Screening of differently expressed miRNAs and the mechanism of miR-3907 in meibomian gland carcinoma

Abstract

Objective: To screen the differently expressed microRNAs (miRNAs) and to explore the effect and mechanism of microRNA-3907 (miR-3907) in meibomian gland carcinoma (MGC). Methods: Experimental research. MGC tissues and para-carcinoma tissues of patients diagnosed with MGC by histopathology were collected from July 2011 to January 2019 in Tianjin Medical University Eye Hospital. The miRNA microarray analysis of MGC and para-carcinoma tissue samples from 5 patients was performed. miR-3907 with a significant up-regulation was selected as a research object. Bioinformatics predicted and dual-luciferase gene reporter assay verified miR-3907 target genes. The protein expression levels of target genes in 18 MGC tissues and 6 para-carcinoma tissue samples were determined by immunohistochemical staining. miR-3907 over-expression, miR-3907 knock-down, target gene knock-down and miR-3907 knock-down with target gene knock-down were respectively performed in MGC cell. The mRNA and protein expressions were validated by real-time PCR and Western blotting after transfection. The cell proliferation and migration ability was detected by cell counting kit-8 and scratch experiment after transfection. The main statistical methods were Fisher's exact test, independent sample t test, two-factor repeated measure analysis of variance. Results: There were 22 differently up-regulated miRNAs and 5 differently down-regulated miRNAs in MGC tissues, of which miR-3907 was significantly up-regulated. Thrombospondin-1 (THBS1) was a target gene of miR-3907 according to bioinformatics and dual-luciferase gene reporter assay. The positive expression rate of THBS1 protein in para-carcinoma tissues (6/6) was significantly higher than that in MGC tissues (5/18), and the difference was statistically significant (P=0.003). Compared with the negative control group, the proliferation ability of the miR-3907 over-expression group was increased at 48 h and 72 h (F=3.70, 2.65; both P<0.01), and the migration rate at 24 h was significantly higher (54.6%±3.4% vs. 34.2%±0.6%; t=8.34, P<0.01). Compared with the negative control group, the proliferation ability of the miR-3907 knock-down group was decreased at 24 h, 48 h and 72 h (F=3.10, 2.17, 3.09; all P<0.05), and the migration rate at 24 h was significantly lower (40.8%±2.8% vs. 69.7%±2.7%; t=10.42, P<0.01). Compared with the negative control group, the THBS1 knock-down group promoted cell proliferation at 24 h, 48 h and 72 h (F=3.84, 3.79, 2.24; all P<0.05), and the migration rate at 24 h was significantly increased (82.5%±1.9% vs. 37.6%±5.1%; t=11.74, P<0.01). Compared with the control group, the miR-3907 knock-down with THBS1 knock-down group increased proliferation at 24 h and 48 h (F=3.97, 3.31; both P<0.05), and the migration healing rate at 24 h was significantly increased (56.9%±2.2% vs. 41.9%±4.3%; t=3.53, P<0.05). Conclusions: There are differently expressed miRNAs between MGC and para-carcinoma tissues, which may be related to the occurrence and development of MGC. miR-3907 in MGC tissues has a significant difference from that in para-carcinoma tissues. Moreover, miR-3907 can play a role in promoting proliferation and migration of MGC by inhibiting the expression of THBS1.