scholarly journals Analysis of Selenoprotein Expression in Response to Dietary Selenium Deficiency During Pregnancy Indicates Tissue Specific Differential Expression in Mothers and Sex Specific Changes in the Fetus and Offspring

2020 ◽  
Vol 21 (6) ◽  
pp. 2210 ◽  
Author(s):  
Pierre Hofstee ◽  
James S.M. Cuffe ◽  
Anthony V. Perkins
Keyword(s):  
Gene Expression ◽  
Rna Extraction ◽  
Selenium Deficiency ◽  
Sexually Dimorphic ◽  
Foetal Development ◽  
Dietary Selenium ◽  
Pregnant Mice ◽  
Low Selenium ◽  
The Impact ◽  
Age And Sex

The human selenoproteome is comprised of ~25 genes, which incorporate selenium, in the form of selenocysteine, into their structure. Since it is well known that selenium is important to maternal health and foetal development during pregnancy, this study aimed at defining the impact of selenium deficiency on maternal, placental, foetal and offspring selenoprotein gene expression. Female C57BL/6 mice were randomly allocated to control (>190 μg/kg) or low selenium (<50 μg/kg) diets four weeks prior to mating and throughout gestation. At embryonic day (E)18.5, pregnant mice were sacrificed followed by collection of maternal and foetal tissues. A subset of mice littered down, and offspring were monitored from postnatal day (PN) 8, weaned at PN24 and sacrificed at PN180, followed by tissue collection. Following RNA extraction, the expression of 14 selenoproteins was assessed with qPCR in liver, kidneys, muscle and placenta. Selenium deficiency downregulated expression (Ptrt < 0.05) of many selenoproteins in maternal tissues and the placenta. However, foetal selenoprotein expression was upregulated (Ptrt < 0.05) in all tissues, especially the kidneys. This was not reflected at PN180; however, a sexually dimorphic relationship in selenoprotein expression was observed in offspring. This study demonstrates the selenoproteome is sensitive to dietary selenium levels, which may be exacerbated by pregnancy. We concluded that transcriptional regulation of selenoproteins is complex and multifaceted, with expression exhibiting tissue-, age- and sex-specificities.

scholarly journals The impact of RNA extraction method on accurate RNA sequencing from formalin-fixed paraffin-embedded tissues

BMC Cancer ◽  
2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Michal Marczyk ◽  
Chunxiao Fu ◽  
Rosanna Lau ◽  
Lili Du ◽  
Alexander J. Trevarton ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Gene Expression ◽  
Rna Sequencing ◽  
Rna Extraction ◽  
Formalin Fixed Paraffin ◽  
Ffpe Samples ◽  
Formalin Fixed Paraffin Embedded ◽  
The Impact ◽  
Formalin Fixed

Abstract Background Utilization of RNA sequencing methods to measure gene expression from archival formalin-fixed paraffin-embedded (FFPE) tumor samples in translational research and clinical trials requires reliable interpretation of the impact of pre-analytical variables on the data obtained, particularly the methods used to preserve samples and to purify RNA. Methods Matched tissue samples from 12 breast cancers were fresh frozen (FF) and preserved in RNAlater or fixed in formalin and processed as FFPE tissue. Total RNA was extracted and purified from FF samples using the Qiagen RNeasy kit, and in duplicate from FFPE tissue sections using three different kits (Norgen, Qiagen and Roche). All RNA samples underwent whole transcriptome RNA sequencing (wtRNAseq) and targeted RNA sequencing for 31 transcripts included in a signature of sensitivity to endocrine therapy. We assessed the effect of RNA extraction kit on the reliability of gene expression levels using linear mixed-effects model analysis, concordance correlation coefficient (CCC) and differential analysis. All protein-coding genes in the wtRNAseq and three gene expression signatures for breast cancer were assessed for concordance. Results Despite variable quality of the RNA extracted from FFPE samples by different kits, all had similar concordance of overall gene expression from wtRNAseq between matched FF and FFPE samples (median CCC 0.63–0.66) and between technical replicates (median expression difference 0.13–0.22). More than half of genes were differentially expressed between FF and FFPE, but with low fold change (median |LFC| 0.31–0.34). Two out of three breast cancer signatures studied were highly robust in all samples using any kit, whereas the third signature was similarly discordant irrespective of the kit used. The targeted RNAseq assay was concordant between FFPE and FF samples using any of the kits (CCC 0.91–0.96). Conclusions The selection of kit to purify RNA from FFPE did not influence the overall quality of results from wtRNAseq, thus variable reproducibility of gene signatures probably relates to the reliability of individual gene selected and possibly to the algorithm. Targeted RNAseq showed promising performance for clinical deployment of quantitative assays in breast cancer from FFPE samples, although numerical scores were not identical to those from wtRNAseq and would require calibration.

scholarly journals Selenoprotein Gene Expression in Thyroid and Pituitary of Young Pigs Is Not Affected by Dietary Selenium Deficiency or Excess

2009 ◽  
Vol 139 (6) ◽  
pp. 1061-1066 ◽  
Author(s):  
Ji-Chang Zhou ◽  
Hua Zhao ◽  
Jun-Gang Li ◽  
Xin-Jie Xia ◽  
Kang-Ning Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Selenium Deficiency ◽  
Dietary Selenium ◽  
Selenoprotein Gene ◽  
Young Pigs

scholarly journals Impact of RNA degradation on measurements of gene expression

2014 ◽  
Author(s):  
Irene Gallego Romero ◽  
Athma A. Pai ◽  
Jenny Tung ◽  
Yoav Gilad
Keyword(s):  
Gene Expression ◽  
Expression Profiling ◽  
Rna Extraction ◽  
Rna Degradation ◽  
Significant Loss ◽  
Whole Genome ◽  
Expression Levels ◽  
Rna Quality ◽  
The Impact ◽  
Gene Expression Levels

The use of low quality RNA samples in whole-genome gene expression profiling remains controversial. It is unclear if transcript degradation in low quality RNA samples occurs uniformly, in which case the effects of degradation can be normalized, or whether different transcripts are degraded at different rates, potentially biasing measurements of expression levels. This concern has rendered the use of low quality RNA samples in whole-genome expression profiling problematic. Yet, low quality samples are at times the sole means of addressing specific questions – e.g., samples collected in the course of fieldwork. We sought to quantify the impact of variation in RNA quality on estimates of gene expression levels based on RNA-seq data. To do so, we collected expression data from tissue samples that were allowed to decay for varying amounts of time prior to RNA extraction. The RNA samples we collected spanned the entire range of RNA Integrity Number (RIN) values (a quality metric commonly used to assess RNA quality). We observed widespread effects of RNA quality on measurements of gene expression levels, as well as a slight but significant loss of library complexity in more degraded samples. While standard normalizations failed to account for the effects of degradation, we found that a simple linear model that controls for the effects of RIN can correct for the majority of these effects. We conclude that in instances where RIN and the effect of interest are not associated, this approach can help recover biologically meaningful signals in data from degraded RNA samples.

40 Gene Expression Profiling of In Vitro-Produced Blastocysts Derived from In Vitro-Matured Bovine Oocytes Vitrified/Warmed in Media Supplemented with a Biopolymer Produced by an Antarctic Bacterium

2018 ◽  
Vol 30 (1) ◽  
pp. 159 ◽  
Author(s):  
N. Arcarons ◽  
M. Vendrell ◽  
M. Yeste ◽  
M. E. Mercadé ◽  
M. López-Béjar ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Patterns ◽  
Rna Extraction ◽  
Mrna Abundance ◽  
The Other ◽  
Developmental Competence ◽  
Crystal Formation ◽  
Control Analysis ◽  
The Impact

Previous reports have demonstrated the beneficial effect of antifreeze glycoprotein supplementation during oocyte vitrification on preventing ice crystal formation and thus enhancing developmental competence after vitrification-warming. Pseudomonas sp. ID1, a bacterium isolated from marine sediment from Antarctica, produces an exopolysaccharide, M1 EPS, as a cold adaptation mechanism. Despite numerous studies on structural and morphological damages induced by cryopreservation in oocytes, few studies have focused on the impact of vitrification on the expression pattern of genes during early embryo development. In the present study, the expression patterns of 6 genes (BAX, BCL2-like 1, DNMT3A, UBE2A, SCLC2A3, and HDAC1) were investigated in Day 8 blastocysts resulting from in vitro-matured oocytes vitrified/warmed in media supplemented with various concentrations of M1 EPS. After 21 h of IVM, 1,062 oocytes were vitrified/warmed in media supplemented with 0, 0.001, 0.01, 0.1, and 1 mg mL−1 M1 EPS. At 24 h of IVM, oocytes were in vitro fertilized and in vitro cultured and the resulting blastocysts were harvested at Day 8 for RNA extraction and qPCR analysis. Fresh, non-vitrified oocytes were used as a control. Analysis of gene expression was performed through Kruskall-Wallis test and followed by Mann-Whitney test, and the level of significance was set at P ≤ 0.05. No significant differences were detected in relative mRNA abundance for SLC2A3, UBE2A, or HDAC-1 between blastocysts derived from vitrified oocytes, regardless of M1 EPS treatment. Expression of DNMT3A was significantly higher in embryos obtained from oocytes vitrified and warmed with 0.1 mg mL−1 M1 EPS compared with other treatment groups. However, no differences in DNMT3A expression were observed when the other vitrified groups were compared. The relative abundance of BAX transcript in embryos from oocytes vitrified in media supplemented with 0.1 mg mL−1 M1 EPS was higher than that in 0 or 0.001 mg mL−1 groups. Embryos from 0.01 and 0.1 mg mL−1 groups showed higher BCL2-like 1 mRNA abundance than those from the 0, 0.001, and 1 mg mL−1 groups. Whereas blastocysts from oocytes vitrified with 0.01 mg mL−1 M1 EPS exhibited the lowest BAX:BCL2-like 1 ratio, no significant differences in BAX:BCL2-like 1 ratio were observed between the other treatments. The significantly lower BAX:BCL2 ratio observed in blastocysts obtained from oocytes vitrified with 0.01 mg mL−1 M1 EPS could be indicative for a better embryo quality. Although optimizing the use of M1 EPS may benefit oocyte cryopreservation protocols, further research is required to clarify the exact mechanism through which it exerts its protective role. This study was supported by the Spanish Ministry of Science and Innovation (Project AGL2016-79802-P and grant CTQ2014-59632-R).

High throughput, highly multiplexed transcriptional profiling of cytologic preparations from buccal mucosa

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 14104-14104
Author(s):  
R. B. Everson ◽  
D. Bangsi ◽  
L. L. Darga ◽  
M. R. Bell ◽  
V. P. Pidlaoan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cell ◽  
Smoking Status ◽  
Transcriptional Profiling ◽  
Rna Extraction ◽  
Rna Isolation ◽  
Buccal Cells ◽  
Cross Sectional ◽  
Molecular Therapeutics ◽  
The Impact

14104 Background: Alterations in gene expression can be sensitive and informative indicators that a biologically effective exposure occurred. For example, serial specimens could provide evidence that a dose with a biological effect was reached in Phase I testing. Sampling buccal mucosal can provide epithelial cell specimens rapidly with little discomfort. However, ribonucleases in saliva rapidly degrade epithelial cell RNA, prohibiting analysis by standard array techniques. Using buccal cells, we conducted a pilot study to optimize procedures for RNA isolation and analysis and determined influences of gender, race, and cigarette smoking on gene expression. Methods: Buccal cells were collected by scraping the inner cheek with a previously described standardized plastic tool (BioTechniques. 2004;36:484–487) made available Avrum Spira of Boston University. Preliminary experiments compared RNA extraction procedures, including methods based on trizol (Invitrogen, Carlsbad, CA), RNeasy columns (Qiagen, Valencia, CA), and the High Pure RNA Paraffin Kit (Roche, Indiana, USA). After standardization of the method, specimens were obtained from 64 subjects using a blocked design sampling equal numbers of subjects by gender, African American and white race, and smoking status. Gene expression levels were determined using the cDNA-mediated annealing, selection, extension, and ligation assay (Illumina, Inc.), which measures expression of over 500 genes per analysis. Results: The Roche extraction method provided the highest yield of RNA and was used for subsequent assays. Technical replicates were highly reproducible. Preliminary analyses revealed that using P=0.05, 38 genes were expressed differentially by gender, 20 by race, and 10 by smoking status. The genes most differentially expressed by gender included IRF1, MET, STAT1, RAP1GDS1; race CD9, CCNA2, CEACAM1, FVT1; smoking CD44, NQO1, SKI, SRC. Conclusions: Highly multiplexed gene expression analysis of buccal cells are feasible. Demographic characteristics of study subjects can be important, but they do not heavily influence levels for many genes. Results indicate the assays could be provide useful information in cross-sectional or serial studies of the impact of molecular therapeutics. No significant financial relationships to disclose.

scholarly journals Sex-specific responses to cold in a very cold-tolerant, northern Drosophila species

Heredity ◽  
2021 ◽  
Author(s):  
Darren J. Parker ◽  
Tapio Envall ◽  
Michael G. Ritchie ◽  
Maaria Kankare
Keyword(s):  
Gene Expression ◽  
Cold Tolerance ◽  
Cold Treatment ◽  
Sexually Dimorphic ◽  
Immune Genes ◽  
Cold Tolerant ◽  
Males And Females ◽  
The Impact ◽  
Drosophila Montana ◽  
Harsh Conditions

AbstractOrganisms can plastically alter resource allocation in response to changing environmental factors. For example, in harsh conditions, organisms are expected to shift investment from reproduction toward survival; however, the factors and mechanisms that govern the magnitude of such shifts are relatively poorly studied. Here we compared the impact of cold on males and females of the highly cold-tolerant species Drosophila montana at the phenotypic and transcriptomic levels. Although both sexes showed similar changes in cold tolerance and gene expression in response to cold treatment, indicating that the majority of changes are concordant between the sexes, we identified a clear reduction in sexually dimorphic gene expression, suggesting that preparing for the colder season involves reducing investment in sex-specific traits. This reduction was larger in males than females, as expected if male sexual traits are more condition-dependent than female traits, as predicted by theory. Gene expression changes were primarily associated with shifts in metabolic profile, which likely play a role in increasing cold tolerance. Finally, we found that the expression of immune genes was reduced following cold treatment, suggesting that reduced investment in costly immune function may be important in helping flies survive colder periods.

scholarly journals The effects of aneuploidy on gene expression in a dosage series of maize chromosome arm 1L

2017 ◽  
Author(s):  
◽  
Adam Johnson
Keyword(s):  
Gene Expression ◽  
Chromosome Region ◽  
Rna Extraction ◽  
Leaf Tissue ◽  
Dosage Level ◽  
Expression Of Genes ◽  
Chromosome Arm ◽  
Cascade Effects ◽  
The Impact

Aneuploidy is a class of genetic conditions involving an unbalanced number of chromosomes. The most familiar human aneuploid condition is trisomy 21, called Down syndrome. Aneuploid conditions necessarily involve a change in the dosage of those genes which are located on the varied chromosome. However, the dosage level of a gene does not automatically correspond to the amount of RNA or protein that will be produced in vivo. Based on previously published studies, the impact of chromosome dosage changes on the transcription of single genes may be direct, inverse, or anywhere in between; and genes may be impacted anywhere in the genome, not just on the varied chromosome. Using a maize model system, a dosage series of plants was produced in which sibling plants are identical, except for the copy number of chromosome arm 1L. These plants were grown until 45 days postgermination, at which point leaf tissue was collected for RNA extraction. This dosage series included 5 dosage levels for comparison: diploid, trisomic, tetrasomic, haploid, and disomic haploid. A second dosage series was grown up to day 55, and included diploid, monosomic, and trisomic. Using RNA sequencing, expression levels for all genes were determined. The results were analyzed in aggregate, allowing for a view of effects on the level of the whole transcriptome. Results suggest that dosage of genes on the varied chromosome region has some correlation with expression of those genes, though the change compared to a diploid is often partial. Inverse relationships between chromosome dosage and RNA expression of genes elsewhere in the genome are seen to occur. Both direct and inverse reactions were amplified by increased levels of genomic imbalance. The kinetics of interacting proteins and other cellular components, as described in the gene balance hypothesis, may be the mechanism leading to these responses. Using the same methods of analysis, similar phenomena were observed in aneuploid/euploid comparisons in other organisms. Partial dosage compensation and inverse effects were observed in published datasets from aneuploid yeast and mouse. A set of trisomics in Arabidopsis displayed the same effects, though to a different extent in different trisomies. Using a published database of transcription factors, the responses of genes to dosage changes of their regulators was analyzed. A number of cascade effects were observed, in which inverse relationships of transcription factor dosage and target gene expression occurred sequentially, disrupting normal regulation of several genes in a network by changing the dosage of a single component.

scholarly journals Sex-specific responses to cold in a very cold-tolerant, northern Drosophila species

2020 ◽  
Author(s):  
Darren J. Parker ◽  
Tapio Envall ◽  
Michael G. Ritchie ◽  
Maaria Kankare
Keyword(s):  
Gene Expression ◽  
Cold Tolerance ◽  
Cold Treatment ◽  
Sexually Dimorphic ◽  
Immune Genes ◽  
Cold Tolerant ◽  
Males And Females ◽  
The Impact ◽  
Drosophila Montana ◽  
Harsh Conditions

AbstractOrganisms can plastically alter resource allocation in response to changing environmental factors. For example, in harsh conditions organisms are expected to shift investment from reproduction towards survival, however, the factors and mechanisms that govern the magnitude of such shifts are relatively poorly studied. Here we compared the impact of cold on males and females of the highly cold-tolerant species Drosophila montana at the phenotypic and transcriptomic levels. Although both sexes showed similar changes in cold tolerance and gene expression in response to cold treatment, indicating that the majority of changes are concordant between the sexes, we identified a clear reduction in sexually dimorphic gene expression, suggesting that preparing for colder season also involves reducing investment in sex-specific traits. This reduction was larger in males than females, as expected if male sexual traits are more condition-dependent than female traits, as predicted by theory. Gene expression changes were primarily associated with shifts in metabolic profile which likely play a role in increasing cold tolerance. Finally, we found that the expression of immune genes was reduced following cold treatment, suggesting that reduced investment in immunity may be important in helping flies survive colder periods.

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