制首乌含药血清对人乳腺癌T-47D细胞增殖及ER表达的影响
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篇名: 制首乌含药血清对人乳腺癌T-47D细胞增殖及ER表达的影响
TITLE:
摘要: 目的:研究制首乌含药血清对人乳腺癌T-47D细胞增殖及雌激素受体(ER)表达的影响,探讨其植物雌激素(PE)样作用。方法:将性未成熟SD大鼠随机分为空白组,戊酸雌二醇(Ev)组(阳性对照,0.12 mg/kg),制首乌低、高剂量组(0.75、3 g/kg,以生药量计)以及制首乌低、高剂量+Ev联用组(剂量同各药单用组),每组10只。空白组大鼠灌胃等体积水,各给药组大鼠灌胃相应药物,早晚各1次,连续4 d。末次给药2 h后采血,制备空白血清和含药血清。将T-47D细胞随机分为空白组,Ev组,制首乌低、高剂量组以及制首乌低、高剂量+Ev联用组,分置于含20%空白或相应含药血清的培养基中培养,采用CCK-8法检测各组细胞的增殖率(PR),采用Western blotting法和逆转录-聚合酶链反应法检测ER-α、ER-β蛋白及其mRNA的表达情况。结果:与空白组比较,各给药组细胞的PR[各给药组(24 h),除制首乌高剂量+Ev联用组外的其余各给药组(48、72 h)]均显著升高;且制首乌高剂量组(72 h)显著高于Ev组,制首乌低剂量组+Ev联用组(72 h)显著高于同剂量制首乌单用组,而制首乌高剂量+Ev联用组(72 h)显著低于同剂量制首乌单用组(P<0.05或P<0.01)。Ev组、制首乌高剂量组和制首乌低剂量+Ev联用组细胞中ER-α蛋白,各给药组细胞中ER-α mRNA和ER-β蛋白以及Ev组、制首乌低剂量组和制首乌+Ev联用组细胞中ER-β mRNA的相对表达量均显著升高;Ev组细胞中ER-α蛋白及其mRNA的相对表达量均显著高于制首乌单用和联用组;Ev组细胞中ER-β蛋白及其mRNA的相对表达量均显著低于制首乌低剂量+Ev联用组,但ER-β mRNA的相对表达量显著高于制首乌单用组和制首乌高剂量+Ev联用组;制首乌低剂量+Ev联用组细胞中ER-α、ER-β蛋白及其mRNA以及制首乌高剂量+Ev联用组细胞中ER-β mRNA的相对表达量均显著高于同剂量制首乌单用组,而制首乌高剂量+Ev联用组细胞中ER-α蛋白及其mRNA相对表达量均显著低于同剂量制首乌单用组(P<0.05或P<0.01)。结论:制首乌含药血清可促进人乳腺癌T-47D细胞的增殖,并可通过促进ER-α和ER-β蛋白及其mRNA的表达来发挥PE样作用。但上述作用弱于雌激素,且两者联合可能会拮抗雌激素的作用。
ABSTRACT: OBJECTIVE: To study the effects of processed Polygonum multiflorum containing serum on the proliferation and the expression of estrogen receptor (ER) of human breast cancer T-47D cells, and to investigate its phytoestrogen (PE)-like effect. METHODS: Sexually immature SD rats were randomly divided into estradiol valerate (Ev) group (positive control, 0.12 mg/kg), processed P. multiflorum low-dose and high-dose groups (0.75, 3 g/kg, by crude drug), low-dose and high-dose processed P. multiflorum+Ev groups (same dose as single drug group), with 10 rats in each group. Blank group was given constant volume of water intragastrically, and administration groups were given relevant medicine intragastrically; once day and night, for consecutive 4 days. Two hours after last administration, blank serum and containing serum were prepared. T-47D cells were also randomly divided into blank group, Ev group, low-dose and high-dose processed P. multiflorum groups, low-dose and high-dose processed P. multiflorum+Ev groups, and then were cultured in medium which contained 20% blank serum or drug containing serum. CCK-8 assay was used to detect proliferation rate (PR). Western blotting assay and RT-PCR were used to detect the protein and mRNA expression of ER-α and ER-β. RESULTS: Compared with blank group, PR of administration groups [each administration group (24 h), other administration groups (48, 72 h) except for high-dose processed P. multiflorum+Ev group] were increased significantly; high-dose processed P. multiflorum group (72 h) was significantly higher than Ev group, and low-dose processed P. multiflorum+Ev group (72 h) was significantly higher than the same-dose processed P. multiflorum group; high-dose processed P. multiflorum+Ev group (72 h) was significantly lower than the same-dose processed P. multiflorum group (P<0.05 or P<0.01). Relative protein expression of ER-α in Ev group, high-dose processed P. multiflorum group and low-dose processed P. multiflorum+Ev group, relative mRNA expression of ER-α and protein expression of ER-β in administration groups, relative mRNA expression of ER-β in Ev group, low-dose processed P. multiflorum group and processed P. multiflorum+Ev groups were all increased significantly. Relative protein and mRNA expression of ER-α in Ev group were significantly higher than processed P. multiflorum groups and combination groups. Relative protein and mRNA expression of ER-β in Ev group were significantly lower than low-dose processed P. multiflorum+Ev group, but relative mRNA expression of ER-β was significantly higher than processed P. multiflorum groups and high-dose processed P. multiflorum+Ev group. Relative protein and mRNA expression of ER-α and ER-β in low-dose processed P. multiflorum+Ev group as well as relative mRNA expression of ER-β in high-dose processed P. multiflorum+Ev group were significantly higher than the same-dose processed P. multiflorum group. Relative protein and mRNA expression of ER-α in high-dose processed P. multiflorum+Ev group were significantly lower than the same-dose processed P. multiflorum group (P<0.05 or P<0.01). CONCLUSIONS: The processed P. multiflorum containing serum can promote the proliferation of human breast cancer T-47D cells, and play the PE-like role through promoting protein and mRNA expression of ER-α and ER-β. However, the above effects are weaker than estrogen, and the combination of the two may antagonize the effect of estrogen.
期刊: 2019年第30卷第22期
作者: 朱璨,王嫣,李尧锋,田敏,唐文超,杨长福,王和生
AUTHORS: ZHU Can,WANG Yan,LI Yaofeng,TIAN Min,TANG Wenchao,YANG Changfu,WANG Hesheng
关键字: 制首乌;含药血清;T-47D细胞;细胞增殖;雌激素受体;植物雌激素样作用
KEYWORDS: Processed Polygonum multiflorum; Containing serum; T-47D cells; Cell proliferation; Estrogen receptor; Phytoestrogen-like effect
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