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Functional differences between neonatal and adult fibroblasts and keratinocytes: Donor age affects epithelial-mesenchymal crosstalk in vitro.

ABSTRACT: Clinical evidence suggests that healing is faster and almost scarless at an early neonatal age in comparison with that in adults. In this study, the phenotypes of neonatal and adult dermal fibroblasts and keratinocytes (nestin, smooth muscle actin, keratin types 8, 14 and 19, and fibronectin) were compared. Furthermore, functional assays (proliferation, migration, scratch wound closure) including mutual epithelial?mesenchymal interactions were also performed to complete the series of experiments. Positivity for nestin and ? smooth muscle actin was higher in neonatal fibroblasts (NFs) when compared with their adult counterparts (adult fibroblasts; AFs). Although the proliferation of NFs and AFs was similar, they significantly differed in their migration potential. The keratinocyte experiments revealed small, poorly differentiated cells (positive for keratins 8, 14 and 19) in primary cultures isolated from neonatal tissues. Moreover, the neonatal keratinocytes exhibited significantly faster rates of healing the experimentally induced in vitro defects in comparison with adult cells. Notably, the epithelial/mesenchymal interaction studies showed that NFs in co-culture with adult keratinocytes significantly stimulated the adult epithelial cells to acquire the phenotype of small, non-confluent cells expressing markers of poor differentiation. These results indicate the important differences between neonatal and adult cells that may be associated with improved wound healing during the early neonatal period.

SUBMITTER: Mateu R

PROVIDER: S-EPMC5029973 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Functional differences between neonatal and adult fibroblasts and keratinocytes: Donor age affects epithelial-mesenchymal crosstalk in vitro.

Mateu Rosana R Živicová Veronika V Krejčí Eliška Drobná ED Grim Miloš M Strnad Hynek H Vlček Čestmír Č Kolář Michal M Lacina Lukáš L Gál Peter P Borský Jiří J Smetana Karel K Dvořánková Barbora B

International journal of molecular medicine 20160811 4

Clinical evidence suggests that healing is faster and almost scarless at an early neonatal age in comparison with that in adults. In this study, the phenotypes of neonatal and adult dermal fibroblasts and keratinocytes (nestin, smooth muscle actin, keratin types 8, 14 and 19, and fibronectin) were compared. Furthermore, functional assays (proliferation, migration, scratch wound closure) including mutual epithelial‑mesenchymal interactions were also performed to complete the series of experiments ...[more]

PMID: 27513730

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Project description:Background:Keloids are benign fibroproliferative skin lesions that are difficult to treat and become a lifetime predicament for patients. Several treatment modalities have been put forth, but as yet no satisfactory approach to the prevention or treatment of keloids has been identified. The process of epithelial-to-mesenchymal transition (EMT) has been implicated in keloid scarring, as keloid keratinocytes display an EMT-like phenotype. This study investigated the potential of pirfenidone, an antifibrotic agent, to counteract EMT-like alterations in keloid keratinocytes, including gene expression, cell migratory and proliferative functions. Methods:Normal and keloid keratinocytes were isolated from discarded normal skin tissues and from resected keloid tissues, respectively. Cells were quiesced for 24 h without epidermal growth factor DS-Qi1MCDigital and were exposed to transforming growth factor-beta1 (TGF-?1; 10 ng/mL), with or without pirfenidone (400 ?g/mL), for an additional 24 h. The effects of pirfenidone on cytotoxicity, cell migration, cell proliferation, and on expression of genes and proteins involved in EMT were assayed. Statistical significance was determined by two-way ANOVA using Sigma Plot. Results:We found that pirfenidone did not elicit any cytotoxic effect at concentrations up to 1000 ?g/mL. A statistically significant dose-dependent decrease in basal cell proliferation rate was noted in both normal and keloid keratinocytes when exposed to pirfenidone at concentrations ranging from 200 to 1000 ?g/mL. Pirfenidone significantly decreased basal cell migration in both normal and keloid keratinocytes, but a significant decrease in TGF-?1-induced cell migration was seen only in keloid keratinocytes. Significant inhibition of the expression of TGF-?1-induced core EMT genes, namely hyaluronan synthase 2, vimentin, cadherin-11, and wingless-type MMTV integration site family, member 5A along with fibronectin-1, was observed in both normal and keloid keratinocytes treated with pirfenidone. In addition, the protein levels of vimentin and fibronectin were significantly reduced by pirfenidone (400 ?g/mL) in both normal and keloid keratinocytes. Conclusions:For the first time, this study shows the efficacy of pirfenidone in inhibiting the EMT-like phenotype in keratinocytes derived from keloids, suggesting that pirfenidone may counteract a critical contributor of keloid progression and recurrence.

Dataset Information

Functional differences between neonatal and adult fibroblasts and keratinocytes: Donor age affects epithelial-mesenchymal crosstalk in vitro.

Publications

Functional differences between neonatal and adult fibroblasts and keratinocytes: Donor age affects epithelial-mesenchymal crosstalk in vitro.

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