This experiment was conducted to study the effects of tea saponin on proliferation and milk fat syn⁃thesis key enzymes of bovine mammary epithelial cells. Mammary epithelial cells were obtained by the method of enzyme digestion. After immunofluorescence identification was done, mammary epithelial cells were cul⁃tured in tea saponin solution of different concentrations [0 (control), 0.05, 0.25, 0.50, 1.00, 5.00, 10.00, 20.00, 40.00, 60.00, 80.00 and 100.00 μg/mL]. 1) Thiazolyl blue tetrazolium bromide (MTT) method was used to detect the effects of tea saponin on the proliferation of cells. 2) Enzyme⁃linked immunosorbent as⁃say ( ELISA) kit was used to measure acetyl⁃CoA carboxylase ( ACACA) , fatty acid synthase ( FASN) and stearoyl⁃coenzyme A desaturase ( SCD ) contents in cells. 3 ) Fluorescence⁃based quantitative real⁃time PCR method was used to measure the relative expression level of mRNA of ACACA, FASN and SCD. The results showed as follows:1) compared with control group, there was no significant effect of tea saponin at concen⁃tration of 0.05 to 5.00 μg/mL ( P>0.05) , but the cell proliferation was significantly inhibited by tea saponin at concentration of 10.00 to 100.00 μg/mL ( P<0.05); 2) the contents of FASN, ACACA and SCD in cells didn’ t significantly change among control group and 0.50, 5.00 and 20.00 μg/mL concentration groups ( P>0.05); 3) compared with control group, the relative expression level of SCD mRNA in 0. 50, 5. 00 and 20.00 μg/mL concentration groups was significantly decreased ( P<0.05) . In conclusion, tea saponin plays a role in inhibition of proliferation and the relative expression level of fat synthesis key enzyme gene SCD mRNA of bovine mammary epithelial cells.%本试验旨在研究茶皂素对乳腺上皮细胞增殖、乳脂合成关键酶的影响。采用酶消化法分离获取乳腺上皮细胞，经免疫荧光鉴定后，分别添加不同浓度[0（对照）、0．05、0．25、0．50、1．00、5．00、10．00、20．00、40．00、60．00、80．00、100．00μg／mL]的茶皂素溶液培养，然后进行如下操作：1）采用噻唑蓝（ MTT）法检测不同浓度的茶皂素对乳腺上皮细胞增殖的影响；2）采用酶联免疫分析（ ELISA）试剂盒测定细胞中乙酰辅酶A羧化酶（ ACACA）、脂肪酸合成酶（ FASN）、硬脂酰辅酶A去饱和酶（ SCD）的含量；3）采用实时荧光定量PCR的方法测定FASN、ACACA、SCD基因mRNA相对表达水平。结果显示：1）茶皂素浓度为0．05～20．00μg／mL对细胞增殖无显著影响（P＞0．05），浓度为20．00～100．00μg／mL时显著抑制细胞增殖（P＜0．05）；2）细胞FASN、ACACA、SCD的含量在对照组和0．50、5．00、20．00μg／mL浓度组间无显著差异（ P＞0．05）；3）细胞与茶皂素共育24、48 h后，与对照组相比，0．50、5．00、20．00μg／mL 浓度组SCD基因mRNA相对表达水平显著降低（ P＜0．05）。综合以上，茶皂素对奶牛乳腺上皮细胞增殖及乳脂合成关键酶SCD基因mRNA表达具有一定抑制作用。
展开▼
