Wang Gui-hua, Lu Mei-hong, Wang Jing-chun, Wang Feng, Ding Wei-feng, Wang Yue-guo, Ju Shao-qing, Wang Hui-min
Laboratory Medicine Center, Affiliated Hospital of Nantong University, Nantong 226001, China.
Zhonghua Zhong Liu Za Zhi. 2013 Apr;35(4):249-55. doi: 10.3760/cma.j.issn.0253-3766.2013.04.003.
To investigate the effects of a proliferation-inducing ligand (APRlL) on colorectal cancer (CRC) cell growth and migration, and to observe the role of APRIL in CRC biological behavior.
The siRNA plasmid vector targeting APRIL gene (APRIL-siRNA) was transfected into human colorectal cancer SW480 cells and recombinant human APRIL (rhAPRIL) was used to stimulate human colorectal cancer HCT-116 cells. Cell proliferation activity was analyzed using cell counting kit-8 (CCK-8), cell cycle was detected by flow cytometry, and the protein expression of cyclin D1, p21 and Bcl-2 was detected by Western blot analysis. Tumor cell migration and invasion were measured by Transwell chambers. RT-PCR was applied to examine the mRNA expression level of MMP-2 and MMP-9. APRIL-siRNA was used to transfect directly SW480 cells, which were injected subcutaneously into nude mice, then the tumor growth and metastasis were observed.
Cell proliferation ability of APRIL-siRNA-transfected SW480 cells was drastically repressed, and the percentage of G0/G1 phase cells was significantly increased (t = 4.12, P < 0.05), accompanied with depressed cyclin D1, Bcl-2 expression and elevated p21 expression. Cell proliferation ability of rhAPRIL-stimulated HCT-116 cells was promoted with a decreased G0/G1 phase ratio (t = 3.31, P < 0.05). cyclin D1 and Bcl-2 protein expression was up-regulated while p21 was down-regulated by rhAPRIL stimulation. Metastatic and invasive capacities of APRIL-siRNA-transfected SW480 cells were significantly inhibited compared with their respective controls (both P < 0.05), accompanied with the deregulated MMP-2 and MMP-9 mRNA expression. Metastatic and invasive capacities of rhAPRIL-stimulated HCT-116 cells were promoted with up-regulated MMP-2 and MMP-9 mRNA expression(both P < 0.05). Tumor growth in the group transfected with APRIL-siRNA appeared to be slower than that in the control groups and the expression of MMP-2, MMP-9 in tumor tissues was depressed in the APRIL-siRNA group.
APRIL facilitates tumor growth and metastasis, and is associated with carcinogenesis and prognosis. Our findings suggest that APRIL might be used as a novel target for the intervention and therapy of colorectal cancer.
探讨增殖诱导配体（APRIL）对结直肠癌（CRC）细胞生长和迁移的影响，观察APRIL在CRC生物学行为中的作用。
将靶向APRIL基因的小干扰RNA质粒载体（APRIL-siRNA）转染至人结直肠癌SW480细胞，并用重组人APRIL（rhAPRIL）刺激人结直肠癌HCT-116细胞。采用细胞计数试剂盒-8（CCK-8）分析细胞增殖活性，通过流式细胞术检测细胞周期，采用蛋白质免疫印迹法检测细胞周期蛋白D1、p21和Bcl-2的蛋白表达。用Transwell小室检测肿瘤细胞的迁移和侵袭能力。应用逆转录-聚合酶链反应（RT-PCR）检测基质金属蛋白酶-2（MMP-2）和基质金属蛋白酶-9（MMP-9）的mRNA表达水平。将APRIL-siRNA直接转染SW480细胞后皮下注射到裸鼠体内，观察肿瘤生长和转移情况。
转染APRIL-siRNA的SW480细胞增殖能力明显受到抑制，G0/G1期细胞百分比显著增加（t = 4.12，P < 0.05），同时细胞周期蛋白D1、Bcl-2表达降低，p21表达升高。rhAPRIL刺激的HCT-116细胞增殖能力增强，G0/G1期比例降低（t = 3.31，P < 0.05）。rhAPRIL刺激使细胞周期蛋白D1和Bcl-2蛋白表达上调，p21表达下调。与各自对照组相比，转染APRIL-siRNA的SW480细胞的转移和侵袭能力明显受到抑制（均P < 0.05），同时MMP-2和MMP-9 mRNA表达失调。rhAPRIL刺激的HCT-116细胞的转移和侵袭能力增强，MMP-2和MMP-9 mRNA表达上调（均P < 0.05）。转染APRIL-siRNA组的肿瘤生长似乎比对照组慢，且APRIL-siRNA组肿瘤组织中MMP-2、MMP-9的表达降低。
APRIL促进肿瘤生长和转移，与肿瘤发生及预后相关。我们的研究结果提示，APRIL可能作为结直肠癌干预和治疗的新靶点。
