牡丹籽乙醇提取物对UVB诱导HaCaT细胞光老化的保护作用及机制研究

韩婕珺; 洪铮怡; 龚天贵; 王斌; 张蓝月

doi:10.13386/j.issn1002-0306.2023080212

牡丹籽乙醇提取物对UVB诱导HaCaT细胞光老化的保护作用及机制研究

Protective Effect and Mechanism of Paeonia suffruticosa Seed Ethanol Extract on Photoaging of HaCaT Cells Induced by UVB

摘要

摘要: 目的：研究牡丹籽乙醇提取物对UVB诱导人永生化角质形成细胞（HaCaT）光老化的保护作用及机制。方法：通过超高效液相色谱-四级杆飞行时间质谱（UPLC-Q-TOF-MS）法分析牡丹籽乙醇提取物存在的抗光老化活性成分，采用UVB刺激HaCaT细胞建立光老化细胞模型，噻唑蓝比色（MTT）法测定细胞活力，细胞划痕法检测牡丹籽乙醇提取物对细胞迁移能力的影响，通过酶联免疫吸附试验（ELISA）检测影响衰老的白细胞介素-1（Interleukin-1，IL-1）、白细胞介素-6（Interleukin-6，IL-6）、白细胞介素-22（Interleukin-22，IL-22）、干扰素-γ（Interferon-γ，IFN-γ）和转化生长因子-β（Tansforming growth factor-β，TGF-β）等细胞因子，以活性氧（Reactive oxygen species，ROS）和核因子-κB（Nuclear factor-κB，NF-κB）的水平为指标，研究不同浓度的牡丹籽乙醇提取物对HaCaT细胞的抗光老化活性及其机制。结果：含量最多的前五种成分为单硬脂酸甘油酯（10.73%）、芥酸酰胺（3.17%）、5-（2R,3S）-6-羟基-2-（4-羟基苯基）-4-（E）-2-（4-羟基苯基）乙烯基-2,3-二氢1-苯并呋喃-3-基苯-1,3-二醇（2.20%）、α,α-海藻糖（1.90%）和芍药内酯苷（1.45%）。经不同浓度牡丹籽乙醇提取物处理后的HaCaT细胞活力均在80%以上，其中浓度为6.25 μg/mL的牡丹籽乙醇提取物对HaCaT细胞无显著毒性，并且浓度为12.5 μg/mL（H组）和6.25 μg/mL（L组）的牡丹籽乙醇提取物均能抑制HaCaT细胞的迁移能力，H组能显著降低细胞IL-1、IL-6、IL-22、IFN-γ和TGF-β水平（P<0.05）从而抵抗皮肤衰老，L组能减少ROS产生，降低NF-κB蛋白因子的含量（P<0.05），从而有效地抑制HaCaT细胞的光老化反应。结论：牡丹籽乙醇提取物具有抗光老化作用，可以改善UVB诱导的氧化应激和炎症反应。

Abstract: Objective: To study the protective effect and mechanism of ethanol extract from Paeonia suffruticosa seed on photoaging of HaCaT cells induced by UVB. Method: The anti-photoaging active ingredients in the ethanol extract of Paeonia suffruticosa seed were analyzed using ultra-performance liquid chromatography/quadrupole time-of-flight-tandem mass spectrometry (UPLC-Q-TOF-MS), and the photoaging cell model was established by stimulating HaCaT cells with UVB. Cell viability was determined by the multiple table tournament (MTT) method. The effect of ethanol extract from Paeonia suffruticosa seed on cell migration was detected by cell scratch assay. The related cytokines affecting senescence such as interleukin-1 (IL-1), interleukin-6 (IL-6), interleukin-22 (IL-22), interferon-γ (IFN-γ), and tansforming growth factor-β (TGF-β) were detected by enzyme-linked immunosorbent (ELISA) experiment, and the levels of reactive oxygen species (ROS) and nuclear factor-κ-gene binding (NF-κB) were used as indicators to study the anti-photoaging activity and mechanism of Paeonia suffruticosa seed ethanol extract at different concentrations on HaCaT cells. Results: The top five components with the highest content were 1-stearoylglycerol (10.73%), erucamide (3.17%), 5-(2R,3S)-6-hydroxy-2-(4-hydroxyphenyl)-4-(E)-2-(4-hydroxyphenyl)ethenyl-2,3-dihydro-1-benzofuran-3-ylbenzene-1,3-diol (2.20%), α,α-trehalose (1.90%) and albiflorin (1.45%). After treatment with different concentrations of Paeonia suffruticosa seed ethanol extract, the viability of HaCaT cells was above 80%. The concentration of 6.25 μg/mL of Paeonia suffruticosa seed ethanol extract showed no significant cytotoxicity to HaCaT cells. Both the concentrations of 12.5 μg/mL (H group) and 6.25 μg/mL (L group) of Paeonia suffruticosa seed ethanol extract could inhibit the migration ability of HaCaT cells. The H group significantly reduced the levels of IL-1, IL-6, IL-22, IFN-γ, and TGF-β in cells (P<0.05), thereby resisting skin aging. The L group reduced ROS production and decreased the content of NF-κB protein factor (P<0.05), effectively inhibiting the photoaging response of HaCaT cells. Conclusion: Ethanol extract from Paeonia suffruticosa seed can improve oxidative stress and inflammation induced by UVB, and has the effect of anti-photoaging.