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Project description:Purpose: In lung tissue of mice with experimental PH, we identified an upregulated lncRNA 5031425E22 which mapped to a human ortholog KMT2E-AS1. Across mammals, this lncRNA gene sits adjacent (head-to-head) to the chromosomal location of the histone lysine N-methyltransferase 2E gene (KMT2E), a member of a family of regulators controlling histone 3 lysine 4 trimethylation (H3K4Me3) and chromatin remodeling. We found that both mouse lncRNA E22 and human KMT2E-AS1 were increased in multiple in vivo rodent and human instances of PH. This study was designed to compare the RNA profile of human pulmonary arterial endothelial cells (HPAECs) treated with siRNA targeting KMT2E-AS1, KMT2E and negative control respectively. Method: Primary HPAECs were grown in EBM-2 basal medium supplemented with EGM-2 MV BulletKit (Lonza). Experiments were performed at passages 5 to 8. Cultured HPAECs were transfected with siRNAs targeting either negative control, KMT2E-AS1 or KMT2E, then exposed to hypoxic stress (0.2% oxygen) for 24 hours. Cells were harvested for total RNA extraction, and total RNA samples received ribosomal depletion and directional (stranded) RNA library prep. mRNA and lncRNA profiles were generated by deep sequencing, using NextSeq P3 flow cell (2 x 101bp, 40M reads/sample). RNA-Seq reads were mapped against the human genome build hg19 using STAR aligner and the read counts for each gene were calculated using featureCounts. Gene expression was normalized by DESeq2 with variance-stabilizing transformation (VST) and the low expression genes with total counts across all samples less than 10 were excluded. The differential gene expression analysis was performed using DESeq2 with the adjusted p-value cutoff of 0.05. The Benjamini-Hochberg method was used for multiple test correction. GSEA was conducted on differentially expressed genes with the enrichR R package [4] using the "GO_Biological_Process_2021", "Reactome_2022", "KEGG_2021_Human" databases.
2023-11-09 | GSE232797 | GEO
Project description:H3K4me3, mRNA and lncRNA profiling of hypoxic human pulmonary artery endothelial cells treated with KMT2E-AS1 siRNA
| PRJNA973979 | ENA
Project description:Hypoxic reprogramming of vasculature relies on genetic, epigenetic, and metabolic circuitry, but the control points are unknown. In pulmonary arterial hypertension (PAH), a disease driven by hypoxia inducible factor (HIF)-dependent vascular dysfunction, HIF-2α promoted expression of neighboring genes, long noncoding RNA (lncRNA) histone lysine N-methyltransferase 2E-antisense 1 (KMT2E-AS1) and histone lysine N-methyltransferase 2E (KMT2E). KMT2E-AS1 stabilized KMT2E protein to increase epigenetic histone 3 lysine 4 trimethylation (H3K4me3), driving HIF-2α-dependent metabolic and pathogenic endothelial activity. This lncRNA axis also increased HIF-2α expression across epigenetic, transcriptional, and posttranscriptional contexts, thus promoting a positive feedback loop to further augment HIF-2α activity. We identified a genetic association between rs73184087, a single-nucleotide variant (SNV) within a KMT2E intron, and disease risk in PAH discovery and replication patient cohorts and in a global meta-analysis. This SNV displayed allele (G)-specific association with HIF-2α, engaged in long-range chromatin interactions, and induced the lncRNA-KMT2E tandem in hypoxic (G/G) cells. In vivo, KMT2E-AS1 deficiency protected against PAH in mice, as did pharmacologic inhibition of histone methylation in rats. Conversely, forced lncRNA expression promoted more severe PH. Thus, the KMT2E-AS1/KMT2E pair orchestrates across convergent multi-ome landscapes to mediate HIF-2α pathobiology and represents a key clinical target in pulmonary hypertension.
| S-EPMC10947529 | biostudies-literature
Project description:mRNA and lncRNA profiling of hypoxic human pulmonary artery endothelial cells treated with siRNA-mediated knockdown of KMT2E-AS1 or KMT2E
| PRJNA973984 | ENA
Project description:The incidence of hypoxic pulmonary hypertension (HPH) is increasing. Accumulating evidence suggests that long noncoding RNAs (lncRNAs) play an important role in HPH, but the functions and mechanism have yet to be fully elucidated. In the present study, we established a HPH rat model with 8 h of hypoxia exposure (10% O2) per day for 21 days. High-throughput sequencing identified 60 differentially expressed (DE) lncRNAs, 20 DE miRNAs and 695 DE mRNAs in rat lung tissue. qRT-PCR verified the accuracy of the results. The DE mRNAs were significantly enriched in immune response, inflammatory response, leukocyte migration, cell cycle, cellular response to interleukin-1, IL-17 signalling pathway, cytokine-cytokine receptor interaction and Toll-like receptor signalling pathway. According to the theory of competing endogenous RNA (ceRNA) networks, lncRNA-miRNA-mRNA network was constructed by Cytoscape software, 16 miRNAs and 144 mRNAs. The results suggested that seven DE lncRNAs (Ly6l, AABR07038849.2, AABR07069008.2, AABR07064873.1, AABR07001382.1, AABR07068161.1 and AABR07060341.2) may serve as molecular sponges of the corresponding miRNAs and play a major role in HPH.
| S-EPMC8390787 | biostudies-literature
Project description:Background:Hypoxia following ischemic stroke is a common cause of brain insults. Mounting evidence suggests that long non-coding RNAs (lncRNAs) play a vital role in regulating certain physiological and pathological processes including ischemic stroke. For the first time, the present study investigated the effects and mechanism of LncRNA MACC1-AS1 on hypoxia-induced human brain microvascular endothelial cells (HBMECs). Methods:LncRNA MACC1-AS1 levels in HBMECs were detected via reverse transcription quantitative polymerase chain reaction (RT-qPCR) assay. Reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), and catalase (CAT), were detected using their respective kits. Flow cytometry and clone formation assay were performed to evaluate the effects of lncRNA MACC1-AS1 on cell apoptosis and cell proliferation respectively. Angiogenesis capacity was evaluated via tube formation assay. Transwell migration assay was performed for assessment of cell migration, Western blot assay was performed for measurement of Twist1 and VE-cadherin level, and permeability assay was performed for evaluation of the cell barrier function. The target gene was predicted via bioinformatics online tool and validated through luciferase reporter assay and RNA pull-down assay. Results:LncRNA MACC1-AS1 was downregulated in hypoxia-induced HBMECs. Overexpression of LncRNA MACC1-AS1 reduced cell apoptosis and oxidative stress, while promoting cell proliferation, migration, and angiogenesis. Moreover, LncRNA MACC1-AS1 overexpression reduced cell permeability and elevated VE-cadherin level, which contributed to maintaining cell barrier function. TWIST1 was validated as the target of miR-6867-5p which was further targeted by lncRNA MACC1-AS1. Thus, LncRNA MACC1-AS1 functions in hypoxia-induced HBMECs by regulating miR-6867-5p/TWIST1. Conclusions:In this study, we found that LncRNA MACC1-AS1 exerted a protective role in hypoxia-induced HBMECs via regulating miR-6867-5p/TWIST1, indicating a new therapeutic strategy for future ischemic stroke therapy.
| S-EPMC7396759 | biostudies-literature
Project description:HPAEC were treated with either siRNA to SOX17 or CRISPR activating or inhibiting the promoter or an enhancer region associated with pulmonary hypertension and sequenced.
2023-07-10 | GSE214742 | GEO
Project description:Transcriptional profiling of human pulmonary artery endothelial (HAPEC) and smooth muscle (PASM) cells comparing control untreated cells with cells transfected with ARSB-siRNA. Goal was to determine the effects of silenced of arylsufatase B on global gene expression in HAPEC and PASM cells.
2018-10-01 | GSE115268 | GEO
Project description:Malignant tumors are a grave threat to human health. Esophageal squamous cell carcinoma (ESCC) is a common gastrointestinal malignant tumor. China has a high incidence of ESCC, and its morbidity and mortality are higher than the global average. Increasingly, studies have shown that long noncoding RNAs (lncRNAs) play a vital function in the occurrence and development of tumors. Although the biological function of FOXP4-AS1 has been demonstrated in various tumors, the potential molecular mechanism of FOXP4-AS1 in ESCC is still poorly understood. The expression of FOXP4 and FOXP4-AS1 was detected in ESCC by quantitative real-time PCR (qRT-PCR) or SP immunohistochemistry (IHC). shRNA was used to silence gene expression. Apoptosis, cell cycle, MTS, colony formation, invasion and migration assays were employed to explore the biological functions of FOXP4 and FOXP4-AS1. The potential molecular mechanism of FOXP4-AS1 in ESCC was determined by dual-luciferase reporter, RNA immunoprecipitation (RIP) and chromatin immunoprecipitation (ChIP). Here, we demonstrated that FOXP4-AS1 was significantly increased in ESCC tissues and cell lines, associated with lymph node metastasis and TNM staging. Cell function experiments showed that FOXP4-AS1 promoted the proliferation, invasion and migration ability of ESCC cells. The expression of FOXP4-AS1 and FOXP4 in ESCC tissues was positively correlated. Further research found that FOXP4-AS1, upregulated in ESCC, promotes FOXP4 expression by enriching MLL2 and H3K4me3 in the FOXP4 promoter through a "molecular scaffold". Moreover, FOXP4, a transcription factor of β-catenin, promotes the transcription of β-catenin and ultimately leads to the malignant progression of ESCC. Finally, FOXP4-AS1 may be a new therapeutic target for ESCC.
| S-EPMC8712759 | biostudies-literature
Project description:To identify CLIC4 effectors by studying proteins expressiosn altered by CLIC4 overexpression in human pulmonary artery endothelial cells.
2018-10-16 | PXD008714 | Pride