Dysregulation Of miRNAs Research Articles

Apolipoprotein E dysfunction in Alzheimer’s disease: a study on miRNA regulation, glial markers, and amyloid pathology

IntroductionApolipoprotein E (ApoE) plays a crucial role in lipid homeostasis, predominantly expressed in astrocytes and to a lesser extent in microglia within the central nervous system (CNS). While the APOE4 allele is the strongest genetic risk factor for late-onset Alzheimer’s disease (AD), its precise role in AD pathogenesis remains elusive. Apoe-knockout (Apoe-ko) mice, mice expressing human APOE4, and human APOE4 carriers exhibit similar deficits in lipid metabolism, cognitive and behavioral functions, and neurodegeneration. The retina, as part of the CNS, has been studied to investigate the underlying mechanisms of AD, including neuroinflammation, amyloid aggregation, and neurodegeneration. This study explores ApoE’s role in AD by analyzing brain and eye samples from Apoe-ko mice, focusing on identifying potential retinal biomarkers associated with ApoE dysfunction.MethodsWe compared female Apoe-ko mice on a regular diet to age-matched C57BL/6J controls at 3 and 9 months. Our investigations included microRNAs (miRNAs), their target messenger RNAs (mRNAs), and selected protein markers, including astroglial (Gfap), microglial/macrophage (Iba1 and Trem2) markers, and amyloid precursor protein (APP)/amyloid-β (Aβ) peptides implicated in AD pathogenesis. We also examined female Apoe-ko mice on a high-fat diet versus a regular diet at 9 months for differential miRNA and mRNA expressions.ResultsOur findings demonstrated that miRNA levels were generally lower in 3-month-old Apoe-ko mice but increased in 9-month-old mice across five distinct brain regions, as well as in eye tissue and tear fluid. A high-fat diet further enhanced miRNA dysregulation in brain and eye tissues, but not in tear fluid. Target mRNAs were generally higher in the neocortex-hippocampus and eye tissue of 3-month-old Apoe-ko mice but decreased with age, except for glial cell mRNAs like Gfap and Aif1. Protein analysis revealed elevated Gfap expression, and increased APP/Aβ peptide accumulation in the neocortex-hippocampus, including brain endothelial cells at the meninges, as well as in the retina of 9-month-old Apoe-ko mice. These findings highlight ApoE’s pivotal role in AD, demonstrating its impact on inflammatory and amyloidogenic/angiogenic miRNA expression, glial homeostasis, and APP/Aβ peptide clearance. The observed upregulation of proinflammatory miR-146a and anti-amyloidogenic/angiogenic miR-15a in 9-month-old Apoe-ko mice suggests their potential as tear-based biomarkers for ApoE dysfunction.

Integrated Analysis of Gene Expression and Immune Cell Infiltration Reveals Dysregulated Genes and miRNAs in Acute Kidney Injury.

Acute Kidney Injury (AKI) is a multifaceted condition characterised by rapid deterioration of renal function, often precipitated by diverse etiologies. A comprehensive understanding of the molecular underpinnings of AKI is pivotal for identifying potential diagnostic markers and therapeutic targets. This study utilised bioinformatics to elucidate gene expression and immune infiltration in AKI. Publicly available mRNA and miRNA datasets were harnessed to discern differentially expressed genes (DEGs) and miRNAs in AKI. The CIBERSORT algorithm was employed to quantify immune cell infiltration in AKI samples. Functional enrichment analyses were conducted to unravel the implicated biological processes. Furthermore, the expression of identified genes and miRNAs was validated by quantitative real-time PCR in an AKI model. Our study revealed significant dysregulation of three genes (Aspn, Clec2h, Tmigd1) and two miRNAs (mmu-miR-21a-3p, mmu-miR-223-3p) in AKI, each with p < 0.0001. These molecular markers are implicated in immune responses, tissue remodelling, and inflammation. We observed notable disturbances in specific immune cells, including activated and immature dendritic cells, M1 macrophages, and subsets of T cells (Treg, Th1, Th17). These alterations correlated significantly with AKI pathology, with dendritic cells and M1 macrophages showing p < 0.01, and T cell subsets demonstrating p < 0.05. These results highlight the intricate involvement of the immune system in AKI and indicate significant enrichment of pathways related to immune response, inflammation, and tissue remodelling, pointing to their pivotal roles in AKI pathophysiology. Our study underscored the significance of immune cell infiltration and dysregulated gene and miRNA expression in AKI. The identified genes (Clec2h, Aspn, and Tmigd1) and miRNAs (mmu-miR-21a-3p and mmu-miR-223-3p) offer potential diagnostic markers and therapeutic avenues for AKI. Subsequent investigations targeting these genes and miRNAs, along with the elucidated pathways, may augment the clinical management and outcomes for AKI patients.

MiR-198 targets TOPORS: implications for oral squamous cell carcinoma pathogenesis.

miRNAs play a critical role in the progression of various diseases, including oral squamous cell carcinoma (OSCC), which represents a major health concern and is one of the leading causes for new cancer cases worldwide. The miRNA dysregulation causes havoc and could be attributed to various factors, with epigenetic silencing of tumor suppressor genes being a major contributor to tumorigenesis. In this study, we have explored the tumor suppressive role of miR-198 in OSCC. The tumor suppressive effect of miR-198 is established using miRNA analysis in OSCC cell lines, patient samples and xenograft nude mice model. The relationship between the miR-198 and TOPORS is explored using bioinformatics analyses, qRT-PCR, dual-luciferase reporter assay, Western blotting and cancer hall marks assays. The hypermethylation of the MIR198 promoter is confirmed using bisulfite sequencing PCR. We have found miR-198 to be upregulated in OSCC cells treated with 5-Azacytidine, a known DNA methyltransferase inhibitor. Upregulation of miR-198 in 5-Azacytidine treated OSCC cells appears to be due to methylation of the MIR198 promoter. Using bioinformatics analysis and dual-luciferase reporter assay, we have identified TOPORS (TOP1 binding arginine/serine rich protein, E3 ubiquitin ligase) as a novel gene target for miR-198. miR-198-mediated repression of TOPORS decreases cell proliferation and anchorage-independent growth and enhances apoptosis of OSCC cells, which is dependent on the presence of the 3'UTR in TOPORS. An inverse correlation between the expression levels of miR-198 and TOPORS is observed in OSCC patient samples, highlighting the biological relevance of their interaction. Delivery of a synthetic miR-198 mimic to OSCC cells results in a significant decrease in xenograft size in nude mice, potentiating its use in therapeutics. These results suggest that miR-198 is epigenetically silenced in OSCC, which promotes tumor growth, in part, by upregulating the levels of TOPORS.

A new quantitative real-time PCR method to measure human miRNAs using the PROMER technology

MicroRNAs (miRNAs) are small, non-coding RNA molecules that play a crucial role in regulating gene expression. Dysregulation of miRNAs is associated with various human diseases, including cancer. Accurate quantification of miRNAs in bodily fluids or tissue biopsy samples is essential for their use as biomarkers in tumor diagnosis, yet current methods remain suboptimal. In this study, we introduce a novel quantitative real-time PCR platform based on PROMER technology, which relies on RNA:DNA perfect matching in the primer-template interaction, followed by RNAse H2 cleavage to generate a 3'-hydroxyl group for new DNA synthesis.We selected 16 miRNAs implicated in lung cancer and evaluated the specificity and performance of the platform using synthesized miRNA mimics. Each miRNA could be measured with high specificity and accuracy, even at copy numbers as low as 1-10, while non-template controls exhibited minimal amplification. Additionally, key miRNAs involved in tumor progression, such as miR-210, miR-331, and miR-505, were accurately quantified using minimal amounts of plasma.In summary, we present a new quantitative real-time PCR method for miRNA detection using PROMER technology. This platform successfully measured miRNA mimics and was further applied to quantify miRNA species in plasma samples from cancer patients. Our findings support the potential clinical application of this method for early cancer diagnosis using liquid biopsy samples.

Role of miRNAs in breast cancer development and progression: Current research.

Breast cancer, a complex and heterogeneous ailment impacting numerous women worldwide, persists as a prominent cause of cancer-related fatalities. MicroRNAs (miRNAs), small non-coding RNAs, have garnered significant attention for their involvement in breast cancer's progression. These molecules post-transcriptionally regulate gene expression, influencing crucial cellular processes including proliferation, differentiation, and apoptosis. This review provides an overview of the current research on the role of miRNAs in breast cancer. It discusses the role of miRNAs in breast cancer, including the different subtypes of breast cancer, their molecular characteristics, and the mechanisms by which miRNAs regulate gene expression in breast cancer cells. Additionally, the review highlights recent studies identifying specific miRNAs that are dysregulated in breast cancer and their potential use as diagnostic and prognostic biomarkers. Furthermore, the review explores the therapeutic potential of miRNAs in breast cancer treatment. Preclinical studies have shown the effectiveness of miRNA-based therapies, such as antagomir and miRNA mimic therapies, in inhibiting tumor growth and metastasis. Emerging areas, including the application of artificial intelligence (AI) to advance miRNA research and the "One Health" approach that integrates human and animal cancer insights, are also discussed. However, challenges remain before these therapies can be fully translated into clinical practice. In conclusion, this review emphasizes the significance of miRNAs in breast cancer research and their potential as innovative diagnostic and therapeutic tools. A deeper understanding of miRNA dysregulation in breast cancer is essential for their successful application in clinical settings. With continued research, miRNA-based approaches hold promise for improving patient outcomes in this devastating disease.

Exploring dysregulated miRNAs in ALS: implications for disease pathogenesis and early diagnosis.

Amyotrophic lateral sclerosis(ALS) is a progressive neurodegenerative disease marked by motor neuron degeneration, leading to muscle weakness and paralysis, with no effective treatments available. Early diagnosis could slow disease progression and optimize treatment. MicroRNAs(miRNAs) are being investigated as potential biomarkers due to their regulatory roles in cellular processes and stability in biofluids. However, variability across studies complicates their diagnostic utility in ALS. This study aims to identify significantly dysregulated miRNAs in ALS through meta-analysis to elucidate disease mechanisms and improve diagnostic strategies. We systematically searched PubMed, Google Scholar, and the Cochrane Library, following predefined inclusion and exclusion criteria. The primary effect measure was the standardized mean difference (SMD) with a 95% confidence interval, analyzed using a random-effects model. Additionally, we used network pharmacology to examine the targets of dysregulated miRNAs and their roles in ALS pathology. Analysing 34 studies, we found significant upregulation of hsa-miR-206, hsa-miR-133b, hsa-miR-23a, and hsa-miR-338-3p, and significant downregulation of hsa-miR-218, hsa-miR-21-5p, and hsa-let-7b-5p in ALS patients. These miRNAs are involved in ALS pathophysiology, including stress granule formation, nuclear pore complex, SMCR8 and Sig1R dysfunction, histone methyltransferase complex alterations, and MAPK signaling perturbation, highlighting their critical role in ALS progression. This study identifies several dysregulated miRNAs in ALS patients, offering insights into their role in the disease and potential as diagnostic biomarkers. These findings enhance our understanding of ALS mechanisms and may inform future diagnostic strategies. Validating these results and exploring miRNA-based interventions are crucial for improving ALS diagnosis and treatment outcomes.

The regulation of miRNAs using curcumin and other polyphenols during the prevention and treatment of Alzheimer's disease.

Alzheimer's disease (AD), a prevalent neurodegenerative disorder, predominantly affects individuals over the age of 65 and poses significant challenges in terms of effective management and treatment. The disease's pathogenesis involves complex molecular pathways including misfolded proteins accumulation, neuroinflammation, and synaptic dysfunction. Recent insights have highlighted the role of microRNAs (miRNAs) as critical regulators within these pathways, where they influence gene expression and contribute to the pathophysiological landscape of AD. Notably, emerging research has demonstrated that polyphenols, including curcumin, might modulate miRNA activity, thus offering a novel approach to mitigate AD symptoms and progression. This review explores the potential mechanisms through which polyphenols regulate miRNA expression and activity, specifically focusing on autophagy enhancement and inflammation reduction in the context of AD. We provide a detailed examination of key studies linking miRNA dysregulation to AD pathogenesis and discuss how polyphenols might correct these aberrations. The findings presented here underscore the therapeutic potential of polyphenols in AD treatment via miRNA modulation, pointing to new directions in disease management strategies and highlighting the need for targeted research into miRNA-based interventions.

Reduced Levels of miR-145-3p Drive Cell Cycle Progression in Advanced High-Grade Serous Ovarian Cancer.

High-grade serous ovarian cancer (HGSOC) is the most lethal form of gynecologic cancer, with limited treatment options and a poor prognosis. Epigenetic factors, such as microRNAs (miRNAs) and DNA methylation, play pivotal roles in cancer progression, yet their specific contributions to HGSOC remain insufficiently understood. In this study, we performed comprehensive high-throughput analyses to identify dysregulated miRNAs in HGSOC and investigate their epigenetic regulation. Analysis of tissue samples from advanced-stage HGSOC patients revealed 20 differentially expressed miRNAs, 11 of which were corroborated via RT-qPCR in patient samples and cancer cell lines. Among these, miR-145-3p was consistently downregulated post-neoadjuvant therapy and was able to distinguish tumoural from control tissues. Further investigation confirmed that DNA methylation controls MIR145 expression. Functional assays showed that overexpression of miR-145-3p significantly reduced cell migration and induced G0/G1 cell cycle arrest by modulating the cyclin D1-CDK4/6 pathway. These findings suggest that miR-145-3p downregulation enhances cell proliferation and motility in HGSOC, implicating its restoration as a potential therapeutic target focused on G1/S phase regulation in the treatment of HGSOC.

Single-Molecule Detection of Serum MicroRNAs for Medulloblastoma with Biphasic Sandwich Hybridization-Assisted Plasmonic Resonant Scattering Imaging.

MicroRNA (miRNA) dysregulation is closely related to the occurrence and progression of medulloblastoma (MB). However, the full potential of serum circulating miRNAs in MB diagnosis is restricted by their ultralow abundance in peripheral blood due to blood-brain barrier. Here, we report the direct preamplification-free detection of aberrant expression of oncogenic miRNAs in serum from MB patients by proposing a simple yet robust single-molecule assay that combines biphasic sandwich hybridization in nucleic acids and the dark-field single-particle plasmonic imaging (B2S2PI). In this strategy, signal DNA was prehybridized with target miRNA in homogeneous solution to form sDNA-RNA complexes. Then the captured DNA strands with rationally adjusted surface densities could efficiently capture the sDNA-RNA complexes to generate a well-separated DNA-RNA sandwich structure. The combination of homogeneous and heterogeneous reactions enabled interface-mediated hybridization reactions to maintain molecular stability with fewer bases, making it suitable for the direct amplification-free assays of short miRNA targets. Labeling the DNA-RNA hybrids with plasmatic gold nanotags allowed nondestructive recognition and imaging of individual miRNA targets under mild conditions with high signal-to-noise ratio. By digitally counting and analyzing the bright plasmonic resonant scattering spots, B2S2PI enabled both the measurement of a low femtomolar concentration of circulating miRNA-21 in 5 μL sample volume within a turnaround of 2 h and the discrimination of single base mismatches. Moreover, B2S2PI was universal for detecting miRNAs with different sequences and secondary structures. Further analysis of clinical serum samples revealed that B2S2PI was capable of accurately distinguishing MB patients from noncancer controls with an area under the curve (AUC) of 0.99, which was superior to that of qRT-PCR. B2S2PI holds promise as a novel alternative means for single-molecule miRNA assay and sheds light on the circulating nucleic acid-based liquid biopsy of intracranial malignant tumors.

Discovering the role of microRNAs and exosomal microRNAs in chest and pulmonary diseases: a spotlight on chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a progressive respiratory condition and ranks as the fourth leading cause of mortality worldwide. Despite extensive research efforts, a reliable diagnostic or prognostic tool for COPD remains elusive. The identification of novel biomarkers may facilitate improved therapeutic strategies for patients suffering from this debilitating disease. MicroRNAs (miRNAs), which are small non-coding RNA molecules, have emerged as promising candidates for the prediction and diagnosis of COPD. Studies have demonstrated that dysregulation of miRNAs influences critical cellular and molecular pathways, including Notch, Wnt, hypoxia-inducible factor-1α, transforming growth factor, Kras, and Smad, which may contribute to the pathogenesis of COPD. Extracellular vesicles, particularly exosomes, merit further investigation due to their capacity to transport various biomolecules such as mRNAs, miRNAs, and proteins between cells. This intercellular communication can significantly impact the progression and severity of COPD by modulating signaling pathways in recipient cells. A deeper exploration of circulating miRNAs and the content of extracellular vesicles may lead to the discovery of novel diagnostic and prognostic biomarkers, ultimately enhancing the management of COPD. The current review focus on the pathogenic role of miRNAs and their exosomal counterparts in chest and respiratory diseases, centering COPD.

Dysregulation of miRNAs in Soft Tissue Sarcomas.

MicroRNAs (miRNAs) are pivotal regulators of gene expression, influencing key cellular processes such as proliferation, differentiation, apoptosis, and metastasis. In the realm of sarcomas-a diverse group of malignant tumors affecting soft tissues and bone sarcomas-miRNAs have emerged as crucial players in tumorigenesis and tumor progression. This review delves into the intricate roles of miRNAs across various soft tissue sarcoma subtypes, including rhabdomyosarcoma, liposarcoma, leiomyosarcoma, synovial sarcoma, fibrosarcoma, angiosarcoma, undifferentiated pleomorphic sarcoma (UPS), and malignant peripheral nerve sheath tumor (MPNST). We explore how dysregulated miRNAs function as oncogenes or tumor suppressors, modulating critical pathways that define the aggressive nature of these cancers. Furthermore, we discuss the diagnostic and prognostic potential of specific miRNAs and highlight their promise as therapeutic targets. By understanding the miRNA-mediated regulatory networks, this review aims to provide a comprehensive overview of current research while pointing towards future directions for miRNA-based therapies. Our findings underscore the potential of miRNAs to transform the landscape of sarcoma treatment, offering hope for more precise, personalized, and effective therapeutic strategies.

MiR-193b-5p and miR-374b-5p Are Aberrantly Expressed in Endometriosis and Suppress Endometrial Cell Migration In Vitro.

(1) Background: Endometriosis is a highly prevalent gynecological disease affecting 10% of women of reproductive age worldwide. miRNAs may play a role in endometriosis, though their exact function remains unclear. This study aimed to identify differentially expressed miRNAs in endometriosis and study their functions in the disease. (2) Methods: Endometrial tissue was collected from women with endometriosis (n = 15) and non-endometriosis controls (n = 17). Dysregulated miRNAs were identified through small RNA-sequencing, and their biological significance was explored by target gene prediction and pathway analysis. Selected miRNAs were examined in paired ectopic endometriomas and eutopic endometrium (n = 10) using qRT-PCR. Their roles in cell migration and proliferation were further examined in vitro using functional assays. To identify potential target genes, we performed mRNA sequencing on transfected cells and the endometrioma cohort. (3) Results: We identified 14 dysregulated miRNAs in the eutopic endometrium of women with endometriosis compared to endometrial tissue from women without endometriosis. Pathway analysis indicated enrichment in cell migration and proliferation-associated pathways. Further ex vivo studies of miR-193b-5p and miR-374b-5p showed that both miRNAs were upregulated in endometrioma. Overexpression of these two miRNAs in vitro inhibited cell migration, and mRNA sequencing revealed several migration-related genes that are targeted by these miRNAs. (4) Conclusions: Our study identified two key endometrial miRNAs that may be involved in the pathogenesis of endometriosis by regulating cell migration.

A Thorough Navigation of miRNA's Blueprint in Crafting Cardiovascular Fate.

Cardiovascular diseases contribute significantly to global morbidity and mortality. MicroRNAs are crucial in the development and progression of these diseases by regulating gene expression in various cells and tissues. Their roles in conditions like atherosclerosis, heart failure, myocardial infarction, and arrhythmias have been widely researched. The present study provides an overview of existing evidence regarding miRNAs' role in cardiovascular disease pathogenesis. Furthermore, the study examines current state-of-the-art technologies used in the study of miRNAs in cardiovascular disease. As a final point, we examine how miRNAs may serve as disease biomarkers, therapeutic targets, and prognostic indicators. In cardiology, microRNAs, small noncoding RNA molecules, are crucial to the posttranscriptional regulation of genes. Their role in regulating cardiac cell differentiation and maturation is critical during the development of the heart. They maintain the cardiac function of an adult heart by contributing to its electrical and contractile activity. By binding to messenger RNA molecules, they inhibit protein translation or degrade mRNA. Several cardiovascular diseases are associated with dysregulation of miRNAs, including arrhythmias, hypertension, atherosclerosis, and heart failure. miRNAs can be used as biomarkers to diagnose and predict diseases as well as therapeutic targets. A variety of state-of-the-art technologies have aided researchers in discovering, profiling, and analyzing miRNAs, including microarray analysis, next-generation sequencing, and others. Developing new diagnostics and therapeutic approaches is becoming more feasible as researchers refine their understanding of miRNA function. Ultimately, this will reduce the burden of cardiovascular disease around the world.

Subcellular mass spectrometric detection unveils hyperglycemic memory in the diabetic heart.

Intensive glycemic control is insufficient to reduce the risk of heart failure in patients with diabetes mellitus. While the hyperglycemic memory in the diabetic cardiomyopathy has been well documented, its underlying mechanisms are not fully understood. The present study tried to investigate whether the dysregulated proteins/biological pathways, which persistently altered in diabetic hearts during normoglycemia, participate in the hyperglycemic memory. Hearts of streptozotocin-induced diabetic mice, with or without intensive glycemic control using slow-release insulin implants, were collected. Proteins from total heart samples and subcellular fractions were assessed by mass spectrometry, Western blotting, and KEGG pathway enrichment analysis. mRNA sequencing was used to determine whether the persistently altered proteins were regulated at the transcriptional or post-transcriptional level. Western blot validation of several proteins with high pathophysiological importance, including MYH7, HMGCS2, PDK4, and BDH1, indicated that mass spectrometry was able to qualitatively, but not quantitatively, reflect the fold changes of certain proteins in diabetes. Pathway analysis revealed that the peroxisome, PPAR pathway, and fatty acid metabolism could be efficiently rescued by glycemic control. However, dysregulation of oxidative phosphorylation and reactive oxygen species persisted even after normalization of hyperglycemia. Notably, mRNA sequencing revealed that dysregulated proteins in the oxidative phosphorylation pathway were not accompanied by coordinated changes in mRNA levels, indicating post-transcriptional regulation. Moreover, literature review and bioinformatics analysis suggested that hyperglycemia-induced persistent alterations of miRNAs targeted genes from the persistently dysregulated oxidative phosphorylation pathway, whereas, oxidative phosphorylation dysfunction-induced ROS regulated miRNA expression, which thereby might sustained the dysregulation of miRNAs. Glycemic control cannot rescue hyperglycemia-induced alterations of subcellular proteins in the diabetic heart, and persistently altered proteins are involved in multiple functional pathways, including oxidative phosphorylation. These findings might provide novel insights into hyperglycemic memory in diabetic cardiomyopathy.

Unlocking the Potential of Circulating miRNAs as Biomarkers in Glioblastoma

Using microRNAs (miRNAs) as potential circulating biomarkers in diagnosing and treating glioblastoma (GBM) has garnered a lot of scientific and clinical impetus in the past decade. As an aggressive primary brain tumor, GBM poses challenges in early detection and effective treatment with significant current diagnostic constraints and limited therapeutic strategies. MiRNA dysregulation is present in GBM. The intricate involvement of miRNAs in altering cell proliferation, invasion, and immune escape makes them prospective candidates for identifying and monitoring GBM diagnosis and response to treatment. These miRNAs could play a dual role, acting as both potential diagnostic markers and targets for therapy. By modulating the activity of various oncogenic and tumor-suppressive proteins, miRNAs create opportunities for precision medicine and targeted therapies in GBM. This review centers on the critical role and function of circulating miRNA biomarkers in GBM diagnosis and treatment. It highlights their significance in providing insights into disease progression, aiding in early diagnosis, and potential use as targets for novel therapeutic interventions. Ultimately, the study of miRNA would contribute to improving patient outcomes in the challenging landscape of GBM management.

Profiling of the serum MiRNAome in pediatric egyptian patients with wilms tumor.

Wilms tumor (WT) is a pediatric kidney cancer associated with poor outcomes in patients with unfavorable histological features such as anaplasia. Small non-coding RNAs, such as miRNAs, are known to be involved in WT pathogenesis. However, research on the clinical potential of blood-based miRNAs is limited. This study aimed to profile aberrantly expressed miRNAs in WT serum samples, evaluate their potential to differentiate standard-risk patients with favorable histology from those with anaplastic WTs, and assess their clinical value as minimally invasive biomarkers for WT detection. The study used next-generation sequencing (NGS) to analyze miRNA expressions in serum samples from 37 Egyptian children, including 10 healthy individuals, 14 with non-anaplastic WTs (favorable histology FH-WTs), and 13 with anaplastic WTs (unfavorable histology UnFH-WTs). Functional enrichment analysis was conducted to identify critical pathways and biological processes affected by dysregulated miRNAs, and a network was created for the most promising miRNA-target interactions linked to WT. The study identified a distinct miRNA expression signature of 45 miRNAs (3 upregulated and 42 downregulated) in WT serum samples compared to healthy controls, with 29 miRNAs exclusively dysregulated in FH-WTs and 6 miRNAs dysregulated solely in UnFH-WTs. These dysregulated miRNAs displayed significant enrichment in cancer-related pathways, such as PI3K/AKT, FOXO, and MAPK signaling. In relation to WT clinicopathological features, decreased levels of hsa-miR-2355-3p showed a significant positive correlation with clinical stage (r = 0.6597, p = 0.0006) and WT metastasis (r = 0.439, p = 0.021). The ROC curve analysis revealed that multiple dysregulated miRNAs in WT, specifically hsa-miR-7-5p, hsa-miR-146a-5p, hsa-miR-378a-3p, and hsa-miR-483-5p, exhibited high diagnostic potential for WT, with AUC values exceeding 0.86. Among WT histopathology types, the hsa-miR-1180-3p showed a 2.3 log2fold difference in expression between UnFH-WTs and FH-WTs, indicating its potential as a biomarker with 92% sensitivity and 85% specificity for identifying UnFH-WTs. Its target genes were enriched in pathways related to cell division and cell cycle regulation. In conclusion, hsa-miR-1180-3p could be a reliable blood-based biomarker for distinguishing WT histopathological types, and further research is needed to validate its clinical value.

Circulatory microRNAs and proinflammatory cytokines as predictors of lupus nephritis.

Lupus nephritis (LN) is one of the most prevalent severe organ manifestations of systemic lupus erythematosus (SLE), impacting 70% of SLE patients. MicroRNAs (miRNAs), are small non-coding RNA molecules which influence the expression of approximately one-third of human genes after the process of transcription. Dysregulation of miRNAs was documented in numerous disorders, including SLE and LN. Cytokines are the orchestrators of the immune response in autoimmune diseases. Our study aims to explore the variation in the levels of circulating miRNAs and proinflammatory cytokines as potential diagnostic biomarkers among LN and SLE patients without LN in comparison to controls. The study involved 20 LN patients, 20 SLE patients without LN, and 10 healthy controls. Serum levels of IL-12 and IL-21 in addition to miR-124, miR-146a, miR-199a, and miR-21 were assessed using the enzyme-linked immunosorbent assay (ELISA) for cytokines and quantitative real-time PCR for miRNAs. A significant downregulation in miR-124 (p<0.001) and a significant overexpression of miR-146a (p=0.005) were found in SLE patients without LN in comparison to controls. In comparison to SLE patients without LN and the control group, miR-199a, miR-21, and miR-146a were significantly upregulated in LN patients (p=<0.001) with high diagnostic values of these miRNAs in discriminating LN from SLE patients without LN according to Receiver operating curve (ROC) analysis. Logistic regression analysis revealed that only miR-199a is an independent predictor of LN (OR 1.69; 95% CI: 1.1-2.6). The expression of miR-124 was reduced in LN patients in comparison to the control but increased in LN patients in comparison to SLE patients without LN. However, there was no statistically significant difference in either scenario. In comparison to both SLE patients without LN and controls, LN patients exhibited the highest serum levels of IL-12 and IL-21, with no statistically significant difference. Regression analysis revealed that only miR-146a was associated with creatinine levels and SLEDAI score (p= 0.009 and 0.03, respectively), while miR-124 was associated with hemoglobin level (p=0.03). MiR-199a is an independent predictor for LN and might be used as a diagnostic biomarker for this disease. MiR-146a might play an important role in LN pathophysiology.

5178 TNFα-dependent altered miRNA expression regulates endometrial stromal cell proliferation

Abstract Disclosure: I. Chowdhury: None. S. Banerjee: None. W. Xu: None. A. Driss: None. C. Nezhat: None. N. Sidell: None. R.N. Taylor: None. W.E. Thompson: None. Abstract: Aberrant upregulation of cytokines, including tumor necrosis factor α (TNFα) and activation of nuclear factor κB (NF-κB) with disbalance of microRNAs (miRNAs) are often found in endometrium of endometriosis subjects, which causes directly or indirectly pelvic inflammation, pain, and infertility, Although, the pathophysiology of endometriosis is not clearly established yet, inflammation is an essential feature. MicroRNAs are emerging as potential regulatory factors in cellular processes such as cell growth, survival, migration, and invasion. In the current study, we examined the direct effects of exogenous recombinant TNFα dependent regulation of miRNAs involved in proliferation in primary cultures of normal endometrial stromal cells (NESC) and compared with the untreated cells derived from endometriosis-ESC (EESC). NanoString nCounter-based assays and quantitative real-time polymerase chain reaction (RT-PCR) were used to identify and validate differentially expressed miRNAs linked to cell proliferation in EESC compared to NESC and TNFα treated NESCs. The expression level of miRNAs linked to cell cycle was downregulated significantly in EESCs compared to NESCs. Interestingly, a dose and time-dependent exogenous TNFα treatment in NESCs showed a similar cell cycle regulator miRNAs expression pattern with EESCs. In further studies, protein analysis by Western Blots of selective cell cycle regulators such as PCNA, Ki67, CDK2, CDK4, CDK6, p21, and p27, and cell proliferation assay demonstrated an inverse correlation between cell cycle regulators and miRNA expression pattern both in EESC and TNFα treated NESCs. These findings suggest that TNFα dependent dysregulation of miRNAs in EESCs may promote proliferation of endometriosis. Further research is needed to investigate the detailed mechanisms by which TNFα regulates miRNA expression and determine these findings' potential therapeutic implications. Acknowledgments: This study was supported in part by National Institutes of Health Grants 1SC3 GM113751, 1SC1 GM130544, U01 HD66439, 1R01HD057235, U54 CA118948, HD41749, S21MD000101, and G12-RR03034. This investigation was conducted in a facility constructed with support from Research Facilities Improvement Grant #C06 RR18386 from NIH/NCRR. Presentation: 6/2/2024

Crosstalk between MIR-96 and IRS/PI3K/AKT/VEGF cascade in hRPE cells; A potential target for preventing diabetic retinopathy.

Regulation of visual system function demands precise gene regulation. Dysregulation of miRNAs, as key regulators of gene expression in retinal cells, contributes to different eye disorders such as diabetic retinopathy (DR), macular edema, and glaucoma. MIR-96, a member of the MIR-183 cluster family, is widely expressed in the retina, and its alteration is associated with neovascular eye diseases. MIR-96 regulates protein cascades in inflammatory and insulin signaling pathways, but further investigation is required to understand its potential effects on related genes. For this purpose, we identified a series of key target genes for MIR-96 based on gene and protein interaction networks and utilized text-mining resources. To examine the MIR-96 impact on candidate gene expression, we overexpressed MIR-96 via adeno-associated virus (AAV)-based plasmids in human retinal pigment epithelial (RPE) cells. Based on Real-Time PCR results, the relative expression of the selected genes responded differently to overexpressed MIR-96. While the expression levels of IRS2, FOXO1, and ERK2 (MAPK1) were significantly decreased, the SERPINF1 gene exhibited high expression simultaneously. pAAV-delivered MIR-96 had no adverse effect on the viability of human RPE cells. The data showed that changes in insulin receptor substrate-2 (IRS2) expression play a role in disrupted retinal insulin signaling and contribute to the development of diabetic complications. Considered collectively, our findings suggest that altered MIR-96 and its impact on IRS/PI3K/AKT/VEGF axis regulation contribute to DR progression. Therefore, further investigation of the IRS/PI3K/AKT/VEGF axis is recommended as a potential target for DR treatment.

MicroRNA expression signature in gastrointestinal stromal tumour & their molecular & histological features.

Background & objectives In gastrointestinal stromal tumour (GIST), not only genetic abnormalities are responsible for adverse clinical events, but epigenetic modifications also play a crucial role. MicroRNA (miRNA) dysregulation plays a significant role in carcinogenesis as miRNAs serve as natural silencer for their targets. Our study aimed to explore the miRNAs expression and its association with molecular and histopathological characteristics of GIST. Methods Fifty GIST samples, including 45 formalin fixed paraffin embedded (FFPE) and fresh tissues were included. Peripheral non-tumour tissues were used as controls. All the cases were confirmed using immunohistochemistry. RNA was extracted using miRNA-specific kit, and the expression was performed using RT-qPCR. The data were evaluated using AriaMx software version 1.5 (Agilent, US). MiRNAs expression was analyzed by using the relative quantification method (ΔΔCT). Results miR-221, miR-222, miR-494 and miR-34a showed significant down-regulation in tumours relative to non-tumour tissues. The expression levels of these miRNAs were significantly down-regulated in c-KIT (proto-oncogene encoding the tyrosine kinase transmembrane receptor)-positive tumours compared to c-KIT-negative. Further analysis revealed that reduced expression was associated with spindle subtypes and gastric localization. However, there was no significant correlation with other histological features. Additionally, miR-221/222, and miR-494 were down-regulated in most of the KIT exon 11 mutant subtypes, while miRNA-34a was associated with platelet derived growth factor receptor alpha (PDGFRA) mutations. Interpretation & conclusions The present study showed that the down-regulation of these miRNAs may help better molecular classification and characterization of GISTs. Our results offer new insight into the association between miRNAs and histological features, enabling a more thorough understanding of GISTs at the molecular level.