Circular RNA hsa_circ_0007990 as a blood biomarker for unruptured intracranial aneurysm with aneurysm wall enhancement

Abstract

Background: Circular RNAs (circRNAs) may involve the formation and rupture of intracranial aneurysms (IA). Inflammation plays a vital role in the development and progression of IA, which can be reflected by aneurysm wall enhancement (AWE) on high-resolution vessel wall magnetic resonance imaging (HR-VWI). This study aims to evaluate the role of circRNAs as the blood inflammatory biomarker for unruptured IA (UIA) patients with AWE on HR-VWI.

Methods: We analyzed the circRNA expression profiles in the peripheral blood samples among subjects from saccular UIA with AWE, UIA without AWE, and healthy controls by the circRNA microarray. The differential expression of hsa_circ_0007990 was assessed. We constructed the hsa_circ_0007990-microRNA-mRNA network and the regulatory axis of hub genes associated with the AWE in UIA.

Results: Eighteen patients harboring saccular UIAs with HR VWI and five healthy controls were included. We found 412 differentially expressed circRNAs between UIA patients and healthy controls by circRNA microarray. Two hundred thirty-one circRNAs were significantly differentially expressed in UIA patients with AWE compared with those without AWE. Twelve upregulated circRNAs were associated with AWE of UIA, including hsa_circ_0007990, hsa_circ_0114507, hsa_circ_0020460, hsa_circ_0053944, hsa_circ_0000758, hsa_circ_0000034, hsa_circ_0009127, hsa_circ_0052793, hsa_circ_0000301 and hsa_circ_0000729. The expression of hsa_circ_0007990 was increased gradually in the healthy control, UIA without AWE, and UIA with AWE confirmed by RT-PCR (P<0.001). We predicted 4 RNA binding proteins (Ago2, DGCR8, EIF4A3, PTB) and period circadian regulator 1 as an encoding protein with hsa_circ_0007990. The hsa_circ_0007990-microRNA-mRNA network containing five microRNAs (miR-4717-5p, miR-1275, miR-150-3p, miR-18a-5p, miR-18b-5p), and 97 mRNAs was constructed. The five hub genes (hypoxia-inducible factor 1 subunit alpha, estrogen receptor 1, forkhead box O1, insulin-like growth factor 1, CREB binding protein) were involved in the inflammatory response.

Conclusion: Differentially expressed blood circRNAs associated with AWE on HR-VWI may be the novel inflammatory biomarkers for assessing UIA patients. The mechanism of hsa_circRNA_0007990 for UIA progression needs to investigate further.

Keywords: aneurysm wall enhancement; blood marker; circular RNA; inflammation; intracranial aneurysm; vessel wall imaging.

Figure 1

Differentially expressed circRNAs in peripheral blood identified by circRNAs microarray. (A) The hierarchical clustering of differentially expressed circRNAs between the unruptured intracranial aneurysm (UIA) and healthy control groups. Group A represented healthy control; group B represented the UIA group. (B) The scatterplot is used to assess the circRNA expression variation between the UIA and healthy control group of samples. The circRNAs above the top green line and below the green bottom line indicate more than a 1.3-fold change of circRNAs between the two compared samples. (C) Volcano Plots are used to visualize differential expression between the UIA and healthy control groups. The red point in the plot represents the differentially expressed circRNAs with statistical significance. (D) The hierarchical clustering of differentially expressed circRNAs between the UIA with aneurysm wall enhancement (AWE) and UIA without AWE. Group B1 represented UIA with AWE of samples; group B2 represented UIA without AWE of samples. (E) The scatterplot is used for assessing the circRNA expression variation between the UIA with AWE and UIA without AWE. The circRNAs above the top green line and below the green bottom line indicate more than a 1.3-fold change of circRNAs between the two compared samples. (F) Volcano Plots are used for visualizing differential expression between the UIA with AWE and UIA without AWE. The red point in the plot represents the differentially expressed circRNAs with statistical significance. (G) The genomic origin of the differentially expressed circRNAs among three groups of samples (UIA with AWE, UIA without AWE, and healthy controls). (H) A total of 12 significantly differentially expressed peripheral blood circRNAs among the three compared groups (UIA with AWE, UIA without AWE, and healthy controls) were identified by circRNA microarray.

Figure 2

Upregulated expression of hsa_circ_0007990 associated with aneurysm wall enhancement confirmed by RT-PCR. The data are the mean ± SD, **p < 0.01, ***p < 0.001.

Figure 3

Schematic diagram of hsa_circ_0007990. The red bar represents the microRNA binding sites, the blue bar represents proteins binding with hsa_circRNA_0007990, and the light green bar represents the open reading frame.

Figure 4

RNA binding proteins with hsa_circ_0007990. According to the CircInteractome database, the 3′start flanking intron upstream of hsa_circRNA_0007990 has a potential binding region with DGCR8 protein, and the 3′start flanking intron downstream of hsa_circRNA_0007990 has a potential binding region with Ago2, EIF4A3, and PTB protein.

Figure 5

Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) functional enrichment analysis of the predicted target genes from has_circRNA_0007990. (A) KEGG. (B) GO-molecular functions. (C) GO-biological processes. (D) GO-cell components.

Figure 6

The circRNA-miRNA-mRNA regulatory network of hsa_circRNA_0007990. (A) Protein-protein interaction network obtained on STRING website. (B) Relationship network diagram of hub genes from protein-protein interaction network. (C) The reconstructed circRNA-microRNA-hub genes network.