Overexpression Of miR-21 Research Articles

2336P MiR-155 overexpression regulates epithelial-mesenchymal transition through repressing Quaking in lung cancer cells

2336P MiR-155 overexpression regulates epithelial-mesenchymal transition through repressing Quaking in lung cancer cells

MicroRNA-1 protects the endothelium in acute lung injury.

Acute lung injury (ALI) and its most severe form, acute respiratory distress syndrome (ARDS), cause severe endothelial dysfunction in the lung, and vascular endothelial growth factor (VEGF) is elevated in ARDS. We found that the levels of a VEGF-regulated microRNA, microRNA-1 (miR-1), were reduced in the lung endothelium after acute injury. Pulmonary endothelial cell-specific (EC-specific) overexpression of miR-1 protected the lung against cell death and barrier dysfunction in both murine and human models and increased the survival of mice after pneumonia-induced ALI. miR-1 had an intrinsic protective effect in pulmonary and other types of ECs; it inhibited apoptosis and necroptosis pathways and decreased capillary leak by protecting adherens and tight junctions. Comparative gene expression analysis and RISC recruitment assays identified miR-1 targets in the context of injury, including phosphodiesterase 5A (PDE5A), angiopoietin-2 (ANGPT2), CNKSR family member 3 (CNKSR3), and TNF-α-induced protein 2 (TNFAIP2). We validated miR-1-mediated regulation of ANGPT2 in both mouse and human ECs and found that in a 119-patient pneumonia cohort, miR-1 correlated inversely with ANGPT2. These findings illustrate a previously unknown role of miR-1 as a cytoprotective orchestrator of endothelial responses to acute injury with prognostic and therapeutic potential.

Nuclear miR-150 enhances hepatic lipid accumulation by targeting RNA transcripts overlapping the PLIN2 promoter

Alcohol-associated liver disease is a prevalent chronic liver disease caused by excessive ethanol consumption. This study aims to investigate the role of miR-150 in regulating hepatic lipid homeostasis in alcoholic fatty liver (AFL). miR-150 was mainly distributed in the nucleus of hepatocytes and correlated with the degree of liver injury. The decreased expression of miR-150 observed in AFL was a compensatory response to ethanol-induced hepatic steatosis. Overexpression of miR-150 facilitated hepatic lipid accumulation in cellulo and exacerbated ethanol-induced liver steatosis invivo. In silico analysis identified perilipin-2 (PLIN2) as a potential target gene of miR-150. miR-150 activated PLIN2 transcription by directly binding the RNA transcripts overlapping PLIN2 promoter and facilitating the recruitment of DNA helicase DHX9 and RNA polymeraseⅡ. Overall, our study provides fresh insights into the homeostasis regulation of hepatic steatosis induced by ethanol and identifies miR-150 as a pro-steatosis effector driving transcriptional PLIN2 gene activation.

MiR-21 attenuates FAS-mediated cardiomyocyte apoptosis by regulating HIPK3 expression.

MicroRNA-21 (miR-21) plays an anti-apoptotic role following ischemia-reperfusion (I/R) injury (IRI) in vivo; however, its underlying mechanism remains unclear. This study explored the effects of miR-21 and homeodomain interacting protein kinase 3 (HIPK3) on cardiomyocyte apoptosis induced by hypoxia/reoxygenation (H/R) in vitro. To this end, the rat cardiomyocyte H9C2 cell line was exposed to H/R and the roles of miR-21 and HIPK3 in regulating cell viability and apoptosis were evaluated by cell counting kit-8 assay, terminal-deoxynucleotidyl-transferase -mediated dUTP nick end labeling, and flow cytometry. Immunofluorescence and western blotting were performed to detect the expression/phosphorylation of apoptosis-related proteins. miR-21 expression was measured with quantitative real-time polymerase chain reaction. The putative interaction between miR-21 and HIPK3 was evaluated using the luciferase reporter assay. Our results showed that: (i) miR-21 overexpression or HIPK3 downregulation significantly attenuated H9C2 cells apoptosis after H/R; (ii) suppression of miR-21 expression promoted apoptosis; (iii) miR-21 overexpression inhibited HIPK3 expression; (iv) HIPK3 was the direct and main target of miR-21; (v) miR-21/HIPK3 formed part of a reciprocal, negative feedback loop; and (vi) HIPK3 downregulation decreased FAS-mediated apoptosis by inhibiting the phosphorylation of FADD, which subsequently inhibited the expression of BAX and cleaved caspase-3 and increased the expression of BCL2. Our study indicates that miR-21 attenuates FAS-mediated cardiomyocyte apoptosis by regulating HIPK3 expression, which could eventually have important clinical implications for patients with acute myocardial infarction.

MicroRNA-155 downregulates long noncoding RNA prostate cancer-associated transcript 29 in hepatocellular carcinoma to suppress cancer cell invasion and migration.

Long noncoding RNA (lncRNA) prostate cancer-associated transcript 29 (PCAT29) is known to suppress several cancers, but its participation in hepatocellular carcinoma (HCC) remains elusive. This study tried to explore PCAT29 function in HCC. In this study, a total of 62 HCC patients were enrolled. Tissue samples were collected from all 62 patients to isolate RNA samples. Quantitative reverse transcription polymerase chain reaction was applied for the expression analysis of PCAT29 and microRNA-155 (miR-155) in these tissue samples. The 62 HCC patients were followed up for 5 years to explore the prognostic value of PCAT29 for HCC. Correlations were analyzed using linear regression. IntaRNA 2.0 was used to predict the interaction between PCAT29 and miR-155. The role of PCAT29 and miR-155 in regulating HCC cell invasion and migration was evaluated by Transwell assay. We found that PCAT29 expression was downregulated in HCC and miR-155 expression was upregulated in HCC compared to nontumor samples (p < 0.001). Downregulation of PCAT29 was found to be closely associated with poor survival of HCC patients. MiR-155 was inversely correlated with PCAT29. It was predicted that miR-155 could target PCAT29. In HCC cells, miR-155 overexpression resulted in reduced PCAT29 expression (p < 0.05). MiR-155 counteracted the inhibitory effects of PCAT29 overexpression on HCC cell migration and invasion. These results suggest that PCAT29 may be a potential prognostic biomarker and a novel therapeutic target for treating HCC.

MicroRNA-375 Is Induced during Astrocyte-to-Neuron Reprogramming and Promotes Survival of Reprogrammed Neurons when Overexpressed.

While astrocyte-to-neuron (AtN) reprogramming holds great promise in regenerative medicine, the molecular mechanisms that govern this unique biological process remain elusive. To understand the function of miRNAs during the AtN reprogramming process, we performed RNA-seq of both mRNAs and miRNAs on human astrocyte (HA) cultures upon NeuroD1 overexpression. Bioinformatics analyses showed that NeuroD1 not only activated essential neuronal genes to initiate the reprogramming process but also induced miRNA changes in HA. Among the upregulated miRNAs, we identified miR-375 and its targets, neuronal ELAVL genes (nELAVLs), which encode a family of RNA-binding proteins and were also upregulated by NeuroD1. We further showed that manipulating the miR-375 level regulated nELAVLs' expression during NeuroD1-mediated reprogramming. Interestingly, miR-375/nELAVLs were also induced by the reprogramming factors Neurog2 and ASCL1 in HA, suggesting a conserved function to neuronal reprogramming, and by NeuroD1 in the mouse astrocyte culture and spinal cord. Functionally, we showed that miR-375 overexpression improved NeuroD1-mediated reprogramming efficiency by promoting cell survival at early stages in HA and did not appear to compromise the maturation of the reprogrammed neurons. Lastly, overexpression of miR-375-refractory ELAVL4 induced apoptosis and reversed the cell survival-promoting effect of miR-375 during AtN reprogramming. Together, we demonstrated a neuroprotective role of miR-375 during NeuroD1-mediated AtN reprogramming.

Small Nucleolar RNA Host Gene 9 Promotes the Development of Breast Cancer by Regulating the miR-326/Wnt5a/β-Catenin Axis

Breast cancer (BC) is a highly prevalent aggressive malignancy in women worldwide, and the search for key targets in its pathogenesis is a focus of research. Long non-coding RNAs (lncRNAs) play an important role in many cancers, including breast cancer. This study aimed to investigate the role of lncRNA SNHG9 in BC. The expression of SNHG9 in BC cells was found to be higher than that of human mammary epithelium. SNHG9 was found to inhibit the proliferation, migration, and invasion of cells and promote apoptosis. It was also found that SNHG9 regulates the miR-326/Wnt5a/β-catenin axis to promote the development of BC. Dual luciferase reporter and RNA pull-down assays confirmed the interaction between SNHG9, Wnt5a, and miR-326. Western blot analysis indicated that the expression of Wnt5a, β-catenin, c-myc, and cyclin D1 decreased significantly after the silencing of SNHG9 and the overexpression of miR-326. On the contrary, Wnt5a, β-catenin, c-myc, and cyclin D1 proteins were significantly up-regulated after inhibiting miR-326 expression. These findings suggest that SNHG9 is a promising target for BC therapy.

Identification and functional validation of SRC and RAPGEF1 as new direct targets of miR-203, involved in regulation of epidermal homeostasis

The epidermis is mostly composed of keratinocytes and forms a protecting barrier against external aggressions and dehydration. Epidermal homeostasis is maintained by a fine-tuned balance between keratinocyte proliferation and differentiation. In the regulation of this process, the keratinocyte-specific miR-203 microRNA is of the outmost importance as it promotes differentiation, notably by directly targeting and down-regulating mRNA expression of genes involved in keratinocyte proliferation, such as ΔNp63, Skp2 and Msi2. We aimed at identifying new miR-203 targets involved in the regulation of keratinocyte proliferation/differentiation balance. To this end, a transcriptome analysis of human primary keratinocytes overexpressing miR-203 was performed and revealed that miR-203 overexpression inhibited functions like proliferation, mitosis and cell cycling, and activated differentiation, apoptosis and cell death. Among the down-regulated genes, 24 putative target mRNAs were identified and 8 of them were related to proliferation. We demonstrated that SRC and RAPGEF1 were direct targets of miR-203. Moreover, both were down-regulated during epidermal morphogenesis in a 3D reconstructed skin model, while miR-203 was up-regulated. Finally silencing experiments showed that SRC or RAPGEF1 contributed to keratinocyte proliferation and regulated their differentiation. Preliminary results suggest their involvement in skin carcinoma hyperproliferation. Altogether this data indicates that RAPGEF1 and SRC could be new mediators of miR-203 in epidermal homeostasis regulation.

Microrna 138 Upregulation is Associated with Decreasing Levels of CCND1 Gene Expression and Promoting Cell Death in Human Prostate Cancer Cell Lines

Background: This research intended to discover the significance of miR-138 (microRNA 138) on the expression profile, proliferation, and the associated regulatory mechanisms in prostate cancer (PCa). Methods: Thirty-five specimens of prostate were studied to evaluate the expression level of miR138 by RT-qPCR (Quantitative reverse transcription polymerase chain reaction). Bioinformatics analysis was performed to search for the target genes of miR-138; and ABL1 (ABL proto-oncogene 1, non-receptor tyrosine kinase), CCND1 (cyclin D1), CCND3 (cyclin D3), VIM (vimentin), TWIST1 (twist family bHLH transcription factor 1), HIF1A (hypoxia-inducible factor 1 subunit alpha), and TERT (telomerase reverse transcriptase) genes were selected. Then, the biological role of miR-138 and CCND1 in the progression of PCa was investigated using RT-qPCR and luciferase reporter gene assay. Finally, overexpression of miR-138 on the proliferation in PCa cell lines was analyzed using the MTT (3-(4, 5-dimethylthiazol-2-Yl)-2, 5-diphenyltetrazolium bromide, Sigma, Germany) assay. Results: RT-qPCR showed that the expression of miR-138 downregulated in PCa tissues and cell lines. Bioinformatics analysis and RT-qPCR assay demonstrated that CCND1 expression level was negatively correlated with miR-138 in PCa tissues and the PC3 cell line. Moreover, CCND1 was predicted to be the target gene of miR138 in the PC3 cell line based on the results of luciferase reporter gene assay. Substantially, over-expression of miR138-5p mimic could inhibit the expression level of CCND1 gene in PC3 cell lines. Lastly, over-expression of miR-138 inhibited the proliferative capacities in PC3 and DU-145 cells. Conclusion: Our research introduces miR-138 as a negative regulator of CCND1 in the progression of PCa with an inhibitory impact on the proliferation rate of PCa cell lines. This regulatory mechanism could be utilized for the design and target selection of remedial miRNA-based approaches.

MiR-128–3p increases the radiosensitivity in nasopharyngeal carcinoma via regulating vascular endothelial growth factor C

PurposeThis study aims to investigate the role of miR-128–3p in the radiosensitivity of nasopharyngeal carcinoma (NPC) and its underlying mechanism. Methods6–10B cells were transfected with miR-128–3p mimic, pcDNA-VEGFC, and the corresponding negative control. C666–1 cells were transfected with miR-128–3p inhibitor, sh-VEGFC, and the corresponding negative control. RT-qPCR was used to determine the miR-128–3p and VEGFC mRNA expression level. Dual-luciferase assay was used to investigate the relationship between miR-128–3p and VEGFC. The protein levels of VEGFC, H2AX, γ-H2AX, p-P50, p-P65, p-IκB, and the apoptosis markers Bcl-2, caspase3, caspase9, and Bax were detected by Western blot. The proliferation activity was detected by CCK-8, and cell DNA damage was assessed by comet assay. The apoptosis rate was detected by flow cytometry. The growth of NPC in vivo was observed in mice through xenotransplantation. TUNEL staining was used to detect cell apoptosis in tumor tissues. ResultsmiR-128–3p was targeted and was negatively regulated with VEGFC. Overexpression of miR-128–3p or knockdown VEGFC significantly inhibited the proliferation of 6–10B and C666–1 cells, induced DNA damage and apoptosis and promoted the radiosensitivity of cells. Knocking down miR-128–3p or up-regulated VEGFC promoted the proliferation of C666–1 and 6–10B cells, reduced cell DNA damage and apoptosis, and enhanced cell resistance to radiotherapy. Overexpression of miR-128–3p reversed the effect of VEGFC on 6–10B cells and inhibited P50/P65/IKB signal pathway. In vivo, experiments in mice confirmed that miR-128–3p significantly inhibited NPC proliferation and promoted DNA damage and apoptosis by targeting VEGFC. ConclusionThe miR-128–3p pathway is a novel therapy target to overcome radiation resistance in NPC.

Vitamin A regulates dermal papilla cell proliferation and apoptosis under heat stress via IGF1 and Wnt10b signaling

Heat stress (HS) negatively affects the development of hair follicles. The present study investigated the effect of vitamin A (VA) on the development of rabbit dermal papilla cells (DPCs) under HS and the underlying regulatory mechanisms. Addition of 0.4 mg/L VA to the culture medium significantly enhanced cell proliferation (P < 0.001) and inhibited the apoptosis of DPCs (P < 0.01). VA decreased the proportion of DPCs in G0/G1 stage of the cell cycle under HS along with the expression of caspase 3, heat shock protein 70 (HSP70), and microRNA 195 (miR-195) (P < 0.05). VA also activated the insulin-like growth factor 1 (IGF1) and Wnt10b/β-catenin signaling pathways. The results of the dual luciferase reporter assay showed that IGF1 expression was modulated by miR-195–5p. Over-expression of miR-195–5p in DPCs with HS+VA treatment significantly reduced cell viability and IGF1 signaling (P < 0.01) and increased apoptosis (P < 0.01) compared with the HS+VA group. The positive effects of VA on proliferation and apoptosis of DPCs under HS were significantly attenu-ated by blocking Wnt10b and β-catenin signaling with IWP-2 and XAV-939, respectively. These results demonstrate that VA can promote hair follicle development following HS via modulation of miR-195/IGF1 and Wnt10b/β-catenin signaling pathways.

TAZ upregulates MIR-224 to inhibit oxidative stress response in multiple myeloma.

Oxidative stress within the bone marrow niche of multiple myeloma contributes to disease progression and drug resistance. Recent studies have associated the Hippo pathway with miRNA biogenesis and oxidative stress in solid tumors. Oxidative stress and miRNA pathway inter-relates in several cancers. Our group recently showed that TAZ functions as a tumor suppressor in MM. However, the role of TAZ in oxidative stress in MM is unknown. We sought to examine the role of TAZ in myeloma cells' response to BM oxidative stress. We postulated that TAZ might be associated with an oxidative stress phenotype and distinct miRNA signature in MM. Using human myeloma cell lines and clinical samples, we demonstrate that TAZ promotes myeloma cells' sensitivity to oxidative stress and anticancer-induced cytotoxicity by inducing miR-224 to repress the NRF2 antioxidant program in MM. We show that low expression of TAZ protein confers an oxidative stress-resistant phenotype in MM. Furthermore, we provide evidence that overexpression of miR-224 in myeloma cells expressing low amounts of TAZ protein inhibits cell growth and enhances sensitivity to anti-myeloma therapeutics. Our findings uncover a potential role for TAZ in oxidative stress response in MM via the miR-224-NRF2 molecular pathway. This provides the scientific ground to explore miR-224 as a potential molecular target to modify TAZ expression and enhance myeloma sensitivity to treatment.

Downregulated FTO Promotes MicroRNA-155-mediated Inflammatory Response in Cerebral Ischemia/Reperfusion Injury.

This study aimed to elucidate the mechanism for alteration of m6A RNA modification in cerebral ischemia/reperfusion(I/R) injury and identify novel therapeutic targets. A rat cerebral I/R injury model was established by middle cerebral artery occlusion (MCAO) followed by reperfusion. Changes in m6A RNA modification were evaluated by colorimetric quantification. The expression of the m6A methyltransferases METTL3, METTL14, and WTAP, and the demethylases FTO and ALKBH5 were determined using qPCR and western blot analyses. FTO was overexpressed in brain tissues via intracerebroventricular injection of adenoviruses encoding FTO. The protective effect of FTO on m6A RNA modification and cerebral I/R injury was assessed. MeRIP assays were used to detect the impact of FTO overexpression on m6A modification of pri-miR-155; qPCR analysis was used to identify its maturation. Finally, the role of miR-155 overexpression in the protective effects of FTO on cerebral I/R injury was examined. m6A levels of total RNA were increased, and m6A methyltransferase FTO expression was decreased in post-I/R injury cerebral tissues. FTO overexpression reversed the increase in m6A RNA modification and attenuated cerebral I/R injury. Furthermore, FTO overexpression increased the m6A modification of pri-miR-155 and enhanced its maturation to form miR-155. Notably, miR-155 overexpression blunted FTO's protective effect against cerebral I/R injury. We propose that downregulation of FTO expression contributes to increased m6A RNA modification in cerebral I/R injury. FTO overexpression reverses increased total m6A RNA modification and exerts a protective effect against cerebral I/R injury via downregulating m6A modification of pri-miR-155 to inhibit its maturation process.

Modifications of The Human Liver Cancer Cells through microRNA-145-Mediated Targeting of CDCA3.

Owing to the lethality of liver cancer, it is considered as one of the devastating types of cancers across the globe. Consistently, the study was designed to elucidate the role and to explore the therapeutic implications of miR-145 in human liver cancer. In the current experimental study, gene expression was determined by RT-PCR analysis. Transfection of cancer cells was carried out using Lipofectamine 2000. The cell proliferation of liver cancer cells was estimated by MTT assay. Clonogenic assay was performed for analysis of colony forming potential of cancer cells. Flow cytometry was done to analyze the cell cycle phase distribution of cancer cells. Transwell chamber assay was performed to assess the motility of cancer cells. Western blotting was done to estimate the expression levels of proteins. Dual luciferase assay was performed for interaction analysis of miR-145 with CDCA3. The miR-145 expression was found to be downregulated in liver cancer cells. The transfection mediated overexpression of miR-145 inhibited the cancer cell proliferation and when miR-145 inhibitor was transfected, cancer cells showed higher proliferation rates. Enrichment of miR-145 levels led to cell cycle arrest at G2/M phase by inhibiting cyclin B1. miR-145 also restricted the migration and invasion of cancer cells. CDCA3 was shown to be the intracellular target of miR-145 and it was found that the inhibitory effects of miR-145 were modulated through CDCA3, intracellularly. The current study clearly revealed that there is a need to investigate the regulatory role of different molecular entities like microRNAs in cancer development to better understand mechanics behind this pathogenesis and design more effective combating strategies against cancer.

Identification of the epigenetic mechanism for colorectal cancer stem cell regulation through the comprehensive microRNA expression profiling of human colorectal cancer tissues.

42 Background: MicroRNAs (miRNAs) are key regulators of cancer stem cell (CSC) functions, including self-renewal and differentiation. However, little is known about miRNAs that regulate such properties in human colorectal cancer. Methods: We compared the expression levels of 754 miRNAs between paired samples of EpCAM+/CD44+ cancer cells (enriched in CSCs) and EpCAM+/CD44neg cancer cells (non-tumorigenic cancer cells (NTCs)) sorted in parallel from human primary colorectal cancers by multiplex semi-quantitative PCR. Results: Comparison of the expression levels of miRNAs in the CSCs and NTCs resulted in the identification of 10 miRNAs upregulated (miR-221, miR-218, miR-500 and miR-195) or downregulated (miR-185, miR-19a, miR-337, miR-10a, miR-21 and miR-24) in CSCs compared to NTCs. Among them, miR-221 was most highly expressed in CSCs; and its expression level was very low in NTCs and normal colon epithelial cells (n=6, p&lt;0.05). Kaplan-Myer estimates and univariate and multivariate analyses showed that higher miR-221 expression was an independent prognosis factor for poorer outcome. Constitutive overexpression of miR-221 enhanced organoid-forming capacity of both conventional colorectal carcinoma cell lines and patient-derived xenografts (PDX) in vitro. Importantly, constitutive downregulation of miR-221 suppressed organoid-forming capacity in vitro and substantially reduced the tumorigenic capacity of CSC populations from PDX lines in vivo. miR-221 directly targeted Quaking (QKI) 5, the most abundant splicing isoform of the human QKI gene, and suppressed its protein level. Knockdown of miR-221 suppressed the clonogenicity of colorectal CSCs in vitro in a QKI-expression dependent manner and ability to form tumors in vivo. Conclusions: Our study reveals a mechanistic link between miR-221 and QKI and highlights their key role in regulating CSC properties in human colorectal cancer and proposes that miR-221 will be a promising marker and a specific therapeutic target for human colorectal CSCs.

Circulating miR-21 Overexpression Correlates with PDCD4 and IL-10 in Systemic Lupus Erythematosus (SLE): A Promising Diagnostic and Prognostic Biomarker.

Systemic Lupus Erythematosus (SLE) is a chronic autoimmune condition that affects multiple organs significantly impacts morbidity and mortality. The development of SLE is influenced by genetic predisposition and dysregulated immune response. Our objective was to investigate miR-21, IL-10, and PDCD4 expression in SLE patient plasma and analyze their correlations and potential diagnostic and prognostic values. The study included 100 healthy subjects, 50 newly diagnosed (ND), and 50 under-treatment (UT) SLE patients. The patients were observed for 24 weeks to track relapses. miR-21 and PDCD4 gene expression levels were measured using real-time RT-PCR, and IL-10 production was measured using ELISA. miR-21 and IL-10 expression levels were significantly greater in SLE patients than in healthy subjects, with the highest levels observed in ND patients. PDCD4 expression was also significantly greater in SLE patients than in subjects, with the highest levels observed in UT patients. ROC curve analyses and Cox-Mantel Log-rank tests indicated miR-21, PDCD4, and IL-10 as proper diagnostic and prognostic biomarkers for SLE. The study also revealed a significant positive correlation between miR-21 and PDCD4 and IL-10 levels in SLE patients. The studies suggest that dysregulation of miR-21, PDCD4, and IL-10 in patients with SLE may contribute to disease development and provides new diagnostic and prognostic markers. Additionally, the observed correlation between miR-21, PDCD4, and IL-10 levels in SLE patients signifies a potential interplay between these molecules.

MiR-21 Targets ASPP2 to Inhibit Apoptosis via CHOP-Mediated Signaling in Helicobacter pylori-Infected Gastric Cancer Cells.

Helicobacter pylori (H. pylori) infection affects cell survival pathways, including apoptosis and proliferation in host cells, and disruption of this balance is the key event in the development of H. pylori-induced gastric cancer (HPGC). H. pylori infection induces alterations in microRNAs expression that may be involved in GC development. Bioinformatic analysis showed that microRNA-21 (miR-21) is significantly upregulated in HPGC. Furthermore, quantitative proteomics and in silico prediction were employed to identify potential targets of miR-21. Following functional enrichment and clustered interaction network analyses, five candidates of miR-21 targets, PDCD4, ASPP2, DAXX, PIK3R1, and MAP3K1, were found across three functional clusters in association with cell death and survival, cellular movement, and cellular growth and proliferation. ASPP2 is inhibited by H. pylori-induced miR-21 overexpression. Moreover, ASPP2 levels are inversely correlated with miR-21 levels in HPGC tumor tissues. Thus, ASPP2 was identified as a miR-21 target in HPGC. Here, we observed that H. pylori-induced ASPP2 suppression enhances resistance to apoptosis in GC cells using apoptosis assays. Using protein interaction network and coimmunoprecipitation assay, we identified CHOP as a direct mediator of the ASPP2 proapoptotic activity in H. pylori-infected GC cells. Mechanistically, ASPP2 suppression promotes p300-mediated CHOP degradation, in turn inhibiting CHOP-mediated transcription of Noxa, Bak, and suppression of Bcl-2 to enact antiapoptosis in the GC cells after H. pylori infection. Clinicopathological analysis revealed correlations between decreased ASPP2 expression and higher HPGC risk and poor prognosis. In summary, the discovery of H. pylori-induced antiapoptosis via miR-21-mediated suppression of ASPP2/CHOP-mediated signaling provides a novel perspective for developing HPGC management and treatment.

MiR‑122/SENP1 axis confers stemness and chemoresistance to liver cancer through Wnt/β‑catenin signaling.

The property of inherent stemness of tumor cells coupled with the development of chemoresistance results in a poor prognosis for patients with liver cancer. Therefore, the present study focused on microRNA (miR)-122, a potential tumor suppressor, the expression of which has been previously shown to be significantly decreased and negatively associated with cancer cell stemness in liver cancer. The present study aimed to identify the molecular targets of miR-122 whilst uncovering the mechanism underlying chemoresistance and stemness of HepG2 cells in liver cancer. Bioinformatics online tools, such as ENCORI, coupled with dual-luciferase reporter assays in HepG2 cells, were used to identify and validate small ubiquitin-like modifier (SUMO) specific peptidase 1 (SENP1) as a potential target of miR-122 in liver cancer. The liver cancer stem cell population was determined using sphere formation assays and flow cytometry, whilst stem cell markers (Oct3/4, Nanog, B lymphoma Mo-MLV insertion region 1 homolog and Notch1) were detected by reverse transcription-quantitative PCR. Chemoresistance, cell proliferation and migratory ability of HepG2 cells were monitored using Cell Counting Kit-8, colony formation and Transwell assays, respectively. The overexpression of miR-122 by mimic transfection led to a significant decrease in the number spheres, downregulation of stem cell marker expression, the number of CD24+ cells, drug-resistance protein levels (P-glycoprotein and multidrug resistance protein), impaired chemoresistance, proliferation and migration of HepG2 cells. The transfection of SENP1 overexpression vector resulted in contrasting functions to miR-122 mimics, by partially reversing the effects induced by miR-122 mimic transfection in HepG2 cells. Wnt/β-catenin signaling has been proven to be involved in cancer stemness and malignant behavior. Western blotting analysis in HepG2 cells showed that the expression levels of both Wnt1 and β-catenin were significantly reduced after overexpressing miR-122, but increased after overexpressing SENP1. Co-transfection with the SENP1 overexpression vector reversed the suppression induced by the miR-122 mimics on Wnt1 and β-catenin expression. Co-immunoprecipitation, SUMOylation and half-life assays showed SENP1 interacted with β-catenin and decreased the SUMOylation of β-catenin, thereby enhancing its stability. Finally, tumor xenograft analyses revealed that HepG2 cells transfected with Agomir-122 exerted significantly lower tumor initiation frequency and growth rate, and a superior response to DOX in vivo, compared with those transfected with Agomir NC. Taken together, data from the present study miR-122/SENP1 axis can regulate β-catenin stability through de-SUMOylation, thereby promoting stemness and chemoresistance in liver cancer.

The Simultaneous Effects of miR-145-5p and hsa-let-7a-3p on Colorectal Tumorigenesis: In Vitro Evidence.

MicroRNAs (miRNAs) are a group of small regulatory non-coding RNAs, which are dysregulated through tumor progression. let-7 and MIR-145 are both tumor suppressor microRNAs that are downregulated in a wide array of cancers including colorectal cancer (CRC). This study was aimed to investigate the effect of simultaneous replacement of these two tumor suppressor miRNAs on proliferation, apoptosis, and migration of CRC cells. HCT-116 with lower expression levels of hsa-let-7a-3p and MIR-145-5p was selected for functional investigations. The cells were cultured and transfected with hsa-let-7a and MIR-145, separately and in combination. Cell viability and apoptosis rates were assessed by MTT assay and flow cytometry, respectively. Cell cycle status was further evaluated using flow cytometry and qRT-PCR was employed to evaluate gene expression. The obtained results showed that exogenous overexpression of MIR-145 and hsa-let-7a in HCT-116 cells could cooperatively decrease CRC cell proliferation and induce sub-G1 cell cycle arrest. Moreover, hsa-let-7a and MIR-145 co-transfection significantly increased apoptosis induction compared to separate transfected cells and control through modulating the expression levels of apoptosis-related genes including Bax, Bcl-2, P53, Caspase-3, Caspase-8, and Caspase-9. Furthermore, qRT-PCR results illustrated that hsa-let-7a and MIR-145 combination more effectively downregulated MMP-9 and MMP-2 expression, as the important modulators of metastasis, compared to the controls. Taken together, considering that exogenous overexpression of MIR-145 and hsa-let-7a showed cooperative anti-cancer effects on CRC cells, their combination may be considered as a novel therapeutic strategy for the treatment of CRC.

MIR-107/HMGB1/FGF-2 axis responds to excessive mechanical stretch to promote rapid repair of vascular endothelial cells

The increase of vascular wall tension can lead to endothelial injury during hypertension, but its potential mechanism remains to be studied. Our results of previous study showed that HUVECs could induce changes in HMGB1/RAGE to resist abnormal mechanical environments in pathological mechanical stretching. In this study, we applied two different kinds of mechanical tension to endothelial cells using the in vitro mechanical loading system FlexCell-5000T and focused on exploring the expression of miR-107 related pathways in HUVECs with excessive mechanical tension. The results showed that miR-107 negatively regulated the expression of the HMGB1/RAGE axis under excessive mechanical tension. Excessive mechanical stretching reduced the expression of miR-107 in HUVECs, and increased the expression of the HMGB1/RAGE axis. When miR-107 analog was transfected into HUVECs with lipo3000 reagent, the overexpression of miR-107 slowed down the increase of the HMGB1/RAGE axis caused by excessive mechanical stretching. At the same time, the overexpression of miR-107 inhibited the proliferation and migration of HUVECs to a certain extent. On the contrary, when miR-107 was silent, the proliferation and migration of HUVECs showed an upward trend. In addition, the study also showed that under excessive mechanical tension, miR-107 could regulate the expression of FGF-2 by HMGB1. In conclusion, these findings suggest that pathological mechanical stretching promote resistance to abnormal mechanical stimulation on HUVECs through miR-107/HMGB1/RAGE/FGF-2 pathway, thus promote vascular repair after endothelial injury. The suggest that miR-107 is a potential therapeutic target for hypertension.