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MiR-1307-5p targeting TRAF3 upregulates the MAPK/NF-?B pathway and promotes lung adenocarcinoma proliferation

ABSTRACT: Background Non-small cell lung cancer (NSCLC) includes lung adenocarcinoma (LUAD) and lung squamous cell carcinoma (LUSC). MicroRNA (miRNA) plays an important role in the regulation of post-transcriptional gene expression in animals and plants, especially in lung adenocarcinoma. Methods MiR-1307-5p is an miRNA with significant differences screened by the second generation of high-throughput sequencing in the early stage of our research group. In the current study, a series of in vitro and in vivo experiments were carried out. MiR-1307-5p mimic, miR-1307-5p inhibitor, and NC were transfected into A549 and H1299 lung adenocarcinoma cells. The correlation between miR-1307-5p and clinicopathological features in pathological samples was analyzed using a lung adenocarcinoma tissue microarray, and miR-1307-5p expression was detected by qPCR. CCK-8, EdU, colony formation, scratch test, and Transwell assays were used to observe cell proliferation and migration. Double luciferase assay, western blot, qPCR, and immunohistochemistry were employed in confirming the target relationship between miR-1307-5p and TRAF3. Western blotting was used to analyze the relationship between miR-1307-5p and the NF-?B/MAPK pathway. Finally, the effect of miR-1307-5p on tumor growth was studied using a subcutaneous tumorigenesis model in nude mice. Results Increased miR-1307-5p expression was significantly related to decreased overall survival rate of lung adenocarcinoma patients, revealing miR-1307-5p as a potential oncogene in lung adenocarcinoma. MiR-1307-5p mimic significantly promoted while miR-1307-5p inhibitor reduced the growth and proliferation of A549 and H1299 cells. MiR-1307-5p overexpression significantly enhanced the migration ability while miR-1307-5p inhibition reduced the migration ability of A549 and H1299 cells. Target binding of miR-1307-5p to TRAF3 was confirmed by double luciferase assay, western blot, qPCR, and immunohistochemistry. miR-1307-5p caused degradation of TRAF3 mRNA and protein. MiR-1307-5p targeted TRAF3 and activated the NF-?B/MAPK pathway. TRAF3 colocalized with p65 and the localization of TRAF3 and p65 changed in each treatment group. Tumor volume of the lv-miR-1307-5p group was significantly larger than that of the lv-NC group, and that of the lv-miR-1307-5p-inhibitor group was significantly smaller than that of the lv-NC group. Conclusion In conclusion, miR-1307-5p targets TRAF3 and activates the NF-?B/MAPK pathway to promote proliferation in lung adenocarcinoma.

SUBMITTER: Du X

PROVIDER: S-EPMC7552495 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Project description:ObjectiveTo explore the effect of miR-1307-5p which specifically inhibits transforming growth factor beta-induced gene (TGFBI) on the biologic behavior of osteoarthritis (OA) chondrocytes.MethodsWe detected miR-1307-5p and TGFBI expression in the cartilage tissue specimens of OA patients and mice, respectively. RNA22 was applied to predict the target gene of miR-1307-5p, and we further verified the relationship by performing a dual luciferase reporter experiment. Enzyme-linked immunosorbent assay was used to measure the expression of matrix metalloproteinase inhibitor-1 (TIMP-1), interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in the culture medium of mouse chondrocytes. Quantitative reverse transcription-polymerase chain reaction and western blot were used to measure the expression of Bax and Bcl-2. MTT method was applied to detect the proliferation activity of chondrocytes, while flow cytometry was implemented to detect the apoptosis of chondrocytes.ResultsThe expression of miR-1307-5p in cartilage tissue specimens of OA patients was up-regulated, while TGFBI expression was down-regulated. Compared with normal mice cartilage tissue specimens, the expression of miR-1307-5p in cartilage tissue specimens of OA mouse was increased, while TGFBI expression was decreased (both P<0.05). The results of the dual luciferase reporter experiment showed that TGFBI was a target gene of miR-1307-5p. In cell experiments, compared with the normal group, TIMP-1 and Bcl-2 expression, and cell proliferation activities in all model groups were decreased. IL-1β, IL-6, TNF-α, Bax expression, and cell apoptosis rates were increased (all P<0.05). Compared with the blank group, TIMP-1 and Bcl-2 expression, and cell proliferation activities in the miR-1307-5p inhibitor group and the TGFBI group were increased, while IL-1β, IL-6, TNF-α, and Bax expression, and cell apoptosis rates were decreased (all P<0.05). The changes in all indicators in the miR-1307-5p mimic group were opposite to those of the miR-1307-5p inhibitor group (all P<0.05). There were no significant differences concerning all indicators between the blank group and the NC group, and between the blank group and the miR-1307-5p mimic + TGFBI group (all P>0.05).ConclusionThe suppression of miR-1307-5p expression can increase TGFBI expression, promoting the proliferation of chondrocytes in OA mice, while inhibiting their apoptosis.

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MiR-1307-5p targeting TRAF3 upregulates the MAPK/NF-?B pathway and promotes lung adenocarcinoma proliferation

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