Expression Levels Of H19 Research Articles

LncRNA H19-induced AMPKα/LKB1 Complex Is Required for Dihydroartemisinin to Regulate Lipid Droplet Metabolism in Hepatic Stellate Cells

Background: Recently, long non-coding RNA-H19 was reported to mediate fatty acids and bile acids metabolism in liver tissue and play important role in fatty liver and cholestatic liver injury. However, little was known about H19 function and mechanism in liver fibrosis. Methods: ICR mice were intoxicated with CCl4 for evaluating H19 and Dihydroartemisinin effects in vivo. HSC-LX2 was cultured and multiple molecular experiments including real-time PCR, western blot, immunofluorescence, oil red staining, nile red staining, dual-luciferase reporter assay, fluorescence in situ hybridization, RNA immunoprecipitation and co-immunoprecipitation were used to elucidate the underlying mechanisms. Results: Decreased expression of H19 significantly ameliorated histopathological feature of liver fibrosis and inhibited HSC activation. This role of H19 was associated with lipid droplet metabolism dependent on AMPKα pathway in HSC. Interestingly, Dihydroartemisinin (DHA) inhibited H19 expression level in a dose-dependent manner. DHA decreased HIF-1α expression level to prevent HIF-1α binding to the promoter of H19. Mechanically, H19 drove AMPKα to interact with LKB1, resulting in the formation of the LKB1/AMPKα complex to facilitate phosphorylation of LKB1 towards AMPKα, causing obvious LDs consumption in HSC, but which was inhibited by DHA. Conclusions: Collectively, H19-induced AMPKα/LKB1 complex was required for DHA to regulate LD metabolism in activated HSC. This study highlighted a new molecular mechanism of DHA against liver fibrosis. Funding Statement: This study was supported by the National Natural Science Foundation of China (81270514, 31571455, 31401210, 31600653, and 81600483), the Natural Science Foundation of Jiangsu Province (BK20140955), the Open Project Program of Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica (No. JKLPSE201804), the Project of the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD) and the Postgraduate Research& Practice Innovation Program of Jiangsu Province (KYCX19_1262). Declaration of Interests: The authors claim that they have no conflicts of interest. Ethics Approval Statement: All experimental procedures were approved by the Institutional and Local Committee on the Care and Use of Animals of Nanjing University of Chinese Medicine, and all animals received humane care according to the National Institutes of Health (USA) guidelines.

Gene Expression of the Circulating Long Noncoding RNA H19 and HOTAIR in Egyptian Colorectal Cancer Patients.

Background: HOX transcript antisense RNA (HOTAIR) and H19 are two long noncoding RNAs that play vital functions in the development of colorectal cancer (CRC). Subjects and Methods: The expression level of HOTAIR and H19 in the sera samples of Egyptian CRC patients along with normal controls was evaluated by quantitative real-time PCR. The possible correlations with the biochemical and clinicopathological characteristics were determined. Results: The expression of HOTAIR and H19 showed 7.55- and 11.38-fold increased levels, respectively, in CRC patients compared to the controls (p < 0.001). Furthermore, HOTAIR expression in CRC patients with regional lymph node metastasis was significantly higher when compared with CRC patients without regional lymph node metastasis (p = 0.034). HOTAIR and H19 expression showed no significant correlation with tumor site or carcinoembryonic antigen concentration. The sensitivity and specificity of HOTAIR and H19 in the detection of CRC cases were calculated as 92.9% and 100%, respectively. Conclusion: HOTAIR and H19 expression levels are upregulated in Egyptian CRC patients, and therefore can be considered noninvasive diagnostic biomarkers with high sensitivity and specificity.

Effect of long chain non-coding RNA H19 on the migration and invasion of oral cancer cells and its molecular mechanism

To investigate the effect of the long chain non-coding RNA H19 (lncRNA H19) on the invasion and migration of oral cancer cells and its related molecular mechanism. The expression levels of lncRNA H19, miR-107, and cyclin-dependent kinase 6 (CDK6) in the immortalized oral epithelial cell line HIOEC and the oral cancer cell line CAL27 were detected by real-time quantitative polymerase chain reaction. CAL27 cells were transfected with siRNA H19, miR-107 mimics, pcDNA H19, or anti-miR-107, and the effects of H19 and miR-107 on the invasion and migration of cells were examined via Transwell assay. The TargetScan database predicted the targeting of H19, miR-107, and CDK6. Double luciferase reporter gene assay was performed to detect interactions among H19, miR-107, and CDK6. Western blot analysis was conducted to examine the effects of H19 and miR-107 on the protein level of the target gene CDK6. Compared with that in HIOEC cells, the expression of H19 was significantly increased in CAL27 cells (P<0.05). After transfection with siRNA H19, the expression of H19 decreased, and the invasion and migration ability of CAL27 cells were inhibited (P<0.05). H19 could bind specifically to the 3'-UTR of miR-107 to modulate the expression of miR-107. Compared with that in HIOEC cells, the expression of miR-107 significantly decreased in CAL27 cells (P<0.05). The expression of miR-107 increased after transfection with siRNA H19, and anti-mir-107 co-transfection could promote the invasion and migration ability of siRNA H19 in CAL27 cells (P<0.05). Compared with that in HIOEC cells, CDK6 expression significantly increased in CAL27 cells (P<0.05), and the expression level of the gene was coregulated by H19 and miR-107 (P<0.05). lncRNA H19 plays an important role in the development of oral cancer. It can regulate the invasion and migration of oral cancer cells by targeting the miR-107/CDK6 signaling axis.

Exosome-Transmitted lncRNA H19 Inhibits the Growth of Pituitary Adenoma.

Our previous study demonstrated that the expression of long noncoding RNA (lncRNA) H19 was frequently downregulated in human primary pituitary adenomas and negatively correlated with tumor progression. However, the role of exosomal lncRNA H19 in the inhibition of pituitary tumor growth remains unclear. To investigate whether exosomal H19 could be transported across the cell membrane to exert its inhibitory effect on pituitary tumor growth. Empty lentivirus GH3 cells with or without H19 overexpression were used to establish a xenograft model. Isolated exosomes were identified by transmission electron microscopy, nanoparticle tracking, and Western blotting. The expression levels of serum exosomal H19 from 200 healthy subjects and 206 patients with various subtypes of pituitary tumors were detected by ultracentrifugation and quantitative real-time PCR. The growth of distal tumor cells was inhibited by transferring exosomal H19, which could be transported through cell membrane and exert its inhibitory effect. Cabergoline increased H19 expression and played a synergic therapeutic effect with exosomal H19. Exosomal H19 inhibited phosphorylation of the mTORC1 substrate 4E-BP1. Of note, the expression level of exosomal H19 in the patients with all subtypes of pituitary tumors was significantly lower than that in the healthy subjects. The change of plasma exosomal H19 level may be correlated with the prognosis or drug response of the patients. Exosomal H19 inhibits the growth of distal pituitary tumors through inhibiting 4E-BP1 phosphorylation. Plasma exosomal H19 may serve as an important biomarker for predicting medical responses of patients with prolactinomas.

Long non-coding RNAs H19, MALAT1 and MIAT as potential novel biomarkers for diagnosis of acute myocardial infarction

In this study, we evaluated the potential of peripheral blood mononuclear cells (PBMC) derived long non-coding RNAs (lncRNAs) as biomarkers for acute myocardial infarction (AMI). To assess the value of PBMCs-derived lncRNAs levels in predicting clinical outcomes in AMI. We measured the PBMC-derived levels of 10 individual lncRNAs which are known to be relevant to cardiovascular disease in PBMCs from 132 AMI patients and 104 healthy participants using quantitative RT-PCR. For AMI group, blood sample were obtained from patients after the onset of AMI. Out of the 10 lncRNAs tested, the mRNA level of lncRNA H19, MIAT and MALAT1 were significantly higher in AMI patients than in healthy control (2.3 ± 0.2 vs. 1.0 ± 0.1, p < 0.001, 1.5±0.1 vs. 1.0±0.1, p = 0.002, 1.8±0.2 vs. 1.0±0.1, p < 0.001, respectively). Receiver operating characteristic curve analyses showed that PBMC-derived H19 had significant diagnostic value for AMI (AUC, 0.753; 95% CI, 0.689˜0.817).Multivariate logistic regression analysis showed that H19 as a dangerous risk for AMI (OR = 2.498, 95% CI, 1.321–4.726, p = 0.005). In addition, the lncRNA H19 alteration was inversely associated with a number of cardiovascular protective factors, and positively associated with cardiovascular risk factors, such as high-density lipoprotein (HDL) (r=-0.198, p = 0.010), lipoprotein A (r=-0.153, p = 0.049), white blood cell counting (r=0.301, p < 0.001) and cardiac ejection fraction (r=-0.157, p = 0.042). Moreover, lncRNA H19 was positively correlated with cardiac biomarkers, i.e. troponinT (r=0.344,p < 0.001), CK (r=0.261, p = 0.001) and CKMB (r=0.24, p = 0.002).Hence, elevated expression level of PBMC-derived H19, MIAT and MALAT1 may be considered as novel biomarkers of AMI.

Abstract 758: Long non-coding RNAH19 as a competing endogenous RNA of KIT in the progression of GIST

Abstract Background: Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor with a wide range of biological behaviors, and activating mutations of KIT are particularly crucial to its development. Long non-coding RNAs (lncRNAs) have been identified as an oncogene in multiple cancer types, but data in GIST are missing. The aim of the study was to evaluate the contribution of lncRNA H19 to the malignant course of GISTs. Materials and Methods: RNA-seq analysis of lncRNA expression in five paired GIST and ANTs were analyzed to screen for differentially expressed lncRNAs. 274 human lncRNAs analyzed, exhibited significantly differential expressions with 186 genes being up-regulated and 88 genes being down-regulated compared to ANT (fold change&amp;gt;5, p&amp;lt;0.05). Unsupervised hierarchical clustering identified 48 lncRNAs and lncRNA H19 was the most overexpressed novel one. To assess the clinical importance of lncRNA H19 upregulation in GISTs, based on the expression levels 92 GIST patients were categorized into two groups: high (n=46) and low (n=46) lncRNA H19 expression. Results: After identifying lncRNA H19 as the most important one, validation in the expanded cohort confirmed the frequent upregulation of lncRNA H19 in GIST. Functional assays revealed that lncRNA H19 promotes GIST cell proliferation and metastasis. We could demonstrate that lncRNA H19 was predicted to sponge 19 miRNAs with six potential binding sites shared with KIT 3’-UTR using bio-informative database analysis. It was confirmed that lncRNA H19 was positively correlated with the expression of KIT, and the knockdown of lncRNA H19 led to the depression of KIT and its downstream pathway. Using luciferase reporter assays, lncRNA H19 was observed to sponge several miRNAs including miR-18a/b-5p, miR-19a/b-3p, miR-193a/b-3p, miR-216b-5p and mIR-3666. The combination of lncRNA H19 and KIT inhibition showed increased growth inhibition in GIST cell lines. Taken together, the data indicate that lncRNA H19 functions as a competing endogenous RNA (ceRNA) of KIT and promotes GIST progression. It could be shown that the expression levels of lncRNA H19 were significantly correlated with the progression markers of GIST, tumor size (P&amp;lt;0.001), mitosis count (P&amp;lt;0.001), and the modified NIH risk criteria (P&amp;lt;0.001) Conclusion: Our results indicate that lncRNA H19 can affect imatinib-induced apoptotic cell death and play a role in facilitating GIST progression by modulating the expression of KIT through the ceRNA network. High lncRNA H19 expression is associated with worse prognosis in patients with GIST and lncRNA H19 may serve as a new predictor of prognosis. Furthermore, targeting lncRNA H19/KIT ceRNA network appears to be a promising treatment for GIST. Citation Format: Peter Hohenberger, Lin Tu, Wenyi Zhao, Ming Wang Wang, Zizheng Zhang, Heike Allgayer, Hui Cao. Long non-coding RNAH19 as a competing endogenous RNA of KIT in the progression of GIST [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 758.

Angelica polysaccharide promotes proliferation and osteoblast differentiation of mesenchymal stem cells by regulation of long non-coding RNA H19: An animal study.

ObjectivesOsteoporosis is a systemic bone metabolic disease, which often occurs among the elderly. Angelica polysaccharide (AP) is the main component of angelica sinensis, and is widely used for treating various diseases. However, the effects of AP on osteoporosis have not been investigated. This study aimed to uncover the functions of AP in mesenchymal stem cell (MSC) proliferation and osteoblast differentiation.MethodsMSCs were treated with different concentrations of AP, and then cell viability, Cyclin D1 protein level, and the osteogenic markers of runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), alkaline phosphatase (ALP), bone morphogenetic protein 2 (BMP-2) were examined by Cell Counting Kit-8 (CCK-8) and western blot assays, respectively. The effect of AP on the main signalling pathways of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) and Wnt/β-catenin was determined by western blot. Following this, si-H19#1 and si-H19#2 were transfected into MSCs, and the effects of H19 on cell proliferation and osteoblast differentiation in MSCs were studied. Finally, in vivo experimentation explored bone mineral density, bone mineral content, and the ash weight and dry weight of femoral bone.ResultsThe results revealed that AP significantly promoted cell viability, upregulated cyclin D1 and increased RUNX2, OCN, ALP, and BMP-2 protein levels in MSCs. Moreover, we found that AP notably activated PI3K/AKT and Wnt/β-catenin signalling pathways in MSCs. Additionally, the relative expression level of H19 was upregulated by AP in a dose-dependent manner. The promoting effects of AP on cell proliferation and osteoblast differentiation were reversed by H19 knockdown. Moreover, in vivo experimentation further confirmed the promoting effect of AP on bone formation.ConclusionThese data indicate that AP could promote MSC proliferation and osteoblast differentiation by regulating H19.Cite this article: X. Xie, M. Liu, Q. Meng. Angelica polysaccharide promotes proliferation and osteoblast differentiation of mesenchymal stem cells by regulation of long non-coding RNA H19: An animal study. Bone Joint Res 2019;8:323–332. DOI: 10.1302/2046-3758.87.BJR-2018-0223.R2.

Downregulation of H19 decreases the radioresistance in esophageal squamous cell carcinoma cells.

Background: Radiotherapy is one of the most common treatments for esophageal squamous cell carcinoma (ESCC). Radioresistance is a major obstacle that limits the efficacy of radiotherapy. H19 has been considered as a factor affecting radioresistance, whereas the specific mechanism of H19 in ESCC radioresistance remains to be further elucidated.Purpose: The objective of this study was to identify the relationship between H19 and radioresistance. The findings are expected to provide new insights into the treatment of radioresistant ESCC.Methods: The expression levels of H19 in ESCC was analyzed using the online database starBase. The Oncomine database was used to further verify the association between H19 expression and patient age, gender, and tumor stage. The overall survival rates of ESCC patients were analyzed using the KM plotter database. Clonogenic survival was conducted to identify the value of survival fraction. The optical density values were obtained via MTS assays. Cells migration and stemness were observed through Transwell and sphere formation assays. The expression levels of H19, miR-22-3p and WNT1 were analyzed using qPCR.Results: In our study, we firstly screened the H19 according to the online database starBase, and then the Oncomine database and KM plotter database showed that H19 expression was significantly upregulated in the ESCC tissues and associated with poor prognosis. Secondly, an ESCC radioresistant cell line, KYSE150R was established. Clonogenic survival showed that radiation decreased the value of survival fraction. MTS assays suggested that optical density values in KYSE150R cells were significantly higher than that in KYSE150 cells. Transwell and sphere formation assays showed radiation enhanced cell migration and stemness in ESCC cells. In addition, qPCR showed that H19 was upregulated in KYSE150R cells, and survival fraction assays showed that knockdown of H19 decreased the survival fraction values. MTS assays, migration and invasion assays suggested that H19 inhibited cells proliferation, migration and stemness in radioresistant KYSE150 cells. Moreover, qPCR assay showed that miR-22-3p expression levels was downregulated, but WNT1 was upregulated in KYSE150R cells as well as protein levels. Luciferase activity assay further showed that miR-22-3p inhibits the WNT1 expression.Conclusion: Our results demonstrate that H19 knockdown downregulates the WNT1 via upregulating miR-22-3p expression, which leads to the inhibition of cells proliferation, migration and stemness in the radioresistant ESCC cells.

LncRNA H19 promotes epithelial mesenchymal transition and metastasis of esophageal cancer via STAT3/EZH2 axis.

Long non-coding RNA H19 (lncRNA H19) has been widely reported in esophageal cancer (EC), and previous study had found that lncRNAH19 was up-regulated in EC and promoted cell proliferation and metastasis. However, the mechanism still needs further studied. Levels of lncRNA H19 were analyzed by qRT-PCR in matched samples from 30 patients. Expression levels of lncRNA H19, let-7, STAT3 and EZH2 were additionally identified by qRT-PCR and western blotting in five EC cell lines. The effects of lncRNA H19 on cell proliferation, migration, invasion and apoptosis in cell lines were performed by MTT assay, colony formation assay, Transwell assay and flow cytometry in vitro, and tumor formation was detected by xenograft nude mice model in vivo. The expression level of STAT3, EZH2, β-catenin, and EMT and metastasis related molecules such as E-cadherin, N-cadherin, Snail-1 and MMP-9 was assessed by qRT-PCR and western blotting. Finally, luciferase reporter assay and RIP assay were used to verify the interaction between lncRNA H19 and let-7c, and their subsequent regulation of STAT3. Knockdown of lncRNA H19 repressed cell proliferation, migration and invasion as well as EMT and metastasis via STAT3-EZH2-β-catenin pathway, while lncRNA H19 regulated STAT3 negatively regulated let-7c in EC cell lines. lncRNA H19 facilitates EMT and metastasis of EC through let-7c/STAT3/EZH2/β-catenin axis.

Long non-coding RNA H19 promotes the proliferation and invasion of lung cancer cells and regulates the expression of E-cadherin, N-cadherin, and vimentin.

BackgroundThis study aimed to explore the effect of long non-coding RNA (LncRNA) H19 on the proliferation and invasion of lung carcinoma cells A549, and to determine its molecular targets.MethodsA549 cells were with either LncRNA H19 or LncRNA H19 shRNA, and the expression levels of LncRNA H19 were evaluated by quantitative real-time PCR (RT-PCR). We measured cell proliferation using the CCK-8 assay, cell counting assays, and colony formation assay in response to shLncRNA H19-2. Cell migration and invasion were assessed by wound healing assay and Transwell assay, respectively. The mRNA and protein expression levels of E-cadherin, N-cadherin, and vimentin were determined by RT-PCR and western blot, respectively.ResultsThe three LncRNA H19 shRNAs used in our study significantly reduced the expression levels of LncRNA H19 in A549 cells (P<0.05). Moreover, LncRNA H19 shRNA 2 (shLncRNA-2) was the most potent inhibitor of LncRNA H19 expression, and was selected for further experimentation. Transfection with shLncRNA H19-2 significantly decreased the proliferation, migration, and invasion of A549 cells, while overexpression of LncRNA H19 had the opposite effect in these cells (P<0.05). In response to shLncRNA H19-2, the expression levels of E-cadherin were notably elevated (P<0.05), while the expression levels of N-cadherin and vimentin were decreased (P<0.05). In contrast, overexpression of LncRNA H19 induced the expression of E-cadherin, and blocked the expression of N-cadherin, and vimentin (P<0.05).ConclusionOur results suggest that LncRNA H19 mediates the proliferation and invasion of lung cancer cells via upregulation of N-cadherin and vimentin, and downregulation of E-cadherin.

Increased H19 Long Non-coding RNA Expression in Schwann Cells in Peripheral Neuropathic Pain.

Neuropathic pain is an intractable chronic pain condition caused by damage to the somatosensory system. Although non-coding RNAs such as microRNAs are important regulators of neuropathic pain, the role of long non-coding RNAs (lncRNAs) is poorly understood. This study used a rat model of neuropathic pain induced by lumbar fifth spinal nerve ligation (SNL). Microarray analysis of lncRNAs in the lumbar fifth dorsal root ganglion was performed at day 14 after SNL. Expression levels of H19 were examined by using quantitative PCR. In situ hybridization was used to determine the distribution of H19 at day 14 after SNL. Schwann cells were isolated from peripheral nerves at day 14 after SNL. H19 lncRNA was greatly increased in the L5 dorsal root ganglion at day 14 after SNL and was significantly higher at and after day 4. In the dorsal root ganglion, H19 was detected mainly in non-neuronal cells but not in primary sensory neurons. Consistent with this, H19 expression was upregulated in Schwann cells isolated from peripheral nerves after SNL. Increased H19 lncRNA in Schwann cells might be involved in neuropathic pain.

The expression profiling of circulating miR-204, miR-182, and lncRNA H19 as novel potential biomarkers for the progression of peptic ulcer to gastric cancer.

Deregulation of noncoding RNAs, microRNAs (miRNAs) and long noncoding RNA (lncRNA), are implicated in the initiation and progression of gastric cancer (GC). This study is a pilot case-control study carried out on 75 subjects, 40 of them were Helicobacter pylori-gastric ulcer patients and 35 were GC patients recruited from the Gastrointestinal Endoscopy Unit in Al-Kasr Al-Aini Hospital, Cairo University in Egypt. Real-time PCR was performed to evaluate the expression level of serum miR-204, miR-182, and lncRNA H19 in patients with peptic ulcer-progressed GC vs nonprogressed peptic ulcer patients. Fibroblast growth factor 18 (FGF-18)/FGF receptor 2 (FGFR2) expression and their downstream immunological and inflammatory signaling markers were assessed and their association with the addressed noncoding RNAs investigated. As regards miR-204 and miR-182, they were significantly increased (12.5 and 2.6 folds, respectively) in GU samples, compared with those of healthy control levels. The elevated levels of these miRNAs were significantly de-escalated in GC samples compared with GU and the fold decrease valued 2.2 fold for miR-204 and 1.8 folds for miR-182. On the other hand, the significant escalation in the level of lnRNA H19 in GU recorded a 16.6 fold increase and further elevation in its levels was evident in GC samples. The herein assessed miRNAs are correlated with disease duration and FGFR2 with miR-182 being significantly correlated with all inflammatory markers, TAC, INF-γ, matrix metallopeptidase 9, and FGF-18. In terms of diagnostic accuracy of assessed miRNAs (stages III to IV), the receiver operating characteristic analysis indicated that serum lncRNA H19 showed the highest diagnostic accuracy (95.5%), specificity (100%), and sensitivity (90.9%), compared with miR-204 and miR-182, which showed the same specificity (60%), sensitivity (72.7%), and diagnostic accuracy (68.8%). Our findings conclude that lnRNA H19, miR-204, and miR-182 may function as novel prospective plasma biomarkers to detect GC and its progression from H. pylori-peptic ulcer, which would be helpful to improve the theranostics of GC.

A negative feedback loop of H19/miR-675/EGR1 is involved in diabetic nephropathy by downregulating the expression of the vitamin D receptor.

We aimed to explore the regulatory relationship among the long noncoding RNA H19, micorRNA-675 (miR-675), the vitamin D (VD) receptor (VDR), and the early growth response protein 1 (EGR1)in the pathogenesis of diabetic nephropathy (DN) among patients with diabetes mellitus (DM). Expression levels of H19, miR-675, VDR, and EGR in patients or CIHP-1/HEK 293 cells were measured via quantitative reverse-transcription polymerase chain reaction and western blot analysis. Computational analysis and luciferase assays were performed to determine EGR1 as a target gene of miR-675. The relative expression of miR-675 was higher in the presence of H19, whereas the expression of both VDR and EGR1 messenger RNA was decreased in the presence of H19 or miR-675. However, relative expression of H19 and miR-675 was increased, whereas VDR expression was suppressed upon the treatment of 1,25-dihydroxyvitamin D3 or EGR1. VDR was identified as a target gene of miR-675. The H19 promoter and EGR1 increased the luciferase activity of cells transfected with wild-type VDR. Compared with DM patients free of DN, the levels of H19 and miR-675 were increased in the DN(+) group, whereas the levels of VDR and EGR1 were decreased. In summary, the above results indicate the presence of a negative feedback loop in the pathological mechanism of DN, where H19 downregulates the expression of VDR by upregulating the expression of miR-675, whereas reduced VDR expression subsequently reduced the expression of EGR1. Moreover, reduced EGR1 expression inhibits H19 expression, thus forming a negative feedback loop required to maintain the homeostasis of VDR and to reduce the incidence of DN.

H19 regulates trophoblastic spheroid adhesion by competitively binding to let-7.

Integrin β3 (ITGB3), which is the target gene of the miRNA let-7 that can be antagonized by long noncoding RNA (lncRNA) H19, is well known to have a critical role in endometrium receptivity. However, the regulation of ITGB3 in cell–cell or cell–extracellular matrix adhesion and invasion for the maintenance of early pregnancy remains unknown. This study aimed to explore the role of the H19/let-7/ITGB3 axis in regulating trophoblastic spheroid adhesion and in vitro invasion ability using the HTR-8/SVneo cell line and to investigate the expression levels of lncRNA H19 and ITGB3 in human products of conception. The in vitro knockdown of H19 resulted in decreased expression of ITGB3 at the mRNA and protein levels and reduced the adhesion and invasion ability. In the embryonic chorion tissue of spontaneous abortion (SA), the expressions of H19 and ITGB3 at both the mRNA and protein levels decreased. The results of quantitative RT-PCR, Western blot analysis, dual-luciferase report gene and functional miRNA let-7 rescue experiments, adhesion assay and in vitro transwell invasion assay confirmed that H19 regulated trophoblastic spheroid adhesion with endometrial stromal cells through the H19/let-7/ITGB3 axis, thereby providing an improved understanding of the molecular mechanism of SA.

Effect of supplemetation of Zebularine and Scriptaid on efficiency of in vitro developmental competence of ovine somatic cell nuclear transferred embryos

Somatic cell nuclear transfer (SCNT) technology has been applied in the construction of disease model, production of transgenic animals, therapeutic cloning, and other fields. However, the cloning efficiency remains limited. In our study, to improve SCNT efficiency, brilliant cresyl blue (BCB) staining were chosen to select recipient oocytes. In addition, DNA methyltransferase inhibitor Zebularine (5 nmol/L) and histone deacetylase inhibitor Scriptaid (0.2 μmol/L) were jointly used to treat sheep donor cumulus cells and reconstructed embryo. Moreover, the expression levels of embryonic development-related genes (OCT4, SOX2, H19, IGF2 and Dnmt1) of reconstructed embryo were also detected. Using BCB + oocytes as recipient cell, donor cumulus cells and reconstructed embryos were treated with 5 nmol/L Zebularine and 0.2 μmol/L Scriptaid, the blastocyst rate in Zeb + SCR-SCNT group (28.25%) was significantly higher than SCNT (21.16%) (p < 0.05). Furthermore, results showed that expression levels of OCT4, SOX2, H19, IGF2 and Dnmt1 genes in Zeb + SCR-SCNT embryos were more similar to IVF embryos. Our study proved that 5 nmol/L Zebularine and 0.2 μmol/L Scriptaid treating with sheep donor cumulus cells and reconstructed embryos improved SCNT blastocyst rate and relieve the abnormal expression of embryonic developmental related genes.

The overexpression of lncRNA H19 as a diagnostic marker for coronary artery disease.

Our study aimed to investigate the diagnostic value of lncRNA H19 for coronary artery disease (CAD) and to explore its possible mechanisms. A total of 30 CAD patients and 30 healthy individuals, as well as patients with different cardiovascular diseases, were included in this study. Blood was drawn from each participant to prepare serum samples, and the expression of lncRNA H19 was detected using qRT-PCR. The ROC curve analysis was used to analyze the diagnostic value of H19 for CAD. The effects of patients' basic information and lifestyle on H19 expression were analyzed. The plasma level of TGF-β1 was measured by ELISA. The H19 overexpression in the human primary coronary artery endothelial cell (HCAEC) line was constructed, and the effects of H19 overexpression on the TGF-β1 expression were analyzed using Western blot. The results of H19 expression were specifically upregulated in patients with CAD but not in healthy individuals and patients with other types of cardiovascular diseases. The ROC curve analysis showed that the H19 expression level could be used to predict CAD accurately. Gender, age, and patients' lifestyle had no significant effects on H19 expression, but H19 expression was higher in patients with a longer course of disease in comparison with the controls. H19 expression was positively correlated with the serum level of TGF-β1, and H19 overexpression significantly increased TGF-β1 protein level in HCAEC. H19 overexpression participates in the pathogenesis of CAD by increasing the expression level of TGF-β1, and H19 expression level may serve as a diagnostic marker for CAD.

Progesterone ameliorates the endometrial polyp by modulating the signaling pathway of Wnt and β-catenin via regulating the expression of H19 and miR-152.

MicroRNAs and long noncoding RNAs are believed to play important roles in the pathogenesis of various diseases. This study aimed to explore the potential mechanism of the involvement of H19 and miR-152 in an endometrial polyp. Luciferase assay was conducted to determine the effect of progesterone. Real-time polymerase chain reaction (PCR) and western blot were performed to detect the influence of progesterone on miR-152 and Wnt1. MTT assay and flow cytometry (FCM) were utilized to detect the effect of progesterone on cell proliferation and apoptosis. In silicon analysis, luciferase assay, real-time PCR, and immunohistochemistry (IHC) were performed to explore the regulatory relationship between H19 and miR-152 or miR-152 and Wnt1. Progesterone dose-dependently increased the H19 expression level through driving the promoter efficiency of H19. Then, progesterone upregulated Wnt1 level and downregulated miR-152 in a dose-dependent manner in ECC1 and HEC1A cells. Administration of progesterone inhibited cell viability and promoted cell apoptosis. H19 negatively regulated miR-152 expression by binding to miR-152. Furthermore, Wnt1 was identified as a virtual target gene of miR-152 and was inhibited by miR-152. Progesterone receptors mRNA and miR-152 were lowly expressed in participants with an endometrial polyp, while the levels of H19 and Wnt1 were much higher in the endometrial polyp group compared with normal controls. H19 negatively regulated miR-152 and miR-152 negatively regulated Wnt1, with the negative correlation coefficients being -0.500 and -0.500, respectively. Using IHC, it was found that Wnt1 and Bcl-2 protein were highly expressed in the endometrial polyp group compared with normal controls. The results suggested that H19 was associated with endometrial polyp via mediating cell proliferation and apoptosis.

LncRNA H19 Suppresses Metastasis of Follicular Thyroid Carcinoma &lt;i&gt;via&lt;/i&gt; the IGF1/JAK/STAT Pathway

Long noncoding RNAs (lncRNAs), which are important functional regulators in cancer, have emerged as critical molecular regulators in various biological processes. However, the mechanisms by which H19 modulates follicular thyroid cancer (FTC) development remain largely unknown. Here, we found that the H19 expression level was lower in tumour tissues than in adjacent normal tissues of FTC patients by using next-generation sequencing technology. The results of functional experiments showed that overexpression of H19 inhibited the proliferation, migration, invasion and angiogenic ability of FTC cells. By using Gene Ontology (GO) and KEGG pathway enrichment analyses, we found that the IGF1 signalling pathway was one of the most enriched pathways within the set of upregulated mRNAs, suggesting that it may be involved in the pathogenesis and development of FTC. In this mechanism, H19 recruited the transcription factor Pax5 to the promoter region of suppressors of cytokine signalling 3 (SOCS3), which activated SOCS3 transcription. Moreover, H19 functioned as a competing endogenous RNA (ceRNA) acting as a sponge for miR-29-3p, which activated IGF2BP1. Collectively, these data indicate that H19 is a tumour suppressor in the development of FTC and that it may contribute to the improvement of FTC diagnosis and therapy. Funding Statement: This work was supported by the grants from the Science Research Project of the Department of Education of Liaoning Province (No. L2016011) and the Doctoral Scientific Research Fund of the Second Hospital of Dalian Medical University (DY2YBSQD201501). Declaration of Interests: The authors declare that they have no conflict of interest. Ethics Approval Statement: Samples were obtained from patients who provided informed consent, in accordance with the ethical standards of the Second Hospital of Dalian Medical University (Dalian, China). All of the animal experiments were approved by the responsible governmental animal ethics committee and complied with the ARRIVE guidelines.

Long non-coding RNA H19 is associated with polycystic ovary syndrome in Chinese women: a preliminary study.

Polycystic ovary syndrome (PCOS) represents a serious reproductive and endocrine condition and is associated with high incidence rates. H19 is a compelling long noncoding RNA (lncRNA) which carries out a range of biological functions. However, prior to this study, little was known as to whether there was an association between lncRNA H19 and PCOS. In the current study, we used quantitative real-time polymerase chain reaction (qRT-PCR) to determine lncRNA H19 expression levels in peripheral blood leukocytes from patients with PCOS and compared this data with that derived from normal controls. We also screened data for potential relationships between lncRNA H19 and a range of endocrine variables in PCOS. The expression of lncRNA H19 was significantly higher in cases of PCOS than in controls. Individuals exhibiting higher expression levels of lncRNA H19 were associated with a significantly higher risk of PCOS than those with lower expression levels. Moreover, lncRNA H19 expression was positively correlated with fasting plasma glucose levels; this was the case with both raw data, and after adjustment for age and BMI in the PCOS group. However, lncRNA H19 expression showed no significant correlation with total testosterone or insulin resistance in either PCOS cases or the controls. In conclusion, we demonstrate the first evidence to indicate that lncRNA H19 is associated with PCOS, suggesting that elevated lncRNA H19 levels are a risk factor for PCOS. For susceptible individuals, lncRNA H19 may represent a useful biomarker of the early stages of endocrine and metabolic disorders in PCOS.

Targeted inhibition of long non-coding RNA H19 blocks anaplastic thyroid carcinoma growth and metastasis

ABSTRACT Long non-coding RNA H19 (H19) is highly expressed in cancers and is considered to highly correlate with the extent of malignant degree. The present study was performed to determine the expression levels of H19 in anaplastic thyroid carcinoma (ATC) tissues and the role of H19 in ATC 8505C cells in vitro and in vivo. Expression of H19 was detected in 19 ATC and 19 normal thyroid tissues by real-time quantitative polymerase chain reaction. Utilizing the siRNA or short hairpin RNA (shRNA) directed against human H19 (H19 siRNA or shRNA H19) depleted H19 in ATC 8505C cells and characterized the outcomes. The results showed that H19 was overexpressed in ATC tissues. Targeting H19 inhibited proliferation, migration, and invasion and induced apoptosis in 8505C cells in vitro and inhibited tumorigenesis and metastasis in vivo. Therefore, the H19 might be an effective target for ATC molecular therapy.