miRNAs In Peripheral Blood Research Articles

MiR-142-3p Regulates Airway Inflammation Through PTEN/AKT in Children and Mice with Asthma

ABSTRACT Background Asthma is the most common chronic pulmonary disease in children. MicroRNAs (miRNAs) play a regulatory role in the occurrence and development of asthma. We aimed to explore the differential expression of miRNAs in the peripheral blood of children with asthma and identify a miRNA that can alleviate asthma inflammation. Methods We used high-throughput sequencing to analyze differences in peripheral blood miRNA between children with acute asthma and healthy children, followed by target gene prediction and functional enrichment analysis. We inhibited miR-142-3p’s expression in asthmatic mice to observe asthma symptoms. Inflammatory changes in lung tissue were assessed using hematoxylin and eosin staining and ELISA. Subsequently, the target gene of miR-142-3p was identified through a dual-luciferase reporter assay, and PTEN and AKT expression levels in mice lung tissue were determined using qPCR and western blot. Results Fifty one differentially expressed miRNAs were identified. Inhibition of miR-142-3p expression in asthmatic mice reversed the downregulation of PTEN and activation of AKT in lung tissue, while also significantly alleviating symptoms and pulmonary inflammation in the asthmatic mice. Conclusion miRNAs were differentially expressed in the peripheral blood of children with asthma. miR-142-3p regulates airway inflammation via the PTEN/AKT pathway.

Peripheral Blood miRNA Expression in Patients with Essential Hypertension in the Han Chinese Population in Hefei, China.

Primary hypertension is a significant risk factor for cardiovascular diseases. However, the pathogenesis of primary hypertension involves multiple biological processes, including the nervous system, circulatory system, endocrine system, and more. Despite extensive research, there is no clear understanding of the regulatory mechanism underlying its pathogenesis. In recent years, miRNAs have gained attention as a regulatory factor capable of modulating the expression of related molecules through gene silencing. Therefore, exploring differentially expressed miRNAs in patients with essential hypertension (EH) may offer a novel approach for future diagnosis and treatment of EH. This study included a total of twenty Han Chinese population samples from Hefei, China. The samples consisted of 10 healthy individuals and 10 patients with EH. Statistical analysis was conducted to analyze the general information of the two-sample groups. High-throughput sequencing and base identification were performed to obtain the original sequencing sequences. These sequences were then annotated using various databases including Rfam, cDNA sequences, species repetitive sequences library, and miRBase database. The number of miRNA species contained in the samples was measured. Next, TPM values were calculated to determine the expression level of each miRNA. The bioinformatics of the differentiated miRNAs were analyzed using the OECloud tool, and RPM values were calculated. Furthermore, the reliability of the expression was analyzed by calculating the area under the Roc curve using the OECloud tools. Statistical analysis revealed no significant differences between the two samples in terms of age distribution, gender composition, smoking history, and alcohol consumption history (P > 0.05). However, there was a notable presence of family genetic history and high BMI in the EH population (P < 0.05). The sequencing results identified a total of 245 miRNAs, out of which 16 miRNAs exhibited differential expression. Among the highly expressed miRNAs were let-7d-5p, miR-101-3p, miR-122-5p, miR-122b-3p, miR-192-5p, and miR-6722-3p. On the other hand, the lowly expressed miRNAs included miR-103a-3p, miR-16-5p, miR-181a-2-3p, miR-200a-3p, miR-200b-3p, miR-200c-3p, miR-221-3p, miR-30d-5p, miR-342-5p, and miR-543. This study initially identified 16 miRNAs that are aberrantly expressed and function in various processes associated with the onset and progression of essential hypertension. These miRNAs have the potential to be targeted for future diagnosis and treatment of EH. However, further samples are required to provide additional support for this study.

Ischemic Stroke with Comorbid Cancer Has Specific miRNA-mRNA Networks in Blood That Vary by Ischemic Stroke Mechanism.

Approximately half of ischemic strokes (IS) in cancer patients are cryptogenic, with many presumed cardioembolic. We evaluated whether there were specific miRNA and mRNA transcriptome architectures in peripheral blood of IS patients with and without comorbid cancer, and between cardioembolic versus noncardioembolic IS etiologies in comorbid cancer. We studied patients with cancer and IS (CS; n = 42), stroke only (SO; n = 41), and cancer only (n = 28), and vascular risk factor-matched controls (n = 30). mRNA-Seq and miRNA-Seq data, analyzed with linear regression models, identified differentially expressed genes in CS versus SO and in cardioembolic versus noncardioembolic CS, and miRNA-mRNA regulatory pairs. Network-level analyses identified stroke etiology-specific responses in CS. A total of 2,085 mRNAs and 31 miRNAs were differentially expressed between CS and SO. In CS, 122 and 35 miRNA-mRNA regulatory pairs, and 5 and 3 coexpressed gene modules, were associated with cardioembolic and noncardioembolic CS, respectively. Complement, growth factor, and immune/inflammatory pathways showed differences between IS etiologies in CS. A 15-gene biomarker panel assembled from a derivation cohort (n = 50) correctly classified 81% of CS and 71% of SO participants in a validation cohort (n = 33). Another 15-gene panel correctly identified etiologies for 13 of 13 CS-cardioembolic and 11 of 11 CS-noncardioembolic participants upon cross-validation; 11 of 16 CS-cryptogenic participants were predicted cardioembolic. We discovered unique mRNA and miRNA transcriptome architecture in CS and SO, and in CS with different IS etiologies. Cardioembolic and noncardioembolic etiologies in CS showed unique coexpression networks and potential master regulators. These may help distinguish CS from SO and identify IS etiology in cryptogenic CS patients. ANN NEUROL 2024;96:565-581.

The Expression of microRNAs and Their Involvement in Recurrent Pregnancy Loss.

Background: Recurrent pregnancy loss refers to the spontaneous demise of two or more pregnancies before the 24 weeks of gestation. In almost half of the cases of recurrent miscarriages, the causes remain unknown since there is no reliable way of prognosis, early diagnosis, or treatment. Recent research has detected differential expression of certain miRNAs in reproductive system pathologies. Methods: The aim of the present review is to focus on microRNAs and their relationship with idiopathic recurrent miscarriages and to correlate miRNA expression with recurrent miscarriage and examine their potential role as biomarkers. Pubmed/Medline and Scopus databases were searched up to 31st January 2024 with terms related to recurrent pregnancy loss and miRNAs. Results: In total, 21 studies were selected for the review. A total of 75 different miRNAs were identified, showing a statistically significant differential expression. Around 40 miRNAs had increased expression, such as miR-520, miR-184 and miR-100-5p, 21 decreased, such as let-7c, and 14 had either increased or decreased expression depending on the study, such as miR-21. Conclusions: The dysregulation of miRNA expression is strongly associated with recurrent miscarriages. The circulating in the peripheral blood miRNAs, miR-100-5p and let-7c, might be utilized as biomarkers and establish a valuable non-invasive prognostic and diagnostic tool in the future.

Interface-constrained catalytic hairpin assembly permits highly sensitive SERS signaling of miRNA.

The rapid and precise monitoring of peripheral blood miRNA levels holds paramount importance for disease diagnosis and treatment monitoring. In this study, we propose an innovative research strategy that combines the catalytic hairpin assembly reaction with SERS signal congregation and enhancement. This combination can significantly enhance the stability of SERS detection, enabling stable and efficient detection of miRNA. Specifically, our paper-based SERS detection platform incorporates a streptavidin-modified substrate, biotin-labeled catalytic hairpin assembly reaction probes, 4-ATP, and primer-co-modified gold nanoparticles. In the presence of miRNA, the 4-ATP and primer-co-modified gold nanoparticles can specifically recognize the miRNA and interact with the biotin-labeled CHA probes to initiate an interfacial catalytic hairpin assembly reaction. This enzyme-free high-efficiency catalytic process can accumulate a large amount of biotin on the gold nanoparticles, which then bind to the streptavidin on the substrate with the assistance of the driving liquid, forming red gold nanoparticle stripes. These provide a multitude of hotspots for SERS, enabling enhanced signal detection. This innovative design achieves a low detection limit of 3.47 fM while maintaining excellent stability and repeatability. This conceptually innovative detection platform offers new technological possibilities and solutions for clinical miRNA detection.

The plasma miRNome and venous thromboembolism in high-grade glioma: miRNA Sequencing of a nested case-control cohort.

Patients with high-grade gliomas are at high risk of venous thromboembolism (VTE). MicroRNAs (miRNAs) are small non-coding RNAs with multiple roles in tumour biology, haemostasis and platelet function. Their association with VTE risk in high-grade glioma has not been comprehensively mapped so far. We thus conducted a nested case-control study within 152 patients with WHO grade IV glioma that had been part of a prospective cohort study on VTE risk factors. At inclusion a single blood draw was taken, and patients were thereafter followed for a maximum of 2 years. During that time, 24 patients (16%) developed VTE. Of the other 128 patients, we randomly selected 24 age- and sex-matched controls. After quality control, the final group size was 21 patients with VTE during follow-up and 23 without VTE. Small RNA next-generation sequencing of plasma was performed. We observed that hsa-miR-451a was globally the most abundant miRNA. Notably, 51% of all miRNAs showed a correlation with platelet count. The analysis of miRNAs differentially regulated in VTE patients-with and without platelet adjustment-identified potential VTE biomarker candidates such as has-miR-221-3p. Therewith, we here provide one of the largest and deepest peripheral blood miRNA datasets of high-grade glioma patients so far, in which we identified first VTE biomarker candidates that can serve as the starting point for future research.

Risk factors and related miRNA phenotypes of chronic pain after thoracoscopic surgery in lung adenocarcinoma patients.

Chronic postsurgical pain may have a substantial impact on patient's quality of life, and has highly heterogenous presentation amongst sufferers. We aimed to explore the risk factors relating to chronic pain and the related miRNA phenotypes in patients with lung adenocarcinoma after video-assisted thoracoscopic lobectomy to identify potential biomarkers. Our prospective study involved a total of 289 patients with early invasive adenocarcinoma undergoing thoracoscopic lobotomy and a follow-up period of 3 months after surgery. Blood was collected the day before surgery for miRNA detection and patient information including operation duration, duration of continuous drainage of the chest, leukocyte count before and after operation, and postoperative pain scores were recorded. Using clinical and biochemical information for each patient, the risk factors for chronic postsurgical pain and related miRNA phenotypes were screened. We found that chronic postsurgical pain was associated with higher body mass index; greater preoperative history of chronic pain; longer postoperative drainage tube retention duration; higher numerical rating scale scores one, two, and three days after surgery; and changes in miRNA expression, namely lower expression of miRNA 146a-3p and higher expression of miRNA 550a-3p and miRNA 3613-3p in peripheral blood (p < 0.05). Of these factors, patient body mass index, preoperative history of chronic pain, average numerical rating scale score after operation, and preoperative peripheral blood miRNA 550a-3P expression were independent risk factors for the development of chronic postsurgical pain. Identification of individual risk markers may aid the development and selection of appropriate preventive and control measures.

Correlation Between MicroRNA by Extracellular Vesicle Mediated and Antiviral Effects of Interferon Omega in Feline Peripheral Blood.

Feline interferon omega (IFN-ω) has been proven to have high antiviral activity; however, its in-depth antiviral effects remain unknown. Extracellular vesicles (EVs) have been demonstrated to participate in the regulation of the immune response pathway for the body through various active substances, especially through the microRNA (miRNA) carried by them. In this study, we isolated EVs from feline peripheral blood by differential centrifugation, and further found that the content of IFN-ω in EVs increased continuously within 24 h after IFN-ω treatment, and a large number of miRNAs were significantly downregulated in EVs within 12 h after IFN-ω treatment. These significantly differentially expressed miRNAs were important for regulating changes in antiviral cytokines. This study reveals for the first time the correlation between EVs-mediated miRNA in feline peripheral blood and IFN-ω on antiviral immune response, which may provide strong data support for the development of novel antiviral nanomedicine and the research of the antiviral effects of IFN-ω.

MicroRNA -21 expression as an auxiliary diagnostic biomarker of acute brucellosis.

This study aimed to measure the expression levels of peripheral blood miRNAs in brucellosis and their involvement in the different phases of the brucellosis. The expression levels of miRNAs including miR-210, miR-155, miR-150, miR-146a, miR-139-3p, miR-125a-5p, miR-29 and miR-21 were quantified in 57 brucellosis patients subgrouped into acute, under treatment & relapse phase and 30 healthy controls (HCs) using real-time polymerase chain reaction (RT-PCR). The receiver operating characteristic (ROC) analysis curve analysis was performed to find a biomarker for discrimination of different phases of brucellosis. The expression of miR-155, miR-146a, miR-125a-5p, miR-29, and miR-21 was found to be elevated in the acute brucellosis patients compared to HCs. miR-29 changed in under-treatment patients, while miR-139-3p and miR-125a-5p showed alterations in relapse cases. The ROC curve analysis depicted the potential involvement of miR-21 in the pathogenesis of acute brucellosis. The expression level of miR-21 is significantly augmented in acute brucellosis and has the potential to be a contributing diagnostic factor for acute infection.

MicroRNAs modulate immunological and inflammatory responses in Holstein cattle naturally infected with Mycobacterium avium subsp. paratuberculosis

MicroRNAs (miRNAs) regulate the post-transcriptional expression of genes by binding to their target mRNAs. In this study, whole miRNA sequencing was used to compare the expression of miRNAs in ileocecal valve (ICV) and peripheral blood (PB) samples of cows with focal or diffuse paratuberculosis (PTB)-associated lesions in gut tissues versus (vs) control cows without lesions. Among the eight miRNAs differentially expressed in the PB samples from cows with diffuse lesions vs controls, three (miR-19a, miR-144, miR32) were also down-regulated in cows with diffuse vs focal lesions. In the ICV samples, we identified a total of 4, 5, and 18 miRNAs differentially expressed in cows with focal lesions vs controls, diffuse lesions vs controls, and diffuse vs focal lesions, respectively. The differential expression of five microRNAs (miR-19a, miR-144, miR-2425-3p, miR-139, miR-101) was confirmed by RT-qPCR. Next, mRNA target prediction was performed for each differentially expressed miRNA. A functional analysis using the predicted gene targets revealed a significant enrichment of the RNA polymerase and MAPK signaling pathways in the comparison of cows with focal vs no lesions and with diffuse vs focal lesions, respectively. The identified miRNAs could be used for the development of novel diagnostic and therapeutical tools for PTB control.

A pH-Mediated Highly Selective System Enabling Simultaneous Analysis of Circulating RNAs Carried by Extracellular Vesicles and Lipoproteins.

Extracellular vesicles (EVs) and lipoproteins (LPPs) serve as important carriers of circulating miRNAs in peripheral blood, offering immense potential for disease diagnosis and therapeutic interventions. Due to their shared physicochemical attributes, EVs and LPPs are frequently coisolated, potentially leading to misunderstandings regarding their distinct functional roles in physiological and pathological processes. Here, we report a highly selective magnetic system based on the pH-mediated affinity displayed by cibacron blue (CB) toward EVs and LPPs, enabling successful separation and collection of these two nanoparticles without cross-contamination for subsequent circulating RNA analysis. First, we found that CB-modified magnetic beads (CBMBs) exhibit a strong affinity toward LPP particles while displaying little interaction with EVs in standard samples under physiological pH conditions. We further demonstrate that the affinity between CB molecules and bionanoparticles in plasma samples is highly pH-dependent. Specifically, CBMBs show affinities for both LPP and EV particles under neutral and acidic conditions. However, at basic pH levels, CB molecules selectively bind only to LPP particles. Consequently, the remaining EV particles present in plasma are subsequently isolated by using titanium dioxide-modified beads (TiMBs) through phospholipid affinity. The simultaneous analysis of the transcriptomic contents of EV and LPP reveals clear differences in their small RNA profiles, with the differentially expressed RNAs reflecting distinct biological processes. Significantly, in a proof-of-concept study, we successfully demonstrated a strong correlation between miRNAs carried by both EV and LPP particles with the occurrence of ocular neovascularization during the progression of diabetic retinopathy. The involved miRNAs may serve as potential biomarkers for DR diagnostics and severity classification. To sum up, this pH-mediated separation system is not only user-friendly but also highly compatible, rendering it a potent tool for probing the molecular compositions, biomarkers, and underlying biological mechanisms of EVs and LPPs.

Circulating Exosomal miRNA Signature As a Potential Risk Factor for DNA Virus Infection in Patients with Multiple Myeloma

Introduction DNA virus infection is a significant cause of morbidity and mortality in patients with multiple myeloma (MM). Monocyte dysfunction in MM patients plays a central role in infectious complications, but the precise molecular mechanism underlying the reduced monocyte resistance to viruses and risk factors for infection in newly diagnosed MM patients remains to be elucidated. Methods To determine the role of exosomal miRNAs in DNA infection of MM patients, we analyzed miRNAs in BM exosomes from MM patients (Exo-miRs) using high-throughput small RNA sequencing. To further evaluate the effect of these five upregulated Exo-miRs in antiviral response, we transfected the host cells with chemically synthesized miRNAs mimics to overexpress miRNAs. We utilized qPCR and WB to characterize the fate the cGAS-STING signaling pathway of THP-1 after overexpress miRNAs. The efficacy and safety of these Exo-miRs in innate antiviral immunity were evaluated using 5TGM1 C57BL/KaLwRij mice models. Additionally, miRNAs copy number as well as the viral susceptibility of newly diagnosed MM patients were determined in 82 samples from newly diagnosed biopsy proven MM patients. Results Screening of miRNAs enriched in exosomes delivered by primary MM cells identified five miRNAs as inhibitory factors that negatively regulate the cGAS-STING antiviral immune response. When engraftments of 5TGM1-Luc cells via tail vein for eight weeks in C57BL/KaLwRij mice resulted in MM progression with typical MM phenotypes, model mice were primed with Exo-miR antagomiRs oligonucleotides mixture through tail vein injection three times every 24 h followed by Murine cytomegalovirus (MCMV) challenge for 36 h. Consistent with the immunosuppression in MM patients, 5TGM1 C57BL/KaLwRij mice showed lower IFN-β in sera. Notably, intravenous administration of antagomiRs against miRNAs resulted in a marked increase of IFN-β levels in serum and IFN-I expression in organs (Figure 1). To investigate the potential of the miR-score as a prognostic determinant of vulnerability to DNA virus infection, we compared the miR-score in plasma exosomes collected from 82 newly diagnosed biopsy proven MM patients. We classified the viral susceptibility of newly diagnosed MM patients into two groups: I, DNA virus infection; II, no DNA virus infection. In human, a pronounced inverse correlation between IFN-β production by DNA virus in PBMCs and the microRNA (miR)-score of the exosomes within the plasma. The patients with DNA virus infection showed significantly enhanced circulatory expression of miRNAs than without DNA virus infection (p&amp;lt;0.001) patients. The diagnostic potential of miRNAs was assessed by plotting ROC curve, 5 miRNAs showed optimum sensitivity and specificity（AUC 0.854, P&amp;lt;0.0001, Sensitivity 87.5%, Specificity 74.1%, Cut-off values 5.234)（Figure 2）. Conclusions These findings identify a mechanism by which MM cells can suppress the host's innate immunity and potentially lead to immunocompromise in MM patients. Furthermore, the aberrant expression of specific miRNAs in MM patients emerges as a potential therapeutic target in MM cells. By quantifying miRNA scores based on abundant exosomal miRNAs in peripheral blood, the risk of DNA virus infection in MM patients can be assessed. Our findings shed light on a previously unknown molecular mechanism underlying immunosuppression in MM patients who have not received antineoplastic therapy, and also reveal a distinct miRNA signature that serves as a crucial determinant of the susceptibility of newly diagnosed MM patients to DNA virus infections.

Dynamic expression analysis of peripheral blood derived small extracellular vesicle miRNAs in sepsis progression.

Immune disorders caused by sepsis have recently drawn much attention. We sought to dynamically monitor the expression of small extracellular vesicle (sEV) miRNAs in peripheral blood during sepsis to explore these miRNAs as potential biomarkers for monitoring immune function in sepsis patients. This study included patients with sepsis. Blood samples were obtained from 10 patients on the first through 10th days, the 12th day and the 14th day since sepsis onset, resulting in 120 collected samples. Serum sEVs were extracted from peripheral venous blood, and levels of MIR497HG, miR-195, miR-497, and PD-L1 in serum sEVs were detected by qPCR, and clinical information was recorded. Our study revealed that the levels of MIR497HG, miR-195, miR-497 and PD-L1 in serum sEVs showed periodic changes; the time from peak to trough was approximately 4-5 days. The levels of sEV MIR497HG and miR-195 had a positive linear relationship with SOFA score (r values were -0.181 and -0.189; p values were 0.048 and 0.039, respectively). The recorded quantities of sEV MIR497HG, miR-195 and PD-L1 showed a substantial correlation with ARDS. ROC curve analysis revealed that sEV MIR497HG, miR-195 and miR-497 could predict the 28-day mortality of sepsis patients with an AUC of 0.66, 0.68 and 0.72, respectively. Levels of sEVs MIR497HG, miR-195, miR-497 and PD-L1 showed periodic changes with the immune status of sepsis, which provides a new exploration direction for immune function biomarkers and immunotherapy timing in sepsis patients.

Dynamic peripheral blood microRNA expression landscape during the peri-implantation stage in women with successful pregnancy achieved by single frozen-thawed blastocyst transfer.

What are the dynamic expression features of plasma microRNAs (miRNAs) during the peri-implantation period in women with successful pregnancy via single frozen-thawed blastocyst transfer? There is a significant change in the plasma miRNA expression profile before and after blastocyst transfer, during the window of implantation. The expression of miRNAs in peripheral blood has indicative functions during the peri-implantation period. Nevertheless, the dynamic expression profile of circulating miRNAs during the peri-implantation stage in women with a successful pregnancy has not been studied. Seventy-six women treated for infertility with a single frozen-thawed blastocyst transfer in a natural cycle were included in this study. Among them, 57 women had implantation success and a live birth, while 19 patients experienced implantation failure. Peripheral blood samples were collected at five different time points throughout the peri-implantation period, including D0 (ovulation day), D3, D5, D7, and D9 in this cycle of embryo transfer. The plasma miRNAs in women with blastocyst transfer were isolated, sequenced, and analyzed. Peripheral blood samples were collected in EDTA tubes and stored at -80°C until further use. miRNAs were isolated from blood, cDNA libraries were constructed, and the resulting sequences were mapped to the human genome. The plasma miRNAs were initially analyzed in a screening cohort (n = 34) with successful pregnancy. Trajectory analysis, including a global test and pairwise comparisons, was performed to detect dynamic differentially expressed (DE) miRNAs. Fuzzy c-means clustering was conducted for all dynamic DE miRNAs. The correlation between DE miRNAs and clinical characteristics of patients was investigated using a linear mixed model. Target genes of the miRNAs were predicted, and functional annotation analysis was performed. The expression of DE miRNAs was also identified in a validation set consisting of women with successful (n = 23) and unsuccessful (n = 19) pregnancies. Following small RNA sequencing, a total of 2656 miRNAs were determined as valid read values. After trajectory analysis, 26 DE miRNAs (false discovery rate < 0.05) were identified by the global test, while pairwise comparisons in addition identified 20 DE miRNAs. A total of seven distinct clusters representing different temporal patterns of miRNA expression were discovered. Nineteen DE miRNAs were further identified to be associated with at least one clinical trait. Endometrium thickness and progesterone level showed a correlation with multiple DE miRNAs (including two of the same miRNAs, hsa-miR-1-3p and hsa-miR-6741-3p). Moreover, the 19 DE miRNAs were predicted to have 403 gene targets, and there were 51 (12.7%) predicted genes likely involved in both decidualization and embryo implantation. Functional annotation for predicted targets of those clinically related DE miRNAs suggested the involvement of vascular endothelial growth factor and Wnt signaling pathways, as well as responses to hormones, immune responses, and cell adhesion-related signaling pathways during the peri-implantation stage. The raw miRNA sequence data reported in this article have been deposited in the Genome Sequence Archive (GSA-Human: HRA005227) and are publicly accessible at https://ngdc.cncb.ac.cn/gsa-human/browse/HRA005227. Although the RNA sequencing results revealed the global dynamic changes of miRNA expression, further experiments examining the clinical significance of the identified DE miRNAs in embryo implantation outcome and the relevant regulatory mechanisms involved are warranted. Understanding the dynamic landscape of the miRNA transcriptome could shed light on the physiological mechanisms involved from ovulation to the post-implantation stage, as well as identifying biomarkers that characterize stage-related biological process. The study was funded by the Major clinical research project of Tangdu Hospital (2021LCYJ004) and the Discipline Platform Improvement Plan of Tangdu Hospital (2020XKPT003). The funders had no influence on the study design, data collection, and analysis, decision to publish, or preparation of the article. There are no conflicts of interest to declare.

Intelligent dual-drive DNA nanosensor for ultrasensitive detection of prostate cancer-related circulating microRNA-200c

Rapid and accurate detection of the dynamic changes of prostate cancer-related miRNAs in peripheral blood is of great significance for prostate monitoring. Herein, we constructed an intelligent dual-drive DNA nanosensor (DD-DNS) for molecularly determination of microRNA-200c (miR-200c), a potential monitoring marker for diagnosis and treatment of prostate cancer. Unlike common detection technologies, the most distinctive feature of DD-DNS is the ingenious combination of enzyme-catalyzed chain amplification reaction and non-enzyme-catalyzed chain replacement reaction to realize the complementation of performance and improvement of detection capability. This improvement can significantly improve the detection sensitivity and environmental adaptability, which is more conducive to the detection of actual samples. In the presence of miR-200c, miR-200c activates the polymerase/nickase cycle reaction, resulting in exponential levels of cycle primers being produced. As a programmed process, cyclic primers catalyze the hairpin assembly reaction to achieve a surge in fluorescence signals. As a novel amplification strategy, the detection limit of this method can reach 5.5 fM, which fully meets the requirements of clinical quantitative analysis. Thus, miRNA detection technology based on DD-DNS shows superior detection performance and stability, which is conducive to carrying out extensive disease screening and treatment monitoring in a variety of complex situations.

Dynamic changes in radiological parameters, immune cells, selected miRNAs, and cytokine levels in peripheral blood of patients with severe COVID‑19.

The present study aimed to investigate the dynamic changes in peripheral blood leucocyte subpopulations, cytokine and miRNA levels, and changes in computed tomography (CT) scores in patients with severe coronavirus disease 2019 (COVID-19) (n=14) and age-matched non-COVID-19 volunteers (n=17), which were included as a reference control group. All data were collected on the day of patient admission (day 0) and on the 7th, 14th and 28th days of follow-up while CT of the lungs was performed on weeks 2, 8, 24 and 48. On day 0, lymphopenia and leucopenia were detected in most patients with COVID-19, as well as an increase in the percentage of banded neutrophils, B cells, and CD4+ Treg cells, and a decrease in the content of PD-1low T cells, classical, plasmacytoid, and regulatory dendritic cells. On day 7, the percentage of T and natural killer cells decreased with a concurrent increase in B cells, but returned to the initial level after treatment discharge. The content of different T and dendritic cell subsets among CD45+ cells increased during two weeks and remained elevated, suggesting the activation of an adaptive immune response. The increase of PD-1-positive subpopulations of T and non-T cells and regulatory CD4 T cells in patients with COVID-19 during the observation period suggests the development of an inflammation control mechanism. The levels of interferon γ-induced protein 10 (IP-10), tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 decreased on day 7, but increased again on days 14 and 28. C-reactive protein and granulocyte colony-stimulating factor (G-CSF) levels decreased gradually throughout the observation period. The relative expression levels of microRNA (miR)-21-5p, miR-221-3p, miR-27a-3p, miR-146a-5p, miR-133a-3p, and miR-126-3p were significantly higher at the beginning of hospitalization compared to non-COVID-19 volunteers. The plasma levels of all miRs, except for miR-126-3p, normalized within one week of treatment. At week 48, CT scores were most prominently correlated with the content of lymphocytes, senescent memory T cells, CD127+ T cells and CD57+ T cells, and increased concentrations of G-CSF, IP-10, and macrophage inflammatory protein-1α.

Abstract TP210: Small Extracellular Vesicle-derived Micrornas Differentiate Ischemic Stroke And Intracerebral Hemorrhage: A Pilot Study

Investigating the short and long-term signaling mechanisms via small extracellular vesicles (sEVs) in peripheral blood following human Ischemic Stroke (IS) and Intracerebral Hemorrhage (ICH) is of great interest. The sEV’s cargo can induce distant responses which can be used to derive potential novel therapeutic targets and biomarkers to differentiate the two brain pathologies. Thus, we sequenced the miRNA cargo derived from peripheral blood sEVs in IS (n=3), ICH (n=3), and Vascular Risk Factor-matched Control (VRFC; n=3) subjects. Subjects were divided into contrast groups (IS vs VRFC, ICH vs VRFC, ICH vs IS), and log2 transformed expression underwent Kruskal-Wallis tests to identify differentially expressed (DE; p&lt;0.05) miRNAs. We found 55 DE miRNAs in IS vs VRFC, 38 in ICH vs VRFC, and 45 in ICH vs IS ( Fig. 1A ). The combination of these miRNAs differentiated the three groups on Principal Components Analysis ( Fig. 1B ). IS associated miRNA included miR-30a, miR-30b, and miR-144. miR-30a is involved in hematopoietic stem cell self-renewal and can impair B cell differentiation. miR-30b may be an immune suppressor via Notch1. In male mice, miR-144 is protective against atherosclerosis. ICH associated miRNA included miR-195, miR-1-3p, and miR-20b-5p. miR-195 can inhibit the pro-inflammatory roles of macrophages. miR-1-3p is involved in cardiomyocyte development, can target TLR1 (Toll-Like Receptor 1), and may regulate autophagy. miR-20b-5p reduces Amyloid Precursor Protein (APP) mRNA and protein levels; vascular accumulation of APP is one cause of Lobar ICH. We show differential expression of sEV-derived miRNAs in peripheral blood of human IS and ICH patients that are involved in relevant signaling processes. sEV cargo profiles pose a largely underexplored intercellular signaling mechanism in IS and ICH with the potential to better characterize long distance signaling from injured brain to peripheral blood leukocytes in these brain disorders.

Establishment of a schizophrenia classifier based on peripheral blood signatures and investigation of pathogenic miRNA-mRNA regulation

To date, the diagnosis of schizophrenia (SCZ) mainly relies on patients' or guardians' self-reports and clinical observation, and the pathogenesis of SCZ remains elusive. In this study, we sought to develop a reliable classifier for diagnosing SCZ patients and provide clues to the etiology and pathogenesis of SCZ. Based on the high throughput sequencing analysis of peripheral blood miRNA expression profile and weighted gene co-expression network analysis (WGCNA) in our previous study, we selected eleven hub miRNAs for validation by qRT-PCR in 51 SCZ patients and 51 controls. miR-939-5p, miR-4732-3p let-7d-3p, and miR-142-3p were confirmed to be significantly up-regulated, and miR-30e-3p and miR-23a-3p were down-regulated in SCZ patients. miR-30e-3p with the most considerable fold change and statistically significance was selected for targeting validation. We first performed bioinformatics prediction followed by qRT-PCR and verified the up-regulation of potential target mRNAs (ABI1, NMT1, HMGB1) expression. Next, we found that the expression level of ABI1 was significantly up-regulated in SH-SY5Y cells transfected with miR-30e-3p mimics. Lastly, we conducted a luciferase assay in 293T cells confirming that miR-30e-3p could directly bind with the 3′untranslated region (3′-UTR) of ABI1, revealing that miR-30e-3p might play a role in the polymerization of neuronal actin and the reconstruction of the cytoskeleton via the downstream regulation of ABI1. In addition, we constructed a classifier by a series of bioinformatics algorithms and evaluated its diagnostic performance. It appears that the classifier consists of miRNAs and mRNAs possess a better discrimination performance than individual miRNA or mRNA in SCZ.

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.

Involvement of miR-495 overexpression in the pathogenesis of bronchopulmonary dysplasia in preterm infants via the targeting of NEDD4L-ENaC pathway.

Bronchopulmonary dysplasia (BPD) is a severe pulmonary complication causing morbidity and mortality in preterm infants. A key histopathological feature of BPD is late lung growth retardation, in which the process of alveolarization is hindered and the mechanism of which is unclear. Emerging evidence indicates that microRNAs (miRNAs) promote the development of BPD via the inhibition of their target genes. MiR-495 has been reported to be involved in various lung diseases. However, the physiological function of miR-495 in BPD has not yet been fully understood. Differentially expressed miRNAs in peripheral blood of patients with BPD were compared with those of normal controls. A dual-luciferase reporter assay was performed to identify the target genes of miR-495. A BPD neonatal rat model was established by injecting lipopolysaccharide (LPS) in the amniotic sac of pregnant rats. The morphology of the lungs was observed using hematoxylin and eosin (HE) staining. The expression of miR-495, neural precursor cell expressed developmentally down-regulated 4-like (NEDD4L), and epithelial Na+ channel (ENaC) was tested using quantitative reverse transcription-polymerase chain reaction (qRT-PCR), Western blot analysis, and immunofluorescent (IF) staining. The expression of miR-495 was significantly increased in the peripheral blood samples of premature infants with BPD and verified using qRT-PCR. NEDD4L was proven to be the target gene of miR-495. Additionally, miR-495 expression was also increased in the lungs of rat pups with BPD at postnatal day (P) 3 compared with the control group. qRT-PCR and Western blot results showed that NEDD4L expression was decreased while ENaC expression was increased at the transcriptional and translational levels. IF staining results showed that NEDD4L level was decreased while ENaC level was increased in the LPS-induced BPD rat model, which was consistent with abnormal changes in alveolar structure. The aberrant overexpression of miR-495 may contribute to the development of BPD by targeting NEDD4L-ENaC pathway, implying an imbalance in lung fluid clearance.