盒A是高迁移率群盒1（HMGB1）的高度保守DNA结合结构域，在许多细胞模型中已被证明能够逆转细胞衰老和老化特征。我们研究了盒A的激活是否会影响干细胞特性。作为间充质细胞模型，我们选择了人皮肤乳头（DP）细胞和原代人白色前脂肪细胞（HWPc）。我们使用盒A过表达质粒来诱导细胞中盒A的表达。分别通过实时聚合酶链反应和免疫荧光分析测量了DP细胞和HWPc中干细胞标记物POU类5同源框1伪基因5（OCT4, HGNC: 9221）、Nanog同源框（NANOG, HGNC: 20857）和SRY-盒转录因子2（SOX2, HGNC: 11195）的mRNA和蛋白水平。盒A过表达质粒的转染效率分别为DP细胞和HWPc的80%和50%。转染后72小时，两种细胞类型的增殖率显著增加。与转染空质粒的细胞相比，盒A转染的DP细胞和HWPc中OCT4、NANOG和SOX2的mRNA和蛋白表达水平显著增加。免疫荧光分析证实，盒A能够诱导DP细胞和HWPc中OCT4、NANOG和SOX2的蛋白表达。OCT4和SOX2在核和细胞质中均有表达，而NANOG则主要定位于盒A转染细胞的细胞核中。我们的发现表明，盒A可能有助于增强干细胞性质，这可能对改善干细胞功能并应对衰老过程和疾病具有重要意义。这项研究对再生医学应用可能具有重要影响。版权所有©2023，国际抗癌研究学会（George J. Delinasios博士），保留所有权利。

Box A is a highly conserved DNA-binding domain of high-mobility group box 1 (HMGB1) and has been shown to reverse senescence and aging features in many cell models. We investigated whether the activation of box A can influence stem cell properties.Human dermal papilla (DP) cells and primary human white pre-adipocytes (HWPc) were employed as mesenchymal cell models. Box A-overexpressing plasmids were used to induce cellular box A expression. mRNA and protein levels of stemness markers POU class 5 homeobox 1 pseudogene 5 (OCT4, HGNC: 9221), Nanog homeobox (NANOG, HGNC: 20857), and SRY-box transcription factor 2 (SOX2, HGNC:11195) in DP cells and HWPc were measured by real-time polymerase chain reaction and immunofluorescence analysis, respectively.Transfection efficiency of box A-overexpressing plasmid was 80% and 50% in DP cells and HWPc, respectively. The proliferative rate of both cell types significantly increased 72 h after transfection. Levels of OCT4, NANOG and SOX2 mRNA and protein expression were significantly increased in box A-transfected DP cells and HWPc compared to empty plasmid-transfected cells. Immunofluorescence analysis confirmed the induction of OCT4, NANOG and SOX2 protein expression in response to box A in DP cells and HWPc. OCT4 and SOX2 were expressed in both the nuclear and cytoplasmic compartments, while NANOG was intensely located in the nucleus of box A-transfected cells.Our findings suggest that box A may potentially enhance stemness, which may have significant benefits in improving stem cell function due to aging processes and disease. This research may have implications for regenerative medicine applications.Copyright © 2023, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.