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. 2020 Feb 13:12:1135-1149.
doi: 10.2147/CMAR.S237439. eCollection 2020.

lncRNA ZFAS1 Is Involved in the Proliferation, Invasion and Metastasis of Prostate Cancer Cells Through Competitively Binding to miR-135a-5p

Jiaqiang Pan #  1 , Xingyan Xu #  2 , Guangliang Wang  3
Affiliations

lncRNA ZFAS1 Is Involved in the Proliferation, Invasion and Metastasis of Prostate Cancer Cells Through Competitively Binding to miR-135a-5p

Jiaqiang Pan et al. Cancer Manag Res. .

Abstract

Background: Prostate cancer (PCa) is a common malignant tumor in men. lncRNA ZFAS1 plays a carcinogenic role in many types of cancer; however, its potential role in PCa remains unclear. The current study aimed to determine the expression and function of ZFAS1 in PC.

Methods: The ZFAS1 expression in PC tissues and cells was determined by quantitative polymerase chain reaction (qPCR). SiZFAS1, miR-135a-5p mimic and miR-135a-5p inhibitor were transfected into PCa cells. The direct target of ZFAS1 was predicted by Starbase and verified by dual-luciferase reporter. Cell viability, proliferation, apoptosis, migration and invasion of the PCa cells were determined by cell counting kit-8, clone formation assay, flow cytometer, scratch and Transwell assay, respectively. The expression levels of related proteins and mRNAs were determined by Western blotting and qPCR.

Results: ZFAS1 expression was up-regulated in PCa cells and tissues. ZFAS1 could competitively bind to miR-135a-5p in PCa cells, and down-regulation of ZFAS1 inhibited cell viability, proliferation, migration, invasion of PCa cells and the occurrence of epithelial-mesenchymal transformation (EMT) and promoted apoptosis of PCa cells and increased the miR-135a-5p expression. Moreover, the function of miR-135a-5p mimic in PCa cells was consistent with ZFAS1 knockdown, while the function of miR-135a-5p inhibitor was opposite to that of miR-135a-5p mimic in PCa cells. The results showed that knocking down ZFAS1 could attenuate the effects of miR-135a-5p inhibitor on cell proliferation, invasion and EMT of PCa cells.

Conclusion: Knocking down ZFAS1 could inhibit the proliferation, invasion and metastasis of PCa cells through regulating miR-135a-5p expression.

Keywords: epithelial-mesenchymal transition; lncRNA ZFAS1; miR-135a-5p; prostate cancer.

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Conflict of interest statement

The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
Expression and effect of long non-coding RNA zinc finger antisense 1 (lncRNA ZFAS1) in prostate cancer (PCa) tissues and cell lines. (A) Expression of ZFAS1 in tissues from patients with PCa, quantitative polymerase chain reaction (qPCR) was performed. (B) Expression of ZFAS1 in RWPE-1 cells and different PCa cell lines (PC3, DU145, 22RV1 and LNCAP) was detected by qPCR. (C) The siRNA (siZFAS1) was used to construct ZFAS1 knockdown PC3 cells, and the knockdown efficiency was detected by qPCR. (D) The siRNA was used to construct ZFAS1 knockdown DU145 cells, and the knockdown efficiency was detected by qPCR. (E) Cell counting kit-8 kit (CCK-8) assay showed that siZFAS1 inhibited cell viability of PC3 cells. (F) CCK-8 assay showed that siZFAS1 inhibited cell viability of DU145 cells. (G) Clone formation assay showed that siZFAS1 decreased colony number of PC3 cells. (H) Clone formation assay showed that siZFAS1 decreased colony number of DU145 cells. ##P<0.01 vs RWPE-1; *P<0.05 vs siNC, **P<0.01 vs siNC.
Figure 2
Figure 2
siZFAS1 regulated apoptosis and migration of PCa cells. (A) Flow cytometry showed that PC3 cells transfected with siZFAS1 increased apoptosis rate. (B) Flow cytometry showed that DU145 cells transfected with siZFAS1 increased apoptosis rate. (C) Inhibition of siZFAS1 on the migration of PC3 cells was observed by scratch assay. (D) Inhibition of siZFAS1 on the migration of DU145 cells was observed by scratch assay. **P<0.01 vs siNC.
Figure 3
Figure 3
siZFAS1 regulated invasion of PCa cells and expressions of epithelial-mesenchymal transformation (EMT)-related proteins. (A) Inhibition of siZFAS1 on the invasion of PC3 cells was observed by Transwell assay. (B) Inhibition of siZFAS1 on the invasion of DU145 cells was observed by Transwell assay. (C) Western blotting results indicated that siZFAS1 down-regulated the expressions of N-cadherin and Snail, and up-regulated the E-cadherin expression of PC3 cells. (D) Western blotting results indicated that siZFAS1 down-regulated the N-cadherin and Snail expressions, and up-regulated the E-cadherin expression of DU145 cells. **P<0.01 vs siNC.
Figure 4
Figure 4
The relationship between ZFAS1 and miR-135a-5p. (A) StarBase predicted that ZFAS1 could bind to miR-135a-5p. (B) Dual-luciferase assay on PC3 cells co-transfected with ZFAS1-WT and miR-135a-5p mimic. (C) Dual-luciferase assay on DU145 cells co-transfected with ZFAS1-WT and miR-135a-5p mimic. (D) Expression of miR-135a-5p in tissues from patients with PC, quantitative polymerase chain reaction (qPCR) was performed. (E) Expression of miR-135a-5p in RWPE-1 cells and different PCa cell lines (PC3, DU145, 22RV1 and LNCAP) was detected by qPCR. (F) siZFAS1 up-regulated the miR-135a-5p expression of PC3 cells, as detected by qPCR. (G) siZFAS1 up-regulated the miR-135a-5p expression of DU145 cells, as detected by qPCR. ##P<0.01 vs Blank; **P<0.01 vs siNC; ^^P<0.01 vs RWPE-1.
Figure 5
Figure 5
Effects of miR-135a-5p and siZFAS1 on PCa cells. (A) PC3 cells were transfected with miR-135a-5p mimic (M), miR-135a-5p mimic control (MC), miR-135a-5p inhibitor (I), miR-135a-5p inhibitor control (IC) or siZFAS1, transfection efficiency was detected by qPCR. (B) DU145 cells were transfected with miR-135a-5p mimic (M), miR-135a-5p mimic control (MC), miR-135a-5p inhibitor (I), miR-135a-5p inhibitor control (IC) or siZFAS1, transfection efficiency was detected by qPCR. (C) CCK-8 assay showed that miR-135a-5p mimic decreased cell viability of PC3 cells, miR-135a-5p inhibitor increased cell viability, and siZFAS1 partially reversed the miR-135a-5p inhibitor effect. (D) CCK-8 assay showed that miR-135a-5p mimic decreased cell viability of DU145 cells, miR-135a-5p inhibitor increased cell viability, and siZFAS1 partially reversed the miR-135a-5p inhibitor effect. (E) The images of the clone formation assay of PC3 cell. (F) The images of the clone formation assay of DU145 cell. (G) Clone formation assay showed that miR-135a-5p mimic decreased colony number of PC3 cells, miR-135a-5p inhibitor increased colony number, and siZFAS1 partially reversed the miR-135a-5p inhibitor effect. (H) Clone formation assay showed that miR-135a-5p mimic decreased colony number of DU145 cells, miR-135a-5p inhibitor increased colony number, and siZFAS1 partially reversed the miR-135a-5p inhibitor effect. **P<0.01 vs MC; ##P<0.01 vs IC; ^^P<0.01 vs I.
Figure 6
Figure 6
Effects of miR-135a-5p and siZFAS1 on PCa cell apoptosis and migration. (A) Flow cytometry showed that miR-135a-5p mimic promoted apoptosis of PC3 cells, miR-135a-5p inhibitor inhibited apoptosis, and siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. (B) Flow cytometry showed that miR-135a-5p mimic promoted apoptosis of DU145 cells, miR-135a-5p inhibitor inhibited apoptosis, and siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. (C) Scratch assay showed that miR-135a-5p mimic decreased migration of PC3 cells, miR-135a-5p inhibitor increased migration, and siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. (D) Scratch assay showed that miR-135a-5p mimic decreased migration of DU145 cells, miR-135a-5p inhibitor increased migration, and siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. **P<0.01 vs MC; #P<0.05 vs IC, ##P<0.01 vs IC; ^^P<0.01 vs I.
Figure 7
Figure 7
Effects of miR-135a-5p and siZFAS1 on PCa cell invasion and the EMT-related protein expressions. (A) Inhibition of miR-135a-5p mimic on the invasion of PC3 cells was observed by Transwell assay, and miR-135a-5p inhibitor promoted invasion, siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. (B) Inhibition of miR-135a-5p mimic on the invasion of DU145 cells was observed by Transwell assay, and miR-135a-5p inhibitor promoted invasion, siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. (C) Western blotting results indicated that miR-135a-5p mimic inhibited the occurrence of EMT on PC3 cells, miR-135a-5p inhibitor promoted EMT, and siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. (D) Western blotting results indicated that miR-135a-5p mimic inhibited the occurrence of EMT on DU145 cells, miR-135a-5p inhibitor promoted EMT, and siZFAS1 partially reversed the effect by miR-135a-5p inhibitor. **P<0.01 vs MC; ##P<0.01 vs IC; ^^P<0.01 vs I.
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References

    1. Siegel RL, Miller KD, Jemal A. Cancer statistics, 2017. CA Cancer J Clin. 2017;67(1):7–30. doi:10.3322/caac.21387 - DOI - PubMed
    1. Stuchbery R, Kurganovs NJ, McCoy PJ, et al. Target acquired: progress and promise of targeted therapeutics in the treatment of prostate cancer. Curr Cancer Drug Targets. 2015;15(5):394–405. doi:10.2174/1568009615666150416113453 - DOI - PubMed
    1. Jones C, Fam MM, Davies BJ. Expanded criteria for active surveillance in prostate cancer: a review of the current data. Transl Androl Urol. 2018;7(2):221–227. doi:10.21037/tau - DOI - PMC - PubMed
    1. Thorstenson A, Garmo H, Adolfsson J, Bratt O. Cancer specific mortality in men diagnosed with prostate cancer before age 50 years: a nationwide population based study. J Urol. 2017;197(1):61–66. doi:10.1016/j.juro.2016.06.080 - DOI - PubMed
    1. Feldman AS, Meyer CP, Sanchez A, et al. Morbidity and mortality of locally advanced prostate cancer: a population based analysis comparing radical prostatectomy versus external beam radiation. J Urol. 2017;198(5):1061–1068. doi:10.1016/j.juro.2017.05.073 - DOI - PubMed