Spinal muscular atrophy (SMA) is a severe neurodegenerative disorder caused by deficiency of survival of motor neuron (SMN). While significant progress has been made in SMA therapy by rescuing SMN expression, limited knowledge about SMN downstream genes has hindered the development of alternative therapies. Here, we conducted whole-transcriptome sequencing of spinal cord, heart, and liver tissues of a severe SMA mouse model at early postnatal ages to explore critical coding and non-coding RNAs (ncRNAs). A large number of differentially expressed RNAs (DE-RNAs) were obtained, including 2,771 mRNAs, 382 microRNAs (miRNAs), 1,633 long ncRNAs, and 1,519 circular RNAs. Through in-depth data mining, we unveiled deregulation of miR-34a in all tissues. Analysis of competitive endogenous RNA networks of DE-RNAs identified multiple novel targets of miR-34a including Spag5 mRNA, lncRNA00138536, and circRNA007386. Further in vitro studies using mouse myoblast and human cardiomyocyte cell lines showed that knockdown of SMN upregulated miR-34a-5p and overexpression of miR-34a-5p alone disrupted cell-cycle progression through regulating its targets, recapitulating gene expression patterns observed in cardiac tissue of SMA mice. Our results identified a critical miRNA involved in SMA pathology, which sheds insights into the molecular basis of widespread tissue abnormalities observed in severe forms of SMA.

Non-coding RNAs (ncRNAs) have gained significant attention in recent years due to advancements in biotechnology, particularly high-throughput total RNA sequencing. These developments have led to new understandings of non-coding biology, revealing that approximately 80% of non-coding regions in the genome possesses biochemical functionality. Among ncRNAs, circular RNAs (circRNAs), first identified in 1976, have emerged as a prominent research field. CircRNAs are abundant in most human cell types, evolutionary conserved, highly stable, and formed by back-splicing events which generate covalently closed ends. Notably, circRNAs exhibit high expression levels in neural tissue and perform diverse biochemical functions, including acting as molecular sponges for microRNAs, interacting with RNA-binding proteins to regulate their availability and activity, modulating transcription and splicing, and even translating into functional peptides in some cases. Recent advancements in computational and experimental methods have enhanced our ability to identify and validate circRNAs, providing valuable insights into their biological roles. This review focuses on recent developments in circRNA research as they related to neuropsychiatric and neurodegenerative conditions. We also explore their potential applications in clinical diagnostics, therapeutics, and future research directions. CircRNAs remain a relatively underexplored area of non-coding biology, particularly in the context of neurological disorders. However, emerging evidence supports their role as critical players in the etiology and molecular mechanisms of conditions such as schizophrenia, bipolar disorder, major depressive disorder, Alzheimer's disease, and Parkinson's disease. These findings suggest that circRNAs may provide a novel framework contributing to the molecular dysfunctions underpinning these complex neurological conditions.

Stroke refers to an abrupt neurological deficit, caused by an acute focal injury of the central nervous system via infarction or hemorrhage due to impaired vascularity, and remains among the leading causes of disability and death worldwide. Stroke is often preceded by an episode of neuronal deficit termed transient ischemic attack (TIA), which presents an effective opportunity for mitigating the risk of an eminent acute ischemic stroke (AIS). Circulating non-coding RNAs (ncRNAs) have emerged as important biomarkers for stroke, but PIWI-interacting RNAs (piRNAs), a class of small regulatory ncRNAs, have not been previously explored as diagnostic or prognostic biomarkers for stroke.

We conducted comprehensive circulating piRNA profiling of AIS and TIA patients using RNA-seq on serum samples collected within 24 h of clinical diagnosis. The study cohort was divided into discovery and cross-validation datasets to identify replicated piRNAs using stringent analysis cut-offs. The expression levels of the panel of differentially regulated piRNAs between AIS and TIA patients were also compared with healthy controls.

We identified a panel of 10 differentially regulated piRNAs between AIS and TIA patients; hsa-piR-28272, -piR-32972, -piR-28247, -piR-24553, -piR-24552, -piR-28275, -piR-28707 and -piR-32882 were upregulated, while hsa-piR-23058 and -piR-23136 were downregulated in AIS patients. Moreover, these 10 piRNAs were also differentially expressed in AIS patients compared to healthy controls. In addition, we investigated the potential gene targets of the dysregulated piRNAs and their plausible involvement in pathophysiological processes affected in stroke.

The imbalances in the circulating piRnome of AIS and TIA patients presented herein provide important insights into the roles of piRNAs following ischemic brain injury and potentially provide opportunities to mitigate stroke-induced mortality and morbidity.

Environmental factors affect not only paternal condition but may translate into the following generations where sperm-mediated small RNAs (sRNAs) can contribute to the transmission of paternal effects. sRNAs play a key role in the male germ line in genome maintenance and repair, and particularly in response to environmental stress and the resulting increase in transposable element (TE) activity. Here, we investigated how the social environment (high competition, low competition) of male zebrafish Danio rerio affects sRNAs in sperm and how these are linked to gene expression and TE activity in their offspring. In a first experiment, we collected sperm samples after exposing males to each social environment for 2 weeks to test for differentially expressed sperm micro- (miRNA) and piwi-interacting RNAs (piRNA). In a separate experiment, we performed in vitro fertilisations after one 2-week period using a split-clutch design to control for maternal effects and collected embryos at 24 h to test for differentially expressed genes and TEs. We developed new computational prediction tools to link sperm sRNAs with differentially expressed TEs and genes in the embryos. Our results support the idea that the molecular stress response in the male germ line has significant down-stream effects on the molecular pathways, and we provide a direct link between sRNAs, TEs and gene expression.

Porcine epidemic diarrhea virus (PEDV) is a primary cause of viral diarrhea in neonatal piglets, leading to substantial economic losses in the swine industry globally. It primarily targets epithelial cells of the small intestine, compromising intestinal function and resulting in the death of affected animals. As mitochondria are essential for maintaining gut health, this study investigates the effects of PEDV infection on mitochondrial function in small intestinal epithelial cells and its subsequent impacts. Using small RNA sequencing, fluorescence in situ hybridization, dual luciferase reporter assay, gene overexpression, and silencing experiments, we investigated the mitochondrial structural and functional impairments induced by PEDV infection in jejunum epithelial cells of piglets and characterized the regulatory pattern of miRNAs in mitochondria of jejunum epithelial cells during PEDV infection. The results indicate that PEDV infection leads to the upregulation and mitochondrial localization of the nuclear-encoded microRNA, miR-34c, which in turn suppresses COX1 expression. The activation of the miR-34c/COX1 axis diminishes mitochondrial complex III, IV, and V activities, depletes ATP, lowers mitochondrial oxygen consumption, induces mitochondrial depolarization, increases the accumulation of mitochondrial reactive oxygen species (mtROS), and stimulates mitophagy. Furthermore, we confirm that CREB3L1 acts as an upstream transcription factor regulating the miR-34c/COX1 axis during PEDV infection, modulating mitochondrial damage in the epithelial cells of the jejunum. These findings demonstrate for the first time that PEDV infection activates the miR-34c/COX1 axis via the transcription factor CREB3L1 and regulates the nuclear-mitochondrial communication and mitochondrial fate, providing a new perspective on the pathogenesis of PEDV.IMPORTANCEThis study reveals the mechanism by which the porcine epidemic diarrhea virus (PEDV) disrupts mitochondrial function in piglets, enhancing viral pathogenicity. By demonstrating how PEDV infection upregulates miR-34c, leading to COX1 suppression and subsequent mitochondrial dysfunction, the research highlights a novel aspect of viral manipulation of host cellular mechanisms. These findings provide a deeper understanding of the PEDV pathogenesis and identify potential targets for therapeutic intervention, advancing efforts to mitigate the economic impact of PEDV on the swine industry.

Mastitis caused by Staphylococcus aureus (S. aureus) is one of the most intractable problems for the dairy industry, causing significantly reduced milk yields and early slaughter of cows worldwide. MicroRNAs (miRNAs) can post-transcriptionally regulate gene expression and studies in recent years have shown the importance of miRNA-associated gene regulation in S. aureus-induced mastitis.

In this study, to investigate the role of miR-223 in mastitis, we performed experiments to overexpress and suppress miR-223 in an immortalized bovine mammary epithelial cell line (MAC-T) infected with S. aureus. Overexpression of miR-223 in MAC-T cells repressed cell apoptosis and necrosis induced by S. aureus infection, whereas suppression of miR-223 had the opposite effect. Transcriptome expression profiling with weighted gene co-expression network analysis (WGCNA) and gene set variation analysis (GSVA) showed that miR-223 affects apoptosis and inflammation-related pathways. Furthermore, differentially expressed (DE) genes were evaluated, and genes exhibiting contrasting expression trends in the miR-223 overexpressed and suppressed groups were assessed as potential target genes of miR-223. Potential target genes, including CDC25B, PTPRF, DCTN1, and DPP9, were observed to be associated with apoptosis and necroptosis. Finally, through integrative analysis of genome-wide association study (GWAS) data and the animal quantitative trait loci (QTL) database, we determined that target genes of miR-223 were significantly enriched in single-nucleotide polymorphisms (SNP) and QTLs related to somatic cell count (SCC) and mastitis.

In summary, miR-223 has an inhibitory effect on S. aureus-induced cell apoptosis and necrosis by regulating PTPRF, DCTN1, and DPP9. These genes were significantly enriched in QTL regions associated with bovine mastitis resistance, underscoring their relevance in genetic regulation of disease resilience. Our findings provide critical genetic markers for enhancing mastitis resistance, particularly S. aureus-induced mastitis, through selective breeding. This work offers valuable insights for developing cattle with improved resistance to mastitis via targeted genetic selection.

Cancer stem cells (CSCs) represent a small, yet pivotal subpopulation of tumor cells that play significant roles in tumor initiation, progression and therapeutic resistance. Circular RNAs (circRNAs) are a distinct class of RNAs characterized by their closed‑loop structures, lacking 5' to 3'ends. There is growing evidence that circRNAs are integral to the development and regulation of CSCs. Aberrant expression of circRNAs in CSCs can contribute to oncogenic properties and drug resistance. Specifically, oncogenic circRNAs modulate CSC behavior via key signaling pathways, thereby promoting CSC self‑renewal and maintenance, as well as tumor progression. This review summarizes the latest research on the functional roles and regulatory mechanisms of circRNAs in CSC behavior and discusses potential applications and challenges of targeting circRNAs in CSCs. Understanding the intricate interactions between circRNAs and CSCs may lead to novel therapeutic strategies that effectively combat treatment resistance and improve patient outcomes.

Effective osteoarthritis treatments that enhance the anabolic/regenerative capacity of chondrocytes are needed. Studying cartilage development processes may inform us of approaches to control chondrocyte differentiation and anabolism and, ultimately, how to effectively treat OA. MicroRNAs are broad-acting epigenetic regulators known to affect many skeletal processes. Previous reports from our group indicated that miR-181a-1 is upregulated during chondrocyte differentiation. The goal of this study was to determine how the entire miR-181a/b-1 cluster regulates in vitro chondrogenesis.

Precursor miR-181a/b-1 was over-expressed in cartilage progenitor cells using lentiviral technology Transduced cartilage progenitor cells were cultured as micromass pellets in hypoxic conditions and stimulated to undergo chondrogenic differentiation for five weeks. Bulk RNA-sequencing and immunostaining was applied to evaluate chondrogenic differentiation and matrix production.

Immunostaining of cartilage pellet sections showed that miR-181a/b-1 increased mature type II collagen and decreased expression of the chondroprogenitor type IIA collagen isoform. Bulk RNA-Seq at day 7 of chondrogenesis revealed upregulation of pro-anabolic genes such as COL2A1, COL9A2/3, COL11A2 and SNORC. Of the genes significantly downregulated by miR-181a/b-1, aquaporin 9 (AQP9) was the top hit which decreased in expression by over 14-fold. While a predicted target of miR-181a/b, our data showed that this miRNA cluster likely suppresses AQP9 via an indirect targeting mechanism.

Our findings demonstrate a pro-differentiation/anabolic function for miR-181a/b-1 during in vitro chondrogenesis that may be due, in part, to suppression of AQP9. Future studies are needed to elucidate the role of this membrane channel protein in regulating chondrocyte differentiation and homeostasis.

Background/Objectives: microRNAs are small non-coding RNAs that regulate gene expression by inducing mRNA degradation or inhibiting translation. A growing body of evidence suggests that miRNAs may be utilized as anti-cancer therapeutics by targeting expression of key genes involved in cancerous transformation and progression. Renal cell cancer (RCC) is the most common kidney malignancy. The most efficient RCC treatments involve blockers of immune checkpoints, including antibodies targeting PD-L1 (Programmed Death Ligand 1). Interestingly, recent studies revealed the cross-kingdom horizontal transfer of plant miRNAs into mammalian cells, contributing to the modulation of gene expression by food ingestion. Here, we hypothesized that PD-L1 expression may be modulated by miRNAs originating from edible plants. Methods: To verify this hypothesis, we performed bioinformatic analysis to identify mes-miR395e from Manihot esculenta (cassava) as a promising candidate miRNA that could target PD-L1. To verify PD-L1 regulation mediated by the predicted plant miRNA, synthetic mes-miR395 mimics were transfected into cell lines derived from RCC tumors, followed by evaluation of PD-L1 expression using qPCR and Western blot. Results: Transfection of mes-miR395e mimics into RCC-derived cell lines confirmed that this miRNA decreases expression of PD-L1 in RCC cells at both mRNA and protein levels. Conclusions: This preliminary study shows the promise of plant miRNA as potential adjuvants supporting RCC treatment.

Fine particulate matter (PM2.5) is often linked to a range of respiratory diseases and cellular damage. Although studies have shown that the expression profiles of microRNAs (miRNAs) are altered during lung damage brought on by PM2.5, the underlying functions of miRNAs remain poorly understood. In this research, we explored the role of PM2.5-induced apoptosis in detail and focused on the miRNA (miR-212-5p) that regulates apoptosis. Through a dual-luciferase assay, a direct targeting connection between laminin subunits γ2 (LAMC2) and α3 (LAMA3) and miR-212-5p was successfully demonstrated. This study focused on revealing the negative regulatory relationship between miR-212-5p and LAMC2 and LAMA3, providing important clues for a deeper understanding of the relevant physiological and pathological mechanisms. The present study showed that LAMC2 and LAMA3 positively regulate the PI3K-AKT pathway and negatively regulate the NF-κB pathway, which directly leads to significant changes in apoptosis rates. This study reveals a previously unrecognized molecular mechanism by showing that miR-212-5p directly targets LAMC2 and LAMA3 and thus associates with PM2.5-induced apoptosis via the PI3K/AKT/NF-κB pathway. These findings not only redefine the role of miR-212-5p in apoptosis but also open up new avenues for future research.

Small RNAs play critical roles in gene regulation in diverse processes across organisms. Crosslinking, ligation, and analyses of sequence hybrid (CLASH) experiments have shown PIWI and Argonaute proteins bind to diverse mRNA targets, raising questions about their functional relevance and the degree of flexibility in target recognition. As crosslinking-induced mutations (CIMs) provides nucleotide-resolution of RNA binding sites, we developed MUTACLASH to systematically analyze CIMs in piRNA and miRNA CLASH data in C. elegans . We found CIMs are enriched at the nucleotide positions of mRNA corresponding to the center of targeting piRNAs and miRNAs. Notably, CIMs are also enriched at nucleotides with local pairing mismatches to piRNA. In addition, distinct patterns of CIMs are observed between canonical and non-canonical base pairing interactions, suggesting that the worm PIWI Argonaute PRG-1 adopts distinct conformations for canonical vs. non-canonical interactions. Critically, non-canonical miRNA or piRNA binding sites with CIMs exhibit more regulatory effects than those without CIMs, demonstrating CIM analysis as a valuable approach in assessing functional significance of small RNA targeting sites in CLASH data. Together, our analyses reveal the landscapes of Argonaute crosslinking sites on mRNAs and highlight MUTACLASH as an advanced tool in analyzing CLASH data.

Understanding the essential role of miRNA in regulating cell invasion in glioblastoma opens up new avenues for targeted therapeutic interventions in the future. By screening out eligible miRNA expression data sets from the GEO database, the WGCNA package based on the R language is further used to construct a co-expression network model of the chip data set, to identify modules related to disease states and perform pivotal miRNA screening on the related modules. The target relationship between miRNA and TRPM8 was verified by bioinformatics and luciferase gene report, and the effect of miRNA overexpression on TRPM8 protein level was analyzed by Western blot. The result of miR-433 overexpression on the invasion ability of glioblastoma cells in vitro was examined by scratch test and Transwell invasion test. The results of this study indicate that the selected target miR-433 has a strong binding relationship with TRPM8 and can effectively regulate its expression. Furthermore, overexpression of miR-433 was found to inhibit the invasion ability of glioblastoma cells by targeting TRPM8. These data demonstrate that miR-433 can target TRPM8 to inhibit glioblastoma cell invasion.

Acute myocardial infarction (AMI) is one of the critical health conditions often caused by the rupture of unstable coronary artery plaque, triggering a series of events, such as platelet activation, thrombus formation, coronary artery blockage, lasted severe ischemia, and hypoxia in cardiomyocytes, and culminating in cell death. Platelet-derived microvesicles (PMVs) act as intermediates for cellular communication. Nevertheless, the role of PMVs in myocardial infarction remains unclear. Initially, AMI-related messenger ribose nucleic acid (mRNA) and micro RNA (miRNA) datasets from the Gene Expression Omnibus (GEO) database were analyzed, specifically focusing on the expressed genes associated with Ferroptosis. Further, a miRNA-mRNA regulatory network specific to AMI was constructed. Then, the effect of PMVs on cardiomyocyte survival was further confirmed through in vitro experiments. High ACSL1 expression was observed in the platelets of AMI patients. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that ACSL1, located in the mitochondria, played a key role in the PPAR signaling pathway. The elevated ACSL1 expression in a co-culture model of PMVs and AC16 cardiomyocytes significantly increased the AC16 cell Ferroptosis. Further, we validated that the platelet ACSL1 expression could be regulated by hsa-miR-449a. Together, these findings suggested that platelet ACSL1 could trigger myocardial cell death via PMV transport. In addition, this research provided a theoretical framework for attenuating myocardial cell Ferroptosis in patients with acute myocardial infarction.

The miRNA binds to AGO's seed region, prompting the exploration of small molecules that can offset miRNA repression of target mRNA. This miRNA-181c-5p was found to be upregulated in the chronic traumatic encephalopathy, a prevalent neurodegenerative disease in contact sports and military personals. The research aimed to identify compounds that disrupt the AGO-assisted loop formation between miRNA-181c-5p and ATM, consequently repressing the translation of ATM. Target genes from commonly three databases (DIANA-microT-CDS, miRDB, RNA22 and TargetScan) were subjected to functional annotation and clustering analysis using DAVID bioinformatics tool. Haddock server were employed to make miRNA-181c-5p:ATM-AGO complex. A total of 2594 small molecules were screened using Glide XP based on their highest binding affinity towards the complex, through a three-phase docking approach. The top 5 compounds (DB00674-Galantamine, DB00371-Meprobamate, DB00694-Daunorubicin, DB00837-Progabide, and DB00851-Dacarbazine) were further analyzed for stability in the miRNA-181c-5p:ATM-AGO-ligand complex interaction using GROMACS (version 2023.2). Hence, these findings suggest that these molecules hold potential for facilitating AGO-assisted repression of ATM gene translation.

The important role of PIWI-interacting RNAs (piRNAs) in tumors has garnered increasing attention. However, research on their role in lung adenocarcinoma (LUAD) remains limited. Elevated levels of piRNA-137463 have been linked to poor prognosis in LUAD patients. Inhibition of piRNA-137463 curbed the proliferation, migration, and invasion of LUAD cells, enhanced T cell cytotoxicity through increased IFN-γ secretion, disrupted cholesterol metabolism, and reduced intracellular cholesterol, lipid raft content, and PD-L1 expression in LUAD cells. Bioinformatic prediction identified a potential interaction between piRNA-137463 and lncRNA LOC100128494. Inhibiting piRNA-137463 increased the stability and expression of LOC100128494, which further modulated insulin-induced gene 1 protein (INSIG1) levels via a competitive endogenous RNA network involving LOC100128494 and miR-24-3p. Notably, the effect of piRNA-137463 in LUAD cells is dependent on the expression of LOC100128494 and INSIG1. Inhibiting the expression of piRNA-137463 with AntagopiRNA-137463 suppressed tumor growth and metastasis via LOC100128494 in nude mice and enhanced the response of LUAD to anti-PD-1 therapy in immune-competent mice. In summary, this study elucidates the role of piRNA-137463 in the reprogramming of cholesterol metabolism, which drives the progression of LUAD, thereby identifying a new target for the comprehensive clinical management of LUAD.

Wing dimorphism is regarded as an important phenotypic plasticity involved in the migration and reproduction of aphids. However, the signal transduction and regulatory mechanism of wing dimorphism in aphids are still unclear. Herein, the optimal environmental conditions were first explored for inducing winged offspring of green peach aphid, and the short photoperiod was the most important environmental cue to regulate wing dimorphism. Compared to 16 L:8 D photoperiod, the proportion of winged offspring increased to 90% under 8 L:16 D photoperiod. Subsequently, 5 differentially expressed microRNAs (miRNAs) in aphids treated with long and short photoperiods were identified using small RNA sequencing, and a novel miR-3040 was identified as a vital miRNA involved in photoperiod-mediated wing dimorphism. More specifically, the inhibition of miR-3040 expression could reduce the proportion of winged offspring induced by short photoperiod, whereas its activation increased the proportion of winged offspring under long photoperiod. Meanwhile, the expression level of miR-3040 in winged aphids was about 2.5 times that of wingless aphids, and the activation or inhibition of miR-3040 expression could cause wing deformity, revealing the dual-role regulator of miR-3040 in wing dimorphism and wing development. In summary, the current study identified the key environmental cue for wing dimorphism in green peach aphid, and the first to demonstrate the dual-role regulator of miR-3040 in photoperiod-mediated wing dimorphism and wing development.

Piwi-interacting RNAs (piRNA)s are non-coding small RNAs that post-transcriptionally affect gene expression and regulation. Through complementary seed region binding with transposable elements (TEs), piRNAs protect the genome from transposition. A tool to link piRNAs with complementary TE targets will improve our understanding of the role of piRNAs in genome maintenance and gene regulation. Existing tools such as TEsmall can process sRNA-seq datasets to produce differentially expressed piRNAs, and piRScan developed for nematodes can link piRNAs and TEs but it requires knowledge about the target region of interest and works backwards.

We developed FishPi to predict the pairings between piRNA and TEs for available genomes from zebrafish, medaka and tilapia, with full user customisation of parameters including orientation of piRNA, mismatches in the piRNA seed binding to TE and scored output lists of piRNA-TE matches. FishPi works with individual piRNAs or a list of piRNA sequences in fasta format. The software focuses on the piRNA-TE seed region and analyses reference TEs for piRNA complementarity. TE type is examined, counted and stored to a dictionary, with genomic loci recorded. Any updates to piRNA-TE binding rules can easily be incorporated by changing the seed-region options in the graphic user-interface. FishPi provides a graphic interface using tkinter for the user to input piRNA sequences to generate comprehensive reports on piRNA-TE interactions. FishPi can easily be adapted to genomes from other species and taxa opening the interpretation of piRNA functionality to a wide community.

Users will gain insight into genome mobility and FishPi will help further our understanding of the biological role of piRNAs and their interaction with TEs in a similar way that public databases have improved the access to and the understanding of the role of small RNAs.

The interaction relationship between miRNAs and genes is important as miRNAs play a crucial role in regulating gene expression. In the literature, several databases have been constructed to curate known miRNA target genes, which are valuable resources but likely only represent a small fraction of all miRNA-gene interactions. In this study, we constructed machine learning models to predict miRNA target genes that have not been previously reported. Using the miRNA and gene expression data from TCGA, we performed a correlation analysis between all miRNAs and all genes across multiple cancer types. The correlations served as features to describe each miRNA-gene pair. Using the existing databases of curated miRNA targets, we labeled the miRNA-gene pairs, and trained machine learning models to predict novel miRNA-gene interactions. For the miRNA-gene pairs that were consistently predicted across the models, we called them significant miRNA-gene pairs. Using held-out miRNA target databases and a literature survey, we validated 5.5% of the predicted significant miRNA-gene pairs. The remaining predicted miRNA-gene pairs could serve as hypotheses for experimental validation. Additionally, we explored several additional datasets that provided gene expression data before and after a specific miRNA perturbation and observed consistency between the correlation direction of predicted miRNA-gene pairs and their regulatory patterns. Together, this analysis revealed a novel framework for uncovering previously unidentified miRNA-gene relationships, enhancing the collective comprehension of miRNA-mediated gene regulation.

MiR-30c and fatty acid synthase (fas) both play important roles in physiological processes such as lipid synthesis and fat metabolism. Predictive analysis revealed that fas is a target gene of miR-30c with multiple seed sites. Seed sites are useful to predict miRNA targeting relationships; however, detailed analyses of seed sites in fish genomes remain poorly studied. In this study, the regulatory relationship between miR-30c and fas, number and effect of seed regions, and mechanism by which miR-30c regulates lipid metabolism were evaluated in blunt snout bream (Megalobrama amblycephala). Four miR-30c target sites for fas were identified using various prediction tools. miR-30c mimics were transfected into 293 T cells, and dual-luciferase reporter assays were used to evaluate the roles of different fas target sites. When a single target site was mutated, relative luciferase activity was higher than that in the control group, with different activity levels depending on the mutation site. When multiple target sites were mutated, relative luciferase activity increased significantly as the number of mutation sites increased and was the highest when the four sites were mutated simultaneously. The miR-30c agomir was injected into the abdominal cavity of M. amblycephala at various concentrations for analyses of physiological and biochemical parameters in the liver and blood and the expression of genes related to lipid metabolism in the liver. Total cholesterol, free fatty acid, triglyceride, and low density lipoprotein levels were significantly lower after miR-30c agomir injection comparing to the control (P < 0.05). Additionally, the expression levels of genes related to lipid metabolism were significantly lower after miR-30c agomir injection than in the control (P < 0.05). In summary, this study identified four specific miR-30c target sites in the 3' UTR of fas mRNA; the effects of these sites are cumulative, and the redundancy ensures the accurate regulation of fas during evolution. In addition, miR-30c has a negative regulatory effect on fas and regulates lipid metabolism via various genes related to this process. Therefore, the regulation of miR-30c can effectively ameliorate the side effects of a high-fat diet on liver function in M. amblycephala.

With the rapid increase in the number of implant operations, the incidence of bone infections has increased. Methicillin-resistant Staphylococcus aureus (S. aureus) and other emerging fully drug-resistant strains make the management of bone infections even more challenging. Bone infections are mainly caused by S. aureus and require extensive surgical intervention and long-term antibiotic therapy. The host autophagy response is critical to the elimination of S. aureus infections. In this study, we demonstrate that a circular RNA (circRNA), circSyk, is a potential biological target for the treatment of S. aureus-induced bone infection. Most importantly, S. aureus regulates circSyk to block autophagy and promote bone destruction via the circSyk/miR-5106/Sik3 axis in a nonclassical pathway, which is involved in the S. aureus infection process through a competitive endogenous RNA network. In summary, this study proposes a novel perspective on the immune escape of S. aureus in bone infections, based on circRNA.

Although the therapeutic potential of microRNA-mediated gene regulation has been investigated, its precise functional regulatory mechanism in neuropathic pain remains incompletely understood. In this study, we elucidate that miR-216a-3p serves as a critical noncoding RNA involved in the modulation of trigeminal-mediated neuropathic pain. By conducting RNA-seq and qPCR analysis, we observed a notable decrease of miR-216a-3p in the injured trigeminal ganglia (TG) of male rats. Intra-TG administration of miR-216a-3p agomir or lentiviral-mediated overexpression of miR-216a-3p specifically in sensory neurons of injured TGs alleviated established neuropathic pain behaviors, while downregulation of miR-216a-3p (pharmacologically or genetically) in naive rats induced pain behaviors. Moreover, nerve injury significantly elevated the histone H3 lysine-27 (H3K27) trimethylation (H3K27me3) levels in the ipsilateral TG, thereby suppressing the SRY-box TF 10 (SOX10) binding to the miR-216a-3p promoter and resulting in the reduction of miR-216a-3p. Inhibiting the enzymes responsible for catalyzing H3K27me3 restored the nerve injury-induced reduction in miR-216a-3p expression and markedly ameliorated neuropathic pain behaviors. Furthermore, miR-216a-3p targeted stromal interaction molecule 1 (STIM1), and the decreased miR-216a-3p associated with neuropathic pain caused a significant upregulation in the protein abundance of STIM1. Conversely, overexpression of miR-216a-3p in the injured TG suppressed the upregulation of STIM1 expression and reversed the mechanical allodynia. Together, the mechanistic understanding of H3K27me3-dependent SOX10/miR-216a-3p/STIM1 signaling axial in sensory neurons may facilitate the discovery of innovative therapeutic strategies for neuropathic pain management.

Plant-derived microRNAs (miRNAs) have attracted significant attention for their potential in cross-kingdom gene regulation, but the mechanisms of their entry, stability, and function in animal bodies need further investigation. We provided an in-depth analysis of tissue-specific miRNA expression in dairy cows, identifying 347 miRNAs, including 16 novel candidates, across 21 normal tissues. Our findings revealed that specific miRNAs, such as miR-192, miR-143, miR-148a, miR-486, and miR-21-5p, showed distinct tissue enrichment. In addition, a total of 167 maize-derived miRNAs were identified in dairy cow tissues, particularly in the rumen, mammary glands, serum, and exosomes. These exogenous miRNAs, which are abundant and conserved among plants, may be absorbed by the SLC46A2 transporter in the rumen epithelium during feeding and distributed to other tissues via exosomal encapsulation. The maize-derived miR166a-3p was highly abundant. Transfection experiments confirmed that miR166a-3p reduces the expression of proliferation markers (PCNA, Cyclin D, and Cyclin E) and the anti-apoptotic gene Bcl2, while upregulating the pro-apoptotic gene Bax. Moreover, exosomes derived from bovine serum were found to mediate these effects, as miR166a-3p-enriched exosomes inhibited cell proliferation and promoted apoptosis, further supporting the cross-kingdom role of plant-derived miRNAs in regulating biological processes. This study enhances the understanding of miRNA regulatory mechanisms, particularly the absorption and systemic transport of plant-derived miRNAs in dairy cows. The findings underscore the potential for using exogenous miRNAs, like miR166a-3p, in agricultural and medical contexts, warranting further investigation into their functions and cross-species interactions.

Background/Objectives: Chronic venous insufficiency (CVI), a chronic vascular dysfunction, is a common health problem that causes serious complications such as painful varicose veins and even skin ulcers. Identifying the underlying genetic and epigenetic factors is important for improving the quality of life of individuals with CVI. In the literature, many genes, variants, and miRNAs associated with CVI have been identified through genomic and transcriptomic studies. Despite molecular pathogenesis studies, how the genes associated with CVI are regulated by miRNAs and the effect of variants in binding regions on expression levels are still not fully understood. In this study, previously identified genes, variants, and miRNAs associated with CVI, common variants in the mRNA-miRNA binding regions, were investigated using in silico analyses. Methods: For this purpose, miRNA research tools, MBS (miRNA binding site) database, genome browsers, and the eQTL Calculator in the GTEx portal were used. Results: We identified SNVs associated with CVI that may play a direct role in the miRNA-mediated regulation of the ZNF664, COL1A2, HFE, MDN, MTHFR, SRPX, TDRD5, TSPYL4, VEGFA, and APOE genes. In addition, when the common SNVs in the mRNA binding region of 75 unique CVI related-miRNAs in five candidate genes associated with CVI were examined, seven miRNAs associated with the expression profiles of ABCA1, PIEZO1, and CASZ1 genes were identified. Conclusions: In conclusion, the relationship between genetic markers identified in the literature that play a role in the pathogenesis of the CVI and the expression profiles was evaluated for the first time in the mRNA-miRNA interaction axis.

The beet webworm, Loxostege sticticalis, is a typical migratory pest. Although miRNAs participate in many physiological functions, little is known about the functions of miRNAs in olfactory regulation. In this study, 1120 (869 known and 251 novel) miRNAs were identified in the antennae of L. sticticalis by using high-throughput sequencing technology. Among the known miRNAs, 189 from 49 families were insect-specific, indicating that these miRNAs might play unique roles in insects. Furthermore, based on the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses, we found that 3647 and 1393 miRNAs were associated with localization and the regulation of localization, respectively, and 80 miRNAs were enriched in the neuroactive ligand-receptor interaction pathway. These miRNAs might be involved in the olfactory system of L. sticticalis. Notably, qRT-PCR showed that most of the tested miRNAs presented similar expression patterns compared with the RNA-seq data and that miR-87-3, novel-miR-78, and novel-miR-142 were significantly differentially expressed in the antennae of males and females. In addition, 21 miRNAs were predicted to target 23 olfactory genes, including 10 odorant-binding proteins (OBPs), 3 chemosensory proteins (CSPs), 4 odorant receptors (ORs), 1 ionotropic receptor (IR), and 5 gustatory receptors (GRs). The olfactory-related miRNAs exhibited low-abundance transcripts, except undef-miR-55 and undef-miR-523, and gender-biased expression was not observed for olfactory-related miRNAs. Our findings provide an overview of the potential miRNAs involved in olfactory regulation, which may provide important information on the function of miRNAs in the insect olfactory system.

Different neuron types develop characteristic axonal and dendritic arborizations that determine their inputs, outputs, and functions. Expression of fate-determinant transcription factors is essential for specification of their distinct identities. However, the mechanisms downstream of fate-determinant factors coordinating different aspects of neuron identity are not understood. Specifically, how distinct projection neurons develop appropriate dendritic arbors that determine their inputs is unknown. Here, we investigate this question in corticospinal and callosal projection neurons. We identified a mechanism linking the corticospinal/corticofugal identity gene Fezf2 with the regulation of dendritic development. We show that miR-193b∼365 microRNA cluster is regulated by Fezf2 and enriched in corticospinal neurons. miR-193b∼365 represses mitogen-activated protein kinase 8 (MAPK8) to regulate corticospinal dendritic development. miR-193b∼365 overexpression in callosal neurons abnormally reduces MAPK8 signal and dendritic complexity. Our findings show that regulation of MAPK8 via miR-193b∼365 cluster regulates dendritic development, providing a mechanism that coordinates projection neuron identity, specified by Fezf2, and neuron-specific dendritic morphology.

The ovary is a central organ in the reproductive system that produces oocytes and synthesizes and secretes steroid hormones. Healthy development and regular cyclical change in the ovary is crucial for regulating reproductive processes. However, the key genes and metabolites that regulate ovarian development and pregnancy have not been fully elucidated. This study conducted high-throughput RNA sequencing and untargeted metabolite profiling of the ovarian tissues from Chenghua pigs at four stages, including postnatal day 3 (D3), puberty at the age of about 125 days (Pub), sexual maturity at the age of about 365 days (Y1), and 105 days after pregnancy at the age of about 360 days (Pre).

A total of 9,264 and 1,593 differentially expressed genes (DEGs) were identified during ovarian development and pregnancy. Several key genes involved in ovarian development, including SQLE, HMGCS1, MSMO1, SCARB1, CYP11A1, HSD3B1, HSD17B1, and SERPINE1 were identified. Similarly, LUM, FN1, PLAUR, SELP, SDC1, and VCAN were considered to be associated with pregnancy maintenance. Overexpression of HSD17B1 in granulosa cells significantly upregulated estrogen synthesis-related genes (HSD3B1, CYP11A1, and STAR); meanwhile, overexpression of PLAUR promotes granulosa cell proliferation. Furthermore, 66, 24, 77, and 7 differentially expressed miRNAs (DEMis) were found, leading to the selection of key miRNAs such as ssc-miR-206, ssc-miR-107, ssc-miR-429, ssc-miR-210, and ssc-miR-133a-3p by differential miRNA-targeted mRNA interaction network; meanwhile, ssc-miR-133a-3p was validated to have a targeting relationship with KCNA1 by dual-luciferase reporter systems assay. At the metabolic levels, androstenedione, 17a-hydroxyprogesterone, dehydroepiandrosterone, and progesterone were identified, with their synthesis regulated by these DEGs in the ovarian steroidogenesis pathway. Furthermore, treatment of cells with androstenedione upregulated the expression of HSD3B1, CYP11A1, and STAR.

This study revealed the dynamic changes in the transcriptome and metabolome of pig ovaries across developmental stages and gestation, indicating that it may provide new theoretical insights for improving sow fertility.

Studies have correlated living close to major roads with Alzheimer's disease (AD) risk. However, the mechanisms responsible for this link remain unclear.

We exposed olfactory mucosa (OM) cells of healthy individuals and AD patients to diesel emissions (DE). Cytotoxicity of exposure was assessed, mRNA, miRNA expression, and DNA methylation analyses were performed. The discovered altered pathways were validated using data from the human population-based Rotterdam Study.

DE exposure resulted in an almost four-fold higher response in AD OM cells, indicating increased susceptibility to DE effects. Methylation analysis detected different DNA methylation patterns, revealing new exposure targets. Findings were validated by analyzing data from the Rotterdam Study cohort and demonstrated a key role of nuclear factor erythroid 2-related factor 2 signaling in responses to air pollutants.

This study identifies air pollution exposure biomarkers and pinpoints key pathways activated by exposure. The data suggest that AD individuals may face heightened risks due to impaired cellular defenses.

Healthy and AD olfactory cells respond differently to DE exposure. AD cells are highly susceptible to DE exposure. The NRF2 oxidative stress response is highly activated upon air pollution exposure. DE-exposed AD cells activate the unfolded protein response pathway. Key findings are also confirmed in a population-based study.

Epstein-Barr virus (EBV) infection in humans is ubiquitous and associated with various diseases. Remodeling of the immune microenvironment is the primary cause of EBV infection and pathogenesis; however, the underlying mechanism has not been fully elucidated. In this study, we used whole-transcriptome RNA-Seq to detect mRNAs, long non-coding RNAs (lncRNA), and microRNA (miRNA) profiles in the control group, 3 days, and 28 days after EBV infection, based on the tree shrew model that we reported previously. First, we estimated the proportion of 22 cell types in each sample using CIBERSORT software and identified 18 high-confidence DElncRNAs related to immune microenvironment regulation after EBV infection. Functional enrichment analysis of these differentially expressed lncRNAs primarily focused on the autophagy, endocytosis, and ferroptosis signalling pathways. Moreover, EBV infection affects miRNA expression patterns, and many miRNAs are silenced. Finally, three competing endogenous RNA regulatory networks were built using lncRNAs that significantly correlated with immune cell types, miRNAs that responded to EBV infection, and potentially targeted the mRNA of the miRNAs. Among them, MRPL42-AS-5 might act as an hsa-miR-296-5p "sponge" and compete with target mRNAs, thus increasing mRNA expression level, which could induce immune cell infiltration through the cellular senescence signalling pathway against EBV infection. Overall, we conducted a complete transcriptomic analysis of EBV infection in vivo for the first time and provided a novel perspective for further investigation of EBV-host interactions.

Current medical research has been demonstrating the roles of miRNAs in a variety of cellular mechanisms, lending credence to the association between miRNA dysregulation and multiple diseases. Understanding the mechanisms of miRNA is critical for developing effective diagnostic and therapeutic strategies. miRNA-mRNA interactions emerge as the most important mechanism to be understood despite their experimental validation constraints. Accordingly, several computational models have been developed to predict miRNA-mRNA interactions, albeit presenting limited predictive capabilities, poor characterisation of miRNA-mRNA interactions, and low usability. To address these drawbacks, we developed PRIMITI, a PRedictive model for the Identification of novel miRNA-Target mRNA Interactions. PRIMITI is a novel machine learning model that utilises CLIP-seq and expression data to characterise functional target sites in 3'-untranslated regions (3'-UTRs) and predict miRNA-target mRNA repression activity. The model was trained using a reliable negative sample selection approach and the robust extreme gradient boosting (XGBoost) model, which was coupled with newly introduced features, including sequence and genetic variation information. PRIMITI achieved an area under the receiver operating characteristic (ROC) curve (AUC) up to 0.96 for a prediction of functional miRNA-target site binding and 0.96 for a prediction of miRNA-target mRNA repression activity on cross-validation and an independent blind test. Additionally, the model outperformed state-of-the-art methods in recovering miRNA-target repressions in an unseen microarray dataset and in a collection of validated miRNA-mRNA interactions, highlighting its utility for preliminary screening. PRIMITI is available on a reliable, scalable, and user-friendly web server at https://biosig.lab.uq.edu.au/primiti.

Piwi-interacting RNAs (piRNAs) are a class of small non-coding RNAs (ncRNAs) that plays important roles in many biological processes and major cancer diagnosis and treatment, thus becoming a hot research topic. This study aims to provide an in-depth review of computational piRNA-related research, including databases and computational models. Herein, we perform literature analysis and use comparative evaluation methods to summarize and analyze three aspects of computational piRNA-related research: (i) computational models for piRNA-related molecular identification tasks, (ii) computational models for piRNA-disease association prediction tasks, and (iii) computational resources and evaluation metrics for these tasks. This study shows that computational piRNA-related research has significantly progressed, exhibiting promising performance in recent years, whereas they also suffer from the emerging challenges of inconsistent naming systems and the lack of data. Different from other reviews on piRNA-related identification tasks that focus on the organization of datasets and computational methods, we pay more attention to the analysis of computational models, algorithms, and performances that aim to provide valuable references for computational piRNA-related identification tasks. This study will benefit the theoretical development and practical application of piRNAs by better understanding computational models and resources to investigate the biological functions and clinical implications of piRNA.

The global rise of oropharyngeal cancers (OPC) associated with the human papillomavirus (HPV) type 16 necessitates a deeper understanding of their underlying molecular mechanisms. Our study utilised RNA-sequencing data from The Cancer Genome Atlas (TCGA) to identify and analyse differentially expressed (DE) long non-coding RNAs (lncRNAs), microRNAs (miRNAs), and messenger RNAs (mRNAs) in HPV16-positive OPC, and to elucidate the interplay within the lncRNA/miRNA/mRNA regulatory network. We revealed 1929 DE lncRNAs and identified a significant expression shift in 37 of these, suggesting a regulatory 'sponge' function for miRNAs that modulate cellular processes. Notably, the lncRNA Linc00911 exhibited decreased expression in HPV16-positive OPC, a change directly attributable to HPV oncogenes E6 and E7 as confirmed by RT-qPCR in cell lines and patient samples. Our comprehensive analysis presents an expansive landscape of ncRNA-mRNA interactions, offering a resource for the ongoing pursuit of elucidating the molecular underpinnings of HPV-driven OPC.

Despite the recent therapeutic advances in neurological disorders, curative therapy remains a serious challenge in many cases. Even though recent years have witnessed the development of gene therapy from among the different therapeutic approaches affecting pathophysiological mechanisms, intriguing aspects exist regarding the effectiveness, safety, and mechanism of action of gene therapies. Micro ribonucleic acid (microRNA-miRNA), as a fundamental gene regulator, regulates messenger ribonucleic acid (mRNA) by directly binding through the 3'-untranslated region (3'-UTR). MicroRNA-219 is a specific brain-enriched miRNA associated with neurodevelopmental disorders that play crucial roles in the differentiation of oligodendrocyte progenitorcells, promotion of oligodendrocyte maturation, remyelination, and cognitive functions to the extent that it can be considered a potential therapeutic option for demyelination in multiple sclerosis and spinal cord injury and reverse chronic inflammation pains. Additionally, miR-219 regulates the circadian clock, influencing the duration of the circadian clock period. This regulation can impact mood stability and is associated with phase fluctuations in bipolar patients. Furthermore, miR-219 also plays a role in modulating tau toxicity, which is relevant to the pathophysiology of Alzheimer's disease and schizophrenia. Finally, it reportedly has protective effects against seizures and Parkinson's disease, as well as neoplasms, by inhibiting proliferation, suppressing invasion, and inducing cell death in tumor cells. Exploring the miR-219 molecular pathways and their therapeutic effects on central nervous system disorders and the mechanisms involved, the present review study aims to illustrate how this information may change the future of gene therapy.

Despite all the progress in treating SARS-CoV-2, escape mutants to current therapies remain a constant concern. Promising alternative treatments for current and future coronaviruses are those that limit escape mutants by inhibiting multiple pathogenic targets, analogous to the current strategies for treating HCV and HIV. With increasing popularity and ease of manufacturing of RNA technologies for vaccines and drugs, therapeutic microRNAs represent a promising option. In the present work, miR-24-3p was identified to inhibit SARS-CoV-2 entry, replication, and production; furthermore, this inhibition was retained against common mutations improving SARS-CoV-2 fitness. To determine the mechanism of action, bioinformatic tools were employed, identifying numerous potential effectors promoting infection targeted by miR-24-3p. Of these targets, several key host proteins for priming and facilitating SARS-CoV-2 entry were identified: furin, NRP1, NRP2, and SREBP2. With further experimental analysis, we show that miR-24-3p directly downregulates these viral entry factors to impede infection when producing virions and when infecting the target cell. Furthermore, we compare the findings with coronavirus, HCoV-229E, which relies on different factors strengthening the miR-24-3p mechanism. Taken together, the following work suggests that miR-24-3p could be an avenue to treat current coronaviruses and those likely to emerge.

The excision of specific tRNA-derived small RNAs (tsRNAs), yRNA-derived small RNAs (ysRNAs) and ribosomal RNA-derived small RNAs (rsRNAs) is now well established. Several reports have suggested many of these fragments function much like traditional microRNAs (miRNAs). That said, whereas the expressions of the majority of appreciably expressed miRNAs in HCT116 colon cancer cells are significantly decreased in individual knockouts (KOs) of DROSHA, DGCR8, XPO5, and DICER, on average, only 3.5% of tsRNA, ysRNA, and rsRNA expressions are impaired. Conversely, tsRNA, ysRNA, and rsRNA expressions are significantly increased in each of these KOs as compared to WT. As such, although DICER has been suggested to be involved with the expression of specific tsRNAs, ysRNAs, and rsRNAs, our study finds no evidence supporting the involvement of any of these canonical miRNA biogenesis enzymes in their expressions.

Metabolism-disorder-induced liver diseases have become increasingly prevalent worldwide and are clinically linked to obesity and type 2 diabetes. In addition, a large number of previous literature studies have indicated that plasma miR-130b is a promising biomarker for the early diagnosis and treatment of obesity. However, whether miRNA-130b that was positively correlated with obesity resulted in hepatic inflammation needs to be further studied. Therefore, the study aims to determine the effect of microvesicle-shuttled miRNA-130b (miR-130b-MV) on the hepatic inflammation and its potential mechanism in high-fat diet-induced obese mice. Three-week-old C57BL/6 mice were fed a high-fat diet for eight weeks. Then, the obese mice received tail vein injections of MV-packaged scrambled control microRNA (miR-SC-MV) or miR-130b-MV every other day for 10 days. Compared with the control group, the miR-130b-MV injection significantly reduced the body weight while increasing the ratio of liver wet weight to total body weight. In addition, the miR-130b-MV injection significantly activated the hepatic inflammation by increasing the expression of proinflammatory genes, although the plasma concentrations of IL-6 and TNF-α were only slightly increased. Furthermore, the miR-130b-MV injection significantly increased the hepatic miR-130b expression while significantly suppressing the protein expression and phosphorylation of GR, a potential target of miR-130b. Moreover, the miR-130b overexpression results in a decrease in the expression of endogenous GR protein and a decrease in the activity of the luciferase reporter of GR 3'-UTR. In addition, the miR-130b-MV injection significantly upregulated NF-kB (p50) in both the cytoplasm and nucleus, showing enhanced proinflammation response. The above results demonstrated that miR-130b-MV activated the hepatic inflammation by inhibiting GR-mediated immunosuppression in high-fat diet-induced obese mice, suggesting a novel mechanism underlying the obesity-induced hepatic inflammation, and the inhibition of miR-130b may serve as a new molecular therapeutic target for the prevention and treatment of hepatic inflammation.

Chickens play a crucial role as the primary global source of eggs and poultry, and the quality of rooster semen significantly impacts poultry reproductive efficiency. Therefore, it is imperative to comprehend the regulatory mechanisms underlying sperm development.

In this study, we established transcriptome profiles of lncRNAs, miRNAs, and mRNAs in 3 testis tissues and 3 epididymis tissues from "Jing Hong No.1" roosters at 24, 35, and 64 weeks of age. Using the data, we conducted whole transcriptome analysis and constructed a ceRNA network. We detected 10 differentially expressed mRNAs (DEmRNAs), 33 differentially expressed lncRNAs (DElncRNAs), and 10 differentially expressed miRNAs (DEmiRNAs) in the testis, as well as 149 DEmRNAs, 12 DElncRNAs, and 10 DEmiRNAs in the epididymis. These genes were found to be involved in cell differentiation and development, as well as various signaling pathways such as GnRH, MAPK, TGF-β, mTOR, VEGF, and calcium ion pathways. Subsequently, we constructed two competing endogenous RNA (ceRNA) networks comprising DEmRNAs, DElncRNAs, and DEmiRNAs. Furthermore, we identified four crucial lncRNA-mRNA-miRNA interactions that govern specific biological processes in the chicken reproductive system: MSTRG.2423.1-gga-miR-1563-PPP3CA and MSTRG.10064.2-gga-miR-32-5p-GPR12 regulating sperm motility in the testis; MSTRG.152556.1-gga-miR-9-3p-GREM1/THYN1 governing immunomodulation in the epididymis; and MSTRG.124708.1-gga-miR-375-NDUFB9/YBX1 controlling epididymal sperm maturation and motility.

Whole transcriptome sequencing of chicken testis and epididymis screened several key genes and ceRNA regulatory networks, which may be involved in the regulation of epididymal immunity, spermatogenesis and sperm viability through the pathways of MAPK, TGF-β, mTOR, and calcium ion. These findings contribute to our comprehensive understanding of the intricate molecular processes underlying rooster spermatogenesis, maturation and motility.

In recent years, the red swamp crayfish (Procambarus clarkii, P. clarkii) farming industry has suffered huge economic losses due to the pathogenic bacterium Spiroplasma eriocheiris (S. eriocheiris). To elucidate the immune response mechanism and identify hub immune genes as well as their associated microRNAs that regulate the host response of P. clarkii against S. eriocheiris infection, we conducted a comprehensive analysis on P. clarkii hemocyte mRNA and microRNA (miRNA) transcriptomes at different infection stages using third- and second-generation sequencing technologies. In full-length transcriptome functional annotation, 8155 unigenes were annotated, and 1168 potential new transcripts were predicted. In the mRNA transcriptome, a total of 3168 differentially expressed genes were identified at different infection stages, including 1492 upregulated and 1676 downregulated genes (duplicate genes excluded). Transcriptome analysis revealed 880 differentially expressed genes involved in multiple pathways and processes such as endocytosis, autophagy, lysosome, mTOR signaling, phagosome, and the Fanconi anemia pathway. Mfuzz analysis was employed to integrate and cluster the differential expression trends of genes across the three infection stages. In the miRNA transcriptome, 234 miRNAs and 966 predicted target genes were identified, with 86 differentially expressed miRNAs identified across the three time periods. A significant difference (P < 0.05) was observed for miRNAs including pcl-miR-146-3p, pcl-miR-74-3p, pcl-miR-225-5p, and pcl-miR-68-5p. These miRNAs are involved in multiple immune and autophagy-related pathways and have regulatory effects on immune genes including Vps26, lqf, and ERK-A. Based on the differentially expressed immune-related genes, we constructed a protein-protein interaction (PPI) network, which revealed the interactions among hub genes including Rac1, Akt1, Rho1, and Egfr. We also constructed a miRNA-gene interaction network in immune and autophagy-related processes, highlighting the potential regulatory effects of miRNAs including pcl-miR-183-5p, pcl-miR-146-3p, pcl-miR-176-5p, and pcl-miR-225-5p on proteins including LST8, SNAP29, Rab-7A, and ERK-A. To conclude, this study has identified hub immune genes and corresponding regulatory miRNAs in P. clarkii hemocytes in response to S. eriocheiris infection and explored the roles of these genes in selected pathways and processes. These findings are expected to provide further insights into the molecular mechanisms that confer resistance to S. eriocheiris infection in P. clarkii.

Centrally mediated abdominal pain syndrome (CAPS) has generated a heavy disease burden worldwide. This study aimed to explore the serum exosomal microRNAs (miRNAs) as potential diagnostic biomarkers for CAPS. From September 2022 to October 2023, 97 patients with CAPS and 96 healthy subjects were enrolled. Differentially expressed serum exosomal miRNAs between patients with CAPS and healthy controls were identified by high-throughput sequencing and quantitative real-time polymerase chain reaction. The receiver operating characteristic curves and multivariate logistic regression analysis were used to evaluate the diagnostic value of the serum exosomal miRNAs. MiR-6850-5p, miR-194-5p, miR-199a-3p, and miR-4525 were significantly downregulated in serum exomes of CAPS patients compared with healthy controls, which yielded the area under curve (AUC) values of .914 (95% confidence interval (CI), .873-.954), .767 (95% CI, .695-.839), .617 (95% CI, .527-.708), and .561 (95% CI, .465-.656), respectively, to distinguish CAPS patients from healthy subjects. And AUC of the integration of the above 4 miRNAs was .931 (95% CI, .896-.966). Multivariate logistic regression indicated that hsa-miR-6850-5p (odds ratio (OR) = .046, P < .001), anxiety (OR = 7.670, P = .025), and depression (OR = 22.967, P = .008) were the independent predictors of CAPS. Serum exosomal miR-6850-5p is a promising diagnostic biomarker for CAPS. PERSPECTIVE: This study may be the first to explore serum exosomal miRNAs as new diagnostic biomarkers for CAPS, and the findings may help clinicians to access comprehensive understanding and accurate diagnosis of CAPS.