Induction of senescence pathways in Kindler syndrome primary keratinocytes. (2nd April 2013)

• Record Type: Journal Article
• Authors: Piccinni, E.; Di Zenzo, G.; Maurelli, R.; Dellambra, E.; Teson, M.; Has, C.; Zambruno, G.; Castiglia, D.
• Is Part Of: British journal of dermatology. Volume 168:Number 5(2013:May)
• Page Start: 1019
• Page End: 1026
• Publication Date: 2013-04-02
• Subjects: Dermatology -- Periodicals; Skin -- Diseases -- Periodicals; 616.5
• DOI: 10.1111/bjd.12184
• Languages: English
• ISSNs: 0007-0963

Abstract

Summary: Background Individuals with Kindler syndrome (KS) have loss‐of‐function mutations in the FERMT1 gene that encodes the focal adhesion component kindlin‐1. The major clinical manifestation of KS is epidermal atrophy (premature skin ageing). This phenotypic feature is thought to be related to the decreased proliferation rate of KS keratinocytes; nevertheless, molecular mediators of such abnormal behaviour have not been fully elucidated. Objectives To investigate how kindlin‐1 deficiency affects the proliferative potential of primary human keratinocytes. Methods We serially cultivated nine primary KS keratinocyte strains until senescence and determined their lifespan and colony‐forming efficiency (CFE) at each serial passage. The expression of molecular markers of stemness and cellular senescence were investigated by immunoblotting using cell extracts of primary keratinocyte cultures from patients with KS and healthy donors. In another set of experiments, kindlin‐1 downregulation in normal keratinocytes was obtained by small interfering RNA (siRNA) technology. Results We found that KS keratinocytes exhibited a precocious senescence and strongly reduced clonogenic potential. Moreover, KS cultures showed a strikingly increased percentage of aborted colonies (paraclones) already at early passages indicating an early depletion of stem cells. Immunoblotting analysis of KS keratinocyte extracts showed reduced levels of the stemness markers p63 and Bmi‐1, upregulation of p16 and scant amounts of hypophosphorylated Rb protein, which indicated cell cycle‐arrested status. Treatment of normal human primary keratinocytes with siRNA targeting kindlin‐1 proved that its deficiency was directly responsible for p63, Bmi‐1 and pRb downregulation and p16 induction. Conclusions Our data directly implicate kindlin‐1 in preventing premature senescence of keratinocytes. Abstract : What's already known about this topic? Kindler syndrome (KS) is a genodermatosis caused by mutations in the FERMT1/KIND1 gene encoding kindlin‐1. A clinically diagnostic feature of KS is premature skin ageing. Kindlin‐1 deficiency associates with functional abnormalities of keratinocytes. Epidermal atrophy is thought to reflect a reduced proliferation rate of KS keratinocytes both in vivo and in vitro . What does this study add? KS primary keratinocyte cultures are depleted in stem cells, due to premature senescence. The reduced clonogenicity of KS keratinocytes correlates with both upregulation of proteins associated with cell senescence and downregulation of markers identifying stem cells. Our findings underline the crucial contribution of kindlin‐1 deficiency in the premature ageing of primary human keratinocytes.