Mechanism of Panax notoginseng saponins modulation of miR-214-3p/NR1I3 affecting the pharmacodynamics and pharmacokinetics of warfarin.

Background: With the prevalence of dietary supplements, the use of combinations of herbs and drugs is gradually increasing, together with the risk of drug interactions. In our clinical work, we unexpectedly found that the combination of Panax notoginseng and warfarin, which are herbs that activate blood circulation and remove blood stasis, showed antagonistic effects instead. The purpose of this study was to evaluate the drug interaction between Panax notoginseng saponins (PNS) and warfarin, the main active ingredient of Panax notoginseng, and to explore the interaction mechanism. Methods: The effects and mechanisms of PNS on the pharmacodynamics and pharmacokinetics of warfarin were explored mainly in Sprague-Dawley rats and HepG2 cells. Elisa was used to detect the concentrations of coagulation factors, HPLC-MS to detect the blood concentrations of warfarin in rats, immunoblotting was employed to examine protein levels, qRT-PCR to detect mRNA levels, cellular immunofluorescence to detect the localization of NR1I3, and dual luciferase to verify the binding of miR-214-3p and NR1I3. Results: PNS significantly accelerated warfarin metabolism and reduced its efficacy, accompanied by increased expression of NR1I3 and CYP2C9. Interference with NR1I3 rescued the accelerated metabolism of warfarin induce by PNS co-administration. In addition, we demonstrated that PNS significantly reduced miR-214-3p expression, whereas miR-214-3p overexpression reduced NR1I3 and CYP2C9 expression, resulting in a weakened antagonistic effect of PNS on warfarin. Additionally, we found that miR-214-3p bound directly to NR1I3 3'-UTR and significantly downregulated NR1I3 expression. Conclusion: Our study demonstrated that PNS accelerates warfarin metabolism and reduces its pharmacodynamics by downregulating miR-214-3p, leading to increased expression of its target gene NR1I3, these findings provide new insights for clinical drug applications to avoid adverse effects.

Mesenchymal stem cell-derived extracellular vesicles mitigate neuronal damage from intracerebral hemorrhage by modulating ferroptosis.

BACKGROUND: Hemorrhagic stroke is a devastating cerebrovascular event with a high rate of early mortality and long-term disability. The therapeutic potential of mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) for neurological conditions, such as intracerebral hemorrhage (ICH), has garnered considerable interest, has garnered considerable interest, though their mechanisms of action remain poorly understood. METHODS: EVs were isolated from human umbilical cord MSCs, and SPECT/CT was used to track the 99mTc-labeled EVs in a mouse model of ICH. A series of comprehensive evaluations, including magnetic resonance imaging (MRI), histological study, RNA sequencing (RNA-Seq), or miRNA microarray, were performed to investigate the therapeutic action and mechanisms of MSC-EVs in both cellular and animal models of ICH. RESULTS: Our findings show that intravenous injection of MSC-EVs exhibits a marked affinity for the ICH-affected brain regions and cortical neurons. EV infusion alleviates the pathological changes observed in MRI due to ICH and reduces damage to ipsilateral cortical neurons. RNA-Seq analysis reveals that EV treatment modulates key pathways involved in the neuronal system and metal ion transport in mice subjected to ICH. These data were supported by the attenuation of neuronal ferroptosis in neurons treated with Hemin and in ICH mice following EV therapy. Additionally, miRNA microarray analysis depicted the EV-miRNAs targeting genes associated with ferroptosis, and miR-214-3p was identified as a regulator of neuronal ferroptosis in the ICH cellular model. CONCLUSIONS: MSC-EVs offer neuroprotective effects against ICH-induced neuronal damage by modulating ferroptosis highlighting their therapeutic potential for combating neuronal ferroptosis in brain disorders.

Morinda Officinalis Polysaccharides Inhibit Osteoclast Differentiation by Regulating miR-214-3p/NEDD4L in Postmenopausal Osteoporosis Mice.

To investigate the potential mechanism of Morinda officinalis F. C. How polysaccharides (MOPs) in regulating osteoclast differentiation and apoptosis through miR-214-3p and its target protein. Ovariectomy was performed in 8-week female C57BL6 mice to establish the postmenopausal osteoporosis (PMOP) model. Mice were treated immediately with 500 mg/kg of MOPs (prevention group); others were treated 2 weeks after operation (treatment group). Left femur bone mineral density (BMD) was examined. RAW264.7 cells were administered with receptor activator of NF-κB ligand (RANKL) to establish the osteoclast (OC) model and treated with serum containing 1 or 2 g/kg of MOPs. Apoptosis-related indexes, miR-214-3p, and Expressed Developmentally Down-regulated 4-Like (NEDD4L) were detected by western blot, quantitative real-time-reverse transcription polymerase chain reaction (qRT-PCR), and flow cytometry. OC received a miR-214-3p inhibitor or NEDD4L small interfering RNA (siRNA). MOPs reversed the PMOP-induced changes in bones. Compared with the RANKL group, MOPs increased the apoptosis and related markers in OCs. MOPs decreased the femur miR-214-3p of PMOP mice (P < 0.001). Higher concentrations of MOPs reversed the upregulation of miR-214 mRNA in OCs (P < 0.001). miR-214-3p inhibitor increased the expression of Bax and CC3 (P < 0.01) and decreased the expression of Bcl-2 (P < 0.05). NEDD4L is targeted by miR-214. NEDD4L was upregulated in the RANKL + MOPs group (P < 0.01). miR-214-3p inhibitor increased the upregulation of NEDD4L induced by MOPs (P < 0.05). siRNA NEDD4L significantly reversed the inhibition of MOPs on osteoclast differentiation with miR-214-3p inhibitor (P < 0.01). MOPs effectively prevent PMOP by inhibiting osteoclastogenesis and inducing OC apoptosis through the miR-214-3p/NEDD4L pathway.

Delivery of miR-214 via extracellular vesicles downregulates Xbp1 expression and pro-inflammatory cytokine genes in macrophages.

Aim: Tumor-infiltrating macrophages are tumor-promoting and show activation of the unfolded protein response (UPR). The transcription factor X-box binding protein 1 (XBP1) is a conserved element of the UPR. Upon activation, the UPR mediates the transcriptional activation of pro-inflammatory cytokines and immune suppressive factors, hence contributing to immune dysregulation in the tumor microenvironment (TME). miR-214 is a short non-coding miRNA that targets the 3'-UTR of the Xbp1 transcript. Here, we tested a new method to efficiently deliver miR-214 to macrophages as a potential new therapeutic approach. Methods: We generated miR-214-laden extracellular vesicles (iEV-214) in a murine B cell and demonstrated that iEV-214 were enriched in miR-214 between 1,500 - 2,000 fold relative to control iEVs. Results: Bone marrow-derived macrophages (BMDM) treated with iEV-214 for 24 h underwent a specific enrichment in miR-214, suggesting transfer of the miR-214 payload from the iEVs to macrophages. iEV-214 treatment of BMDM markedly reduced (> 50%) Xbp1 transcription under endoplasmic reticulum stress conditions compared to controls. Immune-related genes downstream of XBP1s (Il-6, Il-23p19, and Arg1) were also reduced by 69%, 51%, and 34%, respectively. Conclusions: Together, these data permit to conclude that iEV-214 are an efficient strategy to downregulate the expression of Xbp1 mRNA and downstream genes in macrophages. We propose miRNA-laden iEVs are a new approach to target macrophages and control immune dysregulation in the TME.

MiR-214_L-1R+4 regulate gossypol-induced immune response through MyD88-dependent signaling pathway in Cyprinus carpio.

MicroRNAs (miRNAs) have been demonstrated to act as crucial modulators with considerable impacts on the immune system. Cottonseed meal is often used as a protein source in aqua feed, cottonseed meal contains gossypol, which is harmful to animals. However, there is a lack of research on the role of miRNAs in fish exposed to gossypol stress. To determine the regulatory effects of miRNAs on gossypol toxicity, Cyprinus carpio were given to oral administration of 20 mg/kg gossypol for 7 days, and the gossypol concentration in the tissues was tested. Then, we detected spleen index, histology, immune enzyme activities of fish induced by gossypol. The results of miRNA sequencing revealed 8 differentially expressed miRNAs in gossypol group, and miR-214_L-1R+4 was found involved in immune response induced by gossypol. The potential targets of miR-214_L-1R+4 were predicted, and found a putative miR-214_L-1R+4 binding site in the 3'UTR of MyD88a. Furthermore, dual-luciferase reporter assays displayed miR-214_L-1R+4 decreased MyD88a expression through binding to the 3'UTR of MyD88a. Moreover, miR-214_L-1R+4 antagomir were intraperitoneally administered to C. carpio, down-regulated miR-214_L-1R+4 could increase MyD88a expression, as well as inflammatory cytokines and anti-inflammatory cytokines expression. These findings revealed that miR-214_L-1R+4 via the MyD88-dependent signaling pathway modulate the immune response to gossypol in C. carpio spleen.

LncRNA ZFAS1 promotes invasion of medullary thyroid carcinoma by enhancing EPAS1 expression via miR-214-3p/UCHL1 axis.

lncRNA ZFAS1 was identified to facilitate thyroid cancer, but its role in medullary thyroid carcinoma (MTC) remains unknown. This study aimed to unravel the potential function of this lncRNA in MTC by investigating the involvement of the lncRNA ZFAS1 in a ceRNA network that regulates MTC invasion. Proliferation, invasion, and migration of cells were evaluated using EdU staining and Transwell assays. Immunoprecipitation (IP) assays, dual-fluorescence reporter, and RNA IP assays were employed to examine the binding interaction among genes. Nude mice were used to explore the role of lncRNA ZFAS1 in MTC in vivo. ZFAS1 and EPAS1 were upregulated in MTC. Silencing ZFAS1 inhibited MTC cell proliferation and invasion under hypoxic conditions, which reduced EPAS1 protein levels. UCHL1 knockdown increased EPAS1 ubiquitination. ZFAS1 positively regulated UCHL1 expression by binding to miR-214-3p. Finally, silencing ZFAS1 significantly repressed tumor formation and metastasis in MTC. LncRNA ZFAS1 promotes invasion of MTC by upregulating EPAS1 expression via the miR-214-3p/UCHL1 axis.

Luteolin protects mouse hippocampal neuronal cells against isoflurane-induced neurotoxicity through miR-214/PTEN/Akt pathway.

Isoflurane is one of the most commonly used anaesthetic agents in surgery procedures. During the past decades, isoflurane has been found to cause impairment in neurological capabilities in new-borns and elderly patients. Luteolin is a flavonoid that has been documented to possess a neuroprotective effect. Here we investigated the putative neuroprotective effects of luteolin on isoflurane-induced neurotoxicity in mouse hippocampal neuronal HT22 cells and explored the potential mechanisms. We demonstrated that luteolin improved mitochondrial dysfunction and reduced oxidative stress and apoptosis in isoflurane-treated HT22 cells, and thus inhibiting the isoflurane-induced neuronal injury. Further investigations showed that isoflurane exposure caused miR-214 downregulation, which could be mitigated by treatment with luteolin. Knockdown of miR-214 attenuated the neuroprotection of luteolin on isoflurane-induced neuronal injury. More importantly, luteolin inhibited isoflurane-caused regulation of the PTEN/Akt pathway, while miR-214 knockdown altered the regulatory effect of luteolin on the PTEN/Akt pathway. Furthermore, the effects of miR-214 knockdown on the neuroprotection of luteolin could also be prevented by knockdown of PTEN, implying that the neuroprotective effect of luteolin was mediated by miR-214/PTEN/Akt signaling pathway. These findings provided evidence for the potential application of luteolin in preventing isoflurane-induced neurotoxicity.

MiR-214-3p regulates Piezo1, lysyl oxidases and mitochondrial function in human cardiac fibroblasts.

Cardiac fibroblasts are pivotal regulators of cardiac homeostasis and are essential in the repair of the heart after myocardial infarction (MI), but their function can also become dysregulated, leading to adverse cardiac remodelling involving both fibrosis and hypertrophy. MicroRNAs (miRNAs) are noncoding RNAs that target mRNAs to prevent their translation, with specific miRNAs showing differential expression and regulation in cardiovascular disease. Here, we show that miR-214-3p is enriched in the fibroblast fraction of the murine heart, and its levels are increased with cardiac remodelling associated with heart failure, or in the acute phase after experimental MI. Tandem mass tagging proteomics and in-silico network analyses were used to explore protein targets regulated by miR-214-3p in cultured human cardiac fibroblasts from multiple donors. Overexpression of miR-214-3p by miRNA mimics resulted in decreased expression and activity of the Piezo1 mechanosensitive cation channel, increased expression of the entire lysyl oxidase (LOX) family of collagen cross-linking enzymes, and decreased expression of an array of mitochondrial proteins, including mitofusin-2 (MFN2), resulting in mitochondrial dysfunction, as measured by citrate synthase and Seahorse mitochondrial respiration assays. Collectively, our data suggest that miR-214-3p is an important regulator of cardiac fibroblast phenotypes and functions key to cardiac remodelling, and that this miRNA represents a potential therapeutic target in cardiovascular disease.

LINC01535 promotes hepatocellular carcinoma proliferation and metastasis by regulating the miR-214-3p/VASP axis.

Background: Emerging evidence has indicated that long noncoding RNAs (lncRNAs) are associated with the development and progression of several carcinomas, including hepatocellular carcinoma (HCC). However, the role of LINC01535 in HCC is still unknown. Materials and methods: In this study, RNA-seq, CCK-8, colony formation, wound healing, Transwell and tumor xenograft assays were used to explore the function of LINC01535 in the proliferation and metastasis of HCC in vitro and in vivo. Fluorescence in situ hybridization (FISH) assay, bioinformatics analysis, dual-luciferase assay, quantitative real-time polymerase chain reaction (qRT-PCR), and western blot analysis were used to reveal the interactions of LINC01535, miR-214-3p and VASP. Results: LINC01535 was overexpressed in HCC tissues and HCC cell lines. Gain- and loss-of-function studies revealed that LINC01535 could promote HCC cell proliferation, migration and invasion both in vitro and in vivo. In addition, upregulation of LINC01535 significantly decreased the expression of microRNA-214-3p (miR-214-3p), which was found closely associated with suppressing tumor progression. Moreover, VASP was identified as a direct downstream target gene of miR-214-3p. LINC01535 positively regulated VASP expression by sponging miR-214-3p, and VASP overexpression activated the PI3K/AKT signaling pathway and stimulated epithelial-to-mesenchymal transition (EMT) in HCC. Conclusions: Our study first found that LINC01535 promoted HCC progression by regulating its downstream target, the miR-214-3p/VASP axis, via the PI3K/AKT signaling pathway. The function and novel regulatory mechanism of LINC01535 may provide a valuable target for the diagnosis and treatment of HCC patients.

miR-214-PTEN pathway is a potential mechanism for stress-induced immunosuppression affecting chicken immune response to avian influenza virus vaccine.

Stress-induced immunosuppression (SIIS) is one of common problems in the intensive poultry industry, affecting the effect of vaccine immunization and leading to high incidences of diseases. In this study, the expression characteristics and regulatory mechanisms of miR-214 in the processes of SIIS and its influence on the immune response to avian influenza virus (AIV) vaccine in chicken were explored. The qRT-PCR results showed that serum circulating miR-214 was significantly differentially expressed (especially on 2, 5, and 28 days post immunization (dpi)) in the processes, so had the potential as a molecular marker. MiR-214 expressions from multiple tissues were closely associated with the changes in circulating miR-214 expression levels. MiR-214-PTEN regulatory network was a potential key regulatory mechanism for the heart, bursa of Fabricius, and glandular stomach to participate in the process of SIIS affecting AIV immune response. This study can provide references for further understanding of stress affecting immune response.

Elevated microRNA-214-3p level ameliorates neuroinflammation after spinal cord ischemia-reperfusion injury by inhibiting Nmb/Cav3.2 pathway.

BACKGROUND: Neuromedin B (Nmb) plays a pivotal role in the transmission of neuroinflammation, particularly during spinal cord ischemia-reperfusion injury (SCII). However, the detailed molecular mechanisms underlying this process remain elusive. METHODS: The SCII model was established by clamping the abdominal aorta of male Sprague-Dawley (SD) rats for 60 min. The protein expression levels of Nmb, Cav3.2, and IL-1ß were detected by Western blotting, while miR-214-3p expression was quantified by qRT-PCR. The targeted regulation between miR-214-3p and Nmb was investigated using a dual-luciferase reporter gene assay. The cellular localization of Nmb and Cav3.2 with cell-specific markers was visualized by immunofluorescence staining. The specific roles of miR-214-3p on the Nmb/Cav3.2 interactions in SCII-injured rats were explored by intrathecal injection of Cav3.2-siRNA, PD168368 (a specific NmbR inhibitor) and synthetic miR-214-3p agomir and antagomir in separate experiments. Additionally, hind-limb motor function was evaluated using the modified Tarlov scores. RESULTS: Compared to the Sham group, the protein expression levels of Nmb, Cav3.2, and the proinflammatory factor Interleukin(IL)-1ß were significantly elevated at 24 h post-SCII. Intrathecal injection of PD168368 and Cav3.2-siRNA significantly suppressed the expression of Cav3.2 and IL-1ß compared to the SCII group. The miRDB database and dual-luciferase reporter gene assay identified Nmb as a direct target of miR-214-3p. As expected, in vivo overexpression of miR-214-3p by agomir-214-3p pretreatment significantly inhibited the increases in Nmb, Cav3.2 and IL-1ß expression and improved lower limb motor function in SCII-injured rats, while antagomiR-214-3p pretreatment reversed these effects. CONCLUSIONS: Nmb protein levels positively correlated with Cav3.2 expression in SCII rats. Upregulating miR-214-3p ameliorated hind-limb motor function and protected against neuroinflammation via inhibiting the aberrant Nmb/Cav3.2 interactions and downstream IL-1ß release. These findings provide novel therapeutic targets for clinical prevention and treatment of SCII.

CircCOL1A1 promotes proliferation, migration, and invasion of colorectal cancer (CRC) cells and glutamine metabolism through GLS1 up-regulation by sponging miR-214-3p.

BACKGROUND: Circular ribose nucleic acids (circRNAs), an abundant type of noncoding RNAs, are widely expressed in eukaryotic cells and exert a significant impact on the initiation and progression of various disorders, including different types of cancer. However, the specific role of various circRNAs in colorectal cancer (CRC) pathology is still not fully understood. METHODS: The initial step involved the use of quantitative reverse transcription polymerase chain reaction (RT-qPCR) to assess the expression levels of circRNAs and messenger RNA (mRNA) in CRC cell lines and tissues. Subsequently, functional analyses of circCOL1A1 knockdown were conducted in vitro and in vivo through cell counting kit (CCK)-8, colony formation and transwell assays, as well as xenograft mouse model of tumor formation. Molecular expression and interactions were investigated using luciferase reporter assays, Western blot analysis, RNA immunoprecipitation (RIP), and immunohistochemical staining. RESULTS: The RT-qPCR results revealed elevated levels of circCOL1A1 expressions in CRC tissues and cell lines as compared to the normal counterparts. In addition, circCOL1A1 expression level was found to be correlated with TNM stage, lymph node metastases, distant metastases, and invasion. Knockdown of circCOL1A1 resulted in impaired invasion, migration, and proliferation of CRC cells, and suppressed tumor generation in the animal model. We further demonstrated that circCOL1A1 could act as a sponge for miR-214-3p, suppressing miR-214-3p activity and leading to the upregulation of GLS1 protein to promote glutamine metabolism. CONCLUSION: These findings suggest that circCOL1A1 functions as an oncogenic molecule to promote CRC progression via miR-214-3p/GLS1 axis, hinting on the potential of circCOL1A1 as a therapeutic target for CRC.

Depression of LncRNA DANCR alleviates tubular injury in diabetic nephropathy by regulating KLF5 through sponge miR-214-5p.

OBJECTIVE: Diabetic nephropathy (DN) manifests a critical aspect in the form of renal tubular injury. The current research aimed to determine the function and mechanism of long non-coding ribonucleic acid (LncRNA) differentiation antagonising non-protein coding RNA (DANCR), with a focus on its impact on renal tubular injury. METHODS: Quantitative reverse transcription polymerase chain reaction was employed to analyze the RNA levels of DANCR in the serum of patients with DN or human proximal tubular epithelial cells (human kidney 2 [HK2]). The diagnostic significance of DANCR was assessed using a receiver operating characteristic curve. A DN model was established by inducing HK-2 cells with high glucose (HG). Cell proliferation, apoptosis, and the levels of inflammatory factors, reactive oxygen species (ROS), and malondialdehyde (MDA) were detected using the Cell Counting Kit - 8, flow cytometry, and enzyme-linked immunosorbent assay. The interaction between microRNA (miR)-214-5p and DANCR or Krüppel-like factor 5 (KLF5) was investigated using RNA immunoprecipitation and dual-luciferase reporter assays. RESULTS: Elevated levels of DANCR were observed in the serum of patients with DN and HG-inducted HK-2 cells (P < 0.05). DANCR levels effectively identified patients with DN from patients with type 2 diabetes mellitus. Silencing of DANCR protected against HG-induced tubular injury by restoring cell proliferation, inhibiting apoptosis, and reducing the secretion of inflammatory factors and oxidative stress production (P < 0.05). DANCR functions as a sponge for miR-214-5p, and the mitigation of DANCR silencing on HG-induced renal tubular injury was partially attenuated with reduced miR-214-5p (P < 0.05). Additionally, KLF5 was identified as the target of miR-214-5p. CONCLUSION: DANCR was identified as diagnostic potential for DN and the alleviation of renal tubular injury via the miR-214-5p/KLF5 axis, following DANCR silencing, introduces a novel perspective and approach to mitigating DN.

LINC00942 Alleviates NaAsO2-induced Apoptosis by Promoting GSH Synthesis Through Targeting miR-214-5p.

The mechanism of arsenic-induced liver toxicity is not fully understood. This study aimed to investigate the role of LINC00942 in arsenic-induced hepatotoxicity by regulating miR-214-5p. As the exposure dose of NaAsO2 gradually increases, cell viability, intracellular GSH content, ΔΨm, and the protein levels of GCLC and GCLM were reduced significantly. Apoptosis rate, ROS, and expression of apoptosis-related and NF-κB pathway proteins increased. The expression of LINC00942 was increased, while the expression of miR-214-5p was decreased. After suppressing LINC00942 levels, NaAsO2 exposure further decreased cell viability, intracellular GSH content, ΔΨm, GCLC protein, and miR-214-5p expression. The apoptosis rate, ROS, and apoptosis-related and NF-κB pathway proteins further increased. miR-214-5p is targeted and negatively regulated by LINC00942. After miR-214-5p was overexpressed, NaAsO2 further decreased cell viability, intracellular GSH content, ΔΨm, and GCLC protein expression compared to NaAsO2 exposure. The apoptosis rate, ROS, apoptosis-related and NF-κB pathway proteins p65, and IKKß were higher than those exposed to NaAsO2. LINC00942 inhibitor along with miR-214-5p inhibitor combined with NaAsO2 treatment resulted in increased cell viability, GSH, Bcl-2, and GCLC protein expression and decreased apoptosis rate, apoptosis related, p65, IKKß protein, and ΔΨm, as compared to the combined NaAsO2 and si LINC00942 group. NaAsO2 exposure induces oxidative damage and apoptosis in LX-2 cells by activating NF-κB and inhibiting GSH synthesis. During this process, the expression level of LINC00942 increases, targeting to reduce the level of miR-214-5p, then weakening the effect of NaAsO2 on NF-κB, thereby alleviating cellular oxidative damage and playing a protective role.

The Rejuvenation and Functional Restoration of Aged Adipose Stem Cells by DUXAP10 Knockdown via the Regulation of the miR-214-3p/RASSF5 Axis.

Adipose stem cell (ASC)-based therapies provide an encouraging option for tissue repair and regeneration. However, the function of these cells declines with aging, which limits their clinical transformation. Recent studies have outlined the involvement of long non-coding RNAs in stem cell aging. Here, we reanalyzed our published RNA sequencing (RNA-seq) data profiling differences between ASCs from young and old donors and identified a lncRNA named double homeobox A pseudogene 10 (DUXAP10) as significantly accumulated in aged ASCs. Knocking down DUXAP10 promoted stem cell proliferation and migration and halted cell senescence and the secretion of proinflammatory cytokines. In addition, DUXAP10 was located in the cytoplasm and functioned as a decoy for miR-214-3p. miR-214-3p was downregulated in aged ASCs, and its overexpression rejuvenated aged ASCs and reversed the harm caused by DUXAP10. Furthermore, Ras Association Domain Family Member 5 (RASSF5) was the target of miR-214-3p and was upregulated in aged ASCs. Overexpressing DUXAP10 and inhibiting miR-214-3p both enhanced RASSF5 content in ASCs, while DUXAP10 knockdown promoted the therapeutic ability of aged ASCs for skin wound healing. Overall, this study offers new insights into the mechanism of age-related ASC dysfunction and names DUXAP10 and miR-214-3p as potential targets for energizing aged stem cells.

Dysregulation and antimetastatic function of circLRIG1 modulated by miR-214-3p/LRIG1 axis in bladder carcinoma.

CircLRIG1, a newly discovered circRNA, has yet to have its potential function and biological processes reported. This study explored the role of circLRIG1 in the development and progression of bladder carcinoma and its potential molecular mechanisms. Techniques such as qRT-PCR, Western blot, various cellular assays, and in vivo models were used to investigate mRNA and protein levels, cell behavior, molecular interactions, and tumor growth. The results showed that both circLRIG1 and LRIG1 were significantly reduced in bladder carcinoma tissues and cell lines. Low circLRIG1 expression was associated with poor patient prognosis. Overexpressing circLRIG1 inhibited bladder carcinoma cell growth, migration, and invasion, promoted apoptosis, and decreased tumor growth and metastasis in vivo. Importantly, circLRIG1 was found to sponge miR-214-3p, enhancing LRIG1 expression, and its overexpression also modulated protein levels of E-cadherin, N-cadherin, Vimentin, and LRIG1. Similar effects were observed with LRIG1 overexpression. Notably, a positive correlation was found between circLRIG1 and LRIG1 expression in bladder carcinoma tissues. Additionally, the tumor-suppressing effect of circLRIG1 was reversed by overexpressing miR-214-3p or silencing LRIG1. The study concludes that circLRIG1 suppresses bladder carcinoma progression by enhancing LRIG1 expression via sponging miR-214-3p, providing a potential strategy for early diagnosis and treatment of bladder carcinoma.

MicroRNAs dysregulated in multiple sclerosis affect the differentiation of CG-4 cells, an oligodendrocyte progenitor cell line.

Introduction: Demyelination is one of the hallmarks of multiple sclerosis (MS). While remyelination occurs during the disease, it is incomplete from the start and strongly decreases with its progression, mainly due to the harm to oligodendrocyte progenitor cells (OPCs), causing irreversible neurological deficits and contributing to neurodegeneration. Therapeutic strategies promoting remyelination are still very preliminary and lacking within the current treatment panel for MS. Methods: In a previous study, we identified 21 microRNAs dysregulated mostly in the CSF of relapsing and/or remitting MS patients. In this study we transfected the mimics/inhibitors of several of these microRNAs separately in an OPC cell line, called CG-4. We aimed (1) to phenotypically characterize their effect on OPC differentiation and (2) to identify corroborating potential mRNA targets via immunocytochemistry, RT-qPCR analysis, RNA sequencing, and Gene Ontology enrichment analysis. Results: We observed that the majority of 13 transfected microRNA mimics decreased the differentiation of CG-4 cells. We demonstrate, by RNA sequencing and independent RT-qPCR analyses, that miR-33-3p, miR-34c-5p, and miR-124-5p arrest OPC differentiation at a late progenitor stage and miR-145-5p at a premyelinating stage as evidenced by the downregulation of premyelinating oligodendrocyte (OL) [Tcf7l2, Cnp (except for miR-145-5p)] and mature OL (Plp1, Mbp, and Mobp) markers, whereas only miR-214-3p promotes OPC differentiation. We further propose a comprehensive exploration of their change in cell fate through Gene Ontology enrichment analysis. We finally confirm by RT-qPCR analyses the downregulation of several predicted mRNA targets for each microRNA that possibly support their effect on OPC differentiation by very distinctive mechanisms, of which some are still unexplored in OPC/OL physiology. Conclusion: miR-33-3p, miR-34c-5p, and miR-124-5p arrest OPC differentiation at a late progenitor stage and miR-145-5p at a premyelinating stage, whereas miR-214-3p promotes the differentiation of CG-4 cells. We propose several potential mRNA targets and hypothetical mechanisms by which each microRNA exerts its effect. We hereby open new perspectives in the research on OPC differentiation and the pathophysiology of demyelination/remyelination, and possibly even in the search for new remyelinating therapeutic strategies in the scope of MS.

The lncRNA lnc_AABR07044470.1 promotes the mitochondrial-damaged inflammatory response to neuronal injury via miR-214-3p/PERM1 axis in acute ischemic stroke.

PURPOSE: We investigated the role of lnc_AABR07044470.1 on the occurrence and development of acute ischemic stroke (AIS) and neuronal injury by targeting the miR-214-3p/PERM1 axis to find a novel clinical drug target and prediction and treatment of AIS. METHODS: The mouse AIS animal model was used in vivo experiments and hypoxia/reoxygenation cell model in vitro was established. Firstly, infarction volume and pathological changes of mouse hippocampal neurons were detected using HE staining. Secondly, rat primary neuron apoptosis was detected by flow cytometry assay. The numbers of neuron, microglia and astrocytes were detected using immunofluorescence (IF). Furthermore, binding detection was performed by bioinformatics database and double luciferase reporter assay. Lnc_AABR07044470.1 localization was performed using fluorescence in situ hybridization (FISH).Lnc_AABR07044470.1, miR-214-3pand PERM1mRNA expression was performed using RT-qPCR. NLRP3, ASC, Caspase-1 and PERM1 protein expression was performed using Western blotting. IL-1ß was detected by ELISA assay. RESULTS: Mouse four-vessel occlusion could easily establish the animal model, and AIS animal model had an obvious time-dependence. HE staining showed that, compared with the sham group, infarction volume and pathological changes of mouse hippocampal neurons were deteriorated in the model group. Furthermore, compared with the sham group, neurons were significantly reduced, while microglia and astrocytes were significantly activated. Moreover, the bioinformatics prediction and detection of double luciferase reporter confirmed the binding site of lnc_AABR07044470.1 to miR-214-3p and miR-214-3p to Perm1. lnc_AABR07044470.1 and PERM1 expression was significantly down-regulated and miR-214-3pexpression was significantly up-regulated in AIS animal model in vivo. At the same time, the expression of inflammasome NLRP3, ASC, Caspase-1 and pro-inflammatory factor IL-1ß was significantly up-regulated in vivo and in vitro. The over-expression of lnc_AABR07044470.1 and miR-214-3p inhibitor could inhibit the neuron apoptosis and the expression of inflammasome NLRP3, ASC, Caspase-1 and pro-inflammatory factor IL-1ß and up-regulate the expression of PERM1 in vitro. Finally, over-expression of lnc_AABR07044470.1 and miR-214-3p inhibitor transfected cell model was significant in relieving the AIS and neuronal injury. CONCLUSION: Lnc_AABR07044470.1 promotes inflammatory response to neuronal injury via miR-214-3p/PERM1 axis in AIS.

The novel circRNA circ_0045881 inhibits cell proliferation and invasion by targeting mir-214-3p in triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is the most lethal subtype of breast cancer (BC). The circRNA-miRNA‒mRNA axis is a promising biomarker for the early diagnosis and prognosis of BC. However, the critical circRNA mediators involved in TNBC progression and the underlying regulatory mechanism involved remain largely unclear. METHODS: In this study, we carried out a circRNA microarray analysis of 6 TNBC patients and performed a gene ontology (GO) analysis. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis was used to characterize important circRNAs involved in TNBC progression. The interaction between circRNAs and miRNAs was determined by dual luciferase and RNA immunoprecipitation (RIP) assays. Moreover, Transwell, wound healing and Cell Counting Kit-8 (CCK8) assays were performed with altered circRNA or miRNA expression in MDA-MB-231 and BT-549 cells to investigate the roles of these genes in cell invasion, migration and proliferation. RESULTS: A total of 78 circRNAs were differentially expressed in TNBC tissues, and the hsa_circ_0045881 level was significantly decreased in TNBC tissues and cells. Lentivirus-mediated hsa_circ_0045881 overexpression in MDA-MB-231 and BT-549 cells significantly reduced cell invasion and migration capacity. Additionally, hsa_circ_0045881 interacted with miR-214-3p in MDA-MB-231 cells. miR-214-3p mimics in MDA-MB-231 and BT-549 cells significantly enhanced cell invasion, migration and proliferation, but the other combinations of inhibitors had opposite effects on cell activity. CONCLUSIONS: Our data indicated that the circRNA has_circ_0045881 plays key roles in TNBC progression and that hsa_circ_0045881 might act as a sponge for miR-214-3p to modulate its levels in TNBC cells, thereby regulating cell invasion, metastasis and proliferation. hsa_circ_004588 might be a potential prognostic marker and therapeutic target for TNBC.

Exosomes from Hypoxia-treated Mesenchymal Stem Cells: Promoting Neuroprotection in Ischemic Stroke Through miR-214-3p/PTEN Mechanism.

Stroke stands as the second leading cause of death globally, surpassed only by ischemic heart disease. It accounts for 9% of total worldwide deaths. Given the swiftly evolving landscape, medical professionals and researchers are devoting increased attention to identifying more effective and safer treatments. Recent years have witnessed a focus on exosomes derived from mesenchymal stem cells cultivated under hypoxic conditions, referred to as Hypo-Exo. These specialized exosomes contain an abundance of components that facilitate the restoration of ischemic tissue, surpassing the content found in normal exosomes. Despite advancements, the precise role of Hypo-Exo in cases of cerebral ischemia remains enigmatic. Therefore, this study was designed to shed light on the potential efficacy of Hypo-Exo in stroke treatment. Our investigations unveiled promising outcomes, as the administration of Hypo-Exo led to improved behavioral deficits and reduced infarct areas in mice affected by ischemic conditions. Notably, these positive effects were hindered when Hypo-Exo loaded with anti-miR-214-3p were introduced, implying that the neuroprotective attributes of Hypo-Exo are reliant on miR-214-3p. This conclusion was substantiated by the high levels of miR-214-3p detected within Hypo-Exo. Furthermore, our examination of the ischemic penumbra zone revealed a gradual and sustained escalation in PTEN expression, a phenomenon effectively countered by Hypo-Exo treatment. Collectively, our findings suggest the existence of a regulatory pathway centered on miR-214-3p within Hypo-Exo. This pathway exerts a downregulating influence on the PTEN/Akt signaling pathway, thereby contributing to the amelioration of neurological function subsequent to ischemia-reperfusion events.