Abstract
BackgroundCervical cancer is a serious disease with complicated pathogenesis and thus there is an urgent need to find novel targets for the treatment. Recently, long non-coding RNAs (lncRNAs) have emerged as critical factors in tumorigenesis. In this study, we aimed to investigate the mechanism of HAND2 antisense RNA 1 (HAND2-AS1) on the invasion and metastasis of cervical cancer cells.MethodsThe expression patterns of HAND2-AS1, microRNA-330-5p (miR-330-5p) and leucine zipper down-regulated in cancer 1 (LDOC1) in cervical cancer were characterized by RT-qPCR and western blot analysis. Dual luciferase reporter assay and RIP were applied to verify relationship between HAND2-AS1, miR-330-5p and LDOC1. Fluorescence in situ hybridization (FISH) was used to detect the subcellular localization of HAND2-AS1. Besides, viability, invasion and migration ability of HeLa cells were investigated by cell counting kit-8 (CCK-8) and Transwell assays respectively. Hematoxylin–eosin staining was performed for lymph node metastasis detection. In addition, the tumor growth in nude mice was evaluated.ResultsLow expression of HAND2-AS1 and LDOC1, and high expression of miR-330-5p were detected in cervical cancer tissues and cells. It was found that binding of HAND2-AS1 to miR-330-5p results in upregulation of LDOC1 expression. Also, overexpressed HAND2-AS1 and LDOC1 or down-regulated miR-330-5p inhibited expression of proliferation-associated proteins Ki-67, PCNA, migration-associated proteins N-cad and invasion-related proteins MMP-2, MMP-9 as well as lymph node metastasis. Moreover, HAND2-AS1 inhibited tumor formation and lymph node metastasis by binding to miR-330-5p in vivo.ConclusionHAND2-AS1 promotes LDOC1 expression by competitively binding to miR-330-5p and consequently inhibiting cervical cancer cell invasion and metastasis. This could facilitate development of therapeutic strategies against cervical cancer.
Highlights
Cervical cancer is a serious disease with complicated pathogenesis and there is an urgent need to find novel targets for the treatment
The sections were incubated in 3% hydrogen peroxide for 15 min, blocked with goat serum at 37 °C for 20 min and incubated with primary rabbit anti-leucine zipper down-regulated in cancer 1 (LDOC1) antibody (1:1000, ab86126, Abcam Inc., Cambridge, MA, USA) overnight at 4 °C
reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis of tissue samples obtained from 68 patients showed that HAND2-AS1 was down-regulated in cervical cancer tissues when compared to adjacent tissues (Fig. 1c)
Summary
Cervical cancer is a serious disease with complicated pathogenesis and there is an urgent need to find novel targets for the treatment. We aimed to investigate the mechanism of HAND2 antisense RNA 1 (HAND2-AS1) on the invasion and metastasis of cervical cancer cells. LncRNAs have an antagonistic effect on the post-transcriptional regulation of microRNAs (miRNAs or miRs) in gene expression and play a vital role during disease development [5]. A recent study has shown that HAND2-AS1 represses colorectal cancer progression by upregulating miR-1275-mediated KLF14 expression [8]. LncRNA WT1-AS inhibits the aggressiveness of cervical cancer cell via regulating p53 expression via sponging miR-330-5p [15]. All this evidence supports this study investigating the possible interplay between HAND2-AS1 and miR-330-5p in the development of cervical cancer
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