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. 2016 Aug 5:16:605.
doi: 10.1186/s12885-016-2628-z.

MicroRNA-21 promotes proliferation, migration, and invasion of colorectal cancer, and tumor growth associated with down-regulation of sec23a expression

Chenli Li  1 , Lingxu Zhao  1 , Yuan Chen  1 , Tiantian He  1 , Xiaowan Chen  1 , Jiating Mao  1 , Chunmei Li  1 , Jianxin Lyu  2 , Qing H Meng  3
Affiliations

MicroRNA-21 promotes proliferation, migration, and invasion of colorectal cancer, and tumor growth associated with down-regulation of sec23a expression

Chenli Li et al. BMC Cancer. .

Abstract

Background: MicroRNA-21 (miR-21) is up-regulated in many cancers, including colorectal cancer (CRC). Nevertheless, the function of miR-21 in CRC and the mechanism underlying that function is still unclear.

Methods: After analyzing the expression of miR-21 and Sec23A in CRC cell lines, we transfected the highest miR-21 expressing cell line, SW-480, with a plasmid containing an miR-21 inhibitor and the lowest miR-21 expressing cell line, DLD-1, with a plasmid containing an miR-21 mimic and measured the effects on the expression of Sec23A and on cell proliferation, migration, and invasion. We also evaluated the effect of knocking down Sec23A on miR-21 expression and its effects on cell proliferation, migration, and invasion. Finally, we assessed the effect of miR-21 in a xenograft tumor model in mice. Tumor tissues from these mice were subjected to immunohistochemical staining to detect the expression of Sec23A.

Results: Genetic deletion of miR-21 suppressed the proliferation, migration, and invasion of SW-480 cells, while over-expression of miR-21 promoted proliferation, migration, and invasion of DLD-1 cells. Inhibition of miR-21 increased the expression of Sec23A protein in SW-480 cells while over-expression of miR-21 significantly suppressed the expression of Sec23A protein and Sec23A mRNA in DLD-1 cells. Knockdown of Sec23A increased the expression of miR-21 in SW480 and DLD-1 cells and their proliferation (DLD-1 only), migration, and invasion. Over-expression of miR-21 promoted tumor growth in BALB/c nude mice and suppressed tumor expression of Sec23A.

Conclusion: These findings provide novel insight into the molecular functions of miR-21 in CRC, which may serve as a potential interesting target.

Keywords: Colorectal cancer; Proliferation; Sec23A; Tumor growth; miR-21.

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Figures

Fig. 1
Fig. 1
miR-21 is overexpressed in CRC cell lines. a Expression of miR-21 in untreated HT-29, SW-480, and DLD-1 cell lines. b The expression of miR-21 in SW-480 cells after transfection with miR-21 inhibitor, negative control (NC) or cells without treatment (MOCK). c The expression of miR-21 in DLD-1 cells after transfection with miR-21 mimic, negative control (NC) or cells without treatment (MOCK). * p < 0.05, ** p < 0.01, *** p < 0.001
Fig. 2
Fig. 2
MiR-21 overexpression stimulates proliferation, migration, and invasion of CRC cells. a-c Inhibition of miR-21 in SW-480 by miR-21 inhibitor reduced cell proliferation, migration, and invasion relative to negative control (NC) and mock-treated cells (MOCK). d-f Up-regulation of miR-21 in DLD-1 by miR-21 mimic increased proliferation, migration, and invasion relative to negative control and mock-treated cells. In panels b, c, e, and f, photos on the left are representative images of migrated (b, e) and invaded cells on matrigel membranes (c, f), while the graphs on the right indicate quantification of cells. ** p < 0.01
Fig. 3
Fig. 3
MiR-21 inhibits expression of Sec23A in CRC cell lines. a Inhibition of miR-21 expression promoted the expression of Sec23A protein in SW-480 cells relative to negative control (NC) and mock-treated cells (MOCK). b There was no significant difference in expression of Sec23A mRNA in SW-480 cells relative to negative control and mock-treated cells. c-d Over-expression of miR-21 by miR-21 mimic decreased the expression of Sec23A protein and mRNA in DLD-1 cells relative to negative control and mock-treated cells. Protein expression in these cells was determined by densitometric analysis. * p < 0.05, ** p < 0.01
Fig. 4
Fig. 4
Sec23A knockdown increases expression of miR-21 in CRC cell lines. a Expression of Sec23A mRNA in untreated HT-29, SW-480, and DLD-1 cell lines. b-c Sec23A mRNA and protein expression was suppressed in Sec23A knockdown SW-480 cells. d-e Sec23A mRNA and protein expression was suppressed in Sec23A knockdown DLD-1 cells. Protein expression in these cells was quantified by densitometric analysis. f-g MiR-21 expression was increased in Sec23A knockdown SW-480 and DLD-1 cells. ** p < 0.01
Fig. 5
Fig. 5
Downregulation of Sec23A promotes the proliferation, migration, and invasion of CRC cells. a-c Sec23A knockdown increased migration, and invasion of SW-480 cells relative to negative controls (sh-NC). d-f Sec23A knockdown increased proliferation, migration, and invasion of DLD-1 cells relative to negative controls. In panels b, c, e, and f, photos on the left are representative images of migrated (b, e) and invaded cells on Matrigel membranes (c, f) while the graphs on the right indicate quantification of cells. * p < 0.05, ** p < 0.01
Fig. 6
Fig. 6
Over-expression of miR-21 enhances DLD-1 tumor growth in BALB/c nude mice. a Mice inoculated with miR-21 over-expressing DLD-1 cells grew larger tumors than mice inoculated with negative controls (NC) or mock-treated cells (MOCK). b Tumor-bearing mice before dissection. c The dissected tumors. d Tumors dissected from mice inoculated with miR-21 over-expressing DLD-1 cells weighed significantly more than tumors from mice inoculated with negative control or mock-treated cells. * p < 0.05, ** p < 0.01
Fig. 7
Fig. 7
Greater expression of miR-21 in DLD-1 tumors is associated with lower levels of Sec23A protein expression. a Tumors from mice inoculated with miR-21 over-expressing DLD-1 cells expressed higher levels of miR-21 than tumors from mice inoculated with negative controls (NC) or mock-treated cells (MOCK). b Tumors from mice inoculated with miR-21 over-expressing DLD-1 cells expressed lower levels of Sec23A than tumors from mice inoculated with negative controls or mock-treated cells. c The expression of Sec23A in tumor tissues was determined by immunohistochemical staining; representative images are shown (original magnification, ×200). * p < 0.05, ** p < 0.01
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