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Parthenogenetically activated oocytes cannot develop to term in mammals due to lack of paternal gene expression. Disruption of imprinted gene expression and DNA methylation status in parthenogenetic fetuses has been reported in mice and pigs, but not in rabbits. In this study, the genomic imprinting status of the paternally expressed genes Neuronatin ( NNAT ), Nucleosome assembly protein 1-like 5 ( NAP1L5 ), and Makorin ring finger protein 3 ( MKRN3 ) was compared between rabbit parthenogenetic (PA) and normally fertilized fetuses (Con) using quantitative real-time PCR (qRT-PCR) and bisulfite sequencing PCR (BSP). The results revealed a significantly reduced expression of NNAT , NAP1L5 , and MKRN3 in rabbit PA fetuses compared with Con fetuses ( p < 0.05). In addition, the BSP results demonstrated hypermethylation in the differentially methylated regions (DMRs) of NNAT , NAP1L5, and MKRN3 in rabbit PA fetuses. Taken together, these results suggest that hypermethylation of DMRs is associated with decreased NNAT , NAP1L5, and MKRN3 expression, which may be responsible for developmental failure of rabbit PA fetuses. [ABSTRACT FROM AUTHOR]

To understand if the genomic imprinting status of the donor cells is altered during the process of SCNT (somatic cell nuclear transfer), cloned pigs were produced by SCNT using PEF (porcine embryonic fibroblast) and P-PEF (parthenogenetic-PEF) cells as donors. Then, the gene expression and methylation patterns of H19 , IGF2 , NNAT and MEST were compared between PEF vs. C-PEF (cloned-PEF), P-PEF vs. CP-PEF (cloned-P-PEF), respectively. Taken together, the results revealed that there was no significant difference in the expression of imprinted genes and conserved genomic imprints between the donor and cloned cells. [ABSTRACT FROM AUTHOR]

Purpose: To determine the expression patterns of imprinted genes and their methylation status in aborted cloned porcine fetuses and placentas. Methods: RNA and DNA were prepared from fetuses and placentas that were produced by SCNT and controls from artificial insemination. The expression of 18 imprinted genes was determined by quantitative real-time PCR (q-PCR). Bisulfite sequencing PCR (BSP) was conducted to determine the methylation status of PRE- 1 short interspersed repetitive element (SINE), satellite DNA and H19 differentially methylated region 3 (DMR3). Results: The weight, imprinted gene expression and genome-wide DNA methylation patterns were compared between the mid-gestation aborted and normal control samples. The results showed hypermethylation of PRE- 1 and satellite sequences, the aberrant expression of imprinted genes, and the hypomethylation of H19 DMR3 occurred in mid-gestation aborted fetuses and placentas. Conclusions: Cloned pigs generated by somatic cell nuclear transfer (SCNT) showed a greater ratio of early abortion during mid-gestation than did normal controls because of the incomplete epigenetic reprogramming of the donor cells. Altered expression of imprinted genes and the hypermethylation profile of the repetitive regions ( PRE- 1 and satellite DNA) may be associated with defective development and early abortion of cloned pigs, emphasizing the importance of epigenetics during pregnancy and implications thereof for patient-specific embryonic stem cells for human therapeutic cloning and improvement of human assisted reproduction. [ABSTRACT FROM AUTHOR]

A previous report suggested that the expression of ten-eleven translocation (TET) proteins is abnormal in certain cancers. Quercetin has been demonstrated as anti-cancer role in cancer development. In order to explore the inhibitory effect and mechanism of quercetin on uveal melanoma cells, the expression of TET proteins was analyzed in the present study. Our results suggest that the expression of TET1 was increased following treatment with quercetin in OCM-1, SK-MEL-1, and B16 cells. In addition, quercetin treatment induced apoptosis and inhibited migration and invasion. To further investigate the association of the expression of TET1 with cell growth, apoptosis, migration, and invasion, cell lines in which TET1 was knocked-down or overexpressed were constructed. The results showed that the increased expression of TET1-induced apoptosis, increased 5-hydroxymethylcytosine (5 hmC). and inhibited invasion. Our bioinformatics studies indicated that TET1 is a target gene of microRNA-17 (miR-17) Our results showed that inhibition of the expression of miR-17 resulted in increased TET1 expression in OCM-1 cells. Furthermore, our results indicated that quercetin treatment increased TET1 expression and inhibited melanoma growth in nude mice. Taken together, our results suggest that quercetin can regulate cell proliferation and apoptosis through TET1 via miR-17 in melanoma cells. [ABSTRACT FROM AUTHOR]

Aberrant expression of imprinted genes is the main reason for developmental retardation in mammalian parthenogenetic fetuses. Mesoderm specific transcript ( MEST ) is a maternally imprinted gene that is linked to cancer and is necessary for normal early embryonic development. Tissue and isoform-specific imprinting of MEST have been identified in humans and mice, but have not yet been identified in pigs. In this study, the three isoforms of porcine MEST were identified using the GenBank and Ensembl sequence databases. Then, we determined MEST isoform-specific mRNA expression and methylation levels by quantitative real-time PCR (qRT-PCR) and bisulfite sequencing PCR analysis (BSP) between porcine parthenogenetic (PA) and control (Con) fetuses, respectively. Altogether, our results demonstrated that the expression of MEST-1A and MEST-1B has no evident differences between PA and Con groups; however, there is no expression of MEST-1C in PA fetuses. In addition, the results of BSP showed that the hypermethylation of exon 1c of MEST-1C was observed in PA samples. Thus, we suggested that MEST-1C was isoform-specific imprinting, which may be contributed to differential methylation status of exon 1c in porcine fetuses. [ABSTRACT FROM AUTHOR]

Background: Galangin has been extensively studied as the antitumor agent in various cancers. However, the effect of galangin in hepatocellular carcinoma (HCC) remains elusive. Methods: Using RNA sequencing, the differential expression of lncRNA in human HCC cell line with highly metastatic potential (MHCC97H) cells treated with galangin was investigated. Furthermore, H19 expression pattern was also determined in MHCC97H cells following treatment with galangin. In addition, knockdown and overexpression of H19 was performed to analyze the effect of the expression pattern of H19 on cell apoptosis, cell cycle, migration, and invasion in HCC cells. Moreover, the in vivo effect of galangin on tumor development was also determined in nude mice. In order to analyze loss expression of H19 in vivo, clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/Cas9) was used. Results: Total of 50 lncRNAs were significantly differentially expressed in MHCC97H cells treated with galangin. Besides, the expression of H19 was markedly reduced following treatment with galangin in MHCC97H cells. Compared to the Control group, the galangin‐treated group inhibited cell migration and invasion. Knockdown of H19 expression showed increased cell apoptosis and decreased invasion. In addition, RNA‐seq data also identified 161 mRNA which was significantly differentially expressed following treatment with galangin. To further determine the underlying mechanism, p53 protein was analyzed. Notably, the results indicated that knockdown of H19 and miR675 induced the expression of p53, eventually promoting cell apoptosis in MHCC97H cells. These results indicated that galangin promoted cell apoptosis through reduced the expression of H19 and miR675 in MHCC97H cells. The in vivo result showed that compared to the Con, tumor growth was remarkably suppressed with loss expression of H19. Conclusion: Our data suggested that galangin has a crucial role in hepatocarcinogenesis through regulating the expression pattern of H19. [ABSTRACT FROM AUTHOR]

Aberrant epigenetic modification, including N6-methylation of adenosine (m6A), has been frequently reported in embryos derived from parthenogenetic activation (PA). However, the role of Igf2bp1 expression pattern in m6A modification and the mechanism through which Igf2bp1 function is regulated in PA embryos remains elusive. Therefore, in this study, using si-Igf2bp1 and betaine (N,N,N-trimethylglycine, a major methyl donor), we investigated the effect of Igf2bp1 expression in m6A modification on the development of PA embryos. The results indicated that the down-regulation of Igf2bp1 reduced the cleavage and blastula rates of PA embryos. Moreover, m6A expression level was markedly down-regulated following microinjection with si-Igf2bp1. However, the treatment with betaine could significantly restore the m6A level. Further bioinformatics analysis revealed Igf2bp1 as the putative target of microRNA 670 (miR-670). Thus, to confirm this finding, mimics and inhibitor of miR-670 were microinjected into PA embryos. The results demonstrated that miR-670 inhibitor augmented the expression of Igf2bp1 and rescued cleavage and blastula rates. In addition, the miR-670 inhibitor promoted the m6A expression level. TUNEL assay revealed a loss of expression of Igf2bp1 induced cell apoptosis in PA embryos. Taken together, these results demonstrated that miR-670-3p functions as the regulator of Igf2bp1 expression and plays a crucial role in PA development through m6A modification. [ABSTRACT FROM AUTHOR]

Abstract Ten-eleven translocation (TET) proteins have been shown to be abnormally expressed in different cancers. To investigate the expression pattern of TET proteins in HepG2 cells, sodium ascorbate was used to treat HepG2 cells. Our results showed that TET1 , TET2 and TET3 expression was increased after sodium ascorbate treatment. The TET proteins catalyze the oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), thus, 5mC and 5hmC levels were examined. The results suggested that 5hmC was increased after sodium ascorbate treatment. To further determine the biological function of the TET proteins, si-TET1, si-TET2 and si-TET3 were transfected into HepG2 cells. The results showed that a knock down of TET3 expression stimulated cell proliferation of HepG2 cells. To further understand the effects of TET3 expression on cell proliferation, sodium ascorbate was added to the cells after transfection with si-TET3. The results demonstrated that sodium ascorbate could rescue TET3 expression and inhibit cell proliferation. Taken together, these results indicate that TET3 expression regulated cell proliferation, which is associated with 5hmC in HepG2 cells. Highlights • Sodium ascorbate stimulated TET proteins expression. • Sodium ascorbate inhibited cell proliferation • TET3 regulated cell proliferation through 5hmC. [ABSTRACT FROM AUTHOR]

It has been suggested that the imprinted gene, H19, plays a crucial role in the development of cancer. In the present study, we attempted to treat the abnormal expression and methylation status of H19 in A549 cells using valproic acid (VPA), ascorbic acid (Vc), and 5-aza-Cytidine (5-Aza). The results suggested that VPA administration could alter the expression pattern of H19, while the hypomethylation status of H19 DMR was unchanged. Furthermore, overexpression of HDAC1 and DNMT1 was associated with decreased expression of H19 in VPA-treated cells. Western blot results showed that the expression of p53 protein was increased following treatment with VPA. In addition, we also investigated cellular apoptosis and the cell cycle of treated cells. Flow cytometry data indicated that VPA could increase the occurrence of cell apoptosis in A549 cells. Taken together, our results suggest that H19 expression was suppressed by VPA through HDAC1 and DNMT1 and decreased H19 expression correlated with cell apoptosis in A549 cells. [ABSTRACT FROM AUTHOR]

Abstract: Shadoo is a glycoprotein expressed in the adult brain that is an interacting protein of prion protein; however, its function remains to be determined. To elucidate its role in prion pathogenesis, we generated transgenic mice overexpressing wild-type (wt) Shadoo driven by the murine PrP promoter. Expression of the murine Sprn transgene significantly increased brain Shadoo protein levels in all three mouse lines generated. Following infection with mouse-adapted scrapie strain 22L, all transgenic mice tested exhibited characteristics of scrapie disease. Importantly, there was no correlation between the expression level or incubation time of Shadoo with disease phenotype. We therefore conclude that Shadoo has little or no influence on the outcome of transmissible spongiform encephalopathy (TSE) disease in transgenic mice. [Copyright &y& Elsevier]

Background: Long noncoding RNAs (lncRNAs) can act as microRNA (miRNA) sponges to regulate protein-coding gene expression; therefore, lncRNAs are considered a major part of the competitive endogenous RNA (ceRNA) network and have attracted growing attention. The present study explored the regulatory mechanisms and functional roles of lncRNAs as ceRNAs in hepatocellular carcinoma (HCC) and their potential impact on HCC patient prognosis. Methods: In this study, we systematically studied the expression profiles and prognostic value of lncRNA, miRNA, and mRNA from a total of 838 HCC patients from five HCC cohorts (TCGA, GSE54236, GSE76427, GSE64041 and GSE14520). The TCGA, GSE54236 and GSE76427 HCC cohorts were utilized to establish a prognosis-related network of dysregulated ceRNAs by bioinformatics methods. The GSE64041 and GSE14520 HCC cohorts were utilized to verify the expression of candidate genes. Results: In total, 721 lncRNAs, 73 miRNAs, and 1563 mRNAs were aberrantly expressed in HCC samples. A ceRNA network including 26 lncRNAs, four miRNAs, and six mRNAs specific to HCC was established. The survival analysis showed that four lncRNAs (MYCNOS, DLX6-AS1, LINC00221, and CRNDE) and two mRNAs (CCNB1 and SHCBP1) were prognostic biomarkers for patients with HCC in both the TCGA and GEO databases. Conclusion: The proposed ceRNA network may help elucidate the regulatory mechanism by which lncRNAs function as ceRNAs and contribute to the pathogenesis of HCC. Importantly, the candidate lncRNAs, miRNAs, and mRNAs involved in the ceRNA network can be further evaluated as potential therapeutic targets and prognostic biomarkers for HCC. [ABSTRACT FROM AUTHOR]