Exploring hair aging through ex vivo induction of DNA damage and senescence

Summary: Abstract Body: In hair follicle (HF) aging hair density and thinning occur, caused by prolonged telogen and/or catagen phases and HF miniaturization. While models for extrinsic aging exist, there is a lack of models for intrinsic cell aging mechanisms such as DNA damage, repair defects, and cellular senescence. Here, we investigated whether the DNA-intercalating nucleotide BrdU, known to induce intrinsic skin aging ex vivo, could similarly affect aging and senescence in human HFs ex vivo. Microdissected HFs from 3 healthy donors were treated with different BrdU concentrations (10µM-1mM) and analyzed by quantitative (immuno-) histomorphometry. None of the tested concentrations increased LDH release or melanin clumping. Instead, BrdU increased the number of cells expressing the DNA damage marker γH2AX or the cell cycle inhibitor p21. Furthermore, BrdU tendentially enhanced expression of the senescence-associated secretory phenotype indicator CXCL10. Most of these effects occurred in a dose-dependent manner in the HF epithelium. Next we assessed how these changes would translate into HF function. BrdU inhibited hair shaft production and reduced hair shaft quality, demonstrated by a significantly and tendentially decreased pre-cortical hair matrix (HM) expression of Keratin 85 (K85) and K86, respectively, and significantly lower levels of keratin associated protein 3.3 (KRTAP3.3) in the inner root sheath. Furthermore, BrdU induced premature catagen development accompanied by a significant reduction in the hair cycle score and HM keratinocyte proliferation (Ki-67+ cells). Our findings show that BrdU accelerates intrinsic aging and senescence in human HFs ex vivo without causing cytotoxicity. Additionally, our assay serves as a tool to study chronological aging mechanisms and test potential senolytic compounds, potentially reducing HF aging also in vivo. Janin Edelkamp¹, Cecilia Colombero¹, Joana Viola-Söhnlein¹, Sabine Gruedl², Onur Egriboz^{1, 3}, Thomas Welss², Francisco Jimenez⁴, Karin I. Pappelbaum¹, Marta Bertolini¹ 1. QIMA Life Sciences, QIMA Monasterium, Münster, Germany. 2. Henkel AG & Co KGaA, Düsseldorf, NRW, Germany. 3. DWI Labs, Deriworks A.S., Istanbul, Turkey. 4. Mediteknia Clinic, Las Palmas de Gran Canaria, Spain. Translational Studies: Cell and Molecular Biology