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. 2019 Mar 11;17(1):76.
doi: 10.1186/s12967-019-1830-6.

Circular RNA hsa_circ_0055538 regulates the malignant biological behavior of oral squamous cell carcinoma through the p53/Bcl-2/caspase signaling pathway

Wen Su  1   2 , Shuai Sun  2 , Feng Wang  2 , Yuehong Shen  2 , Hongyu Yang  3   4
Affiliations

Circular RNA hsa_circ_0055538 regulates the malignant biological behavior of oral squamous cell carcinoma through the p53/Bcl-2/caspase signaling pathway

Wen Su et al. J Transl Med. .

Retraction in

Abstract

Background: Oral squamous cell carcinoma (OSCC) is a common oral and maxillofacial malignant tumor with high rates of metastasis and mortality. Circular RNAs (circRNAs), a type of non-coding RNA, are involved in the development of a variety of tumors. The roles of circRNAs in OSCC are unclear; in this study, the correlation between the circRNA hsa_circ_0055538, previously identified by high-throughput sequencing, and the biological behavior of OSCC was evaluated VSports手机版. .

Methods: circRNA expression was evaluated using patient tissue samples and various OSCC cell lines. The effects of overexpression and knockdown were evaluated by lentiviral infection and siRNA transfection of the SCC9 and CAL27 cell lines. Migration, invasion, apoptosis, and the expression of proteins in the p53 signaling pathway were evaluated. Infected cells were injected into nude mice to evaluate tumorigenesis. V体育安卓版.

Results: Low hsa_circ_0055538 expression levels were verified in tumor tissues and OSCC cell lines. Clinical data analysis showed that the expression level is related to the degree of tumor differentiation. Lentiviral infection and siRNA transfection of SCC9 and CAL27 cell lines revealed that changes in circRNA expression significantly affected the malignant biological behavior of OSCC cells. Importantly, nude mouse experiments showed that high expression of hsa_circ_0055538 inhibited tumor growth. Finally, hsa_circ_0055538 may affect the development of OSCC via the p53/Bcl-2/caspase signaling pathway V体育ios版. .

Conclusions: Our results indicated that hsa_circ_0055538 is involved in OSCC via the p53 signaling pathway and may be a diagnostic and/or prognostic marker as well as a therapeutic target. VSports最新版本.

Keywords: Circular RNA; Oral tumor; Squamous cell carcinoma; p53/Bcl-2/caspase signaling pathway. V体育平台登录.

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Figures

Fig. 1
Fig. 1
hsa_circ_0055538 is downregulated in OSCC tissue specimens and cell lines. a, b qRT-PCR was used to determine the hsa_circ_0055538 expression level in a tumor tissue specimens from patients with OSCC, relative to that in adjacent normal tissues (n = 44); and b HOK, CAL27, SCC9, SCC15, and SCC25 cell lines. c, d The hsa_circ_0055538 expression level in stable SCC9 (c) and CAL27 (d) cell lines was quantified by qRT-PCR. GFP expression was examined under a fluorescence microscope using stable SCC9 and CAL27 cell lines. BF bright field. e, f qRT-PCR quantification of hsa_circ_0055538 levels in SCC9 (e) and CAL27 (f) cells transfected with hsa_circ_0055538 siRNA. Si and NC refer to OSCC cells transfected with hsa_circ_0055538 siRNA or normal controls. Data are presented as mean ± SEM of three independent experiments. Student’s t test, ***P < 0.001
Fig. 2
Fig. 2
High expression of hsa_circ_0055538 inhibits OSCC cell proliferation. a, b SCC9 (a) and CAL27 (b) cells were infected with an empty vector control (control) or lentivirus harboring hsa_circ_0055538 (test), and the CCK-8 assay was used to measure cell proliferation at different time points after infection. c, d The EdU incorporation assay was used to measure proliferation in control SCC9 (c) and CAL27 (d) cells and those overexpressing hsa_circ_0055538. Data are presented as mean ± SEM of three independent experiments. Student’s t-test, ***P < 0.001, **P < 0.01, *P < 0.05, Scale bar, 20 µm
Fig. 3
Fig. 3
High expression of hsa_circ_0055538 promotes apoptosis in OSCC cells. a, b Annexin V-FITC/PI dual-label flow cytometry was performed to determine the apoptotic rate in SCC9 (a) and CAL27 (b) cells. Data are presented as mean ± SEM of three independent experiments. Student’s t-test, ***P < 0.001
Fig. 4
Fig. 4
High expression of hsa_circ_0055538 inhibits OSCC cell migration and invasion. a, b Wound healing assays were performed using a SCC9 and b CAL27 cells infected with the empty vector control or lentivirus harboring hsa_circ_0055538. The scratch area was measured at 0 and 24 h, and the percentage of closure at 24 h was calculated. c, d Transwell assays were performed to quantify the migration and invasion ability of SCC9 and CAL27 cells infected with the empty vector control or lentivirus harboring hsa_circ_0055538. c Cells were seeded into the upper chamber (uncoated); after 24 h, those that crossed to the lower chamber were used for imaging and quantification: SCC9 (top), CAL27 (bottom). The proportion of migrating cells was quantified using SCC9 (left) and CAL27 (right) cells. d Cells were seeded into the Matrigel-coated upper chamber; after 48 h, those that passed across the coated chamber were used for imaging and quantification: SCC9 (top), CAL27 (bottom). The proportion of invading cells was quantified in SCC9 (left) and CAL27 (right) cells. Data are presented as mean ± SEM of three independent experiments. Student’s t-test, ***P < 0.001, **P < 0.01
Fig. 5
Fig. 5
Expression of hsa_circ_0055538 affects the levels of key proteins involved in the p53/Bcl-2/caspase signaling pathway. a, b SCC9 and CAL27 cells were subjected to either the overexpression a or knockdown b of hsa_circ_0055538. Cell extracts were used for immunoblotting, and the levels of key proteins related to the p53/Bcl-2/caspase signaling pathway were detected by western blotting. c, d SCC9 (left) and CAL27 (right) cells were subjected to either the overexpression c or knockdown d of hsa_circ_0055538. The mRNA levels of the key genes related to the p53/Bcl-2/caspase signaling pathway (corresponding to proteins in a and b) in SCC9 and CAL27 cells were detected by qRT-PCR. e, f SCC9 (e) and CAL27 (f) cells overexpressing p53 after knockdown of hsa_circ_0055538 (Si + p53 AT) were subjected to CCK-8 assay to measure cell proliferation at different time points. The mRNA level of the RMND5A was detected by qRT-PCR. Data are presented as mean ± SEM of three independent experiments. Student’s t-test, ***P < 0.001, **P < 0.01
Fig. 6
Fig. 6
Expression of hsa_circ_0055538 affects the levels of key proteins involved in the p53/Bcl-2/caspase signaling pathway. a, b Wound healing assays were performed using a SCC9 and b CAL27 cells overexpressing p53 after knockdown of hsa_circ_0055538. The scratch area was measured at 0 and 24 h, and the percentage of closure at 24 h was calculated. c Invasion assays were performed using SCC9 (top) and CAL27 (bottom) cells overexpressing p53 after knockdown of hsa_circ_0055538. Cells were seeded into the Matrigel-coated upper chamber; after 48 h, those that passed across the coated chamber were imaged and quantified. The proportion of invading cells was quantified in SCC9 (left) and CAL27 (right) cells. d SCC9 and CAL27 cells were subjected to either the overexpression (left) or knockdown (right) of hsa_circ_0055538. The mRNA level of the RMND5A was detected by qRT-PCR. Data are presented as mean ± SEM of three independent experiments. Student’s t-test, ***P < 0.001, **P < 0.01
Fig. 7
Fig. 7
High expression of hsa_circ_0055538 inhibits tumorigenesis. a Tumors obtained in nude mice. b Tumor growth curve showing changes in tumor volume following injection. c Tumor weight plots of the empty vector control (control) and hsa_circ_0055538 overexpression groups (test). d P53 signaling pathway associated protein levels in nude mouse tumor specimens were detected by western blotting. e HE staining of tumor specimens in test group and control group
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