miR-325-3p Regulates Epithelial-mesenchymal Transition, Invasion and Metastasis of Gastric Cancer via Targeting CLDN1 Gene
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摘要:目的
探讨miR-325-3p靶向CLDN1基因对胃癌上皮间质转化和侵袭转移的影响。
方法选取人胃黏膜上皮细胞株GES-1以及人胃癌细胞株HGC27、SGC-7901、MKN-45和MGC-803，并检测细胞中miR-325-3p和CLDN1的表达。双荧光素酶报告实验验证miR-325-3p和CLDN1的靶向关系，干预胃癌细胞中miR-325-3p和CLDN1的表达，qRT-PCR和Western blot检测细胞中N-cadherin、Vimentin和MMP2的表达，CCK-8检测细胞增殖活力，Transwell和流式细胞仪分别检测细胞侵袭和凋亡能力。
结果相对于GES-1细胞，MGC-803细胞中miR-325-3p表达降低而CLDN1表达增高（均P < 0.05），双荧光素酶报告实验证实CLDN1为miR-325-3p的靶基因。过表达miR-325-3p能够抑制胃癌细胞的增殖、侵袭和上皮间质转化，促进胃癌细胞凋亡，抑制miR-325-3p则能够促进胃癌细胞的增殖、侵袭和上皮间质转化，抑制胃癌细胞凋亡（均P < 0.05）。而过表达CLDN1则能够逆转miR-325-3p过表达对胃癌细胞生物学行为的影响。
结论miR-325-3p能够靶向抑制CLDN1进而抑制胃癌细胞的侵袭转移和上皮间质转化，促进胃癌细胞凋亡，miR-325-3p有望成为治疗胃癌的新靶点。
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关键词:
- 胃癌 /
- CLDN1 /
- miR-325-3p /
- 上皮间质转化 /
- 侵袭转移
Abstract:ObjectiveTo investigate the effects of miR-325-3p on the EMT, invasion and metastasis of gastric cancer cells by targeting CLDN1 gene.
MethodsWe selected human gastric epithelial cell lines GES-1 and gastric cancer cell lines HGC27, SGC-7901, MKN-45 and MGC-803, and detected the expression of miR-325-3p and CLDN1. The targeting relation between miR-325-3p and CLDN1 were verified by dual luciferase report experiments, and the expression of miR-325-3p and CLDN1 in gastric cancer cells were intervened. qRT-PCR and Western blot were adopted to detect N-cadherin, vimentin and MMP2 expression in cells. CCK-8 assay, Transwell assay, flow cytometry were utilized to detect cell proliferation activity, invasion and apoptosis, respectively.
ResultsCompared with GES-1 cells, miR-325-3p expression was decreased while CLDN1 expression was increased in MGC-803 cells (P < 0.05). CLDN1 was a target gene of miR-325-3p. Overexpression of miR-325-3p could inhibit the proliferation, invasion and EMT of gastric cancer cells, while promote the apoptosis of gastric cancer cells. However, the inhibition of miR-325-3p had the opposite effect (P < 0.05). The overexpression of CLDN1 could reverse the effect of miR-325-3p overexpression on the biological behavior of gastric cancer cells.
ConclusionmiR-325-3p can suppress CLDN1, inhibit the invasion, metastasis and EMT while promote the apoptosis of gastric cancer cells. miR-325-3p is expected to be a new target in gastric cancer treatment.
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Key words:
- Gastric cancer /
- CLDN1 /
- miR-325-3p /
- Epithelial-mesenchymal transition /
- Invasion and metastasis
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Competing interests: The authors declare that they have no competing interests.作者贡献：杜记涛：实验方案设计、文章撰写和修改曹建、赵稳：实验实施、数据统计万相斌：文章撰写、数据核查李智：实验方案设计、文章审核和修改
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图 1 不同癌组织和细胞中miR-325-3p和CLDN1的表达
Figure 1 Expression of miR-325-3p and CLDN1 in different cancer tissues and cell lines
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图 2 miR-325-3p与CLDN1的靶向关系验证
Figure 2 Validation of targeting relation between miR-325-3p and CLDN1
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图 3 过表达miR-325-3p后各组细胞凋亡(A, B)和增殖(C)能力检测结果
Figure 3 Cell apoptosis(A, B) and proliferation(C) of each group after miR-325-3p was overexpressed
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图 4 过表达miR-325-3p后各组细胞侵袭能力(A)和EMT相关因子mRNA(B)及蛋白(C)的表达
Figure 4 Cell invasion(A) and EMT-related factors mRNA(B) and protein(C) expression in each group after miR-325-3p was overexpressed
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图 5 过表达CLDN1后各组细胞增殖(A, B)和凋亡(C)能力检测结果
Figure 5 Cell proliferation(A, B) and apoptosis(C) of each group after CLDN1 was overexpressed
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图 6 过表达CLDN1后各组细胞侵袭能力(A)和EMT相关因子mRNA(B)及蛋白(C)的表达
Figure 6 Cell invasion(A) and EMT-related factors mRNA(B) and protein(C) expression in each group after CLDN1 was overexpressed
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