中华皮肤科杂志 ›› 2026, Vol. 59 ›› Issue (3): 244-252.doi: 10.35541/cjd.20240511

• 论著 • 上一篇    下一篇

CCNA2与CDK1过表达对中波紫外线辐射的HaCaT细胞生长周期和细胞凋亡蛋白表达的影响

汪振娟    刘辉煌    孙迎    朱玉婷    傅钰    杨英   

  1. 遵义医科大学第二附属医院皮肤科，遵义  563000 
  • 收稿日期:2024-09-26 修回日期:2025-06-16 发布日期:2026-03-03
  • 通讯作者: 杨英 E-mail:3894446271@qq.com
  • 基金资助:
    遵义市科技与大数据局科学技术基金项目［遵市科合HZ字（2022）406号］；遵义医科大学第二附属医院硕士启动资金项目（SQ-2021-01）

Effects of CCNA2 and CDK1 overexpression on the cell cycle- and apoptosis-related protein expression in HaCaT cells exposed to ultraviolet B radiation

Wang Zhenjuan, Liu Huihuang, Sun Ying, Zhu Yuting, Fu Yu, Yang Ying   

  1. Department of Dermatology, the Second Affiliated Hospital of Zunyi Medical University, Zunyi 563000, China
  • Received:2024-09-26 Revised:2025-06-16 Published:2026-03-03
  • Contact: Yang Ying E-mail:3894446271@qq.com
  • Supported by:
    Zunyi Science and Technology and Big Data Bureau Science and Technology Fund Project （HZ-［2022］No.406）; Master's Start-up Fund Project of the Second Affiliated Hospital of Zunyi Medical University （SQ-2021-01）

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摘要： 【摘要】 目的 探索细胞周期蛋白A2（CCNA2）和细胞周期依赖性激酶1（CDK1）对中波紫外线（UVB）辐射的HaCaT细胞生长周期和细胞凋亡蛋白表达的影响。方法 检索基因表达数据库（GEO）收录的日光性皮炎（SD）患者转录组测序数据集GSE86406，经基因差异表达分析、京都基因和基因组数据库富集分析、蛋白-蛋白相互作用网络分析，筛选出SD致病机制中关键基因CCNA2、CDK1，并进一步在体外实验中探索其对人角质形成细胞功能的影响。将人角质形成细胞HaCaT分为正常对照（NC）组（仅正常培养）和UVB组（累积辐射剂量30 mJ/cm2），蛋白质印迹法检测CCNA2和CDK1表达。分别过表达及联合过表达CCNA2、CDK1后行UVB照射建模（CCNA2 + UVB组、CDK1 + UVB组、CCNA2 + CDK1 + UVB组），以NC组和UVB组作为对照，流式细胞仪检测细胞周期变化，蛋白质印迹法检测细胞增殖相关蛋白抗原Ki-67、增殖细胞核抗原（PCNA）、凋亡相关蛋白B细胞淋巴瘤2（Bcl-2）、Bcl-2相关X蛋白（Bax）、胱天蛋白酶3（caspase-3）、裂解的胱天蛋白酶3（cleaved-caspase-3）表达，免疫共沉淀检测CCNA2和CDK1的相互作用。结果 人SD样本GEO测序数据分析显示，SD组CCNA2与CDK1表达低于健康对照组（log2FC = -1.16、-1.13，t = -6.71、-5.47，均P＜0.001）。蛋白质印迹法检测显示，UVB组细胞周期蛋白CCNA2、CDK1表达低于NC组（t = 5.93、6.39，P = 0.004、0.003）。与HaCaT细胞UVB组（G1：65.66% ± 0.30%，S：27.56% ± 0.43%，G2：6.77% ± 0.28%）相比，CCNA2 + UVB组（G1：69.31% ± 0.61%，S：21.53% ± 0.44%，G2：9.16% ± 0.58%）和CCNA2 + CDK1 + UVB组（G1：69.15% ± 0.30%，S：20.02% ± 0.11%，G2：10.83% ± 0.34%）G1期、G2期细胞百分比升高，S期降低，均P＜0.001；而CDK1 + UVB组（G1：56.16% ± 0.27%，S：36.36% ± 0.28%，G2：7.48% ± 0.34%）G1期细胞百分比下降（P＜0.001），S期升高（P＜0.001），G2期差异无统计学意义（P = 0.102）。蛋白质印迹实验显示，与UVB组（蛋白相对定量，Ki-67：0.92 ± 0.18，PCNA：0.54 ± 0.10，Bax：1.98 ± 0.29，Bcl-2：0.46 ± 0.10，cleaved-caspase-3：2.05 ± 0.20）相比，CCNA2 + UVB组、CDK1 + UVB组Ki-67表达上调（1.75 ± 0.18、1.70 ± 0.13，均P＜0.05），PCNA表达差异无统计学意义（0.99 ± 0.13、1.00 ± 0.01，均P > 0.05） ，Bax表达下调（1.18 ± 0.67、0.79 ± 0.19，均P＜0.05），Bcl-2表达上调（0.88 ± 0.06、0.86 ± 0.07，均P＜0.05），CCNA2 + UVB组cleaved-caspase-3表达差异无统计学意义（1.55 ± 0.04，P > 0.05），CDK1 + UVB组cleaved-caspase-3表达下调（0.93 ± 0.07，P＜0.001）；与CCNA2 + UVB组和/或CDK1 + UVB组相比，CCNA2 + CDK1 + UVB组Ki-67、Bcl-2表达进一步上调（2.15 ± 0.14、0.91 ± 0.02，均P < 0.01），Bax、cleaved-caspase-3表达进一步下调（0.73 ± 0.05、0.71 ± 0.08，均P＜0.01），PCNA表达上调（1.33 ± 0.12，P＜0.01）。免疫共沉淀实验显示，CCNA2与CDK1有相互作用。结论 CCNA2可与CDK1相互作用，协同促进角质形成细胞的周期调节、增殖，并抑制其凋亡发生。

关键词: 皮炎, 日光性皮炎, 细胞周期蛋白A2, 细胞周期依赖性激酶1, 细胞周期, 细胞增殖, 细胞凋亡

Abstract: 【Abstract】 Objective To investigate the effects of cyclin A2 （CCNA2） and cyclin-dependent kinase 1 （CDK1） on cell cycle- and apoptosis-related protein expression in HaCaT cells exposed to ultraviolet B （UVB） radiation. Methods The transcriptome sequencing dataset GSE86406 from patients with solar dermatitis （SD） was obtained from the Gene Expression Omnibus （GEO） database. Through differential gene expression analysis, Kyoto Encyclopedia of Genes and Genomes enrichment analysis, and protein-protein interaction network analysis, CCNA2 and CDK1 were identified to be key genes involved in the pathogenesis of SD. Their effects on the function of human keratinocytes were further explored in vitro. Human keratinocyte HaCaT cells were divided into a normal control （NC） group （cultured under standard conditions） and a UVB group （irradiated at a cumulative dose of 30 mJ/cm2）. The expression of CCNA2 and CDK1 was determined by Western blot analysis. CCNA2 and CDK1 were overexpressed separately or co-overexpressed in HaCaT cells, followed by UVB irradiation （i.e., a CCNA2 + UVB group, a CDK1 + UVB group, and a CCNA2 + CDK1 + UVB group）, and the NC group and UVB group served as controls. Flow cytometry was performed to analyze changes in the cell cycle, while Western blot analysis was employed to determine the expression of proliferation-related proteins （Ki-67 and proliferating cell nuclear antigen ［PCNA］） and apoptosis-related proteins （B-cell lymphoma 2 ［Bcl-2］, Bcl-2-associated X protein ［Bax］, caspase-3, and cleaved caspase-3）. Co-immunoprecipitation assay was performed to investigate the interaction between CCNA2 and CDK1. Results Analysis of GEO sequencing data from human SD samples showed that the expression of CCNA2 and CDK1 was significantly lower in the SD group than in the healthy control group （log2FC = -1.16, -1.13, t = -6.71, -5.47, respectively, both P < 0.001）. Western blot analysis revealed that CCNA2 and CDK1 protein expression was significantly lower in the UVB group than in the NC group （t = 5.93, 6.39, P = 0.004, 0.003, respectively）. Compared with the UVB group （G1 phase: 65.66% ± 0.30%, S phase: 27.56% ± 0.43%, G2 phase: 6.77% ± 0.28%）, the CCNA2 + UVB group （G1 phase: 69.31% ± 0.61%, S phase: 21.53% ± 0.44%, G2 phase: 9.16% ± 0.58%） and the CCNA2 + CDK1 + UVB group （G1 phase: 69.15% ± 0.30%, S phase: 20.02% ± 0.11%, G2 phase: 10.83% ± 0.34%） showed increased percentages of HaCaT cells in G1 and G2 phases and decreased percentages of HaCaT cells in S phase （all P < 0.001）, while the CDK1 + UVB group （G1 phase: 56.16% ± 0.27%, S phase: 36.36% ± 0.28%, G2 phase: 7.48% ± 0.34%） exhibited decreased percentages of HaCaT cells in G1 phase （P < 0.001）, increased percentages of HaCaT cells in S phase （P < 0.001）, and no significant difference in the percentages of HaCaT cells in G2 phase （P = 0.102）. As Western blot analysis further revealed, compared with the UVB group （Ki-67: 0.92 ± 0.18; PCNA: 0.54 ± 0.10; Bax: 1.98 ± 0.29; Bcl-2: 0.46 ± 0.10; cleaved caspase-3: 2.05 ± 0.20）, both the CCNA2 + UVB group and CDK1 + UVB group exhibited increased Ki-67 expression （1.75 ± 0.18, 1.70 ± 0.13, respectively, both P < 0.05）, no significant change in PCNA expression （0.99 ± 0.13, 1.00 ± 0.01, respectively, both P > 0.05）, reduced Bax expression （1.18 ± 0.67, 0.79 ± 0.19, respectively, both P < 0.05）, and increased Bcl-2 expression （0.88 ± 0.06, 0.86 ± 0.07, respectively, both P < 0.05）, the CCNA2 + UVB group showed no significant change in cleaved caspase-3 expression （1.55 ± 0.04, P > 0.05）, while the CDK1 + UVB group revealed reduced cleaved caspase-3 expression （0.93 ± 0.07, P < 0.001）; moreover, compared with the CCNA2 + UVB group and/or CDK1 + UVB group, the CCNA2 + CDK1 + UVB group showed further upregulation of Ki-67 and Bcl-2 expression （2.15 ± 0.14, 0.91 ± 0.02, respectively, both P < 0.01）, further downregulation of Bax and cleaved caspase-3 expression （0.73 ± 0.05, 0.71 ± 0.08, respectively, both P < 0.01）, and upregulation of PCNA expression （1.33 ± 0.12, P < 0.01）. Co-immunoprecipitation assay confirmed the interaction between CCNA2 and CDK1. Conclusion CCNA2 could interact with CDK1 to cooperatively promote the cell cycle progression and proliferation of keratinocytes, but inhibit their apoptosis.

Key words: Dermatitis, Solar dermatitis, CCNA2, CDK1, Cell cycle, Cell proliferation, Apoptosis

引用本文

汪振娟 刘辉煌 孙迎 朱玉婷 傅钰 杨英. CCNA2与CDK1过表达对中波紫外线辐射的HaCaT细胞生长周期和细胞凋亡蛋白表达的影响[J]. 中华皮肤科杂志, 2026,59(3):244-252. doi:10.35541/cjd.20240511

Wang Zhenjuan, Liu Huihuang, Sun Ying, Zhu Yuting, Fu Yu, Yang Ying. Effects of CCNA2 and CDK1 overexpression on the cell cycle- and apoptosis-related protein expression in HaCaT cells exposed to ultraviolet B radiation[J]. Chinese Journal of Dermatology, 2026, 59(3): 244-252.doi:10.35541/cjd.20240511