The Total Flavonoids of Clerodendrum bungei Suppress A549 Cells Proliferation,Migration,and Invasion by Impacting Wnt/β-Catenin Signaling

Objectives: The objective of this study is to evaluate the effect of the total flavonoids of Clerodendrum bungei(TFCB) on the proliferation,invasion, and metastasis of A549 lung cancer cells through the Wnt signaling pathway. Materials and Methods: A549 cells were transfected with a β-catenin overexpression plasmid and the empty vector pcDNA3.1. The A549 cells were divided into six groups: normal A549 cell group, normal A549 cells with TFCB group, vector control group, vector with TFCB group,(3-catenin overexpression group, and β-catenin with TFCB group. We used the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay to detect cell proliferation, a scratch test was used to observe cell migration, and a transwell experiment was employed to evaluate cell invasion. Proteins related to the Wnt pathway were detected with Western blot analysis, including β-catenin, GSK-3 β, P-GSK-3 β, c-Myc, and CyclinD1. Results: The proliferation, invasion,and metastasis of A549 cells were significantly enhanced after being transfected with the β-catenin overexpression plasmid(P< 0.05 or 0.01),accompanied by increased expression of β-catenin, C-Myc, CyclinD1 and reduced expression of Gsk-3 β and P-GSK-3 β. Treatment of cells with TFCB resulted in inhibition of cell proliferation, migration, and invasion; downregulated expression of β-catenin, C-Myc, and CyclinD1;and upregulated expression of GSK-3 β and P-GSK-3 β,especially in the β-catenin overexpression group. Conclusion: TFCB has the potential to inhibit the Wnt/β-catenin pathway by prohibiting the overexpression of β-catenin and regulating its downstream factors.

臭牡丹总黄酮对A549细胞增殖迁移侵袭力及Wnt通路相关蛋白的影响

目的:观察臭牡丹总黄酮对β-catenin过表达A549细胞增殖、迁移、侵袭力以及Wnt通路相关蛋白的影响。方法:选取构建β-catenin真核过表达载体并转染A549细胞,将其分为6组,分别为未转染细胞、未转染细胞+臭牡丹总黄酮、空载组、空载+臭牡丹总黄酮、β-catenin过表达组、β-catenin过表达+臭牡丹总黄酮。MTT法检测各组细胞的相对存活率,划痕实验检测各组细胞迁移能力,Transwell小室侵袭实验检测各组细胞的侵袭力。Western Blot检测各组Wnt通路相关蛋白β-catenin、GSK-3β、P-GSK-3β、c-myc、CyclinD1的表达。结果:转染β-catenin过表达的A549细胞增殖能力、迁移能力、侵袭力均明显增强(P<0.05),并显著上调wnt通路相关因子β-catenin、C-Myc,CyclinD的蛋白表达,下调P-GSK-3β的蛋白表达(P<0.05)。采用臭牡丹总黄酮干预后,能明显降低各组细胞的存活率,降低细胞向划痕区域迁移的能力,减少侵袭小室穿过基膜的细胞数(P<0.05)。并下调β-catenin、C-Myc、CyclinD1表达(P<0.05),上调P-GSK-3β表达(P<0.05)。臭牡丹总黄酮的干预作用对转染β-catenin过表达细胞尤为明显。结论:转染β-catenin过表达的A549细胞其增殖、迁移、侵袭力均明显增强,能激活Wnt/β-catenin通路。臭牡丹总黄酮可抑制A549细胞的增殖、迁移、侵袭力,其机制可能是通过抑制细胞中β-catenin的高表达从而调控下游的一系列因子而起到抑制Wnt/β-catenin通路的作用。

臭牡丹中糖苷类化学成分研究

目的:研究臭牡丹Clerodendrum bungei Steud.中的糖苷类成分。方法:采用硅胶、ODS和半制备液相色谱等方法对臭牡丹70%乙醇提取物的乙酸乙酯部位进行分离纯化,并通过1D-NMR、MS等现代波谱技术对化合物结构进行鉴定。结果:从臭牡丹70%乙醇提取物的乙酸乙酯部位分离得到11个糖苷类化合物,分别鉴定为:金合欢素-7-O-β-D-葡萄糖醛酸苷(1)、田蓟苷(2)、香叶木苷(3)、芹菜素-7-O-β-D-芦丁糖苷(4)、金合欢素-7-O-β-D-芦丁糖苷(5)、香叶木素-7-O-(2″-O-β-D-葡萄糖醛酸,6″-O-α-L-鼠李糖)-β-D-葡萄糖苷(6)、金合欢素-7-O-(2″-O-β-D-葡萄糖醛酸,6″-O-α-L-鼠李糖)-β-D-葡萄糖苷(7)、芹菜素-7-O-(2″-O-β-D-葡萄糖醛酸,6″-O-α-L-鼠李糖)-β-D-葡萄糖苷(8)、松果菊苷(9)、毛蕊花糖苷(10)、异毛蕊花糖苷(11)。结论:其中,化合物1~9为首次从该植物中分离得到。

臭牡丹总黄酮通过调控Wnt/β-catenin通路影响A549细胞上皮间质转化研究

目的探讨臭牡丹总黄酮对人肺癌A549细胞增殖、迁移和侵袭的作用及其机制。方法将pcDNA3.1+空载体与S45位点突变的β-链蛋白(S45A-β-catenin)真核过表达载体转染A549细胞,给予臭牡丹总黄酮干预。MTT法检测细胞的相对存活率,划痕实验检测细胞迁移能力,Transwell小室侵袭实验检测细胞的侵袭能力,Western blotting法检测各组细胞β-catenin、上皮细胞钙黏蛋白(E-cadherin)、波形蛋白(vimentin)、转录因子Snail 2(Slug)、糖原合成酶激酶-3β(GSK-3β)、p-GSK-3β的表达水平。结果转染S45A-β-catenin质粒后,A549细胞增殖、迁移、侵袭能力均明显增强,β-catenin、vimentin蛋白表达水平显著上调,E-cadherin、GSK-3β、P-GSK-3β蛋白的表达水平显著下调。臭牡丹总黄酮可抑制A549细胞的增殖、迁移与侵袭能力,对转染S45A-β-catenin质粒组的细胞作用尤为明显。对转染空载质粒的A549细胞,可上调E-cadherin、GSK-3β、P-GSK-3β表达,下调β-catenin、vimentin表达;对转染S45A-β-catenin质粒的A549细胞,可下调β-catenin、vimentin表达。结论抑制细胞中β-catenin的高表达并调控下游的一系列因子,从而影响Wnt/β-catenin通路诱导的上皮间质转化(EMT)现象可能是臭牡丹总黄酮抑制肺癌侵袭、转移的重要作用机制之一。