Pulsed Electromagnetic Fields Combined With Adipose-Derived Stem Cells Protect Ischemic Myocardium by Regulating miR-20a-5p/E2F1/p73 Signaling.

Myocardial infarction (MI) is a serious threat to human health. Although monotherapy with pulsed electromagnetic fields (PEMFs) or adipose-derived stem cells (ADSCs) has been reported to have positive effect on the treatment of MI, a satisfactory outcome has not yet been achieved. In recent years, combination therapy has attracted widespread interest. Herein, we explored the synergistic therapeutic effect of combination therapy with PEMFs and ADSCs on MI and found that the combination of PEMFs and ADSCs effectively reduced infarct size, inhibited cardiomyocyte apoptosis and protected the cardiac function in mice with MI. In addition, bioinformatics analysis and RT-qPCR showed that the combination therapy could affect apoptosis by regulating the expression of miR-20a-5p. A dual-luciferase reporter gene assay also confirmed that the miR-20a-5p could target E2F transcription factor 1 (E2F1) and inhibit cardiomyocyte apoptosis by regulating the E2F1/p73 signaling pathway. Therefore, our study systematically demonstrated the effectiveness of combination therapy on the inhibition of cardiomyocyte apoptosis by regulating the miR-20a-5p/E2F1/p73 signaling pathway in mice with MI. Thus, our study underscored the effectiveness of the combination of PEMFs and ADSCs and identified miR-20a-5p as a promising therapeutic target for the treatment of MI in the future.

Regulatory mechanism of miR-20a-5p in neuronal damage and inflammation in lipopolysaccharide-induced BV2 cells and MPTP-HCl-induced Parkinson's disease mice.

BACKGROUND: Parkinson's disease (PD) is a prevailing neurodegenerative disorder increasingly affecting the elderly population. The involvement of microRNAs (miRNAs) in PD has been confirmed. We sought to explore the molecular mechanism of miR-20a-5p in PD. METHODS: Lipopolysaccharide (LPS)-induced BV2 cell model and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP-HCl)-induced PD mouse model were established. miR-20a-5p, inducible nitric oxide synthase (iNOS), interleukin (IL)-6, tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-ß1, and IL-10 expression in BV2 cells was examined by reverse transcription - quantitative polymerase chain reaction. Cell viability was assessed by MTT assay. The apoptotic rate and levels of Bcl-2, Bax, cleaved caspase-3, and signal transducer and activator of transmission (STAT)3 were examined by flow cytometry and Western blot. Bioinformatics software predicted the potential binding sites of miR-20a-5p and STAT3. Dual-luciferase experiment verified the binding relationship. Iba1-positive and tyrosine hydroxylase (TH)-positive cell numbers in substantia nigra pars compacta were detected by immunohistochemistry. The effect of miR-20a-5p on motor function in MPTP-induced PD mice was detected by Rota-rod test, Pole test, Traction test and Beam-crossing task. RESULTS: miR-20a-5p was under-expressed in LPS-induced BV2 cells. Overexpression of miR-20a-5p increased the viability of LPS-induced BV2 cells and reduced apoptosis rates. Moreover, overexpression of miR-20a-5p reduced cleaved caspase-3, Bax, iNOS, IL-6, and TNF-α and increased Bcl-2 and TGF-ß1, and IL-10. miR-20a-5p targeted STAT3. STAT3 overexpression partially reversed miR-20a-5p overexpression-mediated effects on LPS-induced BV2 cell viability, apoptosis, and inflammatory responses. miR-20a-5p overexpression inhibited MPTP-induced STAT3 and α-synuclein levels, microglia activation, and inflammatory response, and reduced the loss of TH-positive cells in mice. miR-20a-5p overexpression ameliorated MPTP-induced dyskinesia in PD model mice. CONCLUSION: miR-20a-5p alleviates neuronal damage and suppresses inflammation by targeting STAT3 in PD.

MiR-20a-5p alleviates kidney ischemia/reperfusion injury by targeting ACSL4-dependent ferroptosis.

Ischemia/reperfusion injury (IRI) is prone to occur after kidney transplantation, leading to delayed graft function (DGF). MicroRNAs play a crucial role in the pathogenesis of ischemia/reperfusion-induced acute kidney injury, and miR-20a-5p was found to be the most significantly upregulated gene in a DGF patient cohort. However, the roles of microRNAs in transplanted kidneys remain largely unknown. In this study, we found that miR-20a-5p was upregulated in the kidneys of acute kidney injury mice and in patients with DGF. We identified early growth response-1 as a critical upstream target and verified the binding of early growth response-1 to a predicted sequence in the promoter region of the miR-20a-5p gene. Functionally, the miR-20a-5p mimic attenuated IRI and postischemic renal fibrosis, whereas the miR-20a-5p inhibitor delivery aggravated IRI and fibrosis. Importantly, delivery of the miR-20a-5p mimic or inhibitor in the donor kidneys attenuated or aggravated renal loss and structural damage in cold storage transplantation injury. Furthermore, our study identified miR-20a-5p as a negative regulator of acyl-CoA synthetase long-chain family member 4 (ACSL4) by targeting the 3' untranslated region of ACSL4 mRNA, thereby inhibiting ACSL4-dependent ferroptosis. Our results suggest a potential therapeutic application of miR-20a-5p in kidney transplantation through the inhibition of ACSL4-dependent ferroptosis.

Non-coding RNAs are correlated to TGF-ß receptor type 2 in patients with colorectal cancer.

BACKGROUND: Multiple molecular expression alterations, particularly in non-coding RNAs, play fundamental roles in the regulations of cellular processes and may relate to the occurrence and progression of colorectal cancer (CRC). In the present study, we investigated the associations between TGFBR2, miR20a-5p and long non-coding RNA (lncRNA) LAMTOR5-AS1 in CRC patients. METHODS: Colorectal cancer and adjacent normal tissue samples (n = 34) were prepared from CRC patients. The associations between TGFBR2, miR20a-5p and lncRNA LAMTOR5-AS1 were predicted using bioinformatics tools. The expression levels of TGFBR2, miR20a-5p and lncRNA LAMTOR5-AS1 were measured using a quantitative real-time polymerase chain reaction technique. The TGFBR2 protein values were measured by western blotting. The clinical importance of lncRNA LAMTOR5-AS1 was assessed using receiver operating characteristic curve. RESULTS: The up-regulated levels of TGFBR2 (p = 0.02), TGFBR2 protein (p = 0.008) and lncRNA LAMTOR5-AS1 (p = 0.02) were significantly observed in CRC tissues compared to the adjacent normal tissues. The miR20a-5p expression level (p = 0.009) was downregulated in CRC tissues. In addition, the miR20a-5p expression level was inversely correlated to the TGFBR2 gene (r2  = 0.88, p < 0.0001), protein (r2  = 0.95, p < 0.0001) and lncRNA LAMTOR5-AS1 gene (r2  = 0.93, p < 0.0001) expression levels. Based on the area under curve, the increase of lncRNA LAMTOR5-AS1 expression level with a sensitivity of 64.52% and specificity of 65.52% was considered in CRC patients. CONCLUSIONS: We propose that miR20a-5p is inversely related to long non-coding RNA (lncRNA) LAMTOR5-AS, such that it may be involved in the regulation of TGFBR2 expression level in CRC patients.

Dynamic effects of miR-20a-5p on hippocampal ripple energy after status epilepticus in rats.

To determine the dynamic effects of miR-20a-5p on hippocampal ripple energy in rats after status epilepticus (SE). A lithium pilocarpine (LiCl-PILO)-induced rat model of status epilepticus (SE) was established, and the rats were divided into the normal control (Control, CTL), epileptic control (PILO), valproic acid (VPA + PILO), miR-20a-5p overexpression lentivirus vector (miR + PILO), sponges blocking lentivirus vector (Sponges + PILO), and scramble sequence negative control (Scramble + PILO) groups (n = 6). Electroencephalograms (EEGs) were used to analyze changes in hippocampal ripple energy before and after SE. Quantitative polymerase chain reaction (q-PCR) analysis showed that miR-20a-5p levels gradually increased after miR-20a-5p overexpression lentivirus vector injection into the lateral ventricle, and the miR-20a-5p levels were significantly higher than that in CTL group on days 7 and 36 (P < 0.001). The miR-20a-5p levels decreased significantly on days 7 and 36 after blocking by sponges lentivirus vector injected into the lateral ventricle (P < 0.001). After injection of PILO, the average ripple energy expression in each group gradually increased, and reached the peak before chloral hydrate injection (compared with 1 day before SE, P < 0.05). The ripple energy in the VPA + PILO and Sponges + PILO groups was significantly lower than that in the PILO group at 60 min and 70 min after PILO injection and before chloral hydrate injection (P < 0.05), and maintained lower until 2 h after chloral hydrate injection in VPA + PILO (P < 0.05). Compared with the VPA + PILO group, the mean ripple energy of the Sponges + PILO group had no difference at all time points (P ≥ 0.05). After SE, ripple distribution of space and energy is closely related to the occurrence of epilepsy. Inhibition of miR20a-5p expression can downregulate ripple oscillation energy during seizure.

Plasmatic MicroRNAs and Treatment Outcomes of Patients with Metastatic Castration-Resistant Prostate Cancer: A Hospital-Based Cohort Study and In Silico Analysis.

Prostate cancer (PCa) is one of the most common malignancies among men worldwide. Inevitably, all advanced PCa patients develop metastatic castration-resistant prostate cancer (mCRPC), an aggressive phase of the disease. Treating mCRPC is challenging, and prognostic tools are needed for disease management. MicroRNA (miRNA) deregulation has been reported in PCa, constituting potential non-invasive prognostic biomarkers. As such, this study aimed to evaluate the prognostic potential of nine miRNAs in the liquid biopsies (plasma) of mCRPC patients treated with second-generation androgen receptor axis-targeted (ARAT) agents, abiraterone acetate (AbA) and enzalutamide (ENZ). Low expression levels of miR-16-5p and miR-145-5p in mCRPC patients treated with AbA were significantly associated with lower progression-free survival (PFS). The two miRNAs were the only predictors of the risk of disease progression in AbA-stratified analyses. Low miR-20a-5p levels in mCRPC patients with Gleason scores of <8 were associated with worse overall survival (OS). The transcript seems to predict the risk of death regardless of the ARAT agent. According to the in silico analyses, miR-16-5p, miR-145-5p, and miR-20a-5p seem to be implicated in several processes, namely, cell cycle, proliferation, migration, survival, metabolism, and angiogenesis, suggesting an epigenetic mechanism related to treatment outcome. These miRNAs may represent attractive prognostic tools to be used in mCRPC management, as well as a step further in the identification of new potential therapeutic targets, to use in combination with ARAT for an improved treatment outcome. Despite the promising results, real-world validation is necessary.

LncRNA CALML3-AS1 suppresses papillary thyroid cancer progression via sponging miR-20a-5p/RBM38 axis.

BACKGROUND: The incidence and mortality of thyroid cancer (TC) has been steadily rising in the past decades. It is imperative to have a better understanding of the molecular mechanisms underlying TC development and identify novel therapeutic targets. This study characterized the role of lncRNA CALML3-AS1 (CALML3-AS1) in the development of papillary thyroid cancer (PTC). METHOD: Related mRNAs expression were validated in the tumor and adjacent normal tissues from 52 PTC patients and PTC cell lines by qRT-PCR. Expression of RBM38 was detected by Western blot. We have also conducted CCK-8 and colony formation assays were used to detect the effect of CALML3-AS1 on cell proliferation, Transwell assay was utilized to evaluate cell migration and invasion, apoptosis detected by flow cytometry assay, RNA pull-down and luciferase assays were performed to validate gene predictions. RESULTS: The results indicated that the expression of both CALML3A-S1 and RBM38 were significantly downregulated in PTC tissues (p < 0.01), while the expression of miR-20a-5p was increased in PTC (p < 0.01). Functionally, CALML3-AS1 overexpression inhibited PTC cell proliferation in vitro and in vivo. Mechanistically, CALML 3-AS1 sponged miR-20a-5p, which in turn leads to the suppression of RBM38 expression and PTC progression. CONCLUSIONS: CALML3-AS1 functions as a ceRNA for miR-20a-5p in the regulation of the expression of RBM38 in PTC. Higher level of CALML3-AS1 serves as a good prognostic indicator of survival in PTC patients. Targeting CALML3-AS1/ miR-20a-5p/RBM38 axis may represent a novel therapeutic strategy in the treatment of PTC.

Knockdown of lncRNA H19 alleviates ox-LDL-induced HCAECs inflammation and injury by mediating miR-20a-5p/HDAC4 axis.

BACKGROUND: Coronary artery disease (CAD) seriously disturbs the life of people. LncRNA H19 is reported to promote the progression of CAD; Nevertheless, the detailed mechanism by which H19 modulates CAD development is unclear. METHODS: Clinical samples of CAD patients were collected, meanwhile we established in vitro and in vivo models of CAD by treating HCAECs with ox-LDL and feeding ApoE-/- mice with high fat diets (HFD). MTT assay was adopted to assess the cell viability. Transwell detection was applied to test the migration, and apoptosis was tested by flow cytometry. The levels of inflammatory cytokines were examined by ELISA. The relation among H19, miR-20a-5p and HDAC4 was explored by dual luciferase reporter and RIP assay. RESULTS: H19 and HDAC4 levels were elevated, while miR-20a-5p was reduced in plasma of CAD patients and ox-LDL-treated HCAECs. ox-LDL increased H19 level and induced apoptosis and inflammation in HCAECs, while silencing of H19 rescued this phenomenon. In addition, the level of H19 was negatively correlated with miR-20a-5p, and miR-20a-5p inhibitor restored the effect of H19 silencing on HCAECs function. HDAC4 was the downstream mRNA of miR-20a-5p, and miR-20a-5p upregulation reversed ox-LDL-induced HCAECs injury through targeting HDAC4. Furthermore, H19 silencing significantly alleviated the coronary atherosclerotic plaques and inhibited the inflammatory responses in vivo. CONCLUSIONS: We proved that knockdown of H19 alleviated ox-LDL-induced HCAECs injury via miR-20a-5p/HDAC4 axis, which might provide a new tactics against CAD.

Combined high-throughput library screening and next generation RNA sequencing uncover microRNAs controlling human cardiac fibroblast biology.

BACKGROUND: Myocardial fibrosis is a hallmark of the failing heart, contributing to the most common causes of deaths worldwide. Several microRNAs (miRNAs, miRs) controlling cardiac fibrosis were identified in recent years; however, a more global approach to identify miRNAs involved in fibrosis is missing. METHODS AND RESULTS: Functional miRNA mimic library screens were applied in human cardiac fibroblasts (HCFs) to identify annotated miRNAs inducing proliferation. In parallel, miRNA deep sequencing was performed after subjecting HCFs to proliferating and resting stimuli, additionally enabling discovery of novel miRNAs. In-depth in vitro analysis confirmed the pro-fibrotic nature of selected, highly conserved miRNAs miR-20a-5p and miR-132-3p. To determine downstream cellular pathways and their role in the fibrotic response, targets of the annotated miRNA candidates were modulated by synthetic siRNA. We here provide evidence that repression of autophagy and detoxification of reactive oxygen species by miR-20a-5p and miR-132-3p explain some of their pro-fibrotic nature on a mechanistic level. CONCLUSION: We here identified both miR-20a-5p and miR-132-3p as crucial regulators of fibrotic pathways in an in vitro model of human cardiac fibroblast biology.

VEGF-mediated angiogenesis in retinopathy of prematurity is co-regulated by miR-17-5p and miR-20a-5p.

The microRNAs miR-17-5p and miR-20a-5p play important roles on angiogenesis; however, it is arguable whether they regulate the formation of retinal blood vessels in retinopathy of prematurity (ROP). We used a mouse model of oxygen-induced retinopathy (OIR) to simulate the development of retinas in mice suffering from ROP, and the expression levels of miR-20a-5p, miR-17-5p, hypoxia-inducible factor 1-alpha (HIF-1α), and vascular endothelial growth factor (VEGF) were measured by RT-qPCR and Western blotting. Cell proliferation, apoptosis, and angiogenesis in the OIR model mice were measured using MTT assays, flow cytometry, and Matrigel assays, respectively. The interaction between HIF-1α/VEGF and miR-20a-5p/miR-17-5p were further validated using dual-luciferase reporter assays, biotin-labeled RNA-pulldown, and RNA immunoprecipitation (RIP) assays. In our OIR model, retinal angiogenesis in the mice was associated with down-regulation of miR-20a-5p and miR-17-5p, as well as up-regulation of HIF-1α and VEGF. In addition, the miR-20a-5p and miR-17-5p inhibited cell proliferation and angiogenesis through regulating HIF-1α and VEGF in the retinal cells of the OIR model mice. Moreover, it was found that miR-20a-5p and miR-17-5p bind to HIF-1α and VEGF at the 3'UTR, and there was a combined effect between miR-20a-5p and miR-17-5p on the regulation of HIF-1α and VEGF. It is worth noting that miR-17-5p and miR-20a-5p can preferentially regulate HIF-1α, then act on VEGF, thereby affecting the angiogenesis associated with ROP.

Long non-coding RNA SNHG16 regulates E2F1 expression by sponging miR-20a-5p and aggravating proliferative diabetic retinopathy.

Long non-coding RNAs (lncRNAs) were reported to be related to microvascular dysfunction in diabetic retinopathy (DR), but the potential mechanism remains unknown. This study was designed to elucidate the effects of lncRNA small nucleolar RNA host gene 1 (SNHG16) in proliferative DR progression. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to measure the levels of SNHG16 and miR-20a-5p from peripheral blood samples of different participants. Pearson's correlation analysis was applied to the plasma data to detect correlations between SNHG16 and miR-20a-5p. Finally, the interactions of miR-20a-5p and SNHG16 or E2F1 were assessed by luciferase reporter assays. SNHG16 and E2F1 were increased and miR-20a-5p was decreased in proliferative DR both in vivo and in vitro when compared with control or non-proliferative DR. E2F1 was identified as the target of miR-20a-5p. The miR-20a-5p interacted with SNHG16 and E2F1 and was controlled by SNHG16. The regulation of SNHG16 on E2F1 was mediated by miR-20a-5p. Cells transfected with SNHG16 overexpression plasmid markedly increased cell apoptosis and vessel-like formation, whereas the miR-20a-5p mimic partially reversed these effects. Transfection with gene silencing E2F1 plasmid rescued SNHG16 overexpression-aggravated proliferative DR. This study indicated that SNHG16 regulated E2F1 expression by sponging miR-20a-5p and aggravating proliferative DR.

Long non-coding RNA H19 promotes myoblast fibrogenesis via regulating the miR-20a-5p-Tgfbr2 axis.

Emerging evidence has indicated long non-coding RNAs (lncRNAs) play important roles in diverse biological processes, including fibrosis. Here, we report that lncRNA H19 is able to promote skeletal muscle fibrosis. lnc-H19 was identified to be highly expressed in skeletal muscle fibrosis in vivo and in vitro; while lnc-H19 knockdown attenuated fibrosis in vitro. The knockdown of lnc-H19 was proved to inhibit the activation of the TGFß/Smad pathway in C2C12 myoblasts by sponging miR-20a-5p to regulate Tgfbr2 expression through the competing endogenous RNA function. Our study elucidates the roles of the lnc-H19-miR-20a-5p-Tgfbr2 axis in regulating the TGFß/Smad pathway of myoblast fibrogenesis, which might provide a promising therapeutic target for skeletal muscle fibrosis.

miR-19b-3p and miR-20a-5p are associated with the levels of antiphospholipid antibodies in patients with antiphospholipid syndrome.

Monocytes play a key role in pathophysiology of antiphospholipid syndrome (APS), nevertheless it is unclear if microRNA expression is associated with particular APS features. Identify whether miR-19b-3p and miR-20a-5p expression in monocytes are associated with hallmarks of the APS. Fifty-seven APS patients and 18 healthy controls were studied. Expression of miR-19b-3p and miR-20a-5p was measured in monocytes by RT-qPCR. Both miR-19b-3p (AUC = 0.835, 95% CI 0.733-0.938; P < 0.001) and miR-20a-5p (AUC = 0.857, 0.757-0.957; P < 0.001) discriminated APS patients from healthy individuals. A cut-off point of 1.98 for miR-19-3p and 2.18 for miR-20a-5p showed that APS patients with low microRNA expression had higher levels of IgM and IgG anticardiolipin antibodies than patients with high microRNA expression. In addition, APS patients with low microRNA expression had higher IgG anti-ß2 glycoprotein I antibody levels than their counterparts with high microRNA expression. Finally, miR-19b-3p and miR-20a-5p expression levels were significantly higher in APS patients using oral anticoagulants. Monocyte expression of miR-19b-3p and miR-20a-5p is low in APS, and patients with the lowest microRNA expression presented the highest levels of antiphospholipid antibodies.

HIF1A/miR-20a-5p/TGFß1 axis modulates adipose-derived stem cells in a paracrine manner to affect the angiogenesis of human dermal microvascular endothelial cells.

Angiogenic cytokines secreted by the adipose-derived stem cells (ADSCs) might promote the angiogenesis of endothelial cells. In the present study, we hypothesize that miR-20a targets TGFB1 to modulate the transforming growth factor ß1 (TGFß1) secretion by ADSCs, therefore affecting the angiogenesis. We found that hypoxia-inducible factor 1A (HIF1A) and TGFß1 expressions were increased by hypoxia, accompanied with promoted ADSC cell viability. Incubation with conditioned medium from ADSCs treated with hypoxia significantly enhanced the angiogenesis capacity of human dermal microvascular endothelial cells (HDMECs), while TGFB1-silenced ADSCs medium significantly reverses HDMECs angiogenesis. miR-20a suppresses the expression of TGFB1 and secretion of TGFß1 by ADSCs via binding to its 3'untranslated region, therefore modulating the HDMEC angiogenesis via affecting the paracrine from ADSCs; the effects of miR-20a-overexpressed conditioned medium on HDMEC angiogenesis were significantly reversed by TGFB1-overexpressed conditioned medium. Finally, HIF1A suppressed the expression of miR-20a via targeting its promoter region, subsequently promoting the paracrine from ADSCs and HDMEC angiogenesis.

miR-20a-5p is enriched in hypoxia-derived tubular exosomes and protects against acute tubular injury.

Exosomes have been shown to effectively regulate the biological functions of target cells. Here, we investigated the protective effect and mechanism of hypoxia-induced renal tubular epithelial cells (TECs)-derived exosomes on acute tubular injury. We found that in vitro hypoxia-induced tubular exosomes (Hy-EXOs) were protective in acute tubular injury by promoting TECs proliferation and improving mitochondrial functions. By using exosome miRNA sequencing, we identified miR-20a-5p was abundant and was a key mechanism for the protective effect of Hy-EXOs on tubular injury as up-regulation of miR-20a-5p enhanced but down-regulation of miR-20a-5p inhibited the protective effect of Hy-EXOs on tubular injury under hypoxia conditions. Further study in a mouse model of ischemia-reperfusion-induced acute kidney injury (IRI-AKI) also confirmed this notion as pre-treating mice with the miR-20a-5p agomir 48 h prior to AKI induction was capable of inhibiting IRI-AKI by lowering serum levels of creatinine and urea nitrogen, and attenuating the severity of tubular necrosis, F4/80(+) macrophages infiltration and vascular rarefaction. Mechanistically, the protective effect of miR-20a-5p on acute kidney injury (AKI) was associated with inhibition of TECs mitochondrial injury and apoptosis in vitro and in vivo. In conclusion, miR-20a-5p is enriched in hypoxia-derived tubular exosomes and protects against acute tubular injury. Results from the present study also reveal that miR-20a-5p may represent as a novel therapy for AKI.

MiR-20a-5p suppressed TGF-ß1-triggered apoptosis of human bronchial epithelial BEAS-2B cells by targeting STAT3.

Apoptosis of bronchial epithelial cells contributes to lung diseases, including asthma. Although miR-20a-5p is reportedly downregulated in the bronchial epithelia of asthmatic patients, its function and mechanism still need to be explored. Here, we explored how miR-20a-5p affects human bronchial epithelial cells stimulated with transforming growth factor (TGF)-ß1. Using qRT-PCR, we observed downregulated miR-20a-5p levels in these cells. After transfecting miR-20a-5p mimics or inhibitors into human bronchial epithelium BEAS-2B cells, a Cell Counting Kit-8 assay and flow cytometry analysis showed that the mimics mitigated suppression of cell viability and acceleration of apoptosis that was triggered by TGF-ß1, whereas the inhibitors exerted the opposite effects. TGF-ß1 induced a decrease in expression of Bcl-2 and an increase in expression of Bax, both of which were inhibited by miR-20a-5p mimics and further enhanced by miR-20a-5p inhibitors. Further study verified that miR-20a-5p targeted the signal transducer and activator of transcription 3 (STAT3) and the STAT3 level was inversely related to the miR-20a-5p level. Furthermore, STAT3 overexpression partly counteracted the miR-20a-5p-induced anti-apoptotic effect in TGF-ß1-treated BEAS-2B cells. In summary, this study suggested that miR-20a-5p restrained apoptosis in TGF-ß1-stimulated BEAS-2B cells by targeting STAT3. MiR-20a-5p thus may be a novel therapeutic target for asthma treatment.

miR-20a-5p promotes pulmonary artery smooth muscle cell proliferation and migration by targeting ABCA1.

BACKGROUND: The function of miR-20a-5p in pulmonary artery smooth muscle cells (PASMCs) and the underlying mechanism remains largely unknown. METHODS: C57BL/6J mice and PASMCs were used for constructing pulmonary artery hypertension (PAH) animal and cell models, respectively. Reverse transcription polymerase chain reaction (RT-PCR) was employed to detect miR-20a-5p and ATP-binding cassette subfamily A member 1 (ABCA1) messenger RNA expression. CCK-8, Transwell, and TUNEL experiments were used to determine PASMCs proliferation, migration, and apoptosis. The relationship between miR-20a-5p and ABCA1 was detected by luciferase reporter experiment, Western blot analysis, and qRT-PCR. RESULTS: miR-20a-5p was remarkably elevated in PASMCs of PAH mice and human PASMCs treated by hypoxia, while ABCA1 was remarkably decreased. After transfection of miR-20a-5p mimics, PASMCs proliferation and migration were promoted and PASMCs apoptosis was suppressed. ABCA1 was confirmed to be a target of miR-20a-5p and restoration of ABCA1 reversed the function of miR-20a-5p. CONCLUSION: miR-20a-5p enhances the proliferation and migration of PASMCs to promote the development of PAH via targeting ABCA1.

miR-20a-5p regulates pulmonary surfactant gene expression in alveolar type II cells.

MicroRNA (miRNA) critically controls gene expression in many biological processes, including lung growth and pulmonary surfactant biosynthesis. The present study was conducted to investigate whether miR-20a-5p had such regulatory functions on alveolar type II (AT-II) cells. To accomplish this, miR-20a-5p-overexpressed and miR-20a-5p-inhibited adenoviral vectors were constructed and transfected into cultured AT-II cells that were isolated from rat foetal lungs of 19 days' gestation. Transfection efficiency was confirmed by observing the fluorescence of green fluorescent protein (GFP) carried by the viral vector, whereas miR-20a-5p levels were verified by real-time PCR. The CCK-8 assay was used to compare the proliferation ability of AT-II cells that had over- or underexpressed miR-20a-5p. The expression of surfactant-associated proteins (SPs) and phosphatase and tensin homolog (PTEN) was measured by real-time PCR and Western blotting. In AT-II cells, transfection resulted in over- or under-regulation of miR-20a-5p. While overexpression of miR-20a-5p promoted pulmonary surfactant gene expression, its underexpression inhibited it. Consistent with its role in negatively regulating the pulmonary surfactant gene, an opposite pattern was observed for miR-20a-5p regulation of PTEN. As a result, when miR-20a-5p was rendered overexpressed, PTEN was down-regulated. By contrast, when miR-20a-5p was underexpressed, PTEN was up-regulated. Neither overexpression nor underexpression of miR-20a-5p altered the cell proliferation. miR-20a-5p plays no role in proliferation of foetal AT-II cells but is a critical regulator of surfactant gene expression. The latter appears to be achieved through a regulatory process that implicates expression of PTEN.

Silencing of circ_0009910 inhibits acute myeloid leukemia cell growth through increasing miR-20a-5p.

Acute myeloid leukemia (AML) is the most common acute leukemia in adults, which is aggressive cancer. CircRNAs are abundantly expressed in the hematologic malignancy cells. In this study, we aimed to investigate the expression profiling of circRNAs in AML. We performed circRNA-sequencing to identify differentially expressed circRNAs in bone marrow samples from AML patients and iron-deficiency anemia (control). Furthermore, we found that circ_0009910 was significantly upregulated in AML patients compared with iron-deficiency anemia patients. High circ_0009910 expression predicted a poor risk and outcome of AML patients. Further experiments in vitro and in vivo demonstrated that knockdown of circ_0009910 inhibited AML cell proliferation and induced apoptosis through sponging miR-20a-5p. Our findings firstly identify that circ_0009910 is significantly upregulated in AML bone marrow samples and might serve as a novel outcome biomarker for AML. Both circ_0009910 and miR-20a-5p may be potential therapeutic targets for future AML treatment.

LncRNA PVT1 triggers Cyto-protective autophagy and promotes pancreatic ductal adenocarcinoma development via the miR-20a-5p/ULK1 Axis.

BACKGROUND: Defective autophagy is thought to contribute to the pathogenesis of many diseases, including cancer. Human plasmacytoma variant translocation 1 (PVT1) is an oncogenic long non-coding RNA that has been identified as a prognostic biomarker in pancreatic ductal adenocarcinoma, but how PVT1 operates in the regulation of autophagy in pancreatic ductal adenocarcinoma (PDA) is unclear. METHODS: PVT1 expression level was detected by quantitative real-time polymerase chain reaction (qRT-PCR) and hybridization in situ (ISH). Western blot or qRT-PCR was performed to assess the ULK1 protein or mRNA level. Autophagy was explored via autophagic flux detection under a confocal microscope and autophagic vacuoles investigation under a transmission electron microscopy (TEM). The biological role of PVT1 in autophagy and PDA development was determined by gain-of-function and loss-of-function assays. RESULTS: We found that PVT1 levels paralleled those of ULK1 protein in PDA cancer tissues. PVT1 promoted cyto-protective autophagy and cell growth by targeting ULK1 both in vitro and in vivo. Moreover, high PVT1 expression was associated with poor prognosis. Furthermore, we found that PVT1 acted as sponge to regulate miR-20a-5p and thus affected ULK1 expression and the development of pancreatic ductal adenocarcinoma. CONCLUSIONS: The present study demonstrates that the "PVT1/miR-20a-5p/ULK1/autophagy" pathway modulates the development of pancreatic ductal adenocarcinoma and may be a novel target for developing therapeutic strategies for pancreatic ductal adenocarcinoma.