Differentially Expressed miRNAs Research Articles

Transcriptome and small RNA analysis unveils novel insights into the C4 gene regulation in sugarcane.

This study reveals miRNA indirect regulation of C4 genes in sugarcane through transcription factors, highlighting potential key regulators like SsHAM3a. C4 photosynthesis is crucial for the high productivity and biomass of sugarcane, however, the miRNA regulation of C4 genes in sugarcane remains elusive. We have identified 384 miRNAs along the leaf gradients, including 293 known miRNAs and 91 novel miRNAs. Among these, 86 unique miRNAs exhibited differential expression patterns, and we identified 3511 potential expressed targets of these differentially expressed miRNAs (DEmiRNAs). Analyses using Pearson correlation coefficient (PCC) and Gene Ontology (GO) enrichment revealed that targets of miRNAs with positive correlations are integral to chlorophyll-related photosynthetic processes. In contrast, negatively correlated pairs are primarily associated with metabolic functions. It is worth noting that no C4 genes were predicted as targets of DEmiRNAs. Our application of weighted gene co-expression network analysis (WGCNA) led to a gene regulatory network (GRN) suggesting miRNAs might indirectly regulate C4 genes via transcription factors (TFs). The GRAS TF SsHAM3a emerged as a potential regulator of C4 genes, targeted by miR171y and miR171am, and exhibiting a negative correlation with miRNA expression along the leaf gradient. This study sheds light on the complex involvement of miRNAs in regulating C4 genes, offering a foundation for future research into enhancing sugarcane's photosynthetic efficiency.

Plasma microRNA expression in adolescents and young adults with endometriosis: the importance of hormone use

IntroductionPrior studies have investigated the diagnostic potential of microRNA (miRNA) expression profiles for endometriosis. However, the vast majority of previous studies have only included adult women. Therefore, we sought to investigate differential expression of miRNAs among adolescents and young adults with endometriosis.MethodsThe Women's Health Study: from Adolescence to Adulthood (A2A) is an ongoing WERF EPHect compliant longitudinal cohort. Our analysis included 64 patients with surgically-confirmed endometriosis (96% rASRM stage I/II) and 118 females never diagnosed with endometriosis frequency matched on age (median = 21 years) and hormone use at blood draw. MicroRNA measurement was separated into discovery (10 cases and 10 controls) and internal replication (54 cases and 108 controls) phases. The levels of 754 plasma miRNAs were assayed in the discovery phase using PCR with rigorous internal control measures, with the relative expression of miRNA among cases vs. controls calculated using the 2−ΔΔCt method. miRNAs that were significant in univariate analyses stratified by hormone use were included in the internal replication phase. The internal replication phase was split 2:1 into a training and testing set and utilized FirePlex miRNA assay to assess 63 miRNAs in neural network analyses. The testing set of the validation phase was utilized to calculate the area under the curve (AUC) of the best fit models from the training set including hormone use as a covariate.ResultsIn the discovery phase, 49 miRNAs were differentially expressed between endometriosis cases and controls. The associations of the 49 miRNAs differed by hormone use at the time of blood draw. Neural network analysis in the testing set of the internal replication phase determined a final model comprising 5 miRNAs (miR-542-3p, let-7b-3p, miR-548i, miR-769-5p, miR-30c-1-3p), yielding AUC = 0.77 (95% CI: 0.67–0.87, p < 0.001). Sensitivity in the testing dataset improved (83.3% vs. 72.2%) while the specificity decreased (58.3% vs. 72.2%) compared to the training set.ConclusionThe results suggest that miR-542-3p, let-7b-3p, miR-548i, miR-769-5p, miR-30c-1-3p may be dysregulated among adolescent and young adults with endometriosis. Hormone use was a significant modifier of miRNA dysregulation and should be considered rigorously in miRNA diagnostic studies.

Differential gene expression and miRNA regulatory network in coronary slow flow

Coronary slow flow (CSF) is characterized by slow progression of coronary angiography without epicardial stenosis. The aim of this study was to explore the potential biomarkers and regulatory mechanism for CSF. Peripheral blood mononuclear cells from 3 cases of CSF and 3 healthy controls were collected for high-throughput sequencing of mRNA and miRNA, respectively. The differentially expressed mRNAs (DE-mRNAs) and miRNAs (DE-miRNAs) was identified. A total of 117 DE-mRNAs and 32 DE-miRNAs were obtained and they were mainly enriched in immune and inflammatory responses. Twenty-six DE-mRNAs were the predicted target genes for miRNAs by RAID, and then the regulatory network of 15 miRNAs were constructed. In addition, through the PPI network, we identified the three genes (FPR1, FPR2 and CXCR4) with larger degrees as hub genes. Among them, FPR1 was regulated by hsa-miR-342-3p, hsa-let-7c-5p and hsa-miR-197-3p and participated in the immune response. Finally, we validated the differential expression of hub genes and key miRNAs between 20 CSF and 20 control. Moreover, we found that miR-342-3p has a targeted regulatory relationship with FPR1, and their expression is negatively correlated. Then we established a hypoxia/reoxygenation (H/R) HUVEC model and detected FPR1, cell proliferation and apoptosis. Transfection with miR-342-3p mimics can significantly promote the proliferation of HUVEC under H/R conditions. FPR1 were associated with CSF as a biomarker and may be regulated by miR-342-3p potential biomarkers.

Integrative analysis of the lncRNA-miRNA-mRNA interactions in smooth muscle cell phenotypic transitions

Objectives: We previously found that the pluripotency factor OCT4 is reactivated in smooth muscle cells (SMC) in human and mouse atherosclerotic plaques and plays an atheroprotective role. Loss of OCT4 in SMC in vitro was associated with decreases in SMC migration. However, molecular mechanisms responsible for atheroprotective SMC-OCT4-dependent effects remain unknown.Methods: Since studies in embryonic stem cells demonstrated that OCT4 regulates long non-coding RNAs (lncRNAs) and microRNAs (miRNAs), making them candidates for OCT4 effect mediators, we applied an in vitro approach to investigate the interactions between OCT4-regulated lncRNAs, mRNAs, and miRNAs in SMC. We used OCT4 deficient mouse aortic SMC (MASMC) treated with the pro-atherogenic oxidized phospholipid POVPC, which, as we previously demonstrated, suppresses SMC contractile markers and induces SMC migration. Differential expression of lncRNAs, mRNAs, and miRNAs was obtained by lncRNA/mRNA expression array and small-RNA microarray. Long non-coding RNA to mRNA associations were predicted based on their genomic proximity and association with vascular diseases. Given a recently discovered crosstalk between miRNA and lncRNA, we also investigated the association of miRNAs with upregulated/downregulated lncRNA-mRNA pairs.Results: POVPC treatment in SMC resulted in upregulating genes related to the axon guidance and focal adhesion pathways. Knockdown of Oct4 resulted in differential regulation of pathways associated with phagocytosis. Importantly, these results were consistent with our data showing that OCT4 deficiency attenuated POVPC-induced SMC migration and led to increased phagocytosis. Next, we identified several up- or downregulated lncRNA associated with upregulation of the specific mRNA unique for the OCT4 deficient SMC, including upregulation of ENSMUST00000140952-Hoxb5/6 and ENSMUST00000155531-Zfp652 along with downregulation of ENSMUST00000173605-Parp9 and, ENSMUST00000137236-Zmym1. Finally, we found that many of the downregulated miRNAs were associated with cell migration, including miR-196a-1 and miR-10a, targets of upregulated ENSMUST00000140952, and miR-155 and miR-122, targets of upregulated ENSMUST00000155531. Oppositely, the upregulated miRNAs were anti-migratory and pro-phagocytic, such as miR-10a/b and miR-15a/b, targets of downregulated ENSMUST00000173605, and miR-146a/b and miR-15b targets of ENSMUST00000137236.Conclusion: Our integrative analyses of the lncRNA-miRNA-mRNA interactions in SMC indicated novel potential OCT4-dependent mechanisms that may play a role in SMC phenotypic transitions.

Identification of microRNAs and their target genes associated with chasmogamous and cleistogamous flower development in Viola prionantha.

Differentially expressed microRNAs were found associated with the development of chasmogamous and cleistogamous flowers in Viola prionantha, revealing potential roles of microRNAs in the developmental evolution of dimorphic flowers. In Viola prionantha, chasmogamous (CH) flowers are induced by short daylight, while cleistogamous (CL) flowers are triggered by long daylight. How environmental factors and microRNAs (miRNAs) affect dimorphic flower formation remains unknown. In this study, small RNA sequencing was performed on CH and CL floral buds at different developmental stages in V. prionantha, differentially expressed miRNAs (DEmiRNAs) were identified, and their target genes were predicted. In CL flowers, Viola prionantha miR393 (vpr-miR393a/b) and vpr-miRN3366 were highly expressed, while in CH flowers, vpr-miRN2005, vpr-miR172e-2, vpr-miR166m-3, vpr-miR396f-2, and vpr-miR482d-2 were highly expressed. In the auxin-activated signaling pathway, vpr-miR393a/b and vpr-miRN2005 could target Vpr-TIR1/AFB and Vpr-ARF2, respectively, and other DEmiRNAs could target genes involved in the regulation of transcription, e.g., Vpr-AP2-7. Moreover, Vpr-UFO and Vpr-YAB5, the main regulators in petal and stamen development, were co-expressed with Vpr-TIR1/AFB and Vpr-ARF2 and showed lower expression in CL flowers than in CH flowers. Some V. prionantha genes relating to the stress/defense responses were co-expressed with Vpr-TIR1/AFB, Vpr-ARF2, and Vpr-AP2-7 and highly expressed in CL flowers. Therefore, in V. prionantha, CH-CL flower development may be regulated by the identified DEmiRNAs and their target genes, thus providing the first insight into the formation of dimorphic flowers in Viola.

Biomarker profiling to determine clinical impact of microRNAs in cognitive disorders

Alzheimer’s disease (AD) and post-stroke cognitive impairment (PSCI) are the leading causes of progressive dementia related to neurodegenerative and cerebrovascular injuries in elderly populations. Despite decades of research, patients with these conditions still lack minimally invasive, low-cost, and effective diagnostic and treatment methods. MicroRNAs (miRNAs) play a vital role in AD and PSCI pathology. As they are easily obtained from patients, miRNAs are promising candidates for the diagnosis and treatment of these two disorders. In this study, we performed complete sequencing analysis of miRNAs from 24 participants, split evenly into the PSCI, post-stroke non-cognitive impairment (PSNCI), AD, and normal control (NC) groups. To screen for differentially expressed miRNAs (DE-miRNAs) in patients, we predicted their target genes using bioinformatics analysis. Our analyses identified miRNAs that can distinguish between the investigated disorders; several of them were novel and never previously reported. Their target genes play key roles in multiple signaling pathways that have potential to be modified as a clinical treatment. In conclusion, our study demonstrates the potential of miRNAs and their key target genes in disease management. Further in-depth investigations with larger sample sizes will contribute to the development of precise treatments for AD and PSCI.

MicroRNA Regulatory Pattern in Diabetic Mouse Cortex at Different Stages Following Ischemic Stroke.

After ischemic stroke, microRNAs (miRNAs) participate in various processes, including immune responses, inflammation, and angiogenesis. Diabetes is a key factor increasing the risk of ischemic stroke; however, the regulatory pattern of miRNAs at different stages of diabetic stroke remains unclear. This study comprehensively analyzed the miRNA expression profiles in diabetic mice at 1, 3, and 7 days post-reperfusion following the middle cerebral artery occlusion (MCAO). We identified differentially expressed (DE) miRNAs in diabetic stroke and found significant dysregulation of some novel miRNAs (novel_mir310, novel_mir89, and novel_mir396) post-stroke. These DEmiRNAs were involved in apoptosis and the formation of tight junctions. Finally, we identified three groups of time-dependent DE miRNAs (miR-6240, miR-135b-3p, and miR-672-5p). These have the potential to serve as biomarkers of diabetic stroke. These findings provide a new perspective for future research, emphasizing the dynamic changes in miRNA expression after diabetic stroke and offering potential candidates as biomarkers for future clinical applications.

MicroRNA Profiling Revealed the Mechanism of Enhanced Cold Resistance by Grafting in Melon (Cucumis melo L.).

Grafting is widely used to improve the resistance to abiotic stresses in cucurbit plants, but the effect and molecular mechanism of grafting on cold stress are still unknown in melon. In this study, phenotypic characteristics, physiological indexes, small-RNA sequencing and expression analyses were performed on grafted plants with pumpkin rootstock (PG) and self-grafted plants (SG) to explore the mechanism of changed cold tolerance by grafting in melon. Compared with SG plants, the cold tolerance was obviously enhanced, the malondialdehyde (MDA) content was significantly decreased and the activities of antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT; peroxidase, POD) were significantly increased in PG plants. Depend on differentially expressed miRNA (DEM) identification and expression pattern analyses, cme-miR156b, cme-miR156f and chr07_30026 were thought to play a key role in enhancing low-temperature resistance resulting from grafting. Subsequently, 24, 37 and 17 target genes of cme-miR156b, cme-miR156f and chr07_30026 were respectively predicted, and 21 target genes were co-regulated by cme-miR156b and cme-miR156f. Among these 57 unique target genes, the putative promoter of 13 target genes contained the low-temperature responsive (LTR) cis-acting element. The results of qRT-PCR indicated that six target genes (MELO3C002370, MELO3C009217, MELO3C018972, MELO3C016713, MELO3C012858 and MELO3C000732) displayed the opposite expression pattern to their corresponding miRNAs. Furthermore, MELO3C002370, MELO3C016713 and MELO3C012858 were significantly downregulated in cold-resistant cultivars and upregulated in cold-sensitive varieties after cold stimulus, and they acted as the key negative regulators of low-temperature response in melon. This study revealed three key miRNAs and three putative target genes involved in the cold tolerance of melon and provided a molecular basis underlying how grafting improved the low-temperature resistance of melon plants.

M1-type microglia-derived exosomes contribute to blood–brain barrier damage

BackgroundAs a key substance for intercellular communication, exosomes could be a potential strategy for stroke treatment. Activated microglia disrupt the integrity of blood–brain barrier (BBB) to facilitate the stroke process. Hence, this study was designed to investigate the effect of microglia-derived exosomes on BBB cell model injury and to explore the underlying molecular mechanisms. MethodsM1 polarization of BV2 cells was induced with LPS and their derived exosomes were isolated. Astrocytes were cultured in primary culture and constructed with End3 cells as a BBB cell model. After co-culture with exosomes, the BBB cell model was examined for changes in TEER, permeability, and expression of BBB-related proteins (Claudin-1, Occludin, ZO-1 and JAM). Resting and M1-type BV2 cell-derived exosomes perform small RNA sequences and differentially expressed miRNAs (DE-miRNAs) are identified by bioinformatics. ResultsM1-type BV2 cell-derived exosomes decreased End3 cell viability, and increased their apoptotic ratio. Moreover, M1 type BV2 cell-derived exosomes dramatically enhanced the permeability of BBB cell model, and diminished the TEER and BBB-related protein (Claudin-1, Occludin, ZO-1) expression. Notably, resting BV2 cell-derived exosomes had no effect on the integrity of BBB cell model. Sequencing results indicated that 71 DE-miRNAs were present in M1 BV2 cell-derived exosomes, and their targets mediated neurological development and signaling pathways such as MAPK and cAMP. RT-qPCR confirmed the differential expression of mmu-miR-125a-5p, mmu-miR-122b-3p, mmu-miR-139-3p, mmu-miR-330-3p, mmu-miR-3057-5p and mmu-miR-342-3p consistent with the small RNA sequence. Furthermore, Creb1, Jun, Mtor, Frk, Pabpc1 and Sdc1 are the most well-connected proteins in the PPI network. ConclusionM1-type microglia-derived exosomes contribute to the injury of BBB cell model, which has the involvement of miRNAs. Our findings provide new perspectives and potential mechanisms for future M1 microglia-derived exosomes as therapeutic targets in stroke.

Whole transcriptome sequencing analyses of islets reveal ncRNA regulatory networks underlying impaired insulin secretion and increased β-cell mass in high fat diet-induced diabetes mellitus.

Our study aims to identify novel non-coding RNA-mRNA regulatory networks associated with β-cell dysfunction and compensatory responses in obesity-related diabetes. Glucose metabolism, islet architecture and secretion, and insulin sensitivity were characterized in C57BL/6J mice fed on a 60% high-fat diet (HFD) or control for 24 weeks. Islets were isolated for whole transcriptome sequencing to identify differentially expressed (DE) mRNAs, miRNAs, IncRNAs, and circRNAs. Regulatory networks involving miRNA-mRNA, lncRNA-mRNA, and lncRNA-miRNA-mRNA were constructed and functions were assessed through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Despite compensatory hyperinsulinemia and a significant increase in β-cell mass with a slow rate of proliferation, HFD mice exhibited impaired glucose tolerance. In isolated islets, insulin secretion in response to glucose and palmitic acid deteriorated after 24 weeks of HFD. Whole transcriptomic sequencing identified a total of 1324 DE mRNAs, 14 DE miRNAs, 179 DE lncRNAs, and 680 DE circRNAs. Our transcriptomic dataset unveiled several core regulatory axes involved in the impaired insulin secretion in HFD mice, such as miR-6948-5p/Cacna1c, miR-6964-3p/Cacna1b, miR-3572-5p/Hk2, miR-3572-5p/Cckar and miR-677-5p/Camk2d. Additionally, proliferative and apoptotic targets, including miR-216a-3p/FKBP5, miR-670-3p/Foxo3, miR-677-5p/RIPK1, miR-802-3p/Smad2 and ENSMUST00000176781/Caspase9 possibly contribute to the increased β-cell mass in HFD islets. Furthermore, competing endogenous RNAs (ceRNA) regulatory network involving 7 DE miRNAs, 15 DE lncRNAs and 38 DE mRNAs might also participate in the development of HFD-induced diabetes. The comprehensive whole transcriptomic sequencing revealed novel non-coding RNA-mRNA regulatory networks associated with impaired insulin secretion and increased β-cell mass in obesity-related diabetes.

Oxidative metabolism induces differential miRNA expression in mitochondrial disease LCLS

Oxidative metabolism induces differential miRNA expression in mitochondrial disease LCLS

MicroRNA expression as a prognostic biomarker of tongue squamous cell carcinoma (TSCC): a systematic review and meta-analysis

BackgroundRecent studies have indicated that microRNA (miRNA) expression in tumour tissues has prognostic significance in Tongue squamous cell carcinoma (TSCC) patients. This study explored the possible prognostic value of miRNAs for TSCC based on published research.MethodsA comprehensive literature search of multiple databases was conducted according to predefined eligibility criteria. Data were extracted from the included studies by two researchers, and HR results were determined based on Kaplan‒Meier curves according to the Tierney method. The Newcastle‒Ottawa Scale (NOS) and GRADE (Grading of Recommendations Assessment, Development, and Evaluation) pro-GDT were applied to assess the quality of all studies. Publication bias was estimated by funnel plot, Egger’s rank correlation test and sensitivity analysis.ResultsEleven studies (891patients) were included, of which 6 reported up-regulated miRNAs and 7 mentioned down-regulated miRNAs. The pooled hazard ratio (HR) from the prognostic indicator overall survival (OS) was 1.34 (1.25–1.44), p < 0.00001, indicating a significant difference in miRNA expression between TSCC patients with better or worse prognosis.ConclusionMiRNAs may have high prognostic value and could be used as prognostic biomarkers of TSCC.

A Comprehensive Interaction Network Constructed Using miRNAs and mRNAs Provides New Insights into Potato Tuberization under High Temperatures.

High temperatures delay tuberization and decrease potato (Solanum tuberosum L.) yields. However, the molecular mechanisms and regulatory networks underlying tuberization under high temperatures remain largely unknown. Here, we performed the mRNA and miRNA sequencing of leaves and stems to identify genes and regulatory networks involved in tuberization under high temperatures. A total of 2804 and 5001 differentially expressed genes (DEGs) under high-temperature stress were identified in leaves and stems, respectively. These genes were significantly enriched in gene ontology terms regarding meristem development, the sucrose biosynthetic process, and response to heat. Meanwhile, 101 and 75 differentially expressed miRNAs (DEmiRNAs) were identified in leaves and stems, respectively. We constructed an interaction network between DEmiRNAs and DEGs, identifying 118 and 150 DEmiRNA-DEG pairs in leaves and stems, respectively. We found three miRNA-mRNA candidate modules involved in tuberization under high temperatures, including stu-miR8030-5p/StCPY714, stu-miR7981f-p5/StAGL8a, and stu-miR10532A/StAGL8b. Our study constructed an interaction network between miRNAs and target genes and proposes candidate miRNA-gene modules that regulate tuber formation under high temperatures. Our study provides new insights for revealing the regulatory mechanism of the high-temperature inhibition of tuberization and also provides gene resources for improving the heat tolerance in potatoes.

Study on miRNA expression and regulatory network of Amygdalus mongolica oil and amygdalin in the treatment of pulmonary fibrosis

IntroductionAmygdalus mongolica (Maxim.) Ricker is a traditional Chinese medicine known for its ability to relieve cough and expectorate phlegm and is commonly used to treat dry throat, dry cough, and bronchitis. A. mongolica oil and amygdalin were the active components isolated from the seed kernel seeds of A. mongolica. The main objective of this study was to investigate the effects of A. mongolica oil and amygdalin on bleomycin-induced pulmonary fibrosis (PF) in rats by integrating pharmacodynamic, transcriptomics, and bioinformatics, to determine the changes of miRNAs in both against PF, and to explore the underlying mechanisms. MethodsA rat model of PF was induced by administering bleomycin through the trachea. The rats were randomized into several groups: a control group (CON), an untreated model group (MOD), an active control group treated with pirfenidone (PFD), and groups receiving different doses of A. mongolica oil (5 g/kg [OIL-L], 10 g/kg [OIL-M], and 15 g/kg [OIL-H]), or amygdalin (20 mg/kg [AMY-L], 40 mg/kg [AMY-M], and 80 mg/kg [AMY-H]). After 28 days, samples were collected for subsequent pharmacodynamic and transcriptomic analyses. ResultsThe results demonstrated that A. mongolica oil and amygdalin effectively reduced the abnormally elevated levels of IL-1β, IL-6, HA, LN and COL-IV in PF rats and alleviated bleomycin-induced pulmonary fibrotic changes. Furthermore, compared to the MOD group, the expression levels of 27 miRNAs were up-regulated and 23 miRNAs were down-regulated in the OIL 10 g/kg group. Among the up-regulated miRNAs, two novel differentially expressed miRNAs (DE-miRNAs) were identified. In the AMY 20 mg/kg group, 12 miRNAs were up-regulated and 10 were down-regulated when compared to the MOD group. These DE-miRNAs were found to be enriched in pathways such as the PI3K-Akt signalling pathway, MAPK signalling pathway, and calcium signalling pathway. The microRNAs miR-301b-3p, miR-667-5p, miR-363-3p, and miR-540-3p may be associated with both the anti-PF mechanism of A. mongolica oil and the anti-PF mechanism of amygdalin. DiscussionNumerous studies have demonstrated the significance of various pathways, including PI3K-Akt, MAPK, and calcium signalling pathways, in the mechanism of PF and the development of anti-PF medications. Meanwhile, multiple miRNAs, including miR-301b-3p, miR-667-5p, miR-363-3p, and miR-540-3p, have been found to be associated with lung disease and fibrosis. ConclusionsThe present experiments confirmed the effectiveness of A. mongolica oil and amygdalin in reducing inflammation and fibrosis in bleomycin-induced PF rats, with particularly favorable outcomes observed in the A. mongolica oil 10 g/kg versus amygdalin 20 mg/kg. Meanwhile, miRNAs miR-301b-3p, miR-667-5p, miR-363-3p and miR-540-3p were found to be potentially associated with both A. mongolica oil and amygdalin anti-PF mechanisms. Based on our findings, we suggest that these miRNAs might play key roles in the anti-PF mechanism of A. mongolica. The individual miRNAs require further investigation.

Withaferin A alters the expression of microRNAs 146a-5p and 34a-5p and associated hub genes in MDA-MB-231 cells.

Triple-negative breast cancer (TNBC) is a highly metastatic subtype of breast cancer. Due to the absence of obvious therapeutic targets, microRNAs (miRNAs) provide possible hope to treat TNBC. Withaferin A (WA), a steroidal lactone, possesses potential anticancer activity with lesser side effects. The present study identifies hub genes (CDKN3, TRAF6, CCND1, JAK1, MET, AXIN2, JAG1, VEGFA, BRCA1, E2F3, WNT1, CDK6, KRAS, MYB, MYCN, TGFβR2, NOTCH1, SIRT1, MYCN, NOTCH2, WNT3A) from the list of predicted targets of the differentially expressed miRNAs (DEMs) in WA-treated MDA-MB-231 cells using in silico protein-protein interaction network analysis. CCND1, CDK6, and TRAF6 hub genes were predicted as targets of miR-34a-5p and miR-146a-5p, respectively. The study found the lower expression of miR-34a-5p and miR-146a-5p in MDA-MB-231 cells, and further, it was observed that WA treatment effectively restored the lost expression of miR-34a-5p and miR-146a-5p in MDA-MB-231 cells. An anti-correlation expression pattern was found among the miR-34a-5p and miR-146a-5p and the respective target hub genes in WA-treated TNBC cells. In conclusion, WA might exert anti-cancer effect in TNBC cells by inducing miR-34a-5p and miR-146a-5p expressions and decreasing CCND1, CDK6, and TARF6 target hub genes in TNBC cells.

Unveiling the Role of Novel miRNA PC-5P-12969 in Alleviating Alzheimer's Disease.

The intricate and complex molecular mechanisms that underlie the progression of Alzheimer's disease (AD) have prompted a concerted and vigorous research endeavor aimed at uncovering potential avenues for therapeutic intervention. This study aims to elucidate the role of miRNA PC-5P-12969 in the pathogenesis of AD. We assessed the differential expression of miRNA PC-5P-12969 in postmortem AD brains, AD animal and cell models using real-time reverse-transcriptase RT-PCR, we also checked the gene and protein expression of GSK3α and APP. Our investigation revealed a notable upregulation of miRNA PC-5P-12969 in postmortem brains of AD patients, in transgenic mouse models of AD, and in mutant APP overexpressing-HT22 cells. Additionally, our findings indicate that overexpression of miRNA PC-5P-12969 exerts a protective effect on cell survival, while concurrently mitigating apoptotic cell death. Further-more, we established a robust and specific interaction between miRNA PC-5P-12969 and GSK3α. Our luciferase reporter assays provided confirmation of the binding between miRNA PC-5P-12969 and the 3'-UTR of the GSK3α gene. Manipulation of miRNA PC-5P-12969 levels in cellular models of AD yielded noteworthy alterations in the gene and protein expression levels of both GSK3α and APP. Remarkably, the manipulation of miRNA PC-5P-12969 levels yielded significant enhancements in mitochondrial respiration and ATP production, concurrently with a reduction in mitochondrial fragmentation, thus unveiling a potential regulatory role of miRNA PC-5P-12969 in these vital cellular processes. In summary, this study sheds light on the crucial role of miRNA PC-5P-12969 and its direct interaction with GSK3α in the context of AD.

Let-7 family regulates HaCaT cell proliferation and apoptosis via the ΔNp63/PI3K/AKT pathway.

We evaluated the expression profiles of differentially expressed miRNAs (DEmiRNAs) involved in human fetal skin development via high-throughput sequencing to explore the expression difference and the regulatory role of miRNA in different stages of fetal skin development. Analysis of expression profiles of miRNAs involved collecting embryo samples via high-throughput sequencing, then bioinformatics analyses were performed to validate DEmiRNAs. A total of 363 miRNAs were differentially expressed during the early and mid-pregnancy of development, and upregulated DEmiRNAs were mainly concentrated in the let-7 family. The transfection of let-7b-5p slowed down HaCaT cell proliferation and promoted apoptosis, as evidenced by the cell counting kit-8 assay, quantitative real-time polymerase chain reaction, and flow cytometry. The double luciferin reporter assay also confirmed let-7b-5p and ΔNp63 downregulation through the combination with the 3'-untranslated region of ΔNp63. Moreover, treatment with a let-7b-5p inhibitor upregulated ΔNp63 and activated the phosphoinositide 3-kinase (PI3K)-protein kinase B (AKT) signaling pathway. The let-7b-5p caused a converse effect on HaCaT cells because of Np63 upregulation. Let-7b-5p regulates skin development by targeting ΔNp63 via PI3K-AKT signaling, contributing to future studies on skin development and clinical scar-free healing.

Human and Viral microRNA Expression in Acute and Chronic HIV Infections

Human and viral microRNAs (miRNAs) are involved in the regulation of gene transcription, and the establishment of their profiles in acute (AHI) and chronic (CHI) HIV infections may shed light on the pathogenetic events related to different phases of HIV disease. Next-generation sequencing (NGS) of miRNA libraries was performed, and the reads were used to analyze miRNA differential expression in the plasma with AHI and CHI. Functional analysis was then undertaken to investigate the biological processes characterizing the two phases of HIV infection. Except for hsa-miR-122-5p, which was found in 3.39% AHI vs. 0.18% CHI, the most represented human miRNAs were similarly represented in AHI and CHI. However, when considering the overall detected miRNAs in AHI and CHI, 15 displayed differential expression (FDR p &lt; 0.05). Functional analysis identified 163 target mRNAs involved in promoting angiogenesis activation in AHI versus CHI through the action of hsa-miR10b-5p, hsa-miR1290, hsa-miR1-3p, and hsa-miR296-5p. The viral miRNAs detected, all belonging to herpesviruses, accounted for only 0.014% of total reads. The present data suggest that AHI patients exhibit strong innate immune activation through the upregulation of hsa-miR-122-5p and early activation of angiogenesis. More specific investigations are needed to study the role of viral miRNAs in HIV pathogenesis.

Abstract 5690: Urinary microRNA expression in pancreatic cancer with cachexia

Abstract Background: Cancer cachexia (CC) is a multifactorial syndrome characterized by appetite loss, weight loss and skeletal muscle wasting. It increases functional impairment, treatment-related toxicity, and a lower quality of life. Importantly, CC is one of the leading causes of cancer-related deaths, and its occurrence is particularly prevalent in advanced stages of pancreatic cancer (PC). The syndrome is thought to be associated with systemic inflammation and increased energy expenditure, yet the specific mechanisms remain to be fully understood. Although timely detection and intervention might be crucial in preventing CC’s progression, early diagnosis of this syndrome remains challenging due to the absence of suitable biomarkers. Here, we analyzed urinary microRNAs (miRNAs) as a potential noninvasive biomarkers for detecting CC in patients (pts) with PC. Methods: Clinical information including presence or absence of CC and urine samples were collected from pts with PC treated at National Cancer Center Hospital, Tokyo, Japan from March to October 2022. The diagnosis of CC was made according to the following definitions by Fearon. K et al. (1) weight loss of ≥5% within 6 months; (2) weight loss of ≥2% in pts with a BMI of &amp;lt;20 kg/m2; or (3) weight loss of ≥2% in pts with sarcopenia. In this study, pts who met the definition of CC within 6 months of enrollment were considered to have CC.Urinary miRNA profiles from pts with PC were sequenced by NGS and analyzed using DESeq2 to identify differentially expressed miRNAs (DEMs) between pts with CC and those without. DEMs were further explored for associated pathways and target genes using miRPathDB v.2.0. Results: Of the 64 subjects, 52 pts were included in the analysis after excluding three pts with low urine sample read coverage and nine pts with no information on CC status. Among those, 38 pts were classified as having CC. Patients’ background was as follows (CC/no CC); median age 70 [range: 44-80]/67 [57-74], male and female 19/7 pts, respectively, cStage {I (6/0), II (1/0), III (17/4), IV (14/10)} pts, median BMI (male 22.1 [17.7-27.2]/21.7 [18.2-28.2], female 21.7 [14.6-25.2]/21.9 [16.6-27.4]) kg/m2. Among 307 miRNAs detected in the urine of pts with PC, 49 miRNAs had previously been reported in CC research. The miR-141-5p was differentially expressed between pts with CC and those without CC (Wald test pBH = 0.079). It was down-regulated in pts with CC (Mann-Whitney U test p = 6.4e-4) and most significantly associated with the FoxO signaling pathway (ORA pBH = 1.84e-4), followed by the mTOR signaling pathway (ORA pBH = 0.008). Conclusions: Our study indicates that miR-141-5p holds potential as a noninvasive biomarker for the detection of CC in pts with PC. Although miR-141-5p has not been previously reported in CC, its significant association with the FoxO signaling pathway, which is one of the major pathways regulating skeletal muscle atrophy, underscores the relevance of miR-141-5p in the context of CC. Citation Format: Mao Okada, Shunsuke Kondo, Yukiko Igawa, Tatsuya Yoshida, Milos Havelka, Mika Mizunuma, Yuki Ichikawa, Noboru Yamamoto. Urinary microRNA expression in pancreatic cancer with cachexia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5690.

Abstract 2277: Understanding the impact of clonal hematopoiesis on multiple myeloma: Insights from exosomal RNA analysis and survival analysis

Abstract Clonal Hematopoiesis of Indeterminate Potential (CHIP) is a trait characterized by the accumulation of somatic mutations in hematopoietic stem cells in certain adults. It has been observed that the presence of CHIP can have significant implications for the development and prognosis of various blood cancers. Multiple myeloma (MM), a cancer involving the abnormal proliferation of malignant plasma cells in the bone marrow, is one such condition. Exosomes, a type of extracellular vesicles, play a role in cell communication by carrying information between cells. Analysis of UK Biobank data confirmed the poor prognosis of multiple myeloma patients with CHIP and identified a potential of paracrine effect on plasma cancer cells. To gain deeper insights into this paracrine effect on plasma cells, we attempted to analyze cell-to-cell interaction, such as the expression of miRNAs in exosomes, through exosome RNA sequencing. In our research, exosomes were extracted from bone marrow samples of 30 multiple myeloma patients, and exosome RNA sequencing was performed. Differential expression of miRNAs was identified. A total of 14 down-regulated miRNAs were identified based on adjusted p value &amp;lt; 0.05, log2 fold change &amp;lt; -1. We found that hsa-let-7f-5p, hsa-let-7a-5p, and hsa-miR-148a-3p, which are mainly miRNAs targeting MAP kinase, were significantly down-regulated in the CHIP cases. A computational tool was used to predict and integrate the target genes and pathways regulated by these DE miRNAs. The results suggested that certain inflammatory and oncogenic pathways including the MAPK signaling pathways, were not effectively suppressed in cases with CHIP (KEGG, p &amp;lt; 0.005). In addition, the chemokine signaling pathway and cytokine-cytokine receptor interaction (KEGG, p &amp;lt; 0.005) related to the paracrine effect, integrins in angiogenesis (PID, p &amp;lt; 0.005) related to poor prognosis and metastasis of cancer were also not adequately inhibited in the CHIP cases. Using UK Biobank data, we called somatic CHIP variants excluding those with VAF &amp;lt; 0.02 and analyzed them against ICD-10 codes. Hazard ratios (HR) were computed through Cox proportional regression with covariates. CHIP presence significantly correlated with MM diagnosis (HR: 1.64, CI: 1.35-2, p &amp;lt; 0.001), notably in TET2 mutation carriers (HR: 2.35, CI: 1.63-3.4, p &amp;lt; 0.001). TET2 mutations intensified MGUS to MM progression (HR: 5.0, CI: 2.27-11.0, p &amp;lt; 0.001). CHIP in MM impacted EFS (HR: 1.61, CI: 1.25-2.1, p &amp;lt; 0.001), with TET2 mutation carriers facing higher hazards (HR: 1.66, CI: 1.05-2.6, p = 0.031). CHIP, especially TET2 mutations, significantly contribute to adverse MM outcomes and MGUS to MM evolution. Our findings provide insight into how CHIP influences the tumorigenesis and cancer progression of MM. Citation Format: Jeongmin Park, GangPyo Ryu, Daniel Nachun, Maggie Maurer, Siddhartha Jaiswal, Dong-Yeop Shin, Ja Min Byun, Junshik Hong, Youngil Koh, Sung-Soo Yoon. Understanding the impact of clonal hematopoiesis on multiple myeloma: Insights from exosomal RNA analysis and survival analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2277.