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目的建立猪肾近曲小管上皮细胞(LLC-PK1)的高糖损伤模型,观察α-亚麻酸(ALA)对高糖损伤LLC-PK1细胞的保护作用并探讨其作用机制。方法 CCK-8试剂盒测定葡萄糖对LLC-PK1细胞增殖的影响,流式细胞术测定不同浓度ALA干预高糖损伤LLC-PK1的凋亡率和活性氧(ROS)含量。结果高糖环境可以抑制体外培养的LLC-PK1细胞的增殖,形成体外高糖损伤模型;经适当浓度(50～100μmol/L)的ALA干预后,前干预组和持续干预组细胞的凋亡率显著低于阳性对照组(P<0.05);当ALA浓度为10～100μmol/L时,持续干预组LLC-PK1细胞内ROS含量显著低于阳性对照组(P<0.05),当ALA浓度为50μmol/L时,前干预组LLC-PK1细胞内ROS含量显著低于阳性对照组(P<0.05)。结论高糖损伤LLC-PK1模型为研究DN肾小管上皮细胞的防治干预提供了良好的体外研究平台,ALA有望成为预防肾小管高糖损伤的保护剂,减少活性氧的产生可能是ALA保护肾小管上皮细胞的作用机制之一。
OBJECTIVE: To establish a high glucose injury model of porcine renal proximal tubule epithelial cells (LLC-PK1) and to observe the protective effect of α-linolenic acid (ALA) on LLC-PK1 cells injured by high glucose and to explore its mechanism. Methods CCK-8 kit was used to determine the effect of glucose on the proliferation of LLC-PK1 cells. Flow cytometry was used to determine the apoptotic rate and reactive oxygen species (ROS) of LLC-PK1 cells treated with different concentrations of ALA. Results High glucose environment inhibited the proliferation of LLC-PK1 cells in vitro and formed a model of high glucose injury in vitro. After intervention with ALA at the appropriate concentration (50-100 μmol / L), the apoptosis rate of cells in the pre-intervention group and the sustained intervention group (P <0.05). When the concentration of ALA was 10 ~ 100μmol / L, the content of ROS in LLC-PK1 cells in continuous intervention group was significantly lower than that in positive control group (P <0.05). When ALA concentration was 50μmol / L, the level of ROS in LLC-PK1 cells in the pre-intervention group was significantly lower than that in the positive control group (P <0.05). Conclusion The high glucose-induced LLC-PK1 model provides a good in vitro research platform for the study of DN tubular epithelial cells in prevention and treatment. ALA is expected to become a protective agent against renal injury induced by high glucose. Reducing reactive oxygen species may be one of the mechanisms by which ALA protects renal tubules Epithelial cells one of the mechanisms.