Department of Pharmacology and Physiology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand.
Center of Excellence in Cancer Cell and Molecular Biology, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand.
In Vivo. 2023 Sep-Oct;37(5):2006-2017. doi: 10.21873/invivo.13298.
BACKGROUND/AIM: 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.
背景/目的：Box A 是高迁移率族蛋白 1（HMGB1）的一个高度保守的 DNA 结合域，已在许多细胞模型中显示出逆转衰老和老化特征的作用。我们研究了 Box A 的激活是否会影响干细胞特性。
人真皮乳头（DP）细胞和原代人白色前脂肪细胞（HWPc）被用作间充质细胞模型。使用 Box A 过表达质粒诱导细胞 Box A 表达。通过实时聚合酶链反应和免疫荧光分析分别测量 DP 细胞和 HWPc 中干性标志物 POU 类 5 同源框 1 假基因 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。
Box A 过表达质粒在 DP 细胞和 HWPc 中的转染效率分别为 80%和 50%。两种细胞类型的增殖率在转染后 72 小时显著增加。与空质粒转染细胞相比，Box A 转染的 DP 细胞和 HWPc 中 OCT4、NANOG 和 SOX2 的 mRNA 和蛋白表达水平均显著升高。免疫荧光分析证实了 Box A 在 DP 细胞和 HWPc 中诱导 OCT4、NANOG 和 SOX2 蛋白表达。OCT4 和 SOX2 表达于核和细胞质区室，而 NANOG 则强烈定位于 Box A 转染细胞的核内。
我们的研究结果表明，Box A 可能增强干细胞特性，这可能由于衰老过程和疾病导致的干细胞功能下降而具有显著的益处。这项研究可能对再生医学应用具有重要意义。
