通关藤苷H抗结肠癌作用的研究

Study on the effect of Tenacissoside H on colon cancer

目的探讨通关藤苷H(TSH)对人结肠癌LoVo细胞增殖、迁移和凋亡的影响,及其对高尔基磷蛋白3(GOLPH3)的调控作用。方法实验1:分为低、中低、中、高剂量组和对照1组,分别给予0.1,1.0,10.0和100.0μg·mL^(-1) TSH及等量0.9%NaCl干预,处理24,48和72 h;实验2:分为(1)对照2组,0.9%NaCl;(2)TSH组,TSH;(3)空载组,p-CMV-control+0.9%NaCl;(4)GOLPH3组,p-CMV-2-GOLPH3+0.9%NaCl;(5)实验1组,p-CMV-control+TSH;(6)实验2组,p-CMV-2-GOLPH3+TSH;本部分实验的TSH质量浓度均为25μg·mL^(-1),干预24 h,检测细胞增殖活力、GOLPH3及转录活化因子3(ATF3)的表达水平、迁移能力和凋亡情况。结果TSH呈浓度依赖性抑制LoVo细胞增殖。TSH组和对照2组的ATF3蛋白相对表达水平分别为0.08±0.02和1.00±0.15。空载组、TSH组、GOLPH3组、实验1组、实验2组和对照2组的GOLPH3蛋白相对表达量分别为1.00±0.27,0.08±0.06,3.82±0.52,0.17±0.12,3.58±0.55和1.00±0.12,迁移细胞数分别为(301±26),(47±12),(372±55),(39±14),(284±31)和(293±64)个,细胞凋亡率分别为(1.11±0.65)%,(31.77±3.47)%,(0.34±0.35)%,(30.06±4.86)%,(2.27±1.19)%和(0.51±0.54)%。TSH组的GOLPH3、ATF3表达水平、细胞迁移数和细胞凋亡率与对照2组比较,差异均有统计学意义(P<0.05,P<0.01);GOLPH3组的GOLPH3的蛋白表达水平与空载组比较,差异有统计学意义(P<0.05);实验2组中GOLPH3的蛋白表达水平与实验1组比较,差异有统计学意义(P<0.05);实验1组的细胞迁移数、细胞凋亡率与空载组比较,差异均有统计学意义(均P<0.01);实验2组的细胞迁移数、细胞凋亡率与实验1组比较,差异均有统计学意义(均P<0.01)。结论TSH可通过调控GOLPH3的表达,抑制人结肠癌LoVo细胞体外增殖和细胞迁移,并诱导凋亡。

Objective To investigate the effect of Tenacissoside H(TSH)on the proliferation,migration and apoptosis as well as its regulation on Golgi phosphoprotein 3(GOLPH3)expression in human colon cancer LoVo cells.Methods Experiment 1:They were divided into low,middle-low,middle and high dose groups and control 1 group,which were treated with 0.1,1.0,10.0 and 100.0μg·mL^(-1) TSH and the same amount of 0.9%NaCl respectively for 24,48 and 72 hours.Experiment 2:They were divided into(1)control 2 group,0.9%NaCl;(2)TSH group,TSH;(3)empty vector group,p-CMV-control+0.9%NaCl;(4)GOLPH3 group,p-CMV-2-GOLPH3+0.9%NaCl;(5)experimental 1 group,p-CMV-control+TSH;(6)experimental 2 group,p-CMV-2-GOLPH3+TSH;the concentration of TSH in this part of experiment was 25μg·m L^(-1),treatment for 24 h,the cell proliferation activity,the expression of GOLPH3 and Activating transcription factor 3(ATF3),migration ability and apoptosis were detected.Results TSH inhibited LoVo cell proliferation in a concentration dependent manner.The expression levels of ATF3 in TSH group and control 2 group were 0.08±0.02and 1.00±0.15.In empty vector,TSH,GOLPH3,experimental 1,experimental 2 and control 2 groups,the relative expression levels of GOLPH3 protein were 1.00±0.27,0.08±0.06,3.82±0.52,0.17±0.12,3.58±0.55 and1.00±0.12;the number of migrating cells were(301±26),(47±12),(372±55),(39±14),(284±31)and(293±64);the apoptosis rates were(1.11±0.65)%,(31.77±3.47)%,(0.34±0.35)%,(30.06±4.86)%,(2.27±1.19)%and(0.51±0.54)%.The expression levels of GOLPH3 and ATF3,cell migration and apoptosis rate in TSH group were compared with those in control 2 group and the differences were statistically significant(P<0.05,P<0.01).The protein expression levels of GOLPH3 in GOLPH3 group was compared with that in empty vector group and the difference was statistically significant(P<0.05).The protein expression levels of GOLPH3 in experimental 2 group was compared with that in experimental 1 group and the difference was statistically significant(P<0.05).The number of cell migration and apoptosis rate in experimental 1 group was compared with that in empty vector group and the difference was statistically significant(all P<0.01).The number of cell migration and apoptosis rate in experimental 2 group was compared with that in experimental 1 group and the difference was statistically significant(all P<0.01).Conclusion TSH can inhibit the proliferation and migration of human colon cancer LoVo cells in vitro and induce apoptosis by regulating the expression of GOLPH3.