Wei K, Shi J, Xiao Y, Wang W, Yang Q, Chen C
Anhui Provincial Key Laboratory of Cancer Translational Medicine, Bengbu Medical College, Bengbu 233000, China.
Key Laboratory of Cancer Research and Clinical Laboratory Diagnosis, Bengbu Medical College, Bengbu 233000, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2023 Jul 20;43(7):1081-1092. doi: 10.12122/j.issn.1673-4254.2023.07.04.
To investigate the regulatory effects of miR-30e-5p on biological behaviors of colorectal cancer cells and the role of PTEN/CXCL12 axis in mediating these effects.
Bioinformatic analysis was performed to explore the differential expression of miR-30e-5p between colorectal cancer tissues and normal tissues. RT-qPCR was used to detect the differential expression of miR-30e-5p in intestinal epithelial cells and colorectal cancer cells. Bioinformatics and dual luciferase assay were used to predict and validate the targeting relationship between miR-30e-5p and PTEN. Human and murine colorectal cancer cell lines were transfected with miR-30e-5p mimics, miR-30e-5p inhibitor, miR-30e-5p mimics+LV-PTEN, or miR-30e-5p inhibitor + si-PTEN. The changes in biological behaviors of the cells were detected using plate clone formation assay, CCK-8 assay, flow cytometry, scratch healing and Transwell assays. PTEN and CXCL12 expressions in the cancer cells were detected by Western blotting. The effects of miR-30e-5p inhibitor on colorectal carcinogenesis and development were observed in nude mice.
Bioinformatic analysis showed that miR-30e-5p expression was significantly elevated in colorectal cancer tissues compared with the adjacent tissue ( < 0.01). Higher miR-30e-5p expression was detected in colorectal cancer cell lines than in intestinal epithelial cells ( < 0.01). Dual luciferase assay confirmed the targeting relationship between miR-30e-5p and PTEN ( < 0.05). Transfection with miR-30e-5p mimics significantly enhanced proliferation and metastasis and inhibited apoptosis of the colorectal cancer cells ( < 0.05), and co-transfection with LV-PTEN obviously reversed these changes ( < 0.05). MiR-30e-5p mimics significantly inhibited PTEN expression and enhanced CXCL12 expression in the cancer cells ( < 0.01), and miR-30e-5p inhibitor produced the opposite effect. Transfection with miR-30e-5p inhibitor caused cell cycle arrest in the cancer cells, which was reversed by co-transfection with si-PTEN ( < 0.05). In the experiments, the colorectal cancer cells transfected with miR-30e-5p inhibitor showed significantly lowered tumorigenesis.
Overexpression of miR-30e-5p promotes the malignant behaviors of colorectal cancer cells by downregulating PTEN to activate the CXCL12 axis.
探讨miR-30e-5p对大肠癌细胞生物学行为的调控作用及PTEN/CXCL12轴在介导这些作用中的作用。
进行生物信息学分析以探究miR-30e-5p在结直肠癌组织与正常组织中的差异表达。采用RT-qPCR检测miR-30e-5p在肠上皮细胞和大肠癌细胞中的差异表达。利用生物信息学和双荧光素酶报告基因检测法预测并验证miR-30e-5p与PTEN之间的靶向关系。将人源和鼠源大肠癌细胞系分别转染miR-30e-5p模拟物、miR-30e-5p抑制剂、miR-30e-5p模拟物+LV-PTEN或miR-30e-5p抑制剂+si-PTEN。采用平板克隆形成实验、CCK-8实验、流式细胞术、划痕愈合实验和Transwell实验检测细胞生物学行为的变化。通过蛋白质免疫印迹法检测癌细胞中PTEN和CXCL12的表达。在裸鼠中观察miR-30e-5p抑制剂对结直肠癌发生发展的影响。
生物信息学分析显示，与癌旁组织相比，miR-30e-5p在结直肠癌组织中的表达显著升高（<0.01）。在大肠癌细胞系中检测到的miR-30e-5p表达高于肠上皮细胞（<0.01）。双荧光素酶报告基因检测法证实了miR-30e-5p与PTEN之间的靶向关系（<0.05）。转染miR-30e-5p模拟物显著增强了大肠癌细胞的增殖和转移能力，并抑制了其凋亡（<0.05），与LV-PTEN共转染明显逆转了这些变化（<0.05）。miR-30e-5p模拟物显著抑制癌细胞中PTEN的表达并增强CXCL12的表达（<0.01），而miR-30e-5p抑制剂则产生相反的效果。转染miR-30e-5p抑制剂导致癌细胞发生细胞周期阻滞，与si-PTEN共转染可逆转这一现象（<0.05）。在体内实验中，转染miR-30e-5p抑制剂的大肠癌细胞的致瘤性显著降低。
miR-30e-5p的过表达通过下调PTEN激活CXCL12轴，促进大肠癌细胞的恶性行为。
