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Berberine Inhibits the Expression of SCT through miR-214-3p Stimulation in Breast Cancer Cells.

ABSTRACT: In this study, we aimed to evaluate the suppressive abilities of berberine (BBR) on MCF-7 and MDA-MB-231 cells and confirm its underlying mechanisms on miR-214-3p. We first built a panel of 18 miRNAs and 9 lncRNAs that were reported to participate in the mechanism of breast cancer. The RT-qPCR results suggested that BBR illustrated a dosage-dependent pattern in the stimulation to miR-214-3p in both MCF-7 and MDA-MB-231 cells. Then, we performed gain-and-lose function tests to validate the role of miR-214-3p contributing to the anticancer effects of BBR. Both BBR and miR-214-3p mimic reduced the cell viability, repressed migration and invasion capacities, increased rates of total apoptotic cells and ratio of Bax/Bcl-2, and increased the percentage of G2/M cells of MCF-7 and MDA-MB-231?cells by colony formation and CKK8 assay, scratch wound healing and gelatin-based 3D conformation assay, transwell invasion assay, and cell cycle analysis, respectively. However, miR-214-3p inhibitor counteracted all these effects of BBR. Based on the bioinformatics analysis and dual-luciferase reporter test, we identified binding sites between SCT and miR-214-3p. We further confirmed that BBR massively and dose-dependently reduced the mRNA expression and protein levels of SCT in both MCF-7 and MDA-231 cells. We testified that both miR-214-3p mimic and BBR could decrease the mRNA expression and protein levels of SCT, while miR-214-3p inhibitor weakened these reductions. In conclusion, BBR suppressed MCF-7 and MDA-MB-231 breast cancer cells by upregulating miR-214-3p and increasing its inhibition to SCT.

SUBMITTER: Zhu C

PROVIDER: S-EPMC7719527 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Berberine Inhibits the Expression of SCT through miR-214-3p Stimulation in Breast Cancer Cells.

Zhu Congyuan C Li Jianping J Hua Yuming Y Wang Jingli J Wang Ke K Sun Jingqiu J

Evidence-based complementary and alternative medicine : eCAM 20201129

In this study, we aimed to evaluate the suppressive abilities of berberine (BBR) on MCF-7 and MDA-MB-231 cells and confirm its underlying mechanisms on miR-214-3p. We first built a panel of 18 miRNAs and 9 lncRNAs that were reported to participate in the mechanism of breast cancer. The RT-qPCR results suggested that BBR illustrated a dosage-dependent pattern in the stimulation to miR-214-3p in both MCF-7 and MDA-MB-231 cells. Then, we performed gain-and-lose function tests to validate the role o ...[more]

PMID: 33312221

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Project description:BackgroundCervical cancer (CC) is one of the most common gynaecological malignancies all around the world. The mechanisms of cervical carcinoma formation remain under close scrutiny. The long non-coding RNAs (lncRNA) and microRNAs (miRNAs) play important roles in controlling gene expression and promoting the development and progression of cervical cancer by acting as competitive endogenous RNA (ceRNA). However, the roles of lncRNA associated with ceRNAs in cervical carcinogenesis remains unknown. In this study, the expression of long non-coding RNA HOTAIR was investigated in HPV16 positive cervical cancer cells, the candidate miRNAs and target genes were identified to clarify putative ceRNAs of HOTAIR/miRNA in cervical cancer cells.MethodsThe proliferation ability of cells was measured by CCK8 and EdU incorporation assays and cell apoptosis was analyzed by flow cytometry. The expression of HOTAIR, miR-214-3p, HPV16 E7 mRNA were detected by qRT-PCR. As for searching for the interaction between miR-214-3p and HOTAIR, the binding sites for miR-214-3p on HOTAIR was predicted by starbase v2.0 database, then dual-luciferase assay was used to verify the binding sites. In addition, Gene Ontology (GO) and protein-protein interaction (PPI) network analysis of target genes of miR-214-3p were performed with bioinformatics analysis. The potential signal pathway regulated by HOTAIR/miR-214-3p was predicted by KEGG enrichment analysis and confirmed by qPCR and WB analysis in cervical cancer cells.ResultsOur results showed that expression of HOTAIR was up-regulated, while that of miR-214-3p was down-regulated in HPV16-positive cervical cancer cells. The expression status of HPV16 E7 played an important role in regulating expression of HOTAIR or miR-214-3p in cervical cancer cells. HOTAIR knockdown could significantly inhibited cell proliferate ability and promote cellular apoptosis, whereas the inhibition of miR-214-3p expression partially reversed such results. Bioinformatics analysis identified 1451 genes as target genes of miR-214-3p. The Gene ontology (GO) and KEGG Pathway enrichment analysis showed that these target genes were mainly related to regulation of cell communication, protein binding, enzyme binding and transferase activity, and Wnt ligand biogenesis. Pathway enrichment analysis results showed that the predicted target genes were significantly enriched in Wnt/β-catenin signaling pathway. Finally, our results confirmed that miR-214-3p could significantly inhibit β-catenin expression in HPV16 positive cancer cells by qPCR and WB analysis.ConclusionHOTAIR could act as a ceRNA through binding to miR-214-3p, promote cell proliferation and inhibit the apoptosis of HPV16 positive cervical cancer. HOTAIR/miR-214-3p/Wnt/β-catenin signal pathway might played important regulated roles in HPV16 positive cervical cancer. Our results provided new insight into defining novel biomarkers for cervical cancer.

Project description:BACKGROUND:Breast cancer (BC) is a common malignant tumor with poor prognosis. Angiogenesis is related to the growth and progression of solid tumors and associated with prognosis. ZLM-7, SP1, VEGFA and miR-212-3p were associated with BC angiogenesis and proliferation, however the detailed mechanism was not clear. This study aimed to reveal the regulatory mechanism of angiogenesis of BC. METHODS:BC cell lines were treated with 10 nM ZLM-7 for 8 h. We detected protein expression level by western blot and RNA expression level by qRT-PCR. Overexpression or inhibition of miR-212-3p is performed using miR-212-3p mimics or miR-212-3p inhibitor, Sp1 overexpression using pcDNA3.1 vector. Angiogenesis was analyzed by co-culturing BC cell lines and HUVEC cells. To evaluate regulatory relationship between miR-212-3p and Sp1, dual luciferase assay was performed. Besides, the direct interaction between Sp1 and VEGFA was analyzed by ChIP. Migration and invasion were analyzed by transwell assay and proliferation was detected by clone formation assay. In mice xenograft model developed using BC cells, we also detected angiogenesis marker CD31 through immunohistochemistry. RESULTS:ZLM-7 up-regulated miR-212-3p and inhibited invasion, migration, proliferation and angiogenesis of BC, while miR-212-3p inhibitor antagonized such effects. Binding sequence was revealed between miR-212-3p and Sp1, and expression of Sp1 was inhibited by miR-212-3p on both protein and mRNA level. Sp1 could interact with VEGFA and promoted its expression. Overexpression of miR-212-3p inhibited migration, invasion, proliferation and angiogenesis of BC cell lines, while Sp1 overexpression showed the opposite effect and could antagonize these effects of miR-212-3p overexpression. ZLM-7 decreased VEGFA expression, which was rescued by co-transfection with miR-212-3p inhibitor. Similar, ZLM-7 could inhibit tumor growth and angiogenesis through the miR-212-3p/Sp1/VEGFA axis in vivo. CONCLUSIONS:ZLM-7 could directly up-regulate miR-212-3p in BC. MiR-212-3p could inhibit VEGFA expression through Sp1, thereby inhibiting angiogenesis and progression of BC.

Dataset Information

Berberine Inhibits the Expression of SCT through miR-214-3p Stimulation in Breast Cancer Cells.

Publications

Berberine Inhibits the Expression of SCT through miR-214-3p Stimulation in Breast Cancer Cells.

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