竹节参皂苷Ⅳa抑制THP-1源性巨噬细胞泡沫化的研究

Chikusetsu saponinⅣa mitigate foam production of THP-1 derived macrophages

目的研究竹节参皂苷Ⅳa(CsⅣa)抑制THP-1源性巨噬细胞泡沫化的作用机制。方法培养THP-1源性巨噬细胞,分为对照组、模型组和低、高剂量实验组。对照组常规培养,模型组细胞用50μg·mL^(-1)氧化低密度脂蛋白(ox-LDL)处理,低、高剂量实验组分别用25,50μmol·L^(-1)的CsⅣa处理1 h,再用50μg·mL^(-1)ox-LDL处理。用细胞计数法-8(CCK-8)检测THP-1源性泡沫细胞活力,用油红O法检测巨噬细胞泡沫化程度,用酶联免疫吸附(ELISA)法检测肿瘤坏死因子-α(TNF-α)和白细胞介素-1β(IL^(-1)β)水平,用逆转录聚合酶链反应(RT-PCR)法检测微管相关蛋白轻链3(LC3)、p62、Beclin1、TNF-α、IL^(-1)β的mRNA水平,用蛋白质印迹法检测LC3、p62、Beclin1蛋白水平。结果对照组、模型组和低、高剂量实验组TNF-α分别为(0.68±0.07),(1.24±0.13),(0.95±0.05)和(0.80±0.03)pg·mL^(-1),IL^(-1)β为(1.60±0.09),(2.14±0.14)和(1.90±0.16),(1.73±0.13)pg·mL^(-1),Beclin 1蛋白表达水平分别为1.48±0.09,0.83±0.07,1.45±0.05和1.62±0.19,p62蛋白表达水平分别为0.52±0.07,0.82±0.08,0.72±0.04和0.41±0.04,LC3Ⅱ/Ⅰ蛋白表达水平分别为3.12±0.23,3.06±0.25,3.66±0.42和4.05±0.23。以上数据,模型组与对照组相比,差异均有统计学意义(均P<0.05),低、高剂量实验组与模型组相比,差异均有统计学意义(均P<0.05)。结论CsⅣa治疗动脉粥样硬化的作用机制可能与促进巨噬细胞自噬、抑制泡沫化的发生、降低炎性因子有关。

Objective To investigate the mechanism of Chikuseksu saponinⅣa(CsⅣa)inhibiting the foam of THP-1 derived macrophages.Methods THP-1-derived macrophages were cultured,divided into control group,model group and low,high dose experimental groups.Control group was conventionally cultured;model group was treated with oxidized low-density lipoprotein(ox-LDL);low,high dose experimental groups were treated with 25,50μmol·L^(-1) CsⅣa for 1 h,and then treated with 50μg·mL^(-1) ox-LDL.Cell counting kit-8(CCK-8)was used to detect the viability of THP-1 derived foam cells.The level of macrophage foam was detected by oil red O;enzyme-linked immunosorbent assay(ELISA)was used to detect the secretion levels of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL^(-1)β).Reverse transcription-polymerase chain reaction(RT-PCR)was used to detect the mRNA levels of microtubule-associated protein light chain 3(LC3),p62,Beclin-1,TNF-αand IL^(-1)β.Western blot was used to detect the protein levels of LC3,p62 and Beclin-1.Results TNF-αlevels in control group,model group and low,high dose experimental groups were(0.68±0.07),(1.24±0.13),(0.95±0.05),(0.80±0.03)pg·m L^(-1),IL^(-1)βlevels were(1.60±0.09),(2.14±0.14),(1.90±0.16),(1.73±0.13)pg·m L^(-1);Beclin 1 protein expression levels were 1.48±0.09,0.83±0.07,1.45±0.05,1.62±0.19;p62 protein expression levels were 0.52±0.07,0.82±0.08,0.72±0.04,0.41±0.04;LC3Ⅱ/Ⅰprotein expression levels were 3.12±0.23,3.06±0.25,3.66±0.42,4.05±0.23,respectively.In the above data,there were statistically significant differences between model group and control group(all P<0.05),and between low,high dose experimental groups and model group(all P<0.05).Conclusion The mechanism of CsⅣa in the treatment of atherosclerosis may be related to promoting macrophage autophagy,inhibiting the foaming of foam and reducing inflammatory factors.