miR-362-3p 的表达及其靶基因与胆囊癌恶性特征的关系
| 作者: |
1掲伟,
1林江飞,
1肖强胜,
1聂晚频
1 中南大学湘雅三医院 普通外科，湖南 长沙 410006 |
| 通讯: |
聂晚频
Email: niewanpin@163.com |
| DOI: | 10.3978/.2018.08.008 |
| 基金: | 中南大学湘雅三医院科学研究资助项目（2018-S351）。 |
摘要
目的：探讨 miR-362-3p 在胆囊癌中的表达及功能。
方法：用 qRT-PCR 检测手术切除的 44 例胆囊癌患者手术标本中 miR-362-3p 的表达，并分析其表达与胆囊癌临床病理特征及预后的关系。miR-362-3p 模拟物转染胆囊癌细胞后，分别用 MTT 法、细胞划痕试验及 Transwell 侵袭试验观察细胞增殖、迁移及侵袭的改变。通过生物信息学方法及双荧光素酶报告基因试验分析 miR-362-3p 的靶基因，并采用补救试验验证。
结果：miR-362-3p 在胆囊癌组织的表达明显低于相应癌旁组织（P<0.05）。miR-362-3p 的低表达与肿瘤 TNM 分期、淋巴结转移及远处转移明显有关（均 P<0.05）。低表达 miR-362-3p 患者总体生存率较高表达 miR-362-3p 患者明显降低（P<0.05）。转染 miR-362-3p 模拟物后，胆囊癌细胞的增殖，迁移及侵袭能力明显减弱（均 P<0.05）。Nemo 样激酶（NLK）被确定为 miR-362-3p 的潜在靶基因，转染NLK 过表达载体后，miR-362-3p 模拟物对胆囊癌细胞的上述作用被明显逆转（均 P<0.05）。
结论：miR-362-3p 在胆囊癌中表达下调，下调的 miR-362-3p 减少了对靶基因 NLK 的抑制，从而促进了胆囊癌细胞的增殖、迁移和侵袭。
关键词：
胆囊肿瘤；微 RNAs；蛋白激酶类；细胞增殖；肿瘤侵润
方法：用 qRT-PCR 检测手术切除的 44 例胆囊癌患者手术标本中 miR-362-3p 的表达，并分析其表达与胆囊癌临床病理特征及预后的关系。miR-362-3p 模拟物转染胆囊癌细胞后，分别用 MTT 法、细胞划痕试验及 Transwell 侵袭试验观察细胞增殖、迁移及侵袭的改变。通过生物信息学方法及双荧光素酶报告基因试验分析 miR-362-3p 的靶基因，并采用补救试验验证。
结果：miR-362-3p 在胆囊癌组织的表达明显低于相应癌旁组织（P<0.05）。miR-362-3p 的低表达与肿瘤 TNM 分期、淋巴结转移及远处转移明显有关（均 P<0.05）。低表达 miR-362-3p 患者总体生存率较高表达 miR-362-3p 患者明显降低（P<0.05）。转染 miR-362-3p 模拟物后，胆囊癌细胞的增殖，迁移及侵袭能力明显减弱（均 P<0.05）。Nemo 样激酶（NLK）被确定为 miR-362-3p 的潜在靶基因，转染NLK 过表达载体后，miR-362-3p 模拟物对胆囊癌细胞的上述作用被明显逆转（均 P<0.05）。
结论：miR-362-3p 在胆囊癌中表达下调，下调的 miR-362-3p 减少了对靶基因 NLK 的抑制，从而促进了胆囊癌细胞的增殖、迁移和侵袭。
Relations of miR-362-3p expression and its target gene with malignant profiles of gallbladder cancer
CorrespondingAuthor:NIE Wanpin Email: niewanpin@163.com
Abstract
Objective: To detect the expression of miR-362-3p in gallbladder cancer and its functions.
Methods: The expressions of miR-362-3p in the surgical specimens from 44 gallbladder cancer patients were determined by qRT-PCR, and the relations of miR-362-3p expression with clinicopathological characteristics and prognosis of the patients were analyzed. In gallbladder cancer cells aft er transfection with miR-362-3p mimics, the changes in proliferation, migration and invasion were examined by MTT colorimetry, wound healing assay, and Transwell assay respectively. Th e target gene of miR-362-3p was analyzed by bioinformatics analysis and dual luciferase reporter gene assay, and then was verifi ed through recovery tests.
Results: Th e expression of miR-362-3p was signifi cantly decreased in gallbladder cancer tissue compared with tumor adjacent tissue (P<0.05). The decreased miR-362-3p expression was significantly associated with tumor TNM stage, lymph node metastasis and distant metastasis (all P<0.05). The overall survival rate in patients with low miR-362-3p expression was significantly lower than that in patients with high miR-362-3p expression (P<0.05). The abilities of proliferation, migration and invasion were significantly reduced in gallbladder cancer cells after transfection with miR-362-3p mimics (all P<0.05). Nemo like kinase (NLK) was found potentially to be the target gene of miR-362-3p. After transfection with NLK overexpression vectors, the above effects exerted by miR-362-3p mimics in gallbladder cancer cells were significantly reversed (all P<0.05).
Conclusion: The expression of miR-362-3p is down-regulated in gallbladder cancer, and the down-regulated miR-362-3p expression may reduce the inhibition on its target gene NLK, and thereby promote the proliferation, migration and invasion of gallbladder cancer cells.
Keywords：
Gallbladder Neoplasms; MicroRNAs; Protein Kinases; Cell Proliferation; Neoplasm Invasiveness
Methods: The expressions of miR-362-3p in the surgical specimens from 44 gallbladder cancer patients were determined by qRT-PCR, and the relations of miR-362-3p expression with clinicopathological characteristics and prognosis of the patients were analyzed. In gallbladder cancer cells aft er transfection with miR-362-3p mimics, the changes in proliferation, migration and invasion were examined by MTT colorimetry, wound healing assay, and Transwell assay respectively. Th e target gene of miR-362-3p was analyzed by bioinformatics analysis and dual luciferase reporter gene assay, and then was verifi ed through recovery tests.
Results: Th e expression of miR-362-3p was signifi cantly decreased in gallbladder cancer tissue compared with tumor adjacent tissue (P<0.05). The decreased miR-362-3p expression was significantly associated with tumor TNM stage, lymph node metastasis and distant metastasis (all P<0.05). The overall survival rate in patients with low miR-362-3p expression was significantly lower than that in patients with high miR-362-3p expression (P<0.05). The abilities of proliferation, migration and invasion were significantly reduced in gallbladder cancer cells after transfection with miR-362-3p mimics (all P<0.05). Nemo like kinase (NLK) was found potentially to be the target gene of miR-362-3p. After transfection with NLK overexpression vectors, the above effects exerted by miR-362-3p mimics in gallbladder cancer cells were significantly reversed (all P<0.05).
Conclusion: The expression of miR-362-3p is down-regulated in gallbladder cancer, and the down-regulated miR-362-3p expression may reduce the inhibition on its target gene NLK, and thereby promote the proliferation, migration and invasion of gallbladder cancer cells.