lncRNA H19 acts as a ceRNA to regulate the expression of CTGF by targeting miR-19b in polycystic ovary syndrome

The etiology of polycystic ovary syndrome (PCOS) is complex and the pathogenesis is not fully understood. Some studies have shown that dysregulation of ovarian granulosa cells may be related to abnormal follicles and excessive androgen in women with PCOS. Our team has also confirmed the high expression status of H19 in PCOS patients in the early stage. However, the relationship between H19 and miR-19b in the development of PCOS is still unknown. Therefore, we used bioinformatics to predict the binding sites of human H19 and miR-19b, and of miR-19b and CTGF genes. After the silencing and overexpression of H19, real-time polymerase chain reaction (PCR) was used to detect the expressions of H19, miR-19b, and CTGF. Western blotting was used to detect CTGF protein. Proliferation of KGN cells after H19 silencing was detected by CCK8. Flow cytometry was used to detect the apoptosis of KGN cells after H19 silencing. After the overexpression of H19, it was found that the expression of miR-19b gene decreased and the expression of CTGF increased, whereas silencing of H19 did the opposite. In addition, H19 could promote cell proliferation and decrease cell apoptosis. Finally, luciferase reporter assays showed that the 3′-end sequences of lncRNA H19 and CTGF contained the binding site of miR-19b. In conclusion, our study indicated that lncRNA H19 acted as a ceRNA to bind to miR-19b via a "sponge" to regulate the effect of CTGF on KGN cells, which may play a vital role in PCOS.

Long non-coding RNA-H19 promotes proliferation and migration of non-small cell lung cancer cells through sponging microRNA-760 to regulate nanog gene expression / 第二军医大学学报

ObjectiveTo explore the role of long non-coding RNA (lncRNA)-H19 in the proliferation and migration of non-small cell lung cancer (NSCLC) cells and the molecular mechanisms. MethodsThe expressions of lncRNA-H19 in 20 NSCLC tissues and paired non-tumor tissues, which were collected from Changhai Hospital of Naval Medical University (Second Military Medical University) from Oct. 2015 to May 2016, were detected by real-time quantitative PCR (qPCR). We also examined lncRNA-H19 expressions in NSCLC cell lines A549 and NCI-H1299 and normal lung epithelial cell line BEAS-2B by qPCR. The proliferation and migration of A549 cells overexpressing lncRNA-H19 were detected by CCK-8 assay and Transwell assay, respectively. Bioinformatics analysis and duel-luciferase reporter assay were conducted to predict and confirm the interaction between microRNA (miRNA)-760 and lncRNA-H19. Western blotting analysis and RT-qPCR were performed to observe the influence of lncRNA-H19 overexpression on the expression of miRNA-760 and target gene nanog. MiRNA-760 was overexpressed in A549 cells, and its role in lncRNA-H19 promoting proliferation and migration of NSCLC cells was observed. Results The expressions of lncRNA-H19 in NSCLC tissues and A549 and NCI-H1299 cells were significantly upregulated compared with those in normal tissues and BEAS-2B cells (all P<0.01). Overexpression of lncRNA-H19 significantly improved the proliferation ability (P<0.05) and migration ability (P<0.01) of A549 cells compared with the negative control group. The results of starBase v3.0 showed that lncRNA-H19 could specifically adsorb miRNA-760, and duel-luciferase reporter assay showed that lncRNA-H19 directly bound to miRNA-760. Compared with the negative control group, overexpression of lncRNA-H19 significantly inhibited miRNA-760 expression in A549 cells (P<0.05) and promoted the expression of the downstream gene nanog at mRNA and protein levels (all P<0.01). Overexpression of miRNA-760 significantly inhibited lncRNA-H19-induced proliferation and migration of A549 cells (all P <0.05). ConclusionLncRNA-H19 can promote the proliferation and migration of NSCLC cells through sponging miRNA-760 to regulate nanog gene expression.

The long non-coding RNA H19 induces hypoxia/reoxygenation injury by up-regulating autophagy in the hepatoma carcinoma cells

BACKGROUND: Long non-coding RNA H19 (H19) plays an important role by regulating protein expression in different tissues and organs of the body. However, whether H19 induces hypoxia/reoxygenation (h/R) injury via increase of autophagy in the hepatoma carcinoma cells is unknown. RESULTS: H19 was expressed in the hepatoma carcinoma cells (Hep G2 and HCCLM3 cells) and its expression was most in 8 h/24R. The knockdown of H19 and 3-MA (an autophagy inhibitor) protected against h/R-induced apoptosis, cell damage, the expression of cleaved caspase-3 and cleaved caspase-9, the release of cytochrome c (Cyt c). The knockdown of H19 and 3-MA also decreased the autophagic vesicles (AVs) and the expression of Beclin-1 and the ration of LC3-II/LC3-I, and increased cell viability, the expression of Bcl-2 and P62 and the phosphorylation of PI3K, Akt and mTOR. In addition, chloroquine (CQ, an inhibitor of autophagy flux) markedly decreased formation of autophagy flux (the ration of LC3-II/LC3-I). The results of the knockdown of H19 group were similar to those of the 3-MA (or CQ) group. Rapamycin (a mTOR inhibitor, an autophagy activator) further down-regulated h/R-induced decrease of the phosphorylated PI3K, Akt and mTOR. The knockdown of H19 cancelled the effect of rapamycin. The overexpression of H19 further expanded h/R-induced increase of the ration of LC3-II/LC3-I and decrease of the phosphorylated PI3K, Akt and mTOR. CONCLUSIONS: Our results suggest that the long non-coding RNA H19 induces h/R injury by up-regulation of autophagy via activation of PI3K-Akt-mTOR pathway in the hepatoma carcinoma cells.

The expression and effect of long non-coding RNA H19 in BPD neonatal rats / 国际儿科学杂志

Objective To investigate the expression of long non-coding RNA H19 (LncRNA H19)and its regulation of histone methyltransferase 2 (enhancer of zeste homolog 2,EZH2) in the lung tissue of neonatal rats with bronchopulmonary dysplasia (BPD),and to lay a foundation for elucidating the pathogenesis of BPD lung epithelium-interstitial transformation (EMT).Methods The BPD model of SD neonatal rats was induced by hyperoxia (inhalation oxygen concentration was 85％) (n =50),and oxygen inhalation concentration of the control group was 21％ (n =50).The two groups were collected at ld,3d,7d,14d and 21d after birth in lung tissue.Immunohistochemistry,Western blot,real-time quantitative PCR and other techniques were used to detect the intracellular localization,and the expression level of EZH2 protein and the mRNA expression level of H19 and EZH2.Results Immunohistochemistry showed that EZH2 protein was located in the nucleus and cytoplasm of alveolar epithelial cells,and the expression of EZH2 protein in the model group was significantly enhanced compared with the control group.Similarly,the results of Western blot demonstrated that the expression of EZH2 protein in the model group increased from ld (control group:0.196 ± 0.030,model group 0.650 ±0.149) to 21d (control group 0.934 ± 0.215,model group 1.785 ± 0.298) rather than the control group (P ＜ 0.05).Compared with the control group,the mRNA expression level of H19 in the model group increased from 7d (control group 2.591 ± 0.211,model group 3.558 ± 0.093,P ＜ 0.05) and the expression level of EZH2 mRNA started to increase from 3d (control group 1.246 ±0.015,model group 2.148 ± 0.215,P ＜0.05).Moreover,the differences between the two groups were obvious with the time of hyperoxia exposure.Conclusion In the development of BPD,the expression levels of H19 and EZH2 protein in lung tissue is up-regulated,and the peak of H19 expression precedes EZH2,which suggest that H19 might be involved in the pathogenesis of pulmonary dysplasia induced by EZH2-mediated EMT.