regardless of their improvements vs. the 2D cell cultures, typically 3D cell cultures still lack in their ability to provide key functions such as cellular waste removal [7]. Microfabricated cell cultures or on-chip cell cultures are advantageous in this instance as they provide microfluidic channels that enable the flow of nutrients and waste removal as well as the ability to mimic vasculature [51, 52]. Thus microfabricated 3D cell cultures are arguably the *in vitro* method of cell cultures that most closely mimic *in vivo* processes [50]. With respect to skin cell cultures, those that are 3D have the advantages of containing a stratum corneum and thus the ability of testing pharmaceutical products on the stratum corneum

ease of use, cost, and abundant scientific literature surrounding its use [81, 82].

**Advantages Disadvantages**

• Do not always provide an accurate representation of *in vivo* processes [7] • Use of a single cell type, [80] for example with skin cell cultures, the use of keratino-

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

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• No cell-to-cell or cell-to-extracellular matrix interactions [5, 80]

• Typically lack in their ability to provide key functions such as cellular waste

• Cost associated with developing cultures

cytes only [8]

removal [7]

[40, 54]

• Ease of use, cost and abundant scientific literature

• Better understanding of the interactions that take place between cell-to-cell and cell-to-extracellular

• Possess gene expression abilities, thus mimicking *in* 

• Microfabricated cell cultures or on-chip cell cultures provide microfluidic channels that enable the flow of nutrients and waste removal as well as the ability

• 3D skin cultures have a stratum corneum and thus the ability of testing pharmaceutical products on the

• Longer skin cell culture use between 10 and 30 days [40]; more recent human skin equivalent (HSE) cultures can even be used for up to 20 weeks [41] • 3D bioprinting provides greater accuracy in placement of cells and extracellular matrix, potential of imbedding vasculature in the skin construct and

• 3D bioprinted skin models provide more uniform models vs. manually developed skin models [55]

**Table 1.** Summary of the advantages and disadvantages of 2D vs. 3D cell cultures.

stratum corneum as a skin barrier [40]

surrounding use [81, 82]

matrix [7]

*vivo* processes [83]

to mimic vasculature [51, 52]

have great plasticity [54]

• Better imitation of *in vivo* processes [7]

2D cell cultures

3D cell cultures

For safety and efficacy testing of medications for psoriasis, commercially available 3D *in vitro* models also exist. Two examples of such models are the Psoriasis Skin Model by Creative Bioarray [77] and the model by MatTek Corporation [78]. According to Creative Bioarray [77], their skin model more closely mimics *in vivo* psoriatic skin vs. typical cell cultures as it allows for cell differentiation and can be used for testing of pharmaceutical products. The psoriasis model by MatTek Corporation can also be used for testing of dermatological products as it a 3D model composed of psoriatic fibroblasts while using normal human epidermal keratinocytes and is capable of mimicking *in vivo* psoriatic responses such as basal cell proliferation [78]. These examples were presented for a better understanding of the methods used in developing psoriatic human skin equivalents as well as the commercially available models for testing the safety and efficacy of dermatological products used for psoriasis.
