The ROC skin model: A robust skin equivalent for permeation and live cell imaging studies

Jakobsen, Nicoline Dorothea; Kaiser, Katharina; Ebbesen, Morten Frendø; Lauritsen, Line; Gjerstorff, Morten F.; Kuntsche, Judith; Brewer, Jonathan R.

doi:10.1016/j.ejps.2022.106282

Cited by 4 publications

(2 citation statements)

References 32 publications

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“…There are several limitations to the current study. Complete differentiation of keratinocytes into corneocytes in native human skin take on average 14 days, during which the dead cell layer of your skin turns over [ 17 , 20 , 54 , 55 ]. Therefore, most studies employ 14–21-day cultures of keratinocyte on the 3D surface at the ALI.…”

Section: Discussionmentioning

confidence: 99%

Keratin Promotes Differentiation of Keratinocytes Seeded on Collagen/Keratin Hydrogels

et al. 2022

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Keratinocytes undergo a complex process of differentiation to form the stratified stratum corneum layer of the skin. In most biomimetic skin models, a 3D hydrogel fabricated out of collagen type I is used to mimic human skin. However, native skin also contains keratin, which makes up 90% of the epidermis and is produced by the keratinocytes present. We hypothesized that the addition of keratin (KTN) in our collagen hydrogel may aid in the process of keratinocyte differentiation compared to a pure collagen hydrogel. Keratinocytes were seeded on top of a 100% collagen or 50/50 C/KTN hydrogel cultured in either calcium-free (Ca-free) or calcium+ (Ca+) media. Our study demonstrates that the addition of keratin and calcium in the media increased lysosomal activity by measuring the glucocerebrosidase (GBA) activity and lysosomal distribution length, an indication of greater keratinocyte differentiation. We also found that the presence of KTN in the hydrogel also increased the expression of involucrin, a differentiation marker, compared to a pure collagen hydrogel. We demonstrate that a combination (i.e., containing both collagen and kerateine or “C/KTN”) hydrogel was able to increase keratinocyte differentiation compared to a pure collagen hydrogel, and the addition of calcium further increased the differentiation of keratinocytes. This multi-protein hydrogel shows promise in future models or treatments to increase keratinocyte differentiation into the stratum corneum.

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Section: Discussionmentioning

confidence: 99%

Keratin Promotes Differentiation of Keratinocytes Seeded on Collagen/Keratin Hydrogels

et al. 2022

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“…Phenotypically similar to basal keratinocytes, REKs endogenously express an abundant level of the gap junction forming protein Cx43 ( Jakobsen et al, 2022 ; Maher et al, 2005 ). To investigate the subcellular localization and functional capacity of Cx31.1 in keratinocytes, we expressed FLAG- and GFP-tagged Cx31.1 in GJIC-competent Cx43-enriched and Cx43-ablated REKs (Cx43 KO).…”

Section: Resultsmentioning

confidence: 99%

Cx31.1 can selectively intermix with co-expressed connexins to facilitate its assembly into gap junctions

Leighton,

Wong,

Lucaciu

et al. 2024

Journal of Cell Science

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Connexins are channel forming proteins that function to facilitate gap junctional intercellular communication. Here we use dual cell voltage clamp and dye transfer studies to corroborate past findings that Cx31.1 is defective in gap junction channel formation; illustrating that Cx31.1 alone does not form functional gap junction channels in connexin-deficient cells. Rather Cx31.1 transiently localizes to the secretory pathway with a subpopulation reaching the cell surface which is rarely seen in puncta reminiscent of gap junctions. Intracellular retained Cx31.1 was subject to degradation as Cx31.1 accumulated in the presence of proteasomal inhibition, had a faster turnover when Cx43 was present, and ultimately reached lysosomes. While intracellularly retained Cx31.1 was found to interact with Cx43, this interaction did not rescue its delivery to the cell surface. Conversely, the co-expression of Cx31 dramatically rescued the assembly of Cx31.1 into gap junctions where gap junction-mediated dye transfer was enhanced. Collectively, our results indicate that the localization and functional status of Cx31.1 is altered through selective interplay with co-expressed connexins, perhaps suggesting Cx31.1 is a key regulator of intercellular signalling in keratinocytes.

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From static to dynamic: The influence of mechanotransduction on skin equivalents analyzed by bioimaging and RNAseq

Kaiser,

Bendixen,

Sørensen

et al. 2024

Materials Today Bio

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The ROC skin model: A robust skin equivalent for permeation and live cell imaging studies

Cited by 4 publications

References 32 publications

Keratin Promotes Differentiation of Keratinocytes Seeded on Collagen/Keratin Hydrogels

Keratin Promotes Differentiation of Keratinocytes Seeded on Collagen/Keratin Hydrogels

Cx31.1 can selectively intermix with co-expressed connexins to facilitate its assembly into gap junctions

From static to dynamic: The influence of mechanotransduction on skin equivalents analyzed by bioimaging and RNAseq

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