Mo Miao, Zhou Mi, Wang Lu, Qi Lin, Zhou Kehua, Liu Long-Fei, Chen Zhi, Zu Xiong-Bing
Department of Urology, Xiangya Hospital, Central South University, Changsha 410008, Hunan, P.R. China.
Department of Urology, Zhejiang Provincial People's Hospital, Hangzhou, Zhejiang 310014, P.R. China.
PLoS One. 2015 Mar 23;10(3):e0119506. doi: 10.1371/journal.pone.0119506. eCollection 2015.
To investigate the effects of CCL21/CCR7 on the proliferation, migration, and invasion of T24 cells and the possible associated mechanisms: expression of MMP-2 and MMP-9, and regulation of BCL-2 and BAX proteins.
T24 cells received corresponding treatments including vehicle control, antibody (20 ng/mL CCR7 antibody and 50 ng/ml CCL21), and 50, 100, and 200 ng/ml CCL21. Proliferation was evaluated by MTT assay; cell migration and invasion were assayed using a transwell chamber. Cell apoptosis was induced by Adriamycin (ADM). The rate of cell apoptosis was examined by flow cytometry using annexin V-FITC/PI staining. Western-blot was used to analyze MMP-2 and MMP-9 and BCL-2 and BAX proteins.
CCL21 promoted T24 cell proliferation in concentration-dependent manner with that 200 ng/mL induced the largest amount of proliferation. Significant differences of cell migration were found between CCL21treatment groups and the control group in both the migration and invasion studies (P < 0.001 for all). The expressions of MMP-2 and MMP-9 proteins were significantly increased after CCL21 treatment (p < 0.05 for all). Protein expression of Bcl-21 follows an ascending trend while the expression of Bax follows a descending trend as the concentration of CCL21 increases. No difference was found between the control group and antibody group for all assessments.
CCL21/CCR7 promoted T24 cell proliferation and enhanced its migration and invasion via the increased expression of MMP-2 and MMP-9. CCL21/CCR7 had antiapoptotic activities on T24 cells via regulation of Bcl-2 and Bax proteins. CCL21/CCR7 may promote bladder cancer development and metastasis.
探讨CCL21/CCR7对T24细胞增殖、迁移和侵袭的影响以及可能的相关机制：基质金属蛋白酶-2（MMP-2）和基质金属蛋白酶-9（MMP-9）的表达，以及B细胞淋巴瘤-2（BCL-2）和Bax蛋白的调控。
T24细胞接受相应处理，包括溶剂对照、抗体（20 ng/mL CCR7抗体和50 ng/ml CCL21），以及50、100和200 ng/ml CCL21。通过MTT法评估增殖；使用Transwell小室检测细胞迁移和侵袭。用阿霉素（ADM）诱导细胞凋亡。采用膜联蛋白V-异硫氰酸荧光素/碘化丙啶（annexin V-FITC/PI）染色，通过流式细胞术检测细胞凋亡率。用蛋白质免疫印迹法分析MMP-2和MMP-9以及BCL-2和Bax蛋白。
CCL21以浓度依赖性方式促进T24细胞增殖，200 ng/mL诱导的增殖量最大。在迁移和侵袭研究中，CCL21处理组与对照组之间的细胞迁移存在显著差异（所有P<0.001）。CCL21处理后，MMP-2和MMP-9蛋白的表达显著增加（所有p<0.05）。随着CCL21浓度增加，Bcl-21蛋白表达呈上升趋势，而Bax表达呈下降趋势。在所有评估中，对照组和抗体组之间未发现差异。
CCL21/CCR7通过增加MMP-2和MMP-9的表达促进T24细胞增殖并增强其迁移和侵袭。CCL21/CCR7通过调控Bcl-2和Bax蛋白对T24细胞具有抗凋亡活性。CCL21/CCR7可能促进膀胱癌的发展和转移。
