Identification and validation of disease severity-related circular RNA in acute pancreatitis

Abstract

Acute pancreatitis arises from the activation of digestive enzymes in pancreatic acinar cells, leading to autodigestion of the pancreas and surrounding tissues. It is a common digestive tract emergency which requires hospitalization, and its incidence is increasing worldwide. In the past decade, several advances have been made in the treatment of acute pancreatitis. However, there is still a lack of efficacious drugs for clinical practice, and the limited value of existing biomarkers for early warning of the severity of acute pancreatitis is a major obstacle. Thus, there is an urgent need to gain a better understanding of the molecular mechanisms of acute pancreatitis. Circular RNAs (circRNAs) are a unique class of RNA molecules that are covalently closed. Ongoing investigations have provided evidence that circRNAs govern downstream target expression by acting as miRNA sponges, functioning as transcription factors, interacting with RNA-binding proteins, and regulating alternative splicing. These mechanisms support the pivotal role of circRNAs in a wide variety of physiological and pathological conditions, such as innate immunity, inflammation, neuronal function, and tumorigenesis. To explore the role of circular RNA in acute pancreatitis, we employed circRNA microarray technology (Arraystar Human circRNA Array V2) to examine the circRNA expression profile in the blood of three acute pancreatitis patients and three healthy controls. Clinical acute pancreatitis samples were obtained from Xiangya Hospital, Central South University. This study was approved by the Ethics Committee of Xiangya hospital (No. 2019010008). Normal control patients were recruited from among individuals who had visited Xiangya Hospital for a routine checkup. Written informed consent was obtained from all participants or their legal representatives for publication of data. The diagnosis and severity classification of acute pancreatitis were performed according to the American Gastroenterological Association guidelines and the Revised Atlanta Classification (RAC). circRNAs with a fold change ≥1.5 and a P value<0.05 were considered to be differentially expressed. As shown in Figure 1A, the two groups presented different expression profiles. We found that 91 circRNAs were significantly differentially expressed in the blood of acute pancreatitis patients, with 10 circRNAs exhibiting increased expression and 81 exhibiting decreased expression (Figure 1B,C). Among the differentially expressed circRNAs in acute pancreatitis, downregulated circRNAs are more prevalent than upregulated circRNAs, and the differential expression is more significant. Therefore, the present study focused on downregulated circRNAs. We selected circRNAs that are downregulated at least 2.5-fold and excluded those with fewer than 1000 bases to ensure the accuracy of qPCR. Based on these criteria, we identified nine circRNAs (circ_0006554, circ_0007798, circRNA_405815, circ_0001847, circ_0069748, circ_0001850, circ_0008417, circ_0002560, and circ_0000008) for validation by qPCR (Applied Biosystems, Foster City, USA) in blood samples from 30 acute pancreatitis patients (10 patients each with mild acute pancreatitis, moderate severe acute pancreatitis, and severe acute pancreatitis) and 15 healthy individuals. The levels of circ_0007798, circ_0001847, and circ_0069748 were significantly lower in acute pancreatitis patients than in normal controls, while the remaining circRNAs were not significantly differentially expressed (Figure 1D). In addition, the levels of circ_0007798 increased gradually with the severity of acute pancreatitis, suggesting that circ_0007798 is associated with the clinical severity of the disease (Figure 1E). Differential circRNAs have been studied for the diagnosis of pancreatic diseases. The expression level of circ_0007798 can be used to grade the severity of acute pancreatitis and provide individualized treatment. Furthermore, homology analysis (NCBI blast) revealed that circ_0007798 has a high degree of conservation between rats and humans according to the basic local alignment search tool. In conclusion, according to the

1. Introduction

Acute pancreatitis arises from the activation of digestive enzymes in pancreatic acinar cells, leading to autodigestion of the pancreas and surrounding tissues. It is a common digestive tract emergency which requires hospitalization, and its incidence is increasing worldwide [1]. In the past decade, several advances have been made in the treatment of acute pancreatitis [2]. However, there is still a lack of efficacious drugs for clinical practice, and the limited value of existing biomarkers for early warning of the severity of acute pancreatitis is a major obstacle. Thus, there is an urgent need to gain a better understanding of the molecular mechanisms of acute pancreatitis.

Circular RNAs (circRNAs) are a unique class of RNA molecules that are covalently closed [3]. Ongoing investigations have provided evidence that circRNAs govern downstream target expression by acting as miRNA sponges, functioning as transcription factors, interacting with RNA-binding proteins, and regulating alternative splicing [4]. These mechanisms support the pivotal role of circRNAs in a wide variety of physiological and pathological conditions, such as innate immunity, inflammation, neuronal function, and tumorigenesis [5].