Characterisation of cell cycle arrest and terminal differentiation in a maximally proliferative human epithelial tissue: Lessons from the human hair follicle matrix

Abstract

Human hair follicle (HF) growth and hair shaft formation require terminal differentiation-associated cell cycle arrest of highly proliferative matrix keratinocytes. However, the regulation of this complex event remains unknown. CIP/KIP family member proteins (p21, p27 and p57) regulate cell cycle progression/arrest, endoreplication, differentiation and apoptosis. Since they have not yet been adequately characterized in the human HF, we asked whether and where CIP/KIP proteins localise in the human hair matrix and pre-cortex in relation to cell cycle activity and HF-specific epithelial cell differentiation that is marked by keratin 85 (K85) protein expression. K85 expression coincided with loss or reduction in cell cycle activity markers, including in situ DNA synthesis (EdU incorporation), Ki-67, phospho-histone H3 and cyclins A and B1, affirming a post-mitotic state of pre-cortical HF keratinocytes. Expression of CIP/KIP proteins was found abundantly within the proliferative hair matrix, concomitant with a role in cell cycle checkpoint control. p21, p27 and cyclin E persisted within post-mitotic keratinocytes of the pre-cortex, whereas p57 protein decreased but became nuclear. These data imply a supportive role for CIP/KIP proteins in maintaining proliferative arrest, differentiation and anti-apoptotic pathways, promoting continuous hair bulb growth and hair shaft formation in anagen VI. Moreover, post-mitotic hair matrix regions contained cells with enlarged nuclei, and DNA in situ hybridisation showed cells that were >2N in the pre-cortex. This suggests that CIP/KIP proteins might counterbalance cyclin E to control further rounds of DNA replication in a cell population that has a propensity to become tetraploid. These data shed new light on the in situ-biography of human hair matrix keratinocytes on their path of active cell cycling, arrest and terminal differentiation, and showcase the human HF as an excellent, clinically relevant model system for cell cycle physiology research of human epithelial cells within their natural tissue habitat.

CDK4/6 inhibition mitigates stem cell damage in a novel model for taxane‐induced alopecia

“…Paclitaxel promoted the extensive accumulation of irregular and shrunken nuclei that localised specifically to the most proliferative region of the hair matrix (Fig A; i.e. predominantly below the critical line of Auber; Purba et al , , 2017a). Docetaxel treatment (24 h) also promoted the formation of irregular nuclear bodies within the hair matrix, albeit not to the extent seen following paclitaxel treatment (Fig B–E).…”

“…Goat anti‐Mouse/Rabbit Secondary Antibodies, Alexa Fluor 488/594 (Invitrogen, #A11001, #A11005 #A11008 #A11037) (1:200). See also supporting references (Purba et al , , ,b).…”

“…by promoting chromosome missegregation/cell division on multipolar spindles (Chen & Horwitz, ; Abal et al , ; Morse et al , ; Weaver, ; Zasadil et al , ). Taxanes are thus presumed to cause hair loss by damaging rapidly dividing matrix keratinocytes and their counterpart stem/progenitor cells, required for healthy hair growth and hair follicle cycling (Paus & Cotsarelis, ; Garza et al , ; Paus et al , ; Purba et al , , ,b; Gao et al , ; Huang et al , ). However, the effects of taxane chemotherapy on the human hair follicle remain to be systematically examined.…”

A combination of pyridine‐2, 4‐dicarboxylic acid diethyl ester and resveratrol stabilizes hypoxia‐inducible factor 1‐alpha and improves hair density in female volunteers

“…The selection of resveratrol to be associated with STX was based on its properties to reduce ROS and inflammation, described in alopecia . We showed that a pre‐treatment with resveratrol protected the hair matrix cells when exposed to a generator of ROS, which is of importance as the hair matrix cells are the key player to produce the hair shaft and must be preserved to guarantee the hair density. In this study, the results highlighted the property of resveratrol to protect the whole hair follicle in vitro , blocking the generation of ROS in the hair cells induced by H 2 O 2 .…”

Visualization of Global RNA Synthesis in a Human (mini-) Organ in Situ by Click Chemistry

“…Comparative analysis of EU incorporation within the matrix, pre-cortex, IRS and DP of the HF bulb [9,10] showed relative compartmental homogeneity in levels of RNA synthesis, as no significant differences in EU signal were seen between these cell populations (Figure 1CD). Supporting this, visualization of proliferation (Ki-67) [9,10] against EU incorporation revealed no distinguishable pattern of RNA synthesis that associated with either cell cycle activity or arrest in this compartment (Figure 1E).…”