The latest progress on miR‐374 and its functional implications in physiological and pathological processes

Abstract Non‐coding RNAs (ncRNAs) have been emerging players in cell development, differentiation, proliferation and apoptosis. Based on their differences in length and structure, they are subdivided into several categories including long non‐coding RNAs (lncRNAs >200nt), stable non‐coding RNAs (60‐300nt), microRNAs (miRs or miRNAs, 18‐24nt), circular RNAs, piwi‐interacting RNAs (26‐31nt) and small interfering RNAs (about 21nt). Therein, miRNAs not only directly regulate gene expression through pairing of nucleotide bases between the miRNA sequence and a specific mRNA that leads to the translational repression or degradation of the target mRNA, but also indirectly affect the function of downstream genes through interactions with lncRNAs and circRNAs. The latest studies have highlighted their importance in physiological and pathological processes. MiR‐374 family member are located at the X‐chromosome inactivation center. In recent years, numerous researches have uncovered that miR‐374 family members play an indispensable regulatory role, such as in reproductive disorders, cell growth and differentiation, calcium handling in the kidney, various cancers and epilepsy. In this review, we mainly focus on the role of miR‐374 family members in multiple physiological and pathological processes. More specifically, we also summarize their promising potential as novel prognostic biomarkers and therapeutic targets from bench to bedside.

mRNAs. 4,5 Because miRNAs imperfectly complement their targets, they are able to interact with tens to hundreds of gene products in various signaling pathways to ultimately regulate cell functions. This also indicates the diversity of miRNAs functions.

MiR-374 family members (including A (a), B (b) and C (c)), as splic-
ing segments of the lncRNA FTX, are located at the X-inactivation center (Xic) of chromosomes Xq13.2 in humans. 8 An increasing number of studies have shown that these members participate in a great diversity of physiological and pathological processes. In this review, we mainly summarize the role of miR-374 family members, a highly conserved miRNA cluster in evolution, in different physiological and pathological processes. More importantly, we also discuss their potential function in the diagnosis and treatment of miR-374 related diseases, especially cancer.

| THE DELIVERY OF MIR-374 AND ITS ROLE IN DE VELOPMENT
In mammalian development, X-chromosome inactivation (XCI) is well-known for epigenetic regulation. 9,10 Mounting studies have shown that female embryos will die without XCI, which corroborates the significance of this epigenetic regulatory mechanism in the development of female embryos. 10,11 The transcriptional silencing of an X chromosome often corrects an imbalance of Xlinked gene dosage. One cis-acting element, named Xic, controls and regulates XCI. There are several transcripts that are located in Xic and can escape XCI, including X-inactive-specific transcript (XIST), TSIX (TSIX transcript, XIST antisense RNA), JPX (JPX transcript, XIST activator), FTX (FTX transcript, XIST regulator) and miRNAs generated from the FTX sequence by cleavage, 12,13 which may lead to gender disparity ( Figure 1). Overwhelming data have implicated that XIST, the master regulator of X-inactivation initiation, is a single and central cis-acting regulator that coordinates imprinted XCI. 11,14 In addition to XIST, FTX is also one of the most abundant transcripts at the stage of preimplantation embryo and is thought to be a positive regulator of XIST. 13,15 It is a well-conserved lncRNA in evolution and includes several conserved miRNAs, such as miR-374, -545 and -421, in its introns. 16 In humans, intron 1 encodes a cluster of 2 miRNAs (MIR-374A and MIR-545), which is absent in rats and mice; intron 5 encodes a cluster of 3 miRNAs (MIR-374B, C and MIR-421) ( Figure 1B). In addition, the sequences of miR-374 family members are highly conserved in different mammalian species, especially in miR-374-5p ( Figure 2).
In recent years, studies have shown that these miRNAs, which are located at Xic, are also imprinted and involved in mammalian development and the generation of paternal sperm. For example, Kobayashi et al 10 reported that miR-374-5p and miR-421-3p mapped adjacent to Xist, were both predominantly expressed in female blastocysts from the Xp chromosome in F1 blastocysts and were indeed imprinted, as determined by allelic expression analysis. In addition, there is another report 17 that miR-374b is markedly decreased in the seminal plasma of azoospermia but is increased in the seminal plasma of asthenozoospermia, as shown by Solexa sequencing and RT-qPCR analysis. The area under the receiver operating characteristic (ROC) curve of miR-374b is 0.839 in the 73 azoospermia cases and 0.813 in the 79 asthenozoospermia cases. The authors point out this is markedly higher than for routine biochemical parameters (0.510-0.622).
Their results revealed that miR-374b level in seminal plasma is a novel, non-invasive approach for diagnosing male infertility.

| THE ROLE OF MIR-374 IN TUMORIG ENE S IS
Most likely because of the advancement of preventive examinations and therapeutic interventions, epidemiological studies have provided strong support for the decrease of the mortality rate due to cancer. Nevertheless, the heterogeneous complex of cancers still remains one of the leading causes of premature death worldwide. 18 Recent reports have verified the presence of ncRNAs, including lncRNA, miRNAs, circular RNAs and so on, in the human cancer. 19,20 We will summarize the role of miR-374 family members in the development and progression of different systematic cancers in this section.

| miR-374 and digestive system carcinoma
The digestive system, as one of the eight systems of the human body, is composed of two parts: the digestive tract and digestive glands. The liver, as one of the important digestive glands, plays an indispensable role in body metabolism, including the storage of glycogen, the synthesis of plasma protein, the decomposition of red blood cells, and the detoxification of toxic substances. Meanwhile, because the liver is a fragile organ, poor protection will lead to the decline of normal function and the development of disease (such as hepatitis and hepatocellular carcinoma). In 2017, Bao et al 21 reported that miR-374 levels were significantly decreased as HBV-related liver fibrosis progressed from S0-S2 to S3-S4. In addition, they identified that miR-374 had a highly diagnostic accuracy in discriminating S0-S2 from S3-S4 using multivariate logistic regression analysis. In recent years, our research group has been unanimously committed to the study of liver disease. In terms of miRNAs, we have previously revealed that the miR-545/374a cluster is up-regulated in HBV-HCC tissue and is significantly correlated with prognosis-related clinical features, including histological grade, metastasis and tumor capsule via the observation of 66 pairs of HBV-related HCC tissue and matched non-cancerous liver tissue specimens. 8 Besides observing the role of miR-374 in liver, some have also investigated its effect on the digestive tract. For example, Wu et al 22 evaluated the potential value of miR-374b as a biomarker in colorectal cancer and found that miR-374b expression was significantly decreased for CRC patients with stage II and stage III disease and may be a novel biomarker for CRC. In addition, Qu et al 23 more comprehensively analysed the function of miR-374b from colon cancer tissues to cell lines. First, they showed that the expression of miR-374b was significantly reduced in colon cancer tissues and cell lines. Second, they investigated the effect and mechanism of miR-374b and found that its overexpression inhibited cell proliferation and invasion, while the role of its knockdown was exactly opposite. Ultimately, they verified that LRH-1 is a direct target of miR-374b via a dual-luciferase reporter assay, and then they showed that miR-374b can suppresses the Wnt signaling pathway through LRH-1 in colon cancer cells.
In recent years, there have also been some reports about the relationship between miR-374 and gastric cancer. Xie J showed that the expression of miR-374b-5p was up-regulated and conducive to gastric cancer cell invasion and metastasis via inhibiting the expression of RECK. 24 Ji et al 25   in primary small cell lung cancer, but they didn't analyze the correlations between its expression and clinical parameters. 30 The latest report 31 confirmed that miR-374 may also participate in the pathogenesis of pituitary gonadotroph adenomas by bioinformatics analysis. However, the author did not discuss the specific mechanism of miR-374 in this disease. In addition to confirming that miR-374 influence the development of tumors, there were some reports that proto-oncogene mutations themselves also affect the expression of miRNAs, including miR-374.

| miR-374 and chemoradiotherapy of tumor
Presently, cancer treatment mainly includes three types: surgical resection, radiotherapy and chemotherapy. In this section, we mainly discussed miR-374 and chemoradiotherapy of tumors. In screened the change of miRNAs with microarray analysis in mouse squamous cell carcinoma line NR-S1, X60 cells (established by irradiating NR-S1 cells with 10 Gy of X-ray radiation once every 2 weeks) and C30 cells (established by irradiating NR-S1 cells with 5 Gy of carbon ion beam radiation once every 2 weeks). They also demonstrated that miR-374c-5p and miR-196a-5p were down-regulated.
When miR-374c-5p was selectively ectopically overexpressed in the human pancreatic cancer cell lines PANC1 and MIA-PaCa-2, these cells were sensitized to carbon ion beam radiation, with no change Therefore, miR-374 may be a novel chemosensitizer and/or radiosensitizer and will be a new potential biomarker for deciding the optimal treatment for cancer.

| miR-374 and epilepsy
Epilepsy is one of the common diseases of the nervous system, and its prevalence is second only to stroke. Accumulated data have shown that miRNAs are involved in various neurological diseases, such as Alzheimer's disease (AD), 56

| miR-374 and other diseases in the nervous system
Hypoxic-ischemic encephalopathy (HIE) is a disease defined as ischemic injury caused by hypoxic asphyxia during the perinatal period. Late diagnosis partially leads to high mortality (approximately 15%-20%) of this diease. Therefore, finding new biomarkers to improve the diagnostic value of neuron-specific enolase (NSE) and S100B protein is especially important. In 2017, Wang et al 66 reported that the expressions of miR-210 and miR-374a, considered to be two important hypoxia-associated miRNAs, were down-regulated in blood samples of HIE newborns compared with those of healthy newborns. Joint analysis of miR-210, miR-374a, S100B protein and NSE help to elevate the diagnostic value and prognostic prediction for HIE by the ROC curves assay.

| THE ROLE OF MIR-374 IN C ARDIOVA SCUL AR DIS E A S E
Studies showed that miR-374 family members can also regulate the pathophysiological process of cardiovascular disease. Early Selenium deficiency has been identified as a causative factor in different kinds of heart failure. Researchers 69 used microarray hybridization to show that there are five up-regulated (>5-fold) miRNAs, which were miR-374, miR-16, miR-199a-5p, miR-195 and miR-30e*, and three down-regulated miRNAs, which were miR-3571, miR-675 and miR-450a*, in rats with selenium deficiency by. And they verified that miR-374 expression was the highest among these up-regulated miRNAs. In the end, they explored that the Wnt/β-catenin signalling pathway was possibly associated with cardiac dysfunction caused by selenium deficiency. However, they did not confirm the targeting relationship between miR-374 and Wnt/β-catenin.
VEGF is a pivotal cytokine that promotes the formation of new blood vessels. However, the VEGF/VEGFR1 signalling pathway regresses cardiac hypertrophy. In contrast, the VEGF/VEGFR2 signaling pathway accelerates cardiac hypertrophy. Lee et al 70 found that miR-374 inhibited the VEGFR1 signalling pathway and activated GPCR signalling pathway by targeting the 3′-UTR of VEGFR1 and cGMP-dependent protein kinase-1 to mediated pro-hypertrophic processes.
In 2016, Licholai et al 71 found that miR-374-5p can maintain vascular integrity by contrasting the different profile of microRNA expression in aneurysmal and unaffected ascending aortic tissue acquired from the same patient. In 2012, Milenkovic 72 observed that miR-374* expression increased after mutagenesis in apoE mice compared to wild-type mice; however, when supplemented with polyphenols in these mice, its expression decreased in apoE. And then they analysed that miR-374* expression presented negative correlations with AKT1.

| THE ROLE OF MIR-374 IN IMMUNE-REL ATED D IS E A S E
CD56 is invariably expressed in normal natural killer cell, 73

a subset of normal T cells and occasionally in T cell acute lymphoid leukemia
(T-ALL). 74 Studies have implied that CD56 is associated with a poor prognosis in lymphoid tumors, including T-ALL. 74  They showed that miR-374b was markedly down-regulated in the T-ALL tissues by microRNA microarray analysis, and the down-regulated miR-374b was greatly associated with worse survival and higher relapse rates in patients with T-ALL. miR-374b overexpression restrained tumorigenicity and cell proliferation, and it accelerated cell apoptosis though targeting Wnt-16 and AKT1, which led to inhibition of AKT signal pathway.
Acute graft-versus-host disease (aGvHD) has a higher mortality rate, the most frequent and serious complication. Expression analy-  Their results confirmed that miR-374a indeed prevented Fas up-regulation by binding with its 3′-UTR to enhance RPE cells survival and protect the cells against oxidative stress. Unterbruner et al 82 found that miR-374a-5p regulate not only the expression of ubiquitin ligase MID1 by binding to the 3′-UTR of the MID1 mRNA but also the mTOR signaling pathway. Therefore, given that dysregulation of MID1 expression is closely associated with multiple diseases including cancer, midline malformation syndromes and neurodegenerative diseases, miR-374a-5p could serve as a potential drug target for future therapy development.

| CON CLUS I ON S AND PER S PEC TIVE S
In conclusion, as shown in Table 1 The therapeutic role of miR-374 family members. In this aspect, we can overexpress or knockdown these members themselves by a variety of methods, such as mimics or inhibitor, ago-or antago-miRNAs, over-expressed or interfering vectors, transgenic or knocking gene and so on. In addition, we can also affect their roles by regulating their targets. However, we found that there were only a small num- different pathological and physiological conditions. This will also be a deficiency in this area.
Therefore, there is still a long way to go until they are used in disease prediction and targeted therapy. So we should accelerate the process of translation of preclinical results into clinic and make them into phase I and II trials to guide clinical diagnosis and treatment. But, this will be a large challenge and hard to pursue in the future. Indeed, the safety (activation of viral delivery systems to immune response, off-target effect of ncRNAs, competition with endogenous miRNAs and multi-targeting of miRNAs) of ncRNAs, as clinical therapeutic targets, needs to be established with certainty.

CO N FLI C T S O F I NTE R E S T
None.