| Objective:Papillary thyroid carcinoma(PTC)is the most common malignant tumor in endocrinology.It is reported that miR-652-3p is down-regulated in PTC tissues,and the molecular mechanism in the progression of PTC is still unclear.This study focuses on miR-652-3p to aim to explore the role and mechanisms of NEAT1/miR-652-3p/UBE2 I in PTC,so as to provide new targets and ideas for the diagnosis and treatment of PTC.Methods:1.q RT-PCR was used to detect the expression of miR-652-3p in PTC tissues and paired adjacent tissues,also in normal human thyroid follicular cell lines and PTC cell lines(TPC-1,K1,and BCPAP).Analysis was performed to explore the correlation between the expression of miR-652-3p and clinicopathological characteristics in PTC patients.CCK-8 assay and colony formation assay were performed to determine cell proliferation,wound healing assay and migration assay were performed to determine cell migration,invasion assay was performed to determine cell invasion,and flow cytometry was performed to evaluate cell apoptosis in PTC cells.PTC cells were transiently transfected with mimics or inhibitors to establish PTC cell models with overexpression of miR-652-3p or knockdown of miR-652-3p respectively.The effects of overexpression and knockdown of miR-652-3p on cell proliferation,migration,invasion,and apoptosis were examined.2.q RT-PCR and Western Blot were used to detect the expression of UBE2 I after overexpression and knockdown of miR-652-3p,and a dual-luciferase reporter gene assay was used to confirm whether there is a binding site between miR-652-3p and UBE2 I,and analyze the correlation between miR-652-3p and UBE2 I in PTC tissue.q RT-PCR was used to detect the expression of UBE2 I in PTC tissues and in PTC cell lines.PTC cells were transiently transfected with si RNA or plasmid to establish PTC cell models with knockdown of UBE2I or overexpression of UBE2I.We examined the effects of knockdown and overexpression of UBE2I on the proliferation,migration,invasion,and apoptosis of PTC cells.PTC cells were co-transfected with miR-652-3p inhibitor and si-UBE2I,and a rescue experiment was carried out to detect whether knockdown of UBE2I could rescue the promotive effects of down-regulation of miR-652-3p on the proliferation,migration,and invasion and to detect whether knockdown of UBE2I could rescue the inhibitory effects on the apoptosis in PTC cells.3.q RT-PCR was used to detect the expression of NEAT1 after overexpression and knockdown of miR-652-3p in PTC cells,and a dual-luciferase reporter gene assay was used to confirm whether there is a binding site between miR-652-3p and UBE2 I,and analyze the correlation between miR-652-3p and NEAT1 in PTC tissue.q RT-PCR was used to detect the expression of NEAT1 in PTC tissues and in PTC cell lines.PTC cells were transiently transfected with si RNA to establish PTC cell models with knockdown of NEAT1.We examined the effects of knockdown of NEAT1 on the proliferation,migration,invasion,and apoptosis of PTC cells.PTC cells were co-transfected with si-NEAT1 and miR-652-3p inhibitor,and a rescue experiment was carried out to detect whether knockdown of miR-652-3p could rescue the inhibitory effects of down-regulation of NEAT1 on the proliferation,migration,and invasion and to detect whether knockdown of miR-652-3p could rescue the promotive effects on the apoptosis in PTC cells.PTC cells were co-transfected with si-NEAT1 and overexpressed UBE2 I plasmids,and a rescue experiment was carried out to detect whether overexpression of UBE2 I could rescue the inhibitory effects of down-regulation of NEAT1 on the proliferation,migration,and invasion and to detect whether overexpression of UBE2 I could rescue the promotive effects on the apoptosis in PTC cells.4.PTC cells were infected with lentivirus to establish PTC cell models with miR-652-3p overexpression,and the effect of miR-652-3p overexpression on the growth of PTC in vivo was investigated by nude mice xenograft experiments,and the expressions of NEAT1 and UBE2 I in the xenograft tissues were detected by q RT-PCR.Results:1.Expression of miR-652-3p in PTC tissues was lower than in paired paracancerous tissues,and also was lower in PTC cell lines(TPC-1,BCPAP,K1)than in normal thyroid follicular cell lines.The low expression of miR-652-3p was positively correlated with lymph node metastasis and extrathyroidal extension.Overexpression of miR-652-3p inhibited the proliferation,migration,and invasion,and promoted apoptosis in PTC cells.Knockdown of miR-652-3p promoted the proliferation,migration,and invasion,and inhibited the apoptosis in PTC cells.2.Overexpression of miR-652-3p down-regulated the expression of UBE2 I,and knockdown of miR-652-3p up-regulated the expression of UBE2 I.The dual-luciferase reporter gene assay confirmed that miR-652-3p could directly bind to the 3'-UTR of UBE2 I.The expression of UBE2 I was negatively correlated with the expression of miR-652-3p in PTC tissues.UBE2 I was highly expressed in PTC tissues and PTC cell lines.Knockdown of UBE2 I inhibited proliferation,migration,and invasion,and promoted apoptosis in PTC cells.Overexpression of UBE2 I promoted proliferation,migration,and invasion,and inhibited apoptosis in PTC cells.Knockdown of UBE2 I rescued the promotive effects of down-regulation of miR-652-3p on the proliferation,migration,and invasion,and rescued the inhibitory effects on the apoptosis in PTC cells.3.Overexpression of miR-652-3p down-regulated the expression of NEAT1,and knockdown of miR-652-3p up-regulated the expression of NEAT1.The dual-luciferase reporter gene assay confirmed that miR-652-3p directly binds to NEAT1.NEAT1 was highly expressed in PTC tissues and in PTC cell lines.Knockdown of NEAT1 inhibited the proliferation,migration,and invasion and promoted apoptosis in PTC cells.Knockdown of miR-652-3p rescued the inhibitory effects of down-regulation of NEAT1 on the proliferation,migration,and invasion,and rescued the promotive effects on the apoptosis in PTC cells.Overexpression of UBE2 I rescued the inhibitory effects of down-regulation of NEAT1 on the proliferation,migration,and invasion,and rescued the promotive effects on the apoptosis in PTC cells.4.Overexpression of miR-652-3p inhibited the growth of transplanted tumors in nude mice and overexpression of miR-652-3p down-regulated the expression of NEAT1 and UBE2 I in the xenograft tissues.Conclusions:1.miR-652-3p is lowly expressed in PTC tissues and cells,and the low expression of miR-652-3p is positively correlated with lymph node metastasis and extrathyroidal extension.miR-652-3p inhibits proliferation,migration,and invasion,and promotes apoptosis in PTC.2.UBE2 I is highly expressed in PTC tissues and cells,and UBE2 I promotes proliferation,migration,and invasion,and inhibits apoptosis in PTC.miR-652-3p negatively regulates UBE2 I to inhibit the proliferation,migration,and invasion and promotes apoptosis in PTC.3.NEAT1 is highly expressed in PTC cells,NEAT1 regulates the expression of UBE2 I by competitively binding to miR-652-3p to promote proliferation,migration,and invasion and inhibits apoptosis in PTC.NEAT1/miR-652-3p/UBE2 I promotes the progression of PTC through a ce RNA mechanism. |
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