石斛碱修复Caco-2+HT29-MTX/ RAW 264.7细胞肠屏障并改善HepG2细胞胰岛素抵抗的双重作用

The Dual Effects of Dendrobine on Repairing Caco-2+HT29-MTX/RAW 264.7 Cell Intestinal Barrier and Improving Insulin Resistance in HepG2 Cells

摘要:

为探讨石斛碱修复肠屏障损伤和改善胰岛素抵抗的作用，研究建立Caco-2/HT29-MTX/RAW264.7共培养细胞模型和胰岛素抵抗HepG2细胞模型，采用跨上皮电阻（TEER）、实时荧光定量PCR和蛋白印迹等技术进行评估。结果表明，相较模型组，石斛碱能维持TEER值在200 Ω·cm2以上，上调紧密连接蛋白Claudin-1（72.19%）、Occludin（82.40%）、ZO-1（75.29%）和黏液蛋白MUC2（70.33%）表达水平，其mRNA表达量分别提高1.47%、 452.93%、2.11%、10.38%。同时降低促炎因子TNF-α（14.14%）和IL-6（68.93%）水平，提升抗炎因子IL-4（243.96%）和IL-10（41.35%）水平。在胰岛素抵抗模型中，石斛碱以剂量依赖方式增加葡萄糖摄取量（10.70 mmol·L-1、11.83 mmol·L-1、13.04 mmol·L-1），提高丙酮激酶（92.14 U·g-1 protein、107.09 U·g-1 protein、122.02 U·g-1 protein）和己糖激酶（29.63 U·g-1 protein、73.99 U·g-1 protein、73.84 U·g-1 protein）活性，并使糖原含量提高1.2、1.4、1.5倍。综上所述，石斛碱能够有效修复肠道屏障损伤并显著改善胰岛素抵抗，为开发降血糖功能食品和药物提供理论基础。

Abstract:

To investigate the effect of Dendrobine (DEN) on repairing intestinal barrier damage and improving insulin resistance, Caco-2/HT29-MTX/RAW264.7 co-culture cell models and insulin resistance HepG2 cell models were established, and evaluated using transepithelial electrical resistance (TEER), real-time quantitative PCR, and Western blot techniques. The results showed that compared to the model group, DEN maintained the TEER value above 200 Ω·cm2, and upregulated the expression levels of tight junction proteins Claudin-1 (72.19% increase), Occludin (82.40% increase), ZO-1 (75.29% increase), and mucin protein MUC2 (70.33% increase). The mRNA expression levels of Claudin-1 (1.47%), Occludin (452.93%), ZO-1 (2.11%), and MUC2 (10.38%) were also significantly increased. In addition, the levels of pro-inflammatory factors TNF-α (14.14%) and IL-6 (68.93%) were significantly reduced, while the levels of anti-inflammatory factors IL-4 (243.96%) and IL-10 (41.35%) were significantly elevated. In the insulin resistance model, DEN increased glucose uptake in a dose-dependent manner (10.70,mmol·L-1 11.83 mmol·L-1, 13.04 mmol·L-1), enhanced the activity of pyruvate kinase (92.14 U·g-1 protein, 107.09 U·g-1 protein, 122.02 U·g-1 protein, ) and hexokinase (29.63 U·g-1 protein, 73.99 U·g-1 protein, 73.99 U·g-1 protein, ), and increased glycogen content by 1.2, 1.4, and 1.5 times, respectively. In conclusion, Dendrobine effectively repaired intestinal barrier damage and significantly improved insulin resistance, providing a theoretical basis for the development of anti-hyperglycemic functional foods and drugs.