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circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p

  • Authors:
    • Rui Wang
    • Yongli He
    • Wuxia Ma
    • Jindong Xu
    • Qi Zhong
    • Cheng Huang
  • View Affiliations / Copyright

    Affiliations: Department of Cardiology, Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, Guangdong 510080, P.R. China, Department of Cardiology, Jinshazhou Hospital of Guangzhou University of Chinese Medicine, Guangzhou, Guangdong 510168, P.R. China, Department of Nursing, Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, Guangdong 510080, P.R. China, Department of Anesthesiology, Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, Guangdong 510080, P.R. China, Department of Cardiology, The First Affiliated Hospital of Hunan University of Medicine, Huaihua, Hunan 418000, P.R. China
    Copyright: © Wang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
  • Article Number: 232
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    Published online on: June 17, 2025
       https://doi.org/10.3892/mmr.2025.13597
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Abstract

The present study aimed to identify differentially expressed circRNAs in hypertrophic cardiac tissues and explored the potential regulatory role and mechanism of one differentially expressed circRNA in myocardial hypertrophy. RNA sequencing was used to identify differentially expressed circRNAs in hypertrophic and control cardiac tissues. CircRNA expression levels were verified by reverse transcription‑quantitative PCR. Isoproterenol (ISO) was used to induce hypertrophy of AC16 cells. The extent of cell hypertrophy was indicated by the cell size, protein/DNA ratio and levels of B‑type natriuretic peptide (BNP) and β‑myosin heavy chain (β‑MHC). The interactions between hsa_circ_0072107 and miR‑516b‑5p, as well as between miR‑516b‑5p and zinc ring finger protein 36 (ZFP36), were confirmed through dual luciferase assays, biotinylated probe pull‑down and anti‑AGO2 RNA immunoprecipitation assays. hsa_circ_0072107 was one of the most upregulated circRNAs in hypertrophic cardiac tissues. hsa_circ_0072107 overexpression and ISO treatment increased cell size, elevated the protein/DNA ratio and increased the levels of BNP and β‑MHC in AC16 cells, indicating that hsa_circ_0072107 aggravates AC16 hypertrophy. These changes induced by ISO treatment could be blocked by the knockdown of hsa_circ_0072107. The dual‑luciferase activity assay indicated that miR‑516b‑5p can bind to hsa_circ_0072107. miR‑516b‑5p binding site mutation blocked the effect of hsa_circ_0072107. ZFP36 is a target gene of miR‑516b‑5p, which suppresses AC16 hypertrophy. hsa_circ_0072107 overexpression alleviated the effect of miR‑516b‑5p overexpression on cell hypertrophy and ZFP36 expression. hsa_circ_0072107 is up‑regulated in hypertrophic cardiac tissues and potentially promotes AC16 hypertrophy and may play its role by acting as a competitive endogenous RNA of miR‑516b‑5p. Thus, hsa_circ_0072107 may be a novel target for the treatment of myocardial hypertrophy.
View Figures

Figure 1

circRNA expression profiles in
interventricular septum specimens from patients with HC compared
with data from NC cardiac tissue. RNA sequencing was performed to
identify differentially expressed circRNAs (n=5). (A) intensity
scatter plot, (B) volcano plot, (C) heatmap and (D) expression
levels of the top 10 differentially expressed circRNAs identified
by RNA-sequencing were verified by PCR (n=10). circRNA, circular
RNA; HC, hypertrophic cardiomyopathy; NC, negative control.

Figure 2

hsa_circ_0072107 knockdown alleviated
the hypertrophy of AC16 cells induced by ISO. (A) Diagrammatic
representation of hsa_circ_0072107. (B) The sequence around the
hsa_circ_0072107 splicing site. The PCR amplification product of
hsa_circ_0072107 was sequenced. (C) The expression of
hsa_circ_0072107 in AC16 was quantified by reverse
transcription-quantitative PCR. hsa_circ_0072107 overexpression and
ISO treatment (D) enlarged cell size (scale bars, 50 µm), (E)
elevated the protein/DNA ratio and (F) escalated BNP and β-MHC
protein levels, whereas the enlargement effect of ISO could be
blocked by hsa_circ_0072107 knockdown. *P<0.05, n=3. ISO,
isoproterenol; BNP, B-type natriuretic peptide; β-MHC, β-myosin
heavy chain vector; si, small interfering; vector group, cells
transfected with empty vector; ov-circ_0072107, cells transfected
with hsa_circ_0072107 overexpression vector; saline, cells treated
with saline; ISO, cells treated with isoproterenol; ISO + si-NC,
ISO treated cells transfected with negative siRNA (si-NC); ISO +
si-circ_0072107, ISO treated cells transfected with siRNA targeting
hsa_circ_0072107 (si-circ_0072107).

Figure 3

miR-516b-5p is a miRNA binding to
hsa_circ_0072107. (A) Venn diagram showed that two miRNAs may bind
to hsa_circ_0072107. (B) The binding site information of
miR-516b-5p and miR-510-5p on hsa_circ_0072107. (C) Reverse
transcription-quantitative PCR validation of transfection
efficiency in cells transfected with miR-NC, miR-516b-5p mimics and
inhibitors. (D) hsa_circ_0072107, miR-516b-5p and miR-510-5p levels
in RNA enriched by circ_0072107 probe or scramble probe during
biotinylated probe pull-down assay. (E) hsa_circ_0072107 level in
miRNA complexes enriched by anti-AGO or IgG during anti-AGO2 RIP
assay. AC16 cells transfected with miR-516b-5p mimic or miR-NC were
used. (F) The effect of miR-516b-5p mimic or miR-NC on the relative
luciferase activity of luciferase reporter with linear circ_0072107
or linear mutcirc_0072107 as the artificial 3′ UTR of
Renilla, respectively. *P<0.05, **P<0.01, n=3. miRNA,
microRNA; AGO2, argonaute 2; RIP, RNA immunoprecipitation; NC,
negative control.

Figure 4

Overexpression of hsa_circ_0072107
with mutant miR-516b-5p binding site cannot induce AC16
hypertrophy. (A) miR-516b-5p expression level in AC16 cells
transfected with empty vector (vector), hsa_circ_0072107
overexpression vector (ov-circ_0072107) and overexpression vector
of hsa_circ_0072107 with mutant miR-516b-5p binding site
(mutov-circ_0072107). No significant difference was observed in (B)
cell size (scale bars, 50 µm), (C) protein/DNA ratio, or (D) levels
of BNP and β-MHC between the vector and mutov-circ_0072107 groups.
miRNA, microRNA.

Figure 5

ZFP36 is a target gene of
miR-516b-5p. (A) Venn diagram identified differentially expressed
target genes of miR-516b-5p. In the GSE32453 dataset, 461
differentially expressed mRNAs were identified in the myocardium of
patients with hypertrophic cardiomyopathy. (B) The binding sites of
miR-516b-5p on ZFP36 3′UTR. (C) ZFP36 mRNA level in RNA enriched by
miR-516b-5p probe or mutant miR-516b-5p probe in a biotinylated
probe pull-down assay. (D) The effect of miR-NC or miR-516b-5p
mimic on the relative luciferase activity of luciferase reporter
using ZFP36 3′UTR or mutant ZFP36 3′UTR (mutZFP36 3′UTR) as the
artificial 3′ UTR of Renilla. (E) The protein level of ZFP36
in AC16 cells transfected with miR-NC, miR-516b-5p, miR-NC
inhibitor, or miR-516b-5p inhibitor. *P<0.05, n=3. ZFP36, zinc
ring finger protein 36; miRNA, microRNA; NC, negative control.

Figure 6

Effect of miR-516b-5p overexpression
on AC16 hypertrophy and ZFP36 expression was blocked by
hsa_circ_0072107 overexpression. Under ISO treatment,
hsa_circ_0072107 overexpression vector (ov-circ_0072107),
miR-516b-5p inhibitor, miR-516b-5p mimic (miR-516b-5p), or
miR-516b-5p mimic plus ov-circ_0072107 was transfected into AC16
cells, respectively and the transfection reagent was used as
negative control (ISO + NC). (A) miR-516b-5p expression level
effects on (B) cell size (scale bars, 50 µm), (C) protein/DNA ratio
and (D) the protein levels of BNP and β-MHC were measured. (E and
F) The protein level of ZFP36 in different groups was detected by
western blotting. (G) Relative ZFP36 levels measured by reverse
transcription-quantitative PCR in different experimental groups.
(H) Western blot analysis of ZFP36 protein expression in different
experimental groups, with quantification of protein expression
levels shown in the histogram. *P<0.05, n=3. miRNA, microRNA;
ZFP36, zinc ring finger protein 36; ISO, isoproterenol; NC,
negative control; vector group, cells transfected with empty
vector; ov-circ_0072107, cells transfected with hsa_circ_0072107
overexpression vector; si-NC group, cells transfected with negative
siRNA; si-circ_0072107 group, cells transfected with siRNA
targeting hsa_circ_0072107 (si-circ_0072107).
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Copy and paste a formatted citation
Spandidos Publications style
Wang R, He Y, Ma W, Xu J, Zhong Q and Huang C: circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p. Mol Med Rep 32: 232, 2025.
APA
Wang, R., He, Y., Ma, W., Xu, J., Zhong, Q., & Huang, C. (2025). circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p. Molecular Medicine Reports, 32, 232. https://doi.org/10.3892/mmr.2025.13597
MLA
Wang, R., He, Y., Ma, W., Xu, J., Zhong, Q., Huang, C."circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p". Molecular Medicine Reports 32.3 (2025): 232.
Chicago
Wang, R., He, Y., Ma, W., Xu, J., Zhong, Q., Huang, C."circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p". Molecular Medicine Reports 32, no. 3 (2025): 232. https://doi.org/10.3892/mmr.2025.13597
Copy and paste a formatted citation
x
Spandidos Publications style
Wang R, He Y, Ma W, Xu J, Zhong Q and Huang C: circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p. Mol Med Rep 32: 232, 2025.
APA
Wang, R., He, Y., Ma, W., Xu, J., Zhong, Q., & Huang, C. (2025). circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p. Molecular Medicine Reports, 32, 232. https://doi.org/10.3892/mmr.2025.13597
MLA
Wang, R., He, Y., Ma, W., Xu, J., Zhong, Q., Huang, C."circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p". Molecular Medicine Reports 32.3 (2025): 232.
Chicago
Wang, R., He, Y., Ma, W., Xu, J., Zhong, Q., Huang, C."circRNA hsa_circ_0072107 aggravates myocardial hypertrophy via its function as a competitive endogenous RNA of miR‑516b‑5p". Molecular Medicine Reports 32, no. 3 (2025): 232. https://doi.org/10.3892/mmr.2025.13597
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