miR-21 Mimic Research Articles

Interferon (IFN)-mediated antiviral activity against the hepatititis C (HCV) through microRNAs ( microRNAs ).

Background: MicroRNAs are a class of small non-coding RNA molecules that function through post-transcriptional regulation of gene expression by a process termed RNA interference (RNAi), and that also have to prominence as critical regulators in a wide array of mechanisms of cell physiology. Aim of the work: The study will attempt to evaluate the expression of several microRNAs in peripheral blood mononuclear cells (PBMCs) from patients with chronic hepatitis C (CHC) at 12 hours after the first injection of pegylated interferon in comparison with healthy controls. Patients and methods: forty patients with chronic hepatitis C virus infection (CHCV), their age ranged between (20-56) years, selected from the National Hepatology and Tropical Medicine Research Institute were included in this study, after 12 hours of the first interferon injection, and twenty healthy individuals were included to serve as controls. All the patients and controls were subjected to the following history, clinical examination, abdominal ultrasonography and collection of samples for routine laboratory investigations. CBCs and Taqman quantitative RT-PCR for MicroRNAs expression analysis of miR- 128a, miR 196a, miR- 196b, miR 296. Results: Our study revealed that the microRNAs had a higher levels of expression in cases of CHCV infection. Conclusion: Ourstudy concluded thatthere’s a highly significant increase in expression levels of IFN-induced microRNAs were observed in patients of microRNAs-128a, 196a, 196b, 296. Recommendations: The future use of miR inhibitors or mimics and / or siRNAs might be useful for the development of diagnostic and therapeutic strategies aimed at the recovering of protective innate responses in HCV infections.

Extracellular vesicles in oral squamous carcinoma carry oncogenic miRNA profile and reprogram monocytes via NF-κB pathway.

Extracellular vesicles (EVs) are carriers of different biomacromolecules that participate in cellular signaling and disease pathogenesis. Although it has been shown that EVs can play an active role in cellular communication and different stages of cancer progression, the role of EVs in oral squamous cell carcinoma (OSCC) cancer pathogenesis, especially in the crosstalk of cancer cells with immune cells is unknown. Here, we present a detailed analysis of findings regarding the profile of EVs in OSCC and the role of EVs and associated miRNAs in the crosstalk of malignant cells with monocytes. We demonstrate that EVs are detectable in significantly higher quantities in the plasma of patients with OSCC. Oncogenic miRNAs (such as miR-21, miR-27) were detectable in high quantities in the circulating EVs and plasma of patients with OSCC. EVs isolated from the circulation of OSCC patients and OSCC cell lines showed comparable miRNA signature, indicating the tumor origin of EVs in the circulation of patients with OSCC. Danger signals such as LPS and ethanol increased the production of EVs. EVs were taken up by monocytes after co-culture. Mechanistically, uptake of EVs derived from oral cancer cells by monocytes caused activation of the inflammatory pathway, NF-κB activation, and establishment of a pro-inflammatory and pro-tumorigenic milieu marked by increased levels of IL-6, CCL2, PEG2 and MMP9 levels. Series of experiments involving the introduction of exogenous oncogenic miR-21 mimic induced a similar pro-inflammatory and pro-tumorigenic profile in monocytes. Inhibiting miR-21 function in monocytes attenuated the pro-inflammatory phenotype of monocytes after EV challenge. These results indicate the role of EV-associated miR-21 in modulating the immune response in monocytes.

Increasing miR-150 and lowering HMGA2 inhibit proliferation and cycle progression of colon cancer in SW480 cells.

High mobility group protein A2 (HMGA2) is a kind of oncogene that regulates cell proliferation and cycle. HMGA2 up-regulation is related to the occurrence of multiple tumors including colorectal cancer. MiR-150 is found down-regulated in colorectal cancer tissue. Bioinformatics analysis shows the complementary targeted relationship between miR-150 and the 3'-UTR of HMGA2. This study explores the role of microRNA-150 (miR-150) in regulating HMGA2 expression, colorectal cancer cell proliferation, and cycle. Colorectal cancer patients were enrolled to collect cancer and para-carcinoma tissues. MiR-150 and HMGA2 expressions were tested in tissue. MiR-150, HMGA2, and Cyclin A levels in colorectal cancer cell line SW480, and normal colorectal epithelial cell line FHC were compared. The targeted relationship between miR-150 and the 3'-UTR of HMGA2 was evaluated by dual luciferase reporter gene assay. SW480 cells were divided into five groups, including miR-control, miR-150 mimic, small interfere normal control (si-NC), si-HMGA2, and miR-150 mimic + si-HMGA2. Cell cycle was determined by using flow cytometry. The cell proliferation was detected by using the cell counting kit 8 (CCK-8) test. HMGA2 expression was significantly increased, while miR-150 levels were significantly declined in colorectal cancer tissue compared with that in para-carcinoma tissue (p<0.05). HMGA2 and Cyclin A levels were higher significantly, whereas miR-150 expression was lower significantly in SW480 cells compared to that in FHC cells (p<0.05). MiR-150 targeted band to the 3'-UTR of HMGA. MiR-150 mimic and/or si-HMGA2 significantly reduced HMGA2 and Cyclin A expressions, blocked cell cycle in the G0/G1 phase, and attenuated cell proliferation. We observed that miR-150 down-regulated Cyclin A expression to block colorectal cancer cell cycle and inhibit proliferation through targeted inhibiting HMGA2.

MS25.05 Targeting the Micro-RNA: A New Therapeutic Venture

MS25.05 Targeting the Micro-RNA: A New Therapeutic Venture

MicroRNA-221 induces autophagy through suppressing HDAC6 expression and promoting apoptosis in pancreatic cancer.

Pancreatic cancer is an aggressive type of cancer with a poor prognosis, short survival rate and high mortality. Therefore, understanding the molecular mechanism underlying the aggressive growth of pancreatic cancer is of importance. An increasing number of studies suggest that numerous microRNAs (miRNAs/miRs) are associated with the tumorigenesis, progression and prognosis of tumors. In a recent study by the present authors, it was revealed that the expression of miR-221 was significantly downregulated in highly aggressive pancreatic cancer cells compared with weakly aggressive pancreatic cancer cells. In addition, miR-221 has been suggested as a novel tumor-associated miRNA, as it is involved in apoptosis, invasion, metastasis and autophagy of tumor cells. However, the function of miR-221 in pancreatic cancer remains yet to be investigated. In the present study, it was revealed that transfection with miR-221 mimic was able to significantly induce apoptosis and autophagy in pancreatic cancer cells compared with the negative control. The protein deacetylase histone deacetylase-6 (HDAC6) has emerged to be an important component in the cellular management of protein aggregates. Studies suggest that HDAC6 serves a function in the clearance of aggresomes, thereby implying a functional association between HDAC6 and autophagy. In the present study, it was revealed that transfection with miR-221 mimic was able to suppress the protein expression of HDAC6 in pancreatic cancer cells compared with the negative control. Immunofluorescence data suggested that the inhibition of HDAC6 was able to induce autophagy in pancreatic cancer cells. Additionally, the results of the present study suggest that the downregulation of miR-221 expression may serve an oncogenic function in the apoptosis and autophagy of pancreatic cancer cells by inducing the expression of HDAC6.

Colon Epithelial MicroRNA Network in Fatty Liver.

Background & Aims Intestinal barrier alterations are associated with fatty liver (FL) and metabolic syndrome (MetS), but microRNA (miR) signaling pathways in MetS-FL pathogenesis remain unclear. This study investigates an epithelial-focused miR network in colorectal cell models based on the previously reported MetS-FL miR trio of hsa-miR-142-3p, hsa-miR-18b, and hsa-miR-890. Methods Each miR mimic construct of MetS-FL miR trio was transfected into human colorectal cells, CRL-1790 or Caco-2. Global miRNome changes posttransfection were profiled (nCounter® Human v3 miRNA, NanoString Technologies). Changes in barrier (transepithelial electrical resistance, TEER) and epithelial cell junction structure (Occludin and Zona Occludens-1/ZO-1 immunofluorescence staining-confocal microscopy) were examined pre- and posttransfection in Caco-2 cell monolayers. A signaling network was constructed from the MetS-FL miR trio, MetS-FL miR-induced colorectal miRNome changes, ZO-1, and Occludin. Results Transfection of CRL-1790 cells with each MetS-FL miR mimic led to global changes in the cellular miRNome profile, with 288 miRs being altered in expression by more than twofold. Eleven miRs with known cytoskeletal and metabolic roles were commonly altered in expression by all three miR mimics. Transfection of Caco-2 cell monolayers with each MetS-FL miR mimic induced barrier-associated TEER variations and led to structural modifications of ZO-1 and Occludin within epithelial cell junctions. Pathway analysis incorporating the MetS-FL miR trio, eleven common target miRs, ZO-1, and Occludin revealed a signaling network centered on TNF and AKT2, which highlights injury, inflammation, and hyperplasia. Conclusions Colon-specific changes in epithelial barriers, cell junction structure, and a miRNome signaling network are described from functional studies of a MetS-FL miR trio signature.

2‐Methoxyestradiol attenuates chronic‐intermittent‐hypoxia‐induced pulmonary hypertension through regulating microRNA‐223

Pulmonary hypertension (PH) is prevalent in patients with obstructive sleep apnea (OSA) syndrome, and coexistence of PH and OSA indicates a worse prognosis and higher mortality. Chronic intermittent hypoxia (CIH) is the key pathogenesis of OSA. Also, microRNA-223 (miR-223) plays a role in the regulation of CIH-induced PH process. However, the detailed mechanism of CIH inducing PH is still unclear. This study aimed to investigate the pathological process of CIH associated PH and explore the potential therapeutic methods. In this study, adult Sprague-Dawley rats were exposed to CIH or normoxic (N) conditions with 2-methoxyestradiol (2-Me) or vehicle treatment for 6 weeks. The results showed that 2-Me treatment reduced the progression of pulmonary angiogenesis in CIH rats, and alleviated proliferation, cellular migration, and reactive oxygen species formation was induced by CIH in pulmonary artery smooth muscle cells (PASMCs). CIH decreased the expression of miR-223, whereas 2-Me reversed the downregulation of miR-223 both in vivo and in vitro. Furthermore, the antiangiogenic effect of 2-Me observed in PASMCs was abrogated by miR-223 inhibitor, while enhanced by miR-223 mimic. These findings suggested that miR-223 played an important role in the process of CIH inducing PH, and 2-Me might reverse CIH-induced PH via upregulating miR-223.

MicroRNA-21 inhibits lipopolysaccharide-induced acute lung injury by targeting nuclear factor-κB.

Acute lung injury (ALI) is a frequent, but severe complication following sepsis in patients with critical illness. The present study aimed to investigate the potential role of microRNA-21 (miR-21) in the regulation of inflammation in the ALI induced by lipopolysaccharide (LPS) in vitro and in vivo. The levels of inflammatory cytokines, tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β and IL-10, and the level of miR-21 expression were measured in the lungs of LPS-induced ALI rats and NR8383 alveolar macrophages (AMs). To confirm the regulatory effect of miR-21 in the inflammatory reactions of ALI, NR8383 cells were transfected with a mimic of miR-21 or an anti-miR-21 inhibitor, and the subsequent changes of the miR-21 level and the levels of inflammatory cytokines were detected. The underlying molecular mechanism was also investigated. LPS-induced ALI in rats resulted in significant overexpression of pro-inflammatory cytokines, TNF-α, IL-6 and IL-1β, and miR-21, but reduced the expression of the anti-inflammatory cytokine IL-10. LPS treatment also led to a higher expression level of miR-21 and increased secretion of pro-inflammatory cytokines in NR8383 cells in a time-dependent manner. Manipulation with the miR-21 mimic significantly suppressed the LPS-mediated induction of TNF-α, IL-6 and IL-1β in NR8383 cells, while that induction was upregulated when miR-21 expression was silenced via transfection with the anti-miR-21 inhibitor. Further mechanism experiments revealed that miR-21 regulates LPS-induced inflammation responses via the Toll-like receptor 4 and nuclear factor-κB (Nf-κB) signaling pathway. miR-21 negatively regulates inflammatory responses in LPS-induced ALI by targeting the NF-κB signaling pathway, providing further insight into the molecular mechanism of ALI progression.

Mir223 restrains autophagy and promotes CNS inflammation by targeting ATG16L1

ABSTRACTMicroglia are innate immune cells in the central nervous system (CNS), that supplies neurons with key factors for executing autophagosomal/lysosomal functions. Macroautophagy/autophagy is a cellular catabolic process that maintains cell balance in response to stress-related stimulation. Abnormal autophagy occurs with many pathologies, such as cancer, and autoimmune and neurodegenerative diseases. Hence, clarification of the mechanisms of autophagy regulation is of utmost importance. Recently, researchers presented microRNAs (miRNAs) as novel and potent modulators of autophagic activity. Here, we found that Mir223 deficiency significantly ameliorated CNS inflammation, demyelination and the clinical symptoms of experimental autoimmune encephalomyelitis (EAE) and increased resting microglia and autophagy in brain microglial cells. In contrast, the autophagy inhibitor 3-methylademine (3-MA) aggravated the clinical symptoms of EAE in wild-type (WT) and Mir223-deficienct mice. Furthermore, it was confirmed that Mir223 deficiency in mice increased the protein expression of ATG16L1 (autophagy related 16-like 1 [S. cerevisiae]) and LC3-II in bone marrow-derived macrophage cells compared with cells from WT mice. Indeed, the cellular level of Atg16l1 was decreased in BV2 cells upon Mir223 overexpression and increased following the introduction of antagomirs. We also showed that the 3’ UTR of Atg16l1 contained functional Mir223-responsive sequences and that overexpression of ATG16L1 returned autophagy to normal levels even in the presence of Mir223 mimics. Collectively, these data indicate that Mir223 is a novel and important regulator of autophagy and that Atg16l1 is a Mir223 target in this process, which may have implications for improving our understanding of the neuroinflammatory process of EAE.Abbreviations: 3-MA: 3-methylademine; ACTB/β-actin: actin, beta; ATG: autophagy related; ATG16L1: autophagy related 16-like 1 (S. cerevisiae); BECN1: beclin 1, autophagy related; CNR2: cannabinoid receptor 2 (macrophage); CNS: central nervous system; CQ: chloroquine; EAE: experimental autoimmune encephalomyelitis; FOXO3: forkhead box O3; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; H&E: hematoxylin and eosin; ITGAM: integrin alpha M; LPS: lipoplysaccharide; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; miRNAs: microRNAs; MS: multiple sclerosis; PPARG: peroxisome proliferator activated receptor gamma; PTPRC: protein tyrosine phosphatase, receptor type, C; RA: rheumatoid arthritis; SQSTM1: sequestosome 1; TB: tuberculosis; TIMM23: translocase of inner mitochondrial membrane 23; TLR: toll-like receptor.

The value of nonhuman primates for safety assessment of monoclonal antibodies

Although the relationship between cigarette smoke and lung cancer has been widely studied, the molecular mechanism for cigarette smoke-induced lung cancer remains largely unclear. The present study investigated the roles of hypoxia-inducible factor (HIF)-1α and miR-21 in the malignant transformation of human bronchial epithelial (HBE) cells induced by cigarette smoke extract (CSE). In case of acute and chronic treatment of HBE cells, CSE increased the levels of HIF-1α, p-Akt, p-NF-κB, and miR-21 and decreased PTEN levels. The increased miR-21 levels induced by CSE were prevented by down-regulation of HIF-1α. Further, elevated miR-21 suppressed PTEN levels, which decreased the levels of p-Akt and p-NF-κB. However, those changes were attenuated in cells co-transfected with HIF-1α siRNA and an miR-21 mimic. Silencing of HIF-1α or NF-κB decreased colony formation and the invasion and migration capacities of CSE-transformed HBE cells; however, up-regulation of miR-21 reversed these effects. These results indicate that the oncogenic capacity of HIF-1α in regulation of miR-21-inhibited PTEN in a manner dependent on the Akt/NF-κB pathway, a process that is involved in the CSE-induced malignant transformation of HBE cells. Thus, the present research has established a new mechanism for cigarette smoke-induced lung carcinogenesis.

K01 – Consumer toxicology: Natural is good, synthetic is bad

Although the relationship between cigarette smoke and lung cancer has been widely studied, the molecular mechanism for cigarette smoke-induced lung cancer remains largely unclear. The present study investigated the roles of hypoxia-inducible factor (HIF)-1α and miR-21 in the malignant transformation of human bronchial epithelial (HBE) cells induced by cigarette smoke extract (CSE). In case of acute and chronic treatment of HBE cells, CSE increased the levels of HIF-1α, p-Akt, p-NF-κB, and miR-21 and decreased PTEN levels. The increased miR-21 levels induced by CSE were prevented by down-regulation of HIF-1α. Further, elevated miR-21 suppressed PTEN levels, which decreased the levels of p-Akt and p-NF-κB. However, those changes were attenuated in cells co-transfected with HIF-1α siRNA and an miR-21 mimic. Silencing of HIF-1α or NF-κB decreased colony formation and the invasion and migration capacities of CSE-transformed HBE cells; however, up-regulation of miR-21 reversed these effects. These results indicate that the oncogenic capacity of HIF-1α in regulation of miR-21-inhibited PTEN in a manner dependent on the Akt/NF-κB pathway, a process that is involved in the CSE-induced malignant transformation of HBE cells. Thus, the present research has established a new mechanism for cigarette smoke-induced lung carcinogenesis.

MicroRNA-155 Suppresses Mesangial Cell Proliferation and TGF-\u03b21 Production via Inhibiting CXCR5-ERK Signaling Pathway in Lupus Nephritis

Increasing evidence shows miR-155 plays an important role in regulating inflammatory processes in systemic lupus erythematosus (SLE), especially in lupus nephritis (LN). Because the chemokine CXCL13 is implicated in the pathogenesis of LN, here we examined whether miR-155 can modulate the activity of CXCL13 or its receptor CXCR5. We determined the expression of CXCL13 in normal and MRL/lpr mice and found elevated levels of CXCL13 in the kidneys of MRL/lpr mice compared with normal kidneys. Besides, CXCL13 expression was mainly detected in the glomerulus, specifically to mesangial areas. We then transfected a miR-155 mimic in human renal mesangial cells (HRMCs) to overexpress miR-155 and detected decreased protein levels of CXCR5 by western blot analysis. Transfection of the miR-155 mimic into CXCL13-treated HRMCs resulted in a significantly reduced proliferation rate of HRMCs as measured by the cell-counting assay and flow cytometry. Moreover, increased intracellular miR-155 also led to decreased phosphorylation of ERK and TGF-β1 production. Together, these results revealed that miR-155 may play a role in the pathogenesis of LN.

Study on the correlation of the effect of microRNA-218 through Survivin on cholangiocarcinoma cells

Objective To study the effect of microRNA-218 (miR-218) on the invasion of cholangiocarcinoma cells by Survivin. Methods Two cholangiocarcinoma cell lines (CCLP1) and (QBC939) were cultured, and transfected with miR-218 mimicss. Negative control group and blank control group were set up. The invasiveness was determined via BD Martigel basement membrane matrix gel invasion Chamber. 3 results were randomly selected from each group. The CCLP1 and QBC939 cells were co transfected with miR-218 mimic and pMIR-Survivin, miR-218 mimic and pMIR-Survivin-mut, respectively. 3 results were randomly selected from each group, and the luciferase activity was detected by enzyme labelling instrument, and the correlation between miR-218 and Survivin was discussed. Results The numbers of CCLP1 cells transfected with miR-218, cells in negative control group, and cells in NC group were (28.33±4.16), (72.00±8.72) and (74.33±4.04) cells, respectively; The numbers of QBC939 cells transfected with miR-218, cells in negative control group, and cells in NC group were (7.33±2.08), (22.00±4.36) and (22.00±4.00) cells, respectively. Results showed that transfection of miR-218 mimics can inhibit the invasion ability of cholangiocarcinoma (The P values of CCLP1 cell line and QBC939 cell line were 0.000, 0.004). Luciferase activity detection results showed that luciferase activity in QBC939 wild type miRNA-218 group was 1.489±0.079, luciferase activity in negative control group was 2.825±0.068, luciferase activity in NC group was 2.747±0.081, luciferase activity in QBC939 mutant miRNA-218 group was 3.078±0.138, luciferase activity in negative control group was 2.944±0.144, the luciferase activity of the NC group was 2.807±0.090, the luciferase activity of the CCLP1 wild type miRNA-218 group was 1.322±0.039, the luciferase activity of the negative control group was 2.362±0.129, the luciferase activity of the NC group was 2.368±0.154, the luciferase activity of the CCLP1 mutant miRNA-218 group was 2.202±0.073, and the negative control group luciferase activity was 2.274±0.250, the activity of luciferase in group NC was 2.202±0.073. The luciferase activity of the wild type 3’untranslated regions (3’UTR) of the reporter gene vector was significantly lower than that of the negative control group and the blank control group (The P values of CCLP1 cell line and QBC939 cell line were 0.000, 0.000). While there was no significant change in the mutation type. Conclusion MiR-218 inhibits the invasion of cholangiocarcinoma cells and plays a negative regulatory role in the development of cholangiocarcinoma. miR-218 can interact with Survivin 3’UTR directly. Thus miR-218 can have a negative regulatory effect on cholangiocarcinoma through the Survivin directly. Key words: Cholangiocarcinoma; MicroRNA-218; Survivin; Cell invasion

MiR-145 eliminates lipopolysaccharides-induced inflammatory injury in human fibroblast-like synoviocyte MH7A cells.

Recently, it has been accepted that miR-based therapy may be beneficial for rheumatoid arthritis (RA). This study aimed to evaluate the potential involvement of miR-145 in RA in vitro. The expression of miR-145 in the human fibroblast-like synoviocyte line MH7A was overexpressed by miR-mimic transfection, after which cells were subjected to lipopolysaccharides (LPS). Cell viability, apoptosis, and the release of pro-inflammatory cytokines were measured. The result showed that the apoptosis and the release of IL-1β, IL-6, IL-8, and TNF-α were significantly induced by LPS. Meanwhile, LPS treatment led to downregulation of miR-145. miR-145 overexpression in LPS-untreated MH7A cells had no impacts on cell apoptosis and inflammation. But, restoring miR-145 expression in LPS-stimulated cells by supplementation of a miR-145 mimic protected MH7A cells against LPS-induced apoptosis and inflammation. Furthermore, miR-145 overexpression in LPS-untreated MH7A cells slightly blocked the PI3K/ATK and mTOR pathways, whereas miR-145 overexpression in LPS-stimulated cells notably repressed the LPS-induced activation of PI3K/ATK and MAPK/mTOR pathways. Our study suggested that miR-145 protected MH7A cells against LPS-induced apoptosis and inflammation by inhibiting the PI3K/AKT and MAPK/mTOR pathways.

The complex landscape of microRNAs in articular cartilage: biology, pathology, and therapeutic targets.

The disabling degenerative disease osteoarthritis (OA) is prevalent among the global population. Articular cartilage degeneration is a central feature of OA; therefore, a better understanding of the mechanisms that maintain cartilage homeostasis is vital for developing effective therapeutic interventions. MicroRNAs (miRs) modulate cell signaling pathways and various processes in articular cartilage via posttranscriptional repression of target genes. As dysregulated miRs frequently alter the homeostasis of articular cartilage, modulating select miRs presents a potential therapeutic opportunity for OA. Here, we review key miRs that have been shown to modulate cartilage-protective or -destructive mechanisms and signaling pathways. Additionally, we use an integrative computational biology approach to provide insight into predicted miR gene targets that may contribute to OA pathogenesis, and highlight the complexity of miR signaling in OA by generating both unique and overlapping gene targets of miRs that mediate protective or destructive effects. Early OA detection would enable effective prevention; thus, miRs are being explored as diagnostic biomarkers. We discuss these ongoing efforts and the applicability of miR mimics and antisense inhibitors as potential OA therapeutics.

SNHG12 Promotes Angiogenesis Following Ischemic Stroke via Regulating miR-150/VEGF Pathway

The promotion of angiogenesis is a promising therapeutic strategy for ischemic stroke. Many long noncoding RNAs (lncRNAs) are related to angiogenesis following ischemic stroke. LncRNA small nucleolar RNA host gene 12 (SNHG12) was upregulated in oxygen-glucose deprivation (OGD)-exposed primary brain microvascular endothelial cells and in microvessel from middle cerebral artery occlusion (MCAO) animal brains. However, the role and underlying mechanism of SNHG12 in ischemic stroke especially associated with angiogenesis process remain unknown. The expression of SNHG12 and miR-150 was determined in OGD-stimulated mouse brain microvascular endothelial (bEnd.3) cells. The role and mechanism of SNHG12 in the angiogenesis after ischemic stroke were investigated using gain- and loss-of function approaches both in OGD-exposed bEnd.3 cells and in MCAO mouse models. We found SNHG12 expression was elevated, whereas miR-150 reduced in OGD-exposed bEnd.3 cells. Upregulation of SNHG12 elevated, and SNHG12 knockdown suppressed the capillary-like tube formation, viability, migration, and VEGF expression in OGD-injured bEnd.3 cells. miR-150 mimic reversed, whereas anti-miR-150 further strengthened the effect of SNHG12 upregulation on the angiogenesis in bEnd.3 cells. Furthermore, we found that SNHG12 functioned as a competing endogenous RNA for miR-150 to regulate VEGF expression. Additionally, overexpression of SNHG12 improved the recovery of neurological function, reduced infarct volume and miR-150 expression, increased vascular density and VEGF expression in the infarct border zone of MCAO mice. In conclusion, SNHG12 promotes the angiogenesis following ischemic stroke via miR-150/VEGF pathway, which further clarified the mechanism of angiogenesis after ischemic stroke and provides a target for the treatment of this disease.

Targeted inhibition of β-catenin by miR-320 and decreased MMP-13 expression in suppressing chondrocyte collagen degradation.

Wnt/β-catenin pathway plays a critical role in modulating embryonic development, cell growth, and differentiation. The over-expression of β-catenin activates this pathway and up-regulates expression of matrix metalloproteinase-13 (MMP-13), and promotes matrix degradation and occurrence of osteoarthritis (OA). This study aims to explore the effect of miR-320 expression in OA chondrocyte and underlying mechanisms. Chondrocyte tissues from OA patients and normal individuals were collected for the detection of expression levels of miR-320, β-catenin, MMP-13, and alpha-1 chain of type II collagen (COL2A1). Dual luciferase reporter assay was performed to test targeted regulation between miR-320 and β-catenin. IL-1β was used to simulate in vitro cultured chondrocytes, which were transfected with miR-320 mimic and/or si-β-catenin, followed by quantification of miR-320, β-catenin, MMP-13, and COL2A1. In chondrocytes of OA patients, expression of microRNA (miR)-320 is decreased. Bioinformatics analysis revealed complementary binding sites between miR-320 and β-catenin. Compared to control group, increasing levels of β-catenin and MMP-13 expression with reduction of miR-320 and COL2A1 expressions were observed in OA chondrocytes. Transfection of miR-320 mimic and/or si-β-catenin depressed expression of β-catenin and MMP-13 inside chondrocytes, accompanied with elevation of COL2A1 expression. MiR-320 expression in OA chondrocyte is decreased, accompanied with up regulation of β-catenin and MMP-13. MiR-320 can inhibit β-catenin and MMP-13 expressions, elevates COL2A1 expression, which provides novel insights for the treatment of osteoarthritis.

MicroRNA-370 inhibits the proliferation, invasion and EMT of gastric cancer cells by directly targeting PAQR4

It has been reported that PAQR4 (Progestin and AdipoQ Receptor 4) expression is closely associated with progression of many cancers and microRNA (miRNA) processing. However, the effects and its precise mechanisms of PAQR4 in gastric cancer (GC) have not been well clarified. Our study aimed to explore the interaction between PAQR4 and miR-370 in GC. In our study, we found that the miR-370 level was significantly down-regulated in GC tissues and cell lines, and the expression of PAQR4 was dramatically increased. Interestingly, the low miR-370 level was closely associated with up-regulated PAQR4 expression in GC tissues. Moreover, introduction of miR-370 dramatically suppressed proliferation, invasion and EMT of GC cells. Whereas, miR-370 knockdown increased the proliferation, invasion and EMT in GC cells. We demonstrated that miR-370 could directly target PAQR4 by using both bioinformatics analysis and luciferase reporter assay. In addition, PAQR4 silencing had the similar effects with miR-370 overexpression on GC cells. Overexpression of PAQR4 in GC cells partially reversed the inhibitory effects of miR-370 mimic. miR-370 inhibited cell proliferation, invasion and EMT of GC cells by directly down-regulating PAQR4 expression, and miR-370 targeting PAQR4 was responsible for inhibition of the proliferation, invasion and EMT of GC cells.

Downregulation of microRNA-9 reduces inflammatory response and fibroblast proliferation in mice with idiopathic pulmonary fibrosis through the ANO1-mediated TGF-β-Smad3 pathway.

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease with increasing occurrence, high death rates and unfavorable treatment regimens. In the current study, we identified the expression of microRNA-9 (miR-9) and anoctamin-1 (ANO1) in IPF mouse models induced by bleomycin, and their effects on inflammation and fibroblast proliferation through the transforming growth factor-β (TGF-β)-Smad3 pathway. To verify the targeting relationship between miR-9 and ANO1, we used bioinformatics prediction and conducted a dual-luciferase reporter gene assay. The underlying regulatory mechanisms of miR-9 and the target gene ANO1 were investigated mainly with the treatment of miR-9 mimic, miR-9 inhibitor, or siRNA against ANO1 in fibroblasts isolated from IPF mice. Enzyme-linked immunosorbent assay was performed to investigate the effect of miR-9 or ANO1 on inflammatory factors. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and flow cytometry were used to detectfibroblast proliferation and apoptosis. Reverse transcription quantitative polymerase chain reaction and western blot analysis were applied to measure the expression of the TGF-β-Smad3 pathway-related genes. The determination of luciferase activity suggested that miR-9 targets ANO1. Upregulation of miR-9 or silencing of ANO1 intensified inflammation in IPF, promoted proliferation and inhibited apoptotic ability of lung fibroblasts. MiR-9 negatively modulated ANO1, and thus activated the TGF-β-Smad3 pathway. These findings suggest that miR-9 can indirectly activate the TGF-β-Smad3 pathway by inhibiting the expression of ANO1, thereby aggravating inflammation, promotes proliferation and suppressing apoptosis of lung fibroblasts in mice models of IPF.

MicroRNA-16 functions as a tumor-suppressor gene in oral squamous cell carcinoma by targeting AKT3 and BCL2L2.

Aberrant expressions of microRNAs have been reported to be strongly associated with the progression and prognosis of various tumors, including oral squamous cell carcinoma (OSCC). Recent studies on miRNA expression profiling have suggested that microRNA‐16 (miR‐16) may be dysregulated in OSCC. However, the tumorigenic roles and mechanisms of miR‐16 in OSCC are still largely unknown. In this study, we demonstrated that miR‐16 was specifically downregulated in both OSCC patients and cancer cell lines. In addition, functional roles of miR‐16 in vitro suggested that the miR‐16 mimic inhibited cell proliferation and induced apoptosis, whereas miR‐16 inhibitor displayed the opposite effects. Luciferase reporter assay and correlation analysis showed that AKT3 and BCL2L2 were directly targeted by miR‐16 and were inversely expressed with miR‐16 in OSCC. Moreover, restoration of AKT3 and BCL2L2 expression could partially reverse the cell proliferation inhibition and apoptosis induction caused by miR‐16. In xenograft nude mice, miR‐16 mimics decreased the expression of AKT3 and BCL2L2 and reduced the tumors volumes and weights, whereas the miR‐16 inhibitor exhibited adverse effects in the derived xenografts. In conclusion, the findings suggested that miR‐16 functions as a tumor suppressor miRNA to inhibit cell proliferation and induce apoptosis in OSCC through decreasing the oncogenes AKT3 and BCL2L2 and that miR‐16 could be a potential therapeutic target for OSCC.