Open Access

lncRNA ROR promotes the progression of renal cell carcinoma through the miR‑206/VEGF axis

  • Authors:
    • Jianguo Shi
    • Datian Zhang
    • Zhenhai Zhong
    • Wen Zhang
  • View Affiliations

  • Published online on: September 2, 2019     https://doi.org/10.3892/mmr.2019.10636
  • Pages: 3782-3792
  • Copyright: © Shi et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Renal cell carcinoma (RCC) is the most common kidney malignancy, responsible for ~80% of all cases in adults. The pathogenesis of RCC is complex, involving alterations at both the genetic and epigenetic levels. Numerous signaling pathways, such as PI3K/Akt/mTOR and Wnt‑β‑catenin have been demonstrated to be associated with the tumorigenesis and development of RCC. Long non‑coding RNAs (lncRNAs) are functional RNA molecules involved in the initiation and progression of cancer, and investigating the effects of lncRNA could facilitate the development of novel treatments. The lncRNA regulator of reprogramming (ROR) is aberrantly expressed in a variety of tumors. However, its underlying mechanisms remain largely unknown. In the present study, ROR was found to be upregulated and microRNA (miR)‑206 was found to be downregulated in RCC tissues and cells. Furthermore, the knockdown of ROR inhibited the proliferation, migration and invasion of RCC cells. It was found that ROR binds to miR‑206, and that ROR‑induced cell proliferation and metastasis were reversed by the overexpression of miR‑206. In addition, the levels of miR‑206 and ROR were negatively correlated in RCC tissues. Furthermore, the overexpression of miR‑206 notably suppressed the proliferation, migration and invasion of RCC cells, and these effects were enhanced by the knockdown of vascular endothelial growth factor (VEGF); cell growth and metastasis induced by miR‑206 inhibitors could be reversed by the knockdown of VEGF. In addition, the expression levels of miR‑206 and VEGF were inversely correlated in RCC samples. In summary, the results of the present study revealed that ROR was upregulated in RCC tissues, which promoted tumor progression by regulating the miR‑206/VEGF axis. The present findings provided a novel insight into the potential functions of ROR in RCC, and the ROR/miR‑206/VEGF pathway may be a promising therapeutic target for the treatment of patients with RCC.

Introduction

Renal cell carcinoma (RCC) is the most prevalent kidney cancer in adults, accounting for ~80% of patients with kidney cancer in European countries between 2006 and 2011 (1). Although multimodal approaches for diagnosis have been previously developed, such as ultrasound and computed tomography technologies, it remains difficult to distinguish between benign and malignant tumors (2). Therefore, it is important to discover novel non-invasive diagnostic and prognostic biomarkers for RCC. In 2015, there was a ~66.8% increase in the number of new cases, and ~23.4% increase in mortality, of renal cancer reported in China (3). The most common subtype of RCC is clear cell RCC, followed by papillary RCC and chromophobe RCC (4). The pathogenesis of RCC is complex, and previous studies have reported that alterations at both the genetic and epigenetic levels contribute to the development of RCC (59); however, the mechanisms underlying the initiation and progression of RCC remain largely unknown.

Long non-coding RNAs (lncRNAs) are a type of RNAs that are >200 nucleotides in length (10). lncRNAs are associated with numerous biological functions, including the regulation of cell proliferation and gene expression (1013). Accumulating evidence has revealed that lncRNAs are associated with the initiation and development of numerous types of cancer, and they may function as oncogenes or tumor suppressors (1416). In addition, the impaired expression of lncRNAs has been detected in tumor cells, suggesting an important role of lncRNAs in carcinogenesis (1621). Aberrant levels of lncRNAs have been reported in RCC (2225). Therefore, investigating the effects of misregulated lncRNAs in RCC may facilitate the development of novel therapies.

The lncRNA regulator of reprogramming (ROR) is involved in carcinogenesis, and previous studies have suggested the role of ROR in cancer (26,27). ROR has been reported to regulate the initiation and progression of tumors through various signaling pathways, such as RAD18 and SOX9 (28,29). Furthermore, our previous study revealed that ROR is a promising biomarker for RCC (30); however, the molecular targets of ROR require further investigation.

MicroRNAs (miRNAs/miRs) are non-coding RNAs and are ~22 nucleotides in length, and are potential downstream targets of lncRNAs (31). Emerging evidence has revealed that the expression of miRNAs is misregulated in cancer, which consequently initiates tumorigenesis (32,33). Additionally, miRNAs, such as miR-122-5p and miR-206, are novel biomarkers for patients with RCC (34); however, the functions of miRNAs in RCC remain unclear.

Vascular endothelial growth factor (VEGF) is a soluble ligand secreted by cells that stimulates the formation of blood vessels, and it is a potent pro-angiogenic factor involved in wound healing and pathogenic processes, including carcinogenesis (35). In the present study, the function of the ROR-mediated miR-206/VEGF signaling pathway in RCC cell growth and metastasis was investigated, which may provide novel insights into the treatment of patients with RCC.

Materials and methods

Clinical samples

A total of 36 paired RCC and para-carcinoma tissues were collected from patients (16 male and 20 female, average age 46±12) who underwent radical nephrectomy in the Department of Urology (The First Affiliated Hospital of Jinzhou Medical University) between June 2014 and July 2015. None of the patients recruited in the present study had received any other treatments prior to surgery. The present study was approved by the Ethics Committee of The First Affiliated Hospital of Jinzhou Medical University. Written informed consent was obtained from each patient. The protocols were approved by the Institutional Review Board of The First Affiliated Hospital of Jinzhou Medical University. All kidney tissues samples were immediately snap-frozen using liquid nitrogen and stored at −80°C until further use.

Cell culture

The human RCC cell lines Caki-1 and Caki-2, and normal human kidney cells HK-2, were purchased from the American Type Culture Collection. Cells were cultured in DMEM containing 10% FBS, 100 U/ml penicillin and 100 µg/ml streptomycin (all from Gibco; Thermo Fisher Scientific, Inc.). Cells were incubated at 37°C in a humidified incubator containing 5% CO2.

Cell transfection

Short hairpin (sh)RNA sequences targeting ROR (sh-ROR), VEGF (sh-VEGF), negative control (sh-NC), miR-206 mimic/inhibitor and miRNA control (miR-NC) were synthesized by Guangzhou RiboBio Co., Ltd. The sequences were: sh-ROR: 5′-GCCTCTGCACTCTTATGGAAGGAGGAAAT-3′; sh-VEGF 5′-GGTGAGAAACCCATTGTTCAGTTCCCTAA-3′; sh-NC: 5′-CGAGGACCGCCTGTCCTGCTTCGCGCAGA-3′. Following annealing, shRNA were integrated into the lentiviral pU6-Luc-Puro vector using Xbal and BamHI restriction sites (Shanghai Genepharma Co. Ltd.). To establish the ROR overexpression model, wild-type (o/e-ROR) or mutant (o/e-NC) ROR fragments were amplified by PCR using Multiplex PCR kit (Qiagen, Germany) according to the manufacture's protocols. The following thermocycling conditions were used: Initial denaturation at 95°C for 30 sec followed by 30 cycles of 95°C for 15 sec, 60°C for 20 sec and 68°C for 1 min. The PCR products were then subcloned into the Nsil/BgIII restriction sites of pcDNA3.1 vector (Invitrogen; Thermo Fisher Scientific, Inc.). 8×105 of Cells were seeded into 6-well plates and cultured in DMEM without antibiotics. Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.) was used for transfection when the cell density reached 60–70%, according to the manufacturer's protocols. A total of 50 pg/µl plasmid was used for each transfection. At 8 h post-transfection, the culture medium was replenished with fresh DMEM containing 10% FBS.

Reverse transcription-quantitative (RT-q)PCR

RT-qPCR was used to evaluate the expression levels of ROR, miR-206 and VEGF. The miRNeasy Mini Kit (Qiagen GmbH) was used for the extraction of miRNAs and total RNA from tissues or cells was extracted using TRIzol reagent (Invitrogen; Thermo Fisher Scientific, Inc.), according to the manufacturer's protocols. The concentration of the RNA extracted was determined using a NanoDrop 1000 spectrophotometer (Thermo Fisher Scientific, Inc.). First-strand complementary DNA was synthesized from total RNA using a PrimeScript™ RT kit (Takara Bio, Inc.) and qPCR was performed using SYBR Green PCR Master Mix (Takara Bio, Inc.), according to the manufacturer's protocols. The reverse transcription reaction was performed using 1 µg RNA diluted in 1 µl nuclease free water, 2 µl first strand buffer, 4 µl MgCl2 solution, 1 µl random primers, 8 µl dNTPs, 1 µl RNase inhibitor and 1 µl reverse transcriptase. The sample was incubated at room temperature for 30 min. After that, 1 cycle of PCR was performed at 42°C for 45 min, 99°C for 5 min and 5°C for 5 min in a PCR cycler. The TaqMan MicroRNA Assay (Applied Biosystems; Thermo Fisher Scientific, Inc.) was performed to evaluate the expression level of miR-206, followed by qPCR using the Applied Biosystem 7500 (Applied Biosystems; Thermo Fisher Scientific, Inc.). U6 small nuclear RNA was used as an internal control for miRNA. The relative expression of mRNA was calculated and normalized to the endogenous expression level of GAPDH. The forward and reverse primer sequences are as follows: ROR forward, 5′-TCCAAACACATCGCCACTCT-3′ and reverse, 5′-TCCTAGGCCATGAGGAGTCA-3′; VEGF forward, 5′-CGAAGTGGTGAAGTTCATGGATG-3′ and reverse, 5′-TTCTGTATCAGTCTTTCCTGGT-3′; GAPDH forward, 5′-GCAAGAGCACAAGAGGAAGA-3′ and reverse, 5′-ACTGTGAGGAGGGGAGATTC-3′; and U6 forward, 5′-CTCGCTTCGGCAGCACATA-3′ and reverse, 5′-AACGATTCACGAATTTGCGT-3. The following thermocycling conditions were used for the qPCR: mRNA; initial denaturation at 95°C for 5 min followed by 45 cycles of 95°C for 15 sec, 60°C for 20 sec and 72°C for 10 sec; miRNA; initial denaturation at 95°C for 30 sec followed by 40 cycles of 95°C for 5 sec, 60°C for 30 sec and 72°C for 10 sec. The data was analyzed using the 2−∆∆Cq method (36).

Western blot analysis

Total protein from tissues or cells was extracted using RIPA buffer (Beyotime Institute of Biotechnology). The protein concentration was evaluated using the bicinchoninic acid method. Equal amounts (50 µg) of protein samples were loaded on to 10% SDS-PAGE gels and transferred onto PVDF membranes. Subsequently, the membranes were blocked in TBST containing 5% skimmed milk for 2 h at room temperature. The membranes were then incubated with primary antibodies against VEGF (1:500; cat. no. MA5-13182; Invitrogen; Thermo Fisher Scientific, Inc.) or GAPDH (1:1,000; cat. no. sc-47724; Santa Cruz Biotechnology, Inc.) at 4°C overnight. Following three washes with TBST, the membranes were incubated with horseradish peroxidase-conjugated secondary antibody (1:5,000; cat. no. sc-2371; Santa Cruz Biotechnology, Inc.) for 1 h at 37°C. Protein bands were visualised using an ECL detection kit (Pierce; Thermo Fisher Scientific, Inc) and quantified by densitometric analysis using ImageJ 1.49 software (National Institutes of Health).

Cell proliferation assays

Transfected cells were harvested 24 h post-transfection and seeded into 96-well plates at a concentration of 5,000 cells/well. Cells were then incubated at 37°C and cell proliferation was determined using the Cell Counting Kit-8 (CCK-8) assay (Dojindo Molecular Technologies, Inc.) at day 1, 2, 3 and 4 according to the manufacturer's protocols. Briefly, 10 µl of CCK-8 solution was added into each well at the indicated time points. Following incubation at 37°C for a further 2 h, the absorbance at 450 nm was measured using a microplate reader (Bio-Rad Laboratories, Inc.).

Transwell assay

The migration and invasion of cells was evaluated using a Transwell assay. For the migration assay, a total of 2×105 cells in FBS-free DMEM were seeded into the upper chamber (BD Biosciences) with an 8 µm pore size. For the invasion assay, cells were inoculated onto a Matrigel-pre-coated (room temperature for 1 h) upper chamber (Sigma-Aldrich; Merck KGaA). Subsequently, 500 µl of culture medium supplemented with 10% FBS (Gibco; Thermo Fisher Scientific, Inc.) was added into the lower chamber. Following overnight incubation at 37°C, cells that had not migrated/invaded were removed using a cotton swab, while the migrated/invaded cells in the lower chamber were fixed with 4% paraformaldehyde at room temperature for 10 mins and stained using 0.5% crystal violet at room temperature for 20 mins. The numbers of migratory/invasive cells were counted in five randomly selected fields using an inverted light microscope (magnification, ×200; Olympus Corporation).

Bioinformatic prediction and luciferase reporter assay

TargetScan 6.2 (www.targetscan.org/) and miRanda 0.10.x (www.microrna.org/microrna/) were employed to predict the potential targets of ROR and miR-206. Wild-type (WT) fragments of the 3′ untranslated region (UTR) of ROR and VEGF containing the potential binding sites of miR-206 were synthesized by Shanghai GenePharma Co., Ltd. and were cloned into pmirGLO Dual-Luciferase miRNA Target Expression Vector using Xhol and Xbal restriction sites (Promega Corporation), according to the manufacturer's protocols. QuikChange Multi Site-Directed Mutagenesis Kit (Stratagene; Agilent) was used to generate the ROR/VEGF-3′UTR-MUT reporter containing mutant miR-206 binding sites. The luciferase vectors were co-transfected with miR-NC or miR-206 mimics/inhibitors (50 pg/µl) into DH5α competent cells using Lipofectamine® 2000 (Thermo Fisher Scientific, Inc.). Luciferase activity was assessed at 48 h post-transfection using a Dual Luciferase Reporter Assay System (Promega Corporation), according to the manufacturer's protocols. The level of firefly luciferase activity was normalized to that of Renilla luciferase.

Statistical analysis

SPSS 17.0 software (SPSS, Inc.) was used for statistical analysis. All experiments were repeated a minimum of three times. Data are presented as the mean ± SD and were analyzed using a Student's t-test or ANOVA. A Student-Newman-Keuls test was performed as a post-hoc test following ANOVA. The association between RNA levels was evaluated using Spearman's correlation analysis. P<0.05 was considered to indicate a statistically significant difference.

Results

ROR is upregulated and miR-206 is downregulated in RCC tissues and cells

The upregulation of ROR was detected in RCC compared with the adjacent normal tissues, which may be associated with poorer prognosis as described in our previous study (33). In addition, the expression of ROR was increased in RCC cells compared with non-RCC cell lines (33). In the present study, the expression level of miR-206 RNA in 36 paired RCC and para-carcinoma samples was determined using RT-qPCR. The present results indicated that miR-206 was significantly downregulated in glioma tissues compared with the control (Fig. 1A). Furthermore, miR-206 RNA was significantly decreased in aggressive RCC, suggesting that downregulation of miR-206 is associated with the development of this disease (Fig. 1B). In addition, the expression levels of ROR and miR-206 were found to be negatively correlated in RCC tissues (Fig. 1C). miR-206 was significantly downregulated in RCC cell lines in comparison with HK-2 cells (Fig. 1D). The present results suggested that the expression levels of ROR and miR-206 were upregulated and downregulated in RCC, respectively, which may be associated with the progression of this disease.

Downregulation of ROR suppresses the proliferation, migration and invasion of RCC cells

To explore the effects of ROR on the proliferation, invasion and migration of RCC cells, the expression of ROR was decreased in Caki-1 and Caki-2 cells. The transfection efficiency was determined using RT-qPCR (Fig. 2A). The results of the CCK-8 assay indicated that the proliferative ability of Caki-1 and Caki-2 cells transfected with sh-ROR was reduced compared with the control (Fig. 2B and C). In addition, Transwell assays indicated that the migration and invasion of sh-ROR-transfected cells was significantly reduced (Fig. 2D-G). These results suggested that the knockdown of ROR inhibited the proliferation, migration and invasion of RCC and may be involved in the development and progression of RCC.

miR-206 is a potential target gene of ROR in RCC cells

To determine whether ROR exerts its functions in RCC by suppressing target miRNAs, the potential binding sites of miR-206 in ROR transcripts were predicted using miRanda (Fig. 3A). Luciferase reporter vectors containing WT (WT-ROR) and mutant ROR (MUT-ROR) sequences of the predicted miR-206 binding sites were constructed. The results revealed that the overexpression of miR-206 significantly attenuated the activity of the luciferase plasmid carrying the WT binding sites, which was not observed in the MUT control (Fig. 3B). In order to further investigate the influence of ROR on the expression of miR-206, Caki-1 and Caki-2 cells were transfected with sh-ROR. Cells transfected with sh-ROR exhibited significantly increased miR-206 expression, which was also detected in cells transfected with the miR-206 mimic (Fig. 3C and D), suggesting that miR-206 may be a novel target of ROR in RCC.

Overexpression of ROR promotes cell proliferation, migration and invasion by regulating miR-206

To investigate whether ROR suppresses the proliferation and metastasis of RCC cells by targeting miR-206, Caki-1 and Caki-2 cells were transfected with o/e-NC, o/e-ROR or co-transfected with o/e-ROR and the miR-206 mimic. The expression of ROR was significantly increased (Fig. 4A) and the level of miR-206 was decreased (Fig. 4B) in Caki-1 and Caki-2 cells transfected with o/e-ROR. Additionally, the overexpression of ROR promoted the proliferation (Fig. 4C and D), migration (Fig. 4E) and invasion (Fig. 4F) of Caki-1 and Caki-2 cells, whereas these effects were significantly reversed by the miR-206 mimic. These results suggested that ROR induced the proliferation, migration and invasion of RCC cells by downregulating miR-206.

VEGF is a target gene of miR-206 in RCC cells

Using the TargetScan database, the complementary sequence between VEGF and miR-206 was identified (Fig. 5A). To investigate whether VEGF was a potential target of miR-206, WT and MUT fragments of VEGF were cloned downstream of the firefly luciferase coding domain. The present results indicated that the overexpression of miR-206 significantly reduced the luciferase activity of the VEGF-WT reporter but not of the VEGF-MUT control (Fig. 5B). To further determine whether miR-206 regulates the expression of VEGF, Caki-1 and Caki-2 cells were transfected with the miR-206 inhibitor. The transfection efficiency was determined by evaluating the level of miR-206 (Fig. 5C). The protein level of VEGF was elevated in cells transfected with the miR-206 inhibitor (Fig. 5D). Furthermore, VEGF was upregulated in RCC tissues compared with the paired para-carcinoma controls (Fig. 5E) and VEGF expression was found to be inversely correlated with miR-206 in RCC samples (Fig. 5F), further suggesting that VEGF may be a target of miR-206 in RCC.

Downregulation of VEGF enhances the effects of miR-206 overexpression and reverses the effects of miR-206 inhibition in RCC cells

To investigate whether the effect of VEGF on the growth and metastasis of RCC cells was regulated by miR-206, Caki-1 and Caki-2 cells were transfected with miR-NC, miR-206 mimic/inhibitor or co-transfected with miR-206 mimic/inhibitor and sh-VEGF. The transfection efficiency of sh-VEGF was determined using RT-qPCR (Fig. 6A). The results revealed that the overexpression of miR-206 suppressed the proliferation (Fig. 6B and C), migration (Fig. 6F) and invasion (Fig. 6H) of Caki-1 and Caki-2 cells, while these effects were increased by the depletion of VEGF. Additionally, the downregulation of miR-206 promoted the proliferation (Fig. 6D and E), migration (Fig. 6G) and invasion (Fig. 6I) of RCC cells, whereas these effects were abrogated following knockdown of VEGF. The present results suggested that miR-206 inhibits the growth of RCC cells by downregulating VEGF. In summary, ROR may regulate the proliferation, migration and invasion of RCC cells via the miR-206/VEGF signaling pathway.

Discussion

lncRNAs are a group of non-coding RNAs of >200 nucleotides in length. Previous studies have revealed the significance of lncRNAs, and accumulating evidence demonstrated that lncRNAs are important regulators of the growth and metastasis of cancer cells (1417). lncRNAs act as oncogenes or suppressing factors in cancer; the dysregulation of lncRNAs is associated with the progression of numerous types of cancer including glioblastoma and astrocytoma (10,1821). The upregulation of long intergenic non-protein coding RNA 01116 was reported to be associated with the overall survival of patients with cancer and metastasis (37). Furthermore, the downregulation of lncRNA-small nucleolar host gene 5 inhibited the proliferation and migration of gastric cancer cells through the miR-32/KLF4 axis (38). The expression level of prostate cancer upregulated lncRNA-1 (PlncRNA-1) was found to be decreased in tumor tissues and the induced expression of PlncRNA-1 was reported to suppress the proliferation and promote the apoptosis of breast cancer cells through the transforming growth factor-β1/D-3-phosphoglycerate dehydrogenase signaling pathway (39). A number of lncRNAs regulate gene expression by interacting with their target miRNAs. For example, the lncRNA H19 imprinted maternally expressed transcript was reported to modulate the proliferation, migration and invasion of gastric cancer cells through downstream miRNAs (40,41). Additionally, the lncRNA BC032469 was found to bind miR-1207-5p and human telomerase reverse transcriptase, inducing the proliferation of cancer cells (42). However, the roles of lncRNAs and their underlying mechanisms in cancer remain largely unknown and require further investigation.

Previous studies have revealed the impaired expression of ROR in prostate and breast cancer (26,27). Furthermore, ROR is involved in the initiation and progression of tumor by regulating numerous signaling pathways, such as RAD18 and SOX9 (28,29). The results of the present study indicated that ROR was significantly upregulated, while miR-206 was downregulated, in RCC tissues, which may be associated with poor prognosis. In addition, the present study suggested a negative correlation between the levels of ROR and miR-206, and miR-206 and VEGF in RCC samples. Therefore, further experiments were conducted to explore the downstream targets of ROR in RCC.

The results of the present study indicated that the knockdown of ROR inhibited the proliferation, migration and invasion of RCC cells. Furthermore, the overexpression of ROR induced the proliferation, migration and invasion of RCC cells, whereas these effects were reversed by the overexpression of miR-206, suggesting that ROR promotes RCC cell growth and metastasis in an miR-206-dependent manner. miRNAs may function as oncogenes or tumor suppressors and are potential targets of lncRNAs (31,32). Consistent with the finding of the present study, previous studies reported impaired levels of miRNAs in various cancer types, including RCC (3234). Furthermore, it was reported that ROR was able to induce the development of osteosarcoma by regulating miR-206 (43).

In addition, luciferase reporter assay revealed that VEGF was a potential target of miR-206, and that the upregulation of miR-206 suppressed RCC cell proliferation, migration and invasion by targeting VEGF. Conversely, the downregulation of miR-206 induced the proliferation, invasion and migration of RCC cells, whereas these effects were abrogated following the depletion of VEGF. Furthermore, VEGF expression was found to be significantly upregulated in RCC tissues compared with the matched non-tumor controls, and was negatively correlated with the level of miR-206. VEGF is a signal protein produced by cells that stimulates the formation of blood vessels and is a potent proangiogenic factor involved in wound healing, and pathogenic processes, including carcinogenesis (35). A previous study reported that the overexpression of VEGF was associated with poor survival for patients with squamous cell carcinoma (44). Furthermore, the downregulation of miR-206 induced the development of breast and laryngeal cancer through the VEGF pathway (45,46). Consistent with these findings, the present study indicated that ROR was upregulated in RCC, which may promote the development of tumors via the miR-206/VEGF signaling pathway. However, there were some limitations to the present study, for example, markers of proliferation and apoptosis were not examined; such markers should be investigated in future studies to support the findings of the present study.

In conclusion, the present study indicated that ROR was a potential oncogene, which could increase the level of VEGF and induce RCC cell proliferation and migration through miR-206. The findings of the present study indicated the important roles of ROR and its underlying mechanisms in the proliferation, migration and invasion of RCC cells. The present study suggested that the ROR/miR-206/VEGF signaling pathway may be a novel therapeutic target for the treatment of patients with RCC.

Acknowledgements

Not applicable.

Funding

The present study was funded by the Natural Science Project of Liaoning Province (grant no. 20180530058).

Availability of data and materials

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.

Authors' contributions

JS initiated and designed the present study. DZ, ZZ and WZ performed the experiments and interpreted the results. All authors read and approved the final manuscript.

Ethics approval and consent to participate

The present study was approved by the Ethics Committee of The First Affiliated Hospital of Jinzhou Medical University. Written informed consent was obtained from each patient for the use of clinical tissues.

Patient consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

References

1 

Dabestani S, Beisland C, Stewart GD, Bensalah K, Gudmundsson E, Lam TB, Gietzmann W, Zakikhani P, Marconi L, Fernandéz-Pello S, et al: Intensive imaging-based follow-up of surgically treated localised renal cell carcinoma does not improve post-recurrence survival: Results from a European multicentre database (RECUR). Eur Urol. 75:261–264. 2019. View Article : Google Scholar : PubMed/NCBI

2 

Sheth S, Scatarige JC, Horton KM, Corl FM and Fishman EK: Current concepts in the diagnosis and management of renal cell carcinoma: Role of multidetector ct and three-dimensional CT. Radiographics. 21:S237–S254. 2001. View Article : Google Scholar : PubMed/NCBI

3 

Chen W, Zheng R, Baade PD, Zhang S, Zeng H, Bray F, Jemal A, Yu XQ and He J: Cancer statistics in China, 2015. CA Cancer J Clin. 66:115–132. 2016. View Article : Google Scholar : PubMed/NCBI

4 

Vera-Badillo FE, Conde E and Duran I: Chromophobe renal cell carcinoma: A review of an uncommon entity. Int J Urol. 19:894–900. 2012. View Article : Google Scholar : PubMed/NCBI

5 

Di Cristofano C, Minervini A, Menicagli M, Salinitri G, Bertacca G, Pefanis G, Masieri L, Lessi F, Collecchi P, Minervini R, et al: Nuclear expression of hypoxia-inducible factor-1alpha in clear cell renal cell carcinoma is involved in tumor progression. Am J Surg Pathol. 31:1875–1881. 2007. View Article : Google Scholar : PubMed/NCBI

6 

Elfiky AA, Aziz SA, Conrad PJ, Siddiqui S, Hackl W, Maira M, Robert CL and Kluger HM: Characterization and targeting of phosphatidylinositol-3 kinase (PI3K) and mammalian target of rapamycin (mTOR) in renal cell cancer. J Transl Med. 9:1332011. View Article : Google Scholar : PubMed/NCBI

7 

Shang D, Liu Y, Ito N, Kamoto T and Ogawa O: Defective Jak- Stat activation in renal cell carcinoma is associated with interferon-alpha resistance. Cancer Sci. 98:1259–1264. 2007. View Article : Google Scholar : PubMed/NCBI

8 

Chappell WH, Steelman LS, Long JM, Kempf RC, Abrams SL, Franklin RA, Bäsecke J, Stivala F, Donia M, Fagone P, et al: Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR inhibitors: Rationale and importance to inhibiting these pathways in human health. Oncotarget. 2:135–164. 2011. View Article : Google Scholar : PubMed/NCBI

9 

Li M, Wang Y, Song Y, Bu R, Yin B, Fei X, Guo Q and Wu B: Expression profiling and clinicopathological significance of DNA methyltransferase 1, 3A and 3B in sporadic human renal cell carcinoma. Int J Clin Exp Pathol. 7:7597–7609. 2014.PubMed/NCBI

10 

Gibb EA, Brown CJ and Lam WL: The functional role of long non-coding RNA in human carcinomas. Mol Cancer. 10:382011. View Article : Google Scholar : PubMed/NCBI

11 

Hsiao J, Yuan TY, Tsai MS, Lu CY, Lin YC, Lee ML, Lin SW, Chang FC, Liu Pimentel H, Olive C, et al: Upregulation of haploinsufficient gene expression in the brain by targeting a long non-coding RNA improves seizure phenotype in a model of Dravet syndrome. EBioMedicine. 9:257–277. 2016. View Article : Google Scholar : PubMed/NCBI

12 

Wang P, Ren Z and Sun P: Overexpression of the long non-coding RNA MEG3 impairs in vitro glioma cell proliferation. J Cell Biochem. 113:1868–1874. 2012. View Article : Google Scholar : PubMed/NCBI

13 

Mills JD, Chen J, Kim WS, Waters PD, Prabowo AS, Aronica E, Halliday GM and Janitz M: Long intervening non-coding RNA 00320 is human brain-specific and highly expressed in the cortical white matter. Neurogenetics. 16:201–213. 2015. View Article : Google Scholar : PubMed/NCBI

14 

Li Z, Dong M, Fan D, Hou P, Li H, Liu L, Lin C, Liu J, Su L, Wu L, et al: LncRNA ANCR down-regulation promotes TGF-β-induced EMT and metastasis in breast cancer. Oncotarget. 8:67329–67343. 2017.PubMed/NCBI

15 

Zhang X, Sun S, Pu JK, Tsang AC, Lee D, Man VO, Lui WM, Wong ST and Leung GK: Long non-coding RNA expression profiles predict clinical phenotypes in glioma. Neurobiol Dis. 48:1–8. 2012. View Article : Google Scholar : PubMed/NCBI

16 

Han L, Zhang K, Shi Z, Zhang J, Zhu J, Zhu S, Zhang A, Jia Z, Wang G, Yu S, et al: LncRNA profile of glioblastoma reveals the potential role of lncRNAs in contributing to glioblastoma pathogenesis. Int J Oncol. 40:2004–2012. 2012.PubMed/NCBI

17 

Vital AL, Tabernero MD, Castrillo A, Rebelo O, Tao H, Gomes F, Nieto AB, Resende Oliveira C, Lopes MC and Orfao A: Gene expression profiles of human glioblastomas are associated with both tumor cytogenetics and histopathology. Neuro Oncol. 12:991–1003. 2010. View Article : Google Scholar : PubMed/NCBI

18 

Zhang XQ, Sun S, Lam KF, Kiang KM, Pu JK, Ho AS, Lui WM, Fung CF, Wong TS and Leung GK: A long non-coding RNA signature in glioblastoma multiforme predicts survival. Neurobiol Dis. 58:123–131. 2013. View Article : Google Scholar : PubMed/NCBI

19 

Amit D, Matouk IJ, Lavon I, Birman T, Galula J, Abu-Lail R, Schneider T, Siegal T, Hochberg A and Fellig Y: Transcriptional targeting of glioblastoma by diphtheria toxin-A driven by both H19 and IGF2-P4 promoters. Int J Clin Exp Med. 5:124–135. 2012.PubMed/NCBI

20 

Zhi F, Wang Q, Xue L, Shao N, Wang R, Deng D, Wang S, Xia X and Yang Y: The use of three long non-coding RNAs as potential prognostic indicators of astrocytoma. PLoS One. 10:e01352422015. View Article : Google Scholar : PubMed/NCBI

21 

Matouk IJ, Mezan S, Mizrahi A, Ohana P, Abu-Lail R, Fellig Y, Degroot N, Galun E and Hochberg A: The oncofetal H19 RNA connection: hypoxia, p53 and cancer. Biochim Biophys Acta. 1803:443–451. 2010. View Article : Google Scholar : PubMed/NCBI

22 

Deng M, Blondeau JJ, Schmidt D, Perner S, Muller SC and Ellinger J: Identification of novel differentially expressed lncRNA and mRNA transcripts in clear cell renal cell carcinoma by expression profiling. Genom Data. 5:173–175. 2015. View Article : Google Scholar : PubMed/NCBI

23 

He HT, Xu M, Kuang Y, Han XY, Wang MQ and Yang Q: Biomarker and competing for endogenous RNA potential of tumor-specific long noncoding RNA in chromophobe renal cell carcinoma. Onco Targets Ther. 9:6399–6406. 2016. View Article : Google Scholar : PubMed/NCBI

24 

Wu Y, Liu J, Zheng Y, You L, Kuang D and Liu T: Suppressed expression of long non-coding RNA HOTAIR inhibits proliferation and tumourigenicity of renal carcinoma cells. Tumour Biol. 35:11887–11894. 2014. View Article : Google Scholar : PubMed/NCBI

25 

Li Y, Wang T, Li Y, Chen D, Yu Z, Jin L, Ni L, Yang S, Mao X, Gui Y and Lai Y: Identification of long-non-coding RNA UCA1 as an oncogene in renal cell carcinoma. Mol Med Rep. 13:3326–3334. 2016. View Article : Google Scholar : PubMed/NCBI

26 

Hou P, Zhao Y, Li Z, Yao R, Ma M, Gao Y, Zhao L, Zhang Y, Huang B and Lu J: LincRNA-ROR induces epithelial-to-mesenchymal transition and contributes to breast cancer tumorigenesis and metastasis. Cell Death Dis. 5:e12872014. View Article : Google Scholar : PubMed/NCBI

27 

Liu T, Chi H, Chen J, Chen C, Huang Y, Xi H, Xue J and Si Y: Curcumin suppresses proliferation and in vitro invasion of human prostate cancer stem cells by ceRNA effect of miR-145 and lncRNA-ROR. Gene. 631:29–38. 2017. View Article : Google Scholar : PubMed/NCBI

28 

Wang L, Yu X, Zhang Z, Pang L, Xu J, Jiang J, Liang W, Chai Y, Hou J and Li F: Linc-ROR promotes esophageal squamous cell carcinoma progression through the derepression of SOX9. J Exp Clin Cancer Res. 36:1822017. View Article : Google Scholar : PubMed/NCBI

29 

Chen Y, Shen Z, Zhi Y, Zhou H, Zhang K, Wang T, Feng B, Chen Y, Song H, Wang R and Chu X: Long non-coding RNA ROR promotes radioresistance in hepatocelluar carcinoma cells by acting as a ceRNA for microRNA-145 to regulate RAD18 expression. Arch Biochem Biophys. 645:117–125. 2018. View Article : Google Scholar : PubMed/NCBI

30 

Shi J, Zhang W, Tian H, Zhang Q and Men T: LncRNA ROR promotes the proliferation of renal cancer and is negatively associated with favorable prognosis. Mol Med Rep. 16:9561–9566. 2017. View Article : Google Scholar : PubMed/NCBI

31 

Xie H, Fu JL and Xie C: MiR-138-5p is downregulated in patients with atrial fibrillation and reverses cardiac fibrotic remodeling via repressing CYP11B2. Eur Rev Med Pharmacol Sci. 22:4642–4647. 2018.PubMed/NCBI

32 

Calin GA and Croce CM: MicroRNA signatures in human cancers. Nat Rev Cancer. 6:857–866. 2006. View Article : Google Scholar : PubMed/NCBI

33 

Liang Z, Feng Q, Xu L, Li S and Zhou L: CREPT regulated by miR-138 promotes breast cancer progression. Biochem Biophys Res Commun. 493:263–269. 2017. View Article : Google Scholar : PubMed/NCBI

34 

Heinemann FG, Tolkach Y, Deng M, Schmidt D, Perner S, Kristiansen G, Müller SC and Ellinger J: Serum miR-122-5p and miR-206 expression: Non-invasive prognostic biomarkers for renal cell carcinoma. Clin Epigenetics. 10:112018. View Article : Google Scholar : PubMed/NCBI

35 

Stacker SA and Achen MG: The VEGF signaling pathway in cancer: The road ahead. Chin J Cancer. 32:297–302. 2013.PubMed/NCBI

36 

Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI

37 

Hu HB, Chen Q and Ding SQ: LncRNA LINC01116 competes with miR-145 for the regulation of ESR1 expression in breast cancer. Eur Rev Med Pharmacol Sci. 22:1987–1993. 2018.PubMed/NCBI

38 

Zhao L, Han T, Li Y, Sun J, Zhang S, Liu Y, Shan B, Zheng D and Shi J: The lncRNA SNHG5/miR-32 axis regulates gastric cancer cell proliferation and migration by targeting KLF4. FASEB J. 31:893–903. 2017. View Article : Google Scholar : PubMed/NCBI

39 

Li Q, Gao H, Zhou S and Liao Y: LncRNA PlncRNA-1 overexpression inhibits the growth of breast cancer by upregulating TGF-β1 and downregulating PHGDH. Breast Cancer. 25:619–625. 2018. View Article : Google Scholar : PubMed/NCBI

40 

Li P, Tong L, Song Y, Sun J, Shi J, Wu Z, Diao Y, Li Y and Wang Z: Long noncoding RNA H19 participates in metformin-mediated inhibition of gastric cancer cell invasion. J Cell Physiol. 234:4515–4527. 2018. View Article : Google Scholar : PubMed/NCBI

41 

Yan J, Zhang Y, She Q, Li X, Peng L, Wang X, Liu S, Shen X, Zhang W, Dong Y, et al: Long Noncoding RNA H19/miR-675 axis promotes gastric cancer via FADD/caspase 8/caspase 3 signaling pathway. Cell Physiol Biochem. 42:2364–2376. 2017. View Article : Google Scholar : PubMed/NCBI

42 

Lu MH, Tang B, Zeng S, Hu CJ, Xie R, Wu YY, Wang SM, He FT and Yang SM: Long noncoding RNA BC032469, a novel competing endogenous RNA, upregulates hTERT expression by sponging miR-1207-5p and promotes proliferation in gastric cancer. Oncogene. 35:3524–3534. 2016. View Article : Google Scholar : PubMed/NCBI

43 

Fei D, Sui G, Lu Y, Tan L, Zhao D and Zhang K: The long non-coding RNA-ROR promotes osteosarcoma progression by targeting miR-206. J Cell Mol Med. 23:1865–1872. 2019. View Article : Google Scholar : PubMed/NCBI

44 

Kyzas PA, Cunha IW and Ioannidis JP: Prognostic significance of vascular endothelial growth factor immunohistochemical expression in head and neck squamous cell carcinoma: A metaanalysis. Clin Cancer Res. 11:1434–1440. 2005. View Article : Google Scholar : PubMed/NCBI

45 

Zhang T, Liu M, Wang C, Lin C, Sun Y and Jin D: Down-regulation of miR-206 promotes proliferation and invasion of laryngeal cancer by regulating VEGF expression. Anticancer Res. 31:3859–3863. 2011.PubMed/NCBI

46 

Liang Z, Bian X and Shim H: Downregulation of microRNA-206 promotes invasion and angiogenesis of triple negative breast cancer. Biochem Biophys Res Commun. 477:461–466. 2016. View Article : Google Scholar : PubMed/NCBI

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October 2019
Volume 20 Issue 4

Print ISSN: 1791-2997
Online ISSN:1791-3004

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Copy and paste a formatted citation
APA
Shi, J., Zhang, D., Zhong, Z., & Zhang, W. (2019). lncRNA ROR promotes the progression of renal cell carcinoma through the miR‑206/VEGF axis. Molecular Medicine Reports, 20, 3782-3792. https://doi.org/10.3892/mmr.2019.10636
MLA
Shi, J., Zhang, D., Zhong, Z., Zhang, W."lncRNA ROR promotes the progression of renal cell carcinoma through the miR‑206/VEGF axis". Molecular Medicine Reports 20.4 (2019): 3782-3792.
Chicago
Shi, J., Zhang, D., Zhong, Z., Zhang, W."lncRNA ROR promotes the progression of renal cell carcinoma through the miR‑206/VEGF axis". Molecular Medicine Reports 20, no. 4 (2019): 3782-3792. https://doi.org/10.3892/mmr.2019.10636