ABSTRACT
Objective To study the effect of LncRNA CRNDE on radiosensitivity of colorectal cells and underlying mechanism.Methods Colorectal cancer HT-29 cells were transfected with CRNDE shRNA and the interference efficiency was determined by Real time PCR.HT-29 cells transfected with CRNDE shRNA or co-transfected with CRNDE shRNA and miR-384 inhibitor were irradiated at 8 Gy dose,then cell proliferation and apoptosis were detected by MTT and flow cytometry assay,respectively,and cell radiosensitivity was evaluated by cloning assay.It was predicted by a bioinformatics software that CRNDE and miR-384 have complementary binding sites,and this was identified by a luciferase reporting system.Results CRNDE shRNA reduced the expression of CRNDE in HT-29 cells (1.00±0.08 vs.0.42±0.06,t=10.051,P＜0.05).Both CRNDE shRNA and radiation inhibited the proliferation and induced apoptosis of HT-29 cells,and their combination treatment had synergistic effect in apoptosis induction [Apoptosis rates:(2.27±0.13)％,(23.58±2.35)％,(26.91±2.81)％,(36.84±3.24)％,F=24.660,P＜0.05;A values:0.45±0.06,0.30±0.02,0.28±0.03,0.20±0.02,F=106.207,P＜0.05].Transfection of CRNDE shRNA increased the radiosensitivity of HT-29 cells with a radiosensitization ratio of 1.374.CRNDE negatively regulated the expression of its target miR-384.The mniR-384 inhibitor antagonized the effect of CRNDE shRNA on proliferation inhibition and apoptosis promotion of radiationtreated colorectal cancer cells.Conclusions Down-regulation of LncRNA expression enhances the radiosensitivity of colorectal cancer cells by regulating miR-384 expression.
ABSTRACT
Objective@#To study the effect of LncRNA CRNDE on radiosensitivity of colorectal cells and underlying mechanism.@*Methods@#Colorectal cancer HT-29 cells were transfected with CRNDE shRNA and the interference efficiency was determined by Real time PCR. HT-29 cells transfected with CRNDE shRNA or co-transfected with CRNDE shRNA and miR-384 inhibitor were irradiated at 8 Gy dose, then cell proliferation and apoptosis were detected by MTT and flow cytometry assay, respectively, and cell radiosensitivity was evaluated by cloning assay. It was predicted by a bioinformatics software that CRNDE and miR-384 have complementary binding sites, and this was identified by a luciferase reporting system.@*Results@#CRNDE shRNA reduced the expression of CRNDE in HT-29 cells(1.00±0.08 vs. 0.42±0.06, t=10.051, P<0.05). Both CRNDE shRNA and radiation inhibited the proliferation and induced apoptosis of HT-29 cells, and their combination treatment had synergistic effect in apoptosis induction [Apoptosis rates: (2.27±0.13)%, (23.58±2.35)%, (26.91±2.81)%, (36.84±3.24)%, F=24.660, P<0.05; A values: 0.45±0.06, 0.30±0.02, 0.28±0.03, 0.20±0.02, F=106.207, P<0.05]. Transfection of CRNDE shRNA increased the radiosensitivity of HT-29 cells with a radiosensitization ratio of 1.374. CRNDE negatively regulated the expression of its target miR-384. The miR-384 inhibitor antagonized the effect of CRNDE shRNA on proliferation inhibition and apoptosis promotion of radiation-treated colorectal cancer cells.@*Conclusions@#Down-regulation of LncRNA expression enhances the radiosensitivity of colorectal cancer cells by regulating miR-384 expression.
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Objective: To investigate whether microRNA-384-5p can interfere with renal fibrosis by regulating BMP7 transcription in renal tubular epithelial cells. Methods: Unilateral ureteral obstructive (UUO) model was established, and fluorescence activated cell sorter (FACS) was used to isolate renal tubular epithelial cells for culture. Plasmid expressing miR-384-5p, antisense miR-384-5p, and blank plasmid were transfected by liposome and injected in UUO mice. Masson staining was used to detect renal fibrosis, and Western blot and real-time PCR were used to detect the expressions of target-genes of miRNA (miR-384-5p, miR-30, miR-92, and miR-128). Results: There was obvious renal fibrosis at 14 day in UUO model group; the expression of BMP7 mRNA in the renal tissue increased obviously while the expression of BMP7 protein was not increased. Although miR-384-5p could not change BMP7 mRNA in the renal tubular epithelial cells, BMP7 protein in the renal tubular epithelial cells that overexpressed miR-384-5p decreased significantly, and BMP7 protein in these cells overexpressing antisense miR-384-5p increased significantly. Immunohistochemistry and quantitative PCR showed that renal tissue fibrosis in UUO mice increased significantly in 14 days, while antisense miR-384-5p could significantly reduce the renal tissue fibrosis. Although antisense-miR-384-5p treatment did not affect the level of BMP7 mRNA, it could significantly increase the expression of BMP7 protein in renal tissue, suggesting that inhibiting miR-384-5p could lessen renal injury in UUO mice. Conclusion: As a key factor regulating the mechanism of renal fibrosis after renal injury, targeting miR-384-5p may become a new method to prevent and treat renal fibrosis.
ABSTRACT
Objective@#To screen the key microRNAs targeting Notch signaling pathway in inner ear and investigate its potential regulating function.@*Methods@#The interaction network and the Core-Notch network, involved with key genes in Notch signal pathway and differential-expressed microRNAs in inner ear, were constructed by bioinformatics methods. The important microRNAs in regulating Notch signaling pathway were screened via topological and GO analysis, followed by in vivo and in vitro investigation.@*Results@#MiRNA-384-5p was identified as a key regulator specifically expressed in mouse brain and inner ear, which could down-regulate Notch1. The Notch1 expression was found significantly down-regulated in miRNA-384-5p-mimic-transfected HeLa cells. The dual-luciferase reporter gene assay further confirmed the effect of miRNA-384-5p on the down-regulation of Notch1 and Dll4 in Notch signaling pathway.@*Conclusions@#The Core-Notch network is constructed to screen microRNAs implicated in inner ear development, and miRNA-384-5p is screened and verified to be target-regulating the Notch signaling pathway, which could be the potential target in the regeneration of impaired hair cells.