Melatonin Improves Oxidative Stress Injury in Retinopathy of Prematurity by Targeting miR-23a-3p/Nrf2.

PURPOSE: Melatonin has promising protective effects for retinopathy. However, its roles in retinopathy of prematurity (ROP) and the underlying mechanisms remain unknown. We aimed to explore its roles and mechanisms in a ROP model. METHODS: Hematoxylin and eosin staining were used to observe the morphology of the retina. Immunofluorescence was used to detect positive (Nrf2+ and VEGF+) cells. Immunohistochemistry was used to detect the level of nuclear expression of PCNA in retinal tissue. Transmission electron microscope (TEM) was used to observe the morphology and structure of pigment cells. qRT-PCR was used to assay the expression of miR-23a-3p, Nrf2, and HO-1. Western blotting was used to detect the expression of Nrf2, HO-1, ß-actin, and Lamin B1. RESULTS: Melatonin or miR-23a-3p antagomir treatment could ameliorate the Oxygen-induced pathological changes, increased the expression of Nrf2 and HO-1, SOD, and GSH-Px, and decreased the expression of VEGF, miR-23a-3p, MDA and the apoptosis in the ROP model. Further target prediction and luciferase reporter assays confirmed the targeted binding relationship between miR-23a-3p and Nrf2. CONCLUSION: Our study showed that melatonin could ameliorate H2O2-induced apoptosis and oxidative stress injury in RGC cells by mediating miR-23a-3p/Nrf2 signaling pathway, thereby improving retinal degeneration.

Unveiling the Regulatory Role of LncRNA MYU in Hypoxia-Induced Angiogenesis via the miR-23a-3p Axis in Endothelial Cells.

Background: Angiogenesis is essential for various physiological and pathological processes, such as embryonic development and cancer cell proliferation, migration, and invasion. Long noncoding RNAs (lncRNAs) play pivotal roles in normal homeostasis and disease processes by regulating gene expression through various mechanisms, including competing endogenous RNAs (ceRNAs) of target microRNAs (miRNAs). The lncRNA MYU is known to promote prostate cancer proliferation via the miR-184/c-Myc regulatory axis and to be upregulated in vascular endothelial cells under hypoxic conditions, which often occurs in solid tumors. In the present study, we investigated whether MYU might affect cancer growth by regulating angiogenesis in vascular endothelial cells under hypoxia. Methods: The expression of MYU-regulated miR-23a-3p and interleukin-8 (IL-8) in HUVEC cell lines was examined using qRT-PCR. The CCK-8 assay, EdU assay, wound-healing assay, and tube-formation assay were used to assess the effects of MYU on cell proliferation, migration, and tube formation of HUVEC cells in vitro. The dual-luciferase reporter assay was performed to examine the effects of miR-23a-3p on MYU and IL-8 expression. Results: We found that the overexpression of MYU and knockdown of miR-23a-3p in human umbilical vein endothelial cells (HUVECs) under hypoxia promoted cell proliferation, migration, and tube formation. Mechanistically, MYU was shown to bind competitively to miR-23a-3p, thereby preventing miR-23a-3p binding to the 3' untranslated region of IL-8 mRNA. In turn, increased production of pro-angiogenic IL-8 promoted HUVEC proliferation, migration, and tube formation under hypoxia. Conclusion: This study identified a new role for lncRNA MYU as a ceRNA for miR-23a-3p and uncovered a novel MYU-miR-23a-3p-IL-8 regulatory axis for angiogenesis. MYU and/or miR-23a-3p may thus represent new targets for the treatment of hypoxia-related diseases by promoting angiogenesis.

Small extracellular vesicles-shuttled miR-23a-3p from mesenchymal stem cells alleviate renal fibrosis and inflammation by inhibiting KLF3/STAT3 axis in diabetic kidney disease.

Human umbilical cord mesenchymal stem cells-derived small extracellular vesicles (MSC-sEV) provide a pragmatic solution as a cell-free therapy for patients with diabetic kidney disease (DKD). However, the underlying protective mechanisms of MSC-sEV remain largely unknown in DKD. Invivo and in vitro analyses demonstrated that MSC-sEV attenuated renal fibrosis and inflammation of DKD. The underlying mechanism of the MSC-sEV-induced therapeutic effect was explored by high-throughput sequencing, which identified the unique enrichment of a set of miRNAs in MSC-sEV compared with human skin fibroblasts-sEV (HSF-sEV). Vitro experiments demonstrated that the protective potential was primarily attributed to miR-23a-3p, one of the most abundant miRNAs in MSC-sEV. Further, overexpression or knockdown analyses revealed that miR-23a-3p, and its target Krüppel-like factor 3 (KLF3) suppressed the STAT3 signaling pathway in high glucose (HG) induced HK-2 cells were essential for the renal-protective property of MSC-sEV. Moreover, we found that miR-23a-3p was packaged into MSC-sEV by RNA Binding Motif Protein X-Linked (RBMX) and transmitted to HG-induced HK-2 cells. Finally, inhibiting miR-23a-3p could mitigate the protective effects of MSC-sEV in db/db mice. These findings suggest that a systemic administration of sEV derived from MSC, have the capacity to incorporate into kidney where they can exert renal-protective potential against HG-induced injury through delivery of miR-23a-3p.

Circular RNA Circ_0122396 Regulates Human Lens Epithelial Cell Progression by Regulating miR-23a-3p and MMP16 in Age-Related Cataract.

BACKGROUND: CircRNA plays a regulatory role in multiple life processes. Circ_0122396 could participate in the regulation of age-related cataract (ARC) progression. However, the precise molecular mechanisms of circ_0122396 In ARC remain enigmatic. METHODS: Circ_0122396, microRNA (miR)-23a-3p, and matrix metalloprotease (MMP)-16 (MMP16) expression levels were detected via quantitative real-time polymerase chain reaction. Western blot was used to detect the levels of MMP16 and apoptosis-related proteins. Cell counting kit-8 analysis and 5-ethynyl-2'-deoxyuridine assay were used to assess human lens epithelial cells (HLECs) proliferation. Flow cytometry was performed to determine cell apoptosis. Levels of malondialdehyde (MDA) and glutathione peroxidase (GSH-PX) were measured using commercial kits. Luciferase reporter assay, RNA immunoprecipitation (RIP) assay, and RNA pull-down assay were used to examine the interaction among circ_0122396, miR-23a-3p, and MMP16. RESULTS: Circ_0122396 and MMP16 were down-regulated while miR-23a-3p was up-regulated in ARC. H2O2 constrained proliferation and GSH-PX level, promotes apoptosis and MDA level in HLECs, and overexpression of circ_0122396 attenuated these effects. miR-23a-3p was a direct target of circ_0122396, and MMP16 was a direct target of miR-23a-3p. The effect of circ_0122396 overexpression on H2O2-induced HLECs was reversed by miR-23a-3p, and MMP16 elevation overturned the impacts of miR-23a-3p in H2O2-induced HLECs. CONCLUSIONS: Circ_0122396 may regulate the progression of ARC via the miR-23a-3p/MMP16 pathway in H2O2-stimulated HLECs, which may serve as a potentially valuable biomarker and novel therapeutic target for ARC.

Unraveling IGFBP3-mediated m6A modification in fracture healing.

BACKGROUND: This study investigates the role of IGFBP3-mediated m6A modification in regulating the miR-23a-3p/SMAD5 axis and its impact on fracture healing, aiming to provide insights into potential therapeutic targets. METHODS: Utilizing fracture-related datasets, we identified m6A modification-related mRNA and predicted miR-23a-3p as a regulator of SMAD5. We established a mouse fracture healing model and conducted experiments, including Micro-CT, RT-qPCR, Alizarin Red staining, and Alkaline phosphatase (ALP) staining, to assess gene expression and osteogenic differentiation. RESULTS: IGFBP3 emerged as a crucial player in fracture healing, stabilizing miR-23a-3p through m6A modification, leading to SMAD5 downregulation. This, in turn, inhibited osteogenic differentiation and delayed fracture healing. Inhibition of IGFBP3 partially reversed through SMAD5 inhibition, restoring osteogenic differentiation and fracture healing in vivo. CONCLUSION: The IGFBP3/miR-23a-3p/SMAD5 axis plays a pivotal role in fracture healing, highlighting the relevance of m6A modification. IGFBP3's role in stabilizing miR-23a-3p expression through m6A modification offers a potential therapeutic target for enhancing fracture healing outcomes.

Naringenin regulates cigarette smoke extract-induced extracellular vesicles from alveolar macrophage to attenuate the mouse lung epithelial ferroptosis through activating EV miR-23a-3p/ACSL4 axis.

BACKGROUND: Alveolar macrophages are one of the momentous regulators in pulmonary inflammatory responses, which can secrete extracellular vesicles (EVs) packing miRNAs. Ferroptosis, an iron-dependent cell death, is associated with cigarette smoke-induced lung injury, and EVs have been reported to regulate ferroptosis by transporting intracellular iron. However, the regulatory mechanism of alveolar macrophage-derived EVs has not been clearly illuminated in smoking-related pulmonary ferroptosis. Despite the known anti-ferroptosis effects of naringenin in lung injury, whether naringenin controls EVs-mediated ferroptosis has not yet been explored. PURPOSE: We explore the effects of EVs from cigarette smoke-stimulated alveolar macrophages in lung epithelial ferroptosis, and elucidate the EV miRNA-mediated pharmacological mechanism of naringenin. STUDY DESIGN AND METHODS: Differential and ultracentrifugation were conducted to extract EVs from different alveolar macrophages treatment groups in vitro. Both intratracheal instilled mice and treated epithelial cells were used to investigate the roles of EVs from alveolar macrophages involved in ferroptosis. Small RNA sequencing analysis was performed to distinguish altered miRNAs in EVs. The ferroptotic effects of EV miRNAs were examined by applying dual-Luciferase reporter assay and miRNA inhibitor transfection experiment. RESULTS: Here, we firstly reported that EVs from cigarette smoke extract-induced alveolar macrophages (CSE-EVs) provoked pulmonary epithelial ferroptosis. The ferroptosis inhibitor ferrostatin-1 treatment reversed these changes in vitro. Moreover, EVs from naringenin and CSE co-treated alveolar macrophages (CSE+Naringenin-EVs) markedly attenuated the lung epithelial ferroptosis compared with CSE-EVs. Notably, we identified miR-23a-3p as the most dramatically changed miRNA among Normal-EVs, CSE-EVs, and CSE+Naringenin-EVs. Further experimental investigation showed that ACSL4, a pro-ferroptotic gene leading to lipid peroxidation, was negatively regulated by miR-23a-3p. The inhibition of miR-23a-3p diminished the efficacy of CSE+Naringenin-EVs. CONCLUSION: Our findings firstly provided evidence that naringenin elevated the EV miR-23a-3p level from CSE-induced alveolar macrophages, thereby inhibiting the mouse lung epithelial ferroptosis via targeting ACSL4, and further complemented the mechanism of cigarette-induced lung injury and the protection of naringenin in a paracrine manner. The administration of miR-23a-3p-enriched EVs has the potential to ameliorate pulmonary ferroptosis.

Exosomal miR-23a-3p derived from human umbilical cord mesenchymal stem cells promotes remyelination in central nervous system demyelinating diseases by targeting Tbr1/Wnt pathway.

Oligodendrocyte precursor cells are present in the adult central nervous system, and their impaired ability to differentiate into myelinating oligodendrocytes can lead to demyelination in patients with multiple sclerosis, accompanied by neurological deficits and cognitive impairment. Exosomes, small vesicles released by cells, are known to facilitate intercellular communication by carrying bioactive molecules. In this study, we utilized exosomes derived from human umbilical cord mesenchymal stem cells (HUMSCs-Exos). We performed sequencing and bioinformatics analysis of exosome-treated cells to demonstrate that HUMSCs-Exos can stimulate myelin gene expression in oigodendrocyte precursor cells. Functional investigations revealed that HUMSCs-Exos activate the Pi3k/Akt pathway and regulate the Tbr1/Wnt signaling molecules through the transfer of miR-23a-3p, promoting oligodendrocytes differentiation and enhancing the expression of myelin-related proteins. In an experimental autoimmune encephalomyelitis model, treatment with HUMSCs-Exos significantly improved neurological function and facilitated remyelination. This study provides cellular and molecular insights into the use of cell-free exosome therapy for central nervous system demyelination associated with multiple sclerosis, demonstrating its great potential for treating demyelinating and neurodegenerative diseases.

Long non-coding RNA FKSG29 regulates oxidative stress and endothelial dysfunction in obstructive sleep apnea.

Altered expressions of pro-/anti-oxidant genes are known to regulate the pathophysiology of obstructive sleep apnea (OSA).We aim to explore the role of a novel long non-coding (lnc) RNA FKSG29 in the development of intermittent hypoxia with re-oxygenation (IHR)-induced endothelial dysfunction in OSA. Gene expression levels of key pro-/anti-oxidant genes, vasoactive genes, and the FKSG29 were measured in peripheral blood mononuclear cells from 12 subjects with primary snoring (PS) and 36 OSA patients. Human monocytic THP-1 cells and human umbilical vein endothelial cells (HUVEC) were used for gene knockout and double luciferase under IHR exposure. Gene expression levels of the FKSG29 lncRNA, NOX2, NOX5, and VEGFA genes were increased in OSA patients versus PS subjects, while SOD2 and VEGFB gene expressions were decreased. Subgroup analysis showed that gene expression of the miR-23a-3p, an endogenous competitive microRNA of the FKSG29, was decreased in sleep-disordered breathing patients with hypertension versus those without hypertension. In vitro IHR experiments showed that knock-down of the FKSG29 reversed IHR-induced ROS overt production, early apoptosis, up-regulations of the HIF1A/HIF2A/NOX2/NOX4/NOX5/VEGFA/VEGFB genes, and down-regulations of the VEGFB/SOD2 genes, while the protective effects of FKSG29 knock-down were abolished by miR-23a-3p knock-down. Dual-luciferase reporter assays confirmed that FKSG29 was a sponge of miR-23a-3p, which regulated IL6R directly. Immunofluorescence stain further demonstrated that FKSGH29 knock-down decreased IHR-induced uptake of oxidized low density lipoprotein and reversed IHR-induced IL6R/STAT3/GATA6/ICAM1/VCAM1 up-regulations. The findings indicate that the combined RNA interference may be a novel therapy for OSA-related endothelial dysfunction via regulating pro-/anti-oxidant imbalance or targeting miR-23a-IL6R-ICAM1/VCAM1 signaling.

M2 macrophage-secreted exosomes promote metastasis and increase vascular permeability in hepatocellular carcinoma.

BACKGROUND: Metastasis is a key feature of malignant tumors and significantly contributes to their high mortality, particularly in hepatocellular carcinoma (HCC). Therefore, it is imperative to explore the mechanism of tumor metastasis. Recently, tumor-associated macrophages (TAMs) have been demonstrated to promote tumor progression, while TAM-derived molecules involved in HCC metastasis warrant further investigation. METHODS: THP-1 was treated with IL-4 (Interleukin-4) and IL-13 (Interleukin-13) for M2 polarized macrophages. Exosomes derived from M2 macrophages were characterized. Then, HCC cells or human umbilical vein endothelial cells (HUVECs) were co-cultured with M2 macrophages or treated with M2 macrophage-secreted exosomes. Next, Transwell®, Scratch assay, tube formation, and endothelial permeability assays were performed. Moreover, RT-PCR, western blotting, immunofluorescence, and ELISA were used to assess mRNA and protein expression levels. Finally, the miRNA expression profiles of exosomes derived from M2 and M0 macrophages were analyzed. RESULTS: M2 macrophage infiltration was correlated with metastasis and a poor prognosis in HCC patients. M2-derived exosomes were absorbed by HCC and HUVEC cells and promoted the epithelial-mesenchymal transition (EMT), vascular permeability, and angiogenesis. Notably, MiR-23a-3p levels were significantly higher in M2-derived exosomes and hnRNPA1 mediated miR-23a-3p packaging into exosomes. Phosphatase and tensin homolog (PTEN) and tight junction protein 1 (TJP1) were the targets of miR-23a-3p, as confirmed by luciferase reporter assays. Lastly, HCC cells co-cultured with M2-derived exosomes secreted more GM-CSF, VEGF, G-CSF, MCP-1, and IL-4, which in turn further recruited M2 macrophages. CONCLUSIONS: Our findings suggest that M2 macrophage-derived miR-23a-3p enhances HCC metastasis by promoting EMT and angiogenesis, as well as increasing vascular permeability. Video Abstract.

MiR-23a-3p alleviates cigarette smoke extract-induced pulmonary vascular endothelial cell apoptosis by targeting DNAJB1 in emphysema.

BACKGROUND: Cigarette smoke (CS) is an important risk factor for chronic obstructive pulmonary disease, including emphysema. MicroRNAs (miRNAs) are important regulators of emphysema progression. However, miR-23a-3p role in emphysema is unclear. METHODS: CS exposure was used to construct emphysema mice models, and cigarette smoke extract (CSE)-induced pulmonary vascular endothelial cells (PMVECs) were used to mimic emphysema cell models. Mouse lung tissue was stained by immunohistochemical staining, hematoxylin and eosin staining, and TUNEL staining. MiR-23a-3p and DnaJ homolog subfamily B member 1 (DNAJB1) levels were tested using quantitative real-time PCR. DNAJB1 and apoptosis-related markers' protein levels were examined via western blot analysis. Cell viability and apoptosis were analyzed by MTT assay and flow cytometry. The interaction between miR-23a-3p and DNAJB1 was evaluated by dual-luciferase reporter assay and RIP assay. RESULTS: MiR-23a-3p was downregulated, and DNAJB1 was upregulated in CS-induced emphysema mice models and CSE-induced PMVECs. MiR-23a-3p overexpression promoted viability and repressed apoptosis in CSE-induced PMVECs. MiR-23a-3p targeted DNAJB1 and negatively regulated DNAJB1 expression. Moreover, DNAJB1 knockdown repressed CSE-induced PMVECs apoptosis, and miR-23a-3p inhibitor reversed this effect. Additionally, miR-23a-3p alleviated lung tissue injury and improved emphysema in mice by reducing DNAJB1 expression. CONCLUSION: MiR-23a-3p alleviated emphysema progression, which could inhibit CSE-induced PMVECs apoptosis by targeting DNAJB1.

Clinical value of miR-23a-3p expression in early diagnosis of diabetic kidney disease.

INTRODUCTION: The objective was to observe the expression of miR-23a-3p in the serum of patients with type 2 diabetic nephropathy (T2DN) and to explore its clinical significance. MATERIALS AND METHODS: 112 patients with type 2 diabetes were divided into a simple diabetes mellitus (NON) group, T2DN microalbuminuria (MIC) group, and T2DN macroalbuminuria (MAC) group, according to the urinary protein-creatinine ratio (uACR). Clinical data were collected, miR-23a-3p levels in serum were measured by quantitative reverse transcription polymerase chain reaction (qRT-PCR), and clinical parameters were measured by an automatic biochemical analyser; the influencing factors of diabetic kidney disease (DKD) and the correlation between miR-23a-3p expression and clinical parameters were analysed. RESULTS: The expression of miR-23a-3p in the serum of the DKD group was lower than that of the normal control (CON) and NON groups. Correlation analysis showed that miR-23a-3p was positively correlated with urinary albumin (Albu), glycosylated haemoglobin (HbA1c), total cholesterol (CHOL), glycated albumin (GA-L), serum creatinine (Scr), fasting blood glucose (GLU), and uric acid (UA), negatively correlated with uACR and high-density lipoprotein cholesterol (HDL-C), but not correlated with urinary creatinine (CREA). The area under the receiver operating characteristic (ROC) curve (AUC) of miR-23a-3p for the diagnosis of DKD was 0.686 [95% confidence interval (CI): 0.599-0.773], with a sensitivity of 64.5% and a specificity of 71.2%; the AUC for differentiating NON from DKD was 0.700 (95% CI: 0.598-0.802), with a sensitivity of 61.8% and a specificity of 77.8%. Multivariate logistic regression analysis showed that serum miR-23a-3p levels were not associated with the development of DKD after adjusting for other levels of influence and were not significant for the differentiation of NON and DKD. CONCLUSION: Serum miR-23a-3p levels are decreased in T2DN patients, and this change becomes more significant with the severity of the disease, which may be a marker for the early diagnosis and progression of T2DN.

M2 Macrophages-Derived Exosomal miRNA-23a-3p Promotes the Progression of Oral Squamous Cell Carcinoma by Targeting PTEN.

Exosomes from tumor cells and immune cells regulate the tumor microenvironment through the biomolecules or microRNAs (miRNAs) they carry. This research aims to investigate the role of miRNA in exosomes derived from tumor-associated macrophages (TAMs) in the progression of oral squamous cell carcinoma (OSCC). RT-qPCR and Western blotting assays were used to determine the expression of genes and proteins in OSCC cells. CCK-8, Scratch assay and invasion-related proteins were utilized to detect the malignant progression of tumor cells. High-throughput sequencing predicted differentially expressed miRNAs in exosomes secreted by M0 and M2 macrophages. Compared with exosomes from M0 macrophages, exosomes from M2 macrophages led to enhanced proliferation and invasion of OSCC cells and inhibited their apoptosis. High-throughput sequencing results show that miR-23a-3p is differentially expressed in exosomes from M0 and M2 macrophages. MiRNA target gene database predicts that phosphatase and tensin homolog (PTEN) are target genes of miR-23a-3p. Further studies revealed that transfection of miR-23a-3p mimics inhibited PTEN expression in vivo and in vitro and promoted the malignant progression of OSCC cells, which was reversed by miR-23a-3p inhibitors. MiR-23a-3p in exosomes derived from M2 macrophages promotes malignant progression of OSCC. PTEN is a potential intracellular target of miR-23a-3p. MiR-23a-3p, an M2 macrophage-associated exosome, is a promising target for the future treatment of OSCC.

Circular RNA hsa_circ_0007444 inhibits ovarian cancer progression through miR-23a-3p/DICER1 axis.

Ovarian cancer is the second leading cause of death in women with gynecological malignancy in China. Circular RNAs are a class of noncoding regulatory RNAs reported to be involved in cancer development and progression. Previous studies, including our own, have indicated that hsa_circ_0007444 is downregulated in ovarian cancer tissues. This study aims to elucidate the function and mechanism of hsa_circ_0007444 in ovarian cancer progression. The expression of hsa_circ_0007444 is determined by quantitative real-time PCR (qRT-PCR). Cell proliferation, invasion, migration and apoptosis are examined by cell counting-kit 8 (CCK-8), transwell and flow cytometry assays. Tumor growth and metastasis are assessed in vivo using Balb/c nude mouse xenograft model and tail vein injection model. And the mechanism of action of hsa_circ_0007444 is analysed by RNA-binding protein immunoprecipitation (RIP), luciferase reporter and rescue assays. hsa_circ_0007444 is downregulated in ovarian cancer tissues and cell lines compared with that in normal ovarian tissues and normal epithelial cell line. Gain- and loss-of-function results indicate that hsa_circ_0007444 inhibits cell proliferation, invasion, migration and increases cell apoptosis of ovarian cancer cells in vitro, and inhibits tumor growth and lung metastasis in vivo. Mechanistically, hsa_circ_0007444 can interact with AGO2 and sponge miR-23a-3p, thereby upregulating DICER1 expression, which is an important tumor suppressor in ovarian cancer. And miR-23a-3p mimics can rescue the inhibitory effect of hsa_circ_0007444 on ovarian cancer cell proliferation, invasion and migration. Therefore, hsa_circ_0007444 can inhibit ovarian cancer progression through the hsa_circ_0007444/miR-23a-3p/DICER1 axis.

ATP2B1-AS1 exacerbates sepsis-induced cell apoptosis and inflammation by regulating miR-23a-3p/TLR4 axis.

BACKGROUND: Sepsis is a life-threatening disease with dominant mortality. Its early diagnosis and treatment can improve prognosis and reduce mortality. Long noncoding RNAs (lncRNAs) ATPase plasma membrane Ca2+ transporting 1 antisense RNA 1 (ATP2B1-AS1) is dysregulated and is involved in the progression of various diseases. Nevertheless, the role of ATP2B1-AS1 in sepsis remains unclear. METHODS: A human monocytic cell line, THP-1 cells, was stimulated to induce a model of sepsis in vitro. The levels of ATP2B1-AS1, miR-23a-3p, and TLR4 were assessed by real-time quantitative polymerase chain reaction. The role of ATP2B1-AS1 in cell apoptosis and inflammation was explored by flow cytometry, Western blot analysis and enzyme-linked immunosorbent serologic assay. The binding sites between ATP2B1-AS1 and miR-23a-3p, and between miR-23a-3p and TLR4 were predicted by BiBiServ and the Encyclopedia of RNA Interactomes (ENCORI) online sites, respectively, and confirmed by the luciferase assay. RESULTS: The level of ATP2B1-AS1 was increased in lipopolysaccharide (LPS)-treated THP-1 cells. LPS increased apoptosis ratio, relative protein expressions of pro-apoptotic factors, and relative messenger RNA (mRNA) level and concentrations of pro-inflammatory cytokines, but decreased the relative expression of anti-apoptosis protein and relative mRNA level and concentrations of anti-inflammatory factor. All these alterations were reversed with transfection of shATP2B1-AS1 into THP-1 cells. Moreover, ATP2B1-AS1 directly bound miR-23a-3p and negatively modulated the level of miR-23a-3p. Meanwhile, TLR4 was directly targeted by miR-23a-3p, and negatively and positively modulated by miR-23a-3p and ATP2B1-AS1, respectively. CONCLUSION: ATP2B1-AS1 aggravated apoptosis and inflammation by modulating miR-23a-3p/TLR4 axis in LPS-treated THP-1 cells.

ATP2B1-AS1 exacerbates sepsis-induced cell apoptosis and inflammation by regulating miR-23a-3p/TLR4 axis

Background: Sepsis is a life-threatening disease with dominant mortality. Its early diagnosis and treatment can improve prognosis and reduce mortality. Long noncoding RNAs (lncRNAs) ATPase plasma membrane Ca2+ transporting 1 antisense RNA 1 (ATP2B1-AS1) is dysregulated and is involved in the progression of various diseases. Nevertheless, the role of ATP2B1-AS1 in sepsis remains unclear. Methods: A human monocytic cell line, THP-1 cells, was stimulated to induce a model of sepsis in vitro. The levels of ATP2B1-AS1, miR-23a-3p, and TLR4 were assessed by real-time quantitative polymerase chain reaction. The role of ATP2B1-AS1 in cell apoptosis and inflammation was explored by flow cytometry, Western blot analysis and enzyme-linked immunosorbent serologic assay. The binding sites between ATP2B1-AS1 and miR-23a-3p, and between miR-23a-3p and TLR4 were predicted by BiBiServ and the Encyclopedia of RNA Interactomes (ENCORI) online sites, respectively, and confirmed by the luciferase assay. Results: The level of ATP2B1-AS1 was increased in lipopolysaccharide (LPS)-treated THP-1 cells. LPS increased apoptosis ratio, relative protein expressions of pro-apoptotic factors, and relative messenger RNA (mRNA) level and concentrations of pro-inflammatory cytokines, but decreased the relative expression of anti-apoptosis protein and relative mRNA level and concentrations of anti-inflammatory factor. All these alterations were reversed with transfection of shATP2B1-AS1 into THP-1 cells. Moreover, ATP2B1-AS1 directly bound miR-23a-3p and negatively modulated the level of miR-23a-3p. Meanwhile, TLR4 was directly targeted by miR-23a-3p, and negatively and positively modulated by miR-23a-3p and ATP2B1-AS1, respectively. Conclusion: ATP2B1-AS1 aggravated apoptosis and inflammation by modulating miR-23a-3p/TLR4 axis in LPS-treated THP-1 cells (AU)

miR-23a-3p promotes the development of colon cancer by inhibiting the expression of NDRG4.

PURPOSE: Previous studies have found that miR-23a-3p, a diagnostic marker for colon cancer (CC), is upregulated in primary CC from stage I/II patients. Nevertheless, the specific functions and molecular mechanisms of miR-23a-3p in colon cancer remain unclear. METHODS: The expression levels of miR-23a-3p and NDRG4 were analyzed by western blot and RT‒qPCR assays. Cell viability and proliferation were measured by CCK8 and colony formation assays. Cell apoptosis was assessed by flow cytometry. Cell migration and invasion were detected by transwell assay. Target binding was detected by luciferase reporter assay. RESULTS: miR-23a-3p was dramatically elevated in CC tissues and cells. In HT29 and SW480 cells, downregulation of miR-23a-3p hampered cell proliferation, migration, and invasion while increasing cell apoptosis. The effects of miR-23a-3p silencing on CC progression were slowed by NDRG4 downregulation. CONCLUSIONS: miR-23a-3p promoted CC progression by modulating the expression of NDRG4. This study demonstrated the mechanism of miR-23a-3p in CC, which may offer a new target for CC therapy.

Effects of Ligusticum Chuanxiong extract on MPP+-induced SH-SY5Y cells in Parkinson’s syndrome model via miR-23a-3p/SNCA axis / 西安交通大学学报(医学版)

【Objective】 To explore the effects of Ligusticum Chuanxiong extract on MPP+-induced SH-SY5Y cell damage and Parkinson’s syndrome. 【Methods】 1-methyl-4phenylpyridine ion (MPP+) interfered with SH-SY5Y to establish a cell model of elderly Parkinson’s syndrome (SH-SY5Y-MPP+). After intervention with Ligusticum Chuanxiong extract, cell proliferation and apoptosis as well as miR-23a-3p and SNCA expressions were detected. In addition, the changes of SH-SY5Y-MPP+ after regulating the expression of miR-23a-3p and SNCA were observed, and the relationship between miR-23a-3p and SNCA was verified by dual luciferase reporter. 【Results】 The cell proliferation capacity of SH-SY5y-MPP+ was significantly lower than that of SH-SY5Y, while the apoptosis rate was higher than that of SH-SY5Y (P<0.05). Under the intervention of Ligusticum Chuanxiong extract, the proliferation ability of SH-SY5Y-MPP+, and Bcl-2 and SNCA protein increased, the apoptosis rate, miR-23a-3p, and Bax proteins decreased (P<0.05). Both silencing miR-23a-3p and increasing SNCA could promote the proliferation of SH-SY5Y-MPP+ and inhibit apoptosis, while increasing miR-23a-3p and silencing SNCA were the opposite (P<0.05). The online target gene prediction website found that miR-23a-3p and SNCA had complementary sites that could bind, and the dual luciferase reporter enzyme showed that the firefly activity of SNCA-wt was significantly inhibited after the miR-23a-3p mimic sequence was transfected (P<0.05). After increasing miR-23a-3p, the expression of SNCA protein in SH-SY5Y-MPP+ decreased, while silencing miR-23a-3p was the opposite (P<0.05). Rescue experiments showed that the intervention effect of Ligusticum Chuanxiong extract on SH-SY5Y-MPP+ was completely reversed by increasing miR-23a-3p or silent SNCA (P>0.05); the effect of increasing miR-23a-3p on SH-SY5Y-MPP+ increased SNCA reversion (P>0.05). 【Conclusion】 Ligusticum Chuanxiong extract can affect the biological behavior changes of SH-SY5Y induced by MPP+ by regulating the miR-23a-3p/SNCA axis, which may be a new direction for the treatment of elderly Parkinson’s syndrome in the future.

Small Extracellular Vesicles Released from Bioglass/Hydrogel Scaffold Promote Vascularized Bone Regeneration by Transferring miR-23a-3p.

Background: The treatment of critical-size bone defect is a great difficulty in orthopedics. Osteogenesis and angiogenesis are critical issue during the process of bone repair and remodeling. Mesenchymal stem cells (MSCs)-derived exosomes have the same therapeutic effect to MSCs-based therapies. The effect of human umbilical cord MSCs-derived sEVs (hUC-MSCs-sEVs) on vascularized bone regeneration and the potential mechanism remains to be investigated. Herein, we aimed to explore the therapeutic effect and the mechanism of hUC-MSCs-sEVs on critical-size bone defect. Methods: To investigate the potential osteogenesis and angiogenesis effects of sEVs in vitro, we extracted sEVs from hUC-MSCs, and then sEVs were co-incubated with BMSCs and HUVECs. We next investigated the effect and potential mechanism of sEVs on the effects of osteogenesis and angiogenesis. We fabricated 3D-printed bioglass scaffold with Gelma/nanoclay hydrogel coatings to load sEVs (BG-gel-sEVs) to ensure in vivo sustained efficacy of sEVs. Finally, the skull defect model was used to evaluate the capacity of vascularized bone regeneration of the composited scaffolds. Results: hUC-MSCs-sEVs facilitated calcium deposition and the endothelial network formation, inducing osteogenic differentiation and angiogenesis by delivering miR-23a-3p to activate PTEN/AKT signaling pathway. Additionally, the BG-gel-sEVs composited scaffold achieved vascularized bone regeneration in vivo. Conclusion: This finding illuminated that hUC-MSCs-sEVs promoted osteogenesis and angiogenesis by delivering miR-23a-3p to activate PTEN/AKT signaling pathway, achieving vascularized bone regeneration.

Circ_0005050 promotes the proliferation of oral squamous cell carcinoma and inhibits the apoptosis by activating JAK/STAT3 signaling pathway.

Oral squamous cell carcinoma (OSCC) is the most predominant type of oral cancer, featured with poor prognosis and high mortality. Circular RNA (circRNA) exerts its function in a variety of human cancers, including OSCC. Circ_0005050, as a novel circRNA, has not been well explored in OSCC so far. This study centered on investigating the impact of circ_0005050 on OSCC cell growth and its molecular mechanism. RNA or protein expression was detected by RT-qPCR or western blot analysis. Functional assays were employed to uncover the changes of OSCC cell biological behaviors. Mechanistic assays were done to verify the underlying mechanism of circ_0005050 in OSCC cells. According to the collected data, circ_0005050 was significantly up-regulated in OSCC cells compared to normal cells. Circ_0005050 depletion hampered proliferative ability of OSCC cells while promoting cell apoptotic ability. As for mechanism analyses, circ_0005050 knockdown led to the reduction of STAT3 expression and JAK/STAT3 signaling pathway activity. Moreover, circ_0005050 competitively bound to miR-23a-3p and miR-625-5p to up-regulate STAT3, thus prompting malignant behaviors of OSCC cells. In conclusion, circ_0005050 regulates miR-23a-3p/miR-625-5p/STAT3 axis to activate JAK/STAT3 signaling pathway, consequently facilitating OSCC cell proliferation and inhibiting cell apoptosis.

Jianpi Huayu decoction inhibits the epithelial-mesenchymal transition of hepatocellular carcinoma cells by suppressing exosomal miR-23a-3p/Smad signaling.

ETHNOPHARMACOLOGICAL RELEVANCE: Jianpi Huayu decoction (JHD) is a traditional Chinese medicinal preparation used to treat a variety of malignant tumors including HCC, although the underlying mechanism remains unknown. Exosomes in the tumor microenvironment mediate intercellular signaling among cancer cells, but precise contributions to hepatocellular carcinoma (HCC) progression are still elusive. AIM OF THE STUDY: In this work, the main objective was to examine the mechanisms underlying anti-tumor effects of JHD and the potential contributions of exosomal signaling. MATERIALS AND METHODS: LC-MS/MS was used for quality control of JDH preparation, while nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM) and western blotting were used for verification of exosomes. In vitro assays included CCK8, wound healing assay, transwell invasion assay, qRT-PCR and western blotting were performed to investigate the effects of JHD on HCC cells and the molecular mechanism. Furthermore, the effects of JHD on subcutaneous tumor model of nude mice were also determined. RESULTS: JHD inhibited the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of cultured HCC cells. Further, exosomes isolated from EMT-induced HCC cells promoted the migration, invasion and EMT of other cultured HCC cells, while exosomes isolated from EMT-induced HCC cells after JHD treatment had little effect. In addition, JHD reduced the expression of exosomal miR-23a-3p in cultured HCC cells. miR-23a-3p was significantly up-regulated in tumor compared with that in adjacent non-cancerous tissues of patients with HCC. HCC patients with high miR-23a-3p expression had poor overall survival after hepatectomy. Meanwhile, miR-23a-3p enhanced HCC cell proliferation, EMT, and expression of Smad signaling proteins. More importantly, overexpression of miR-23a-3p can reverse the inhibition of EMT and Smad signaling pathway caused by JHD treatment. In vivo assays, treatment with JHD also reduced the growth of HCC-derived tumors in nude mice, reduced the expression of miR-23a-3p in serum exosomes and the level of EMT in tumor cells. CONCLUSIONS: the antitumor effects of JHD on HCC are mediated at least in part by inhibition of EMT due to downregulation of exosome-mediated intercellular miR-23a-3p transfer and subsequent blockade of Smad signaling. Disrupting this exosomal miR-23a-3p/Smad signaling pathway may be an effective treatment.