Exon skipping of TGFβRI affects signalling and ECM expression in hypertrophic scar-derived fibroblasts.

Autor:	Raktoe RS; Department of Dermatology, Leiden University Medical Center (LUMC), The Netherlands., Rietveld MH; Department of Dermatology, Leiden University Medical Center (LUMC), The Netherlands., Out-Luiting JJ; Department of Dermatology, Leiden University Medical Center (LUMC), The Netherlands., Kruithof-de Julio M; Department of Urology, LUMC, The Netherlands.; Department of Urology, University of Bern, Switzerland., van Zuijlen PP; Department of Plastic, Reconstructive and Hand Surgery, VU University Medical Center, The Netherlands., van Doorn R; Department of Dermatology, Leiden University Medical Center (LUMC), The Netherlands., Ghalbzouri AE; Department of Dermatology, Leiden University Medical Center (LUMC), The Netherlands.
Jazyk:	angličtina
Zdroj:	Scars, burns & healing [Scars Burn Heal] 2020 May 28; Vol. 6, pp. 2059513120908857. Date of Electronic Publication: 2020 May 28 (Print Publication: 2020).
DOI:	10.1177/2059513120908857
Abstrakt:	Background: In burn patients, wound healing is often accompanied by hypertrophic scar (HS) development, resulting in both functional and aesthetic problems. HSs are characterised by abundant presence of myofibroblasts that contribute to overproduction of extracellular matrix (ECM) that is regulated by the TGF-β signalling pathway. Studies have shown that inhibition of TGF-β receptors in fibrotic diseases reduces the fibrotic load. In the present study, we aim to inactivate ALK5, also known as TGF-β receptor I, in human HS fibroblasts by exon skipping using antisense oligonucleotides (AONs). Methods: HS biopsies were used to isolate and set up fibroblast monocultures. AONs targeting ALK5 were supplemented to the fibroblast cultures to induce exon skipping, while pharmacological ALK5 inhibition was induced using SB431542. AON delivery in HS fibroblasts was examined using immunofluorescence (IF), while TGF-β signalling downstream targets, such as Smad2/3, PAI-1, ACTA2, COL1A1 and COL3A1, were analysed using touchdown polymerase chain reaction (PCR), quantitative PCR (qPCR), IF or western blotting. Results: Our data clearly demonstrate that AONs were successfully delivered in the nuclei of HS fibroblasts and that functional exon skipping of ALK5 took place as confirmed with touchdown PCR and qPCR. In addition, exon skipping affected the expression of ECM-related genes, such as type I/III collagens, PAI-1 and CCN2. Moreover, AON treatment did not affect the migration of HS fibroblasts in a model for wound healing. Conclusion: Exon skipping is a promising tool to modulate the TGF-β signalling pathway in HS. This would open a therapeutic window for the treatment of patients suffering from HSs. Competing Interests: Declaration of conflicting interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. (© The Author(s) 2020.)
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu