TESTING 27
3D skin models for inflammation & pigmentation
n Dr Michel Salmon - StratiCELL, Belgium
Monolayer skin cell cultures (keratinocytes, fibroblasts, melanocytes, etc.) provide us with valuable tools for the evaluation of therapeutic or dermo-cosmetic compounds, particularly in large-scale screening studies. The cost of their use is low, especially if one can work on immortalised lines, in simple and inexpensive culture media. Depending on the nature of the studies, they have limitations and do not always accurately represent the situation encounteredin vivo. A culture of differentiated keratinocytes in a monolayer, for example, is not suitable for evaluating the effect of active compounds on the barrier function of the epidermis, which needs a stratum corneum. Furthermore, they do not allow topical contact with finished products or liposoluble compounds. Their environment is also very simple and different from the skin, in terms of mechanical forces, spatial orientation, pH and oxygen gradient, and interactions with the extracellular matrix. There is therefore a real need for models
that are closer to skin physiology, representative of epidermal differentiation and stratification, and that are more predictive for active ingredients in the preclinical phase, for the purposes of cosmetic objectification or toxicological evaluation. The beginnings of 3D reconstructed skin
models as we know them today originate from the work of James Rheinwald and Howard Green in the late 1970s, who first succeeded in growing a sheet of keratinocytes on a 3T3 murine fibroblast support. However, the model was devoid of a horny layer and not very representative of the skin.1 The first epidermal model with a functional
stratum corneum was published by Michel Prunieras and his team in 1979 on the basis of an emersion keratinocyte culture on an acellular dermal matrix.2 During the 1990s, models that were easier
to manipulate and standardised were developed, notably through the work of Martin Rosdy and LC Clauss, which were cultured at the air-liquid interface on a porous polycarbonate membrane, in a hyper-calcic medium and in the presence of Epidermal Growth Factor (EGF) and vitamin C, among other factors, to promote the formation of a functional epidermal barrier.3
The different types of 3D skin models To date, there are essentially three types of 3D skin models that are compatible with industrial- scale testing, namely reconstituted human epidermis (RHE), human skin equivalent (HSE) and human skin explant, which is taken from human skin during surgery.4
RHEs are most often produced using highly
standardised protocols, in culture inserts of varying sizes on polycarbonate membranes, based on Rosdy’s work mentioned above. They have the advantages of a reasonable unit cost and are commercially available from multiple suppliers (EpiSkin/SkinEthic, EpiDerm/MatTek, epiCS/CellSystems, EPI-001/StratiCELL, EPI- model/LabCyte, etc.). The reconstruction protocol is mainly based on primary keratinocytes derived from neonatal foreskin or abdominal or mammary plasties, but it can easily be applied to keratinocytes derived from iPSC cells, cells from older donors or patients with various pathologies, immortalised keratinocytes (e.g. N-TERT) or genetically modified cells (knock-down, recombinants). Skin equivalents combining a dermal and
an epidermal compartment, are also commercially available (Phenion FT, LabSkin, T-Skin/EpiSkin, EpiDerm FT-200/MatTek), and differ in the nature of the dermal matrix, which may be produced on the basis of collagen, fibrin, other hydrogel-forming proteins, and possibly contain other components such as chitosan and/or hyaluronic acid. The primary advantage of this type of model is that it allows paracrine communication between the cells of the dermis and the epidermis. However, these models are more expensive and time- consuming to produce than reconstructed epidermis models. Finally, ex vivo skin explants from
Th17-driven inflammation in psoriasis
Th17 cytokines Th2 cytokines
Th22-driven hyperplasia in psoriasis
Th22 cytokines
IN VITROMODEL OF SKIN PATHOLOGIES USING CYTOKINE COCKTAILS
Interferon cocktail
Th2-driven atopic dermatitis
Th1 cytokines
Promelanogenic factors
Melanocytorrhagy in vitiligo
Long-chain ceramide deficiency, ichthyosis
Hyperpigmentation in age spots
Figure 1: StratiCELL’s 3D in vitromodels of skin pathologies. November 2020
abdominal or breast surgeries may be acquired from specialised providers or hospital departments, with delivery delays depending on surgical programmes, required specificities, quality criteria and amount of required sample. Even though they are good representative of in vivo skin physiology, they display a high variability of tissue response to treatment or stress, depending on the identity of the donor or the collection site. They are also rapidly found in a pro-inflammatory state linked to survival conditions, and present certain limitations due to preoperative treatment. For example, residual surgical antiseptic compounds exclude ex vivo explants from studies on the skin microbiota. It is also worth emphasising the spectacular advances in cutaneous tissue bio-imprinting
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