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抑制miR-20a-5p通过GJA1抑制结肠癌细胞的上皮-间质转化。

Inhibition of miR-20a-5p Suppresses Epithelial-Mesenchymal Transition of Colorectal Cancer Cells Through GJA1.

作者信息

Zhang Lu, Wang Jun-Bin, Gao Zhen-Yuan, Wu Xiao, Zhou Hai-Rong

机构信息

Department of Medical Oncology, The First Affiliated Hospital of Bengbu Medical College, Bengbu, 233000, China.

出版信息

Mol Biotechnol. 2024 Nov 22. doi: 10.1007/s12033-024-01315-2.

Abstract

This study was designed to clarify the role of GJA1 in colorectal cancer. qPCR was adopted to detect the GJA1 and miR-20a-5p expression levels in tumor tissues and cells; and EdU, Transwell assay, Scratch test to examine the migration, invasion, and proliferation of colorectal cancer cells. The EMT-related protein expression was measured by immunofluorescence and western Blot. The binding relationship between GJA1 and miR-20a-5p was examined using dual luciferase reporting subsystem. In situ hybridization was utilized to examine the miR-20a-5p expression in tumor tissues and metastases. Rescue experiments were performed by simultaneous transfection of sh-GJA1 inhibitor and miR-20a-5p inhibitor. The miR-20a-5p expression was high and the GJA1 expression was low in colorectal cancer tissues and cells. A targeting relationship was found in GJA1 and miR-20a-5p targets. The invasion, migration, and proliferation of colorectal cancer cells can be inhibited by overexpression of GJA1. Meanwhile, overexpression of GJA1 markedly elevated the e-cadherin expression, but reduced the levels of vimentin, α-SMA and n-cadherin expression. miR-20a-5p inhibitor + sh-GJA1 promoted the invasion, migration, and proliferation of colon cancer cells and EMT process. Overall, miR-20a-5p could target GJA1 to down-regulate the GJA1 expression, thereby regulating the EMT response, and ultimately promoting the progression of colorectal cancer.

摘要

本研究旨在阐明GJA1在结直肠癌中的作用。采用qPCR检测肿瘤组织和细胞中GJA1和miR-20a-5p的表达水平;采用EdU、Transwell实验、划痕实验检测结直肠癌细胞的迁移、侵袭和增殖。通过免疫荧光和蛋白质印迹法检测EMT相关蛋白的表达。利用双荧光素酶报告系统检测GJA1与miR-20a-5p之间的结合关系。采用原位杂交检测肿瘤组织和转移灶中miR-20a-5p的表达。通过同时转染sh-GJA1抑制剂和miR-20a-5p抑制剂进行挽救实验。在结直肠癌组织和细胞中,miR-20a-5p表达高而GJA1表达低。发现GJA1与miR-20a-5p靶点之间存在靶向关系。GJA1过表达可抑制结直肠癌细胞的侵袭、迁移和增殖。同时,GJA1过表达显著提高E-钙黏蛋白的表达,但降低波形蛋白、α-平滑肌肌动蛋白和N-钙黏蛋白的表达水平。miR-20a-5p抑制剂+sh-GJA1促进结肠癌细胞的侵袭、迁移和增殖以及EMT进程。总体而言,miR-20a-5p可靶向GJA1下调GJA1表达,从而调节EMT反应,最终促进结直肠癌进展。

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