| 任慧云,冯一伟,许培英.乌头碱调节 C-C基序趋化因子配体 2/C-C基序趋化因子受体 2信号通路对膀胱癌细胞抗肿瘤活性及其作用机制研究[J].安徽医药,2024,28(4):660-665. |
| 乌头碱调节 C-C基序趋化因子配体 2/C-C基序趋化因子受体 2信号通路对膀胱癌细胞抗肿瘤活性及其作用机制研究 |
| The anti-tumor activity and mechanism of aconitine on bladder cancer cells by regulating C-C-motif chemokine ligand 2/C-C-motif chemokine receptor 2 signaling pathway |
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| DOI：10.3969/j.issn.1009-6469.2024.04.005 |
| 中文关键词: 乌头碱 C-C基序趋化因子配体 2/C-C基序趋化因子受体 2 信号通路 膀胱癌 抗肿瘤 表达、，细胞，活、， |
| 英文关键词: Aconitine CCL2/CCR2 Signaling pathway Bladder cancer Anti-tumor |
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| 中文摘要: |
| 目的探讨乌头碱调节 C-C基序趋化因子配体 2(CCL2)/C-C基序趋化因子受体 2(CCR2)信号通路对膀胱癌细胞抗肿瘤活性及其作用机制研究。方法研究起止时间为 2021年 12月至 2022年 10月。采用细胞计数试剂盒( CCK-8)法检测 2.5、5.0、10.0、20.0、30.0 μmol/L乌头碱处理后的人膀胱癌 5637细胞存活率，筛选出合适的乌头碱作用浓度。将 5637细胞分为对照组、乌头碱低剂量( 10 μmol/L)组、乌头碱高剂量( 20 μmol/L)组、乌头碱高剂量(20 μmol/L)+空载组、乌头碱高剂量( 20 μmol/L) +CCL2过表达组，分组处理后，采用 CCK-8法、 5-乙炔基 -2'-脱氧尿苷( Edu)染色法及 Hoechst 33258染色分别检测各组 5637细胞存活率、增殖率、凋亡率；采用 Transwell实验及划痕实验分别检测各组 5637细胞侵袭数、迁移率；采用免疫荧光染色检测各组 5637细胞凋亡相关蛋白 BCL2相关 X蛋白( Bax)和 B淋巴细胞瘤 -2(Bcl-2)表达比值( Bax/Bcl-2)；采用免疫印迹法检测各组 5637细胞 CCL2/CCR2信号通路相关蛋白及上皮间质转化标志蛋白［神经钙黏素( N-cadherin)、锌指 E盒结合同源盒 1(ZEB1)、紧密连接蛋白 1(ZO-1)、上皮钙黏素( E-cadherin)］表达。结果与对照组相比，乌头碱低剂量组、乌头碱高剂量组、乌头碱高剂量 +空载组细胞 CCL2(0.69±0.09、0.20±0.03、0.19±0.04比 1.21±0.13)CCR2(0.78±0.12、0.26±0.06、0.27±0.07比 1.33±0.20)、 Ncadherin(0.65±0.06、0.12±0.02、0.11±0.03比 1.24±0.12)与 ZEB1蛋白存活率、增殖率、侵袭数、迁移率均降低( P<0.05)， Bax/Bcl-2、细胞 ZO-1与 E-cadherin蛋白表达均升高( P<0.05)；乌头碱高剂量组、乌头碱高剂量 +空载组细胞 CCL2、CCR2、N-cad? herin与 ZEB1蛋白表达、存活率、增殖率、侵袭数、迁移率相比乌头碱低剂量组进一步降低( P<0.05)Bax/Bcl-2、细胞 ZO-1与 Ecadherin蛋白表达进一步升高( P<0.05)。与乌头碱高剂量组相比，乌头碱高剂量 +CCL2过表达组CCL2、CCR2、N-cadherin与 ZEB1蛋白表达、存活率、增殖率、侵袭数、迁移率升高( P<0.05)Bax/Bcl-2、细胞 ZO-1与 E-cadherin蛋白表达降低( P<0.05)。结论乌头碱可通过 CCL2/CCR2信号通路而抑制膀胱癌细胞存增殖及侵袭和迁移，促使其凋亡。 |
| 英文摘要: |
| Objective To investigate the anti-tumor activity and mechanism of aconitine on bladder cancer cells by regulating the C-Cmotif chemokine ligand 2 (CCL2)/C-C-motif chemokine receptor 2 (CCR2) signaling pathway.Methods The start and end period of this study was from December 2021 to October 2022. CCK-8 method was used to detect the survival rate of human bladder cancer 5637 cells treated with 2.5, 5, 10, 20, 30 μmol/L aconitine, and the appropriate concentration of aconitine was selected. 5637 cells were randomlyassigned into control group, low-dose aconitine (10 μmol/L) group, high-dose aconitine (20 μmol/L) group, high-dose aconitine (20 μmol/ L)+no load group, high-dose aconitine (20 μmol/L)+CCL2 overexpression group. After grouping and treatment, the survival rate, prolifer?ation rate and apoptosis rate of 5637 cells in each group were detected by CCK-8 method, 5-Ethynyl-2'-deoxyuridine (Edu) stainingmethod and Hoechst 33258 staining. Transwell test and scratch test were used to detect the invasion number and migration rate of 5637cells in each group; immunofluorescence staining was used to detect the expression ratio of apoptosis related protein BCL2-associated X protein (Bax) and B-cell lymphoma-2 (Bcl-2) (Bax/Bcl-2) of 5637 cells in each group; Western blotting was performed to detect the ex?pression of CCL2/CCR2 signaling pathway related proteins and epithelial mesenchymal transformation markers [N-cadherin (N-cad? herin), zinc finger e-box-binding homeobox 1 (ZEB1), zonula occludens-1 (ZO-1), and E-cadherin (E-cadherin)] in 5637 cells of each group. Results Compared with the control group, the expressions of CCL2 (0.69±0.09, 0.20±0.03, 0.19±0.04 vs. 1.21±0.13), CCR2 (0.78±0.12, 0.26±0.06, 0.27±0.07 vs. 1.33±0.20), N-cadherin (0.65±0.06, 0.12±0.02, 0.11±0.03 vs. 1.24±0.12) and ZEB1 proteins, sur?vival rates, proliferation rates, invasion numbers and migration rates of cells in low-dose aconitine group, high-dose aconitine group and high-dose aconitine+no load group decreased (P<0.05), while the expressions of Bax/Bcl-2, ZO-1 and E-cadherin proteins in cells in? creased (P<0.05). Compared with the low-dose aconitine group, the expressions of CCL2, CCR2, N-cadherin and ZEB1 proteins, survival rates, proliferation rates, invasion numbers and migration rates of cells in the high-dose aconitine group, the high-dose aconitine+no load group further reduced (P<0.05), while the expressions of Bax/Bcl-2, ZO-1 and E-cadherin proteins in cells further increased (P<0.05). Compared with the high-dose aconitine group, the expressions of CCL2, CCR2, N-cadherin and ZEB1 proteins, survival rates, prolifera? tion rates, invasion numbers and migration rates of cells in the high-dose aconitine+CCL2 overexpression group increased (P<0.05), while the expressions of Bax/Bcl-2, ZO-1 and E-cadherin proteins decreased (P<0.05).Conclusion Aconitine can inhibit the survival,proliferation, invasion and migration of bladder cancer cells and promote their apoptosis through CCL2/CCR2 signaling pathway. |
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