Changes In miRNA Expression Research Articles

Age‑related brainstem degeneration through microRNA modulation in mice.

Histopathological changes occur in the brainstem during the early stages of Alzheimer's disease (AD), with the pathological changes of the brain lesions ascending progressively in accordance with the Braak staging system. The senescence‑accelerated mouse prone 8 (SAMP8) mouse model has been previously used as a model of age‑dependent neurodegenerative diseases, including AD. In the present study, microRNAs (miRNAs) that were upregulated or downregulated in SAMP8 brainstems were identified using miRNA profiling of samples obtained from miRNA arrays. The preliminary stage of cognitive dysfunction was examined using male 5‑month‑old SAMP8 mice, with age‑matched senescence‑accelerated mouse resistant 1 mice as controls. A Y‑maze alternation test was performed to assess short‑term working memory and miRNA profiling was performed in each region of the dissected brain (brainstem, hippocampus and cerebral cortex). SAMP8 mice tended to be hyperactive, but short‑term working memory was preserved. Two miRNAs were upregulated (miR‑491‑5p and miR‑764‑5p) and two were downregulated (miR‑30e‑3p and miR‑323‑3p) in SAMP8 brainstems. In SAMP8 mice, the expression level of upregulated miRNAs were the highest in the brainstem, wherein age‑related brain degeneration occurs early. It was demonstrated that the order of specific miRNA expression levels corresponded to the progression order of age‑related brain degeneration. Differentially expressed miRNAs regulate multiple processes, including neuronal cell death and neuron formation. Changes in miRNA expression may result in the induction of target proteins during the early stages of neurodegeneration in the brainstem. These findings suggest that studying altered miRNA expression may provide molecular evidence for early age‑related neuropathological changes.

Extension of mRNA poly(A) tails and 3'UTRs during neuronal differentiation exhibits variable association with post-transcriptional dynamics.

Differentiation of neural progenitor cells into mature neuronal phenotypes relies on extensive temporospatial coordination of mRNA expression to support the development of functional brain circuitry. Cleavage and polyadenylation of mRNA has tremendous regulatory capacity through the alteration of mRNA stability and modulation of microRNA (miRNA) function, however the extent of utilization in neuronal development is currently unclear. Here, we employed poly(A) tail sequencing, mRNA sequencing, ribosome profiling and small RNA sequencing to explore the functional relationship between mRNA abundance, translation, poly(A) tail length, alternative polyadenylation (APA) and miRNA expression in an in vitro model of neuronal differentiation. Differential analysis revealed a strong bias towards poly(A) tail and 3'UTR lengthening during differentiation, both of which were positively correlated with changes in mRNA abundance, but not translation. Globally, changes in miRNA expression were predominantly associated with mRNA abundance and translation, however several miRNA-mRNA pairings with potential to regulate poly(A) tail length were identified. Furthermore, 3'UTR lengthening was observed to significantly increase the inclusion of non-conserved miRNA binding sites, potentially enhancing the regulatory capacity of these molecules in mature neuronal cells. Together, our findings suggest poly(A) tail length and APA function as part of a rich post-transcriptional regulatory matrix during neuronal differentiation.

MicroRNAs targeting peroxisome proliferator-activated receptor (PPAR) gene are differentially expressed in low birth weight placentae

IntroductionPeroxisome proliferator-activated receptors (PPARs) are activated by natural ligands like fatty acids and influence placental angiogenesis and pregnancy outcome. However, the underlying molecular mechanisms are not clear. This study aims to investigate the association of maternal and placental fatty acid levels with DNA methylation and microRNA regulation of PPARs in the placentae of women delivering low birth weight (LBW) babies. MethodsThis study includes 100 women delivering normal birth weight (NBW) baby and 70 women delivering LBW baby. Maternal and placental fatty acids levels were estimated by gas chromatograph. Gene promoter methylation and mRNA expression of PPARs was analyzed using Epitect Methyl-II PCR assay kit and RT-PCR respectively. Expression of miRNAs targeting PPAR mRNA were analyzed using a Qiagen miRCURY LNA PCR Array on RT-PCR. ResultsPlacental docosahexaenoic acid (DHA) levels and placental mRNA expression of PPARα and PPARγ were lower (p < 0.05 for all) in the LBW group. Differential expression of miRNAs (upregulated miR-33a-5p and miR-22-5p; downregulated miR-301a-5p, miR-518d-5p, miR-27b-5p, miR-106a-5p, miR-21-5p, miR-548d-5p, miR-17-5p and miR-20a-5p) (p < 0.05 for all) was observed in the LBW group. Maternal and placental polyunsaturated fatty acids and total omega-3 fatty acids were positively associated while saturated fatty acids were negatively associated with expression of miRNAs (p < 0.05 for all). Placental expression of miRNAs were positively associated with birth weight (p < 0.05 for all). DiscussionOur data suggests that maternal fatty acid status is associated with changes in the placental expression of miRNAs targeting PPAR gene in women delivering LBW babies.

Peritoneal Fluid Analysis of Advanced Ovarian Cancers after Hyperthermic Intraperitoneal Chemotherapy.

This study investigated miRNA and cytokine expression changes in peritoneal fluid samples of patients with advanced ovarian cancer (OVCA) after receiving hyperthermic intraperitoneal chemotherapy (HIPEC) during cytoreduction surgery (CRS). We collected samples prior to HIPEC, immediately after HIPEC, and 24/48/72 h after CRS from a total of 6 patients. Cytokine levels were assessed using a multiplex cytokine array, and a miRNA PanelChip Analysis System was used for miRNA detection. Following HIPEC, miR-320a-3p, and miR-663-a were found to be immediately down-regulated but increased after 24 h. Further, significant upregulation post-HIPEC and sustained increases in expression were detected in six other miRNAs, including miR-1290, miR-1972, miR-1254, miR-483-5p, miR-574-3p, and miR-574-5p. We also found significantly increased expression of cytokines, including MCP-1, IL-6, IL-6sR, TIMP-1, RANTES, and G-CSF. The changing expression pattern throughout the study duration included a negative correlation in miR-320a-3p and miR-663-a to cytokines including RANTES, TIMP-1, and IL-6 but a positive correlation in miRNAs to cytokines including MCP-1, IL-6sR, and G-CSF. Our study found miRNAs and cytokines in the peritoneal fluid of OVCA patients demonstrated different expression characteristics following CRS and HIPEC. Both changes in expression demonstrated correlations, but the role of HIPEC remains unknown, prompting the need for research in the future.

The evolutionary relationships of microRNAs in the regulation of glucose and lipid metabolism in human and animals

microRNA (miRNA) is a non-coding RNA type, regulating gene expressions at a post-transcriptional level. Changes in miRNA expressions can cause problems such as lipid metabolism disorder, cardiovascular disease, growth retardation, low birth weight and insulin resistance, etc., in human and animals. In this study, we investigated the evolutionary relationships of 14 miRNAs including hsa-miR27a-5p, hsa-miR149-3p, hsa-miR374c-5p, mmu-miR-678, mmu-miR-698-5p, hsa-miR-199a-3p, hsa-miR122-3p, hsa-miR342-3p, mmu-miR201-5p, hsa-miR429, hsa-miR370-3p, hsa-miR130a-5p, hsa-miR330-3p and hsa-miR770-5p related to different metabolic pathways including cardiovascular diseases and lipid metabolism. For this purpose, miRNAs were retrieved from miRBase database. After, Clustal Omega analyses were performed for alignment, and a phylogenetic tree was constructed via MEGA 11. Phylogenetic tree indicated that 14 miRNA sequences were clustered into four groups. One group consisted of mmu-miR-678, and the other 13 sequences were separated into three groups, revealing a close relationship among miRNAs. Findings from different studies provide a new perspective for potential miRNA-based biomarkers to detect lipid metabolism disorders, cardiovascular diseases as well as related disorders.

Erianin as a Promising Novel Agent in the Treatment of Neuroblastoma: The Anticancer Effects and Underlying Molecular Mechanisms.

Erianin is an active dibenzyl compound isolated from Dendrobium officinale and Dendrobium chrysotoxum and there are very few studies on molecular mechanisms and drug targets of erianin. In addition, there is no study investigating the anti-cancer effect of erianin on neuroblastoma cells. The aim of the study is to investigate the anticancer effect of erianin and the underlying mechanism of this effect on SH-SY5Y cells. The effects of erianin on cell viability, invasion and migration were determined by XTT, matrigel chamber and wound healing evaluation, respectively. Expression changes of miRNAs (microRNA) and apoptosis-related genes were evaluated by RT-PCR, and the apoptosis rate was supported by Annexin V evaluation. Erianin significantly decreased cell proliferation, invasion and migration. Erianin administration caused apoptosis by significantly increasing caspase-7, FADD (Fas-associated protein with death domain), BID (BH3 Interacting Domain Death Agonist) and DR5 (Death receptor 5) gene expressions. While the rate of total apoptotic cells was 45.35 ± 6.80% in SH-SY5Y cells treated with erianin, it was 0.133 ± 0.05% in the control group (p = 0.000). In addition, erianin administration significantly decreased the expressions of hsa-miR-155-5p (p = 0.014) and hsa-miR-223-3p (p = 0.004). Also, our study demonstrated for the first time the relationship between erianin and mi-RNAs in a cancer cell. Our study suggests that erianin may be a natural, safe and easily accessible drug candidate that can be used in the treatment of neuroblastoma.

Characterisation of miRNA Expression in Dental Pulp Cells during Epigenetically-Driven Reparative Processes.

Within regenerative endodontics, exciting opportunities exist for the development of next-generation targeted biomaterials that harness epigenetic machinery, including microRNAs (miRNAs), histone acetylation, and DNA methylation, which are used to control pulpitis and to stimulate repair. Although histone deacetylase inhibitors (HDACi) and DNA methyltransferase inhibitors (DNMTi) induce mineralisation in dental pulp cell (DPC) populations, their interaction with miRNAs during DPC mineralisation is not known. Here, small RNA sequencing and bioinformatic analysis were used to establish a miRNA expression profile for mineralising DPCs in culture. Additionally, the effects of a HDACi, suberoylanilide hydroxamic acid (SAHA), and a DNMTi, 5-aza-2'-deoxycytidine (5-AZA-CdR), on miRNA expression, as well as DPC mineralisation and proliferation, were analysed. Both inhibitors increased mineralisation. However, they reduced cell growth. Epigenetically-enhanced mineralisation was accompanied by widespread changes in miRNA expression. Bioinformatic analysis identified many differentially expressed mature miRNAs that were suggested to have roles in mineralisation and stem cell differentiation, including regulation of the Wnt and MAPK pathways. Selected candidate miRNAs were demonstrated by qRT-PCR to be differentially regulated at various time points in mineralising DPC cultures treated with SAHA or 5-AZA-CdR. These data validated the RNA sequencing analysis and highlighted an increased and dynamic interaction between miRNA and epigenetic modifiers during the DPC reparative processes.

Rab3A/Rab27A System Silencing Ameliorates High Glucose-Induced Injury in Podocytes.

Diabetic nephropathy is a major complication in diabetic patients. Podocytes undergo loss and detachment from the basal membrane. Intra- and intercellular communication through exosomes are key processes for maintaining function, and the Rab3A/Rab27A system is an important counterpart. Previously, we observed significant changes in the Rab3A/Rab27A system in podocytes under glucose overload, demonstrating its important role in podocyte injury. We investigated the implication of silencing the Rab3A/Rab27A system in high glucose-treated podocytes and analysed the effect on differentiation, apoptosis, cytoskeletal organisation, vesicle distribution, and microRNA expression in cells and exosomes. For this, we subjected podocytes to high glucose and transfection through siRNAs, and we isolated extracellular vesicles and performed western blotting, transmission electron microscopy, RT-qPCR, immunofluorescence and flow cytometry assays. We found that silencing RAB3A and RAB27A generally leads to a decrease in podocyte differentiation and cytoskeleton organization and an increase in apoptosis. Moreover, CD63-positive vesicles experienced a pattern distribution change. Under high glucose, Rab3A/Rab27A silencing ameliorates some of these detrimental processes, suggesting a differential influence depending on the presence or absence of cellular stress. We also observed substantial expression changes in miRNAs that were relevant in diabetic nephropathy upon silencing and glucose treatment. Our findings highlight the Rab3A/Rab27A system as a key participant in podocyte injury and vesicular traffic regulation in diabetic nephropathy.

The Role of microRNA-23a-3p in the Progression of Human Aging Process by Targeting FOXO3a

Aging results in deterioration of body functions and, ultimately, death. miRNAs contribute to the regulation of aging. The aim of this study was to explore the contribution of miRNAs to aging and senescence-related changes in gene expression. The expression changes of miRNAs in the blood of people and animal samples collected from different age subjects were examined using Affymetrix miRNA 4.0 microarray and qRT-PCR. MTT assay and flow cytometry were used to examine the effect of miR-23a on cell functions in WI-38 cells. The expression levels of 48 miRNAs, including miR-23a, miR-21, and miR-100, in the blood samples were higher in the middle-aged group than in the young or elderly group. Animal studies further suggested that the expression of miR-23a increased with age. In addition, upregulation of miR-23a dramatically suppressed the cell proliferation and arrested the WI-38 cell cycle in vitro. FOXO3a has been identified as a target gene of miR-23a. MiR-23a downregulated the expression of FOXO3a in WI-38 cells. MiRNAs have different expression levels in different age groups. miR-23a could suppress cell proliferation and arrest the cell cycle in WI-38 cells, which elucidated the mechanism through which miR-23a exerts pivotal role in WI-38 cells by targeting FOXO3a.

Sleep Deprivation Promotes Endothelial Inflammation and Atherogenesis by Reducing Exosomal miR-182-5p.

Insufficient or disrupted sleep increases the risk of cardiovascular disease, including atherosclerosis. However, we know little about the molecular mechanisms by which sleep modulates atherogenesis. This study aimed to explore the potential role of circulating exosomes in endothelial inflammation and atherogenesis under sleep deprivation status and the molecular mechanisms involved. Circulating exosomes were isolated from the plasma of volunteers with or without sleep deprivation and mice subjected to 12-week sleep deprivation or control littermates. miRNA array was performed to determine changes in miRNA expression in circulating exosomes. Although the total circulating exosome levels did not change significantly, the isolated plasma exosomes from sleep-deprived mice or human were a potent inducer of endothelial inflammation and atherogenesis. Through profiling and functional analysis of the global microRNA in the exosomes, we found miR-182-5p is a key exosomal cargo that mediates the proinflammatory effects of exosomes by upregulation of MYD88 (myeloid differentiation factor 88) and activation of NF-ĸB (nuclear factor kappa-B)/NLRP3 pathway in endothelial cells. Moreover, sleep deprivation or the reduction of melatonin directly decreased the synthesis of miR-182-5p and led to the accumulation of reactive oxygen species in small intestinal epithelium. The findings illustrate an important role for circulating exosomes in distant communications, suggesting a new mechanism underlying the link between sleep disorder and cardiovascular disease.

Different miRNAs Related to FBXW7 Mutations or High Mitotic Indices Contribute to Rectal Neuroendocrine Tumors: A Pilot Study.

Recent studies suggest that miRNA may be involved in the development of rectal neuroendocrine tumors (NETs). We explored the frequency of clinicopathologically relevant mutations and miRNA expression in rectal NETs to examine molecular profiles related to prognosis and behavior. Twenty-four eligible specimens with endoscopically excised rectal NETs were selected. Next-generation sequencing and an miRNA expression assay were used to evaluate the expression profile relevant to common genetic mutations in rectal NETs. Kyoto Encyclopedia of Genes and Genomes analysis predicted that the possible target signaling pathways were correlated with dysregulated miRNAs. Nineteen rectal NETs harbored more than one mutation in the 24 cancer-related genes. Seven miRNAs (hsa-miR-769-5p, hsa-miR-221-3p, hsa-miR-34a-5p, hsa-miR-181c-5p, hsa-miR-1246, hsa-miR-324-5p, and hsa-miR-361-3p) were significantly down-regulated in tumors harboring the FBWX7 mutation. Unsupervised hierarchical clustering analysis showed that up-regulation of these seven miRNAs may result in high mitotic indices, indicating the role of miRNAs in tumor progression. Among the down-regulated miRNAs, hsa-miR-769-5p was strongly correlated with extracellular matrix-receptor interaction and lysine degradation. Among the clinicopathological factors, up-regulated hsa-miR-3934-5p was linked to an increased mitotic count. No change in miRNA expression was associated with a tumor size >1 cm, lymphovascular invasion, or Ki-67 index. In summary, we identified different miRNA signatures involved in FBXW7 mutations or high mitotic indices in rectal NETs, which may play a critical role in tumor behavior.

Treatment with benznidazole and pentoxifylline regulates microRNA transcriptomic profile in a murine model of Chagas chronic cardiomyopathy

Chronic Chagas cardiomyopathy (CCC) is one of the leading causes of morbidity and mortality due to cardiovascular disorders in endemic areas of Chagas disease (CD), a neglected tropical illness caused by the protozoan parasite Trypanosoma cruzi. CCC is characterized by parasite persistence and inflammatory response in the heart tissue, which occur parallel to microRNA (miRNA) alterations. Here, we investigated the miRNA transcriptome profiling in the cardiac tissue of chronically T. cruzi-infected mice treated with a suboptimal dose of benznidazole (Bz), the immunomodulator pentoxifylline alone (PTX), or the combination of both (Bz+PTX), following the CCC onset. At 150 days post-infection, Bz, PTX, and Bz+PTX treatment regimens improved electrocardiographic alterations, reducing the percentage of mice afflicted by sinus arrhythmia and second-degree atrioventricular block (AVB2) when compared with the vehicle-treated animals. miRNA Transcriptome profiling revealed considerable changes in the differential expression of miRNAs in the Bz and Bz+PTX treatment groups compared with the control (infected, vehicle-treated) group. The latter showed pathways related to organismal abnormalities, cellular development, skeletal muscle development, cardiac enlargement, and fibrosis, likely associated with CCC. Bz-Treated mice exhibited 68 differentially expressed miRNAs related to signaling pathways like cell cycle, cell death and survival, tissue morphology, and connective tissue function. Finally, the Bz+PTX-treated group revealed 58 differentially expressed miRNAs associated with key signaling pathways related to cellular growth and proliferation, tissue development, cardiac fibrosis, damage, and necrosis/cell death. The T. cruzi-induced upregulation of miR-146b-5p, previously shown in acutely infected mice and in vitro T. cruzi-infected cardiomyocytes, was reversed upon Bz and Bz+PTX treatment regimens when further experimentally validated. Our results further our understanding of molecular pathways related to CCC progression and evaluation of treatment response. Moreover, the differentially expressed miRNAs may serve as drug targets, associated molecular therapy, or biomarkers of treatment outcomes.

Serum miRNA modulations indicate changes in retinal morphology.

Age-related macular degeneration (AMD) is the leading cause of vision loss in the developed world and the detection of its onset and progression are based on retinal morphological assessments. MicroRNA (miRNA) have been explored extensively as biomarkers for a range of neurological diseases including AMD, however differences in experimental design and the complexity of human biology have resulted in little overlap between studies. Using preclinical animal models and clinical samples, this study employs a novel approach to determine a serum signature of AMD progression. Serum miRNAs were extracted from mice exposed to photo-oxidative damage (PD; 0, 1, 3 and 5 days), and clinical samples from patients diagnosed with reticular pseudodrusen or atrophic AMD. The expression of ~800 miRNAs was measured using OpenArray™, and differential abundance from controls was determined using the HTqPCR R package followed by pathway analysis with DAVID. MiRNA expression changes were compared against quantifiable retinal histological indicators. Finally, the overlap of miRNA changes observed in the mouse model and human patient samples was investigated. Differential miRNA abundance was identified at all PD time-points and in clinical samples. Importantly, these were associated with inflammatory pathways and histological changes in the retina. Further, we were able to align findings in the mouse serum to those of clinical patients. In conclusion, serum miRNAs are a valid tool as diagnostics for the early detection of retinal degeneration, as they reflect key changes in retinal health. The combination of pre-clinical animal models and human patient samples led to the identification of a preliminary serum miRNA signature for AMD. This study is an important platform for the future development of a diagnostic serum miRNA panel for the early detection of retinal degeneration.

Effect of Mir-4270 Inhibitor and Mimic on Viability and Stemness in Gastric Cancer Stem-Like Cells Derived from MKN-45 Cell Line.

MicroRNAs (miRNAs) are significant regulatory factors in stem cell proliferation, and change in miRNA expression influences the cancer stem cell viability and gene expression. Herein, we evaluated the effect of the hsa-miR-4270 inhibitor and its mimic on the expression of stem cell markers in gastric cancer (GC) stem-like cells. GC stem-like cells were isolated from the MKN-45 cell line by a non-adherent surface system. The cells were confirmed by differentiation assays using dexamethasone and insulin as adipogenesis-inducing agents and also Staurosporine as a neural-inducing agent. Isolated GC stem-like cells were treated with different concentrations (0, 15, 20, 25, 30, 40, 50, and 60 nM) of hsa-miR-4270 inhibitor and its mimic. The quantity of cell viability was determined by trypan blue method. Transcription of the stem cell marker genes, including CD44, OCT3/4, SOX2, Nanog, and KLF4, was evaluated by real-time RT-PCR. The results showed that GC stem-like cells were differentiated into both adipose cells using dexamethasone and insulin and neural cells by Staurosporine. Treatment of GC stem-like cells with hsa-miR-4270 inhibitor decreased cell viability and downregulated OCT3/4, CD44, and Nanog to 86%, 79%, and 91% respectively. Also, SOX2 and KLF4 were overexpressed to 8.1- and 1.94-folds, respectively. However, hsa-miR-4270 mimic had opposite effects on the cell viability and gene expression of the stem cell markers. The effect of hsa-miR-4270 inhibitor and its mimic on the expression of the stem cell markers in GCSCs indicated that hsa-miR-4270 stimulates the stemness property of GCSCs, likely through stimulating the development of gastric stem cells.

Potential of Circulating miRNAs as Molecular Markers in Mood Disorders and Associated Suicidal Behavior.

Mood disorders are the most prevalent psychiatric disorders associated with significant disability, morbidity, and mortality. The risk of suicide is associated with severe or mixed depressive episodes in patients with mood disorders. However, the risk of suicide increases with the severity of depressive episodes and is often presented with higher incidences in bipolar disorder (BD) patients than in patients with major depression (MDD). Biomarker study in neuropsychiatric disorders is critical for developing better treatment plans by facilitating more accurate diagnosis. At the same time, biomarker discovery also provides more objectivity to develop state-of-the-art personalized medicine with increased accuracy through clinical interventions. Recently, colinear changes in miRNA expression between brain and systemic circulation have added great interest in examining their potential as molecular markers in mental disorders, including MDD, BD, and suicidality. A present understanding of circulating miRNAs in body fluids implicates their role in managing neuropsychiatric conditions. Most notably, their use as prognostic and diagnostic markers and their potential role in treatment response have significantly advanced our knowledge base. The present review discusses circulatory miRNAs and their underlying possibilities to be used as a screening tool for assessing major psychiatric conditions, including MDD, BD, and suicidal behavior.

Differential Expression Analysis of Blood MicroRNA in Identifying Potential Genes Relevant to Alzheimer’s Disease Pathogenesis, Using an Integrated Bioinformatics and Machine Learning Approach

Alzheimer’s disease (AD) is a neurodegenerative disease characterized by cognitive and functional impairment. Recent research has focused on the deregulation of microRNAs (miRNAs) in blood as the potential biomarkers for AD. As such, a differential expression analysis of miRNAs was conducted in this study using an integrated framework that utilized the advantages of statistical and machine learning approaches. Three miRNA candidates that showed the strongest significance and correlation with each other, namely hsa-miR-6501-5p, hsa-miR-4433b-5p, and hsa-miR-143-3p, were identified. The roles and functions of the identified differentiated miRNA candidates with AD development were verified by predicting their target mRNAs, and their networks of interaction in AD pathogenesis were investigated. Pathway analysis showed that the pathways involved in contributing to the development of AD included oxidative phosphorylation, mitochondrial dysfunction, and calcium-mediated signalling. This study supports evidence that the miRNA expression changes in AD and indicates the need for further study in this area.

Comparative Transcriptome Profiling Reveals Key MicroRNAs and Regulatory Mechanisms for Aluminum Tolerance in Olive.

Aluminum toxicity (Al) is one of the major constraints to crop production in acidic soils. MicroRNAs (miRNAs) have emerged as key regulatory molecules at post-transcriptional levels, playing crucial roles in modulating various stress responses in plants. However, miRNAs and their target genes conferring Al tolerance are poorly studied in olive (Olea europaea L.). Here, genome-wide expression changes in miRNAs of the roots from two contrasting olive genotypes Zhonglan (ZL, Al-tolerant) and Frantoio selezione (FS, Al-sensitive) were investigated by high-throughput sequencing approaches. A total of 352 miRNAs were discovered in our dataset, consisting of 196 conserved miRNAs and 156 novel miRNAs. Comparative analyses showed 11 miRNAs have significantly different expression patterns in response to Al stress between ZL and FS. In silico prediction identified 10 putative target gene of these miRNAs, including MYB transcription factors, homeobox-leucine zipper (HD-Zip) proteins, auxin response factors (ARF), ATP-binding cassette (ABC) transporters and potassium efflux antiporter. Further functional classification and enrichment analysis revealed these Al-tolerance associated miRNA-mRNA pairs are mainly involved in transcriptional regulation, hormone signaling, transportation and metabolism. These findings provide new information and perspectives into the regulatory roles of miRNAs and their target for enhancing Al tolerance in olives.

Alterations in microRNA of extracellular vesicles associated with major depression, attention-deficit/hyperactivity and anxiety disorders in adolescents

Extracellular vesicles (EVs) are present in numerous peripheral bodily fluids and function in critical biological processes, including cell-to-cell communication. Most relevant to the present study, EVs contain microRNAs (miRNAs), and initial evidence from the field indicates that miRNAs detected in circulating EVs have been previously associated with mental health disorders. Here, we conducted an exploratory longitudinal and cross-sectional analysis of miRNA expression in serum EVs from adolescent participants. We analyzed data from a larger ongoing cohort study, evaluating 116 adolescent participants at two time points (wave 1 and wave 2) separated by three years. Two separate data analyses were employed: A cross-sectional analysis compared individuals diagnosed with Major Depressive Disorder (MDD), Anxiety disorders (ANX) and Attention deficit/Hyperactivity disorder (ADHD) with individuals without psychiatric diagnosis at each time point. A longitudinal analysis assessed changes in miRNA expression over time between four groups showing different diagnostic trajectories (persistent diagnosis, first incidence, remitted and typically developing/control). Total EVs were isolated, characterized by size distribution and membrane proteins, and miRNAs were isolated and sequenced. We then selected differentially expressed miRNAs for target prediction and pathway enrichment analysis. In the longitudinal analysis, we did not observe any statistically significant results. In the cross-sectional analysis: in the ADHD group, we observed an upregulation of miR-328-3p at wave 1 only; in the MDD group, we observed a downregulation of miR-4433b-5p, miR-584-5p, miR-625-3p, miR-432-5p and miR-409-3p at wave 2 only; and in the ANX group, we observed a downregulation of miR-432-5p, miR-151a-5p and miR-584-5p in ANX cases at wave 2 only. Our results identified previously observed and novel differentially expressed miRNAs and their relationship with three mental health disorders. These data are consistent with the notion that these miRNAs might regulate the expression of genes associated with these traits in genome-wide association studies. The findings support the promise of continued identification of miRNAs contained within peripheral EVs as biomarkers for mental health disorders.

Selection and Validation of miRNA Reference Genes by Quantitative Real-Time PCR Analysis in Paeonia suffruticosa

The miRNA, a kind of endogenous non-coding small RNA, plays an essential role in regulation of gene expression in plants. Quantitative real-time PCR (qRT-PCR) assay is one of the most common methods used for quantification of miRNA expression, and levels of expression are normalized by comparing with reference genes. The present study was intended to identify the appropriate reference genes for normalizing the level of miRNA expression in various developmental stages and tissues such as the bud development process, flower development stages, and different tissues of tree peony of different flowering times. Five algorithms (Delta Ct, geNorm, NormFinder, BestKeeper, and RefFinder) were used for stability analysis. The results showed that mtr-MIR160b-p3 and gma-miR394a-5p were the most stable miRNAs expressed during the bud development process of early-flowering tree peony ‘Feng dan’; PC-5p-19095 was the most stable during the bud development process of late-flowering tree peony ‘Lian he’, followed by gma-miR394a-5p and mtr-MIR160b-p3; the mtr-miR159a was the most stable miRNA expressed in the flower development stages of different tree peony varieties. The PC-3p-871 was the most stable miRNA expressed in different tissues of early-flowering tree peony ‘Feng dan’, followed by PC-5p-4, and PC-5p-4 was the most stable in late-flowering tree peony ‘Lian he’, followed by the mtr-miR168b. The findings of this study provide a reference for studying the changes in miRNA expression, and further exploring the regulatory mechanism of miRNA in tree peony.

Identification and analysis of key microRNAs derived from osteoarthritis synovial fluid exosomes.

Identification and analysis of key microRNAs derived from osteoarthritis synovial fluid exosomes.