Research progress of non-coding RNA in atrial fibrillation

Atrial fibrillation (AF) is a common arrhythmia in clinic, and its incidence is increasing year by year. In today's increasingly prevalent society, ageing poses a huge challenge to global healthcare systems. AF not only affects patients' quality of life, but also causes thrombosis, heart failure and other complications in severe cases. Although there are some measures for the diagnosis and treatment of AF, specific serum markers and targeted therapy are still lacking. In recent years, ncRNAs have become a hot topic in cardiovascular disease research. These ncRNAs are not only involved in the occurrence and development of AF, but also in pathophysiological processes such as myocardial infarction and atherosclerosis, and are potential biomarkers of cardiovascular diseases. We believe that the understanding of the pathophysiological mechanism of AF and the study of diagnosis and treatment targets can form a more systematic diagnosis and treatment framework of AF and provide convenience for individuals with AF and the society.


Introduction
Atrial fibrillation (AF) is a common arrhythmias in clinic, with a high risk of death, stroke, and peripheral embolism, and its incidence has been increasing year by year. Risk factors for AF are closely related to cardiovascular disease, with organic or functional heart problems being more common. In addition, age, gender and genetic factors are also important factors leading to the occurrence of AF (1, 2). AF not only affects life quality of the patients, but also has complications such as thrombosis and heart failure in severe cases. Atrial remodeling is considered to be the basis of the occurrence and development of AF, including structural remodeling, electrical remodeling, neural remodeling, etc (3)(4)(5). The diagnosis of AF mainly depends on electrocardiogram findings, which are often found after complications occur, and there is a certain lag (6). Therefore, biomarkers have potential value in the early diagnosis of AF. Currently, drug therapy for AF patients has poor efficacy and side effects. Radiofrequency ablation is more effective than drug therapy, but the patients are yet able to avoid the operational risks, postoperative recurrence, and high healthcare cost (7)(8)(9). Actively searching for new diagnosis and treatment strategies and exploring the molecular mechanism of AF have great clinical significance and translational prospects.
In recent years, non-coding RNA (ncRNA) has become a research hotspot in cardiovascular diseases. ncRNA mainly includes miRNA, LncRNA and CircRNA. These ncRNAs can not only participate in the occurrence and development of AF, but also play a part in the pathophysiological processes such as myocardial infarction and atherosclerosis, which are potential biomarkers for cardiovascular diseases (10). This article reviews the pathophysiological mechanism of AF, introduces the mechanism and potential value of ncRNAs in AF, and provides a theoretical basis for the diagnosis, treatment and prognosis monitoring of AF ( Figure 1, Tables 1, 2).  Frontiers in Cardiovascular Medicine miRNAs in both non-relapsed and relapsed groups, the results showed that miRNA-451a was down-regulated in relapsed patients, and the recurrence of AF was positively correlated with an increased percentage of scars. It is suggested that low expression of miR-451a may play an important role in the recurrence of AF by controlling fibrosis and progression (70). Akselrod AS et al. found that plasma miR-320a-3p level in patients with AF was higher than that in healthy controls, and the expression level was positively correlated with CHADS-VASc score (28). Sasano T et al. identified 11 candidate miRNAs using high-throughput sequencing and clinical sample validation, and found that miR-214-3p and miR-342-5p had high accuracy in the diagnosis of patients with AF combined with clinicpathological parameter analysis (71). Yang et al. observed genome-wide differential expression profiles of miRNAs in 180 peripheral blood samples and found 14 miRNAs with significant differential expression, among which miR-1266, miR-4279 and miR-4666a-3p were significantly increased in expression, which are potential targets for future diagnosis and treatment of AF (72). About one-third of patients undergoing coronary artery bypass grafting will develop postoperative AF, which seriously affects the prognosis of patients. In order to monitor the occurrence of postoperative AF, Athanasiou et al. prospectively recruited 34 patients after surgery, and compared the myocardial tissue with normal sinus rhythm after surgery, and found 16 differentially expressed miRNAs. The expression of miR-208a was significantly decreased, and the expression of miR-483-5p was significantly increased. It is suggested that these differentially expressed miRNAs can be used to predict the recurrence of AF after coronary artery bypass grafting (73). Kilic et al. recruited 63 patients after coronary artery bypass grafting and monitored their heart rate until discharge. Among them, 20 patients developed postoperative AF, and PCR detected the expression of miR-199a and miR-195. The results showed that the expression of miR-199a significantly decreased in the postoperative AF group, demonstrate its effectiveness as a biomarker for cardiac surgery management (74). By Solexa sequencing 100 patients with AF who underwent catheter ablation and 100 healthy individuals, Wu et al. found that miR-409-3p and miR-432 were significantly reduced in the plasma of patients with AF and are potential markers of AF (75). Xia et al. showed for the first time that plasma miRNA-150 levels in patients with atrial fibrillation are significantly lower than those in healthy individuals, which is a potential biomarker to aid in the diagnosis of atrial fibrillation (76). These studies indicate that miRNAs differentially expressed in plasma of patients with AF and postoperative patients can play an important indicator role in the diagnosis and prognosis monitoring of AF.

miRNAs involved in the regulation of electrical remodeling
Electrical remodeling of atrial muscle is closely related to the occurrence of AF. Electrical remodeling refers to recurrent episodes of AF or continuous atrial stimulation, which leads to progressive shortening of the effective refractory period of the atrium, and the decrease, reversal or disappearance of the physiological frequency adaptation of the atrial refractory period, making AF more likely to be induced and sustained (87,92). AF is caused by abnormal electrical activity of atrial myocardium. During the occurrence of AF, many ion channels also have significant changes, mainly including: L-type Ca 2+ channel, transient outward K + channel, strong inward rectification K + channel (IK1), acetylcholine-activated K + channel (IK, ACh), and ultra-fast delayed rectification K + channel (IKur) (93,94).
Yang et al. found that the expression of miR-328 was increased in the atrial tissue of AF mouse models, and the high expression of miR-328 could reduce the L-type Ca 2+ current and shorten the duration of atrial action potential. Mechanism studies have confirmed that CACNA1C and CACNB1 are the target genes of miR-328, and miR-328 can interact with L-type Ca 2+ channel protein subunits to participate in atrial electroremodeling in AF (51). Nattel et al. found that the expression of miR-26 was down-regulated in the atrial tissues of AF patients, and lowexpressed miR-26 was a potential regulatory gene for the electrophysiological effects of Ca 2+ dependent nuclear factor of activated T cells (NFAT) signaling pathway, and an important participant in the persistence of AF (49). Ricardo et al. found that the high expression of miR-21 in cardiomyocytes of patients with AF was negatively correlated with the expression of CACNA1C and the density of I (Ca, L), suggesting that miR-21 may be involved in the downregulation of L-type Ca 2+ I (Ca, L) induced by chronic AF, and is the key to the persistence of AF (95). Similarly, Qiu et al. found that CACNA1C is a direct target gene of miR-29a-3p, and miR-29a-3p negatively regulates

miRNAs involved in the regulation of structural remodeling
Electrical remodeling is the pathological change in the initial stage of AF, while structural remodeling is the material basis for the long-term maintenance of AF, and it is also the most obvious change of atrium (96,97). Atrial dilatation and fibrosis are the main features of structural remodeling in AF. Atrial fibrosis may lead to slowing of conduction velocity, conduction block to promote reentry and increase susceptibility to AF (98).
Studies have shown that connective tissue growth factor (CTGF) plays an important role in the process of fibrosis. Zhang et al. found that the expression of miR-132 decreased in AF cardiomyocytes. Luciferase assay confirmed that miR-132 could bind to the 3 '-untranslated region of CTGF, thereby inhibiting the expression of CTGF and regulating the fibrosis of cardiac fibroblasts (46). Yang et al. found that overexpression of miR-10a significantly prolonged the duration of AF and decreased Smad7 protein expression. TGF-β1 reversed the inhibitory effect of miR-10a on Smad7, alleviated atrial remodeling, and ultimately inhibited cardiac fibrosis (35). Similarly, Xu et al. found that miR-29b-3p could reduce the degree of atrial fibrosis, and high expression of miR-29b-3p could reduce the expression of fibrosis markers collagen-I and a-SMA, and increase the protein expression of Cx43, thus reversing atrial remodeling (29). Studies have shown that the expression of miR-205-5p is decreased in atrial tissues of patients with AF, and overexpression of miR-205-5p can reduce the expression of TGF-β1, α-SMA, Col III and other fibrosis-related proteins. Mechanism studies have shown that miR-205-5p regulates H3 histone methylation by targeting EHMT2, promotes IGFBP3 expression, and further affects atrial myocyte fibrosis (11). The study found that the expression of miR-29b was low in the atrial tissue of AF rats, overexpression miR-29b can reduce atrial fibrosis, reduce the expression of COL1A1, COL3A1 and TGFβ1, and shorten the duration of AF in rats (14). In addition, the expression of miR-135b was downregulated in AF tissues, while the expression of miR-135b target genes TGFBR1 and TGFBR2 was up-regulated in myocardial fibroblasts. Quercetin can promote miR-135b expression, inhibit TGF-β/Smads pathway, reduce atrial tissue fibrosis and collagen deposition, and thus relieve AF (19). Xu et al. found that miR-101a-3p may prevent AF in rats by targeting EZH2 to inhibit collagen synthesis and atrial fibrosis, which provides a potential target for the prevention of AF (22). miR-1202 was found to negatively regulate atrial fibrosis by targeting nNOS by reducing cell differentiation, collagen deposition, and TGF-β1/Smad2/3 pathway activity (23). Overexpression of miR-133a can inhibit the proliferation and migration of atrial cells, reduce the expression of fibrosis markers and CTGF protein, and improve myocardial fibrosis (24).

miRNAs involved in the regulation of neurohormonal disorders
Autonomic dysfunction is a type of dysfunction that occurs when the balance between sympathetic and parasympathetic nerves is disrupted. cardiac autonomic nerve remodeling (ANR) refers to the changes in the distribution density and spatial arrangement of the autonomic nerve caused by some diseases of the heart (99)(100)(101)(102).
Studies have shown that the contents of tetrahydrobioterin (BH4) and NO are related to nerve regeneration. GCH1 is the rate-limiting enzyme of BH4 synthesis. Hou et al. found that the expression of miR-206 was increased in atrial fibrillation myocarde. High expression of miR-206 could inhibit GCH1, thus affecting the content of BH4 and NO in myocarde (103). In a similar study, miR-206 expression was increased in the the left superior ganglionated plexus (SLGPs). High expression of miR-206 inhibited the expression of superoxide dismutase 1 (SOD1) and increased the levels of reactive oxygen species (ROS) in vitro and in vivo, further exacerbating ANR (45). miR-662 can also regulate the expression of neuropeptides and participate in the occurrence and development of AF after myocardial infarction (17). It was found that the levels of miR-155-5p and miR-24-3p were significantly decreased and the levels of eNOS and NO were increased in patients with AF after ablation compared with those who did not receive ablation therapy (36). Casadei B et al. found that atrial specific upregulation of miR-31 in AF resulted in inhibition of muscular dystrophin (DYS) translation and accelerated degradation of nNOS mRNA, leading to significant reductions in atrial DYS and nNOS protein content and nitric oxide availability. Inhibition of miR-31 restores DYS and nNOS in human AF and normalizes APD and rate dependence of APD (42).

ncRNAs and AF-beyond miRNAs
With the increase of studies on ncRNAs in AF, lncRNAs and circRNAs play an increasingly significant role in AF. Therefore, in addition to miRNAs, this manuscript also discussed the current research content of other ncRNAs in AF. CHA 2 ds2-VASc score was originally used to stratify stroke risk in patients with AF, in order to study whether lncRNAs could improve the predictive ability of CHA 2 ds2-VASc score for stroke. Li et al. added the ability of lncRNA expression level to predict stroke in CHA 2 ds2-VASc scoring model. The results showed that lncRNA H19 plasma expression level was correlated with the risk of stroke in patients with AF, which could significantly improve the ability to predict the risk of stroke in patients with AF, and was a potential prognostic monitoring marker (77). LncRNA GAS5 is significantly down-regulated in the plasma of patients with AF, which is a potential biomarker for the diagnosis and prognosis monitoring of AF (78). Similar studies have found that has_circ_0006314 and hsa_circ_0055387 also have potential predictive value for postoperative AF (79). Fan et al. used GEO database to screen out two different circRNAs. The expression of hsa_circ_0070391 in plasma was up-regulated and hsa_circ_0003935 down-regulated. The area under ROC curve indicated that both of them had high diagnostic efficiency (80). Wang et al. examined plasma circ 8196-RYR2 levels in 136 patients following ablation of AF, suggesting that circ 8196-RYR2 could be used as a new predictor of late recurrence after surgical ablation (81). Another study also show that low expression of circRNA_2773 is a potential diagnostic marker for AF (82).
AF is often accompanied by excessive proliferation of cardiac fibroblasts (CFs). It was found that the expression of HOTAIR was increased in the myocardium of patients with AF, and Ang II significantly increased the activity of atrial fibroblasts. HOTAIR knockdown can significantly inhibit AF cardiac tissue fibrosis by regulating Wnt signaling pathway (9). Knocking down LINC01013 reduced baseline expression of fibrosis markers and their response to TGF-β1. TGF-β1 stimulated atrial fibroblasts to induce the expression of LINC01013, and its knockdown reduced the activation of fibroblasts (56). Plasma H19 levels were significantly higher in patients with AF compared with healthy volunteers. Upregulation of H19 expression contributes to the proliferation and synthesis of extracellular matrix (ECM) related proteins, thereby promoting myocardial fibrosis (53). It was found that the serum TUG1 level was elevated and the expression of miR-29b-3p was low in patients with AF. Pearson correlation analysis showed that TUG1 was negatively correlated with miR-29b-3p expression in AF patients. TUG1 knockdown inhibits vascular endothelium-induced cardiomyocyte proliferation (57). NEAT1 expression was up-regulated in atrial tissues of patients with AF, and was positively correlated with the expression of type I collagen (coll I) and type III collagen (coll III). In addition, the loss of NEAT1 attenuates angiotensin II (Ang II), leading to atrial fibroblast proliferation, migration, and collagen production. These findings suggest that NEAT1 plays an important role in atrial fibrosis and is a new potential molecular target for the treatment of AF (54). In AF patients, LICPAR and TGF-β1 expression were up-regulated and positively correlated. Further analysis showed that Ang II increased LIPCAR, Smad2/3 phosphorylation, and α-smooth muscle actin (α-SMA) levels. Up-regulation of LIPCAR could further promote the promoting effects of Ang II on the phosphorylation levels of LIPCAR, Collagen I, Collagen II, α-SMA and Smad2/3, cell viability and proliferation of atrial fibroblasts. These studies suggest that lncRNA LICPAR regulates atrial fibrosis primarily by regulating the TGF-β/Smad pathway (55). Studies found that downregulation of lncRNA MIAT could significantly relieve AF, increase atrial effective refractory period (AERP), inhibit the expression of fibrosis-related genes coll I, coll III, CTGF, TGF-β1, and effectively reduce AF induced atrial fibrosis (61). PCAT-1 expression was increased in AF patients. PCAT-1 knockdown inhibited the proliferation of AC16 cells. Mechanism studies showed that TGF-β1 was the target of PCAT-1, and its expression in AF tissues was positively correlated with that of PCAT-1. PCAT-1 can promote the proliferation of AF cells by promoting TGF-β1 (58). The expression of GAS5 in myocardium of AF patients was significantly decreased. Overexpression of GAS5 can inhibit the growth of AC16 cells. In addition, further experiments showed that ALK5 was the target of GAS5, and its expression in AF tissue was negatively correlated with that of GAS5. lncRNA GAS5 may inhibit AF cell fibrosis by inhibiting ALK5 (60). The expression of PVT1 in AF patients was increased and positive for coll I and coll III. Overexpression of PVT1 promoted Ang-II-induced atrial fibroblast proliferation, collagen generation, and TGF-β1/Smad signaling activation, while PVT1 knockdown did the opposite. Mechanically, PVT1 acts as a sponge for miR-128-3p and promotes Sp1 expression, thereby activating the TGF-β1/Smad signaling pathway (62).
Hou et al. found that lncRNA TCONS_00075467 may also participate in atrial myocardial electrical remodeling. Interference with TCONS_00075467 can shorten the effective refractory period of the atria in vivo and reduce the duration of L-type calcium current and action potential in vitro (104). Similarly, lncRNA TCONS-00106987 is up-regulated in atrial tissue of patients with AF. Mechanism studies have shown that TCONS_00106987 induces the transcription of its target gene KCNJ2 through miR-26, and increases the inward rectification K + current (IK1). Thus facilitating electrical reconfiguration (59). Studies have shown that interference with lncRNA AK055347 can inhibit the activity of cardiomyocytes, accompanied by the downregulation of Cyp450 and ATP synthase. Mechanism studies have confirmed that AK055347 may regulate the mitochondrial energy production by regulating Cyp450, ATP synthase and MSS51, thus participating in the pathogenesis of AF (64).

Exosome-associated ncRNAs involved in the regulation of AF
In recent years, it has been found that exosome-derived ncRNAs have different expression profiles in various diseases and are a potential non-invasive diagnostic biomarker, which has been widely studied in the medical field. Similarly, exosomes can also be detected in body fluids of patients with atrial fibrillation, and the non-coding RNA carried by them is of great significance for auxiliary diagnosis and prognostic monitoring of AF (105,106).
Wei et al. demonstrated differences in the expression of miRNAs in plasma exosomes in patients with AF. Among them, Xue et al. 10.3389/fcvm.2023.1210762 Frontiers in Cardiovascular Medicine miR-92b-3p, miR-1306-5p and miR-let-7b-3p had significant differences, and gene enrichment analysis showed that these miRNAs and target genes were mainly involved in the occurrence of AF through affecting biological processes such as energy metabolism, lipid metabolism, inflammation and enzyme activity (107). Similar studies have found that miR-483-5p, miR-142-5p and miR-223-3p are also involved in the occurrence and development of AF (108). Joung et al. found that exosomes in the peripheral blood of patients with atrial fibrillation can reduce cardiomyocyte viability, lead to abnormal Ca 2+ channel and induce reactive oxygen species (ROS) production. Highthroughput sequencing found that miR-30a-5p expression was decreased in peripheral blood exosomes of patients with AF, and exosomes with high expression of miR-30a-5p could attenuate pacemaker induced Ca 2+ channel abnormalities (109). Hou et al. screened the differential miRNAs of peripheral blood and exosomes in 40 patients with AF, and found that miR-124-3p was significantly up-regulated, and the high expression of miR-124-3p could improve the viability and proliferation ability of myocardial fibroblasts. Mechanism studies have shown that miR-124-3p can promote the activation and proliferation of fibroblasts through AXIN1 by regulating the WNT/β-catenin signaling pathway (110). Similarly, Exosomal lncRNAs are also potential biomarkers for AF. Joung et al. identified 26 differentially expressed lncrnas in serum exosomes from patients with persistent AF. lncRNAs LOC105377989 and LOC107986997 continued to increase, has significant diagnostic effectiveness for AF, and is a potential biomarker for the diagnosis of AF (106). Lei et al. using GEO database, LINC00636 was found to be an antifibrotic molecule with decreased expression in peripheral blood exosomes of patients with AF. Mechanism studies have shown that LINC00636 can promote the expression of miR-450a-2-3p, thereby inhibiting the expression of MAPK1, and thereby improve cardiac fibrosis in patients with AF (111).

Conclusions
In recent years, with the deepening of research, ncRNAs play an important role in the occurrence and development of AF. Differential expression of ncRNAs in peripheral blood of patients with AF provides a new theoretical basis for auxiliary diagnosis of AF. At the same time, ncRNAs are involved in myocardial cell remodeling and ion channel remodeling, providing a new scheme for the treatment of AF.
This manuscript reviews the research progress of ncRNAs in the occurrence, treatment and potential biomarkers of AF. According to the existing studies, we can find that ncRNAs are closely related to AF and involved in the occurrence and progression of AF, which is worthy of further study and has great clinical significance.

Author contributions
ZX and JZ drafted the manuscript; JL and LW reviewed and edited the manuscript. JD provided ideas. All authors contributed to the article and approved the submitted version.

Funding
This work was supported by the Science and Technology Project of Zhangjiagang City (ZKS2043), Zhangjiagang City Health Youth Science and Technology Project (ZJGQNKJ202113) and Suzhou Science and Technology Development Plan (SKJYD2021006).