Search Results (178)

Cardiovascular remodeling, encompassing vascular remodeling, myocardial remodeling, and fibrosis-associated tissue remodeling, underlies atherosclerosis, pulmonary hypertension, myocardial infarction, myocardial fibrosis, and other cardiovascular diseases. Its regulation has traditionally been studied through transcriptional, inflammatory, metabolic, mechanical, and intercellular signaling mechanisms. Recent advances in epitranscriptomics have identified N6-methyladenosine (m⁶A) RNA methylation as an additional post-transcriptional layer that interacts with microRNA (miRNA) pathways during cardiovascular disease progression. This review summarizes current evidence for m⁶A-miRNA crosstalk in cardiovascular remodeling, focusing on epitranscriptomic checkpoints that regulate miRNA fate, feedback-like regulatory circuits involving miRNAs and the m⁶A machinery, and cell-type-specific programs across endothelial cells, vascular smooth muscle cells, fibroblasts, and cardiomyocytes. We further discuss emerging analytical technologies and translational implications of this regulatory axis. Future studies should clarify causal mechanisms, cell-type and disease-stage specificity, and translational feasibility. Together, this multilayered framework provides a systems-level perspective on how RNA regulatory networks may shape pathological remodeling in cardiovascular disease. Full article

Background/Objectives: N6-methyladenosine (m⁶A) is the most prevalent and functionally significant internal modification within eukaryotic mRNA. While m⁶A is known to be regulated at internal sites by factors such as splice junctions, the mechanisms governing deposition within the cap-proximal region remain poorly understood. This study aims to determine the patterns of m⁶A stoichiometry in cap-proximal regions and to investigate whether the choice of the specific transcription start site (TSS) can affect m⁶A stoichiometry. Methods: We re-analyzed our published single-nucleotide-resolution CROWN-seq data to quantify m⁶A stoichiometry across transcript isoforms with different TSSs, and assessed the relationship between specific TSSs and specific m⁶A sites (“TSS-m⁶A-site pairs”). Results: We established the first single-nucleotide-resolution dataset of m⁶A stoichiometry across the transcriptome in cap proximal regions, including stoichiometry of m⁶A across 5′ isoforms for each gene. We found that m⁶A deposition is markedly inhibited within a narrow cap-proximal region in a distance-sensitive manner. m⁶A sites located close to both 5′ and 3′ exon ends exhibit low methylation due to the overlap between the cap-proximal and 3′ exon-end exclusion zones. Conclusions: We find that the first exon contains a narrow m⁶A exclusion zone at its 5′ end. As a result, cap-proximal m⁶A sites can have different stoichiometries depending on the TSS choice. As the m⁶A site is positioned farther from the TSS, m⁶A stoichiometry increases. These results reveal that TSS switching is a regulatory mechanism for m⁶A stoichiometry in cap-proximal regions and provide a mechanism for fine-tuning gene expression and mRNA fate through isoform-specific m⁶A modification stoichiometry. Full article

Bone marrow-derived mesenchymal stem cells (BMSCs) maintain skeletal homeostasis by balancing adipogenic and osteogenic differentiation, yet clinically used drugs that bias this fate choice and their mechanisms remain incompletely defined. Here, we investigated whether dextromethorphan (DXM), a widely used antitussive, modulated lineage commitment in rat BMSCs and interrogated candidate upstream signaling modules. Rat BMSCs were induced with adipogenic medium or osteogenic medium in the presence of DXM (30 μM). Adipogenesis and osteogenesis were quantified using Oil Red O and Alizarin Red S staining with elution-based quantification, and lineage markers were measured by RT-qPCR. Intracellular Ca²⁺ and ROS were analyzed using flow cytometry, and the levels of p-AKT and p-ERK were assessed through Western blotting analysis. Under adipogenic induction, DXM increased lipid droplet accumulation and the mRNA levels of Pparγ and Fabp4. Although DXM elevated Ca²⁺ and ROS, the chelation of intracellular Ca²⁺ and pharmacological inhibition of Sig-1R/PLC–IP3R signaling, redox/ROS, NMDA receptors, AKT/ERK, Kv channels, bitter taste receptor-related signaling, and mTOR did not attenuate the DXM-enhanced adipogenesis. DXM reduced p-ERK without increasing p-AKT; U0126 lowered basal adipogenesis but did not block the DXM effect. Under osteogenic induction, DXM reduced matrix mineralization and downregulated Runx2 and Bglap mRNA levels, while Wwtr1 mRNA levels were not significantly changed. DXM also partially reversed the osteogenic induction-associated reduction in Mtor mRNA. Separately, under adipogenic induction, rapamycin attenuated baseline adipogenesis but did not prevent the additional lipid accumulation induced by DXM. Collectively, DXM shifted the osteogenic–adipogenic balance toward adipogenesis through a non-canonical mechanism. Full article

Retinitis pigmentosa (RP) is a genetically heterogeneous group of inherited retinal degenerations with primary degeneration of rod photoreceptors followed by secondary cone loss. We investigated whether downregulating Nrl (neural retina leucine zipper), a key transcription factor specifying rod fate, can reprogram rods into a more resilient state. In a transgenic Nrl^N/N mouse in which Nrl was markedly downregulated, the rod phenotype became more like a rod precursor, particularly in the inferior retina. Crossing Nrl^N/N mice with two rod degeneration models, rd1 (Pde6b^rd1/rd1) and rhodopsin P23H knock-in (Rho^P23H/P23H) mice, showed significantly improved photoreceptor survival in double-mutant mice. In addition, AAV-mediated delivery of shRNA targeting Nrl mRNA substantially enhanced photoreceptor survival in rd10 (Pde6b^rd10/rd10) mice. These findings demonstrate that downregulation of Nrl reprograms rods and confers broad resistance to degeneration across multiple RP models. AAV-mediated Nrl knockdown represents a promising mutation-independent therapeutic strategy for autosomal recessive and dominant forms of RP. Full article

Traditional treatments of autoimmune diseases relying on systemic immunosuppression often lack curative potential and have severe side effects. Mesenchymal stem cells (MSCs) are a promising alternative due to their immunomodulatory properties; however, whole-cell therapies have certain limitations. MSC-derived extracellular vesicles (EVs), including small vesicles—exosomes—have emerged as a safe cell-free therapeutic platform capable of crossing biological barriers and delivering bioactive cargo with low immunogenicity. Various types of RNAs abundantly produced by host MSCs represent a key element of EV content. In particular, EVs carry small RNAs, which essentially determine cellular life and fate. Our review provides a comprehensive mechanistic framework for the use of RNA-loaded EVs, specifically those carrying microRNAs (miRNAs), small interfering RNAs (siRNAs), and messenger RNAs (mRNAs), in restoring immune homeostasis. We detail the biogenesis and molecular mechanisms governing sorting of RNA into EVs, along with endogenous and exogenous engineering strategies to enhance therapeutic potency. We examine how RNA-loaded EVs modulate immunological processes like reprogramming of macrophage M1-M2 polarization, Th17/Treg balance, and suppression of inflammatory signaling pathways such as NF-κB and the NLRP3 inflammasome. We address critical translational challenges—EV heterogeneity, manufacturing scalability, and need for standardized quality control—while outlining future opportunities for RNA-loaded EV-based therapeutics. Full article

The m⁶A RNA methylation pathway plays a critical role in host antiviral defense. Host cells employ m⁶A readers such as YTHDF2 to regulate viral RNA fate through diverse mechanisms, including degradation, translational control, and immune recognition. However, we found that YTHDF2 is essential for SARS-CoV-2 replication, suggesting that a virus may exploit this host machinery to its advantage. Through integrative RNA-proteome analysis, we identified the SARS-CoV-2 nucleocapsid (N) transcript as the most heavily m⁶A-modified viral transcript and a direct interactor of YTHDF2. The N protein forms a complex with YTHDF2 in the cytoplasm and redirects this host RNA decay machinery toward host antiviral transcripts. N suppresses ISG15, IFIT1, MX1 and pro-inflammatory cytokines in a largely YTHDF2-dependent manner, an effect that is lost in YTHDF2-knockout cells. These findings reveal a viral immune evasion strategy wherein a viral protein actively hijacks an m⁶A reader to silence antiviral gene expression, establishing the N-YTHDF2 axis as a therapeutic target against SARS-CoV-2 and other coronaviruses. Full article

Pluripotent stem cell (PSC) differentiation is orchestrated by intricate autocrine and paracrine signaling networks. Among these, exosomes, key components of the cellular secretome, are implicated as crucial mediators of intercellular communication via delivery of bioactive molecules, including microRNAs (miRNAs). This study investigated the role of exosomal miRNAs in stem cell differentiation using Dgcr8-deficient mouse embryonic stem cells (mESCs), which are incapable of producing mature miRNAs. Although the differentiation capacity was markedly impaired in these cells, partial restoration was observed following treatment with exosomes derived from differentiating wild-type mESCs. Exosomal miRNA uptake was confirmed, and gene ontology analysis revealed significant enrichment of pathways associated with cell fate determination, morphogenesis, and apoptosis regulation. Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that exosomal miRNAs modulated multiple osteoinductive signaling cascades, notably the MAPK and TGF-β pathways, in Dgcr8-deficient cells. Apoptotic markers were also downregulated, suggesting a protective effect conferred by the exosomal cargo. Collectively, our results suggest that exosome-mediated delivery of miRNAs may represent a fundamental mechanism by which pluripotent stem cells coordinate stress responses and differentiation trajectories, providing novel insights into the regulation of embryogenesis. Full article

In the search for new therapeutic strategies for the treatment of skin cancer, cannabinoids have become the focus of scientific interest. The present study investigated the effects of the phytocannabinoids Δ⁹-tetrahydrocannabinol (THC) and cannabidiol (CBD) on the viability, apoptosis, and mitochondrial function of human melanoma (A375) and cutaneous squamous cell carcinoma (SCC) cells (A431). Both cannabinoids caused a time- and concentration-dependent loss of viability and an upregulation of caspase-3/7 activity, associated with the induction of initiator caspases-8 and -9, PARP cleavage, and an increase in the autophagy marker LC3A/B-II. Inspired by the latest work on the dual role of heme oxygenase-1 (HO-1) in cell fate, the expression of this enzyme was examined and found to be upregulated at the mRNA and protein level by THC and CBD. Inhibition of HO-1 activity by tin protoporphyrin IX (SnPPIX) reduced the loss of viability caused by both cannabinoids, suggesting a cytotoxic rather than cytoprotective mediator role for this enzyme here. At the mitochondrial level, THC and CBD caused a reduction in membrane potential, a release of cytochrome c into the cytosol, and electron microscopically detectable mitochondrial damages. A more detailed functional analysis revealed an inhibition of mitochondrial oxygen consumption rate, accompanied by a decrease in various subunits of mitochondrial oxidative phosphorylation complexes. In conclusion, our data demonstrate a strong cytotoxic effect of THC and CBD on melanoma and cutaneous SCC cells involving mitochondrial apoptosis and mitochondrial dysfunction. Full article

Over the past two decades, it has become clear that gene expression in eukaryotic cells is regulated by diverse RNA molecules. In this process, new RNAs have been discovered, and the roles of their modified molecules have been progressively elucidated. In this review, we first describe how RNA and its modifications function in virus-infected cells. We use adenovirus and several other viruses as models during the early stages of infection, which we believe determines the fate of infected cells. Next, we reviewed the process of identifying the early mRNA transcription initiation sites in adenovirus-infected cells. The results showed that the transcription initiation sites for the E1 and E4 mRNAs—known as adenovirus oncogenes—are highly complex. The same level of complexity in transcription initiation sites has been suggested for oncogenes in several other DNA tumor viruses, including SV40, polyomavirus, and papillomavirus. It is now understood that the transcription of the early adenovirus mRNA involves alternative splicing, rather than constitutive splicing, as we previously demonstrated. Furthermore, recent research indicates that the abnormal alternative splicing of intracellular mRNA may induce cellular carcinogenesis. Finally, we discuss whether alternative splicing plays a role in the carcinogenic effects of DNA tumor viruses, such as adenovirus. Additionally, we discuss that alternative splicing plays a crucial role in adenovirus replication. Full article

Precise control of keratinocyte proliferation and differentiation is critical for oral epithelial regeneration, yet the mechanobiological cues guiding these processes remain incompletely defined. Here, we systematically evaluated how electrospun polycaprolactone (PCL) scaffolds with defined fiber orientations (aligned vs. random) and diameters (600–800 nm, 1.2–1.7 µm, 2.0–2.5 µm) direct gingival keratinocyte fate. Using immortalized human gingival keratinocytes, we assessed cell and nuclear morphology, proliferation dynamics, differentiation marker expression, and the effects of basal keratin (KRT5/KRT14) knockdown. Quantitative morphological analysis revealed scaffold-dependent changes in cell shape: aligned medium-diameter fibers (with fiber diameters of 1.2–1.7 µm) induced pronounced cell and nuclear elongation, whereas random fibers (600–800 nm) promoted larger, more rounded cell and nuclear shapes. Time-resolved EdU assays indicated that aligned scaffolds supported sustained proliferation, whereas random scaffolds elicited a transient proliferative burst followed by a decline. Gene expression analysis (ddPCR) demonstrated that random scaffolds (especially 600–800 nm fibers) upregulated basal keratins (KRT5, KRT14) and early differentiation markers (KRT1, KRT10, KRT4, KRT13) relative to aligned scaffolds. At the protein level, differentiation markers involucrin (IVL) and filaggrin (FLG) were likewise elevated on random scaffolds, corroborating the mRNA findings. Functional KRT5/KRT14 knockdown experiments revealed scaffold-specific dependencies: cells on random scaffolds required these keratins for viability, whereas aligned cultures remained viable upon KRT5/14 loss. Furthermore, KRT5/14 depletion differentially altered downstream differentiation markers (IVL, KRT1) and mechanotransduction markers (LMNB1, YAP1) in a scaffold-dependent manner. Collectively, these findings establish fiber orientation and diameter as key design parameters for controlling keratinocyte fate. As a translational concept, layered scaffolds combining aligned and random fibers may enable spatially controlled proliferation and differentiation in engineered oral epithelia. Full article

Nonsense-mediated mRNA decay (NMD) is a highly conserved RNA quality and quantity surveillance machinery in eukaryotic cells, serving as an important node in the post-transcriptional gene expression. Previous studies using the complete knockout of individual NMD factors in cells or animals reveal that NMD deficiency causes developmental defects and compromises tissue homeostasis. However, because most NMD factors participate in multiple molecular functions, a direct link between NMD and cell fate determination is missing. SMG6 is a core NMD effector and the only endoribonuclease among all NMD factors. The NMD function of SMG6 is exclusively mediated by its PIN (PilT N-terminus) domain. In this study, we engineered a mouse model with the capability of specifically deactivating the SMG6’s PIN domain/endoribonuclease activity (Smg6-PIN^F/F), but not knocking out the complete SMG6 protein. We found that SMG6’s PIN domain is essential for NMD activity in embryonic stem cells (ESCs) and various tissues of adult mice. Furthermore, loss of SMG6’s PIN domain is dispensable for the mouse ESC self-renewal, but severely compromises the differentiation, which consequently causes the mutant mice to die during the process of organogenesis. Through the induced deletion of SMG6’s PIN domain in adult mice, we found that loss of SMG6’s NMD function affects the homeostasis of several mouse tissues, including the testis and the intestine. In sum, our study establishes a mechanistic link between NMD per se and cell fate determination of mouse ESCs, as well as in the tissues of adult mice, where cell fate transitions are actively ongoing. The Smg6-PIN^F/F mouse line could be a valuable strain for elucidating the biology of NMD per se. Full article

Spermatogenesis is a sophisticated process coordinated by germ cells and the somatic microenvironment. Circular RNAs (circRNAs), key components of competitive endogenous RNA (ceRNA) networks, form intricate post-transcriptional regulatory systems by sequestering microRNAs (miRNAs). However, the specific functions of these networks in spermatogenesis, particularly regarding the cell-intrinsic regulatory programs of germ cells, remain poorly understood. To address this, we utilized a unique foxl3 mutant model in medaka (Oryzias latipes), in which XX female germ cells spontaneously transdifferentiate into functional sperm within the ovarian somatic environment. This model enables the functional enrichment of core spermatogenic programs largely independent of male-specific somatic cues. Through whole-transcriptome sequencing and bioinformatic analysis, we identified 58 key circRNAs, 27 core miRNAs, and 2965 mRNAs, and constructed a candidate ceRNA regulatory network mediated by six circRNAs. Under genetically consistent conditions, this study elucidated a putative ceRNA network directly involved in the germ cell-dominant initiation of spermatogenesis, suggesting an essential role of these networks in germ cell fate determination. These findings provide new insights into the regulatory mechanisms of teleost spermatogenesis and offer valuable molecular targets for advancing reproductive medicine and improving breeding efficiency in aquaculture. Full article

Background: Ovarian granulosa cells (GCs) play a pivotal role in folliculogenesis, and their dysfunction is central to disorders such as polycystic ovary syndrome (PCOS) and premature ovarian failure (POF). MicroRNAs (miRNAs) have emerged as crucial post-transcriptional regulators of GC homeostasis. Method: This review synthesizes current evidence by systematically analyzing relevant studies, integrating data from in vitro GC models, animal experiments, human cell lines, and clinical samples to elucidate the specific mechanisms by which miRNAs regulate GCs. Results: miRNAs precisely modulate GC proliferation, apoptosis, steroidogenesis, and oxidative stress responses by targeting key signaling pathways (e.g., PI3K/AKT/mTOR, TGF-β/SMAD) and functional genes (e.g., TP53, CYP19A1). Exosomal miRNAs serve as vital mediators of communication within the follicular microenvironment. To date, nearly 200 miRNAs have been associated with PCOS. Conclusions: miRNAs constitute a decisive regulatory network governing GC fate, offering promising therapeutic targets for PCOS and POF. However, significant challenges remain, primarily miRNA pleiotropy and the lack of follicle-specific delivery systems. Future clinical translation requires rigorous validation in human-relevant models. Full article

People living with HIV-1 (PLWH) are part of the so-called “fragile” populations to which COVID-19 vaccines were/are strongly recommended. The fact that most widely used COVID-19 vaccines rely on the production of a biologically active SARS-CoV-2 Spike protein expressed by synthetic mRNA poses the relevant question of whether and how this vaccination influences the fate of the HIV-1 reservoir. This report presents a detailed analysis of the literature data on the effects of SARS-CoV-2 Spike and COVID-19 vaccines on HIV-1 latently infected cells. Despite being limited in number, the experimental evidences consistently indicate that vaccine mRNA and/or SARS-CoV-2 Spike can effectively reactivate latent HIV-1. This conclusion has been drawn after “in vitro”, “ex vivo”, and “in vivo” assays, and with virus-associated Spike, soluble Spike, or its intracellular expression, as well as with COVID-19 mRNA vaccines. On the other hand, real-world observations on vaccinated PLWH under antiretroviral therapy (ART) provided evidence of HIV-1 reactivation almost exclusively in PLWH with unsuppressed viremia, as measured in terms of size of the HIV-1 reservoir. Although several issues still need to be clarified through urgent additional investigations, these data suggest the possibility that the Spike protein and/or the vaccine mRNA molecules affect the HIV-1 latency in PLWH. Full article

As a type of cell with self-renewal ability and multi-directional differentiation potential, stem cells are closely related to their functions, such as reprogramming transcription factors, histone modifications, and energy metabolism. m⁶A (N⁶-methyladenosine modification) is one of the most abundant modifications in RNA, and dynamic reversible m⁶A modification plays an important role in regulating stem cell function. This review moves beyond listing isolated functions and instead adopts an integrated perspective, viewing m⁶A as a temporal regulator of cellular state transitions. We discuss how m⁶A dynamically regulates stem cell pluripotency, coordinates epigenetic and metabolic reprogramming, and serves as a central hub integrating key signaling pathways (Wnt, PI3K-AKT, JAK-STAT, and Hippo). Finally, using somatic reprogramming as an example, we elucidate the stage-specific role of m⁶A in complex fate transitions. This comprehensive exposition not only clarifies the context-dependent logic of m⁶A regulation but also provides a precise framework for targeting the m⁶A axis in regenerative medicine and cancer therapy. Full article