Chen Xi, Luo XiaoLiang, Qin Jian, Xie HongHui, Xie Hui
Department of Endodontics, Changsha Stomatological Hospital, No. 389 Youyi road, Tianxin District, Changsha City, Hunan Province 410006, China.
Department of Periodontology，Changsha Stomatological Hospital, No. 389 Youyi road,Tianxin District, Changsha City, Hunan Province, 410006, China.
Tissue Cell. 2025 Aug;95:102890. doi: 10.1016/j.tice.2025.102890. Epub 2025 Mar 27.
Stem cell-based therapies, particularly stem cells from human exfoliated deciduous teeth (SHED), are gaining attention for regenerative medicine. Autophagy plays a crucial role in stem cell function by maintaining cellular homeostasis. Understanding how agents like chrysin induce autophagy in SHED cells could improve stem cell maintenance, enhance functionality, and delay aging, thus offering new therapeutic avenues for stem cell-based regenerative treatments.
Flow cytometry as well as multi-lineage differentiation was performed to characterization of SHED cells. The MTT assay was conducted to determine the optimal concentration of chrysin. The expression of autophagy-related proteins, including LC3-II, p62, and Beclin-1, was analyzed using western blotting. To evaluate cell survival and regenerative capacity, the expression of TERT protein was examined. We further assessed the formation of autophagic vesicles using flow cytometry.
SHED cells were positive for CD44 and CD73, while negative for CD31 and CD34. Differentiation assays demonstrate the ability of SHED cells to differentiate into osteoblasts and adipocytes. The MTT indicate an optimal concentration of 20 μM chrysin, potentially showing enhanced cell proliferation. Western blot analysis revealed increased expression of LC3-II and Beclin-1, as autophagy-related protein following chrysin treatment, while a significant decrease was observed in the level of p62 (****P < 0.0001). TERT levels showed a slight decrease in treated group. Chrysin significantly increased the autophagic vesicles, as evidenced by thr percentage of MDC-positive cells (**P < 0.001). Chrysin also induced slight apoptosis in SHED cells(*P < 0.05).
Together, these findings demonstrate that chrysin induces autophagy in SHED cells, suggesting its potential as a modulator of stem cell function.
基于干细胞的疗法，尤其是来自人脱落乳牙的干细胞（SHED），在再生医学中越来越受到关注。自噬通过维持细胞内稳态在干细胞功能中发挥关键作用。了解白杨素等物质如何诱导SHED细胞发生自噬，可能会改善干细胞的维持、增强其功能并延缓衰老，从而为基于干细胞的再生治疗提供新的治疗途径。
采用流式细胞术以及多谱系分化来表征SHED细胞。进行MTT试验以确定白杨素的最佳浓度。使用蛋白质印迹法分析自噬相关蛋白（包括LC3-II、p62和Beclin-1）的表达。为了评估细胞存活和再生能力，检测了TERT蛋白的表达。我们还使用流式细胞术评估了自噬小泡的形成。
SHED细胞CD44和CD73呈阳性，而CD31和CD34呈阴性。分化试验证明SHED细胞具有分化为成骨细胞和脂肪细胞的能力。MTT试验表明白杨素的最佳浓度为20μM，可能显示出增强的细胞增殖。蛋白质印迹分析显示，白杨素处理后，自噬相关蛋白LC3-II和Beclin-1的表达增加，而p62水平显著降低（****P<0.0001）。治疗组TERT水平略有下降。白杨素显著增加了自噬小泡，MDC阳性细胞百分比证明了这一点（**P<0.001）。白杨素还诱导SHED细胞轻微凋亡（*P<0.05）。
总之，这些发现表明白杨素可诱导SHED细胞发生自噬，提示其作为干细胞功能调节剂的潜力。
