EXPRESSION OF MIR-96 IN PATIENTS WITH UTERINE FIBROIDS AND ITS EFFECTS ON PROLIFERATION AND APOPTOSIS OF UTERINE FIBROID CELLS

Fengchun Wang¹, Jingsen Dong^{2, *}

¹School of Nursing, West head of Liantong Road, New District, Zibo City, Shandong Province, zibo vocational institute, Zibo 255314, P.R.China - ²Gynaecology, Jinan Maternity and Child Care Hospital Affiliated to Shandong First Medical University, Jinan 250000, P.R.China

Objective: This paper aimed to investigate miR-96 expression in patients with uterine fibroids (UFs) and its effects on the proliferation and apoptosis of uterine fibroid cells (UFCs).

Methods: Patients with UFs treated in our hospital from October 2018 to October 2019 were enrolled, with human uterine leiomyosarcoma cells (SK-UT-1 cells) and normal human uterine smooth muscle cells (HUSMCs) purchased from BeNa Culture Collection. qRT-PCR was adopted for detecting miR-96 and BRCA1 expression in samples, and Western Blotting (WB) was adopted for detecting BRCA1 protein expression in the cells. Cell proliferation, invasion and apoptosis were detected by MTT assay, Transwell and flow cytometry. 

Results: Compared with adjacent tissues, miR-96 mRNA expression in uterine fibroid tissues remarkably upregulated (P<0.001), and compared with HUSMC cells, the expression in SK-UT-1 cells remarkably upregulated (P<0.001). SK-UT-1 cells were chosen for transfection. According to qRT-PCR, after transfection, miR-96 expression was remarkably lower in the miR-96-inhibitor group but remarkably higher in the miR-96-minics group compared with the NC group (both P<0.001). Cell proliferation remarkably reduced in the miR-96-inhibitor group, but enhanced in the miR-96-minics group compared with the NC group (both P<0.001); the apoptotic rate was remarkably inhibited in the miR-96-minics group (P<0.001); cell invasion remarkably reduced in the miR-96-inhibitor group, but remarkably enhanced in the miR-96-minics group (P<0.001). According to WB, BRCA1 expression remarkably downregulated in SK-UT-1 cells transfected with miR-96-minics, but remarkably upregulated in those transfected with miR-96-inhibitor (P<0.001). BRCA1 protein level in the sh-BRCA1 group remarkably rose (P<0.001), showing that the inhibitory effect of miR-96-minics on the level could be reversed by transfection with sh-BRCA1. According to the functional analysis, after transfection with sh-BRCA1, SK-UT-1 cells had remarkably inhibited proliferation and invasion, but remarkably enhanced apoptosis (all P<0.01). Compared with the NC group, S-phase cells reduced, the expression of pro-apoptotic proteins remarkably upregulated, the expression of Bcl-2, APR3 and Bcl-xl was remarkably inhibited in the sh-BRCA1 group. After transfection with miR-96-minics/sh-BRCA1, S-phase cells rose, the expression of pro-apoptotic proteins was remarkably inhibited compared with the sh-BRCA1 group, and the expression of Bcl-2, APR3 and Bcl-xl remarkably upregulated (P<0.05). 

Conclusion: Highly expressed in patients with UFs, miR-96 can target BRCA1 to inhibit SK-UT-1 cells to proliferate and promote their apoptosis.