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目的研究乳移平含药血清对小鼠乳腺癌4T1细胞增殖与侵袭的影响,并探讨其作用机制。方法 (1)制备不同浓度的乳移平含药血清(10%、20%、30%、40%),并以10%、20%、30%、40%空白血清作对照,采用噻唑蓝(MTT)方法测定含药血清对4T1细胞存活率的影响,筛选最佳作用浓度和最佳作用时间。(2)设空白对照组、TGF-β1对照组、TGF-β1 10%乳移平含药血清组、TGF-β120%乳移平含药血清组;除空白对照组外,其他组细胞都用2 mg/L的TGF-β1预处理3 h,再分别选择空白血清或含药血清作用48 h。采用Transwell侵袭实验检测含药血清对TGF-β1预处理的4T1细胞侵袭率的影响;Western blotting及Real-time PCR检测乳移平含药血清对经TGF-β1预处理的4T1细胞Smad4和Smad7表达的影响。结果 (1)乳移平含药血清显著抑制4T1细胞的增殖活性;选择10%和20%作用浓度,48 h作用时间用于后续实验。(2)乳移平含药血清显著抑制TGF-β1诱导的4T1高侵袭能力,并且存在浓度依赖性;Western blotting和Real-time PCR结果显示,乳移平含药血清明显逆转了TGF-β1诱导的Smad4蛋白及mRNA的高表达,但对TGF-β1诱导的Smad7蛋白及mRNA表达量上调没有影响。结论乳移平含药血清能够抑制4T1的增殖和侵袭能力,其作用机制可能与调控TGF-β/Smads信号通路相关因子的表达有关。
Objective To study the effects of Ru Miping Ping serum on the proliferation and invasion of mouse breast cancer 4T1 cells and to explore its mechanism. Methods (1) Different concentrations of milk serum containing serum (10%, 20%, 30%, 40%) were prepared and compared with 10%, 20%, 30% and 40% MTT) method was used to determine the effect of drug-containing serum on the survival rate of 4T1 cells. The optimum concentration and time of screening were selected. (2) The blank control group, TGF-β1 control group, TGF-β1 10% milk serum containing serum group, TGF-β120% milk serum containing serum group; except the blank control group, the other groups were used 2 mg / L of TGF-β1 pretreatment 3 h, and then were selected blank serum or drug-containing serum for 48 h. Transwell assay was used to detect the effect of serum containing TGF-β1 on the invasion of 4T1 cells. Western blotting and Real-time PCR were used to detect the expression of Smad4 and Smad7 in 4T1 cells pretreated with TGF-β1 Impact. Results (1) The milk serum contained serum significantly inhibited the proliferation activity of 4T1 cells; selected concentration of 10% and 20%, 48 h action time for subsequent experiments. (2) The milk serum contained serum significantly inhibited TGF-β1-induced 4T1 high invasive ability, and there is a concentration-dependent manner; Western blotting and Real-time PCR results show that milk serum containing serum significantly reversed TGF-β1 induced Smad4 protein and mRNA, but had no effect on TGF-β1-induced Smad7 protein and mRNA expression. CONCLUSION: Milk-emulsion-containing serum can inhibit the proliferation and invasion ability of 4T1 and its mechanism may be related to the regulation of TGF-β / Smads signaling pathway related factors.