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Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells

  • Authors:
    • Xuyang Li
    • Jiletu Huge
    • Xiaoling Wang
    • Jing Zhou
    • Xiudan Li
    • Xiaoyan Gao
    • Tiewei Shi
    • Chunying Bai
  • View Affiliations / Copyright

    Affiliations: Key Laboratory of Research on Human Genetic Diseases at Universities of Inner Mongolia Autonomous Region, Chifeng University, Chifeng, Inner Mongolia Autonomous Region 024000, P.R. China, Basic Medicine College, Chifeng University, Chifeng, Inner Mongolia Autonomous Region 024000, P.R. China
    Copyright: © Li et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 221
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    Published online on: April 8, 2026
       https://doi.org/10.3892/ol.2026.15575
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Abstract

The overexpression of 5‑lipoxygenase (5‑LOX) plays a notable role in the development of colorectal cancer (CRC); however, the specific role of 5‑LOX in the migration and invasion of CRC cells through the knockdown and overexpression of 5‑LOX has not been well studied. Epithelial‑mesenchymal transition (EMT) is associated with an invasive phenotype in CRC; however, the relationship between 5‑LOX and EMT in CRC cells remains undefined. In the present study, the effects of 5‑LOX overexpression and silencing on migration, invasion and EMT in CRC cells was investigated. In addition, the mRNA levels of vascular endothelial growth factor (VEGF) and angiogenin were also analyzed in human umbilical vein endothelial cells (HUVECs) when co‑cultured with media acquired from 5‑LOX‑silenced or 5‑LOX‑overexpressed CRC cells. Reverse transcription‑quantitative PCR and western blotting analyses were used to determine mRNA and protein expression levels, respectively. Transwell and wound healing assays were used to determine the invasion and migration ability of CRC cells. Fluorescent images were captured to demonstrate the lentiviral infection in CRC cells. Analysis revealed that the knockdown of 5‑LOX by small interfering RNA significantly inhibited migration, invasion and EMT in HCT116 cells, whereas the overexpression of 5‑LOX by recombinant lentiviruses significantly promoted migration, invasion and EMT in RKO cells. Media from 5‑LOX‑overexpressed RKO cells upregulated the expression of VEGF and angiogenin in HUVECs. These results demonstrated that the overexpression of 5‑LOX promoted EMT in CRC cells, which may play a marked role in facilitating cellular migration and invasion. Furthermore, the tumor microenvironment of 5‑LOX‑overexpressed CRC cells may induce the angiogenesis in HUVECs. The present findings elucidate the role of 5‑LOX in the migration and invasion of CRC cells via the knockdown and overexpression of the 5‑LOX gene.
View Figures

Figure 1

Expression levels of 5-LOX in human
colorectal cancer cell lines. (A) Relative mRNA levels of 5-LOX in
RKO, SW480, HCT116 and HT29 cell lines were measured by reverse
transcription-quantitative polymerase chain reaction. β-actin was
used as an internal control. (B) Protein expression levels of 5-LOX
in RKO, SW480, HCT116 and HT29 cell lines were determined by
western blotting. GAPDH protein was used as an internal control.
Values are expressed as the mean ± standard error of the mean
(n=3). ***P<0.001. 5-LOX, 5-lipoxygenase.

Figure 2

Knockdown efficiency of HCT116 cells
transfected by siRNAs. (A) Relative mRNA levels of 5-LOX in HCT116
cells with three different siRNAs treatments were measured by
reverse transcription-quantitative PCR. β-actin was used as an
internal control. (B) Relative protein expression of 5-LOX in
HCT116 cells with siRNA transfection were determined by western
blotting. GAPDH protein was used as an internal control. Values are
expressed as the mean ± standard error of the mean (n=3).
**P<0.01, ***P<0.001. 5-LOX, 5-lipoxygenase; siRNA, small
interfering RNA; NC, negative control.

Figure 3

Efficiency of lentiviral infection on
5-LOX OE in RKO cells. (A) Bright field and fluorescence images
following the lentiviral infection of RKO cells in OE-Vector and
OE-5-LOX groups. (B) Relative mRNA expression of 5-LOX in RKO cells
with two treatments (OE-Vector and OE-5-LOX) was determined by
reverse transcription-quantitative PCR. (C) Relative protein
expression of 5-LOX in RKO cells with two treatments (OE-Vector and
OE-5-LOX) was determined by western blotting. Values are expressed
as the mean ± standard error of the mean (n=3). ***P<0.001.
OE-Vector, cells infected with empty vector control lentivirus;
OE-5-LOX, cells infected with 5-LOX recombinant lentivirus; GFP,
green fluorescent protein; 5-LOX, 5-lipoxygenase.

Figure 4

Migratory and invasive abilities of
5-LOX-silenced HCT116 cells and 5-LOX-OE RKO cells. HCT-116 cells
transfected with siRNAs were divided into si-NC and si-5-LOX-#2
groups. RKO cells infected with lentivirus were divided into
OE-Vector and OE-5-LOX groups. (A) Representative migrated HCT116
cells stained with crystal violet on the lower side of the membrane
and (B) quantified numbers of migrated HCT116 cells in the
Transwell migration assay. (C) Number and (D) representative images
of invasive HCT116 cells in the Transwell invasion assay. (E)
Representative images and (F) quantified numbers of migrated RKO
cells in the Transwell migration assay. (G) Number of (H) invasive
RKO cells in the Transwell invasion assay. The migration rate of
(I) HCT-116 cells and (J) RKO cells was determined by woundhealing
assay. Values are expressed as the mean ± standard error of the
mean (n=3). **P<0.01, ***P<0.001. OE-Vector, cells infected
with empty vector control lentivirus; OE-5-LOX, cells infected with
5-LOX recombinant lentivirus; siRNA, small interfering RNA; 5-LOX,
5-lipoxygenase; NC, negative control.

Figure 5

Apoptosis rate regulated by 5-LOX
knockdown and overexpression in CRC cells. The apoptotic rates of
(A) siRNA-silenced HCT116 cells (si-NC and si-5-LOX-#2 groups) and
(B) lentivirus-overexpressed RKO cells (OE-Vector and OE-5-LOX
groups) were determined by flow cytometry. **P<0.01,
***P<0.001. OE-Vector, cells infected with empty vector control
lentivirus; OE-5-LOX, cells infected with 5-LOX recombinant
lentivirus; siRNA, small interfering RNA; CRC, colorectal cancer;
5-LOX, 5-lipoxygenase; NC, negative control.

Figure 6

Levels of EMT-associated proteins
regulated by 5-LOX knockdown and overexpression in CRC cells. (A)
The levels of EMT-associated proteins (E-cadherin, N-cadherin,
Snail and Vimentin) were determined by WB in siRNA-silenced HCT116
cells (si-NC and si-5-LOX-#2 groups). (B) The levels of
EMT-associated proteins were determined by WB in
lentivirus-overexpressed RKO cells (OE-Vector and OE-5-LOX groups).
Values are expressed as the mean ± standard error of the mean
(n=3). ***P<0.001. OE-Vector, cells infected with empty vector
control lentivirus; OE-5-LOX, cells infected with 5-LOX recombinant
lentivirus; siRNA, small interfering RNA; CRC, colorectal cancer;
5-LOX, 5-lipoxygenase; NC, negative control; WB, western blot; EMT,
epithelial-mesenchymal transition.

Figure 7

Cell viability and mRNA expression of
VEGF and angiogenin in HUVECs. The cell viability of HUVECs
incubated with collected media from (A) siRNA-silenced HCT116 cells
(si-NC and si-5-LOX-#2 groups) and (B) lentivirus-overexpressed RKO
cells (OE-Vector and OE-5-LOX groups) for 24, 48 and 72 h. mRNA
expression of VEGF and angiogenin in HUVECs were quantified by
RT-qPCR after 24 h incubation with collected media from (C)
siRNA-silenced HCT116 cells and (D) lentivirus-overexpressed RKO
cells. Values are expressed as the mean ± standard error of the
mean (n=3). *P<0.05, **P<0.01, ***P<0.001. OE-Vector,
cells infected with empty vector control lentivirus; OE-5-LOX,
cells infected with 5-LOX recombinant lentivirus; siRNA, small
interfering RNA; 5-LOX, 5-lipoxygenase; NC, negative control;
HUVEC, human umbilical vein endothelial cell.
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Copy and paste a formatted citation
Spandidos Publications style
Li X, Huge J, Wang X, Zhou J, Li X, Gao X, Shi T and Bai C: Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells. Oncol Lett 31: 221, 2026.
APA
Li, X., Huge, J., Wang, X., Zhou, J., Li, X., Gao, X. ... Bai, C. (2026). Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells. Oncology Letters, 31, 221. https://doi.org/10.3892/ol.2026.15575
MLA
Li, X., Huge, J., Wang, X., Zhou, J., Li, X., Gao, X., Shi, T., Bai, C."Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells". Oncology Letters 31.6 (2026): 221.
Chicago
Li, X., Huge, J., Wang, X., Zhou, J., Li, X., Gao, X., Shi, T., Bai, C."Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells". Oncology Letters 31, no. 6 (2026): 221. https://doi.org/10.3892/ol.2026.15575
Copy and paste a formatted citation
x
Spandidos Publications style
Li X, Huge J, Wang X, Zhou J, Li X, Gao X, Shi T and Bai C: Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells. Oncol Lett 31: 221, 2026.
APA
Li, X., Huge, J., Wang, X., Zhou, J., Li, X., Gao, X. ... Bai, C. (2026). Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells. Oncology Letters, 31, 221. https://doi.org/10.3892/ol.2026.15575
MLA
Li, X., Huge, J., Wang, X., Zhou, J., Li, X., Gao, X., Shi, T., Bai, C."Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells". Oncology Letters 31.6 (2026): 221.
Chicago
Li, X., Huge, J., Wang, X., Zhou, J., Li, X., Gao, X., Shi, T., Bai, C."Overexpression and knockdown of 5‑lipoxygenase regulates the migration and invasion of colorectal cancer cells". Oncology Letters 31, no. 6 (2026): 221. https://doi.org/10.3892/ol.2026.15575
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