Clinical Significance of MicroRNAs, Long Non-Coding RNAs, and CircRNAs in Cardiovascular Diseases

Based on recent research, the non-coding genome is essential for controlling genes and genetic programming during development, as well as for health and cardiovascular diseases (CVDs). The microRNAs (miRNAs), lncRNAs (long ncRNAs), and circRNAs (circular RNAs) with significant regulatory and structural roles make up approximately 99% of the human genome, which does not contain proteins. Non-coding RNAs (ncRNA) have been discovered to be essential novel regulators of cardiovascular risk factors and cellular processes, making them significant prospects for advanced diagnostics and prognosis evaluation. Cases of CVDs are rising due to limitations in the current therapeutic approach; most of the treatment options are based on the coding transcripts that encode proteins. Recently, various investigations have shown the role of nc-RNA in the early diagnosis and treatment of CVDs. Furthermore, the development of novel diagnoses and treatments based on miRNAs, lncRNAs, and circRNAs could be more helpful in the clinical management of patients with CVDs. CVDs are classified into various types of heart diseases, including cardiac hypertrophy (CH), heart failure (HF), rheumatic heart disease (RHD), acute coronary syndrome (ACS), myocardial infarction (MI), atherosclerosis (AS), myocardial fibrosis (MF), arrhythmia (ARR), and pulmonary arterial hypertension (PAH). Here, we discuss the biological and clinical importance of miRNAs, lncRNAs, and circRNAs and their expression profiles and manipulation of non-coding transcripts in CVDs, which will deliver an in-depth knowledge of the role of ncRNAs in CVDs for progressing new clinical diagnosis and treatment.


miRNAs and CVDs
The miRNA lin-4 was identified in 1993 in Caenorhabditis [71]. The synthesis of miRNA occurs in the nucleus and is transcribed by RNA polymerase II into coding and noncoding, capping polyadenylated pri-miRNAs. The pre-miRNA produces a hairpin-like structure and is timed by the Drosha nuclear enzyme, then transported to the cytoplasm [12]. The DICER removes the terminal loop of the pri-miRNAs, resulting in a 20-25 nucleotide base pair dsRNA complex. The dsRNA, as attached to the miRNA-linked RISC (RNAinduced silencing complex), targets mRNAs and results in mRNA de-adenylation and translational repression (Figure 3) [13]. In vivo and in vitro studies have shown that miRNA plays a critical role in the regulation of CVDs such as CH, HF, ARR, ACS, MI, AS, RHD, and PAH (Table 2) [14].        Myocardial hypertrophy (MH) is caused by the development of CVDs, including stenosis of the heart valve and hypertension, and causes HF and death [75]. Several miRNAs, including miR-208a, miR-19a/b, miR-34a, miR-145, miR-150, miR-378, and others, are involved in the development of MH [76]. MiR-378 is an anti-MH miRNA and regulates the Igf1r (insulin-like growth factor receptor), Grb2 (growth factor receptor binding protein 2), and Ksr1 (Ras kinase inhibitor 1) [77]. MiR-185 regulates cardiac cell proliferation and is related to the signal transduction mechanism. MiR-34a regulates the Agt9a gene, which is involved in autophagy. The transcription activator p300 is regulated by miR-150 [78]. MiR-1 is involved in the growth and development of cardiomyocytes by reducing the expression of GATA-binding protein 4 (GATA4) and calmodulin Mef2a, which regulate the calcium signal pathway and protein expression and could be targeted for diagnosis and therapy [79].

miRNAs and ACS and MI
ACS (acute coronary syndrome) is developed by reduced blood flow in the heart, an immediate blockage of the coronary arteries, and localized heart necrosis, all contribute to the development of AMI (acute myocardial infractions) [86]. AMI patients have a high level of miR-1 expression. MiR-1, miR-133a, and miR-208a levels have been found to be higher in AMI patients. Cardiac arrest is regulated by miR-208b and miR-499-5p in patients with coronary artery bypass grafting [87]. These two miRNAs are expressed by dysregulated cardiac muscles. A reduced level of expression has been shown in AMI patients. The expression profile of all these miRNA regulations can be used for early diagnosis and treatment [88]. High miR-208 expression levels have been observed in a mouse model with AMI. High-throughput analysis of miRNA expression in patients with AMI can be explored further for sensitive and specific early diagnosis and treatment [89].

miRNAs and Atherosclerosis
The miRNA plays an important role in the generation of atherosclerosis by vascular angiogenesis, endothelial dysfunction, lipid accumulation, local inflammation, calcification, thrombosis, and endothelial dysfunction [90]. Play important in the development of CAD (coronary artery disease), causes significant death at global level. Expression profile of miRNA has been investigated in patients with AS. MiR-33 regulates the AS disease progression by involving the inflammatory response, cell cycle progression, lipid metabolism, and proliferation [91]. In patients with AS, miR-122 is substantially expressed. MiR-122 controls the levels of high-density lipoprotein (HDL) and low-density lipoprotein (LDL).Leukocyte aggregation on endothelial cells is triggered by miR-126-mediated upregulation of VCAM-1 (vascular cell adhesion molecule-1) [92]. Mi-R1 regulates the signalling pathways for MLCK (Myosin Light Chain Kinase) and ERK (Extracellular Signal-Regulated Kinase). MiR-221 and miR-222 control the growth and development of vascular smooth muscle cells (VSMCs). In patients with AS, there is generally less miR-126, miR-1, and miR-221/222 expression [93].

miRNAs and RHD
RHD lesions have primarily been found in the mitral valve. RHD tissue and plasma samples have significant levels of miRNA-1299 and miRNA-1183 expression. MiR-328-3p is found in RHD and AF (atrial fibrillation) patients [94]. MiRNA-432 expression levels have been found to be lower in RHD patients. All these microRNAs could be used for early diagnosis. Further investigations are needed to find out more about miRNA regulations in RHD [95].

LncRNAs and Cardiovascular Diseases
LncRNAs are more complex and heterogeneous in nature in comparison to miRNAs, which regulate gene expression. LncRNAs are involved in CVDs and categorized into various classes based on their structure and functions, including bidirectional lncRNAs, enhancer lncRNAs, sense lncRNAs, antisense lncRNAs, intergenic lncRNAs, and intron lncRNAs [96]. The gene expression level is changed by the interactions of lncRNA with DNA, RNA, proteins, elements of the chromatin modification complex, and transcription factors. Guided lncRNAs can either activate lncRNA processes or suppress gene expression by delocalizing regulatory elements [97][98][99]. Ribonucleoprotein (RNP) complex formation involves the scaffold lncRNAs ( Figure 4). The lncRNAs serve as primary miRNA precursors that are converted into mature miRNAs while the miRNA precursor is suppressed. Longrange gene regulation begins when the lncRNA activates transcription from regulatory areas of the genome. LncRNAs interact with miRNAs and disrupt the RNA molecules' regulatory system ( Figure 5) [97]. lncRNAs also act as a maternal or paternal genomic imprinting expression and help in the development of organisms [100,101]. Regulations and clinical importance of lncRNA in cardiovascular are shown in Table 3.
precursors that are converted into mature miRNAs while the miRNA precursor is suppressed. Long-range gene regulation begins when the lncRNA activates transcription from regulatory areas of the genome. LncRNAs interact with miRNAs and disrupt the RNA molecules' regulatory system ( Figure 5) [97]. lncRNAs also act as a maternal or paternal genomic imprinting expression and help in the development of organisms [100,101]. Regulations and clinical importance of lncRNA in cardiovascular are shown in Table 3.  The integration of various types of cells, the vascular system, and blood vessels are all involved in the generation of the heart [98]. lncRNAs, also known as super-enhancer lncRNAs (SE-lncRNAs), control transcription at the tissue and cell levels. MyoD is an important transcription factor that involves muscle cell differentiation along with other core transcription factors [99]. The CE (core enhancer element) is produced by CERNA, which acts as a positive feedback regulator. It has been recently observed that various     The integration of various types of cells, the vascular system, and blood vessels are all involved in the generation of the heart [98]. lncRNAs, also known as super-enhancer lncRNAs (SE-lncRNAs), control transcription at the tissue and cell levels. MyoD is an important transcription factor that involves muscle cell differentiation along with other core transcription factors [99]. The CE (core enhancer element) is produced by CERNA, which acts as a positive feedback regulator. It has been recently observed that various types of lncRNA are involved in the development of CVDs, including CHRF, Myh7, LIPCAR, MIAT, Carl, LIPCAR, ASB9P1, RP11-218 M11.6, G078882, G064270, G000678, G030563, H19, TUG1, PFL, MIAT, AK081284, HOXA11-ASz, NRON, and GAS5 [96]. H19 is expressed during embryogenesis and CVD but is repressed after birth. miRNA-675 acts as a negative regulator in cardiac hypertrophy. miR-675-3p and miR-675-5p are upregulated in cardiac hypertrophy [100]. Some pro-hypertrophic factors are also involved in CH and are mediated by Ca/calmodulin-dependent protein kinase IIδ (CaMKIIδ). The lncRNA-miRNA-mRNA axis can be a potential target for therapeutic approaches [101]. All these investigations have confirmed that lncRNAs play major roles in cardiovascular biology and diseases (Table 3) [102].

Arterial Hypertension
There are different kinds of lncRNAs that control vascular tone to control the pathophysiology of AH (Table 3). LncRNAs contributor AH controls VSMC dysfunction, and miRNAs 221 and 222, which control lnc-Ang362, control the growth of VSMCs [158,159]. The lncRNAs NR_027032, NR_034083, and NR_104181 act as biomarkers for disease diagnosis in AH. Differential expression of lncRNAs in rats exposed to a peptide hormone (Angiotensin-II) with a variety of functions, including inflammation, fibrosis, vasoconstriction, and hypertrophy/hyperplasia [140]. The silencing of lnc-Ang362 reduced the expression of miRNAs, which led to a reduction in VSMC proliferation. The lncRNA GAS5 (growth arrest-specific 5) regulates vascular remodelling in hypertension and is expressed primarily in ECs and VSMCs [160]. The proliferation of VSMC lncRNAs NR4A3 and AK098656 induces the oxidative stress-induced proliferation of VSMCs. CDKN2B-AS1 antisense lncRNA susceptibility to the development of AH [161].

Coronary Heart Disease
Plaque formation in CVDs is caused by a chronic inflammatory process that narrows the vessels and reduces blood flow, resulting in atherosclerosis and ischemia, which lead to the development of CHD (Table 3) [162]. There are various lncRNAs that are linked to the pathophysiology of CHD and act as biomarkers, including MIAT, MALAT1, ANRIL, LIPCAR, MALAT1, MIAT, and SMILR. HOTTIP, a lincRNA-p21, regulates cell proliferation and apoptosis (Table 3) [163]. The lncRNA BANCR is associated with CHROME (the cholesterol homeostasis regulator of miRNA expression). lncRNA NEXN-AS1 is involved in the mitigation of atherosclerosis. All these RNAs may be targeted for the development of diagnosis and treatment [164].

Acute Myocardial Infarction
AMI is caused by a blockage in blood flow, less oxygen in the blood, and metabolic disorders. There are various lncRNAs involved in potential signalling in AMI (Table 3) [165]. In a rat model, the lncRNA MIAT is involved in myocardium degeneration and regulates adverse heart remodelling [165]. H19 is involved in autophagy, HOTAIR is involved in the cardioprotective activity and sponging with miRNA, KCNQ1OT1 in left ventricle dysfunctions, MALAT1 regulates cardiomyocytes, MDRL in mitochondrial dysfunctions, MEG3 and MEG3 in cardiomyocyte apoptosis, MIAT regulates CH, Mirt1/2 regulates cardiac remodelling [102], NONRATT021972 regulates cardiac activity, PCFL regulates cardiac fibroAll these lncRNAs have shown great importance in the regulation of pathological conditions, diagnosis, and treatments.

Heart Failure
In chronic conditions, the heart does not pump systolic and diastolic properly, which may increase shortness of breath, fatigue, inflammation, and a fast heartbeat [166]. There are various lncRNAs involved in HF, including lncRNAs MHRT, CHAER, CHRF, APF, CARL, aHIF, MIAT, and CHAST [167]. MHRT is a cluster of specific RNA that plays an important role in maintaining the physiological conditions of the heart. Reduced expression of MHRT was observed in HF conditions, isoforms of MHRT, Myh6, and Myh7 are involved in HF [144]. During a cardiac stress response, Brg1 regulates the homeostasis of the heavy chain from -myosin to -myosin. The overexpression of CHRF induces a pathological process and induces apoptosis. The lncRNA CHAER is involved in cardiac remodelling and is expressed during HF in the heart of mice [147]. The long noncoding RNA PCR2 has shown cardioprotective properties in animal models. APF regulates autophagy, CARL regulates angiogenesis, aHIF regulates apoptosis, and MIAT regulates the cardiac fibrosis process in MI [145]. Vascular remodelling in arterial hypertension is regulated by GAS5 and AK0986656 lncRNAs. Studies on all these lncRNAs under investigation could be major targets for diagnosis and treatment before HF (Table 3) [145].

Circular RNA and Cardiovascular Diseases
CircRNAs are SS-RNAs, covalently closed and forming a loop; the 3 and 5 ends of RNA molecules are joined [168]. CircRNAs regulate gene expression and code for proteins; circRNA has been linked to some diseases, such as cancer, diabetes, cardiovascular disease, and others ( Figure 6) [169]. CircRNAs play an important role in the regulation of molecular mechanisms in the cardiovascular system (Table 4). cZNF292 is the firstidentified circRNA in endothelial cells (ECs) and controls angiogenesis [170]. CAD and SMCs (smooth muscle cells) are regulated by circRNA ANRIL; it also regulates proliferation and apoptosis by interfering with the rRNA (ribosomal RNA) maturation process [171]. The vascular SMCs are regulated by circLrp6; another circFoxo3a is involved in cardiac functions, and cardiac muscles are required by circFndc3c after MI. Heart-related circular RNA (HRCR) circRNA and CDR1 act as miRNA sponges [172]. CDR1 is involved in the development of post-myocardial infractions in mice via sponging miR-7. CDR1 is associated with the LV (left ventricular) and RV (right ventricular) and is used as a clinical biomarker for disease diagnosis. CircYOD-1 is used as a biomarker for coronary artery disease [173]. Some circRNAs are observed in paediatric patients with CHD, including HSA circRNA 004183, HSA circRNA 079265, and HSA circRNA 105039, among others. Three circRNAs, namely DNAJC6, TMEM56, and MBOAT2, are identified in patients with hypertrophic cardiomyopathy (HCM). All these biomarkers could be used for diagnosis [174].

circRNA in Atherosclerosis
VSMCs and VEC are involved in the pathogenesis of AS; the circRNA-miRNA-mRNA axis plays a major role in the development of AS [193]. The circRNA hsa-circ-0054633 protects the endothelial cells or tissues from injury by hyperglycemic conditions. Overexpression of circRNA hsa-circ-0054633 induces regulation of miR-218 and regulates the miR-218/HO-1 and miR-218/ROBO1 axis. By regulating the expression of the miR-186/HIF-1 axis, hsa-circ-0010729 increased apoptosis and prevented the proliferation of hypoxiainduced HUVEC (human umbilical vein endothelial cells), reducing endothelial injury [194]. The circRNA circWDR77 prevents the proliferation and migration of VSMCs by regulating miR-124 and FGF-2 (fibroblast growth factor 2). The miR-661/SYK axis activates proliferation and migration by circRNA circ-RUSC2 [195]. All these regulations of circRNA could be major targets for early diagnosis and treatment.

Coronary Heart Disease (CHD)
The CHD is regulated by various types of circRNAs, which play an important role in pathological mechanisms, including circ-SATB2, SM22α, circ-171, and circ-624. Circ-SATB2 has been reported to be upregulated in the proliferative mode of VSMCs and downregulated in contractile VSMCs [169]. There are various lncRNA types that regulate vascular tone to regulate the pathophysiology of AH. Giver AH, a lncRNA, regulates VSMC dysfunction, and miRNAs 221 and 222, which regulate lnc-Ang362, regulate VSMC expansion [196]. These investigations suggest that circ-RNA plays a major role in CHD and could be used as a diagnostic marker.
for disease diagnosis. CircYOD-1 is used as a biomarker for coronary artery disease [173]. Some circRNAs are observed in paediatric patients with CHD, including HSA circRNA 004183, HSA circRNA 079265, and HSA circRNA 105039, among others. Three circRNAs, namely DNAJC6, TMEM56, and MBOAT2, are identified in patients with hypertrophic cardiomyopathy (HCM). All these biomarkers could be used for diagnosis [174]. Figure 6. circRNA biogenesis is shown schematically. The production of circRNAs is possible by a direct backsplicing mechanism in which intronic reverse complementary sequences or RNA-binding protein (RBP) pairing, which brings the sequences close together to promote circularization and drive circularization. CircRNAs can also be produced by a mechanism called lariat-driven, which is characterized by a lariat structure with flanking sequences that help produce circRNAs. circRNAs are exported via the nuclear pore, act as mRNA sponges (CircHRCR, CircZNF609, CircNrg), stabilize protein complexes (USP36, CircLrp6, CircNrg1), and are involved in other functions. Figure 6. circRNA biogenesis is shown schematically. The production of circRNAs is possible by a direct backsplicing mechanism in which intronic reverse complementary sequences or RNA-binding protein (RBP) pairing, which brings the sequences close together to promote circularization and drive circularization. CircRNAs can also be produced by a mechanism called lariat-driven, which is characterized by a lariat structure with flanking sequences that help produce circRNAs. circRNAs are exported via the nuclear pore, act as mRNA sponges (CircHRCR, CircZNF609, CircNrg), stabilize protein complexes (USP36, CircLrp6, CircNrg1), and are involved in other functions.

Cardiac Ischaemia/Reperfusion (I/R) Injury and Myocardial Infarction (MI)
Treatment of MI is based on the pathological conditions of cardiac ischemia (CI) and reperfusion; CI leads to mitochondrial dysfunction and induces apoptosis of cardiac muscle cells. Recent research has shown that the circRNA-miRNA-mRNA axis plays an important role in CI and R injuries and MI [197]. CircumNCX1 increases ROS expression in CI and R injury and regulates apoptosis via the circNCX1-miR-133a-3p-CDIP1 axis [198]. The overexpression of CDIP1 also induces the apoptosis process. The sponging miR-652-3p is regulated by circ-MFACR, which induces the expression of MTP18, which leads to mitochondrial dysfunction and apoptosis of cardiomyocytes [199].

Heart Failure (HF)
HF is mainly caused by MI and cardiac hypertrophy (CH). The development of CH is prevented by circ-HRCR via endogenous regulation of miR-223. Expression of another circRNA, circ-Nfix, induces the regeneration capacity in mice after MI by using the miR-214/Gsk3β axis [200]. The increased expression of circSLC8A1 has been observed in CH, but at the same time, the expression of miR-133a has been downregulated. The diagnosis and therapeutic targets of [201] could be a promising approach to treating CH.

Cardiomyopathies (CM)
CM is caused by a group of cardiac diseases (CD), which dysregulate cardiac functions and lead to HF and high morbidity and mortality reported at the global level [202]. The high-throughput sequencing analysis data has been investigated in patients with dilated DCM (dilated cardiomyopathies), and 9585 circRNAs were differentially regulated. One of the most prevalent circRNAs, CircSLC8A1, is crucial for CM differentiation, cardiac development, and homeostasis [203]. The pathophysiology of CMs is greatly influenced by the cardiac-specific expression of circALPK2, circSPHKAP, circCACNA1D, and circSLC8A1 and acts as a diagnostic biomarker [169]. The highly expressed circRNAs in DCM were regulated by the BTBD7, NHLRC2, FAT1, LYPLAL1, DHX40, DHX40, ICA1, TTN, and PKN2 genes [183]. The circRNAs involved in DCM are described in Table 4. All these investigations have proven the importance of circRNAs in CM [192].

circRNAS in Cardiac Regeneration
Various types of circRNAs are involved in cardiac regeneration (CR), reprogramming, and proliferation of the myocardium ( Table 4). The circNfix regulates proliferation and angiogenesis and improves the cardiac function of cardiomyocytes in a mouse model [204]. Overexpression of CircCDYL promotes the CR, inhibition of CircCDYL induces the proliferation of fibroblasts, CircHIPK3 promotes cellular growth, and CircHIPK3 induces angiogenesis in the myocardium in mice [204]. The diagnosis and therapeutic importance of circRNAs in CVDs need to be explored further.

Clinical Investigations of Cardiovascular Non-Coding RNA Therapies
Clinical investigations of cardiovascular non-coding RNA therapies are under investigation, including CM hypertrophy, altered excitation-contraction coupling, cell death, interstitial fibrosis, and microvascular rarefaction [10][11][12][13][14]96,193,205]. The novel ncRNA therapeutics under clinical investigation are shown in Table 5 [206]. In order to silence the miRNAs responsible for cardiac hypertrophy, a typical strategy has been to use 2 -OMe-modified (2 -O-Methyl (2 -OMe)-4 -thioRNA) antagomiRs or LNAs (locked nucleic acid-modified), with the earliest evidence suggesting in vivo silencing originally stated in mice [207,208]. Several miRNAs that operate on either CMs or fibroblasts have been demonstrated to suppress hypertrophy and remodelling during experimental HF in mice [208]. These include miR-133, miR-199a-5p, miR-21, miR-23a, miR-24, mir-29, miR-34a, and miR-25. Other studies reported success with LNA ASOs, e.g., against mir-34, miR-652, miR-208a, miR-154, miR-29, and miR-21 in pigs. Other cell-type-specific strategies might also be successful. Mice with chronic pressure overload had less cardiac inflammation, hypertrophy, and dysfunction after leucocyte-expressed miR-155 was inhibited [209]. An LNA that targets miR-26a was intravenously administered, and this quickly stimulated angiogenesis, decreased the size of MIs, and enhanced heart performance [207,208]. Osteopontin, a protein that encourages fibrosis and hypertrophy, was the target of an RNA aptamer that prevented or reversed pressure overload-induced coronary artery disease in mice [207]. The inhibition of endothelial miR-24 reduced the size of MI by decreasing endothelial apoptosis. The lncRNA Chast was effectively silenced by GapmeR, which prevented or reduced stress overload-induced pathological cardiac remodelling as one of the first examples of therapeutic lncRNA targeting in the cardiovascular sector [208]. The lncRNA Meg3 reduces cardiac fibrosis and improves diastolic function after chronic pressure overload. Wisper is a lncRNA that is actively involved in preventing heart dysfunction and fibrosis induced by MI, and lncRNA H19 prevents the growth of pulmonary arterial hypertension or aortic aneurysms [205,206]. Various ncRNA medications are under development to address various aspects of hyperlipidemia as a result of the ease with which the liver can be targeted [206]. The LPA (Lipoprotein A) is a significant target that is currently unaffected by small-molecule medications but is a major carrier of oxidized phospholipids in human plasma and a causal risk factor for atherosclerotic CVD and aortic stenosis [205]. Apo(a), the LPA gene's product, is covalently linked to ApoB-100 in lipoprotein(a). Pelacarsen (formerly AKCEA-APO(a)-LRx), an ASO-targeting LPA mRNA, decreases Lp(a) levels by up to 80% and is well tolerated aside from injection-site responses. Aantisense oligonucleotide targeting LPA mRNA (which encodes the main Lp(a) constituent, apolipoprotein(a)) conjugated with triantennary N-acetylgalactosamine to directs the therapy [206]. It is administered subcutaneously two to four times per week. Patients with CVD are now participating in Phase 3 trials for it (NCT04023552) [206]. Olpasiran, a GalNAc-conjugated siRNA that lowers Lp(a), is currently being studied in Phase 2 (NCT04270760), and SLN360, another GalNAc-conjugated siRNA, is being tested in Phase 1 (NCT04606602). Angiopoietin-like 3 (ANGPTL3) is another intriguing hyperlipidemia target, as evidenced by research showing that loss-of-function variations are linked to significantly lower LDL-cholesterol and triglyceride levels and a decreased risk of CHD [206]. The circulating lipoprotein lipase and endothelial lipase inhibitor ANGPTL3, which is mostly made in the liver, influences the uptake of muscle-free fatty acids, adipose tissue lipogenesis, and LDL and residual cholesterol by the liver [206,207]. The latter effects are significant since they are independent of the LDL receptor, indicating that ANGPTL3 reduction should be helpful in FH patients. with mutations in the LDL receptor. In a Phase 2b study (NCT04516291), Vupanorsen, a GalNAc-modified ASO-targeting ANGPTL3 mRNA, was tested in diabetic patients with hepatic steatosis and hypertriglyceridemia [205][206][207][208]. It reduced triglycerides, ApoC3, and residual cholesterol by 38-58%. Finally, ARO-ANG3, a GalNAc-conjugated siRNA that targets ANGPTL3 as well, has been tested in a larger Phase 2 trial (NCT04832971) after being evaluated in a smaller open-label study in heterozygous FH87 [206]. Inhibition of miR-92a with antisense oligonucleotides improves wound healing, speeds up re-endothelialization, and avoids endothelial dysfunction and atherosclerosis in murine and porcine models after MI and hind limb ischaemia. In pigs, administration of an anti-miR-92a LNA ASO via catheter significantly decreased infarct size and enhanced heart function. An LNA ASO targeting miR-132-3p for patients with HF was also reported to have first-in-human data [206].

Future Prospective
Small nucleic acid therapies provide an interesting new CVD treatment option and an opportunity to target disease pathways that have yet to be addressed by small molecule therapeutics. Even though ncRNA biomarker research is developing quickly, there are still a number of difficulties since preanalytical and analytical factors might affect the quality of results. Despite this apparent potential, there remain a number of challenges that must be resolved before ncRNA treatments may be used more widely in clinical settings. Developments in both carrier and modifications to RNA that would allow effective cellular absorption and intracellular stability of the molecules remain a big concern; at least until effective RNA modifications can be made, lipid-based nanocarriers now appear to be the most promising method. The prevention of endogenous activity by inhibitory miRNAs, lncRNAs, and circRNAs is currently possible by using ASO-containing LNA-MN (locked nucleic acid-modified nucleotides). LNAs against miR-15171 and AAV (adenoassociated virus) vectors expressing antisense sequences can be used to induce cardiac activity. Regeneration and cardiac activity could be enhanced by the delivery of miRNAs, lncRNAs, and circRNA mimics, which may improve the development of cardiac activity after MI. The intracardiac injection can be used to mimic miRNAs, lncRNAs, and circRNAs using various types of nanoformulations, including lipids and hydrogels. The application of cell-specific synthetic ncRNA, or ASO, may be helpful in the clinical management of CVDs. Therefore, further efforts are required to develop sensitive, specific ncRNA based on a spatiotemporal delivery approach for therapeutic interest in CVDs. There are several delivery methods used to mimic miRNAs, lncRNAs, and circRNAs, each with its own set of off-target limitations and toxicity. The selection of the material, sample isolation, detection, and processing methods, as well as normalization procedures and the impact of medications and other noncardiac disorders and phenotypes, is some of these variables. The ncRNAbased therapeutic approach is required to overcome off-target toxicity for effective clinical allocation in CVDs. The development of applications and methods related to ncRNA is still under investigation. We need to explore genome-wide analysis; next-generation sequencing and animal model studies will explore more about unknown ncRNAs. We can focus on the conserved nature of ncRNA in humans, cells, and tissue-specific regulations. Moreover, we can focus on circulating ncRNA body fluids, including serum, plasma, and urine, which can be used for diagnosis, prognosis, and treatments based on clinical signs and symptoms in patients with CVDs. CircRNA could be more effective for diagnosis and prognosis. Therefore, the biological and clinical importance of ncRNA in CVD remains unknown. Many unknown functions need to be clarified. Many undiscovered functions are still being developed. Therefore, a thorough investigation of the function of ncRNAs in CVD will lay the groundwork for novel clinical disease treatment initiatives.

Conclusions
We have reviewed significant recent advancements and clinical perspectives of ncRNA research, with special emphasis on major problems related to the conversion of preclinical and basic scientific discoveries into innovative diagnostics and therapies with clinical use. To provide accurate and repeatable results in diagnostics, it is necessary to remove the ncRNA heterogeneity based on technical and analytical factors, use reliable extraction techniques, and standardize the process. This is crucial to automate work processes and set the stage for clinical routine. Large-scale clinical trials are required to accurately evaluate the potential of ncRNAs as clinical therapeutic agents because reported results are often validated. Future clinical applications must carefully evaluate established selection criteria, appropriate clinical outcome metrics, and patient cohorts in which one pathomechanism is the single or main cause of disease development and progression. Future clinical trials will consequently likely focus on patients for whom the extra effort of ncRNA treatment techniques is likely to result in clinical success.

Conflicts of Interest:
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.