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. 2024 Oct 13;19:91. doi: 10.1186/s13062-024-00540-8

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© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/4.0/.

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Fig. 1 — CDK1 is upregulated in stem-cell enriched epidermal cultures. (a) Scheme describing the experimental pipeline for mass spectrometry analysis; abbreviations: EPSC = Epidermal Stem Cell, TAC = Transient Amplifying Cell, TD = Terminally Differentiated cell, LC-MS = Liquid Chromatography Mass Spectrometry; MS = Mass Spectrometry; (b) Volcano plot representation of the proteins identified by the MS proteomics analysis of enriched vs. depleted samples. Significantly downregulated proteins are represented in blue, significantly upregulated proteins are represented in red. On X-axis, Fold-change is reported (log₂(ratio)), while on Y-axis proteins’ significance is depicted (as -Log₁₀ of the corresponding q-value). (c) STRING local Network analysis (LNA) obtained from the MCL cluster. Number of nodes: 8, number of edges: 12, average node degree: 3, avg. local clustering coefficient: 0.792, expected number of edges: 0, PPI enrichment p-value: 2.11 × 10^− 15. (d) qRT-PCR quantification of the mRNA levels of CDK1, H1B, IVL, and SPINK5 on three different NHK primary cultures. Expression levels were normalized per GAPDH. Data are presented as mean +/− SD of N = 3 different independent biological replicates, *P < 0.05 (Student t-test). (e) Western analysis of total cell extracts from cultures generated by sub-confluent NHKs during serial cultivation. Molecular weight indicators are shown. (f) Western analysis of total cell extracts derived from NHK cultures treated with 30 μM CYC (see Methods) or DMSO as control. Molecular weight indicators are shown. (g) Western analysis of total cell extracts derived from NHK cultures treated with 30 μM RO3306 (see Methods) or DMSO as control. Molecular weight indicators are shown. (h) qRT-PCR quantification of the mRNA levels of FOXM1, CDK1, CCNB1, and H1B on cells derived from DMSO-, CYC- or RO3306-treated NHK cultures. Expression levels were normalized per GAPDH and given relative to the control (DMSO) arbitrarily set to 1. Data are presented as mean +/− SD of N = 3 different independent biological replicates, *P < 0.05, **P < 0.01, ***P < 0.001, ns = not significant (Student t-test)