Mesenchymal stem cells-derived exosomes carrying microRNA-30b confer protection against pulmonary fibrosis by downregulating Runx1 via Spred2.

Idiopathic pulmonary fibrosis (IPF) is a chronic pulmonary fibrosis disease that is fatal. Mesenchymal stem cells (MSCs)-secreted exosomes (exos) have been linked to improving PF. Moreover, exosomal microRNAs (miRs) can control the growth of numerous diseases, including lung disorders. Our bioinformatics analysis showed that miR-30b was downregulated in tissue samples from surgical remnants of biopsies or lungs explanted from patients with IPF who underwent pulmonary transplantation. This suggests that miR-30b plays an important role in both the pathogenesis and treatment of IPF. Herein, this research was designed to ascertain the mechanism of MSCs-exos-packaged miR-30b in alleviating PF. The serum was harvested from idiopathic PF (IPF) patients with interstitial pneumonia caused by dermatomyositis and the MLE12 lung epithelial cell fibrosis model was built with TGF-ß1 (10 ng/mL), followed by miR-30b expression determination. TGF-ß1-stimulated MLE12 cells were co-incubated with exos from MSCs with or without Spred2 or Runx1 overexpression, followed by measurement of cell viability and apoptosis. After establishing the IPF mouse model with bleomycin and injecting exos and/or silencing and overexpressing adenovirus vectors, fibrosis evaluation was conducted. In mice and cells, the expression of TGF-ß1, TNF-α, and IL-1ß was tested via ELISA, and the levels of E-cad, ZO-1, α-SMA, and collagen type I via western blot analysis. The promoters of miR-30b, Runx1, and Spred2 were investigated. miR-30b was downregulated in the serum of IPF patients and TGF-ß1-stimulated MLE12 cells. Mechanistically, miR-30b inhibited Spred2 transcription by negatively targeting Runx1. MSCs-exos or MSCs-exo-miR-30b decreased the apoptosis, inflammation, and fibrosis while increasing their viability in TGF-ß1-stimulated MLE12 cells, which was annulled by overexpressing Runx1 or Spred2. Exo-miR-30b decreased Runx1 expression to downregulate Spred2, reducing fibrosis and inflammation in IPF mice. Our results indicated that MSCs-exos-encapsulated miR-30b had a potential function to inhibit PF and part of its function may be achieved by targeting RUNX1 to reduce the Spred2 transcription level. Moreover, this work offered evidence and therapeutic targets for therapeutic strategies for managing clinical PF in patients.

Inhibitory role of bone marrow mesenchymal stem cells-derived exosome in non-small-cell lung cancer: microRNA-30b-5p, EZH2 and PI3K/AKT pathway.

Exosomal microRNA (miRNA) exerts potential roles in non-small-cell lung cancer (NSCLC). The current study elucidated the role of miR-30b-5p shuttled by bone marrow mesenchymal stem cells (BMSCs)-derived exosomes in treating NSCLC. Bioinformatics analysis was performed with NSCLC-related miRNA microarray GSE169587 and mRNA data GSE74706 obtained for collection of the differentially expressed miRNAs and mRNAs. The relationship between miR-30b-5p and EZH2 was predicted and confirmed. Exosomes were isolated from BMSCs and identified. BMSCs-derived exosomes overexpressing miR-30b-5p were used to establish subcutaneous tumorigenesis models to study the effects of miR-30b-5p, EZH2 and PI3K/AKT signalling pathway on tumour growth. A total of 86 BMSC-exo-miRNAs were differentially expressed in NSCLC. Bioinfomatics analysis found that BMSC-exo-miR-30b-5p could regulate NSCLC progression by targeting EZH2, which was verified by in vitro cell experiments. Besides, the target genes of miR-30b-5p were enriched in PI3K/AKT signalling pathway. Animal experiments validated that BMSC-exo-miR-30b-5p promoted NSCLC cell apoptosis and prevented tumorigenesis in nude mice via EZH2/PI3K/AKT axis. Collectively, the inhibitory role of BMSC-derived exosomes-loaded miR-30b-5p in NSCLC was achieved through blocking the EZH2/PI3K/AKT axis.

LncRNA HOTTIP impacts the proliferation and differentiation of fibroblast-like synoviocytes in ankylosing spondylitis through the microRNA-30b-3p/PGK1 axis.

OBJECTIVE: Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) have been reported to exert regulatory effects on biological processes. This study intended to assess the role of the lncRNA HOXA transcript at the distal tip (HOTTIP)/miR-30b-3p/phosphoglycerate kinase 1 (PGK1) axis in ankylosing spondylitis (AS). METHODS: Levels of HOTTIP, miR-30b-3p and PGK1 in AS synovial tissues and cultured AS fibroblast-like synoviocytes (ASFLSs) were assessed. The ASFLSs were identified and, respectively, treated with altered expression of HOTTIP and miR-30b-3p, and then, the proliferation and differentiation of the ASFLSs were assessed. The AS mouse models were established by injection of proteoglycan and Freund's complete adjuvant and then were treated with altered expression of HOTTIP and miR-30b-3p, and the pathological changes and apoptosis of synoviocytes in mice' synovial tissues were measured. The relationship of HOTTIP, miR-30b-3p and PGK1 was verified. RESULTS: HOTTIP and PGK1 were elevated, while miR-30b-3p was reduced in AS synovial tissues and ASFLSs. Elevated miR-30b-3p or inhibited HOTTIP restrained proliferation and differentiation of ASFLSs and also improved the pathological changes and promoted apoptosis of synoviocytes in mice's synovial tissues. PGK1 was a target of miR-30b-3p, and miR-30b-3p could directly bind to HOTTIP. Silencing miR-30b-3p or overexpressing PGK1 reversed the improvement of AS by knocking down HOTTIP or up-regulating miR-30b-3p. CONCLUSION: Our study suggests that reduced HOTTIP ameliorates AS progression by suppressing the proliferation and differentiation of ASFLSs through the interaction of miR-30b-3p and PGK1.

Effect of plasma-derived extracellular vesicles on angiogenesis and the ensuing proliferative diabetic retinopathy through a miR-30b-dependent mechanism.

BACKGROUND/PURPOSE: Proliferative diabetic retinopathy (PDR) is a major diabetic microvascular complication, characterized by pathological angiogenesis. This study sets out to investigate the potential molecular mechanism in the angiogenesis during PDR. METHODS: The expression of microRNA-30b (miR-30b) was quantified in a streptozotocin (STZ)-induced mouse model of PDR. The binding affinity between SIRT1 and miR-30b was then identified and validated. After transduction with In-miR-30b or combined with sh-SIRT1, high-glucose (HG)-induced retinal microvascular endothelial cells (RMECs) were co-cultured with extracellular vesicles (EVs) derived from the plasma of PDR mice (plasma-EVs). The proliferation and angiogenesis of RMECs were then detected in vitro. RESULTS: miR-30b expression was upregulated in the retinal tissue of PDR mice. SIRT1 was a target gene of miR-30b and under the negative regulation by miR-30b in RMECs. In contrast, inhibition of miR-30b resulted in elevated SIRT1 expression, thus alleviating the angiogenesis of RMECs. miR-30b was enriched in the plasma-EVs and could be delivered to RMECs, in which miR-30b exerted pro-angiogenic effects. Furthermore, inhibition of miR-30b arrested the progression of PDR in mice by promoting the expression of SIRT1. CONCLUSION: Collectively, the present study pinpointed the involvement of miR-30b delivered by plasma-EVs in PDR angiogenesis, thus laying the basis for the development of novel therapeutic targets for the treatment of PDR.

LncRNA GATA3­AS1­miR­30b­5p­Tex10 axis modulates tumorigenesis in pancreatic cancer.

Long noncoding RNAs (lncRNAs) have been widely recognized to play an important role in a variety of diseases. Abnormal regulation of lncRNA GATA3­antisense RNA 1 (AS1) occurs in several cancers, but whether it is involved in the progression of pancreatic cancer (PC) remains unknown. The present study aimed to investigate the biological effects of GATA3­AS1 in PC and to explore the underlying molecular mechanisms. Upregulation of GATA3­AS1 was revealed in PC tissues and cell lines. Knockdown of GATA3­AS1 in PANC­1 or AsPC­1 cells markedly reduced cell viability, cell proliferation, and cell invasion abilities, while cell apoptosis was increased. In addition, GATA3­AS1 knockdown suppressed the stemness of PANC­1 and AsPC­1 cells by decreasing the spheroid formation ability. A tumor xenograft in vivo assay demonstrated that GATA3­AS1 knockdown inhibited tumorigenicity of AsPC­1 cells. Furthermore, the microRNA (miR)­30b­5p downregulation and GATA3­AS1 upregulation were revealed in PC tissues and cell lines. Negative correlations were present between GATA3­AS1 and miR­30b­5p and between miR­30b­5p and testis­expressed protein 10 (Tex10) in the PC tissues, while GATA3­AS1 and Tex10 were positively correlated. GATA3­AS1 was then revealed to act as a competing endogenous RNA (ceRNA) for miR­30b­5p in regulating Tex10 expression. Moreover, the miR­30b­5p­Tex10 axis was confirmed to be involved in the regulation of biological effects of GATA3­AS1, including cell viability, cell proliferation, cell invasion, cell apoptosis, and cell stemness, as well as Wnt1/ß­catenin signaling. Collectively, these data indicated that the GATA3­AS1­miR­30b­5p­Tex10 axis modulates tumorigenesis in PC, which may be associated with the Wnt/ß­catenin signaling pathway.

The microRNA cluster miR-30b/-30d prevents tumor cell switch from an epithelial to a mesenchymal-like phenotype in GBC.

As a malignancy of the gastrointestinal tract, gallbladder cancer (GBC) continues to exhibit notable rates of mortality. The current study aimed at investigating the effects associated with miR-30b and miR-30d (miR-30b/-30d) patterns in tumor cells undergoing epithelial-to-mesenchymal transition (EMT) in GBC. It identified that miR-30b and miR-30d, composed as a miRNA cluster, exhibited lower levels in the cancerous tissues from 50 patients with GBC relative to the gallbladder tissues from 35 patients with chronic cholecystitis. As expected, elevated expression of miR-30b/-30d was found to inhibit the EMT process, as evidenced by enhanced E-cadherin and reduced N-cadherin and vimentin in human GBC cells treated with miR-30b mimic, miR-30d mimic, and miR-30b/-30d mimic. Semaphorin-6B (SEMA6B) was identified as a target gene of miR-30b/-30d. Silencing of SEMA6B by its specific small interfering RNA (siRNA) mimicked the effect of miR-30b/-30d upregulation on the GBC cell EMT. Consistently, SEMA6B overexpression promoted this phenotypic switch even in the presence of miR-30b/-30d mimic. The tumorigenicity assay data obtained from nude mice also further supported the notion that miR-30b/-30d inhibited EMT of GBC cells. Thus, based on the key findings of the current study, we concluded that the miR-30b/-30d cluster may provide a potential avenue for targeting mesenchymal-like, invasive tumor cells in GBC.

Regulatory Mechanism of MicroRNA-30b on Neonatal Hypoxic-Ischemic Encephalopathy (HIE).

OBJECTIVE: This study is to investigate the role of microRNA (miR)-30b in the pathogenesis of hypoxic-ischemic encephalopathy (HIE) in neonates. METHODS: Totally 26 cases of neonatal HIE were included in this study. The protein expression levels of CD26P and PAI-1 were detected with ELISA. Serum levels of miR-30b and PAI-1 mRNA was measured by quantitative real-time PCR. Human brain microvascular endothelial cells (HBMECs) were cultured under hypoxic condition, and the intracellular expression levels of miR-30b and PAI-1 were evaluated. Dual-luciferase reporter assay was performed to confirm the interaction between miR-30b and PAI-1. RESULTS: Compared with the control group, both the mRNA and protein expression levels of PAI-1 in the serum were up-regulated in the neonates with HIE, together with up-regulated serum CD26P levels. However, the serum expression level of miR-30b was down-regulated in neonatal HIE. In hypoxia-induced HBMECs, the mRNA and protein expression levels of PAI-1 were significantly up-regulated, while the miR-30b expression level was significantly down-regulated. Dual-luciferase reporter assay showed that PAI-1 was the direct target of miR-30b. CONCLUSION: Neonatal HIE is accompanied with abnormal platelet activation, significantly up-regulated serum PAI-1 expression levels, and down-regulated miR-30b expression. MiR-30b might regulate the disease pathogenesis and immune responses via modulating PAI-1.

MicroRNA-30b participates in the pathological process of hyperuricemia by regulating interleukin-6 receptor.

The present study aimed to examine the expression of hyperuricemia (HUA)-related factors in the body fluids of HUA patients and in renal tissues and body fluids of HUA mice to elucidate the underlying mechanism of HUA and provide theoretical basis for the diagnosis, prevention and treatment of this disease. A total of 51 HUA patients (HUA group), and 36 healthy subjects (control group) were included in the present study. The peripheral blood and urine were collected from all patients and healthy subjects. A total of 20 male Kunming mice were used to construct HUA model, and another 20 mice were used as controls. The kidney tissues, peripheral blood and urine were collected from all mice. ELISA was performed to determine the levels of interleukin-6 receptor (IL-6R) proteins in the serum and urine of human or mice, while western blotting was employed to determine the protein expression in the kidney tissues of mice. Quantitative real-time polymerase chain reaction was used to measure the expression of mRNA and miR-30b in all sample types. Dual luciferase reporter assay was performed to identify the direct interaction between 3'-untranslated region of IL-6R mRNA and miR-30b. The expression of IL-6R mRNA and protein was increased in serum and urine of HUA patients, while the expression of miR-30b was reduced in HUA patients when compared with healthy subjects. The contents of uric acid, urea nitrogen and creatinine in the blood of HUA mice model were significantly elevated. Similarly, the expression of IL-6R mRNA and protein was increased in kidney, serum and urine of HUA mice model, while the expression of miR-30b was reduced in kidney tissues, serum and urine of HUA mice model. Dual luciferase reporter assay showed that miR-30b was able to bind with 3'-UTR seed region of IL-6R mRNA to regulate its expression. These findings demonstrated that the expression of IL-6R in patients and mouse with HUA is elevated, which is related with the down-regulation of miR-30b. Therefore, miR-30b might participate in the pathological process of HUA by regulating IL-6R.

Cannabinoid Receptor 1/miR-30b-5p Axis Governs Macrophage NLRP3 Expression and Inflammasome Activation in Liver Inflammatory Disease.

Nod-like receptor (NLR) family pyrin domain containing 3 (NLRP3) has been regarded as an important initiator or promoter in multiple inflammatory diseases. However, the relationship between cannabinoid receptor 1 (CB1) and macrophage NLRP3 inflammasome and the corresponding molecular mechanism in liver inflammation remain unclear. Mouse liver injury models were induced by carbon tetrachloride (CCl4) or methionine-choline-deficient and high fat (MCDHF) diet. Human liver tissues were obtained from patients with different chronic liver diseases. CB1 expression was increased in liver tissue and macrophages of CCl4- and MCDHF-treated mice, positively correlated with NLRP3. CB1 agonist ACEA (Arachiodonyl-2'-Chloroethylamide) promoted NLRP3 expression and NLRP3 inflammasome activation in macrophages. CB1 blockade with its antagonist AM281 reduced NLRP3 expression, inflammasome activation, and liver inflammation in CCl4- and MCDHF-treated mice. MicroRNA-30b-5p (miR-30b-5p), screened by the intersection of bioinformatics databases and downregulated miRNAs in injured liver, negatively correlated with NLRP3 in mouse and human liver. miR-30b-5p was involved in CB1-mediated activation of NLRP3 inflammasome in macrophages by directly targeting NLRP3. Importantly, administration of miR-30b-5p agomir targeted NLRP3 and attenuated liver inflammation in the injured liver. Altogether, CB1/miR-30b-5p axis modulates NLRP3 expression and NLPR3 inflammasome activation in macrophages during liver inflammation, which provides a potential target for liver disease.

Up-regulation of microRNA-30b/30d cluster represses hepatocyte apoptosis in mice with fulminant hepatic failure by inhibiting CEACAM1.

Recently, impacts of microRNAs have been unraveled in human diseases, and we aimed to confirm the role of miR-30b/30d in fulminant hepatic failure (FHF). Expression of miR-30b/30d and CEACAM1 in serum of FHF patients and healthy people was measured by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis. Mice FHF models were established by injection of D-Galn and lipopolysaccharide, and were treated with miR-30b/30d mimics. Oxidative stress, liver injury, and inflammatory reaction in mouse liver tissues were measured using oxidative stress-related factor kits, hematoxylin-eosin staining and enzyme-linked immunosorbent assay, respectively. Moreover, cell cycle distribution and apoptosis of hepatocytes of mice were determined by flow cytometry, and the target relation between miR-30b/30d and CEACAM1 was confirmed by bioinformatic method and dual luciferase reporter gene assay. MiR-30b/30d expression was positively, and CEACAM1 expression was negatively related to prognosis of FHF patients. Up-regulation of miR-30b/30d attenuated oxidative stress, liver injury, and inflammatory reaction, and improved survival rate of FHF mice. Furthermore, elevated miR-30b/30d ameliorated apoptosis and cell cycle arrest of hepatocytes of FHF mice. CEACAM1 was a target gene of miR-30b/30d. This study highlights that up-regulated miR-30b/30d attenuates the progression of FHF by targeting CEACAM1, which may be helpful to FHF treatment.

Down-regulated microRNA-30b-3p inhibits proliferation, invasion and migration of glioma cells via inactivation of the AKT signaling pathway by up-regulating RECK.

microRNAs (miRNAs) have been found to affect various cancers, and expression of numerous miRNAs is revealed in glioma. However, the role of microRNA-30b-3p (miR-30b-3p) in glioma remains elusive. Therefore, the present study aims to explore the specific mechanism by which miR-30b-3p influence the development of glioma in relation to the AKT signaling pathway. First, glioma cell lines were collected with miR-30b-3p and reversion-inducing cysteine-rich protein with kazal motifs (RECK) expression measured. The functional role of miR-30b-3p and RECK in glioma was determined via gain- and loss-of-function approaches. Subsequently, the expression of invasion- and migration-related factors (MMP-2 and MMP-9) and the AKT signaling pathway-related factors (AKT, p-AKT and PI3K-p85) was detected. Moreover, in vivo experiments were also conducted to investigate how miR-30b-3p influences in vivo tumorigenesis. The results showed that miR-30b-3p was up-regulated and RECK was down-regulated in glioma. RECK was a target gene of miR-30b-3p. Decreased miR-30b-3p and overexpressed RECK led to decreased expression of MMP-2, MMP-9 and p-AKT. Overexpressed RECK and LY294002 could decrease p-AKT and PI3K-p85 expression accompanied with unchanged expression of total protein of AKT. Additionally, proliferation, migration and invasion of glioma cells and tumor formation in nude mice were repressed owing to reduced expression of miR-30b-3p or elevated expression of RECK. In summary, miR-30b-3p inhibition suppresses metastasis of glioma cells by inactivating the AKT signaling pathway via RECK up-regulation, providing a new target for glioma treatment.

Exosomes derived from microRNA-30b-3p-overexpressing mesenchymal stem cells protect against lipopolysaccharide-induced acute lung injury by inhibiting SAA3.

Mesenchymal stem cells (MSCs) have been widely documented for their potential role in the treatment of various clinical disorders, including acute lung injury (ALI). ALI represents a clinical syndrome associated with histopathological diffuse alveolar damage. Recent evidence has demonstrated that exosomes derived from MSCs may serve as a reservoir of anti-apoptotic microRNAs (miRs) conferring protection from certain diseases. Hence, the current study was performed with the aim of investigating whether MSCs-exosomal miR-30b-3p could confer protection against ALI. A bioinformatic analysis and a dual luciferase assay were initially performed to verify that SAA3 was highly-expressed in ALI which was confirmed to be a target gene of miR-30b-3p. Next, the lipopolysaccharide (LPS)-treated type II alveolar epithelial cells (AECs) (MLE-12) were transfected with mimics or inhibitors of miR-30b-3p, or sh-SAA3. It was revealed that LPS induced AEC apoptosis, which could be inhibited by overexpressing miR-30b-3p by down-regulating the expression of SAA3. After co-culture of PKH26-labeled exosomes with MLE-12 cells, we found that the number of PKH26-labeled exosomes endocytosed by MLE-12 cells gradually increased. Furthermore, the LPS-treated MLE-12 cells co-cultured with MSC-exosomes overexpressing miR-30b-3p exhibited increased miR-30b-3p, decreased SAA3 level, as well as increased cell proliferation, accompanied by diminished cell apoptosis in LPS-treated MLE-12 cells. Finally, the protective effect of MSCs-exosomal miR-30b-3p on the AECs in vivo was investigated in an ALI mouse model with tail vein injection of MSC-exosomes with elevated miR-30b-3p, showing that overexpression of miR-30b-3p in MSC-exosomes conferred protective effects against ALI. Taken together, these findings highlighted the potential of MSC-exosomes overexpressing miR-30b-3p in preventing ALI. The exosomes derived from MSCs hold potential as future therapeutic strategies in the treatment of ALI.

Decreased expression of microRNA-30b promotes the development of pulpitis by upregulating the expression of interleukin-6 receptor.

The present study aimed to examine the expression of interleukin-6 receptor (IL-6R) mRNA and protein in pulp tissues, blood and saliva from patients with pulpitis. It also investigated the association between IL-6R and microRNA (miR)-30b, as well as their effects on pulpitis. A total of 28 patients with pulpitis were recruited into the experimental group and 16 subjects with no pulpitis who also underwent tooth extraction were recruited into the control group. Pulp tissues, plasma and saliva were collected from all participants. Reverse transcription-quantitative polymerase chain reaction was used to determine the expression of IL-6R mRNA and miR-30b in all sample types. Western blot analysis was performed to examine the protein expression of IL-6R in pulp tissues, while ELISA was used to determine the contents of IL-6R protein in the plasma and saliva samples. A dual luciferase reporter assay was performed to verify the interactions between IL-6R and miR-30b. The expression of IL-6R mRNA in the pulp tissues, plasma and saliva was significantly increased in patients with pulpitis compared with the control group. Similarly, the IL-6R protein expression in the samples from patients with pulpitis were also significantly increased compared with the control group. Conversely, the expression of miR-30b was significantly reduced in the samples from patients with pulpitis compared with the control group. The dual luciferase reporter assay revealed that miR-30b may bind with the 3'-untranslated seed region of IL-6R mRNA to regulate its expression. The present study demonstrated that the upregulated expression of IL-6R in pulp tissues, plasma and saliva from patients with pulpitis was associated with the downregulation of miR-30b expression. In addition, miR-30b may affect the progression of pulpitis via IL-6R and may be a potential genetic marker for the diagnosis of pulpitis.

Restoration of microRNA-30b expression alleviates vascular calcification through the mTOR signaling pathway and autophagy.

Pathological calcification represents an event that consequently leads to a distinct elevation in the morbidity and mortality of patients with chronic kidney disease (CKD) in addition to strengthening its correlation with hyperphosphatemia. Epigenomic regulation by specific microRNAs (miRNAs) is reported to be involved in ectopic calcification. However, the finer molecular mechanisms governing this event remain unclear. Hence, this study aimed to identify the potential miRNAs involved in vascular calcification (VC) development and progression. Initially, mitochondrial membrane potential (MMP), autophagy-specific markers (LC3II/LC3I and Beclin1) and phenotype-specific markers of osteoblasts (runt-related transcription factor 2 and Msx2) were measured to evaluate autophagy and VC in ß-glycerophosphate-induced vascular smooth muscle cells (VSMCs) with either miR-30b restoration or miR-30b knockdown performed in vitro. The VC in vivo was represented by calcified nodule formation in the aorta of the rats undergoing 5/6 nephrectomy followed by a 1.2% phosphorus diet using Alizarin Red staining. SOX9 was verified as the target of miR-30b according to luciferase activity determination. Restoration of miR-30b was revealed to markedly diminish the expression of SOX9 while acting to inhibit activation of the mTOR signaling pathway. Knockdown of miR-30b reduced MMP and autophagy, elevated VC, and suppressed the presence of rapamycin (an inhibitor of the mTOR signaling pathway). In addition, upregulated expression of miR-30b attenuated VC in vivo. Taken together, the key findings of this study identified the inhibitory role of miR-30b in VC, presenting an enhanced understanding of miRNA as a therapeutic target to curtail progressive VC in hyperphosphatemia of CKD.

LncRNA MALAT1 potentiates autophagy­associated cisplatin resistance by regulating the microRNA­30b/autophagy­related gene 5 axis in gastric cancer.

Gastric cancer (GC) is the fourth most common type of cancer worldwide and chemoresistance is a major obstacle to successful GC treatment. In the present study, reverse transcription­quantitative polymerase chain reaction analysis was used to measure the expression of metastasis­associated lung adenocarcinoma transcript 1 (MALAT1) and microRNA (miR)­30b. Western blot analysis was conducted to detect the protein expression of autophagy­related gene 5 (ATG5), p62 and LC3 (LC3­I and LC3­II). Cell viability and half maximal inhibitory concentration were determined by the Cell Counting Kit­8 assay. The green fluorescent protein (GFP)­LC3­positive cell percentage was determined by the GFP­LC3 puncta experiment. Luciferase reporter and RNA immunoprecipitation assays were used to explore the molecular associations among MALAT1, miR­30b and ATG5. MALAT1 was found to be highly expressed in CDDP­resistant AGS(AGS/CDDP) cells and CDDP­resistant HGC­27 (HGC­27/CDDP) cells. Cell viability was markedly increased in MALAT1­overexpressing AGS/CDDP cells, but was notably reduced in MALAT1­depleted HGC­27/CDDP cells. Moreover, MALAT1 potentiated CDDP resistance by facilitating autophagy in AGS/CDDP and HGC­27/CDDP cells. Further investigations demonstrated that MALAT1 inhibited miR­30b expression by direct interaction. Moreover, miR­30b abolished MALAT1­induced CDDP resistance by inhibiting autophagy in AGS/CDDP and HGC­27/CDDP cells. Furthermore, ATG5 was found to be a target of miR­30b. miR­30b weakened resistance to CDDP by inhibiting autophagy in AGS/CDDP and HGC­27/CDDP cells, while this effect was abrogated by increased ATG5 expression. Additionally, MALAT1 sequestered miR­30b from ATG5 to increase ATG5 expression in AGS/CDDP and HGC­27/CDDP cells. Therefore, MALAT1 potentiated autophagy­related CDDP resistance through suppressing the miR­30b/ATG5 axis in AGS/CDDP and HGC­27/CDDP cells, indicating that it may represent a promising target for the reversal of chemoresistance in GC.

MicroRNA-30b protects myocardial cell function in patients with acute myocardial ischemia by targeting plasminogen activator inhibitor-1.

The aim of the present study was to determine the expression of plasminogen activator inhibitor-1 (PAI-1) and microRNA (miR)-30b in the blood of patients with acute myocardial ischemia (AMI) and in the blood and myocardial tissue of mice with AMI. In addition, the present study aimed to identify the mechanism of action of miR-30b in AMI. A total of 36 patients with AMI were included in the present study and 28 healthy subjects were included as a control. Peripheral blood was collected from all subjects. For animal experiments, mice in the AMI group received an intraperitoneal injection of pituitrin (20 U/kg), whereas mice in the negative control group received an intraperitoneal injection of the same volume of saline. Blood and myocardial tissue was collected from all mice for analysis. Reverse transcription-quantitative polymerase chain reaction was performed to determine the expression of PAI-1 mRNA and miR-30b in the serum and myocardial tissue. An enzyme-linked immunosorbent assay was performed to measure the expression of PAI-1 protein in the serum of humans and mice, whereas western blotting was performed to determine the expression of PAI-1 protein in mouse myocardial tissue. Catalase, glutathione peroxidase and superoxide dismutase activity was measured using an automatic biochemical analyzer. A dual luciferase assay was performed to identify the interactions between PAI-1 mRNA and miR-30b. The results indicated that patients with AMI have higher PAI-1 levels and lower miR-30b expression in the peripheral blood compared with healthy subjects. AMI damaged the myocardium tissue of mice and reduced catalase, glutathione peroxidase and superoxide dismutase activity. Mice that have undergone AMI exhibit increased PAI-1 levels but decreased miR-30b expression in the peripheral blood and myocardial tissues. It was also demonstrated that miR-30b is able to bind to the 3'-untranslated region of PAI-1 mRNA to regulate its expression. The present study demonstrates that patients with AMI exhibit decreased miR-30b expression and elevated PAI-1 expression in the peripheral blood. miR-30b may therefore inhibit the damage to myocardial cells that occurs following AMI and protect myocardial cell function by targeting PAI-1 expression.

RETRACTED: MicroRNA-30b promotes lipopolysaccharide-induced inflammatory injury and alleviates autophagy through JNK and NF-κB pathways in HK-2 cells.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the Editor-in-Chief as there are concerns about the reliability of the results included in the article. The journal was initially contacted by the first author to request the retraction as they reported that results were not reproducible post publication. Also, the author acknowledged that the corresponding authors were not aware of the submission of this article. Given the comments of Dr Elisabeth Bik https://scienceintegritydigest.com/2020/02/21/the-tadpole-paper-mill/ regarding this article, the journal requested the author to provide the raw data. However, the author was not able to fulfil this request.

Regulatory role of microRNA-30b and plasminogen activator inhibitor-1 in the pathogenesis of cognitive impairment.

The present study aimed to investigate the role of plasminogen activator inhibitor-1 (PAI-1) in drug-induced early cognitive impairment and the underlying mechanism concerning microRNA (miR)-30b. A mouse model of cognitive impairment was established by intraperitoneal injection of scopolamine (2 mg/kg body weight) for 13 days. Behavioral performance was assessed using the Morris water maze (MWM) test. The mRNA expression levels of PAI-1 and miR-30b were detected using quantitative polymerase chain reaction (qPCR). The protein expression levels of PAI-1 in the hippocampus and blood were determined using western blot analysis and enzyme-linked immunosorbent assays. The MWM test demonstrated that, on days 3 and 4, the escape latency was significantly elevated in the model mice in comparison with control group (P<0.05). In addition, the length of swimming path was significantly increased (P<0.05), while the number of times of crossing the platform location was significantly reduced in the model mouse group (P<0.05) in comparison with the control group. qPCR demonstrated that the mRNA expression levels of PAI-1 in the model mice was significantly elevated in the hippocampus and blood in comparison with the control group (P<0.01). Furthermore, western blot analysis and enzyme-linked immunosorbent assay demonstrated that the protein expression levels of PAI-1 were significantly elevated in the hippocampus and blood in the model group, in comparison with the control group (P<0.05). Notably, the levels of miR-30b in the hippocampus and blood were significantly decreased in the model mice in comparison with the control group (P<0.01). To conclude, the expression levels of PAI-1 were significantly elevated in mice with scopolamine-induced cognitive impairment, which may be associated with the downregulation of miR-30b. The findings from the present study suggest that miR-30b may be involved in the regulation of PAI-1, which would contribute to the pathogenesis of cognitive impairment.

Anti-apoptotic effect of microRNA-30b in early phase of rat myocardial ischemia-reperfusion injury model.

This study aimed to investigate the effect of microRNA-30b (miR-30b) in rat myocardial ischemic-reperfusion (I/R) injury model. We randomly divided Sprague-Dawley (SD) rats (n = 80) into five groups: 1) control group; 2) miR-30b group; 3) sham-operated group; 4) I/R group, and 5) I/R+miR-30b group. Real-time quantitative polymerase chain reaction, immunohistochemical staining and Western blot analysis were conducted. TUNEL assay was employed for testing cardiomyocyte apoptosis. Our results showed that miR-30b levels were down-regulated in I/R group and I/R + miR-30b group compared with sham-operated group (both P < 0.05). However, miR-30b level in I/R + miR-30b group was higher than I/R group (P < 0.05). Markedly, the apoptotic rate in I/R group showed highest in I/R group (P < 0.05). Additionally, the results illustrated that protein levels of Bcl-2, Bax, and caspase-3 were at higher levels in ischemic regions in I/R group, comparing to sham-operated group (all P < 0.05), while Bcl-2/Bax was reduced (P < 0.05). Bcl-2 level and Bcl-2/Bax were obviously increased in I/R + miR-30b group by comparison with I/R group, and expression levels of Bax and caspase-3 were down-regulated (all P < 0.05). We also found that in I/R + miR-30b group, KRAS level was apparently lower and p-AKT level was higher by comparing with I/R group (both P < 0.05). Our study indicated that miR-30b overexpression had anti-apoptotic effect on early phase of rat myocardial ischemia injury model through targeting KRAS and activating the Ras/Akt pathway.