Long noncoding RNA ZFAS1: A novel anti-apoptotic target in Fuchs endothelial corneal dystrophy.

Fuchs endothelial corneal dystrophy (FECD) is the leading cause of endothelial keratoplasty without efficacious drug treatment. Recent studies have emphasized the involvement of epigenetic regulation in FECD development. Long non-coding RNAs (lncRNAs) are recognized as crucial epigenetic regulators in diverse cellular processes and ocular diseases. In this study, we revealed the expression patterns of lncRNAs using high-throughput sequencing technology in FECD mouse model, and identified 979 significantly dysregulated lncRNAs. By comparing the data from FECD human cell model, we obtained a series of homologous lncRNAs with similar expression patterns, and revealed that these homologous lncRNAs were enriched in FECD related biological functions, with apoptosis (mmu04210) showing the highest enrichment score. In addition, we investigated the role of lncRNA zinc finger antisense 1 (ZFAS1) in apoptotic process. This study would broaden our understanding of epigenetic regulation in FECD development, and provide potential anti-apoptotic targets for FECD therapy.

LncRNA ZFAS1 Alleviated NLRP3 Inflammasome-Mediated Pyroptosis through Regulating miR-96-5p/Smad7 Signaling in Allergic Rhinitis.

INTRODUCTION: The NLR family pyrin domain containing 3 (NLRP3)-mediated pyroptosis was positively correlated with the allergic rhinitis progression and was reported to be regulated by SMAD family member 7 (Smad7). Bioinformatics analysis revealed that Smad7 might be targeted by miR-96-5p, and miR-96-5p might be targeted by long noncoding RNA zinc finger antisense 1 (ZFAS1). However, the effects and regulatory mechanisms of the ZFAS1/miR-96-5p/Smad7 functional axis in allergic rhinitis have not been investigated. METHODS: Human nasal mucosa epithelial cell line RPMI 2650 and C57BL/6 mice were obtained for in vitro and in vivo studies. Dual-luciferase reporter assay and RNA immunoprecipitation were implemented for detecting molecular interactions. Cell counting kit-8 and flow cytometry were used for measuring cell viability and pyroptosis. ELISA was obtained for monitoring cytokine secretion. RT-qPCR and Western blot were examined for determining RNA and protein expression. RESULTS: In vitro studies revealed that ZFAS1 was downregulated in interleukin (IL)-13-treated RPMI 2650 cells, while overexpression of ZFAS1 enhanced cell viability and inhibited NLRP3-mediated pyroptosis and inflammatory response. ZFAS1 directly inhibited miR-96-5p to suppress NLRP3-mediated pyroptosis in IL-13-treated RPMI 2650 cells. MiR-96-5p bound to the 3'-untranslated region of Smad7 and knockdown of Smad7 significantly reversed the effects of miR-96-5p depletion. Moreover, in vivo experiments further confirmed the findings of in vitro studies and showed ZFAS1 overexpression or miR-96-5p inhibition alleviated allergic rhinitis in vivo. CONCLUSION: ZFAS1 downregulated the expression of miR-96-5p to upregulate Smad7 level, which subsequently inhibited NLRP3-mediated pyroptosis and inflammatory response to ameliorate allergic rhinitis.

The protective role of RACK1 in hepatic ischemia‒reperfusion injury-induced ferroptosis.

Although ferroptosis plays a crucial role in hepatic ischemia‒reperfusion injury (IRI), the molecular mechanisms underlying this process remain unclear. We aimed to explore the potential involvement of the receptor for activated C kinase 1 (RACK1) in hepatic IRI-triggered ferroptosis. Using hepatocyte-specific RACK1 knockout mice and alpha mouse liver 12 (AML12) cells, we conducted a series of in vivo and in vitro experiments. We found that RACK1 has a protective effect on hepatic IRI-induced ferroptosis. Specifically, RACK1 was found to interact with AMPKα through its 1-93 amino acid (aa) region, which facilitates the phosphorylation of AMPKα at threonine 172 (Thr172), ultimately exerting an antiferroptotic effect. Furthermore, the long noncoding RNA (lncRNA) ZNFX1 Antisense 1 (ZFAS1) directly binds to aa 181-317 of RACK1. ZFAS1 has a dual impact on RACK1 by promoting its ubiquitin‒proteasome-mediated degradation and inhibiting its expression at the transcriptional level, which indirectly exacerbates hepatic IRI-induced ferroptosis. These findings underscore the protective role of RACK1 in hepatic IRI-induced ferroptosis and showcase its potential as a prophylactic target for hepatic IRI mitigation.

The Expression Analysis of Long Non-coding RNAs Related to Wnt/ß-Catenin Signaling in Pancreatic Cancer Patients.

Background The oncogenic Wnt/ß-catenin signaling plays a critical role in carcinogenesis, prognosis, and resistance to therapy. Pancreatic cancer (PC) has high mortality because of its poor prognosis. Several studies have suggested that lncRNAs are directly involved in the development and progression of PC as well as in Wnt/ß-catenin signaling. In this study, we investigated and compared the expression of Wnt/ß-catenin signaling-related ZFAS1 and HCG11 lncRNAs, and their targets, CTNNB1 and IGF2BP1 genes in the blood of patients with PC and healthy individuals. A total of 47 PC patients and 50 healthy individuals participated in this study. RNA was extracted from the peripheral blood samples of participants, and cDNA was synthesized. The expression level of the selected genes was quantified by real-time PCR. The expression of HCG11 lncRNA and CTNNB1 genes in patients with PC was significantly upregulated compared to healthy individuals, and the expression of the ZFAS1 lncRNA was significantly downregulated. According to the analysis of the ROC curve, the diagnostic powers of ZFAS1 and CTNNB1 in PC were 0.67 and 0.69, respectively. Altogether, the present study suggests a role for ZFAS1 and HCG11 lncRNAs and CTNNB1 and IGF2BP1 in the pathogenesis of pancreatic cancer. Moreover, the peripheral expression of these lncRNAs may be useful as potential biomarkers for PC.

ZNFX1 antisense RNA1 promotes antiviral innate immune responses via modulating ZNFX1 function.

Increasing evidence suggests that natural antisense transcriptional lncRNAs regulate their adjacent coding genes to mediate diverse aspects of biology. Bioinformatics analysis of the previously identified antiviral gene ZNFX1 revealed neighboring lncRNA ZFAS1 transcribed on the opposite strand from ZNFX1. Whether ZFAS1 exerts antiviral function via regulating the dsRNA sensor ZNFX1 is unknown. Here we found that ZFAS1 was upregulated by RNA and DNA viruses and type I IFNs (IFN-I) dependent on Jak-STAT signaling, similar to the transcription regulation of ZNFX1. Knockdown of endogenous ZFAS1 partially facilitated viral infection, while ZFAS1 overexpression showed opposite effects. In addition, mice were more resistant to VSV infection with the delivery of human ZFAS1. We further observed that ZFAS1 knockdown significantly inhibited IFNB1 expression and IFR3 dimerization, whereas ZFAS1 overexpression positively regulated antiviral innate immune pathways. Mechanistically, ZFAS1 positively regulated ZNFX1 expression and antiviral function by enhancing the protein stability of ZNFX1, thereby establishing a positive feedback loop to enhance antiviral immune activation status. In short, ZFAS1 is a positive regulator of antiviral innate immune response via regulating its neighbor gene ZNFX1, adding new mechanistic insight into lncRNA-mediated regulation of signaling in innate immunity.

HDAC3-mediated lncRNA ZFAS1 inhibited IL-13-induced secretion of proinflammatory cytokines in nasal epithelial cells by regulating the miR-7-5p/SIRT1 pathway.

Allergic rhinitis (AR) is a disease that is difficult to cure and accompanies the patient's life. Proinflammatory cytokines (GM-CSF and eotaxin) and MUC5AC are key mediators promoting AR progression. Herein, the function of lncRNA ZFAS1 in AR was investigated. Nasal epithelial cells (NECs) were subjected to 50 ng/mL IL-13 for 24 h to construct an AR cell model. The mRNA and protein expressions were assessed using qRT-PCR and western blot. The levels of GM-CSF, eotaxin, IL-1ß, IL-6, TNF-α and MUC5AC in cell supernatant were examined by ELISA. The binding relationships between HDAC3, ZFAS1, miR-7-5p and SIRT1 were analysed using dual luciferase reporter or ChIP assays. Herein, our results displayed that ZFAS1 and SIRT1 were lowly expressed in AR, while miR-7-5p and HDAC3 were highly expressed. Functional experiments displayed that ZFAS1 overexpression suppressed IL-13-induced proinflammatory cytokines and mucin production in NECs. The highly expressed HDAC3 in AR inhibited ZFAS1 expression by binding with ZFAS1 promoter. In addition, our experiments revealed that ZFAS1 targeted miR-7-5p, and miR-7-5p targeted SIRT1. As expected, miR-7-5p overexpression or SIRT1 silencing abrogated ZFAS1 upregulation's repression on IL-13-induced proinflammatory cytokines and MUC5AC secretory levels in NECs. ZFAS1 suppressed proinflammatory cytokines, inflammatory cytokines, and MUC5AC secretory levels in AR by regulating the miR-7-5p/SIRT1 axis. Thus, our work suggested that ZFAS1 might serve as a novel target for AR treatment and prevention.

LncRNA ZFAS1 promotes laryngeal cancer progression through RBFOX2-mediated MENA alternative splicing.

Laryngeal cancer (LC) is the most common aggressive malignancy of the head and neck. LncRNA ZNFX1 antisense RNA 1 (ZFAS1) displays oncogenic properties in head and neck squamous cell carcinoma, but its regulatory role in laryngeal cancer progression remains obscure. Here, we found that ZFAS1 expression in laryngeal cancer tissues and cells was higher than that in adjacent normal tissues and normal nasopharyngeal epithelial cells. Highly expressed ZFAS1 was associated with advanced lymph node metastasis stages and clinical stages. ZFAS1 overexpression promoted LC cell proliferation, invasion, and N-cadherin and Vimentin expression, and suppressed E-cadherin expression. While ZFAS1 knockdown played an opposite role. Mechanistically, ZFAS1 stabilized RNA binding fox-1 homolog 2 (RBFOX2) protein expression by binding to RBFOX2, and RBFOX2 overexpression reversed the effect of ZFAS1 silence on cell functions. Moreover, highly expressed RBFOX2 led to skipping of MENA exon 11a and generating a pro-invasive isoform (MENAINV ). MENAINV overexpression effectively abolished the inhibitory effect of RBFOX2 knockdown on cell malignant progression. Furthermore, Hep2 cells infected with lentivirus-mediated ZFAS1 shRNA or negative control shRNA were subcutaneously injected into mice to assess the role of ZFAS1 in tumor growth. And the data showed that silencing ZFAS1 in vivo hindered xenograft tumor growth. In conclusion, silencing ZFAS1 alleviated malignant progression of laryngeal cancer cells and mouse xenograft tumor growth by regulating RBFOX2-mediated alternative splicing of MENA.

Astragalus mongholicus Bunge-Curcuma aromatica Salisb. suppresses growth and metastasis of colorectal cancer cells by inhibiting M2 macrophage polarization via a Sp1/ZFAS1/miR-153-3p/CCR5 regulatory axis.

Colorectal cancer (CRC) is regarded as one of the commonest cancer types around the world. Due to the poor understanding on the causes of CRC formation and progression, this study sets out to investigate the physiological mechanisms by which Astragalus mongholicus Bunge-Curcuma aromatica Salisb. (ARCR) regulates CRC growth and metastasis, and the role in which M2 macrophage polarization plays in this process. An orthotopic-transplant model of CRC was established to evaluate the influence of ARCR on the polarization of M2 macrophage and the growth and metastasis of tumors. Next, the binding affinity among Sp1, ZFAS1, miR-153-5p, and CCR5 was identified using multiple assays. Finally, after co-culture of bone marrow-derived macrophages (BMDM) with CRC cell line CT26.WT, the cell proliferative, invasive, and migrated abilities were assessed in gain- or loss-of-function experiments. ARCR inhibited the infiltration of M2 macrophages into tumor microenvironment to suppress the CRC growth and metastasis in vivo. Additionally, ARCR inhibited the transcription of ZFAS1 by reducing Sp1 expression to repress M2 macrophage polarization. Moreover, ZFAS1 competitively binds to miR-153-3p to upregulate the CCR5 expression. Finally, ARCR suppressed the polarization of M2 macrophages to inhibit the tumor growth and tumor metastasis in CRC by mediating the Sp1/ZFAS1/miR-153-3p/CCR5 regulatory axis. Collectively, ARCR appears to suppress the CRC cell growth and metastasis by suppressing M2 macrophage polarization via Sp1/ZFAS1/miR-153-3p/CCR5 regulatory axis. 1. ARCR suppress the CRC cell growth and metastasis 2. ZFAS1 promotes CCR5 expression by competitively binding to miR-153-3p. 3. Sp1 promotes M2 macrophage polarization by activating ZFAS1 via miR-153-3p/CCR5. 4. The study unveiled a protective target against CRC.

LncRNA ZFAS1 regulates the hippocampal neurons injury in epilepsy through the miR-15a-5p/OXSR1/NF-κB pathway.

Long non-coding RNAs (lncRNAs) have been confirmed to be involved in epilepsy development. It has been reported that lncRNA ZFAS1 plays a vital regulatory role in epilepsy progression. Therefore, the role and molecular mechanism of ZFAS1 in epilepsy progression deserve further investigation. Mice status epilepticus (SE) model was constructed, and hippocampal neurons were isolated from mice hippocampus tissues. The expression of ZFAS1, miR-15a-5p and oxidative stress responsive 1 (OXSR1) were determined by quantitative real-time PCR. ELISA assay was used to detect the concentrations of inflammation factors. Cell viability and apoptosis were examined by MTT assay, EdU staining and flow cytometry. Western blot analysis was conducted to measure protein levels, and the productions of SOD and MDA were measured to assess cell oxidative stress. Dual-luciferase reporter assay and RIP assay were employed to validate the relationship between miR-15a-5p and ZFAS1 or OXSR1. LncRNA ZFAS1 was highly expressed in SE mice and SE-stimulated hippocampal neurons. Silenced ZFAS1 promoted viability, while inhibited inflammation, apoptosis and oxidative stress in SE-induced hippocampal neurons. MiR-15a-5p could be targeted by ZFAS1, and its inhibitor also reversed the suppressive effect of ZFAS1 knockdown on SE-induced hippocampal neurons injury. In addition, OXSR1 was a target of miR-15a-5p, and its silencing also could relieve SE-induced hippocampal neurons injury. OXSR1 overexpression reversed the inhibition effect of miR-15a-5p on SE-induced hippocampal neurons injury. Moreover, ZFAS1 positively regulated OXSR1 expression by sponging miR-15a-5p, thereby activating the NF-κB pathway. LncRNA ZFAS1 might contribute to the progression of epilepsy by regulating the miR-15a-5p/OXSR1/NF-κB pathway.

The role of blood lnc-ZFAS1 in acute ischemic stroke: correlation with neurological impairment, inflammation, and survival profiles.

BACKGROUND: Long non-coding RNA zinc finger antisense 1 (lnc-ZFAS1) has been reported to inhibit neuronal damage in acute ischemic stroke (AIS). However, the role of lnc-ZFAS1 in AIS patients remains unclear. Therefore, we assessed the relationship of lnc-ZFAS1 with neurological impairment, inflammation, and prognosis in AIS patients. METHODS: Totally, 241 AIS patients and 120 controls were enrolled. lnc-ZFAS1 in peripheral blood mononuclear cells was evaluated using reverse transcription-quantitative polymerase chain reaction. Besides, a 3-year follow-up was conducted to assess recurrence-free survival (RFS) and overall survival (OS) in AIS patients. RESULTS: lnc-ZFAS1 was reduced in AIS patients compared to that in controls (Z = -10.693, p < 0.001). In AIS patients, lnc-ZFAS1 was negatively correlated with National Institutes of Health Stroke Scale score (rs  = -0.335, p < 0.001), C-reactive protein (rs  = -0.284, p < 0.001), tumor necrosis factor-alpha (rs  = -0.293, p < 0.001), interleukin-1ß (rs  = -0.149, p = 0.021), and interleukin-6 (rs  = -0.161, p = 0.012), but not underlying diseases (all p > 0.05). Besides, lnc-ZFAS1 was divided into high and low levels based on the median expression in AIS patients. Indeed, high lnc-ZFAS1 predicted better RFS (χ2  = 6.222, p = 0.013); the 1-year, 2-year, and 3-year RFS rates were 94.2%, 88.3%, and 85.5%, respectively, in patients with high lnc-ZFAS1, then 87.5%, 79.2%, and 71.6%, respectively, in those with low lnc-ZFAS1. However, lnc-ZFAS1 was not correlated with OS (χ2  = 1.404, p = 0.236); the 1-year, 2-year, and 3-year OS rates were 98.3%, 95.8%, and 94.0%, respectively, in patients with high lnc-ZFAS1, then 96.7%, 93.9%, and 89.6%, respectively, in those with low lnc-ZFAS1. CONCLUSION: Lower lnc-ZFAS1 expression is connected with increased neurological impairment and inflammation as well as worse RFS in AIS patients.

Inhibition of the long non-coding RNA ZFAS1 attenuates ferroptosis by sponging miR-150-5p and activates CCND2 against diabetic cardiomyopathy.

Diabetic cardiomyopathy (DbCM) is responsible for increased morbidity and mortality in patients with diabetes and heart failure. However, the pathogenesis of DbCM has not yet been identified. Here, we investigated the important role of lncRNA-ZFAS1 in the pathological process of DbCM, which is associated with ferroptosis. Microarray data analysis of DbCM in patients or mouse models from GEO revealed the significance of ZFAS1 and the significant downregulation of miR-150-5p and CCND2. Briefly, DbCM was established in high glucose (HG)-treated cardiomyocytes and db/db mice to form in vitro and in vivo models. Ad-ZFAS1, Ad-sh-ZFAS1, mimic miR-150-5p, Ad-CCND2 and Ad-sh-CCND2 were intracoronarily administered to the mouse model or transfected into HG-treated cardiomyocytes to determine whether ZFAS1 regulates miR-150-5p and CCND2 in ferroptosis. The effect of ZFAS1 on the left ventricular myocardial tissues of db/db mice and HG-treated cardiomyocytes, ferroptosis and apoptosis was determined by Masson staining, immunohistochemical staining, Western blotting, monobromobimane staining, immunofluorescence staining and JC-1 staining. The relationships among ZFAS1, miR-150-5p and CCND2 were evaluated using dual-luciferase reporter assays and RNA pull-down assays. Inhibition of ZFAS1 led to reduced collagen deposition, decreased cardiomyocyte apoptosis and ferroptosis, and attenuated DbCM progression. ZFAS1 sponges miR-150-5p to downregulate CCND2 expression. Ad-sh-ZFAS1, miR-150-5p mimic, and Ad-CCND2 transfection attenuated ferroptosis and DbCM development both in vitro and in vivo. However, transfection with Ad-ZFAS1 could reverse the positive effects of miR-150-5p mimic and Ad-CCND2 in vitro and in vivo. lncRNA-ZFAS1 acted as a ceRNA to sponge miR-150-5p and downregulate CCND2 to promote cardiomyocyte ferroptosis and DbCM development. Thus, ZFAS1 inhibition could be a promising therapeutic target for the treatment and prevention of DbCM.

Shedding more light on the role of Midkine in hepatocellular carcinoma: New perspectives on diagnosis and therapy.

One of the most common malignant tumors is hepatocellular carcinoma (HCC). Progression of HCC mainly results from highly complex molecular and pathological pathways. Midkine (MDK) is a growth factor that impacts viability, migration, and other cell activities. Since MDK has been involved in the inflammatory responses, it has been claimed that MDK has a crucial role in HCC. MDK acts as an anti-apoptotic factor, which mediates tumor cell viability. In addition, MDK blocks anoikis to promote metastasis. There is also evidence that MDK is involved in angiogenesis. It has been shown that the application of anti-MDK approaches might be promising in the treatment of HCC. Besides, due to the elevated expression in HCC, MDK has been proposed as a biomarker in the prognosis and diagnosis of HCC. In this review, we will discuss the role of MDK in HCC. It is hoped that the development of new strategies concerning MDK-based therapies will be promising in HCC management.

Transcription factor Sp1 ameliorates sepsis-induced myocardial injury via ZFAS1/Notch signaling in H9C2 cells.

PURPOSE: To investigate whether Sp1 can ameliorate sepsis-induced myocardial injury and explore the potential molecular mechanism. METHODS: The embryonic cardiomyocyte cell line H9C2 and primary cultured mouse neonatal cardiomyocytes (CMNCs) were treated with LPS or phosphate-buffered saline (PBS). A mouse model of LPS-induced sepsis was established using male C57BL/6J mice and their cardiomyocytes were collected. Real-time reverse transcription-PCR (qRT-PCR) assay was used to detect the expression levels of Sp1 and ZFAS1 in cardiomyocytes. Western blotting analysis was used to assess the protein expression levels of Sp1, apoptosis-associated proteins and Notch signaling pathway related proteins. Luciferase assay was used to detect the interaction between Sp1 and ZFAS1. Cell transfection was used to generate H9C2 cells with overexpressed or knocked down of Sp1 or ZFAS1. MTT assay and flow cytometry analysis were used to test the cell proliferation and cell apoptosis ratio. RESULTS: Our data revealed that the expressions of ZFAS1 and Sp1 were significantly reduced in LPS-treated H9C2 cells and primary CMNCs. The downregulation of ZFAS1 and Sp1 were also found in cardiomyocytes obtained from LPS-challenged mice. LPS induced H9C2 cell apoptosis and depressed cell proliferation was ameliorated by ZFAS1 overexpression and aggravated by ZFAS1 knockdown. Mechanistically, Luciferase assay indicated that Sp1 could bind to ZFAS1, and positively regulated ZFAS1 expression. Moreover, Notch signaling pathway participates in H9C2 cell apoptosis mediated by Sp1. CONCLUSION: The present study demonstrates that Sp1 regulates LPS-induced cardiomyocyte apoptosis via ZFAS1/Notch signaling pathway, which may serve as therapeutic targets for sepsis-induced myocardial injury.

lncRNA ZFAS1 promotes ox-LDL induced EndMT through miR-150-5p/Notch3 signaling axis.

EndMT is an active contributor to atherosclerosis pathology, and lncRNAs is widely involved in the occurrence and development of atherosclerosis. The purpose of this study was to investigate the regulatory mechanisms of ZFAS1 in EndMT of atherosclerosis. Here, the ApoE-/- mice were feed with high-fat diet to establish the atherosclerosis model, and HUVECs was stimulated with ox-LDL to induce EndMT. RT-PCR and western blot were used to detect the mRNA and protein expression, respectively. The expression of EndMT markers were detected by immune-fluorescence. The relationships among ZFAS1, miR-150-5p and Notch3 were evaluated by luciferase reporter assay. The role of ZFAS1 in EndMT and its dependence on miR-150-5p/Notch3 axis was further detected by knocking down or over-expressing ZFAS1. We found that ZFAS1 and Notch3 were upregulated while miR-150-5p was downregulated in atherosclerosis mice and ox-LDL-treated HUVECs. The expression of CD31 and vWF were significant decreased, while the α-SMA and vimentin were significant increased in ox-LDL-treated HUVECs, and overexpression of ZFAS1 enhanced the effect of ox-LDL on HUVECs. Further, ZFAS1 functions as a ceRNA to increase Notch3 expression through sponging miR-150-5p, and miR-150-5p mimic or si-Notch3 could reverse LV-ZFAS1-mediated EndMT. In summary, lncRNA ZFAS1 promotes ox-LDL induced HUVECs EndMT through regulating miR-150-5p/Notch3 axis.

PVT1 and ZFAS1 lncRNAs expressions and their biomarker value in gastric cancer tissue sampling among Iranian population.

BACKGROUND: lncRNAs are modulatory factors with critical function in the tumorigenesis pathways, introducing them as promising therapeutic and diagnostic biomarkers for different cancers. This study is thus aimed to evaluate the differences in PVT1 and ZFAS1 gene expression in tumorous tissues as compared with adjacent healthy non-tumorous biopsies of gastric cancer cases. METHODS: One hundred two pairs of tumorous and adjacent non-tumorous biopsies of GC cases were sampled. RNA isolation and cDNA production were carried out. The qRT-PCR was performed to evaluate the expression of PVT1 and ZFAS1 genes. Moreover, the associations between PVT1 or ZFAS1 and clinicopathological characteristics as well as the biomarker roles of the lncRNAs were assessed. RESULTS: The PVT1 and ZFAS1 expressions showed a significant increase and decrease in GC samples as compared with non-cancerous tissues, respectively. PVT1 expression was significantly associated with and lymph-node involvement (p = 0.0007). Moreover, ZFAS1 expression demonstrated a significant association with lymph-node involvement (p = 0.0005), and tumor size >5 cm (p = 0.003). The findings of the ROC curve revealed that PVT1 and ZFAS1 may act as a possible biomarker with AUC of 0.71 and 0.79, specificity of 78.43% and 79.41%, and sensitivity of 55.88% and 64.71%. CONCLUSIONS: Regarding upregulation of PVT1 and downregulation of ZFAS1 in human GC samples, these genes may respectively act as oncogenic and tumor-suppressive factors in GC cases. Furthermore, PVT1 and ZFAS1 can be considered as possible biomarkers for the detection and treatment of GC cases.

Downregulation of lncRNA ZFAS1 and upregulation of microRNA-129 repress endocrine disturbance, increase proliferation and inhibit apoptosis of ovarian granulosa cells in polycystic ovarian syndrome by downregulating HMGB1.

OBJECTIVE: The objective was to find the role of long-non-coding RNA zinc finger antisense 1 (lncRNA ZFAS1)/microRNA (miR)-129/high-mobility group box protein 1 (HMGB1) axis in polycystic ovary syndrome (PCOS). METHODS: Ovarian granulosa cells from non-PCOS patients and PCOS patients were collected, and HMGB1, miR-129 and lncRNA ZFAS1 expression were detected. Ovarian granulosa cells were transfected with si-ZFAS1 or miR-129 mimics to verify their roles in P4 and E2 secretion, and the biological functions of ovarian granulosa cells. RESULTS: LncRNA ZFAS1 and HMGB1 were elevated, while miR-129 was down-regulated in ovarian granulosa cells of PCOS patients. Down-regulated lncRNA ZFAS1 or overexpressed miR-129 could decrease HMGB1 expression, increase P4 and E2 secretion, promote proliferation activity while inhibit apoptosis of ovarian granulosa cells in PCOS. CONCLUSION: LncRNA ZFAS1 could bind to miR-129 to promote HMGB1 expression, thereby affecting the endocrine disturbance, proliferation and apoptosis of ovarian granulosa cells in PCOS.

Long noncoding RNA ZFAS1 promotes tumorigenesis and metastasis in nasopharyngeal carcinoma by sponging miR-892b to up-regulate LPAR1 expression.

OBJECTIVE: In this study, we explored the NPC-specific expression of ZFAS1 and the mechanism of ZFAS1-mediated growth, aggressiveness and tumorigenesis in NPC. METHODS: The expression profile of lncRNAs was detected in NPC tissues and matching para-carcinoma tissues using microarray analysis. LncRNA-miRNA and miRNA-mRNA interaction networks were constructed using the miRcode v11 and TargetScanHuman v7.2 web server and then validated using dual-luciferase assay. Western blot and RT-qPCR were performed to detect protein and RNA expression. The effects of ZFAS1, miR-892b and LPAR1 dysregulation on the proliferative, migratory and invasive abilities of NPC cells were observed using colony formation, cell counting kit-8 (CCK-8) and transwell assays in vitro. In vivo, a xenograft nude mouse model was established to detect the impact of ZFAS1 dysregulation on the tumorigenicity of NPC cells. RESULTS: The expression of multiple lncRNAs, of which ZFAS1 was up-regulated, was dysregulated in NPC tissues. ZFAS1 directly targeted miR-892b, and miR-892b negatively regulated the expression of downstream LPAR1. The proliferation, migration and invasion of NPC cells could be largely enhanced by the downregulation of miR-892b as well as the up-regulation of ZFAS1 and LPAR1, while the overexpression of miR-892b and the downregulation of ZFAS1 and LPAR1 decreased these abilities. In nude mice, the growth of tumour xenografts formed by HONE1 cells was significantly suppressed when ZFAS1 was silenced. CONCLUSION: The study demonstrated that lncRNA ZFAS1 may act as a promoter of tumorigenesis and metastasis in nasopharyngeal carcinoma, by up-regulating the expression of LPAR1 in a miR-892b-dependent manner.

Knockdown of lncRNA ZFAS1-suppressed non-small cell lung cancer progression via targeting the miR-150-5p/HMGA2 signaling.

Non-small cell lung cancer (NSCLC) is the main type of lung malignancy. Early diagnosis and treatments for NSCLC are far from satisfactory due to the limited knowledge of the molecular mechanisms regarding NSCLC progression. Long noncoding RNA (lncRNA) ZNFX1 antisense RNA1 (ZFAS1) has been implicated for its functional role in the progression of malignant tumors. This study aimed to determine the ZFAS1 expression from lung cancer clinical samples and to explore the molecular mechanisms underlying ZFAS1-modulated NSCLC progression. Experimental assays revealed that clinical samples and cell lines of lung malignant tumors showed an upregulation of ZFSA1. ZFAS1 expression was markedly upregulated in the lung tissues from patients with advanced stage of this malignancy. The loss-of-function assays showed that knockdown of ZFAS1-suppressed NSCLC cell proliferative, as well as invasive potentials, increased NSCLC cell apoptotic rates in vitro and also attenuated tumor growth of NSCLC cells in the nude mice. Further experimental evidence showed that ZFAS1 inversely affected miR-150-5p expression and positively affected high-mobility group AT-hook 2 (HMGA2) expression in NSCLC cell lines. MiR-150-5p inhibition or HMGA2 overexpression counteracted the effects of ZFAS1 knockdown on NSCLC cell proliferative, invasive potentials and apoptotic rates. In light of examining the clinical lung cancer samples, miR-150-5p expression was downregulated and the HMGA2 expression was highly expressed in the lung cancer tissues compared with normal ones; the ZFAS1 expression showed a negative correlation with miR-150-5p expression but a positive correlation with HMGA2 expression in lung cancer tissues. To summarize, we, for the first time, demonstrated the inhibitory effects of ZFAS1 knockdown on NSCLC cell progression, and the results from mechanistic studies indicated that ZFAS1-mediated NSCLC progression cells via targeting miR-150-5p/HMGA2 signaling.

Long noncoding RNA ZFAS1 promoting small nucleolar RNA-mediated 2'-O-methylation via NOP58 recruitment in colorectal cancer.

BACKGROUND: Increasing evidence supports the role of small nucleolar RNAs (snoRNAs) and long non-coding RNAs (lncRNAs) as master gene regulators at the epigenetic modification level. However, the underlying mechanism of these functional ncRNAs in colorectal cancer (CRC) has not been well investigated. METHODS: The dysregulated expression profiling of lncRNAs-snoRNAs-mRNAs and their correlations and co-expression enrichment were assessed by GeneChip microarray analysis. The candidate lncRNAs, snoRNAs, and target genes were detected by in situ hybridization (ISH), RT-PCR, qPCR and immunofluorescence (IF) assays. The biological functions of these factors were investigated using in vitro and in vivo studies that included CCK8, trans-well, cell apoptosis, IF assay, western blot method, and the xenograft mice models. rRNA 2'-O-methylation (Me) activities were determined by the RTL-P assay and a novel double-stranded primer based on the single-stranded toehold (DPBST) assay. The underlying molecular mechanisms were explored by bioinformatics and RNA stability, RNA fluorescence ISH, RNA pull-down and translation inhibition assays. RESULTS: To demonstrate the involvement of lncRNA and snoRNAs in 2'-O-Me modification during tumorigenesis, we uncovered a previously unreported mechanism linking the snoRNPs NOP58 regulated by ZFAS1 in control of SNORD12C, SNORD78 mediated rRNA 2'-O-Me activities in CRC initiation and development. Specifically, ZFAS1 exerts its oncogenic functions and significantly up-regulated accompanied by elevated NOP58, SNORD12C/78 expression in CRC cells and tissues. ZFAS1 knockdown suppressed CRC cell proliferation, migration, and increased cell apoptosis, and this inhibitory effect could be reversed by NOP58 overexpression in vitro and in vivo. Mechanistically, the NOP58 protein could be recognized by the specific motif (AAGA or CAGA) of ZFAS1. This event accelerates the assembly of SNORD12C/78 to allow for further guiding of 2'-O-Me at the corresponding Gm3878 and Gm4593 sites. Importantly, silencing SNORD12C or 78 reduced the rRNAs 2'-O-Me activities, which could be rescued by overexpression ZFAS1, and this subsequently inhibits the RNA stability and translation activity of their downstream targets (e.g., EIF4A3 and LAMC2). CONCLUSION: The novel ZFAS1-NOP58-SNORD12C/78-EIF4A3/LAMC2 signaling axis that functions in CRC tumorigenesis provides a better understanding regarding the role of lncRNA-snoRNP-mediated rRNAs 2'-O-Me activities for the prevention and treatment of CRC.

LncRNA ZFAS1 promotes pancreatic adenocarcinoma metastasis via the RHOA/ROCK2 pathway by sponging miR-3924.

BACKGROUND: The mortality and morbidity rates of pancreatic adenocarcinoma have been increasing over the past two decades, and an understanding of the mechanisms underlying pancreatic adenocarcinoma progression is urgently needed. The long non-coding RNA ZFAS1 has been demonstrated to be an oncogene in some cancers, but its function and mechanism in pancreatic adenocarcinoma remain unclear. METHODS: The ZFAS1 expression level in pancreatic adenocarcinoma was predicted by bioinformatic analysis, and the expression level of ZFAS1 in pancreatic adenocarcinoma tissue samples and cell lines was further detected by quantitative real-time PCR and in situ hybridization. The functions of ZFAS1 in pancreatic adenocarcinoma in vitro and in vivo were investigated by further bioinformatic analysis. Dual-luciferase reporter assays were used to confirm the binding of ZFAS1/miR-3924 and miR-3924/ROCK2, and rescue assays were performed to further investigate the underlying mechanism. RESULTS: ZFAS1 overexpression in pancreatic adenocarcinoma was predicted and experimentally verified. ZFAS1 silencing inhibited pancreatic adenocarcinoma metastasis in vitro and in vivo. The competing endogenous RNA mechanism of ZFAS1 was also identified. CONCLUSIONS: Our results demonstrated the promotive effect of ZFAS1 on pancreatic adenocarcinoma metastasis and suggested its potential role as a novel regulator of ROCK2.