NEAT1 Enhances MPP+-Induced Pyroptosis In SH-SY5Y Cells Via Targeting Mir-5047/YAF2 Signaling

Parkinson’s disease (PD) is the second most frequent neurodegenerative disease. The aim of our study is to explore the role and regulatory mechanism of long non-coding RNA (lncRNA) NEAT1 in the MPP+-induced neuron pyroptosis. The levels of miR-5047 and YAF2 mRNA were determined through qRT-PCR. TUNEL staining was carried out to analyze neuronal apoptosis. Luciferase activity assay was accomplished to analyze the combination of miR-5047 and NEAT1 and YAF2 3’-UTR. Besides, the concentrations of IL-1β and IL-18 in supernatant were analyzed by ELISA assay. The levels of protein were examined through Western blot. NEAT1 and YAF2 expression were increased, while miR-5047 level was declined in the SH-SY5Y cells treated with MPP+. NEAT1 was a positively regulator for the SH-SY5Y cells pyroptosis induced by MPP+. In addition, YAF2 was a downstream target of miR-5047. NEAT1 promoted YAF2 expression via inhibiting miR-5047. Importantly, the promotion of NEAT1 to SH-SY5Y cells pyroptosis induced by MPP+ was recused by miR-5047 mimic transfection and YAF2 downregulation. In conclusion, NEAT1 was increased in the SH-SY5Y cells treated with MPP+, and it promoted the MPP+-induced pyroptosis through facilitating YAF2 expression by sponging miR-5047.

Characterization of XR_311113.2 as a MicroRNA Sponge for Pre-ovulatory Ovarian Follicles of Goats via Long Noncoding RNA Profile and Bioinformatics Analysis

Long noncoding RNAs (lncRNAs) were identified recently as a large class of noncoding RNAs (ncRNAs) with a length ≥200 base pairs (bp). The function and mechanism of lncRNAs have been reported in a growing number of species and tissues. In contrast, the regulatory mechanism of lncRNAs in the goat reproductive system has rarely been reported. In the present study, we sequenced and analyzed the lncRNAs using bioinformatics to identify their expression profiles. As a result, 895 lncRNAs were predicted in the pre-ovulatory ovarian follicles of goats. Eighty-eight lncRNAs were differentially expressed in the Macheng black goat when compared with Boer goat. In addition, the lncRNA XR_311113.2 acted as a sponge of chi-miR-424-5p, as assessed via a luciferase activity assay. Taken together, our findings demonstrate that lncRNAs have potential effects in the ovarian follicles of goats and may represent a promising new research field to understand follicular development.

Long noncoding RNA RP11-909N17.2 presages a poor prognosis of non-small cell lung cancer

BACKGROUND: Long non-coding RNAs (lncRNAs) were detected extraordinarily expressed in various tumors and could combine with microRNAs (miRNAs) to play important role in tumor cells. This study is to explore the role of lncRNA RP11-909N17.2 in NSCLC and discuss in what way it functions in NSCLC. METHODS: 120 NSCLC patients were enlisted in this study. Expression levels of lncRNA RP11-909N17.2 and miR-767-3p were detected and the correlation between lncRNA RP11-909N17.2 expression and the clinical data characteristics was analyzed. Prognosis potential of lncRNA RP11-909N17.2 was inferred with Kaplan-Meier and multivariate Cox regression assays. Biological functions of NSCLC cells were accessed by cell counting Kit-8, transwell migration and invasion assay. Mechanism of RP11-909N17.2 action on NSCLC cells was investigated by luciferase activity assay with wide-type or mutation. RESULTS: LncRNA RP11-909N17.2 has an ascendant expression while miR-767-3p has descended one in NSCLC tissue specimens and cells. Over-expression of lncRNA RP11-909N17.2 can shorten the overall survival period of NSCLC patients when compared with low expression. Knockdown of lncRNA RP11-909N17.2 suppressed biology function of NSCLC cell including proliferation, migration, and invasion. CONCLUSION: LncRNA RP11-909N17.2 can be developed into a prognostic index for NSCLC. LncRNA RP11-909N17.2 plays a promoting role in NSCLC cells possibly by binding miR-767-3p as a sponge.

miR-153-3p Targets βII Spectrin to Regulate Formaldehyde-Induced Cardiomyocyte Apoptosis

Background: Formaldehyde (FA) is ubiquitous in the environment and can be transferred to the fetus through placental circulation, causing miscarriage and congenital heart disease (CHD). Studies have shown that βII spectrin is necessary for cardiomyocyte survival and differentiation, and its loss leads to heart development defects and cardiomyocyte apoptosis. Additionally, previous studies have demonstrated that miRNA is essential in heart development and remodeling. However, whether miRNA regulates FA-induced CHD and cardiomyocyte apoptosis remains unclear.Methods: Using commercially available rat embryonic cardiomyocytes and a rat model of fetal cardiomyocyte apoptosis. Real-time quantitative PCR (RT-qPCR) and Western blot were performed to examine the level of miR-153-3p, βII spectrin, caspase 7, cleaved caspase7, Bax, Bcl-2 expression in embryonic cardiomyocytes and a rat model of fetal cardiomyocyte apoptosis. Apoptotic cell populations were evaluated by flow cytometry and Tunel. Luciferase activity assay and RNA pull-down assay were used to detect the interaction between miR-153-3p and βII spectrin. Masson's trichrome staining detects the degree of tissue fibrosis. Fluorescence in situ hybridization (FISH) and Immunohistochemistry were used to detect the expression of miR-153-3p and βII spectrin in tissues.Results: Using commercially available rat embryonic cardiomyocytes and a rat model of fetal cardiomyocyte apoptosis, our studies indicate that miR-153-3p plays a regulatory role by directly targeting βII spectrin to promote cardiomyocyte apoptosis. miR-153-3p mainly regulates cardiomyocyte apoptosis by regulating the expression of caspase7, further elucidating the importance of apoptosis in heart development. Finally, the results with our animal model revealed that targeting the miR-153-3p/βII spectrin pathway effectively regulated FA-induced damage during heart development. Recovery experiments with miR-153-3p antagomir resulted in the reversal of FA-induced cardiomyocyte apoptosis and fetal cardiac fibrosis.Conclusion: This study investigated the molecular mechanism underpinning the role of βII spectrin in FA-induced CHD and the associated upstream miRNA pathway. The study findings suggest that miR-153-3p may provide a potential target for the clinical diagnosis and treatment of CHD.

Down-Regulated hsa-miR-190b-5p is Correlated With BCL11A Expression in Pediatric β-Thalassemia

Background: Transcription factor BCL11A is a key regulator of hemoglobin switching in adult β-thalassemia. Several microRNAs (miRNAs) involve in the pathology of β-thalassemia by regulations of BCL11A. However, the expressions and regulators of BCL11A in pediatric β-thalassemia were unclear. Methods: 18 pediatric β- thalassemia patients and 11 healthy controls were selected in this study. We applied reverse transcript quantitative real time PCR (RT-PCR) to analyze the expression levels of hsa-miR-190b-5p and γ-globin in pediatric β-thalassemia patients and luciferase activity assay to find out the direct regulations of BCL11A . Correlation between hsa-miR-190b-5p and biochemical indicators, BCL11A was assessed by the Pearson’s correlation test. Results: The expression levels of γ-globin in pediatric β-thalassemia patients were significantly increased. Moreover, the expression levels of hsa-miR-190b-5p were significantly down-regulated in pediatric β-thalassemia patients. Furthermore, hsa-miR-190b-5p was negatively correlated with BCL11A expression in pediatric β-thalassemia patients. Through luciferase activity assay, we found that hsa-miR-190b-5p was directly interacted with BCL11A 3’UTR 499-506 regions. Conclusion: Our results suggested that hsa-miR-190b-5p played key roles in regulating of BCL11A expression, which might provide novel therapies in pediatric patients with β-thalassemia .

SINE Insertion in 5’ Flanking of Porcine IGFBP3 May Associated with Gene Expression and Phenotypic Variations

Background: Insulin-like growth factor binding proteins (IGFBPs) have been considered important candidate genes for economic traits due to their involvement in physiological processes related to growth and development. However, most of the current studies on genetic markers of IGFBPs have focused on SNPs, and large fragment insertion mutations such as retrotransposons have rarely been considered.Results: In total twelve retrotransposon insertion polymorphisms (RIPs) were confirmed using bioinformatics prediction combined with the PCR-based amplification. By linkage genetic analysis, IGFBP3-1-RIP and IGFBP3-2-RIP are completely linked, showing only three genotypes, SINE+/+/LINE-/-, SINE-/-/LINE+/+ and SINE+/-/LINE-/+. The age of 100 kg body weight and longissimus muscle thickness of Large white individuals of SINE+/+/LINE-/-genotype were significantly (P<0.05) higher than those of other two genotypes. However, corrected backfat thickness of SINE+/+/LINE-/- individuals were significantly (P<0.05) thinner than those of SINE+/+/LINE-/- genotype. The expression of the IGFBP3 gene in liver and backfat of 30-day Sujiang piglets with SINE+/+/LINE-/- genotype were significantly higher (P<0.05) than those with SINE-/-/LINE+/+ genotype by qPCR. After the core promoter region of the IGFBP3 gene was identified locating within 482bp upstream of ATG by using the dual-luciferase activity assay, further study was conducted to confirm the effect of SINE of IGFBP3-1-RIP and LINE of IGFBP3-2-RIP on the promoter activity of IGFBP3 based on the PGL3-Promoter-Enhancer. The result revealed that only SINE insertion was significantly increased (P<0.05) promoter activity of the IGFBP3 gene, indicating that the SINE may act as an enhancer to regulate the promoter activity of the IGFBP3 gene.Conclusions: Overall, this study identified 12 RIPs in IGFBP gene clusters. Furthermore, SINE insertions in 5’ flanking of IGFBP3 may associated with variations of gene expression and phenotype.

LncRNA DLGAP1-AS2 regulates miR-503/cyclin D1 to promote cell proliferation in non-small cell lung cancer

Background LncRNA DLGAP1-AS2 plays an oncogenic role in glioma, while its role in other cancers is unknown. This study aimed to study the role of DLGAP1-AS2 in non-small cell lung cancer (NSCLC). Methods Expression of DLGAP1-AS2 in NSCLC and paired non-tumor tissues from 64 NSCLC patients and the prognostic value of DLGAP1-AS2 for NSCLC were analyzed by performing a 5-year follow-up study. The interaction between DLGAP1-AS2 and miR-503 was confirmed by dual luciferase reporter assay, and their relationship was explored in NSCLC cells transfected with DLGAP1-AS2 expression vector or miR-503 mimic. The roles of DLGAP1-AS2 and miR-503 in regulating cyclin D1 expression were analyzed by RT-qPCR and Western blot. Cell proliferation was analyzed by CCK-8 assay. Results DLGAP1-AS2 was upregulated in NSCLC and predicted poor survival. Interaction between DLGAP1-AS2 and miR-503 was confirmed by dual luciferase activity assay. Overexpression experiments showed that DLGAP1-AS2 and miR-503 overexpression failed to significantly affect the expression of each other. Interestingly, DLGAP1-AS2 overexpression upregulated cyclin D1, a target of miR-503, increased cell proliferation and reduced the effects of miR-503 overexpression on cyclin D1 expression and cell proliferation. Conclusions DLGAP1-AS2 may regulate miR-503/cyclin D1 to promote cell proliferation in NSCLC.

lncRNA MAGI2-AS3 Exerts Antioncogenic Roles in Hepatocellular Carcinoma via Regulating the miR-519c-3p/TXNIP Axis

Introduction. Our work was aimed to explore the mechanisms of MAGI2 antisense RNA 3 (MAGI2-AS3) in regulating hepatocellular carcinoma (HCC) carcinogenesis. Methods. MAGI2-AS3, microRNA-519c-3p (miR-519c-3p), and thioredoxin interacting protein (TXNIP) levels in HCC were detected by the RT-qPCR method. Cell proliferation and apoptosis rate were measured using Cell Counting Kit-8 assay and flow cytometry assay. Relationship between MAGI2-AS3, TXNIP, and miR-519c-3p were analyzed via luciferase activity assay, RNA pull-down assay, and RNA immunoprecipitation assay. Mouse xenograft models of HCC were conducted to explore the roles of MAGI2-AS3 in vivo. Results. MAGI2-AS3 levels were elevated, and miR-519c-3p decreased in HCC. MAGI2-AS3 overexpression inhibits while its knockdown stimulates HCC cell growth through miR-519c-3p. Moreover, miR-519c-3p overexpression stimulates HCC cell growth. MAGI2-AS3 serves as competing endogenous RNA (ceRNA) of miR-519c-3p to regulate TXNIP in HCC. And, TXNIP upregulation weakened the influence of MAGI2-AS3 knockdown on HCC cell behaviors. Additionally, MAGI2-AS3 overexpression suppressed HCC tumor growth in vivo. Conclusion. MAGI2-AS3 inhibits HCC tumorigenesis through miR-519c-3p/TXNIP axis in vitro and in vivo, indicating MAGI2-AS3 plays a crucial role in HCC development.

miR-223 Inhibits the Polarization and Recruitment of Macrophages via NLRP3/IL-1β Pathway to Meliorate Neuropathic Pain

Background. miRNA is an essential factor in neuropathic pain. However, the underlying mechanism of miRNA in neuropathic pain remains unclear. Objective. To explore the potential role of miR-223 in neuropathic pain in a mice model of chronic sciatic nerve injury. Methods. Mice were divided into the sham group, CCI group, CCI + Lenti-vector group, and CCI + Lenti-miR-223 group. Flow cytometry was used to detect the neuronal apoptosis and the proportion of M1/M2 macrophages in each group. Western blot was used to detect the protein expression levels of ASC, caspase-1, IL-1β, and IL-18 in each group. Luciferase activity assay detects the binding of miR-223 and NLRP3. Macrophage chemotaxis experiments verified the anti-inflammatory effect of miR-223 in vitro. Results. The overexpression of miR-233 significantly reduced the neuropathic pain caused by CCI and reduced the apoptosis and inflammatory factor expression. miR-223 inhibits the expression of NLRP3 by directly binding to the 3′-untranslated region. Overexpression of miR-223 reduces the protein levels of NLRP3, ASC, caspase-1, IL-1β, and IL-18 in the spinal cord of CCI mice, increases the proportion of M2-type macrophages, and reduces the proportion of M1-type macrophages. Conclusion. miR-223 may facilitate the development of neuropathic pain in CCI mice by inhibiting NLRP3-mediated neuroinflammation.

Functional Evaluation of KEL as an Oncogenic Gene in the Progression of Acute Erythroleukemia

Background Acute erythroleukemia (AEL) is an infrequent subtype of acute myeloid leukemia (AML) with worse prognosis. Though the last decade has seen major advances in the novel features and genomic landscape in AEL, there is still a lack of specific therapeutic targets and effective treatment approaches for this disease.MethodsTCGA database was used to screen out the oncogene with specifically aberrant expression in AEL. Protein array was performed to explore the activated signaling pathways and targets that undergo phosphorylation modulation. A series of functional analyses in cell lines and mice models were performed to investigate the biological significance and clinical relevance of KEL regulation in AEL. CHIP, EMSA and dual-luciferase activity assay were performed to explore the upstream regulation mechanism of KEL.ResultsIn this study, the results showed that KEL promoted cell proliferation and its downregulation reversed drug resistance in AEL cells to JQ1. Our findings suggested that KEL contributed to gain of H3K27 acetylation and promoted erythroid differentiation induced by GATA1. Additionally, GATA1 and TAL1 as co-Transcription factors (TFs) modulated the expression of KEL. Maintaining cell viability and differentiation, KEL also played parts in the immune evasion of tumor cells.ConclusionsOur work expands the current knowledge regarding molecular mechanisms involved in cancer onset and progression, offering promising therapeutic target to broaden the treatment options.