MicroRNA let-7g regulates mouse granulosa cell autophagy by targeting insulin-like growth factor 1 receptor

As an important type of somatic cell, granulosa cells play a major role in deciding the fate of follicles. Therefore, analyses of granulosa cell apoptosis and follicular atresia have become hotspots of animal research. Autophagy is a cellular catabolic mechanism that protects cells from stress conditions, including starvation, hypoxia, and accumulation of misfolded proteins. However, the relationship between autophagy and apoptosis in granulosa cells is not well known. Here, we demonstrate that let-7g regulates the mouse granulosa cell autophagy signaling pathway by inhibiting insulin-like growth factor 1 receptor expression and affecting the phosphorylation of protein kinase B/mammalian target of rapamycin. Small interference-mediated knockdown of insulin-like growth factor 1 receptor significantly promoted autophagy signaling of mouse granulosa cells. In contrast, overexpression of insulin-like growth factor 1 receptor in mouse granulosa cells attenuated autophagy activity in the presence of let-7g. In addition, overexpression of let-7g increased the apoptosis rate, as indicated by an increased number of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells. Finally, 3-methyladenine as well as the lysosomal enzyme inhibitor chloroquine partially blocked apoptosis. In summary, this study demonstrates that let-7g regulates autophagy in mouse granulosa cells by targeting insulin-like growth factor 1 receptor and downregulating protein kinase B/mammalian target of rapamycin signaling, and that mouse granulosa cell autophagy induced by let-7g participates in apoptosis.     

The miRNA hsa-miR-6515-3p potentially contributes to lncRNA H19-mediated-lung cancer metastasis

Aberrant expression of long noncoding RNAs (lncRNAs) contributes to all phenotypes of cancer including metastasis, which is a major cause of death in many advanced malignancies. One particular lncRNA, H19, is found to be a crucial player in cancer progression by modulating multiple microRNAs (miRNAs). In this study, we screened miRNAs possibly associated with H19 using lung carcinoma cell lines and patient with lung cancer tissues, and selected one possible hit, hsa-miR-6515-3p, to perform in vitro functional assays. Its inhibition leads to decreased proliferation and migration of SPC-A1 lung cancer cells and is in good correlation with H19-knockdown groups. These results indicate that H19 may be an epigenetic regulator of miR-6515-3p, and its dysregulation may contribute to lung cancer progression and metastasis.     

lncRNA H19 contributes to oxidative damage repair in the early age‐related cataract by regulating miR‐29a/TDG axis

Age‐related cataract (ARC) is caused by the exposure of the lens to UVB which promotes oxidative damage and cell death. This study aimed to explore the role of lncRNA H19 in oxidative damage repair in early ARC. lncRNAs sequencing technique was used to identify different lncRNAs in the lens of early ARC patients. Human lens epithelial cells (HLECs) were exposed to ultraviolet irradiation; and 8‐OHdG ELISA, Cell counting kit 8 (CCK8), EDU, flow cytometry and TUNEL assays were used to detect DNA damage, cell viability, proliferation and apoptosis. Luciferase assay was used to examine the interaction among H19, miR‐29a and thymine DNA glycosylase (TDG) 3'UTR. We found that lncRNA H19 and TDG were highly expressed while miR‐29a was down‐regulated in the three types of early ARC and HLECs exposed to ultraviolet irradiation, compared to respective controls. lncRNA H19 knockdown aggravated oxidative damage, reduced cell viability and proliferation, and promoted apoptosis in HLECs, while lncRNA H19 overexpression led to opposite effects in HLECs. Mechanistically, miR‐29a bound TDG 3'UTR to repress TDG expression. lncRNA H19 up‐regulated the expression of TDG by repressing miR‐29a because it acted as ceRNA through sponging miR‐29a. In conclusion, the interaction among lncRNA H19, miR‐29a and TDG is involved in early ARC. lncRNA H19 could be a useful marker of early ARC and oxidative damage repair pathway of lncRNA H19/miR‐29a/TDG may be a promising target for the treatment of ARC.     

LncRNA H19: A novel oncogene in multiple cancers

Long non-coding RNAs (lncRNAs) are a series of non-coding RNAs that lack open reading frameworks. Accumulating evidence suggests important roles for lncRNAs in various diseases, including cancers. Recently, lncRNA H19 (H19) became a research focus due to its ectopic expression in human malignant tumors, where it functioned as an oncogene. Subsequently, H19 was confirmed to be involved in tumorigenesis and malignant progression in many tumors and had been implicated in promoting cell growth, invasion, migration, epithelial-mesenchymal transition, metastasis, and apoptosis. H19 also sequesters some microRNAs, facilitating a multilayer molecular regulatory mechanism. In this review, we summarize the abnormal overexpression of H19 in human cancers, which suggests wide prospects for further research into the diagnosis and treatment of cancers.     

Knockdown of lncRNA H19 inhibits abnormal differentiation of small intestinal epithelial cells in diabetic mice

Diabetes mellitus (DM) comprises a group of metabolic diseases characterized by insulin deficiency or resistance and hyperglycemia. We previously reported the presence of abnormal differentiation of small intestinal epithelial cells (IECs) in diabetic mice, but the exact mechanism of this phenomenon has not been thoroughly elucidated to date. In this study, we found that H19 was markedly upregulated in IECs of DM mice. H19 knockdown significantly inhibited abnormal differentiation of IECs in DM mice. Bioinformatics analysis identified miR-141-3p as a candidate for H19. Based on luciferase reporter assays, we found that miR-141-3p directly targeted H19. Luciferase reporter assays also showed that miR-141-3p could directly target β-catenin. Furthermore, H19 might act as an endogenous “sponge” by competing for miR-141-3p binding to regulate miRNA targets in vitro and in vivo. In summary, our findings provide the first evidence supporting the role of H19 in IECs of DM mice, and miR-141-3p targets not only protein-coding genes but also the lncRNA H19.     

外泌体介导长链非编码RNA H19促进肝癌细胞增殖与转移

外泌体是由细胞分泌的直径为30～150 nm的小囊泡,含有丰富的mRNA、microRNA和长链非编码RNA(lncRNA)。目前,大多数外泌体研究都集中在mRNA和microRNA,而对lncRNA的生物学功能并不十分清楚。研究表明,肿瘤细胞外泌体 lncRNA H19在肿瘤细胞的增殖、迁移和侵袭中发挥了重要作用。本研究将筛选到的lncRNA H19高表达的肝癌细胞HCCLM3,分别收集其高表达lncRNA H19的外泌体和其下调lncRNA H19表达后的外泌体。然后,将收集到的外泌体分别添加到lncRNA H19低表达的肝癌细胞Hep3B和HepG2孵育液中。孵育24 h后,检测其对肿瘤细胞的增殖、迁移和侵袭能力的影响。结果显示,肝癌细胞HCCLM3可分泌大量的外泌体,且能被其他肿瘤细胞大量摄取;与下调lncRNA H19表达的外泌体相比,lncRNA H19高表达的外泌体能显著增强Hep3B和HepG2细胞的增殖、迁移和侵袭能力。而这一作用可通过激活PI3K/AKT/mTOR通路实现。上述结果表明,lncRNA H19高表达的肝癌细胞以外泌体方式,增强邻近肝癌细胞的增殖、迁移和侵袭能力,促进肝癌的发生与发展。     

Long noncoding RNA H19 participates in metformin-mediated inhibition of gastric cancer cell invasion

Recent research suggests that the first-line oral antidiabetes drug metformin may prevent gastric cancer progression and improve prognosis. Many studies have also shown that long noncoding RNAs (lncRNAs) play important roles in many biological processes. Therefore, we aimed to explore whether lncRNAs participate in the mechanisms by which metformin affects gastric cancer cells. In the current study, we found that metformin significantly inhibited the cellular functions of gastric cancer cells through Cell Counting Kit-8 and invasion assays. We found that lncRNA H19 was greatly downregulated in gastric cancer cells treated with metformin using lncRNA microassays. Based on bioinformatics analyses of the Oncomine and The Cancer Genome Atlas databases, H19 is shown to be overexpressed in gastric cancer tissues, with increased expression of H19 relating to advanced pathological tumor stage and pathological tumor node metastasis stage, indicating that H19 may be associated with the invasive ability of gastric cancer. We knocked down H19 in AGS and SGC7901 cell lines and found that knocked-down H19 could decrease gastric cancer cell invasion and that metformin could not further decrease invasion after the knock down. Moreover, H19 depletion increased AMPK activation and decreased MMP9 expression, and metformin could not further activate AMPK or decrease MMP9 in H19 knocked-down gastric cancer cells. In summary, metformin has a profound antitumor effect on gastric cancer cells, and H19 is a key component in the process of metformin suppressing gastric cancer cell invasion.     

Exosome-mediated transfer of long noncoding RNA H19 induces doxorubicin resistance in breast cancer

Development of the acquired resistance is one major obstacle during chemotherapy for cancer patients. Exosomes mediate intercellular communication and cause environmental changes in tumor progression by transmitting active molecules. In this study, the role of long noncoding RNA H19 within exosomes is elucidated in terms of regulating doxorubicin (DOX) resistance of breast cancer. As a result, increased H19 expression was observed in DOX-resistant breast cancer cells in comparison with the corresponding parental cells. Suppression of H19 significantly lowered DOX resistance by decreasing cell viability, lowering colony-forming ability, and inducing apoptosis. Moreover, extracellular H19 could be moved to sensitive cells via being incorporated into exosomes. Treating sensitive cells with exosomes from resistant cells increased the chemoresistance of DOX, while downregulation of H19 in sensitive cells abated this effect. Taken together, H19 could be delivered by exosomes to sensitive cells, leading to the dissemination of DOX resistance. Our finding highlights the potential of exosomal H19 as a molecular target to reduce DOX resistance.     

LncRNA PTCSC3 inhibits triple-negative breast cancer cell proliferation by downregulating lncRNA H19

Long noncoding RNA (lncRNA) PTCSC3 (hereafter PTCSC3 is used to represent lncRNA PTCSC3) inhibits glioma and thyroid cancer, indicating its potential tumor suppression function in other types of cancers. We explored the potential involvement of PTCSC3 in triple-negative breast cancer (TNBC). In the current study, we found that PTCSC3 was downregulated in tumor tissues of patients with TNBC. PTCSC3 expression was positively correlated with plasma levels of PTCSC3. LncRNA H19 was upregulated and was inversely correlated with PTCSC3 in tumor tissues. PTCSC3 overexpression led to downregulated H19 in TNBC cells, while H19 overexpression did not affect PTCSC3 expression. PTCSC3 inhibited and H19 promoted proliferation of TNBC cells. H19 overexpression attenuated the effects of PTCSC3 overexpression. Cancer cell migration and invasion were not significantly affected by PTCSC3 overexpression. Therefore, lncRNA PTCSC3 inhibits TNBC cell proliferation by downregulating lncRNA H19.     

H19/miR-107/HMGB1 axis sensitizes laryngeal squamous cell carcinoma to cisplatin by suppressing autophagy in vitro and in vivo

Laryngeal squamous cell carcinoma (LSCC) is the most common malignant tumor, which occurs in the head and neck. Current treatments for LSCC are all largely weakened by increasing drug resistance. Our study aimed to investigate the effects of long noncoding RNA (lncRNA) H19 on drug resistance in LSCC. In our study, we found that the level of H19 was sharply upregulated in LSCC tissues and drug-resistant cells compared with the control. Besides, the expression of high-mobility group B1 (HMGB1) was elevated, and microRNA107 (miR-107) was suppressed in drug-resistant cells compared with the control. Further study revealed that the interference of H19 by short hairpin RNA (shRNA) effectively suppressed high autophagy level and obvious drug resistance in drug-resistant cells. Besides that, miR-107 was predicted as a target of H19 and inhibiting effects of H19 shRNA on autophagy and drug resistance were both reversed by miR-107 inhibitor. Moreover, HMGB1 was predicted as a target of miR-107 in LSCC cells and knockdown of HMGB1 was able to suppress autophagy and drug resistance in LSCC cells. In addition, our investigation demonstrated that H19 shRNA exerted an inhibiting effect on autophagy and drug resistance by downregulating HMGB1 by targeting miR-107. Finally, the in vivo experiment revealed that LV-H19 shRNA strongly suppressed drug resistance compared with the usage of cisplatin individually. Taken together, our research indicated an H19–miR-107–HMGB1 axis in regulating the autophagy-induced drug resistance in LSCC in vitro and in vivo, providing novel targets for molecular-targeted therapy and broadening the research for LSCC.     

Escherichia coli O157:H7 induces attaching-effacing lesions in large intestinal mucosal explants from adult cattle

Attaching-effacing (A/E) lesions following natural and experimental infection with Escherichia coli O157:H7 have been seen in neonatal and 3-4-month-old weanling but not older cattle. To test the hypothesis that the adult bovine large intestinal epithelium is resistant to the development of A/E lesions, colonic and rectal mucosal tissue explants from 18-month-old steers were inoculated with E. coli O157:H7 and examined. Epithelial cells of inoculated explants developed A/E lesions at the bacterial attachment sites, providing evidence that the large intestinal mucosal epithelium may be a site of infection that contributes to carriage of E. coli O157:H7 in adult cattle.     

Silencing H19 regulated proliferation,invasion, and autophagy in the placenta by targeting miR-18a-5p

Fetal growth restriction (FGR) is a serious pregnancy complication associated with increased perinatal mortality and morbidity. It may lead to neurodevelopmental impairment and adulthood onset disorders. Recently, long noncoding RNAs (lncRNAs) were found to be associated with the pathogenesis of FGR. Here we report that the lncRNAH19 is significantly decreased in placentae from pregnancies with FGR. Downregulation of H19 leads to reduced proliferation and invasion of extravillous trophoblast cells. This is identified with reduced trophoblast invasion, which has been discovered in FGR. Autophagy is exaggerated in FGR. Downregulation of H19 promotes autophagy via the PI3K/AKT/mTOR and MAPK/ERK/mTOR pathways of extravillous trophoblast cells in FGR. We also found that the expression level of microRNAs miR-18a-5p was negatively correlated with that of H19. H19 can act as an endogenous sponge by directly binding to miR-18a-5p, which targets IRF2. The expression of miR-18a-5p was upregulated, but IRF2 expression was downregulated after the H19 knockdown. In conclusion, our study revealed that H19 downexpressed could inhibit proliferation and invasion, and promote autophagy by targeting miR-18a-5pin HTR8 and JEG3 cells. We propose that aberrant regulation of H19/miR-18a-5p-mediated regulatory pathway may contribute to the molecular mechanism of FGR. We indicated that H19 may be a potential predictive, diagnostic, and therapeutic modality for FGR.     

Novel lncRNA lncRNA001074 participates in the low salinity–induced response in the sea cucumber Apostichopus japonicus by targeting the let-7/NKAα axis

Salinity fluctuations have severe impacts on sea cucumbers and therefore important consequences in sea cucumber farming. The responses of sea cucumbers to salinity changes are reflected in the expression profiles of multiple genes and non-coding RNAs (ncRNAs). The microRNA (let-7) which is a developmental regulator, the ion transporter gene sodium potassium ATPase gene (NKAα), and the long ncRNA lncRNA001074 were previously shown to be involved in responses to salinity changes in various marine species. To better understand the relationship between ncRNAs and target genes, the let-7/NKAα/lncRNA001074 predicted interaction was investigated in this study using luciferase reporter assays and gene knockdowns in the sea cucumber Apostichopus japonicus. The results showed that NKAα was the target gene of let-7 and NKAα expression levels were inversely correlated with let-7 expression based on the luciferase reporter assays and western blots. The let-7 abundance was negatively regulated by lncRNA001074 and NKAα both in vitro and in vivo. Knockdown of lncRNA001074 led to let-7 overexpression. These results demonstrated that lncRNA001074 binds to the 3′-UTR binding site of let-7 in a regulatory manner. Furthermore, the expression profiles of let-7, NKAα, and lncRNA001074 were analyzed in sea cucumbers after the knockdown of each of these genes. The results found that lncRNA001074 competitively bound let-7 to suppress NKAα expression under low salinity conditions. The downregulation of let-7, in conjunction with the upregulation of lncRNA001074 and NKAα, may be essential for the response to low salinity change in sea cucumbers. Therefore, the dynamic balance of the lncRNA001074, NKAα, and let-7 network might be a potential response mechanism to salinity change in sea cucumbers.     

Long noncoding RNA H19 promotes vascular remodeling by sponging let-7a to upregulate the expression of cyclin D1

Vascular remodeling is mainly caused by excessive proliferation of vascular smooth muscle cells (VSMCs). Noncoding RNAs (ncRNAs) have emerged as important regulators in diverse pathological processes. Previous work has shown the functions and mechanisms of long noncoding RNA H19 (LncRNA H19) on VSMCs. As long noncoding RNAs (lncRNAs) are complex in their mechanisms of action, the aim of the study is to identify if there are any other molecular mechanisms of LncRNA H19 on VSMCs. In vivo studies demonstrated that cyclin D1 was overexpressed in neointima of balloon-injured artery. In vitro studies identified that the overexpression of LncRNA H19 promoted VSMCs proliferation and cyclin D1 upregulation. On the contrary, cellular proliferation and expression of cyclin D1 were inhibited in VSMCs after infection with let-7a. Furthermore, luciferase reporter assays and RNA pull-down assays were used to explore the regulatory mechanism, we found that LncRNA H19 functioned as a competing endogenous RNA (ceRNA) by sponging let-7a to promote the expression of the target gene cyclin D1. In conclusion, LncRNA H19 positively regulated cyclin D1 expression through directly binding to let-7a in VSMCs. Our findings provide new insight into the mechanism of LncRNA H19 in VSMCs proliferation and vascular remodeling, and further indicate the implications of LncRNA H19 in the diagnosis and treatment of vascular proliferative diseases.