lncRNA XLOC Research Articles

Transcriptomic profiling reveals salt-responsive long non-coding RNAs and their putative target genes for improving salt tolerance in upland cotton (Gossypium hirsutum)

Salt stress continues to exert notable impacts on the growth and development of cotton plants. In various biological processes of plants and in response to abiotic stress, long noncoding RNAs (lncRNAs) play a crucial regulatory role. In this study, we conducted transcriptome sequencing of cotton plants grown under salt stress and normal water conditions. A total of 519 lncRNAs and 75,376 messenger RNAs (mRNAs) were identified. Compared with mRNAs, lncRNAs have shorter transcript lengths and open reading frames, fewer exons, and lower expression levels. By comparing and analyzing the differentially expressed lncRNAs (DELs) and differentially expressed coding genes (DEGs) after salt treatment, we identified a total of 60 DELs and 217 DEGs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that these target genes are enriched primarily in intracellular organelles and participate in metabolic regulation pathways. Ultimately, lncRNA XLOC_043671, along with its target gene Gohir.D02G125200, and the lncRNA XLOC_034831, along with its target gene Gohir.A12G114800 was identified as a participant in salt tolerance regulation in cotton. RNA-seq and RTqPCR revealed that both these lncRNAs and their target genes exhibited significantly enhanced expression under salt stress. Subsequently, utilizing virus-induced gene silencing (VIGS) technology, we successfully silenced the two lncRNAs in cotton plants, which resulted in reduced expression levels of the corresponding target genes, rendering the plants less salt tolerant than did the control plants. Similarly, we successfully silenced these two target genes in cotton seedlings, and the silenced plants demonstrated a pronounced salt-sensitive phenotype. In summary, our transcriptome analysis revealed a significant number of differentially expressed lncRNAs and their corresponding coding genes. These findings lay the foundation for further investigating the mechanisms of lncRNAs and their target genes in the salt stress response of cotton.

A new tandem repeat-enriched lncRNA XLOC_008672 promotes gastric carcinogenesis by regulating G3BP1 expression.

Aberrant expression of forkhead box transcription factor 1 (FOXM1) plays critical roles in a variety of human malignancies and predicts poor prognosis. However, little is known about the crosstalk between FOXM1 and long noncoding RNAs (lncRNAs) in tumorigenesis. The present study identifies a previously uncharacterized lncRNA XLOC_008672 in gastric cancer (GC), which is regulated by FOXM1 and possesses multiple copies of tandem repetitive sequences. LncRNA microarrays are used to screen differentially expressed lncRNAs in FOXM1 knockdown GC cells, and then the highest fold downregulation lncRNA XLOC_008672 is screened out. Sequence analysis reveals that the new lncRNA contains 62 copies of 37-bp tandem repeats. It is transcriptionally activated by FOXM1 and functions as a downstream effector of FOXM1 in GC cells through invitro and invivo functional assays. Elevated expression of XLOC_008672 is found in GC tissues and indicates worse prognosis. Mechanistically, XLOC_008672 can bind to small nuclear ribonucleoprotein polypeptide A (SNRPA), thereby enhancing mRNA stability of Ras-GTPase-activating protein SH3 domain-binding protein 1 (G3BP1) and, consequently, facilitating GC cell proliferation and migration. Our study discovers a new uncharacterized lncRNA XLOC_008672 involved in GC carcinogenesis and progression. Targeting FOXM1/XLOC_008672/SNRPA/G3BP1 signaling axis might be a promising therapeutic strategy for GC.

Downregulation of lncRNA XLOC_032768 in diabetic patients predicts the occurrence of diabetic nephropathy.

LncRNA XLOC_032768 is reported to prevent renal tubular epithelial cells from cisplatin-induced apoptosis, suggesting its involvement in the development of kidney injury. The present study aimed to explore the role of XLOC_032768 in diabetic nephropathy (DN). The present study enrolled a total of 140 healthy controls (Control group) and 140 patients with type 2 diabetes (Diabetes group). Expression of XLOC_032768 in plasma from these participants was analyzed by performing RT-qPCR. The 140 diabetic patients were followed up for 5 years to monitor the occurrence of diabetic complications. The role of XLOC_032768 in predicting the occurrence of diabetic complications, including DN, diabetic cardiomyopathy (DC), diabetic retinopathy (DR), and diabetic foot (DF) were analyzed by plotting receiver operating characteristic curves and complication-free curves. On the day of admission, plasma levels of XLOC_032768 were not significantly different between Control and Diabetes groups. During follow-up, a total of 22, 15, 13, and 15 cases were diagnosed as DN, DC, DR, and DF, respectively. On the day of diagnosis, plasma levels of XLOC_032768 were only decreased in DN group, but not in other groups, compared to plasma levels of XLOC_032768 on the day of admission. Using plasma levels of XLOC_032768 on the day of admission as a biomarker, potential DN patients were effectively separated from patients with other potential complications and diabetic patients without complications. The 140 diabetic patients were grouped into high and low XLOC_032768 level groups. It was observed that low XLOC_032768 level group showed increased occurrence of DN, but not other complications, compared to high XLOC_032768 level group. Therefore, the downregulation of lncRNA XLOC_032768 in diabetic patients may predict the occurrence of DN.

Long noncoding RNA XLOC_006786 inhibits the proliferation, invasion and metastasis of osteosarcoma cells through NOTCH3 signaling pathway by targeting miR-491-5p.

Recent research has indicated that Long noncoding RNAs (LncRNAs) are crucial in many disorders, especially tumors. However, the exact role of LncRNA XLOC_006786 (LncRNA-SPIDR-2:1) in malignancies, especially in human osteosarcoma, is unclear. The results of RT‒qPCR, western blotting, CCK-8 assays, and Transwell assays showed that LncRNA XLOC_006786 inhibited osteosarcoma cell proliferation, invasion, and migration, indicating that it may be a tumor suppressor gene in osteosarcoma. We found that LncRNA XLOC_006786 negatively regulated NOTCH3, which is an oncogenic gene in osteosarcoma, as we previously reported. Bioinformatics analysis showed that miR-491-5p may be a direct target of LncRNA XLOC_006786, while NOTCH3 is a key target of miR-491-5p. Then, we verified that LncRNA XLOC_006786 could prevent lung metastatic osteosarcoma in vivo. Taken together, our research showed that LncRNA XLOC_006786 suppresses osteosarcoma proliferation, invasion, and metastasis through the NOTCH3 signaling pathway by targeting miR-491-5p.

LncRNA XLOC_015548 affects the proliferation and differentiation of myoblasts via the MAPK signaling pathway.

In recent years, an increasing number of studies have reported that long non-coding RNAs (lncRNAs) play essential regulatory roles in myogenic differentiation. In this study, a specific LncRNA XLOC_015548 (Lnc000280) was identified. However, little research has explored its mechanism of action by constructing XLOC_015548 gene editing cell models. In this study, relevant sequences were obtained according to the RNA-seq results. Subsequently, XLOC_015548 knockdown and over-expression lentiviral vectors were constructed, and the C2C12 myoblast cell line was transfected to prepare the XLOC_015548 gene-edited myoblast model. The in vitro analysis revealed that over-expression of XLOC_015548 significantly promoted the proliferation and differentiation of myoblasts and the formation of myotubes, whereas the opposite result was obtained in the knockdown group. XLOC_015548 regulated myogenic differentiation and affected the expression of myogenic differentiation regulators such as Myod, myogenin, and MyHC. Regarding the signaling pathway, we found that XLOC_015548 correlated with the phosphorylation level of MAPK/MEK/ERK pathway proteins. And the degree of phosphorylation was positively correlated with the protein expression of myogenic differentiation regulators. In conclusion, a new gene-edited myoblast model was constructed based on the lncRNA regulator XLOC_015548. The in vitro cell experiments verified that XLOC_015548 had regulatory effects on muscle growth and myoblast differentiation. These findings provide a laboratory foundation for the clinical application of lncRNAs as regulatory factors in the treatment of disuse muscle atrophy.

Identification and characterization of non-coding RNA networks in infected macrophages revealing the pathogenesis of F. nucleatum-associated diseases

BackgroundF. nucleatum, as an important periodontal pathogen, is not only closely associated with the development of periodontitis, but also implicated in systemic diseases. Macrophages may act as an important mediator in the pathogenic process of F. nucleatum infection. As non-coding RNAs (ncRNAs) have attracted extensive attention as important epigenetic regulatory mechanisms recently, we focus on the competing endogenous RNA (ceRNA) regulatory networks to elucidate the pathogenesis of F. nucleatum-associated diseases.ResultsWe screen abnormally expressed mRNAs, miRNAs, lncRNAs and circRNAs in macrophages after F. nucleatum infection via the whole transcriptome sequencing technology, including 375 mRNAs, 5 miRNAs, 64 lncRNAs, and 180 circRNAs. The accuracy of RNA-seq and microRNA-seq result was further verified by qRT-PCR analysis. GO and KEGG analysis show that the differentially expressed genes were mainly involved in MAPK pathway, Toll-like receptor pathway, NF-κB pathway and apoptosis. KEGG disease analysis reveals that they were closely involved in immune system diseases, cardiovascular disease, cancers, inflammatory bowel disease (IBD) et al. We constructed the underlying lncRNA/circRNA-miRNA-mRNA networks to understand their interaction based on the correlation analysis between the differentially expressed RNAs, and then screen the core non-coding RNAs. In which, AKT2 is controlled by hsa_circ_0078617, hsa_circ_0069227, hsa_circ_0084089, lncRNA NUP210, lncRNA ABCB9, lncRNA DIXDC1, lncRNA ATXN1 and lncRNA XLOC_237387 through miR-150-5p; hsa_circ_0001165, hsa_circ_0008460, hsa_circ_0001118, lncRNA XLOC_237387 and lncRNA ATXN1 were identified as the ceRNAs of hsa-miR-146a-3p and thereby indirectly modulating the expression of MITF.ConclusionsOur data identified promising candidate ncRNAs responsible for regulating immune response in the F. nucleatum-associated diseases, offering new insights regarding the pathogenic mechanism of this pathogen.

CeRNA regulates network and expression and SNP effect on NFKBIA of cashmere fineness

In this study, a total of 1140 Liaoning Cashmere Goats (LCG) were genotyped for single nucleotide polymorphism (SNP) of NFKBIA gene. There are 15 SNPs and 7 genotypes have been found, and G1547A (GG) genotype has been associated with cashmere fineness and cashmere yield. An integrated ceRNA regulatory network of NFKBIA gene was made. To prove NFKBIA and these non-coding RNAs (ncRNAs) may be related to cashmere fineness, we performed qPCR on these ncRNA in LCG coarse type skin (CT-LCG) and LCG fine type skin (FT-LCG). The result of qPCR showed lncRNA XLOC_011060 and ciRNA452 are at high expression level in CT-LCG, all miRNAs appear high expressed in FT-LCG, and mir-93 was the most significant difference between CT-LCG and FT-LCG. In addition, five miRNAs were selected for qPCR in different genotypes. The qPCR results showed that mir-93 might negatively regulate cashmere fineness and mir-17-5p may play a positive role in regulating cashmere fineness of individuals with G1355A (AG) genotype. These results demonstrated that NFKBIA gene is associated with cashmere fineness of LCG and G1547A (GG) genotype is the preferred marker genotype for cashmere fineness.

Identification of Diagnostic Exosomal LncRNA-miRNA-mRNA Biomarkers in Colorectal Cancer Based on the ceRNA Network.

Background: Colorectal cancer (CRC) is currently the fourth most common cancer worldwide. The roles of exosomal competing endogenous RNAs (ceRNAs) in CRC remain unclear. In this study, we constructed an exosomal ceRNA network to identify the core ceRNAs and investigate the diagnostic biomarkers in CRC. Methods and Patients: Serum exosomes were isolated from four CRC patients and two healthy donors by ultracentrifugation, and then subjected to RNA isolation, sequencing and microarray. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) analyses were performed to identify functional enrichment implications of differentially expressed exosomal mRNAs. TargetScan and miRanda were used for identifying the miRNA-mRNA and miRNA-LncRNA interactions. The predicted lncRNAs and mRNAs were intersected with the differentially expressed genes, for which the screening criterion was fold change >1.5 in the microarray. Differentially expressed exosomal miRNAs were identified in the GSE71008 dataset, and differentially expressed mRNAs (DEmRNAs) were further summarized from The Cancer Genome Atlas (TCGA) database. Results: A total of 1186 exosomal DEmRNAs, 2088 exosomal DElncRNAs and 29 exosomal miRNAs were detected in CRC patients compared to the healthy donors. Functional enrichment analysis suggested that exosomal DEmRNAs might participate in pathways related to carcinogenesis and development of cancer. An exosomal ceRNA regulatory network of CRC was constructed based on 40 lncRNAs, two miRNAs, and five mRNAs. Exosomal miR-150-5p and miR-10b-5p expression levels were increased in healthy donors compared with CRC patients in the GSE71008 dataset, and five DEmRNAs (TOMM70A, RBM48, BEND3, RHOBTB1, and ADAMTS2) were significantly upregulated in TCGA database. Two potential exosomal regulatory axes of lncRNA G016261-miR-150-5p-RBM48 and lncRNA XLOC_011677-miR-10b-5p-BEND3 were identified from the network. Conclusion: The current study revealed potential molecular biological regulation pathways and diagnostic biomarkers through the exosomal ceRNA regulatory network.

A study on the roles of long non-coding RNA and circular RNA in the pulmonary injuries induced by polystyrene microplastics

Microplastics (MPs) pollution has become a global concern due to its close relation to the environment and human health. Recently, more and more studies have pointed out the existence of MPs in the air, but its potential inhalation toxicity is unclear. Polystyrene Microplastics (PS-MPs) is one of the representative MPs. Besides, non-coding RNA plays crucial roles in regulating gene expression. Therefore, this study aims to provide new insights into the molecular exploration of PS-MPs inhalation. In this study, Sprague Dawley（SD）rats were treated with 100 nm, 500 nm, 1 μm and 2.5 μm PS-MPs for three days. And then intra-tracheal instillation of saline or 100 nm PS-MPs with 0, 0.5, 1 and 2 mg/200 μL were performed in SD rats every two days for two consecutive weeks. The deposition of PS-MPs was observed through immunofluorescence. Lung histological alternations were observed in haematoxylin and eosin (H&E) staining sections. The expressions of pro-inflammatory cytokines were quantified by ELISA and qPCR. Genome-wide transcriptomic profiling of long noncoding RNAs (lncRNAs), circular RNAs (circRNAs) in rats lung were done by ribosomal RNA depleted RNA sequencing and verified by qRT-PCR. We observed that 100 nm and 1 μm PS-MPs could deposite in the lungs. In addition, pathological examination shows alveolar destruction and bronchial epithelium arranged in a mess in PS-MPs groups. Furthermore, the expressions of pro-inflammatory cytokines IL-6, TNF-α and IL-1β were upregulated in PS-MPs exposed rats. Sequencing results showed that 269 circRNAs and 109 lncRNAs were differentially expressed in lung tissue of the saline and PS-MPs exposed rats. The upregulated expressions of lncRNA XLOC_031479, circRNA 014924 and circRNA 006603 and the downregulated expressions of lncRNA XLOC_014188 and circ003982 were identified by qRT-PCR in MPs group. The identified novel circRNAs and lncRNAs may paly important role in the development of lung inflammation caused by PS-MPs.

Resveratrol inhibits cell apoptosis by suppressing long noncoding RNA (lncRNA) XLOC_014869 during lipopolysaccharide-induced acute lung injury in rats.

BackgroundAcute lung injury (ALI) is a common clinical complication with a high mortality rate. Resveratrol (Res) has been shown to protect against ALI, but the role of long noncoding RNAs (lncRNAs) in this process is still unclear.MethodsMale rats (n=20) aged 7–8 weeks were randomly divided into four groups: control, lipopolysaccharide (LPS), LPS + Res, and LPS + dexamethasone (Dexa). Intragastric administration of Res (0.5 mg/kg) or Dexa (1.5 mg/kg) was performed 1 h before intraperitoneal injection of LPS (5 mg/kg). Lung tissue, serum, and bronchoalveolar lavage fluid were sampled 6 h after LPS treatment for inflammatory factor detection, pathological detection, lncRNA sequencing and bioinformatical analysis, and TdT-mediated dUTP Nick-End Labeling. Quantitative real time polymerase chain reaction and western blotting were used to verify the sequencing results. LPS, Res, and RNA interference were used in rat alveolar epithelial cells experiments to confirm the protective of Res/lncRNA against ALI.ResultsRes pretreatment inhibited lung injury and the increase of inflammatory cytokines induced by LPS. The differentially expressed lncRNAs and mRNAs (P<0.05 and |fold change| >2) were mainly involved in the signaling pathway of immunity, infection, signaling molecules and interactions. Among the lncRNAs and mRNAs, 26 mRNAs and 23 lncRNAs had high levels in lungs treated with LPS but decreased with Res, and 17 mRNAs and 27 lncRNAs were at lower levels in lungs treated with LPS but increased with Res. lncRNA and adjacent mRNA analysis showed that lncRNAs XLOC_014869 and the adjacent gene Fos, and the possible downstream genes Jun and Faslg were increased by LPS, but these changes were attenuated by Res. Pretreatment with Res reduced LPS-induced lung tissue apoptosis. Similarly, Res treatment and knockdown of lncRNA XLOC_014869 reduced LPS-induced apoptosis and the levels of Fos, c-Jun, and Fas-L.ConclusionsRes can inhibit the increase of lncRNAs XLOC_014869 caused by LPS stimulation and inhibit lung cell apoptosis. These effects may be due to lncRNA XLOC_014869 mediation of the pro-apoptotic factors (Fos, c-Jun, and Fas-L).

Effect of lncRNA XLOC_005950 knockout by CRISPR/Cas9 gene editing on energy metabolism and proliferation in osteosarcoma MG63 cells mediated by hsa-miR-542-3p.

Cancer cells use glucose via glycolysis to maintain tumor cell proliferation. However, the effect of long non-coding RNAs (lncRNAs) on glycolysis in osteosarcoma (OS) cells remains unclear. The present study aimed to investigate the involvement of the lncRNA XLOC_005950/hsa-microRNA (miR)-542-3p/phosphofructokinase, muscle (PFKM) axis in the regulation of glucose metabolism, cell proliferation and apoptosis in the progression of OS. lncRNA XLOC_005950, hsa-miR-542-3p and PFKM expression in OS tissues and cells was detected via reverse transcription-quantitative PCR analysis. CRISPR/Cas9 gene editing was used to knockout lncRNA XLOC_005950 expression in MG63 cells. Cell Counting Kit-8 assay, flow cytometry, PFKM activity, and glucose and lactic acid content determination were performed to assess the effects of lncRNA XLOC_005950 knockout and overexpression of hsa-miR-542-3p on the phenotypes of OS cells. The dual-luciferase reporter assay was performed to confirm the targeting associations between lncRNA XLOC_005950, hsa-miR-542-3p and PFKM. The results demonstrated that lncRNA XLOC_005950 expression was upregulated in OS tissues and cells. Functional experiments indicated that lncRNA XLOC_005950 knockout decreased PFKM activity, the intracellular glucose and lactic acid content, and cell proliferation, while increasing apoptosis of OS cells. Furthermore, lncRNA XLOC_005950 knockout upregulated hsa-miR-542-3p expression and downregulated PFKM expression. Overexpression of hsa-miR-542-3p suppressed PFKM expression. Furthermore, lncRNA XLOC_005950, as the molecular sponge of miR-542-3p in OS, modulated the downstream target gene, PFKM. Taken together, the results of the present study suggest that lncRNA XLOC_005950 knockout may inhibit the progression of OS via hsa-miR-542-3p-mediated regulation of PFKM expression.

Transcriptional Profiling Reveals the Regulatory Role of DNER in Promoting Pancreatic Neuroendocrine Neoplasms.

Wnt/β-catenin and NOTCH signaling contribute to the pathogenesis and growth of (PanNENs). The wnt and Notch signaling pathways form an integrated signaling device termed “wntch” and regulate stochastic cell fate decisions, suggesting the essentiality of Wnt/Notch interactions in disease progression. However, the function of Wnt/Notch interactions in PanNENs is unclear. We analyzed RNA sequencing (RNA-seq) data to identify differentially expressed lncRNAs, mRNAs and pathways according to enriched Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways associated with PanNENs. RNA-seq analysis revealed that the levels of the lncRNA XLOC_221242 and the mRNA encoding Delta/Notch-like epidermal growth factor (EGF)-related receptor (DNER) were significantly increased in tumor tissues compared with normal tissues (n = 3). Protein-protein interaction (PPI) prediction combined with transcriptional profiling data analysis revealed that DNER expression levels were positively correlated with those of DNA-binding factor (RBPJ), S phase kinase-associated protein 1 (Skp1), CTNNB1 and Cadherin-2 (CDH2), which promote PanNEN tumorigenesis and progression. These results were consistent with those of immunohistochemical analysis of DNER, RBPJ, SKP1, CTNNB1, and CDH2 expression (n = 15). These findings provide compelling clinical and molecular evidence supporting the conclusion that DNER and the related RBPJ, SKP1, CTNNB1, and CDH2 signaling contribute to PanNEN tumorigenesis and progression by activating wnt/Notch interactions.

Silencing of lncRNA XLOC_035088 Protects Middle Cerebral Artery Occlusion-Induced Ischemic Stroke by Notch1 Signaling.

Ischemic stroke represents one of the leading causes of mortality worldwide and especially in developing countries. It is crucial for finding effective therapeutic targets that protect the brain against ischemic injury. Long noncoding RNAs (lncRNAs) have emerged as major regulators of neurological diseases, and clarifying their roles in cerebral ischemic injury may provide novel targets for the treatment of ischemic stroke. We aimed to investigate the role of lncRNA-XLOC_035088 in middle cerebral artery occlusion (MCAO)-induced rat brain injury and oxygen-glucose deprivation (OGD)-reperfusion treated hippocampal neurons. In our findings, we found that XLOC_035088 expression was significantly upregulated in OGD-reperfusion treated hippocampal neurons and in different brain regions of MCAO-treated rats. XLOC_035088 silencing protected against MCAO-induced ischemic brain injury in vivo and OGD-induced hippocampal neuronal apoptosis in vitro. Intrahippocampal silencing of XLOC_035088 significantly decreased brain XLOC_035088 expression, reduced brain infarct size, and improved neurological function through inhibiting NOTCH1 following derepression of presenilin 2 (PSEN2). Taken together, this study provides evidence that the lncRNA XLOC_035088/PSEN2/Notch1 axis is involved in the pathogenesis of ischemic brain injury, and presents a promising therapeutic route for ischemic stroke.

Expression and Clinical Significance of LcnRNA XLOC_109948 in Patients with Acute Myeloid Leukemia

To detect the expression level of LncRNA XLOC_109948 in bone marrow and serum of patients with acute myeloid Leukemia (AML), to verify the consistency between the expression in bone marrow and serum and to explore the role of LncRNA XLOC_109948 expression in the occurrence a development of AML. Bone marrow and peripheral blood samples were collected from 62 patients with AML, including 36 patients with AML (AML group), 26 AML patients with complete remission (AML-CR group), and peripheral blood from 20 healthy persons (control group) were also collected. The expression level of LncRNA XLOC_109948 was detected by real-time quantitative fluorescence PCR (qRT-PCR), and the relationship between its expression and clinical characteristics was analyzed. The expression of LncRNA XLOC_109948 in bone marrow and serum of AML patients was higher than that of AML patients with complete remission and healthy people (P<0.001). And there was no statistically significant difference between the AML-CR group and control group (P>0.05). The expression of LncRNA XLOC_109948 significantly decreased when AML patients reached to CR, and significantly increased when the disease relapsed (P<0.05). The expression of LncRNA XLOC_109948 significantly correlated with the clinicopathologic parameters of cytogenetics (P<0.05), but not significantly correlated with sex, age, WBC count, blast in bone marrow, FAB classification and other clinical characteristics (P>0.05). The expression of LncRNA XLOC_109948 in bone marrow and serum of AML patients is high, and its expression in time and sequence is consistent between bone marrow and serum, which can reflect the occurrence, development, chemotherapy efficacy and prognosis of AML patients.

LncRNA XLOC_003810 promotes T cell activation and inhibits PD-1/PD-L1 expression in patients with myasthenia gravis-related thymoma.

This study aimed to investigate the effect of long non-coding RNA XLOC_003810 on the activation of CD4+ T cells and expression of PD-1/PD-L1 in patients with myasthenia gravis-related thymoma (MG-T). Thymus specimens and thymic mononuclear cells were obtained from MG and MG-T patients or cardiac surgery patients undergoing thoracotomy who were selected as negative controls (NC). XLOC_003810 expression was examined using quantitative real-time PCR (qRT-PCR). Frequency of CD4+ T cells and proportion of CD4+ PD-1+ T cells and CD14+ PD-L1+ monocytes were quantified by flow cytometry. The release of inflammatory cytokines was measured by qRT-PCR and enzyme-linked immunosorbent assay. Compared with the NC group, expression of XLOC_003810, frequency of CD4+ T cells and the production of inflammatory cytokines were increased in patients with MG and MG-T. XLOC_003810 overexpression significantly increased the frequency of CD4+ T cells, facilitated the production of inflammatory cytokines and decreased the proportion of CD4+ PD-1+ T cells and CD14+ PD-L1+ monocytes in the thymic mononuclear cells. In contrast, XLOC_003810 knockdown exerted the opposite effect. Together, XLOC_003810 promotes T cell activation and inhibits PD-1/PD-L1 pathway in patients with MG-T.

Novel lncRNA XLOC_032768 alleviates cisplatin-induced apoptosis and inflammatory response of renal tubular epithelial cells through TNF-α

The cellular and molecular mechanisms through which cisplatin induces nephrotoxicity have been investigated extensively. However, the role of long non-coding RNAs (lncRNAs) in cisplatin-induced nephrotoxicity is not well known. We explored the functions and underlying mechanisms of a novel lncRNA XLOC_032768 in cisplatin-induced nephrotoxicity. Cisplatin treatment resulted in the apoptosis of the renal tubular epithelial cells and inflammatory response in a mouse model and human renal proximal tubular epithelial cells (HK-2). The differentially expressed genes (DEGs) of the transcriptome data were determined, and the results showed that lncRNA XLOC_032768 expression was significantly repressed by cisplatin treatment. This result was validated by an RT-qPCR experiment on in vivo and in vitro models. The overexpression of XLOC_032768 significantly inhibited the cisplatin-induced apoptosis and inflammatory response in HK-2 cells and mouse exposed to cisplatin. RNA sequencing analysis further confirmed that XLOC_032768 could regulate tumor necrosis factor (TNF)-α in the cisplatin-induced apoptosis of HK-2 cells in trans-manner. TNF-α inhibition also ameliorated cisplatin-induced apoptosis of renal tubular epithelial cells and renal structural damage. As such, XLOC_032768 suppressed cisplatin-induced apoptosis and inflammatory response of renal tubular epithelial cells through TNF-α. LncRNA XLOC_032768 is a potential novel agent to reduce cisplatin-induced nephrotoxicity.

LncRNA XLOC_003810 modulates thymic Th17/Treg balance in myasthenia gravis with thymoma.

Imbalance of T helper 17 (Th17)/regulatory T (Treg) cells is involved in the pathogenesis of myasthenia gravis with thymoma (MG-T). Long non-coding RNAs (lncRNAs) are implicated in the regulation of Th17/Treg balance. This study was designed to explore the role of XLOC_003810, a novel lncRNA, in regulating the Th17/Treg balance in MG-T. The thymic CD4+ T cells were isolated from control subjects and MG-T patients. The Th17/Treg balance was evaluated by determining proportions of Th17 and Treg cells and expression of Th17- and Treg- associated molecules. Lentivirus-mediated silencing and overexpression of XLOC_003810 in CD4+ T cells were performed. The results showed that XLOC_003810 expression was higher in MG-T thymic CD4+ T cells than that in the control group. Furthermore, the ratio of Th17/Treg cells, proportion of Th17 cells and levels of Th17-associated molecules were significantly increased, whereas the proportion of Treg cells and levels of Treg-associated molecules were decreased in MG-T thymic CD4+ T cells. Importantly, the Th17/Treg imbalance in MG-T thymic CD4+ T cells was aggravated by XLOC_003810 overexpression, whereas it was attenuated by XLOC_003810 silencing. Collectively, XLOC_003810 modulates thymic Th17/Treg balance in MG-T patients, providing the scientific basis for the clinical targeted therapy of MG-T.

Novel lncRNA XLOC_032768 protects against renal tubular epithelial cells apoptosis in renal ischemia–reperfusion injury by regulating FNDC3B/TGF-β1

Renal ischemia–reperfusion injury is a leading cause of acute kidney injury, but its underlying mechanism remains poorly understood and effective therapies are still lacking. Here, we identified lncRNA XLOC_032768 as a novel target in renal ischemia–reperfusion injury by analyzing differentially expressed genes of the transcriptome data. PCR results show that XLOC_032768 was markedly downregulated in the kidney during renal ischemia–reperfusion in mice and in cultured kidney cells during hypoxia. Upon induction in vitro, XLOC_032768 overexpression repressed the expression of fibronectin type III domain containing 3B (FNDC3B) and tubular epithelial cells apoptosis. Administration of XLOC_032768 preserved FNDC3B expression and attenuated renal tubular epithelial cells apoptosis, resulting in protection against kidney injury in mice. Knockdown of FNDC3B markedly reduced the expression of TGF-β1 and apoptosis of renal tubular cells. Thus, XLOC_032768/FNDC3B/TGF-β1signaling pathway in ischemia–reperfusion injury may be targeted for therapy.

LncRNA XLOC_006390 promotes pancreatic carcinogenesis and glutamate metabolism by stabilizing c-Myc.

The role of lncRNAs in the regulation of glutamate metabolism and metabolic reprogramming of pancreatic cancer (PC) during nutrient deprivation is largely unknown. Our study found that alpha-ketoglutarate (aKG) levels were significantly reduced in the absence of XLOC_006390. We subsequently confirmed that the decrease in aKG was mainly due to the downregulation of glutamate dehydrogenase 1 (GDH1) at the mRNA level. Therefore, we first screened transcription factors targeting the GDH1 gene promoter and confirmed that c-Myc regulates GDH1 transcription. c-Myc binds to the promoter of GDH1 and activates its transcription. Downregulation of GDH1 mRNA levels by XLOC_006390 deletion could be rescued by overexpression of c-Myc. Overexpression of XLOC_006390 promoted the protein stability of c-Myc by blocking its ubiquitination. Clinically, XLOC_006390 was positively correlated with the mRNA level of GDH1, and c-Myc positively regulated GDH1 gene expression, which was tightly associated with PC patient prognosis. The dysregulated lncRNA/c-Myc axis increased glutamate metabolism, promoting PC progression to a higher stage. Therefore, XLOC_006390/c-Myc may be a potential target for PC, and its abnormal activation also indicates the progression of PC.

Knockdown of lncRNAXLOC_001659 inhibits proliferation and invasion of esophageal squamous cell carcinoma cells.

BACKGROUNDStudies have shown that long non-coding RNAs (lncRNAs) play a key role in almost all key physiological and pathological processes, including different types of malignant tumors. Our previous lncRNA microarray results have shown that lncRNA XLOC_001659 is upregulated in esophageal cancer (EC) tissues, with a fold change of 20.9 relative to normal esophageal tissues. But its effect and the molecular biological mechanisms on proliferation and invasion of EC cells remain unclear.AIMTo investigate the effect of lncRNA XLOC_001659 on esophageal squamous cell carcinoma (ESCC) cells and explore the molecular biological mechanisms involved.METHODSRT-qPCR assay was used to quantify the expression levels of lncRNAXLOC-001659 and miR-490-5p. The proliferative capacity of the cells was determined using CCK8 and colony formation assays, and the effect of lncRNAXLOC-001659 on the invasion of ESCC cells was determined by Transwell assay. Dual-luciferase reporter assay was used to detect the target genes of lncRNAXLOC-001659 and miR-490-5p.RESULTSThe results of RT-qPCR showed that the expression of lncRNAXLOC_001659 was upregulated in ESCC cells. CCK-8 assay showed that knockdown of lncRNAXLOC_001659 significantly inhibited ESCC cell proliferation. Colony formation and Transwell invasion assays showed that knockdown of lncRNAXLOC_001659 or overexpression of miR-490-5p significantly inhibited ESCC cell growth and invasion. Furthermore, lncRNAXLOC_001659 acts as an endogenous sponge by competitively binding to miR-490-5p to downregulate miR-490-5p. Further results confirmed that miR-490-5p targeted PIK3CA, and the recovery of PIK3CA rescued lncRNAXLOC_001659 knockdown or miR-490-5p overexpression-mediated inhibition of cell proliferation and invasion, which suggested the presence of an lncRNAXLOC_001659/miR-490-5p/PIK3CA regulatory axis.CONCLUSIONKnockdown of lncRNA XLOC_001659 inhibits proliferation and invasion of ESCC cells via regulation of miR-490-5p/PIK3CA, suggesting that it may play a role in ESCC tumorigenesis and progression.