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Objective:To observe the expression of miR-142-5p and forkhead transcription protein O subgroup 3 (FOXO3) in CD4 + T cells of experimental autoimmune uveitis (EAU) model rats, and preliminarily explore the targeting relationship between the two and the effect on EAU impact. Methods:Ten Lewis rats were randomly divided into model group and control group. Rats in the model group wree induced an EAU animal model by adoptive immunization. Twenty days after immunization, CD4 + T cells were extracted from the eyeballs and draining lymph nodes of rats in the control group and model group, and divided into control group, model group, mimic-negative control (NC) group, miR-142-5p-mimic group, and small interference (si)-NC group, si-FOXO3 group for in vitro experiments. The miR-142-5p-mimic group and si-FOXO3 group were transfected with miR-142-5p-mimic and si-FOXO3, respectively. Twenty-five Lewis rats were randomly divided into model group, mimic-NC transfected group, miR-142-5p-mimic transfected group, si-NC transfected group, and si-FOXO3 transfected group. The above-mentioned in vitro experimental groups were injected with cells respectively. Slit lamp microscopy and EAU score were performed on 4, 8, 12, 16, 20 days after immunization; on 20 days after immunization, hematoxylin-eosin staining was performed for histopathological grading. Real-time fluorescence quantitative polymerase chain reaction was used to detect the relative expression of miR-142- 5p and FOXO3 mRNA in CD4 + T cells and eye tissues of rats in each group, and helper T cell 17 (Th17) marker interleukin (IL)-17, IL-22, retinoic acid-related orphan receptor gamma (ROR gamma) relative expression level in the supernatant. Bioinformatics website and dual luciferase was used to predict the targeting relationship between miR-142-5p and FOXO3. One-way analysis of variance or t test was used for comparison between groups. Results:All rats in the model group showed symptoms of EAU to varying degrees, and the symptoms became worse with time. Compared with the control group, the relative expression of miR-142-5p mRNA in CD4 + T cells of the model group increased, and the relative expression of FOXO3 mRNA decreased. The differences were statistically significant ( t=7.374, 10.423; P=0.002, 0.001). Compared with the mimic-NC group, the relative expression of miR-142-5p mRNA in the CD4 + T cells of the miR-142-5p-mimic group increased, and the difference was statistically significant ( t=6.540, P=0.003). Compared with the model group, mimic-NC group, and si-NC group, the relative expression of IL-17, IL-22, and RORγ mRNA in CD4 + T cells in the miR- 142-5p-mimic group and si-FOCO3 group increased significantly. The difference was statistically significant ( F=26.110, 6.292, 5.269, 55.660, 10.490, 11.430; P<0.05). Compared with the mimic-NC transfected group, the relative expression of miR-142-5p mRNA in the ocular tissues of the miR-142-5p-mimic transfected rats increased significantly, and the difference was statistically significant ( t=6.690, P<0.05). Compared with the transfected si-NC group, the relative expression of FOXO3 mRNA in the eye tissue of the transfected si-FOXO3 group was significantly decreased, and the difference was statistically significant ( t=17.751, P<0.05). Rats in the mimic-NC transfected group, miR-142-5p-mimic transfected group, si-NC transfected group, and si-FOXO3 transfected group prolonged with time after immunization, and the EAU scores showed an upward trend. The EAU score and histopathological grade of rats in the miR-142-5p-mimic transfected group were higher than those in the mimic-NC transfected group, and the difference was statistically significant ( t=5.633, 6.286; P<0.05). The EAU score and histopathological grade of the rats in the transfected si-FOXO3 group were higher than those in the transfected si-NC group, and the difference was statistically significant ( t=6.852, 6.635; P<0.05). FOXO3 has a targeting relationship with miR-142-5p. Conclusions:In EAU rat CD4 + T cells, the expression of miR-142-5p is up-regulated, while the expression of FOXO3 is down-regulated. miR-142-5p targets the expression of FOXO3 to promote the development of Th17 cell-related inflammatory factors.
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@# Objective: To study the expression of miR-142-5p in lung adenocarcinoma tissues, and to explore its effect on proliferation, invasion, migration and epithelieal-mesenchymal transition (EMT) of H1650 cells and the potential mechanisms. Methods:Atotal of 107 pairs of lung adenocarcinoma tissues and corresponding para-cancerous tissues from patients, who underwent tumor resection and were pathologically confirmed at the Department of Thoracic Surgery, the Fourth Hospital of Hebei Medical University between Jan. 2014 and Jan. 2015, were collected for this study; in addition, human lung adenocarcinoma cell lines (H1650, HCC827, A549, H1975, PC9) and human bronchial epithelial BEAS-2B cells were also used in this study. qPCR was used to detect the expression of miR-142-5p in lung adenocarcinoma tissues and cell lines. The correlation between expression of miR-142-5p and clinical features was analyzed.After transfection with miR-142-5p mimics or miR-negative control (miR-NC) plasmid, the proliferation, invasion and migration of H1650 cells were detected with CCK-8, Transwell invasion assay and Wound healing assay, respectively. The bioinforamtics tool was used to predict the target genes of miR-142-5p, and Luciferase reporter gene assay was performed to validate the regulation of miR-142-5p on target gene. Western blotting (WB) was used to detect the expressions of cyclin-dependent kinase 5 (CDK5) and EMTrelated protein. Results: Compared to Para-cancerous tissues and BEAS-2B cells, the expression of miR-142-5p was lower in lung adenocarcinoma tissues and cell lines (all P<0.01). Of the 107 cases of lung adenocarcinoma tissues, 61 cases (57.01%) showed decreased miR-142-5 expression, which was correlated with the TNM stage and lymph node metastasis (both P<0.01). Transfection of miR-142-5p mimics significantly up-regulated the expression of miR-142-5p and decreased the proliferation, invasion and migration of H1650 cells (all P<0.05 or P<0.01). Bioinformatics showed that CDK5 was a target gene of miR-142-5p. Luciferase reporter gene assay and WB validated that miR-142-5p could significantly down-regulate CDK5 expression in H1650 cells, up-regulate the expression of E-cadherin and down-regulate the expressions of N-cadherin, Twist and Snail in H1650 cells (all P<0.01). Conclusion: miR-142-5p is low expressed in lung adenocarcinoma tissues and cell lines; it suppresses the EMT process to inhibit, invasion and migration of H1650 cells via down-regulating the expression of CDK5.
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OBJECTIVE: To investigate the expression level of miR-142-5p and its potential target gene endothelial PAS domain protein 1(EPAS1) in Stage III colorectal cancer during Transcatheter arterial infusion chemotherapy (TAI). MATERIALS AND METHODS: Illumina high-throughput sequencing was used to obtain miRNA expression profiles of paired tumor and adjacent normal tissues from one patient received TAI 1 week before the operation and another patient directly underwent an operation. The expression levels of miR-142-5p was measured with both high-throughput sequencing and quantitative real time-polymerase chain reaction. RESULTS: The expression levels of miR-142-5p, were significantly reduced in tumor tissues of stage III CRC, then significantly increased in tumor tissues receiving TAI and higher than tumor tissues without TAI. The apoptosis rate of HT-29 colon cancer cells was mildly increased after transfection with pre-miR-142. miR-142-5p could bind directly to the 3´untranslated region of endothelial PAS domain protein 1 and reduce its expression. CONCLUSIONS: miR-142-5p is a potential tumor suppressor in CRC and is upregulated in tumor tissues after TAI, suggesting its potential clinical values for testing the functionality of TAI and predicting the progress of CRC.