LncRNA LINC01857 reduces metastasis and angiogenesis in breast cancer cells via regulating miR-2052/CENPQ axis

Abstract

Long non-coding RNAs have been confirmed closely related to the metastasis and angiogenesis of breast cancer (BC). LINC01857 can promote the growth and metastasis of BC cells. The present work focused on exploring the role of LINC01857 in BC metastasis and angiogenesis and investigating the possible mechanisms. The results showed that LINC01857 and CENPQ were highly expressed in BC tissues and cells, while miR-2052 was contrarily expressed. In vitro study showed that low expression of linc01857 could inhibit the migration ability and vascularization of BC cells, and mir-2052 inhibitor partially restored the effect of si-LINC01857 on the migration ability and vascularization of BC cells. Likewise, inhibition of CENPQ can partially rescue the effects of miR-2052 inhibitor on the migration ability and vascularization of BC cells. In vivo studies showed that down-regulation of LINC01857 notably suppressed tumor growth and angiogenesis in nude mice. The miR-2052 inhibitor partially restored the effects of si-LINC01857. CENPQ suppression partially rescued the effects of the miR-2052 inhibitor. To conclude, LINC01857/miR-2052/CENPQ is the potential novel target for BC treatment.

Keywords: CENPQ; LncRNA LINC01857; MiR-2052; angiogenesis; breast cancer; metastasis.

Figure 1

LINC01857 levels are increased in BC tissues and cells: (a) expression of LINC01857 in total tumor tissues, and the three isoforms (ERBB2−, ERBB2+, and Triple-Neg) were determined by qRT-PCR; (b) VEGF and CD31 levels in BC tissues and (c) cells were detected by qRT-PCR assay; (d) the correlation of VEGF or CD31 with LINC01857 was analyzed by correlation analysis. ^** P < 0.01 and ^*** P < 0.001 vs normal tissues or MCF10A cells.

Figure 2

LINC01857 down-regulation inhibits metastasis of BC cells and angiogenesis of HUVEC: (a) qRT-PCR analysis was performed to evaluate the LINC01857 expressions in MDA-MB-231 transfected with si-LINC01857; (b) angiogenesis experiment was used to evaluate the effects of si-LINC01857 on the tube formation of HUVEC cells; (c) levels of angiogenesis-related proteins, including VEGF and CD31 in TCM-treated HUVEC cells, were evaluated by western blotting assay. The effects of si-LINC01857 on MDA-MB-231 cells metastasis were detected by (d) wound healing and Transwell, (e) migration, and (f) invasion analysis. (g) The levels of metastasis-related proteins, including MMP-2 and MMP-9 in MDA-MB-231 cells transfected with si-LINC0185, were evaluated by western blotting assay. ^** P < 0.01 and ^*** P < 0.001 vs si-NC group.

Figure 3

Regulatory relationship between LINC01857 and miR-2052 in BC: (a) the binding sites between LINC0185 and miR-2052; (b) the interaction between LINC0185 and miR-2052 was evaluated by a dual-luciferase reporter assay. ^*** P < 0.001 vs NC mimic group. MiR-2052 levels in BC (c) tissues and (d) cells were detected by the qRT-PCR assay. ^*** P < 0.001 vs normal tissues or MCF10A cells. (e) MiR-2052 levels in BC cells transfected with si-LINC01857 were determined by qRT-PCR assay. ^*** P < 0.001 vs si-NC group.

Figure 4

MiR-2052 targets CENPQ in BC: (a) the binding sites between miR-2052 and CENPQ; (b) the interaction between miR-2052 and CENPQ was evaluated by a dual-luciferase reporter assay. ^*** P < 0.001 vs NC mimic group. (c) CENPQ levels in BC tissues were detected by the qRT-PCR assay. ^*** P < 0.001 vs normal tissues. CENPQ levels in BC cells were detected by (d) qRT-PCR and (e) western blot assays. ^*** P < 0.001 vs MCF10A cells. (f) CENPQ levels in BC cells transfected with miR-2052 mimic/inhibitor were determined by qRT-PCR assay. ^** P < 0.01 vs NC mimic group and ^### P < 0.001 vs or NC inhibitor group.

Figure 5

MiR-2052/CENPQ mediates LINC01857 functions on angiogenesis and metastasis of BC cells: (a) angiogenesis experiment was used to evaluate the angiogenesis of BC cells after transfection; (b) western blot was used to evaluate the angiogenesis-related proteins. The metastasis of BC cells after transfection was determined by (c) wound healing and Transwell, (d) migration, and (e) invasion analysis. (f) Western blot was used to evaluate the metastasis-related proteins. ^** P < 0.01, ^*** P < 0.001 vs si-NC group, ^## P < 0.01, ^### P < 0.001 vs si-LINC01857 group, ^$ P < 0.05, ^$$ P < 0.01, and ^$$$ P < 0.001 vs si-LINC01857 + miR-2052 inhibitor group.

Figure 6

LINC01857 down-regulation inhibits BC tumor growth in vivo: (a) the representative pictures of mice and tumor tissues are shown (n = 6); (b) histological changes of tumors from each group were determined by HE staining; (c) the volumes and (d) weight of tumors from each group were determined; (e) the protein expressions of VEGF and CD31 in tumors from each group were detected by immunohistochemical analysis. ^*** P < 0.001 vs si-NC group, ^### P < 0.001 vs si-LINC01857 group, and ^$$$ P < 0.001 vs si-LINC01857 + miR-2052 inhibitor group.