Screening of circular RNAs and validation of circANKRD36 associated with inflammation in patients with type 2 diabetes mellitus

Abstract

Circular RNAs (circRNAs) are an abundant class of endogenous non-coding RNAs and are associated with numerous diseases, including cancer, cardiovascular diseases, and type 2 diabetes mellitus (T2DM). However, the association between circRNAs and inflammation or inflammatory cytokines in patients with T2DM remains to be fully elucidated. The purpose of the present study was to investigate the expression profiles of circRNAs in peripheral leucocytes of patients with T2DM and their association with inflammatory cytokines. Peripheral blood from patients with T2DM (n=43) and healthy individuals (n=45) were collected for RNA sequencing and later verification. Reverse transcription-polymerase chain reaction (RT-PCR) and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analyses were used to detect the expression levels of circRNAs. The expression of inflammatory factors, including interleukin (IL)-1, (IL)-6, and tumor necrosis factor (TNF)-α were measured via enzyme-linked immunosorbent assay. Furthermore, the mRNA expression level of ankyrin repeat domain 36 (ANKRD36), a protein located at 2q11.2 that interacts with the GAPDH gene, was measured using RT-qPCR analysis. The circRNA/microRNA (miRNA) interaction was predicted using RegRNA and mirPath software. In total, 220 circRNAs were found to be differentially expressed between patients with T2DM and healthy individuals, of which 107 were upregulated and 113 were downregulated. Among the nine selected circRNAs, circANKRD36 was significantly upregulated in patients with T2DM compared with control subjects (P=0.02). The expression level of circANKRD36 was positively correlated with glucose and glycosylated hemoglobin (r=0.3250, P=0.0047 and r=0.3171, P=0.0056, respectively). The expression level of IL-6 was significantly different between the T2DM group and control group (P=0.028) and was positively correlated with circANKRD36. The difference of circANKRD36 host gene expression between patients with T2DM and healthy controls was significant (P=0.04). Taken together, circANKRD36 may be involved in T2DM and inflammation-associated pathways via interaction with miRNAs, including hsa-miR-3614-3p, hsa-miR-498, and hsa-miR-501-5p. The expression of circANKRD36 was up regulated in peripheral blood leucocytes and was correlated with chronic inflammation in T2DM. Therefore, circANKRD36 can be used as a potential biomarker for screening chronic inflammation in patients with T2DM.

Differentially expressed non-coding RNAs induced by transmissible gastroenteritis virus potentially regulate inflammation and NF-κB pathway in porcine intestinal epithelial cell line

“…We demonstrated that miR-22 was up-regulated by TGEV infection and had an anti-inflammation effect by targeting IL-6 and restraining DDX58, and CCL5, which is correlated with previous reports, but through different mechanisms. cir-cRNAs are an abundant class of endogenous ncRNAs and function as a regulator via various mechanisms such as miRNA sponge, binding to RNA-binding protein, and regulating transcription and translation of gene [44][45][46]. We found that miR-22 could be directly attached to ssc_circ_9380 and mediate TGEV-induced inflammation response, suggesting ssc_circ_9380 acts as the sponge of miR-22.…”

Interactions Among Non-Coding RNAs in Diabetic Nephropathy

“…A back-splicing event during the transcription process that ligates a splice donor site at the upstream to an acceptor sites at downstream of the RNA transcript, produces a covalently closed loop RNA product that does not contain either 5' to 3' polarity nor a poly-adenylated tail, and this is known as the circular RNA (circRNA) (Cao et al, 2018;Fang et al, 2018;Sulaiman et al, 2018;Hu et al, 2019). Due to its circular structure, circRNA is very stable and resistant to degradation by exoribonucleases (Liu et al, 2017;Cao et al, 2018).…”

“…Although circRNAs are known much earlier, its function and potential as biomarkers are not well understood. The most established function of circRNA is that circRNAs can act as a sponge to miRNAs to regulate gene expression (Cao et al, 2018;Fang et al, 2018;Hu et al, 2019), and in some cases, with proteins (Du et al, 2016). In terms of the circRNAs localization, some exonic circRNAs are abundantly present in the cytoplasm (Jeck et al, 2013), but no other information is available for other groups of circRNAs.…”

Islet-expressed circular RNAs are associated with type 2 diabetes status in human primary islets and in peripheral blood

“…CircRNAs may have potential as biomarkers for the development of diabetes or as future molecular targets for novel diabetes therapeutics [14][15][16]. An important pre-requisite for this is the characterisation of circRNA sequences in diabetes-relevant tissues such as pancreatic islets and in more accessible tissues such as peripheral blood.…”