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目的通过体内外实验实验考察薯蓣皂苷(Dio)对肝的毒性。方法①采用MTT法检测薯蓣皂苷对人肝细胞(L-02)活力的影响,并计算IC50值;②以不同浓度的薯蓣皂苷与肝细胞共培养后,检测细胞上清液中的丙氨酸转氨酶(ALT),碱性磷酸酶(ALP)和乳酸脱氢酶(LDH)含量,并采用倒置相差显微镜对细胞的形态进行观察;③采用Hoechst-PI双标染色法观察薯蓣皂苷致肝细胞的凋亡情况;④取KM种小鼠,分为正常组和给药组,每组8只,给药组小鼠每天尾静脉注射10 mg·kg-1薯蓣皂苷,正常组小鼠每天给予等容量生理盐水,连续7 d,于末次给药后1 h处理小鼠,观察血清肝功能和肝组织病理学变化。结果①MTT法显示,薯蓣皂苷对肝细胞活力有明显的抑制作用,且有一定的量效关系,其IC50为2.25μg.ml-1;②10μg.ml-1的薯蓣皂苷能显著升高肝细胞上清中的AST,ALP,LDH,2.5和5μg.ml-1的薯蓣皂苷能升高肝细胞上清中的ALP(P<0.01),细胞形态学检查显示随着薯蓣皂苷浓度剂量的增加,肝细胞不同程度的减少、变圆、甚至死亡;③Ho-echst-PI双标染色结果显示2.5～10μg.ml-1的薯蓣皂苷使肝细胞出现不同程度的凋亡和坏死,且随着浓度增加肝细胞凋亡和坏死的程度增加;④整体实验表明,与正常组相比,给药组小鼠血清中的AST水平出现明显的升高(P<0.05),1只小鼠出现肝细胞广泛水肿,2只小鼠肝组织内偶见巨核肝细胞现象。结论大剂量的薯蓣皂苷对肝脏具有一定的毒性。
Objective To study the toxic effects of diosgenin (Dio) on liver in vitro and in vivo. Methods ① The effects of diosgenin on the activity of human hepatocytes (L-02) were detected by MTT method and the IC50 values were calculated. ② After co-cultured with different concentrations of diosgenin and hepatocytes, the alanine (ALT), lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) were detected by inverted phase contrast microscope. ③Hoechst-PI double staining was used to observe the changes of the content of diosgenin in hepatocytes KM mice were divided into normal group and administration group, with 8 rats in each group. The mice in the treatment group were injected intravenously with 10 mg · kg -1 diosgenin every day, Volume of normal saline for 7 days, the mice were treated 1 h after the last administration, and the changes of serum liver function and pathological changes were observed. Results ① MTT method showed that diosgenin significantly inhibited the viability of hepatocytes and had a dose-response relationship with an IC50 of 2.25 μg · ml-1. ② Diosgenin at 10 μg · ml-1 significantly increased hepatocytes Elemental AST, ALP, LDH, diosgenin of 2.5 and 5μg.ml-1 in Qing could increase ALP in the supernatant of hepatocytes (P <0.01). Morphological examination showed that as the dosage of diosgenin increased, Hoechst-PI double staining showed that 2.5 ~ 10μg.ml-1 of diosgenin in hepatocytes to varying degrees of apoptosis and necrosis, and with increasing concentration of liver The degree of apoptosis and necrosis increased; ④ The whole experiment showed that compared with the normal group, the level of AST in the serum of the mice in the treated group increased significantly (P <0.05), and one mouse showed extensive edema , Occasionally megakaryocyte phenomenon in the liver of 2 mice. Conclusion High-dose diosgenin is toxic to the liver.