Role Of Long Non-coding RNAs Research Articles

Contribution of signaling pathways associated with cellular senescence and regulatory non-coding RNAs to chronic obstructive pulmonary disease

Chronic obstructive pulmonary disease (COPD) is a multifactorial disease of the respiratory system that affects the lung parenchyma and airways; it is one of the leading causes of death in the world, which explains the constant search for new approaches to diagnosis, treatment and prevention of the disease. COPD develops as a result of a complex interaction of molecular genetic factors, a network of epigenetic regulators, and environmental factors that are closely related to lifestyle. The molecular pathogenesis of COPD may include the mechanisms of alteration of the regulation of stressful reactions that prevent cellular senescence.
 Non-coding RNAs play an important role in the regulation of various intracellular signaling pathways and is the most relevant subject of genetic studies of various pathological phenotypes. The expression profile of long non-coding RNA is often disregulated in various diseases. Information on the role of long non-coding RNA in the development of COPD is limited. Long non-coding RNA and the target-genes of signaling pathways involved in cellular senescence form a complex interactive network and may be targets for disease therapy.
 The review presents the data concerning some aspects of the molecular pathogenesis of COPD, as well as role of long non-coding RNAs in the development of the COPD.

Novel insights into the role of long non-coding RNA in the human malaria parasite, Plasmodium falciparum

The complex life cycle of Plasmodium falciparum requires coordinated gene expression regulation to allow host cell invasion, transmission, and immune evasion. Increasing evidence now suggests a major role for epigenetic mechanisms in gene expression in the parasite. In eukaryotes, many lncRNAs have been identified to be pivotal regulators of genome structure and gene expression. To investigate the regulatory roles of lncRNAs in P. falciparum we explore the intergenic lncRNA distribution in nuclear and cytoplasmic subcellular locations. Using nascent RNA expression profiles, we identify a total of 1768 lncRNAs, of which 718 (~41%) are novels in P. falciparum. The subcellular localization and stage-specific expression of several putative lncRNAs are validated using RNA-FISH. Additionally, the genome-wide occupancy of several candidate nuclear lncRNAs is explored using ChIRP. The results reveal that lncRNA occupancy sites are focal and sequence-specific with a particular enrichment for several parasite-specific gene families, including those involved in pathogenesis and sexual differentiation. Genomic and phenotypic analysis of one specific lncRNA demonstrate its importance in sexual differentiation and reproduction. Our findings bring a new level of insight into the role of lncRNAs in pathogenicity, gene regulation and sexual differentiation, opening new avenues for targeted therapeutic strategies against the deadly malaria parasite.

Exploiting Long Non-Coding RNAs and Circular RNAs as Pharmacological Targets in Triple-Negative Breast Cancer Treatment.

Breast cancer is one of the most frequent causes of cancer death among women worldwide. In particular, triple-negative breast cancer (TNBC) represents the most aggressive breast cancer subtype because it is characterized by the absence of molecular targets, thus making it an orphan type of malignancy. The discovery of new molecular druggable targets is mandatory to improve treatment success. In that context, non-coding RNAs represent an opportunity for modulation of cancer. They are RNA molecules with apparently no protein coding potential, which have been already demonstrated to play pivotal roles within cells, being involved in different processes, such as proliferation, cell cycle regulation, apoptosis, migration, and diseases, including cancer. Accordingly, they could be used as targets for future TNBC personalized therapy. Moreover, the peculiar characteristics of non-coding RNAs make them reliable biomarkers to monitor cancer treatment, thus, to monitor recurrence or chemoresistance, which are the most challenging aspects in TNBC. In the present review, we focused on the oncogenic or oncosuppressor role of long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) mostly involved in TNBC, highlighting their mode of action and depicting their potential role as a biomarker and/or as targets of new non-coding RNA-based therapeutics.

Long noncoding RNA CASC11 suppresses sorafenib-triggered ferroptosis via stabilizing SLC7A11 mRNA in hepatocellular carcinoma cells

As a frontline treatment for patients with advanced hepatocellular carcinoma (HCC), sorafenib is an effective drug approved by the Food and Drug Administration (FDA). Ferroptosis, a newly defined programmed cell death process with the hallmark of the accumulation of iron-dependent lipid peroxides, can be induced by sorafenib treatment. Our previous study identified oncogenic roles of long noncoding RNA (lncRNA) Cancer susceptibility candidate 11 (CASC11) in HCC progression. However, the relationship between CASC11 and sorafenib-induced ferroptosis in HCC remains unclear. In the present study, we aim to investigate the role of CASC11 in sorafenib-induced ferroptosis in HCC cell lines and determine the involved molecular mechanisms. Here, we demonstrated that sorafenib decreased CASCL11 expression. Knockdown of CASC11 enhanced sorafenib-induced ferroptosis, while overexpression of CASC11 exerted the opposite effect in HCC cells. Moreover, CASC11 led to the accumulation of intracellular malondialdehyde (MDA), lipid reactive oxygen species (ROS) and Fe2+ while depleting glutathione (GSH), thereby suppressing sorafenib-induced ferroptosis and cell death. Ferrostatin-1 (Ferr-1), a ferroptosis inhibitor, reversed the enhanced anticancer effect of sorafenib caused by the silence of CASC11 in HCC cells. Mechanistically, CASC11 upregulated the expression of solute carrier family 7 member 11 (SLC7A11) which is critical for ferroptosis inhibition. CASC11 associated with and stabilized SLC7A11 mRNA. In summary, our data revealed, for the first time, that CASC11 inhibits the sorafenib-induced ferroptosis in HCC cells via regulating SLC7A11, providing a new basis for clinical therapeutic strategies for patients with HCC.

The roles of long noncoding RNAs in atrial fibrillation

Atrial fibrillation (AF) is a common cardiac arrhythmia that often occurs in patients with structural heart disease and is a significant cause of morbidity and mortality in clinical settings. AF is typically associated with significant changes of both the structure of the atria and the cardiac conduction system. AF can result in reduced heart function, heart failure, and various other complications. Current drug therapy for AF patients is often ineffective and may have adverse effects. Radiofrequency ablation is more effective than traditional drug therapy, but this invasive procedure carries potential risks and may lead to postoperative recurrence, limiting the clinical benefits to some extent. Therefore, in-depth research into the molecular mechanisms of AF and exploration of new treatment strategies based on research findings are prerequisites for improving the treatment of AF and the associated cardiac conditions. Long noncoding RNAs (lncRNAs) are a new class of noncoding RNA (ncRNAs) with a length exceeding 200 nt, which regulate gene expression at multiple levels. Increasing evidence suggests that lncRNAs participate in many pathological processes of AF initiation, development, and maintenance, such as structural remodeling, electrical remodeling, renin-angiotensin system anomalies, and intracellular calcium deregulation s. LncRNAs that play key roles in structural and electrical remodeling may become molecular markers and targets for AF diagnosis and treatment, respectively, while lncRNAs critical to autonomic nervous system remodeling may bring new insights into the prognosis and recurrence of AF. This review article provides a synopsis on the up-to-date research findings relevant to the roles of lncRNAs in AF.

The role of long noncoding RNAs in liquid–liquid phase separation

Long noncoding RNAs (lncRNAs), which are among the most well-characterized noncoding RNAs, have attracted much attention due to their regulatory functions and potential therapeutic options in many types of disease. Liquid–liquid phase separation (LLPS), the formation of droplet condensates, is involved in various cellular processes, but the molecular interactions of lncRNAs in LLPS are unclear. In this review, we describe the research development on LLPS, including descriptions of various methods established to identify LLPS, summarize the physiological and pathological functions of LLPS, identify the molecular interactions of lncRNAs in LLPS, and present the potential applications of leveraging LLPS in the clinic. The aim of this review is to update the knowledge on the association between LLPS and lncRNAs, which might provide a new direction for the treatment of LLPS-mediated disease.

Roles of long non-coding RNAs in angiogenesis-related diseases: Focusing on non-neoplastic aspects

Angiogenesis is a key process in organ and tissue morphogenesis, as well as growth during human development, and is coordinated by pro- and anti-angiogenic factors. When this balance is affected, the related physiological and pathological changes lead to disease. Long non-coding RNAs (lncRNAs) are an important class of non-coding RNAs that do not encode proteins, but play a dynamic role in regulating gene expression. LncRNAs have been reported to be extensively involved in angiogenesis, particularly tumor angiogenesis. The non-tumor aspects have received relatively little attention and summary, but there is a broad space for research and exploration on lncRNA-targeted angiogenesis in this area. In this review, we focus on lncRNAs in angiogenesis-related diseases other than tumors, such as atherosclerosis, myocardial infarction, stroke, diabetic complications, hypertension, osteoporosis, dermatosis, as well as, endocrine, neurological, and other systemic disorders. Moreover, multiple cell types have been implicated in lncRNA-targeted angiogenesis, but only endothelial cells have attracted widespread attention. Thus, we explore the roles of other cells. Finally, we summarize the potential research directions in the area of lncRNAs and angiogenesis that can be undertaken by combining cutting-edge technology and interdisciplinary research, which will provide new insights into the involvement of lncRNAs in angiogenesis-related diseases.

The root of the mystery: the role of long non-coding RNAs in regulating transcription.

The root of the mystery: the role of long non-coding RNAs in regulating transcription.

LncRNA RBAT1 reduces chemosensitivity of cancer cells to carboplatin/paclitaxel by sponging miR‑27b in endometrial carcinoma

BackgroundA recent study reported the role of long non-coding RNA (lncRNA) RBAT1 in promoting the development of retinoblastoma and bladder cancer. However, its function in other cancers is unclear. We then studied the role of RBAT1 in endometrial carcinoma (EC).MethodsThe expression of RBAT1 and miR-27b in EC and paired non-tumor samples from advanced EC patients, as well as in plasma samples of EC patients and healthy controls were detected by RT-qPCR. The direct interaction between RBAT1 and miR-27b, and the subcellular location of RBAT1 were determined by RNA-RNA pulldown assay and subcellular fractionation assay, respectively.ResultsEC tissues showed increased expression levels of RBAT1 and decreased expression levels of miR-27b compared to that in non-tumor tissues. Moreover, EC patients showed higher plasma expression levels of RBAT1 and lower plasma expression levels of miR-27b compared to that in the controls. Drug-resistant (DR) patients showed higher expression levels of RBAT1 and lower expression levels of miR-27b in both EC tissues and plasma samples. RBAT1 was detected in both nuclear and cytoplasm and it directly interacted with miR-27b. RBAT1 and miR-27b did not affect the expression of each other. Upregulation of RBAT1 promoted the expression of multidrug-resistant-related protein (P-gp, MRP1, and BCRP). Overexpression of RBAT1 and inhibition of miR-27b promoted cell viability and impeded cell apoptosis and cell cycle arrest at G0-G1 phase, while knockdown of RBAT1 and overexpression of miR-27b inhibited cell viability and induced cell apoptosis and cell cycle arrest at G0-G1 phase. Moreover, miR-27b could abolish RBAT1-induced effects on cell viability, apoptosis and cell cycle.ConclusionRBAT1 may reduce the chemosensitivity of EC cells to carboplatin/paclitaxel by sponging miR-27b in EC.

Long non-coding RNA CCHE1 modulates LDHA-mediated glycolysis and confers chemoresistance to melanoma cells

Melanoma is considered as the most common metastatic skin cancer with increasing incidence and high mortality globally. The vital roles of long non-coding RNAs (lncRNAs) in the tumorigenesis of melanoma are elucidated by emerging evidence. The lncRNA cervical carcinoma high-expressed 1 (CCHE1) was overexpressed and acted as an oncogene in a variety of cancers, while the function of CCHE1 in melanoma remains unclear. Here, we found that CCHE1 was highly expressed in melanoma and correlated with the poorer survival of melanoma patients. Depletion of CCHE1 inhibited the proliferation, induced cell apoptosis and suppressed in vivo tumor growth. To further understand the functional mechanism of CCHE1, the interacting partners of CCHE1 were identified via RNA pull-down assay followed by mass spectrometry. CCHE1 was found to bind lactate dehydrogenase A (LDHA) and acted as a scaffold to enhance the interaction of LDHA with the fibroblast growth factor receptor type 1 (FGFR1), which consequently enhanced LDHA phosphorylation and activity of LDHA. Inhibiting CCHE1 strikingly suppressed the glycolytic flux of melanoma cells and lactate generation in vivo. Further study demonstrated that CCHE1 desensitized melanoma cells to dacarbazine and inhibition of glycolysis reversed CCHE1-induced chemoresistance. These results uncovered the novel function of CCHE1 in melanoma by reprogramming the glucose metabolism via orchestrating the activity of LDHA.

Long non-coding RNA MIR22HG inhibits the proliferation and migration, and promotes apoptosis by targeting microRNA-9-3p/ SOCS1 axis in small cell lung cancer cells.

This study aims to determine the role of long non-coding RNA (LncRNA) MIR22HG in small cell lung cancer (SCLC), and to explore its relevant mechanism. The expressions of genes and proteins in SCLC cells were examined applying qRT-PCR and western blot. Cell proliferation estimation was implemented utilizing cell counting kit-8 (CCK-8) and colony formation assays; the assessment of cell migration and invasion was operated employing Wound healing and Transwell; apoptosis evaluation was conducted adopting flow cytometric assay. Binding relationships was confirmed by luciferase reporter assay. Moreover, SCLC animal model was established to explore the role of MIR22HG in vivo. It was found that MIR22HG was declined and miR-9-3p was elevated in five SCLC cell lines (NCI-H446, NCI-H69, SHP-77, DMS79 and NCI-H345) in comparison with normal human bronchial epithelial cell line (NHBE). More interestingly, overexpression of MIR22HG resulted in decreased cell viability, declined colony formation, diminished capacities of cell migration and invasion in NCI-H446 and NCI-H345 cells but induced more apoptotic cells. However, these impacts were reversed by miR-9-3p upregulation. Meanwhile, MIR22HG could bind to miR-9-3p and negatively regulate its expression in SCLC. What's more, LncRNA MIR22HG overexpression was also testified to elevate SOCS1 via downregulating miR-9-3p expression. Furthermore, in vivo study further confirmed the role of MIR22HG/miR-9-3p in tumor regulation of SCLC. In conclusion, MIR22HG in SCLC was found to modulate miR-9-3p level and might act as a possible biomarker for SCLC treatment.

C-JUN-induced upregulation of LINC00174 contributes to colorectal cancer proliferation and invasion through accelerating USP21 expression.

Colorectal cancer (CRC) is one of the most common human malignancies due to its invasiveness and metastasis. Recent studies revealed the pivotal roles of long noncoding RNAs (lncRNAs) in tumorigenesis and progressions of various tumors. However, the biological roles and molecular mechanisms of long intergenic noncoding RNA 00174 (LINC00174) in human CRC remain unclear. Here, we report that LINC00174 expression was higher in human CRC tissues and cell lines than in adjacent normal tissues and a colon epithelial cell line (FHC). High expression of LINC00174 was positively correlated with poor overall and disease-free survival in patients with CRC. Loss- and gain-of-function of LINC00174 demonstrated its critical roles in promoting cell proliferation, apoptosis resistance, migration, and invasion of CRC cells in vitro. Moreover, overexpression of LINC00174 enhanced tumor growth in vivo. Mechanistic experiments revealed that LINC00174 could bind to microRNA (miR)-2467-3p and augment the expression and function of ubiquitin-specific peptidase 21 (USP21). Rescue assays found that miR-2467-3p inhibition can offset the actions of LINC00174 or USP21 knockdown in CRC cells. Additionally, transcriptional factor c-JUN transcriptionally activated LINC00174 expression and mediated LINC00174-induced malignant phenotypes of CRC cell lines. Totally, our findings shed light on a new therapeutic strategy in modulating LINC00174/miR-2467-3p, which may interfere with the expression of USP21, and revealed that LINC00174 could be a new therapeutic target or prognostic marker in CRC.

LncRNA ITGB2-AS1 promotes cisplatin resistance of non-small cell lung cancer by inhibiting ferroptosis via activating the FOSL2/NAMPT axis

ABSTRACT Cisplatin resistance is a major therapeutic challenge in non-small cell lung cancer (NSCLC). Herein, the regulatory role of long non-coding RNA (lncRNA) ITGB2-AS1 in regulating NSCLC cisplatin resistance was investigated. NSCLC cisplatin resistance cells were constructed using A549 and H1975 cells. Cell viability and proliferation were detected by MTT assay and colony formation assay, respectively. Cell apoptosis and cell cycle were examined by flow cytometry. GSH, MDA, ROS, and Fe2+ levels were measured by the corresponding kits. The expressions of ferroptosis-negative regulation genes (GPX4 and SLC7A11) were determined by qRT-PCR and western blot. Molecular interactions were analyzed by RNA pull-down, RIP, ChIP, and dual-luciferase reporter assays. The effects of ITGB2-AS1 silencing on NSCLC cisplatin resistance in vivo were elevated by the tumor xenograft experiment. ITGB2-AS1 expression was increased in NSCLC patients and cisplatin-resistant NSCLC cells, which was positively correlated with ferroptosis-negative regulation genes. ITGB2-AS1 knockdown suppressed resistant cell proliferation and promoted cell apoptosis and ferroptosis. ITGB2-AS1 increased NAMPT expression by binding to FOSL2, thereby repressing p53 expression. The ITGB2-AS1 knockdown also inhibited NSCLC cisplatin resistance in vivo. ITGB2-AS1 promoted NSCLC cisplatin resistance by inhibiting p53-mediated ferroptosis via activating the FOSL2/NAMPT axis.

Genome-wide analysis of long noncoding RNAs and their association in regulating the metamorphosis of the Sarcophaga peregrina (Diptera: Sarcophagidae).

The flesh fly, Sarcophaga peregrina (Diptera: Sarcophagidae), is an important hygiene pest, that causes myiasis in humans and other mammals, typically livestock, and as a vector for various parasitic agents, including bacteria, viruses, and parasites. The role of long non-coding RNAs (lncRNAs) in regulating gene expression during metamorphosis of the flesh fly has not been well established. In this study, we performed genome-wide identification and characterization of lncRNAs from the early pupal stage (1-days pupae), mid-term pupal stage (5-days pupae), and late pupal stage (9-days pupae) of S. peregrina by RNA-seq, and a total of 6921 lncRNAs transcripts were identified. RT-qPCR and enrichment analyses revealed the differentially expressed lncRNAs (DE lncRNAs) that might be associated with insect metamorphosis development. Furthermore, functional analysis revealed that the DE lncRNA (SP_lnc5000) could potentially be involved in regulating the metamorphosis of S. peregrina. RNA interference of SP_lnc5000 caused reduced expression of metamorphosis-related genes in 20-hydroxyecdysone (20E) signaling (Br-c, Ftz-F1), cuticle tanning pathway (TH, DOPA), and chitin related pathway (Cht5). Injection of dsSP_lnc5000 in 3rd instar larvae of S. peregrina resulted in deformed pupae, stagnation of pupal-adult metamorphosis, and a decrease in development time of pupal, pupariation rates and eclosion rates. Hematoxylin-eosin staining (H&E), scanning electron microscope (SEM) observation and cuticle hydrocarbons (CHCs) analysis indicated that SP_lnc5000 had crucial roles in the metamorphosis developmental by modulating pupal cuticular development. We established that the lncRNA SP_lnc5000 potentially regulates the metamorphosis of S. peregrina by putatively affecting the structure and composition of the pupal cuticle. This study enhances our understanding of lncRNAs as regulators of metamorphosis in S. peregrina, and provide valuable insights into the identification of potential targets for vector control and the development of effective strategies for controlling the spread of myiasis and parasitic diseases.

Discovery of long non-coding RNAs in Aspergillus flavus response to water activity, CO2 concentration, and temperature changes

Although the role of long non-coding RNAs (lncRNAs) in key biological processes in animals and plants has been confirmed for decades, their identification in fungi remains limited. In this study, we discovered and characterized lncRNAs in Aspergillus flavus in response to changes in water activity, CO2 concentration, and temperature, and predicted their regulatory roles in cellular functions. A total of 472 lncRNAs were identified in the genome of A. flavus, consisting of 470 novel lncRNAs and 2 putative lncRNAs (EFT00053849670 and EFT00053849665). Our analysis of lncRNA expression revealed significant differential expression under stress conditions in A. flavus. Our findings indicate that lncRNAs in A. flavus, particularly down-regulated lncRNAs, may play pivotal regulatory roles in aflatoxin biosynthesis, respiratory activities, cellular survival, and metabolic maintenance under stress conditions. Additionally, we predicted that sense lncRNAs down-regulated by a temperature of 30 °C, osmotic stress, and CO2 concentration might indirectly regulate proline metabolism. Furthermore, subcellular localization analysis revealed that up-and down-regulated lncRNAs are frequently localized in the nucleus under stress conditions, particularly at a water activity of 0.91, while most up-regulated lncRNAs may be located in the cytoplasm under high CO2 concentration.

A predictive molecular signature consisting of lncRNAs associated with cellular senescence for the prognosis of lung adenocarcinoma.

The role of long noncoding RNAs (lncRNAs) has been verified by more and more researches in recent years. However, there are few reports on cellular senescence-associated lncRNAs in lung adenocarcinoma (LUAD). Therefore, to explore the prognostic effect of lncRNAs in LUAD, 279 cellular senescence-related genes, survival information and clinicopathologic parameters were derived from the CellAge database and The Cancer Genome Atlas (TCGA) database. Then, we constructed a novel cellular senescence-associated lncRNAs predictive signature (CS-ALPS) consisting of 6 lncRNAS (AC026355.1, AL365181.2, AF131215.5, C20orf197, GAS6-AS1, GSEC). According to the median of the risk score, 480 samples were divided into high-risk and low-risk groups. Furthermore, the clinicopathological and biological functions, immune characteristics and common drug sensitivity were analyzed between two risk groups. In conclusion, the CS-ALPS can independently forecast the prognosis of LUAD, which reveals the potential molecular mechanism of cellular senescence-associated lncRNAs, and provides appropriate strategies for the clinical treatment of patients with LUAD.

LncRNA NEAT1 promotes IL-6 secretion in monocyte-derived dendritic cells via sponging miR-365a-3p in systemic lupus erythematosus

ABSTRACT Increasing evidence has uncovered the essential roles of long noncoding RNAs (lncRNAs) in biological and pathological functions of dendritic cells (DCs) among patients with systemic lupus erythematosus (SLE). However, whether lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) could modulate DCs, especially in the inflammation of SLE, remains largely unknown. Fifteen SLE patients and fifteen age-matched healthy controls were included, and their monocyte-derived dendritic cells (moDCs) were cultured in vitro. Our research identified that the expression of NEAT1 was significantly increased in moDCs of SLE patients and positively correlated with disease activity. Interleukin 6 (IL-6) from both plasma and secreted supernatants of moDCs was also elevated in the SLE group. In addition, regulation of NEAT1 in moDCs by transfection could lead to the corresponding change in IL-6 generation. While for miR-365a-3p, a micro-RNA that can bind with the 3’ UTR region of IL6 and NEAT1, it may serve as a negative modulator since its overexpression could result in the reduction of IL-6 levels and vice versa. Additionally, the elevation in NEAT1 expression could increase the secretion of IL-6 by specifically binding to miR-365a-3p, reducing the negative modulatory effects of miR-365a-3p on the IL6 target gene, which suggested that elevated NEAT1 expression could function as the competing endogenous RNA (ceRNA). In conclusion, our findings indicate that NEAT1 can efficiently sponge miR-365a-3p to upregulate expression and secretion of IL-6 in moDCs, suggesting that the NEAT1/miR-365a-3p/IL6 axis may be involved in the development of SLE disease.

GAS5 protects against nonalcoholic fatty liver disease via miR-28a-5p/MARCH7/NLRP3 axis-mediated pyroptosis.

Nonalcoholic fatty liver disease (NAFLD) is characterised by hepatic steatosis, inflammation, and insulin resistance. The role of long noncoding RNA (lncRNA)-regulated pyroptosis in NAFLD development remains largely unknown. This study aimed to investigate whether NAFLD development is controlled by lncRNA growth-arrest specific transcript 5 (GAS5)/miR-28a-5p/membrane associated ring-CH-type finger 7 (MARCH7)-mediated pyroptosis using in vivo and in vitro models. First, GAS5 expression was decreased but miR-28a-5p expression was increased in the livers of NAFLD patients, high-fat diet (HFD)-fed mice and leptin-deficient obese (Ob/Ob) mice. Furthermore, GAS5 suppressed while miR-28a-5p promoted NAFLD development, and overexpression of miR-28a-5p reversed the GAS5 overexpression-induced attenuation of NAFLD. Mechanistically, GAS5 served as a sponge of miR-28a-5p, and miR-28a-5p enhanced pyroptosis by targeting the 3' untranslated region (UTR) of the E3 ligase MARCH7 during NAFLD development. MARCH7 interacted with the NOD-like receptor protein 3 (NLRP3) protein, resulting in proteasomal degradation of NLRP3 to inhibit pyroptosis. As expected, MARCH7 knockdown abolished the miR-28a-5p knockdown-induced inhibition of NAFLD development, and the ubiquitin E3 ligase-inactive mutant (W589A/I556A) of MARCH7 failed to inhibit NAFLD development. In conclusion, GAS5 protected against NAFLD development by binding to miR-28a-5p, miR-28a-5p promoted NAFLD development by targeting MARCH7, and MARCH7 ameliorated NAFLD by suppressing NLRP3-mediated pyroptosis. The GAS5/miR-28a-5p/MARCH7/NLRP3 axis plays an important role in NAFLD progression, and it might be a biomarker for NAFLD.

RNA Fluorescence In Situ Hybridization for Long Non-Coding RNA Localization in Human Osteosarcoma Cells.

The important roles of long non-coding RNAs (lncRNAs) in cancer have been studied, such as regulating the proliferation, epithelial-mesenchymal transition (EMT), migration, infiltration, and autophagy of cancer cells. Localization detection of lncRNAs in cells can provide insight into their functions. By designing the lncRNA-specific antisense chain sequence followed by labeling with fluorescent dyes, RNA fluorescence in situ hybridization (FISH) can be applied to detect the cellular localization of lncRNAs. Together with the development of microscopy, the RNA FISH techniques now even allow for visualization of the poorly expressed lncRNAs. This method can not only detect the localization of lncRNAs alone, but also detect the colocalization of other RNAs, DNA, or proteins by using double-color or multicolor immunofluorescence. Here, we have included the detailed experimental operation procedure and precautions of RNA FISH by using lncRNA small nucleolar RNA host gene 6 (SNHG6) in human osteosarcoma cells (143B) as an example, to provide a reference for researchers who want to perform RNA FISH experiments, especially lncRNA FISH.

Long Noncoding RNA MIATNB Regulates Hyperosmotic Stress-induced Corneal Epithelial Cell Injury by Inhibiting Autophagy in Dry Eye Disease

Purpose Dry eye disease (DED) has a complex etiology and the roles of long noncoding RNAs (lncRNAs) in its pathophysiology are not completely understood. Autophagy is a self-eating process important for cell survival and homeostasis. The present study explored the role of myocardial infarction-associated transcript neighbor (MIATNB) long non-coding RNA in hyperosmolarity-induced autophagy and apoptosis in human corneal epithelial cell (HCEC)-based model of dry eye disease. Methods In vitro assays were performed with a human SV40 immortalized corneal epithelial cell line. Different concentrations of NaCl were used to create hyperosmolarity. HCECs were cultured in presence of 70–120 mM NaCl for 24 h to create an in vitro model of dry eye. RT-qPCR was performed to assess the expression of dry eye related LC3B, ATG16L, BECN1, ATG1, ATG7, ATG13, ATG5, ATG10, and ATG101 mRNAs and western blot analysis of LC3B and P62 and RFP -GFP-tagged LC3. Flow cytometry and western blot analysis of caspase 3, BCL2 and BAX were performed to detect apoptosis. Chloroquine (CQ) was used to inhibit autophagy pharmacologically. Results Autophagy flux was activated in HCECs subjected to hyperosmotic stress. Hyperosmolarity activated apoptosis and inhibited HCEC migration and autophagy. Hyperosmolarity upregulated MIATNB expression, while MIATNB knockdown inhibited autophagosome degradation and promoted HCEC apoptosis. Under hyperosmolar conditions, MIATNB knockdown also inhibited the degradation of autophagolysosomes and stimulated HCEC apoptosis. Conclusion MIATNB plays a vital role in dry eye pathogenesis and serves as a bridge between autophagy and apoptosis. Targeting MIATNB for DED treatment should be further evaluated.