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. 2020 Sep 30:12:9351-9362.
doi: 10.2147/CMAR.S263222. eCollection 2020.

CircCFL1/MiR-107 Axis Targeting HMGB1 Promotes the Malignant Progression of Diffuse Large B-Cell Lymphoma Tumors

Xiaowei Chen #  1 , Xiaobin Xie #  2 , Wei Zhou  1
Affiliations

CircCFL1/MiR-107 Axis Targeting HMGB1 Promotes the Malignant Progression of Diffuse Large B-Cell Lymphoma Tumors

Xiaowei Chen et al. Cancer Manag Res. .

Abstract

Objective: The pathogenesis of diffuse large B-cell lymphoma (DLBCL) has not yet been fully elucidated. An increasing number of studies have shown that circular RNAs (circRNAs) play an important role in tumorigenesis and development. The aim of this study was to investigate the effect of CircCFL1 on the malignant progression of DLBCL.

Methods: RT-qPCR was used to detect the expression levels of CircCFL1 and miR-107. A dual-luciferase reporter gene experiment was conducted to verify that CircCFL1 targeted miR-107 and the miR-107 target gene HMGB1. BrdU, transwell, and MTT tests were performed to detect cell invasion and proliferation. Western blot analysis was used to detect the phosphorylation of proteins. Xenograft models were established to confirm the effect of CircCFL1 on DLBCL tumor growth in vivo.

Results: The expression of CircCFL1 in cells transfected with the CircCFL1 overexpression vector was higher than that in the control group. After overexpressing CircCFL1, the expression of miR-107 in cells decreased significantly, and the protein level of HMGB1 increased. The dual-luciferase reporter gene experiment showed that CircCFL1 directly bound to miR-107 and reduced the inhibition of the target gene HMGB1. After CircCFL1 was overexpressed, cell migration and proliferation were enhanced. The tumor volume and weight in the lentivirus CircCFL1 group were higher than those in the lentivirus NC group.

Conclusion: Results showed that the circRNA CircCFL1 could regulate the expression of HMGB1 through miR-107 to promote the proliferation and migration of DLBCL.

Keywords: CircCFL1; HMGB1; circular RNA; lymphoma; migration; proliferation.

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Conflict of interest statement

The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
CircCFL1 promoted the malignant behavior of DLBCL. (A and B) qRT-PCR detection of the expression levels of CircCFL1 in OCI-Ly7 and OCI-Ly3 cell lines. (C and D) Cell proliferation experiment on OCI-Ly7 and OCI-Ly3 after different treatments. CircCFL1 overexpression promoted cell proliferation. (E and F) BrdU assay of the proliferation of OCI-Ly7 and OCI-Ly3 cells. CircCFL1 overexpression promoted cell proliferation. (G and H) Transwell assay experiment on the cell invasion capabilities of OCI-Ly7 and OCI-Ly3 via after different treatments. CircCFL1 overexpression promoted cell invasion. *p < 0.05, **p < 0.01, ***p < 0.001. Magnification: 200×.
Figure 2
Figure 2
Overexpression of CircCFL1 up-regulated the AKT/ERK pathway. (A and B) Detection of proteins related to the AKT/ERK pathway through Western blot analysis. CircCFL1 did not change the total protein expression of AKT, ERK and STAT3. CircCFL1 overexpression promoted the phosphorylation of AKT, ERK, and STAT3. **p < 0.01.
Figure 3
Figure 3
CircCFL1 sponged miR-107. (A) Information on binding sites between CircCFL1 and miR-107. Prediction was performed by using the Starbase (http://starbase.sysu.edu.cn/) website. (B) Double luciferase reporter assay confirmed that CircCFL1 bound to miR-107. (C) MiR-107 expression after CircCFL1 overexpression and knockdown was detected through qPCR. (D) RNA pull-down assay confirmed the binding of CircCFL1 to miR-107. (CircCFL1 as a probe). (E) RNA pull-down assay confirmed the binding of CircCFL1 to miR-107 (miR-107 probe). **p < 0.01, ***p < 0.001.
Figure 4
Figure 4
MiR-107 bound to HMGB1. (A) HMGB1 site information on miR-107 targets. Prediction was performed by using the TargetScan (http://www.targetscan.org/vert_72/) website. (B) Detection of the luciferase activities of CircCFL1-wt and CircCFL1-mut by dual-luciferase reporter gene experiment. (C) RNA pull-down assay was performed to detect whether miR-107 bound to HMGB1. (D) MiR-107 inhibited the expression of HMGB1. (E) HMGB1 expression after CircCFL1 overexpression and knockdown was detected through qPCR. *p < 0.05, **p < 0.01, ***p < 0.001.
Figure 5
Figure 5
Overexpression of miR-107 and knockdown of HMGB1 reversed the promotion of DLBCL by CircCFL1. (A) Detection of HMGB1 expression levels in the OCI-Ly7 cell line. (B) Cell proliferation experiment on OCI-Ly7 after different treatments. (C and D) BrdU assay was performed to detect the proliferation of OCI-Ly7. CircCFL1 overexpression promoted cell proliferation. (E and F) Cell invasion experiment on OCI-Ly7 after different treatments. CircCFL1 overexpression promoted cell invasion. **p < 0.01. Magnification: 200×.
Figure 6
Figure 6
Overexpression of miR-107 and knockdown of HMGB1 reversed the effect of CircCFL1 on the AKT/ERK pathway. (A–G) Detection of proteins related to the AKT/ERK pathway via Western blot analysis. CircCFL1 overexpression promoted the phosphorylation of AKT, ERK, and STAT3. Overexpression of miR-107 and knockdown of HMGB1 reversed the effect of CircCFL1. **p < 0.01.
Figure 7
Figure 7
Overexpression of CircCFL1 promoted the tumor growth of DLBCL. (A) Detection of tumor volume after the injection of OCI-Ly7. Overexpression of CircCFL1 promoted cell proliferation. (B) Detection of tumor weight after the injection of OCI-Ly7. (C) Expression levels of CircCFL1 and miR-107 were detected via RT-qPCR. *p < 0.05, **p < 0.01.
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