Expression Of miRNA Targets Research Articles

Analysis of Polyphyllin II alters microRNA expression profile in lung cancer A549 cells

Non-small cell lung cancer (NSCLC) is a malignant tumor that threatens the whole world, previous studies found that Polyphyllin II (PPII) can suppress NSCLC cells growth, exhibits significant antitumor activity. MicroRNAs (miRNAs) can regulate the expression of other genes and play a crucial role in the prevention and development of cancer. However, the miRNA portrait of ginsenoside PPII-treated NSCLC A549 cells has not yet been studied. In this work, miRNA-seq analysis was used to determine the changes in miRNA expression profile of NSCLC A549 cells PPII-treaded. In addition, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were performed on differentially expressed miRNA target genes. Then, predicted the target genes of miRNA and performed functional enrichment analysis on them. We identified 58 up-regulated and 24 down-regulated miRNAs displaying changes their expression in PPII treated A549 cells were greater than 2-fold. In addition, the expression levels of miRNAs were validated by real-time PCR. Therefore, this work has a certain promoting effect on the study of the anti-cancer mechanism of PPII in lung cells.

The expression profile of brain-derived exosomal miRNAs reveals the key molecules responsible for spontaneous motor function recovery in a rat model with permanent middle cerebral artery occlusion.

The analysis of alterations in the expression and functionality of brain-derived exosomal miRNAs within ischemic stroke lesions provides significant insights into the mechanisms that contribute to disease recovery. We assessed spontaneous motor function in a rat model of permanent middle cerebral artery occlusion (pMCAO) using motor function scores and magnetic resonance imaging (MRI). Brain-derived exosomes from the infarcted brain tissue of the animal model were extracted and high-throughput sequencing of them was performed followed by bioinformatics analysis for differentially expressed miRNAs target genes. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to measure expression levels of differentially expressed miRNAs at various time points. The oxygen-glucose deprivation (OGD) model was established to investigate gene function through the assessment of cell proliferation and apoptosis using EdU proliferation and JC-1 apoptosis assay. The rat model demonstrated a spontaneous recovery of motor function and a reduction in cerebral infarction area from day 1 to day 14 post-operation. Over the course of the recovery period, miR-24-3p, miR-129-1-3p, and miR-212-5p maintained consistent expression levels, reaching their peak on the initial day following surgery. In the cell model, EdU detection indicated that miR-129-1-3p promoted cellular proliferation, while JC-1 detection revealed its suppressive impact on cellular apoptosis. The current research findings indicated the presence of spontaneous motor function restoration in a rat model of ischemic stroke. MiR-24-3p, miR-129-1-3p, and miR-212-5p were identified as pivotal genes in this recovery process, with miR-129-1-3p potentially influencing the restoration of spontaneous motor function in ischemic stroke through the regulation of neuronal proliferation and apoptosis.

Dysregulated MicroRNAs in Chronic Lymphocytic Leukemia.

MiRNAs are a class of non-coding RNAs acting as gene expression regulators by modulating the lifespan of messenger RNA. Commonly referred to as the most frequent leukemia in the Western world, chronic lymphocytic leukemia (CLL) is a lymphoproliferative malignancy characterized by clonal expansion of CD19, CD23, and CD5-positive mature B-cells. While this pathology is regarded as less aggressive and has a variety of treatment options, the cause of its clinical heterogeneity is not yet understood. Moreover, the prognostic markers and treatment recommendations based on predictive markers are limited. This review aims to investigate some miRNAs that are dysregulated and possibly involved in CLL pathogenesis as a starting point for the proposal of new prognostic and predictive markers and, as more agents targeting miRNA expression become available, their potential role as therapeutic targets.

Combinatorial microRNA activity is essential for the transition of pluripotent cells from proliferation into dormancy.

Dormancy is a key feature of stem cell function in adult tissues as well as in embryonic cells in the context of diapause. The establishment of dormancy is an active process that involves extensive transcriptional, epigenetic, and metabolic rewiring. How these processes are coordinated to successfully transition cells to the resting dormant state remains unclear. Here we show that microRNA activity, which is otherwise dispensable for preimplantation development, is essential for the adaptation of early mouse embryos to the dormant state of diapause. In particular, the pluripotent epiblast depends on miRNA activity, the absence of which results in the loss of pluripotent cells. Through the integration of high-sensitivity small RNA expression profiling of individual embryos and protein expression of miRNA targets with public data of protein-protein interactions, we constructed the miRNA-mediated regulatory network of mouse early embryos specific to diapause. We find that individual miRNAs contribute to the combinatorial regulation by the network, and the perturbation of the network compromises embryo survival in diapause. We further identified the nutrient-sensitive transcription factor TFE3 as an upstream regulator of diapause-specific miRNAs, linking cytoplasmic MTOR activity to nuclear miRNA biogenesis. Our results place miRNAs as a critical regulatory layer for the molecular rewiring of early embryos to establish dormancy.

MicroRNA expression profile of chicken liver at different times after Histomonas meleagridis infection

Histomonas meleagridis, an anaerobic intercellular parasite, is known to infect gallinaceous birds, particularly turkeys and chickens. The resurgence of histomonosis in recent times has resulted in significant financial setbacks due to the prohibition of drugs used for disease treatment. Currently, research on about H. meleagridis primarily concentrate on the examination of its virulence, gene expression analysis, and the innate immunity response of the host organism. However, there is a lack of research on differentially expressed miRNAs (DEMs) related to liver infection induced by H. meleagridis. In this study, the weight gain and pathological changes at various post-infection time points were evaluated through animal experiments to determine the peak and early stages of infection. Next, High-throughput sequencing was used to examine the expression profile of liver miRNA at 10 and 15 days post-infection (DPI) in chickens infected with the Chinese JSYZ-F strain of H. meleagridis. A comparison with uninfected controls revealed the presence of 120 and 118 DEMs in the liver of infected chickens at 10 DPI and 15 DPI, respectively, with 74 DEMs being shared between the two time points. Differentially expressed microRNAs (DEMs) were categorized into three groups based on the time post-infection. The first group (L1) includes 45 miRNAs that were differentially expressed only at 10 DPI and were predicted to target 1646 genes. The second group (L2) includes 43 miRNAs that were differentially expressed only at 15 DPI and were predicted to target 2257 genes. The third group (L3) includes 75 miRNAs that were differentially expressed at both 10 DPI and 15 DPI and were predicted to target 1623 genes. At L1, L2, and L3, there were 89, 87, and 41 significantly enriched Gene Ontology (GO) terms, respectively (p<0.05). The analysis of differentially expressed miRNA target genes using KEGG pathways revealed significant enrichment at L1, L2, and L3, with 3, 4, and 5 pathways identified, respectively (p<0.05). This article suggests that the expression of liver miRNA undergoes dynamic alterations due to H. meleagridis and the host. It showed that the expression pattern of L1 class DEMs was more conducive to regulating the development of the inflammatory response, while the L2 class DEMs were more conducive to augmenting the inflammatory response. The observed patterns of miRNA expression associated with inflammation were in line with the liver's inflammatory process following infection. The results of this study provide a basis for conducting a comprehensive analysis of the pathogenic mechanism of H. meleagridis from the perspective of host miRNAs.

MiRNA Expression Analysis of IPEC-J2 Cells Damaged by Soybean 7S Globulin Reveals ssc-miR-221-5p as the Factor Alleviating Cell Damage.

The most significant and sensitive antigen protein that causes diarrhea in weaned pigs is soybean 7S globulin. Therefore, identifying the primary target for minimizing intestinal damage brought on by soybean 7S globulin is crucial. MicroRNA (miRNA) is closely related to intestinal epithelium's homeostasis and integrity. However, the change of miRNAs' expression and the function of miRNAs in Soybean 7S globulin injured-IPEC-J2 cells are still unclear. In this study, the miRNAs' expression profile in soybean 7S globulin-treated IPEC-J2 cells was investigated. Fifteen miRNAs were expressed differently. The differentially expressed miRNA target genes are mainly concentrated in signal release, cell connectivity, transcriptional inhibition, and Hedgehog signaling pathway. Notably, we noticed that the most significantly decreased miRNA was ssc-miR-221-5p after soybean 7S globulin treatment. Therefore, we conducted a preliminary study on the mechanisms of ssc-miR-221-5p in soybean 7S globulin-injured IPEC-J2 cells. Our research indicated that ssc-miR-221-5p may inhibit ROS production to alleviate soybean 7S globulin-induced apoptosis and inflammation in IPEC-J2 cells, thus protecting the cellular mechanical barrier, increasing cell proliferation, and improving cell viability. This study provides a theoretical basis for the prevention and control of diarrhea of weaned piglets.

An optimized microRNA detection platform based on PAM formation-regulated CRISPR/Cas12a activation

MicroRNAs (miRNAs) have emerged as biomarkers for the diagnosis and prognosis of various diseases, such as cancer. Recent advancements in CRISPR/Cas12a-based biosensors in combination with hybridization chain reaction (HCR) make it a promising approach for miRNA detection. To increase the compatibility of HCR and CRISPR/Cas12a, we compared two design strategies of hairpin DNA in HCR. The results showed that different arrangements of the protospacer sequence and protospacer adjacent motif (PAM) in the hairpin DNA could affect the sensing performance. The “PAM Formation” strategy, by which the duplex PAM sites are absent in the hairpin DNA and present in the long duplex DNA after HCR, exhibited advantages in detection sensitivity. By optimizing the probe sequences and reaction conditions, we developed a miRNA detection platform. With the same crRNA, this platform enables the identification of different miRNAs by simply replacing the loop region of the target recognition probe. In addition, the proposed platform can detect single-stranded DNA and distinguishing single or multiple base mutations in the target strand. The application of discriminating the target miRNA expression levels from different cell lines validated the reliability and practicability of the sensor platform, indicating its potential applications in early clinical accurate diagnosis of cancers.

Bioinformatics analysis of the microRNA genes associated with type 2 cardiorenal syndrome

BackgroundMicroRNAs (miRNAs) are important regulatory factors in the normal developmental stages of the heart and kidney. However, it is currently unclear how miRNA is expressed in type 2 cardiorenal syndrome (CRS). This study aimed to detect the differential expression of miRNAs and to clarify the main enrichment pathways of differentially expressed miRNA target genes in type 2 CRS.MethodsFive cases of healthy control (Group 1), eight of chronic heart failure (CHF, Group 2) and seven of type 2 CRS (Group 3) were enrolled, respectively. Total RNA was extracted from the peripheral blood of each group. To predict the miRNA target genes and biological signalling pathways closely related to type 2 CRS, the Agilent miRNA microarray platform was used for miRNA profiling and bioinformatics analysis of the isolated total RNA samples.ResultsAfter the microarray analysis was done to screen for differentially expressed circulating miRNAs among the three different groups of samples, the target genes and bioinformatic pathways of the differential miRNAs were predicted. A total of 38 differential miRNAs (15 up- and 23 down-regulated) were found in Group 3 compared with Group 1, and a total of 42 differential miRNAs (11 up- and 31 down-regulated) were found in Group 3 compared to Group 2. According to the Gene Ontology (GO) function and Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway analysis, the top 10 lists of molecular functions, cellular composition and biological processes, and the top 30 signalling pathways of predicted gene targets of the differentially expressed miRNAs were discriminated among the three groups.ConclusionBetween the patients with CHF and type 2 CRS, miRNAs were differentially expressed. Prediction of target genes of differentially expressed miRNAs and the use of GO function and KEGG pathway analysis may reveal the molecular mechanisms of CRS. Circulating miRNAs may contribute to the diagnosis of CRS, and further and larger studies are needed to enhance the robustness of our findings.

Investigation of the potential of miRNA candidates as non-invasive biomarkers for the diagnosis and follow-up of colorectal cancer

BackgroundColorectal cancer (CRC) is the third most common cancer worldwide, and despite advances in treatment, molecular biomarkers are needed for both early diagnosis and prognosis monitoring. It is known that microRNAs (miRNA), one of the epigenetic mechanisms, are effective in the initiation and development of cancer by regulating the activity of tumor suppressors and/or oncogenes. In this study, the potential of the molecules let-7, miRNA125b, and miRNA30a, which are known to play a role in cellular processes, as biomarkers for colorectal cancer and their molecular mechanisms were investigated in this model. The aim was to evaluate the diagnostic, prognostic, and predictive utility of the target miRNAs in colorectal cancer patients. Material and methodsThe expression changes of miRNAs let-7, miRNA125b, and miRNA30a were investigated by miRNAs isolation and cDNA synthesis from the serum samples of 60 patients diagnosed with CRC or from the serum samples of 20 healthy individuals. The calculation was performed using the quantitative real-time polymerase chain reaction method to determine the expression level. The results were compared with clinical parameters. ResultAn 8-fold decrease in the expression of let-7 and miRNA125b and a 60-fold decrease in the expression of miRNA30a were found in the serum samples of patients diagnosed with colorectal cancer (CRC) compared to the healthy group. A decrease in let-7 was observed in 53.3%, miRNA125b in 58.3%, and miRNA30a in 55% of patients. A significant correlation was found between the reduced expression status and the stage, lymph nodes, local recurrence, and metastasis (p < 0.05). The ROC analysis showed that the miRNA30a level could be a diagnostic biomarker for CRC (p < 0.001). No significant impact of target miRNA expression changes on overall disease survival was observed. ConclusionIt is thought that the target miRNA30a can be used for early diagnosis and screening and that the target miRNA let-7, miRNA125b, and miRNA30a can be used as non-invasive biomarkers for disease follow-up, with larger patient studies being conducted on CRC patients.

Mesenchymal stem cell-regulated miRNA-mRNA landscape in acute-on-chronic liver failure

BackgroundAcute-on-chronic liver failure (ACLF) is a major challenge in the field of hepatology. While mesenchymal stem cell (MSC) therapy can improve the prognosis of patients with ACLF, the molecular mechanisms through which MSCs attenuate ACLF remain poorly understood. We performed global miRNA and mRNA expression profiling via next-generation sequencing of liver tissues from MSC-treated ACLF mice to identify important signaling pathways and major factors implicated in ACLF alleviation by MSCs. MethodsCarbon tetrachloride-induced ACLF mice were treated with saline or mouse bone marrow-derived MSCs. Mouse livers were subjected to miRNA and mRNA sequencing. Related signal transduction pathways were obtained through Gene Set Enrichment Analysis. Functional enrichment, protein-protein interaction, and immune infiltration analyses were performed for the differentially expressed miRNA target genes (DETs). Hub miRNA and mRNA associated with liver injury were analyzed using LASSO regression. The expression levels of hub genes were subjected to Pearson's correlation analysis and verified using RT-qPCR. The biological functions of hub genes were verified in vitro. ResultsThe tricarboxylic acid cycle and peroxisome proliferator-activated receptor pathways were activated in the MSC-treated groups. The proportions of liver-infiltrating NK resting cells, M2 macrophages, follicular helper T cells, and other immune cells were altered after MSC treatment. The expression levels of six miRNAs and 10 transcripts correlated with the degree of liver injury. miR-27a-5p was downregulated in the mouse liver after MSC treatment, while its target gene E2f2 was upregulated. miR-27a-5p inhibited E2F2 expression, suppressed G1/S phase transition and proliferation of hepatocytes, in addition to promoting their apoptosis. ConclusionsThis is the first comprehensive analysis of miRNA and mRNA expression in the liver tissue of ACLF mice after MSC treatment. The results revealed global changes in hepatic pathways and immune subpopulations. The miR-27a-5p/E2F2 axis emerged as a central regulator of the MSC-induced attenuation of ACLF. The current findings improve our understanding of the molecular mechanisms through which MSCs alleviate ACLF.

Integrative bioinformatics analysis of miRNA and mRNA expression profiles identified some potential biomarkers for breast cancer

BackgroundBreast cancer is a common cause of cancer death among women with a complex and heterogeneous picture in histological, molecular and clinical features. The aim of this study was to identify hub gene and their target microRNAs in related pathways for breast cancer.MethodsWe selected screening methods for differentially expressed mRNAs and miRNAs using expression profile data of breast cancer from the cancer genome atlas. Using some databases for annotation, the functional and pathway enrichment for differential expression genes was performed. We selected genes and miRNAs with differential expression pattern. Then we determined target genes for differential expression miRNAs (DEMIs) and intersection between them was selected as differentially expressed miRNA–target genes for breast cancer. In the next step, we constructed miRNA–mRNA regulatory network and protein–protein interaction (PPI) network for more information.ResultsTop 10 DEMIs were identified from miRNA profile. Then, we selected 354 genes as target gene for 10 DEMIs. The miRNA–mRNA and PPI network were constructed, and 10 hub genes and 5 miRNAs identified that some of them are new for breast cancer. Also, miRNA–target genes with differential expressions in this study were all mainly involved in signaling pathways and developmental process.ConclusionThis study identified some candidate biomarkers for breast cancer that they have a potential role in pathways related to breast. These findings can be used for research, early diagnosis and therapeutic goals.

P-673 Women exposed to higher levels of DEHP display altered microRNA expression profile in the pre-ovulatory follicular fluid

Abstract Study question Can higher concentrations of di-2-ethylhexyl phthalate (DEHP) in follicular fluid (FF) lead to an altered microRNA expression profile? Summary answer The present study indicates the upregulation of miR-203a-3p, miR-150-5p and miR-28-3p in the follicular fluid of women with higher DEHP concentrations (FDR&amp;lt;0.05). What is known already Women are exposed daily to endocrine-disrupting chemicals (EDCs) such as phthalates which can be found in hundreds of personal care products and plastic items. In studies of rodents exposed to certain phthalates, high doses have been shown to change hormone levels, disrupt folliculogenesis and cause birth defects. Recently, we reported a strong inverse association between DEHP metabolites and ovarian sensitivity index (OSI) in females undergoing ART treatment. Several mechanisms have been proposed to explain the association between DEHP and fertility in rodents, however not much is known about the mechanisms in humans. Study design, size, duration FF was collected from 96 women undergoing IVF treatment after controlled ovarian stimulation during ovarian puncture. 48 participants from Estonia were recruited at Nova Vita Clinic AS in Tallinn, Estonia in 2019 and 48 participants from Sweden were recruited at Carl von Linnekliniken in Uppsala, Sweden in 2016. In both cohorts, patients signed a written informed consent form. Both cohorts were matched for age, BMI, parity and the number of previous IVF treatments. Participants/materials, setting, methods DEHP metabolite concentrations in the FF samples were measured in our earlier study by LC-MS/MS. Cell free RNA was extracted from FF samples with Qiagen miRNeasy Micro kit, sequencing libraries were prepared with QIAseq miRNA library kit and sequenced on Illumina NextSeq 550 instrument. Differential miRNA expression analysis was performed with the DESeq2 package. Pathway enrichment analysis for differentially expressed miRNA targets was conducted with the miEAA tool integrated with WikiPathways. Main results and the role of chance In total, 465 miRNAs were observed at least twice in ≥ 24 samples. Differential expression analysis was conducted between DEHP-high and DEHP-low groups in combined cohorts using the cohort as a covariate. 16 miRNAs were found to be differentially expressed between DEHP groups with FDR&amp;lt;0.1. Three miRNAs: miR-203a-3p, miR-150-5p and miR-28-3p, were differentially expressed with FDR&amp;lt;0.05 and were more abundant in women with high DEHP-levels in both cohorts. Of these, miR-203a-3p was negatively correlated with OSI (Pearson R = -0.27, p = 0.0089). Six miRNAs (miR-10b-5p, miR-112-5p, miR-132-5p, miR-203-3p, miR-205-5p and miR-27-3p) were found to have predicted targets in the androgen receptor signalling pathway. Limitations, reasons for caution The miRNAs found in this study need to be validated in a larger set of patients as a cohort-dependent difference in the results was observed. There is a need for further functional studies to investigate miRNAs as possible mediators of the association between DEHP and ovarian sensitivity index. Wider implications of the findings The current study presents evidence that DEHP exposure alters the expression of FF miRNAs which are associated with androgen receptor signalling. These results provide novel evidence of the potential influence of endocrine-disrupting chemicals on female reproduction. Trial registration number NA

MicroRNAs Differentially Expressed in Actinic Keratosis and Healthy Skin Scrapings.

Actinic keratosis (AK) is a carcinoma in situ precursor of cutaneous squamous cell carcinoma (cSCC), the second most common cancer affecting the Caucasian population. AK is frequently present in the sun-exposed skin of the elderly population, UV radiation being the main cause of this cancer, and other risk factors contributing to AK incidence. The dysregulation of microRNAs (miRNAs) observed in different cancers leads to an improper expression of miRNA targets involved in several cellular pathways. The TaqMan Array Human MicroRNA Card assay for miRNA expression profiling was performed in pooled AK compared to healthy skin scraping samples from the same patients. Forty-three miRNAs were modulated in the AK samples. The expression of miR-19b (p < 0.05), -31, -34a (p < 0.001), -126, -146a (p < 0.01), -193b, and -222 (p < 0.05) was validated by RT-qPCR. The MirPath tool was used for MiRNA target prediction and enriched pathways. The top DIANA-mirPath pathways regulated by the targets of the 43 miRNAs are TGF-beta signaling, Proteoglycans in cancer, Pathways in cancer, and Adherens junction (7.30 × 10-10 < p < 1.84 × 10-8). Selected genes regulating the KEGG pathways, i.e., TP53, MDM2, CDKN1A, CDK6, and CCND1, were analyzed. MiRNAs modulated in AK regulate different pathways involved in tumorigenesis, indicating miRNA regulation as a critical step in keratinocyte cancer.

Targeting miRNAs and Other Non-Coding RNAs as a Therapeutic Approach: An Update.

Since the discovery of the first microRNAs (miRNAs, miRs), the understanding of miRNA biology has expanded substantially. miRNAs are involved and described as master regulators of the major hallmarks of cancer, including cell differentiation, proliferation, survival, the cell cycle, invasion, and metastasis. Experimental data indicate that cancer phenotypes can be modified by targeting miRNA expression, and because miRNAs act as tumor suppressors or oncogenes (oncomiRs), they have emerged as attractive tools and, more importantly, as a new class of targets for drug development in cancer therapeutics. With the use of miRNA mimics or molecules targeting miRNAs (i.e., small-molecule inhibitors such as anti-miRS), these therapeutics have shown promise in preclinical settings. Some miRNA-targeted therapeutics have been extended to clinical development, such as the mimic of miRNA-34 for treating cancer. Here, we discuss insights into the role of miRNAs and other non-coding RNAs in tumorigenesis and resistance and summarize some recent successful systemic delivery approaches and recent developments in miRNAs as targets for anticancer drug development. Furthermore, we provide a comprehensive overview of mimics and inhibitors that are in clinical trials and finally a list of clinical trials based on miRNAs.

Competing Endogenous RNA Regulatory Networks of hsa_circ_0126672 in Pathophysiology of Coronary Heart Disease.

Coronary heart disease (CHD) is a global health concern, and its molecular origin is not fully elucidated. Dysregulation of ncRNAs has been linked to many metabolic and infectious diseases. This study aimed to explore the role of circRNAs in the pathogenesis of CHD and predicted a candidate circRNA that could be targeted for therapeutic approaches to the disease. circRNAs associated with CHD were identified and CHD gene expression profiles were obtained, and analyzed with GEO2R. In addition, differentially expressed miRNA target genes (miR-DEGs) were identified and subjected to functional enrichment analysis. Networks of circRNA/miRNA/mRNA and the miRNA/affected pathways were constructed. Furthermore, a miRNA/mRNA homology study was performed. We identified that hsa_circ_0126672 was strongly associated with the CHD pathology by competing for endogenous RNA (ceRNA) mechanisms. hsa_circ_0126672 characteristically sponges miR-145-5p, miR-186-5p, miR-548c-3p, miR-7-5p, miR-495-3p, miR-203a-3p, and miR-21. Up-regulation of has_circ_0126672 affected various CHD-related cellular functions, such as atherosclerosis, JAK/STAT, and Apelin signaling pathways. Our results also revealed a perfect and stable interaction for the hybrid of miR-145-5p with NOS1 and RPS6KB1. Finally, miR-145-5p had the highest degree of interaction with the validated small molecules. Henchashsa_circ_0126672 and target miRNAs, notably miR-145-5p, could be good candidates for the diagnosis and therapeutic approaches to CHD.

Identification of microRNA Signatures in Peripheral Blood of Young Women as Potential Biomarkers for Metal Allergy

MicroRNA (miRNA) is a short (19-24 nucleotide) endogenous non-protein RNA that exists in the body and controls the translation process from genes to proteins. It has become useful as a diagnostic tool and a potential treatment target in cancer research. To explore the function of miRNA in contact dermatitis, female participants with a positive metal allergy diagnosis (n = 3) were enrolled along with additional female participants with no medical history of metal allergy (n = 3). A patch test was performed on each participant. Peripheral blood was collected from all the participants before the patch test and at days 3 and 7 after starting the patch test. After total RNA was obtained from peripheral blood leukocytes and cDNA was generated, microarray analysis was performed to analyze the large-scale circulating miRNA profile. Real-time polymerase chain reaction (RT-PCR) was then used to clarify the overall target miRNA expression. Downregulation of hsa-let-7d-5p, hsa-miR-24-3p, hsa-miR-23b-3p, hsa-miR-26b-5p, and hsa-miR-150-5p was found on day 7. Certain miRNAs were confirmed using RT-PCR. These peripheral blood miRNAs could be diagnostic biomarkers for metal allergies.

Identification of miRNAs as Biomarkers of Cardiac Protection in Non-Genetically Modified Primary Human Cardiomyocytes Exposed to Halogenated Hypnotics in an In Vitro Model of Transfection and Ischemia/Reperfusion: A New Model in Translational Anesthesia.

Many clinical studies have identified some circulating micro-RNAs (miRNAs) as potential biomarkers with regard to the cardioprotective effects of halogenated agents administered perioperatively during myocardial conditioning procedures. However, there is a major methodological difficulty in identifying these potential miRNA targets in cardiac cells. We developed an in vitro protocol to analyze the differential expression of target miRNAs at the intracellular level in non-genetically modified primary human cardiomyocytes (HCMs) through their exposure to different hypnotic compounds (i.e., halogenated versus non-halogenated). For this purpose, we performed a validated in vitro model of "ischemia and reperfusion" with the transfection of specific miRNA mimics (MIMICs) designed to simulate naturally occurring mature miRNAs as a functional study. Afterwards, next-generation sequencing (NGS) was used to identify and quantify miRNAs and elucidate their function. The differences in miRNA expression between HCMs exposed to different hypnotic drugs, along with the prediction of functional miRNA targets, were assessed using a meticulous in-house bioinformatics pipeline in order to derive diagnostic biomarkers and possible therapeutic targets. In brief, this methodological procedure was designed to investigate whether the cardioprotective effects of halogenated agents are a phenomenon mediated by either the activation or the suppression of miRNAs targeted by halogenated anesthetics.

Stage- and Subfield-Associated Hippocampal miRNA Expression Patterns after Pilocarpine-Induced Status Epilepticus.

To investigate microRNA (miRNA) expression profiles before and after pilocarpine-induced status epilepticus (SE) in the cornu ammonis (CA) and dentated gyrus (DG) areas of the mouse hippocampus, and to predict the downstream proteins and related pathways based on bioinformatic analysis. An epileptic mouse model was established using a pilocarpine injection. Brain tissues from the CA and DG were collected separately for miRNA analysis. The miRNAs were extracted using a kit, and the expression profiles were generated using the SurePrint G3 Mouse miRNA microarray and validated. The intersecting genes of TargetScan and miRanda were selected to predict the target genes of each miRNA. For gene ontology (GO) studies, the parent-child-intersection (pci) method was used for enrichment analysis, and Benjamini-Hochberg was used for multiple test correction. The Kyoto Encyclopedia of Genes and Genomes (KEGG) was used to detect disease-related pathways among the large list of miRNA-targeted genes. All analyses mentioned above were performed at the time points of control, days 3, 14, and 60 post-SE. Control versus days 3, 14, and 60 post-SE: in the CA area, a total of 131 miRNAs were differentially expressed; 53, 49, and 26 miRNAs were upregulated and 54, 10, and 22 were downregulated, respectively. In the DG area, a total of 171 miRNAs were differentially expressed; furthermore, 36, 32, and 28 miRNAs were upregulated and 78, 58, and 44 were downregulated, respectively. Of these, 92 changed in both the CA and DG, 39 only in the CA, and 79 only in the DG area. The differentially expressed miRNAs target 11-1630 genes. Most of these proteins have multiple functions in epileptogenesis. There were 15 common pathways related to altered miRNAs: nine different pathways in the CA and seven in the DG area. Stage- and subfield-associated hippocampal miRNA expression patterns are closely related to epileptogenesis, although the detailed mechanisms need to be explored in the future.

A miRNA derived from the 17/92 cluster is a novel player in programming pulmonary hypertension in response to gestational hypoxia via NADPH oxidases and the mTOR pathway

Abstract Background Gestational hypoxia can lead to intrauterine growth restriction (IUGR) and programming of cardiovascular diseases in adulthood via different, although not completely understood, epigenetic mechanisms. We have previously shown that reactive oxygen species (ROS) derived from NADPH oxidases contribute to hypoxia induced pulmonary hypertension (PH). However, their role in disease programming by gestational hypoxia is not resolved. Purpose We aimed to elucidate the role of NADPH oxidases and redox sensitive miRNAs in the cardiovascular response of adult offspring to transient gestational hypoxia. Methods Wildtype (WT) and nmf333 mice, lacking a functional p22phox dependent NADPH oxidase, were exposed to gestational hypoxia from E10.5 to E11.5. Embryos and adult offspring were analyzed by functional, immunohistochemical and molecular methods including mRNA and miRNA profiling. miRNA mimics or inhibitors were applied in vivo and in vitro using embryoid bodies (EBs). Heart samples from patients with right heart disease associated with congenital heart disease (CHD) were used to validate target miRNA expression. Results Transient gestational hypoxia was sufficient to induce IUGR in embryos and pulmonary hypertension (PH) characterized by right ventricular hypertrophy, increased right ventricular pressure and pulmonary vascular remodeling in adult offspring from WT mice while p22phox-deficient nmf333 mice were protected. Cardiac mRNA and miRNA profiling followed by qPCR validation identified several single miRNAs derived from the 17/92 miRNA cluster as downregulated by gestational hypoxia and NADPH oxidases in embryos and adult offspring hearts. In vitro analyses using EBs validated a single miRNA from this cluster to be sufficient to induce cardiac differentiation in response to hypoxia and p22phox, and target analyses identified the mTOR pathway as dysregulated. Treatment with an inhibitor targeting this miRNA was sufficient to induce PH in adult mice, while treating offspring derived from gestational hypoxia pregnancies with an miRNA mimic prevented the development of PH in the adult and dysregulation of the mTOR pathway. Importantly, expression of this single miRNA from the miRNA 17/92 cluster was negatively correlated to right ventricular pressure in CHD patients. Conclusions In this study, we demonstrate that gestational hypoxia programs PH in adulthood by upregulating NADPH oxidases and downregulating miRNAs derived from the 17/92 cluster subsequently affecting the mTOR pathway. Since dysregulation of a single miRNA from this cluster induced not only PH but was also associated with right ventricular pressure in CHD patients, these findings might open novel options for diagnosis, prevention and treatment of right heart disease. Funding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): DZHK (German Centre for Cardiovascular Research)

Circulating microRNA expression profiles to identify a potential link between prostate cancer and obesity

Effective diagnostic methods are needed to apply appropriate treatment strategies in patients with aggressive prostate cancer. From this point of view, risk factors that cause prostate cancer or its aggressiveness should be considered. Obesity is a critical risk factor for triggering prostate cancer's metastatic properties. microRNAs are used as biomarkers in diagnosing cancer and obesity depending on their tissue-specific expression patterns. This study investigates the role of obesity in the metastatic profile of prostate cancer depending on the differential expression signatures of selected miRNAs in prostate cancer and obese patients. The roles of miR-100, miR-141 and miR-145 in prostate cancer and obesity are partially known. However, their potential to become circular biomarkers in the blood is not elucidated. There is no previous data on miR-4463 and miR-653 on prostate cancer and obesity association. In this study, the blood samples were taken and obtained serum from 69 patients of 6 subgroups that consisted of one healthy group and five unhealthy groups based on their different prostate cancer or obesity levels. Five selected miRNA expression analyses (miR-100, miR-141, miR-145, miR-4463, and miR-653) were performed through total RNA isolation, which was confirmed via synthetic cel-miR-39 miRNA. Quantitative Real-Time PCR analyzed the expression levels of selected miRNAs. Data analysis was performed via normalising target miRNA expression levels with cel-miR-39. In this study, we found that the relationship between prostate cancer and obesity was investigated at the molecular level. It was suggested that target miR-100 could be a promising biomarker for non-obese and aggressive prostate cancer patients. miR-145 is a more potential biomarker than miR-141 for non-aggressive and non-obese patients. miR-4463 can be used to predict more prostate cancer patients than obese patients. Lastly, miR-653 can be a biomarker for non-aggressive prostate cancer cells.