蔷薇红景天总黄酮对Aβ25-35诱导HT22细胞损伤的保护作用

Protective effect of total flavonoids of Rhodiola rosea on Aβ25-35-induced HT22 cells injury

中文关键词:  蔷薇红景天  总黄酮  β-淀粉样蛋白25-35  小鼠海马神经元细胞  抗炎作用

英文关键词:Rhodiola rosea L.  total flavonoids  amyloid β-protein 25-35  hippocampal neuronal cell line cells  anti-inflammatory effect

中文摘要:

      目的 探讨蔷薇红景天总黄酮提取物(total flavonoids extract, TFE)、纯化物(total flavonoids purification, TFP)及其活性成分草质素对β-淀粉样蛋白25-35 (amyloid β-protein 25-35, Aβ25?35)诱导的小鼠海马神经元(hippocampal neuronal cell line, HT22)细胞损伤的保护作用。方法 采用Aβ25?35诱导HT22细胞损伤, 建立阿尔茨海默病(Alzheimer’s disease, AD)炎症细胞模型, 然后将HT22细胞分为空白组、炎症模型组(15 μmol/L Aβ25?35)、阳性对照盐酸多奈哌齐组(12.5 μmol/L)、不同质量浓度TFE (12.5、25、50 μg/mL)和TFP(6.25、12.5、25 μg/mL)干预组、草质素(5、10、20 μmol/L)干预组, 采用倒置显微镜观察细胞形态; MTT法测定HT22细胞存活率; 采用2,4-二硝基苯肼显色法检测乳酸脱氢酶(lactate dehydrogenase, LDH)的活性; 采用黄嘌呤氧化酶法检测超氧化物歧化酶(superoxide dismutase, SOD)的含量; 膜联蛋白V-异硫氰酸荧光素(fluorescein isothiocyanate, FITC)/碘化丙啶(propidium iodide, PI)双染色法检测细胞凋亡; 酶联免疫法(enzyme linked immunosorbent assay, ELISA)检测细胞培养液上清中肿瘤坏死因子-α (tumor necrosis factor-α, TNF-α)、白细胞介素-6 (interleukin-6, IL-6)、白细胞介素-1β (interleukin-1β, IL-1β)和环氧合酶-2 (cyclooxygenase-2, COX-2)的水平。结果 与空白组比较, 模型组神经元细胞变圆, 细胞核皱缩, 数量减少, 细胞间隙增大, 存活率显著下降(P<0.05), 细胞凋亡率显著提高(P<0.05); 与模型组比较, 蔷薇红景天TFE、TFP和草质素组的高、中、低剂量组均能改善细胞形态, 减少细胞受损, 提高细胞存活率, 显著降低细胞培养液上清中TNF-α、IL-1β、IL-6和COX-2的水平(P<0.05), 且成剂量依赖性。结论 蔷薇红景天TFE和TFP及其活性成分草质素对Aβ25?35诱导HT22细胞损伤具有较好保护作用, 其机制可能与拮抗Aβ25?35的细胞毒性、保护神经元、抑制HT22细胞炎症的发生有关。

英文摘要:

      Objective To investigate the protective effects of the total flavonoids extract (TFE), total flavonoids purification (TFP) and its active component herbacetin of Rhodiola rosea L. on Aβ25?35-induced HT22 cells injury. Methods Alzheimer’s disease (AD) inflammatory cell model was established by Aβ25?35 induced HT22 cells injury, HT22 cells were divided into blank group, inflammatory model group (15 μmol/L Aβ25?35), positive control donepezil hydrochloride group (12.5 μmol/L), different mass concentrations of TFE groups (12.5, 25, 50 μg/mL) and TFP groups (6.25, 12.5, 25 μg/mL), herbacetin groups (5, 10, 20 μmol/L), cell morphology was observed under inverted microscope; the survival rates of HT22 cells were determined by MTT assay; the activities of lactate dehydrogenase (LDH) was detected by 2,4-dinitrophenylhydrazine colorimetry; the content of superoxide dismutase (SOD) was detected by xanthine oxidase method; apoptosis was detected by annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) double staining method; enzyme linked immunosorbent assay (ELISA) was used to detect tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β) and cyclooxygenase-2 (COX-2) levels in cell supernatants. Results Compared with the blank group, the morphology of the neuron cells in the model group became round, nucleus shrank, number decreased, the intercellular space increased, the survival rate decreased significantly (P<0.05), and the apoptosis rate increased significantly (P<0.05); compared with the model group, the high, medium and low dose groups of TFE, TFP and herbacetin of Rhodiola rosea L. can improve the cell morphology, improve the survival rates of damaged cells and significantly reduce the levels of TNF-α, IL-1β, IL-6 and COX-2 in cell culture medium supernatant (P<0.05), which was in a dose-dependent manner. Conclusion TFE, TFP and its active component herbacetin of Rhodiola rosea L. have better protective effects on Aβ25?35-induced HT22 cells, and its mechanism may be related to antagonizing the cytotoxicity of Aβ25?35, protecting neurons, and inhibiting the occurrence of HT22 cell inflammation related.