消癌解毒方诱导人肝癌SMMC-7721细胞miRNA表达变化

目的:探讨消癌解毒方对人肝癌SMMC-7721细胞增殖及微小核糖核酸(microRNA,miR)-25-3p,miR-29a-5p,miR-122-3p,miR-124-3p,miR-182-5p表达谱的影响。方法:将12只雄性大耳白兔随机分为4组,分别为消癌解毒方高、中、低剂量(15.12,7.56,3.78 g·kg-1)组和空白组,各组分别给予消癌解毒方及生理盐水灌胃4 d。4 d后于颈动脉中取血清并配制培养基。用消癌解毒方(15.12,7.56,3.78 g·kg-1)含药血清及空白血清的培养基处理人肝癌细胞株SMMC-7721,噻唑蓝(MTT)比色法检测肿瘤细胞增殖活性、实时荧光定量逆转录聚合酶链式反应(Real-time polymerase chain reaction,Real-time PCR)法验证人肝癌细胞SMMC-7721中miR-25-3p,miR-29a-5p,miR-122-3p,miR-124-3p和miR-182-5p的表达水平。结果:经过12,24,48 h后,消癌解毒方(15.12,7.56,3.78 g·kg-1)含药血清对人肝癌SMMC-7721细胞的增殖均存在抑制作用。随着消癌解毒方含药血清浓度的增加,其对人肝癌细胞SMMC-7721细胞增殖的抑制率也相应增加。其中高剂量组含药血清的抑制效果最好,其干预12,48 h的吸光度A较同期空白组明显降低(P0.05)。高剂量组干预24 h A比同期空白组显著降低(P0.01),该组抑制率为42.86%。Real-time PCR证实消癌解毒方(15.12,7.56,3.78 g·kg-1)含药血清的培养基能诱导miR-25-3p,miR-182-5p表达下调,诱导miR-29a-5p,miR-122-3p,miR-124-3p表达上调。且高剂量组的调控作用较空白组更显著(P0.01)。结论:消癌解毒方可能通过诱导miRNA表达谱的改变而参与抑制人肝癌细胞SMMC-7721增殖作用,但其具体机制仍有待进一步研究。

miRNA Expression of SMMC-7721 Cell Induced by Xiaoai Jiedu Formula

Objective: To study the effect of Xiaoai Jiedu formula on the proliferation of human hepatocarcinoma SMMC-7721 cells and expression profiles of microRNA(miR)-25-3p, miR-29a-5p, miR-122-3p, miR-124-3p and miR-182-5p. Method: Twelve male white rabbits were randomly divided into four groups. They were respectively given high-dose, middle-dose and low-dose Xiaoai Jiedu formula or normal saline for four days. Serum was taken from their carotid blood and then made into culture medium. Cytotoxicity of Xiaoai Jiedu formula against SMMC-7721 cells was measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Expression profiles of miR-25-3p, miR-29a-5p, miR-122-3p, miR-124-3p and miR-182-5p of SMMC-7721 cells were analyzed by a miRNA array and Real-time polymerase chain reaction (Real-time PCR). Result: High-dose, middle-dose and low-dose Xiaoai Jiedu formula could all inhibit the proliferation of SMMC-7721 cells after 12, 24, 48 h. Xiaoai Jiedu formula gradually increased the proliferation inhibition rate of SMMC-7721 cells in a dose-dependent manner, with the increase of drug concentration. The absorbancy of the high-dose group showed statistical differences after 12 h and 48 h compared with blank control group at the same time points (P<0.05). The absorbancy of the high-dose group showed statistically significant differences after 24 h compared with the blank control group at the same time point (P<0.01). The absorbancy of the high dose group after intervention for 24 h was significantly lower than that of blank control group (P<0.01), the group''s inhibition rate was 42.86%. Moreover, Real-time PCR verified the down-regulation of miR-25-3p, miR-182-5p and the up-regulation of miR-29a-5p, miR-122-3p, miR-124-3p induced by Xiaoai Jiedu formula. The regulatory effect of the high-dose group was more significant than that of the blank control group (P<0.01). Conclusion: Xiaoai Jiedu formula may inhibit the proliferation of SMMC-721 by inducing the change in microRNA (miRNA) expression. However, the concrete mechanism remains to be further studied.