Constructing Human Skin Equivalents on Porcine Acellular Peritoneum Extracellular Matrix forIn VitroIrritation Testing

Abstract

The irritancy of topical products has to be investigated to ensure the safety and compliance. Although several reconstructed human epidermal models have been adopted by the Organization for Economic Cooperation and Development (OECD) to replace in vivo animal irritation testing, these models are based on a single cell type and lack dermal components, which may be insufficient to reflect all of the components of irritation. In our study, we investigated the use of acellular porcine peritoneum extracellular matrix as a substrate to construct full-thickness human skin equivalents (HSEs) for use as irritation screening tool. The acellular peritoneum matrix (APM) exhibited excellent skin cell attachment (>80%) and proliferation for human dermal fibroblasts (HDF) and immortalized human keratinocytes (HaCaT). APM-HSEs based on coculture of HDF and HaCaT were prepared. Increased HDF seeding density up to 5 × 10(4)/cm(2) resulted in APM-HSEs with a thicker and more organized epidermis. The epidermis of APM-HSEs expressed keratin 15, a keratinocyte proliferation marker, and involucrin, a differentiation marker, respectively. To assess the use of APM-HSEs for irritation testing, six proficiency chemicals, including three nonirritants (phosphate-buffered saline, polyethylene glycol 400, and isopropanol) and three irritants (1-bromohexane, heptanol, and sodium dodecyl sulfate) were applied. The APM-HSEs were able to discriminate nonirritants from irritants based on the viability. Levels of cytokines (interleukin [IL]-1α, IL-1ra, IL-6, IL-8, and granulocyte macrophage colony-stimulating factor [GM-CSF]) in these treatment groups further assisted the irritancy ranking. In conclusion, we have developed partially differentiated full-thickness APM-HSEs based on acellular porcine peritoneum matrix, and these APM-HSEs demonstrated utility as an in vitro irritation screening tool.

Determination of Chemical Irritation Potential Using a Defined Gene Signature Set on Tissue-Engineered Human Skin Equivalents

“…Elevated IL-6 expression has also been detected in two-dimensional monolayers of cultured primary skin KCs, HaCaT cells, and dermal fibroblasts (Jung et al, 2016;Terunuma et al, 2001). Tsai et al (2016) detected elevated secretion of IL-6 using a full-thickness HSE constructed of HaCaT KCs in response to a number of skin irritants, and Schmalz et al (1998) observed similar findings when HSEs constructed of primary cells were incubated with metal-containing ceramics, whereas Bock et al (2018) found that IL-6 secretion was only increased when MUTZ-3 Langerhans-like cells were incorporated into their HSE on stimulation with irritants. IL-6 was also dramatically increased in a dermal fibroblast onlyepopulated three-dimensional model in response to cadmium chloride and lauryl sulfate (Augustin and Damour, 1995), suggesting that fibroblasts may be an important source of IL-6 in skin.…”

3D Cell Printing of Perfusable Vascularized Human Skin Equivalent Composed of Epidermis, Dermis, and Hypodermis for Better Structural Recapitulation of Native Skin

“…To be more concrete, the degree of maturation of dermis is normally verified by the content of ECM secreted by the fibroblasts in dermis region. The stratified epidermis by differentiated keratinocytes is analyzed through an early differentiation marker of epidermis such as keratin 10 (K10) and late differentiation markers such as involucrin or filaggrin …”

Beyond 2D: 3D bioprinting for skin regeneration

“…Currently, gelatin hydrogels are widely used as bioink for their high biocompatibility . However, the printed construct's disadvantages of severe contraction, fast degradation, and limited lifespan cannot be ignored . Although the chemical cross‐linkers can improve such weakness, the cross‐linkers also have potential toxicities and damage …”

“…5,9,10,44,45 However, the printed construct's disadvantages of severe contraction, fast degradation, and limited lifespan cannot be ignored. 16,46 Although the chemical cross-linkers can improve such weakness, the cross-linkers also have potential toxicities and damage. [47][48][49][50] Despite these obstacles, the developmental prospect of 3D bioprinting technology for skin regeneration is very optimistic.…”