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| LncRNA HCG18 promotes prostate cancer progression by regulating the miR-512-3p/HK-2 axis |
Yaru Zhua,Zhijing Wangb,Haopeng Lia,Zhen Renc,Tong Zia,Xin Qina,Wenhuizi Sund,Xi Chena,Gang Wua,*( )
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aDepartment of Urology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, China
bDepartment of Gastroenterology and Hepatology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, China
cDepartment of Medicine, Dalian University of Technology, Dalian, China
dDepartment of Obstetrics and Gynecology, Tongji Hospital, School of Medicine, Tongji University, Shanghai, China |
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Abstract Objective:Long non-coding RNAs (lncRNAs) play an important role in tumor progression. Numerous studies show that lncRNAs are strongly associated with prostate cancer (PCa) progression. The aim of this study was to investigate the pathway through which lncRNA HCG18 regulates PCa progression by bioinformatics analysis and experiments. Methods:We compared HCG18 expression in PCa versus normal tissue and cells by data and cell lines, followed by comparing the changes in tumor cell proliferation, migration, and invasive ability after knockdown of HCG18. Then we searched for its downstream pathway by database and validated the pathway in vivo and in vitro. Results:HCG18 was highly expressed in PCa and has the ability to promote tumor proliferation, migration, and invasion; knockdown of HCG18 led to a decrease in the ability of cells to do so, which can be reversed by knockdown of miR-512-3p or overexpression of hexokinase 2. Conclusion:Our in vivo and in vitro experiments suggest that HCG18 can play a role in promoting PCa progression by blocking the inhibition of hexokinase 2 by miR-512-3p via sponge adsorption.
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Received: 07 May 2023
Available online: 20 October 2024
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Corresponding Authors:
*E-mail address: 2013wg_urologist@tongji.edu.cn (G. Wu).
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LncRNA HCG18 is markedly increased in PCa and related to cells proliferation, migration, and invasion. (A) LncRNA HCG18 expression levels in PCa samples (n=499) and normal samples (n=52); p=0.039 (data source: ENCORI project); (B) Kaplan-Meier curves for overall survival of PCa patients with different HCG18 expression levels, p=0.024 (data source: GEPIA 2); (C) Relative expression levels of lncRNA HCG18 in different PCa cell lines by qRT-PCR; (D) Analysis of PC-3 and LNCaP cell viability using cell counting kit-8 solution (Dojindo, Tokyo, Japan); (E) Analysis of PC-3 and LNCaP cell migratory capacity using wound healing assay; (F) Analysis of PC-3 and LNCaP cell invasive capacity using transwell assay; (G) The statistical chart of healing experiments comparing cell migration capacity; (H) The statistical chart of transwell assay comparing cell invasion capacity. LncRNA, long non-coding RNA; PCa, prostate cancer; qRT-PCR, quantitative real-time polymerase chain reaction; FPKM, fragment per kilobase of transcript per million mapped reads; NC, negative control; sh-RNA, short hairpin RNA; OD, optical density. ? p<0.05; ?? p<0.01; ??? p<0.001. Scale bars, 100 μm. The databases we used are publicly available and can be reused without restriction through open licenses.
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HK-2 promotes PCa progression and can be regulated by LncRNA HCG18. (A) Metabolic flux analysis of drug-sensitive (C4-2) and drug-resistant (C4-2R) cells; (B) Western blot comparison of HK-2 expression after knockdown of HCG18; (C) Western blot comparison of HK-2 expression after knockdown of HK-2; (D) Analysis of PC-3 and LNCaP cell viability using cell counting kit-8 solution (Dojindo, Tokyo, Japan); (E) Analysis of PC-3 and LNCaP cell migratory capacity using wound healing assay; (F) Analysis of PC-3 and LNCaP cell invasive capacity using transwell assay; (G) The statistical chart of healing experiments comparing cell migration capacity; (H) The statistical chart of transwell assay comparing cell invasion capacity. GAP, glyceraldehyde 3-phosphate; 3PG, 3-phosphoglycerate; DHAP, dihydroxyacetone phosphate; LncRNA, long non-coding RNA; PCa, prostate cancer; OD, optical density. ?p<0.05; ?? p<0.01; ??? p<0.001. Scale bars, 100 μm.
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miR-512-3p correlates with HCG18 and HK-2. (A) Bioinformatic analysis for screening miRNAs; (B) HCG18 was confirmed to bind to miR-512-3p by RNA pull-down assay; (C) The binding site of HK-2 to miR-512-3p; (D) HCG18 was confirmed to bind to HK-2 by the luciferase reporter assay. ns, nonsignificant; oe, overexpress. ? p<0.05; ?? p<0.01.
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miR-512-3p directly interacts with HCG18 and HK-2 and inhibits the progression of prostate cancer cells. (A) Analysis of PC-3 cell migratory capacity using wound healing assay; (B) Analysis of PC-3 cell invasive capacity using transwell assay; (C) Analysis of LNCaP cell migratory capacity using wound healing assay; (D) Analysis of LNCaP cell invasive capacity using transwell assay; (E) The statistical chart of healing experiments comparing cell migration capacity; (F) The statistical chart of transwell assay comparing cell invasion capacity; (G and H) Analysis of PC-3 and LNCaP cell viability using cell counting kit-8 solution (Dojindo, Tokyo, Japan). NC, negative control; OD, optical density. ? p<0.05; ?? p<0.01; ??? p<0.001. Scale bars, 100 μm.
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LncRNA HCG18 promotes PCa progression via the LncRNA HCG18/miR-512-3p/HK-2 axis. (A) Analysis of PC-3 and LNCaP cell invasive capacity using the transwell assay; (B) Analysis of PC-3 and LNCaP cell viability using the cell counting kit-8 solution (Dojindo, Tokyo, Japan); (C) The statistical chart of the transwell assay comparing cell invasion capacity; (D) The statistical chart of healing experiments comparing cell migration capacity; (E) Analysis of PC-3 and LNCaP cell migratory capacity using the wound healing assay. OD, optical density; NC, negative control; PCa, prostate cancer; oe, overexpress. ? p<0.05; ?? p<0.01; ??? p<0.001. Scale bars, 100 μm.
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Animal experiments of nude mice groups under different treatments. (A) Tumor visualization. (B) Tumor weight analysis. NC, negative control. ? p<0.05; ??? p<0.001.
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