Underexpression of 4 Placenta-Associated MicroRNAs in Complete Hydatidiform Moles

2012 ◽  
Vol 22 (6) ◽  
pp. 1075-1080 ◽  
Author(s):  
Quan Na ◽  
Dan Wang ◽  
Weiwei Song
Keyword(s):  
In Situ Hybridization ◽  
Hydatidiform Mole ◽  
Polymerase Chain Reaction Analysis ◽  
Normal Pregnancy ◽  
Differentially Expressed ◽  
Control Group ◽  
Differentially Expressed Mirnas ◽  
Hydatidiform Moles

ObjectiveSeveral placental microRNAs (miRNAs) have been identified as placenta-associated miRNAs with the potential of estimating the condition of the placenta. However, our understanding of these miRNAs is limited. The aim of this study was to determine the expression of 8 placenta-associated miRNAs (miR-512-3p, miR-517a, miR-517b, miR-518b, miR-519a, miR-1185, miR-1283, and miR-1323) in complete hydatidiform mole (CHM).MethodsSamples were obtained from patients with CHM (CHM group, n = 12) and elective terminations of normal pregnancy (control group, n = 20). We detected differentially expressed placenta-associated miRNAs in placenta by quantitative real-time reverse transcriptase–polymerase chain reaction analysis. Subsequently, we assessed the expression location of differentially expressed miRNAs by in situ hybridization analysis.ResultsFour placenta-associated miRNAs (miR-517a, miR-517b, miR-518b, and miR-519a) were underexpressed in the CHM group, compared with the control group (P < 0.01). When further investigating these 4 miRNAs with regard to in vivo localization by in situ hybridization, we found that 2 miRNAs (miR-517b and miR-518b) were detected exclusively in the trophoblast layer, with little signal (if any) observed in villous stroma cells.ConclusionsThe results show that 4 miRNAs (miR-517a, miR-517b, miR-518b, and miR-519a) are deregulated in CHM, which suggests the involvement of these miRNAs in the functions of CHM placenta.

scholarly journals MicroRNA expression profiling involved in Borax-induced anti-tumor effect using gene-chip analysis

2020 ◽  
Author(s):  
Lun Wu ◽  
Ying Wei ◽  
Wen-Bo Zhou ◽  
Jiao Zhou ◽  
Li-Hua Yang ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Hepg2 Cells ◽  
Enrichment Analysis ◽  
Gene Chip ◽  
Differentially Expressed ◽  
Control Group ◽  
Pathway Enrichment Analysis ◽  
Ras Signaling ◽  
Therapeutic Benefits ◽  
Differentially Expressed Mirnas

Abstract Background Borax, a boron compound, which is becoming widely recognized for its biological effects, including antioxidant activity, cytotoxicity, and potential therapeutic benefits. However, the specific molecular mechanisms underlying borax-induced anti-tumor effect still remain to be to further elucidated. MicroRNAs (miRNAs) may play key roles in cellular processes including tumor progression, cell apoptosis and cytotoxicity. Thus, this study aimed to investigate, whether miRNAs were involved in the borax-mediated anti-tumor effect using miRNA profiling of a human liver cancer cell line (HepG2) using gene-chip analysis.Methods Total RNA was extracted and purified from HepG2 cells that were treated with 4 mM borax for either 2 or 24 h. The samples underwent microarray analysis using an Agilent Human miRNA Array. Differentially expressed miRNAs were analysed by volcano plot and heatmap, and were validated using real-time fluorescent quantitative PCR (qPCR).ResultsAmong this, 2- or 24-h exposure to borax significantly altered the expression level of miRNAs in HepG2 cells, 4 or 14 were upregulated and 3 were downregulated compared with the control group, respectively (≥2-fold; P<0.05). GO enrichment analysis and KEGG pathway enrichment analysis revealed that target genes of differentially expressed miRNAs in HepG2 cells predominantly participated in MAPK signaling pathway, TGF-beta signaling pathway, NF-kappa B signaling pathway, etc; in 2-h borax treatment group, while Ras signaling pathway, FoxO signaling pathway, Cellular senescence, etc; involved in 24-h treatment group.Conclusions Result indicates that borax-induced anti-tumor effect may be associated with alterations in miRNAs.

scholarly journals circHMGCS1–016 reshapes immune environment by sponging miR-1236-3p to regulate CD73 and GAL-8 expression in intrahepatic cholangiocarcinoma

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Ya-Ping Xu ◽  
Ze-Ning Dong ◽  
Si-Wei Wang ◽  
Yi-Min Zheng ◽  
Chi Zhang ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Intrahepatic Cholangiocarcinoma ◽  
Molecular Mechanisms ◽  
Prognostic Significance ◽  
Luciferase Reporter ◽  
Mouse Tumor ◽  
Rna Seq ◽  
Potential Biomarker

Abstract Background Accumulating evidence indicates that circRNAs may serve as essential regulators in the progression of several human cancers, but the function and mechanism of circRNAs in intrahepatic cholangiocarcinoma (ICC) are largely unknown. Methods RNA-seq was used to assess differentially expressed circRNAs between 4 ICC and peritumor tissues. Quantitative RT-PCR and in situ hybridization were used to determine the circHMGCS1–016 expression in ICC tissues. The function and mechanism of circHMGCS1–016 were further identified via in vivo experiments. The clinical characteristics and prognostic significance of circHMGCS1–016 were analyzed by a retrospective study. The functions of circHMGCS1–016 were assessed via modifying circRNA expression in ICC cells. Moreover, the molecular mechanisms of circHMGCS1–016 in ICC cells were explored by circRNA precipitation, miRNA immunoprecipitation, SILAC and luciferase reporter assays. Results We identified that compared with peritumor tissues, ICC tissues expressed hsa_circ_0008621 (circHMGCS1–016) high by RNA-seq, which was further identified by qRT-PCR and in situ hybridization. Moreover, the expression of circHMGCS1–016 was revealed to be associated with survival and recurrence of ICC patients. By regulating circHMGCS1–016 expression, we found that elevated circHMGCS1–016 promoted ICC development both in vitro and in vivo. By SILAC and circRNA-pull down, we demonstrated that circHMGCS1–016 induced ICC cell invasion and reshaped the tumor immune microenvironment via the miR-1236-3p/CD73 and GAL-8 axis. In ICC tissues, we uncovered that a high level of circHMGCS1–016 was positively associated with CD73 and GAL-8 expression and negatively related to the CD8+ T cells infiltration, which was further validated by establishing a humanized mouse tumor model. Importantly, we displayed that ICC patients with high levels of circHMGCS1–016 in tumor tissues benefited less from anti-PD1 treatment compared to those with low levels of circHMGCS1–016. Conclusions CircHMGCS1–016 is a forceful contributor in ICC development and immune tolerance via miR-1236-3p/CD73 and GAL-8 axis. CircHMGCS1–016 can be explored as a new potential biomarker and therapeutic target for PD1-resistant ICC.

Use of cross-species in-situ hybridization (ZOO-FISH) to assess chromosome abnormalities in day-6 in-vivo- or in-vitro-produced sheep embryos

2007 ◽  
Vol 15 (3) ◽  
pp. 399-408 ◽  
Author(s):  
Gianfranco Coppola ◽  
Basil Alexander ◽  
Dino Di Berardino ◽  
Elizabeth St John ◽  
Parvathi K. Basrur ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Chromosome Abnormalities

Interactions of human cytomegalovirus with leukocytes in vivo: analysis by in situ hybridization

1987 ◽  
Vol 3 (4) ◽  
pp. 287-297 ◽  
Author(s):  
Lloyd W. Turtinen ◽  
Robin Saltzman ◽  
M.Colin Jordan ◽  
Ashley T. Haase
Keyword(s):  
In Situ Hybridization ◽  
Human Cytomegalovirus ◽  
In Vivo Analysis

Cell-specific metallothionein gene expression in mouse decidua and placentae

Development ◽  
1989 ◽  
Vol 107 (3) ◽  
pp. 611-621 ◽  
Author(s):  
S.K. De ◽  
M.T. McMaster ◽  
S.K. Dey ◽  
G.K. Andrews
Keyword(s):  
Gene Expression ◽  
In Situ Hybridization ◽  
Embryo Implantation ◽  
Normal Pregnancy ◽  
Mrna Levels ◽  
Metallothionein Gene ◽  
Visceral Yolk Sac ◽  
Rna In Situ Hybridization ◽  
Low Levels

Oligodeoxyribonucleotide excess solution hybridization, Northern blot and in situ hybridization were used to analyze metallothionein gene expression in mouse decidua and placentae during gestation. Metallothionein (MT) -I and -II mRNA levels were constitutively elevated, 11- and 13-fold, respectively, relative to the adult liver, in the deciduum (D8), and decreased coordinately about 6-fold during the period of development when the deciduum is replaced by the developing placenta (D10-16). Coincident with this decline, levels of MT mRNA increased dramatically in the visceral yolk sac endoderm. In situ hybridization established that MT-I mRNA was present at low levels in the uterine luminal epithelium (D4), but was elevated at the site of embryo implantation exclusively in the primary decidual zone by D5, and then in the secondary decidual zone (D6-8). Although low levels of MT mRNA were detected in total placental RNA, in situ hybridization revealed constitutively high levels in the outer placental spongiotrophoblasts. Analysis of pulse-labeled proteins from decidua and placentae established that these tissues are active in the synthesis of MT. The constitutively high levels of MT mRNA in decidua were only slightly elevated following injection of cadmium (Cd) and/or zinc (Zn), whereas in placentae they increased several-fold. MT mRNA levels were equally high in decidua and experimentally induced deciduomata (D8) which establishes that decidual MT gene expression is not dependent on the presence of the embryo or some embryo-derived factor. Although the functional role of MT during development is speculative, these results establish the concept that, from the time of implantation to late in gestation, the mouse embryo is surrounded by cells, interposed between the maternal and embryonic environments, which actively express the MT genes. This suggests that MT plays an important role in the establishment and maintenance of normal pregnancy.

P-117 Pre-selected for an award: Bioinformatic analysis of NRF2 in the study of association of NRF2 variant and male infertility related to smoking status

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
D Aydos ◽  
O S Aydos ◽  
Y Yukselten ◽  
A Sunguroglu ◽  
K Aydos
Keyword(s):  
Dna Damage ◽  
Male Infertility ◽  
Mirna Expression ◽  
Smoking Status ◽  
Differentially Expressed ◽  
Control Group ◽  
Functional Variants ◽  
Significant Difference ◽  
Differentially Expressed Mirnas

Abstract Study question Could Nrf2 polymorphism (-617C&gt;A; rs6721961) and oxidative stress (OS)-induced changes of signature seminal plasma (SP) miRNAs related to Nrf2 provide possible biomarkers of male infertility? Summary answer -617C&gt;A SNP is associated with infertility through sperm OS DNA damage and miR-582-5p and miR-20a-5p, differentially represented between spermatozoa of smokers-non-smokers, might regulate Nrf2/ARE axis. What is known already As an extrinsic factor causing OS, smoking decreases male infertility by causing sperm membrane damage and DNA fragmentation. Expression of proteins related to the antioxidant defense system and phase 2 detoxifying enzymes controlled mainly by Nrf2/ARE pathway components is vital in managing OS-induced DNA damage. miRNAs, which multiple of are produced abundantly in male germ cells throughout spermatogenesis, have been detected in SP and contribute to multiple biological processes related to male reproductive events. miRNA-expression alterations may be induced in response to OS and without involving DNA sequence changes, miRNAs can provide additional mechanism of regulating the Nrf2 gene expression. Study design, size, duration Wild-type (WT) and SNP (-617) alleles in the Nrf2 gene were studied in 100 infertile cases and 100 controls and their associations with seminal parameters in relation to smoking status were assessed. In infertile cases, sperm DNA damage level was determined and compared among Nrf2 genotypes. Interactions between differentially expressed miRNAs (DEMIs) in response to smoking and Nrf2/ARE pathway components were visualized on a miRNA-mRNA regulatory network using CluePedia (v1.5.7) plugin of Cytoscape software (v3.8.2). Participants/materials, setting, methods Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was utilized to genotype the Nrf2 SNP (-617). DNA damages were analyzed by Comet assay. DEMIs were identified by a comprehensive bioinformatics analysis using the miRNA expression dataset GSE44134 downloaded from the GEO database. Predicted targets of DEMIs in smokers were identified by mirDIP portal. Known interactions between Nrf2 and its first neighbors were visualized after selecting STRING-actions, miRTarBase and miRecords validated miRNA source files from CluePedia panel. Main results and the role of chance There was significant difference for Nrf2 polymorphism between fertile and infertile males. The A allele was detected more frequently in the patient group; (P = 0.001). The frequencies of the C and A alleles of the Nrf2 were 62% and 38% in patients, and 78% and 44% in control group. The AA genotype was higher in the infertiles; 14% vs. 3% (P = 0.001). In smokers, sperm quality decreased significantly in AA genotype. The risk of DNA damage was highest with 224.58 AU in the AA genotype group, whereas it is the lowest with 164.56 AU in those carrying the CC genotype (P &lt; 0.005). 21 differentially expressed miRNAs (including 7 downregulated and 14 upregulated in smokers) were identified. Among the upregulated DEMIs, miR-582-5p, miR-20a-5p, miR-573, miR-186-5p, miR-499a-5p were found to target the Nrf2 mRNA, suggesting their usage as biomarkers capable of indicating the antioxidant ability of the male reproductive system. The interrelations between Nrf2/Nrf2 direct interactors and DEMIs revealed the regulatory role of hsa-miR-20a-5p in SQSTM1/p62-Keap1-Nrf2 axis linked to selective autophagy. hsa-miR-582-5p was found to regulate the JNK/Jun/caspase-3 pathway, previously shown to be activated in response to OS, in which JUN can activate or suppress the Nrf2 expression. Limitations, reasons for caution Small number of cases while evaluating the effect of smoking weakens our ability to generalize the results. Including other coexisting factors and larger patient groups carrying other functional variants of Nrf2 as well as confirming the results at the protein level would further strengthen the results of the study. Wider implications of the findings This study is the first to report -617C&gt;A polymorphism in the Nrf2 gene in the Turkish population and such a SNP may cause impaired fertility in men, especially in smokers, through oxidative metabolism. Considering these data may be valuable in determining risk groups. Trial registration number N/A

Expression of MMP-8 and MMP-13 Genes in the Periodontal Ligament during Tooth Movement in Rats

2003 ◽  
Vol 82 (8) ◽  
pp. 646-651 ◽  
Author(s):  
I. Takahashi ◽  
M. Nishimura ◽  
K. Onodera ◽  
J.-W. Bae ◽  
H. Mitani ◽  
...  
Keyword(s):  
Gene Expression ◽  
Periodontal Ligament ◽  
Tooth Movement ◽  
Orthodontic Tooth Movement ◽  
Polymerase Chain Reaction Analysis ◽  
Tension And Compression ◽  
Polymerase Chain

Periodontal ligament tissue is remodeled on both the tension and compression sides of moving teeth during orthodontic tooth movement. The present study was designed to clarify the hypothesis that the expression of MMP-8 and MMP-13 mRNA is promoted during the remodeling of periodontal ligament tissue in orthodontic tooth movement. We used the in situ hybridization method and semi-quantitative reverse-transcription/polymerase chain-reaction analysis to elucidate the gene expression of MMP-8 and MMP-13 mRNA. Expression of MMP-8 and MMP-13 mRNA transiently increased on both the compression and tension sides during active tooth movement in vivo. The gene expression of MMP-8 and MMP-13 was induced by tension, while compression indirectly promoted the gene expression of MMP-8 and MMP-13 through soluble factors in vitro. Thus, we concluded that the expression of MMP-8 and MMP-13 is differentially regulated by tension and compression, and plays an important role in the remodeling of the periodontal ligament.

Analysis of HIV-1 expression in vivo with in situ hybridization and the polymerase chain reaction

1992 ◽  
Vol 6 (3) ◽  
pp. 215-221 ◽  
Author(s):  
B. Delord ◽  
M. Ottmann ◽  
M.-H. Schrive ◽  
J.-M. Ragnaud ◽  
J.-M. Seigneurin ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
In Situ Hybridization ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Hiv 1

scholarly journals Nonrandom Transduction of Recombinant Adeno-Associated Virus Vectors in Mouse Hepatocytes In Vivo: Cell Cycling Does Not Influence Hepatocyte Transduction

2000 ◽  
Vol 74 (8) ◽  
pp. 3793-3803 ◽  
Author(s):  
Carol H. Miao ◽  
Hiroyuki Nakai ◽  
Arthur R. Thompson ◽  
Theresa A. Storm ◽  
Winnie Chiu ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Gene Transfer ◽  
Genetic Diseases ◽  
Critical Event ◽  
Dimer Formation ◽  
Adeno Associated Virus ◽  
Virus Vectors ◽  
Preclinical Trials

ABSTRACT Recombinant adeno-associated virus vectors (rAAV) show promise in preclinical trials for the treatment of genetic diseases including hemophilia. Liver-directed gene transfer results in a slow rise in transgene expression, reaching steady-state levels over a period of 5 weeks concomitant with the conversion of the single-stranded rAAV molecules into high-molecular-weight concatemers in about 5% of hepatocytes. Immunohistochemistry and RNA in situ hybridization show that the transgene product is made in about ∼5% of hepatocytes, suggesting that most rAAV-mediated gene expression occurs in hepatocytes containing the double-stranded concatemers. In this study, the mechanism(s) involved in stable transduction in vivo was evaluated. While only ∼5% of hepatocytes are stably transduced, in situ hybridization experiments demonstrated that the vast majority of the hepatocytes take up AAV-DNA genomes after portal vein infusion of the vector. Two different vectors were infused together or staggered by 1, 3, or 5 weeks, and two-color fluorescent in situ hybridization and molecular analyses were performed 5 weeks after the infusion of the second vector. These experiments revealed that a small but changing subpopulation of hepatocytes were permissive to stable transduction. Furthermore, in animals that received a single infusion of two vectors, about one-third of the transduced cells contained heteroconcatemers, suggesting that dimer formation was a critical event in the process of concatemer formation. To determine if the progression through the cell cycle was important for rAAV transduction, animals were continuously infused with 5′-bromo-2′-deoxyuridine (BrdU), starting at the time of administration of a rAAV vector that expressed cytoplasmic β-galactosidase. Colabeling for β-galactosidase and BrdU revealed that there was no preference for transduction of cycling cells. This was further confirmed by demonstrating no increase in rAAV transduction efficiencies in animals whose livers were induced to cycle at the time of or after vector administration. Taken together, our studies suggest that while virtually all hepatocytes take up vector, unknown cellular factors are required for stable transduction, and that dimer formation is a critical event in the transduction pathway. These studies have important implications for understanding the mechanism of integration and may be useful for improving liver gene transfer in vivo.

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