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Proof of Gene Doping in a Mouse Model with a Human Erythropoietin Gene Transferred Using an Adenoviral Vector

ABSTRACT: Gene doping model by hEPO (human erythropoietin) and adenoviral vector was established in the study. This study revealed that the gene expression profile in whole blood on the gene doping model significantly altered, which may be a new marker of hEPO-gene doping

ORGANISM(S): Mus musculus

PROVIDER: GSE178336 | GEO | 2021/08/11

REPOSITORIES: GEO

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Proof of Gene Doping in a Mouse Model with a Human Erythropoietin Gene Transferred Using an Adenoviral Vector

Project description:Proof of Gene Doping in a Mouse Model with a Human Erythropoietin Gene Transferred Using an Adenoviral Vector

| PRJNA738510 | ENA

Proof of Gene Doping in a Mouse Model with a Human Erythropoietin Gene Transferred Using an Adenoviral Vector.

Project description:Despite the World Anti-Doping Agency (WADA) ban on gene doping in the context of advancements in gene therapy, the risk of EPO gene-based doping among athletes is still present. To address this and similar risks, gene-doping tests are being developed in doping control laboratories worldwide. In this regard, the present study was performed with two objectives: to develop a robust gene-doping mouse model with the human EPO gene (hEPO) transferred using recombinant adenovirus (rAdV) as a vector and to develop a detection method to identify gene doping by using this model. The rAdV including the hEPO gene was injected intravenously to transfer the gene to the liver. After injection, the mice showed significantly increased whole-blood red blood cell counts and increased expression of hematopoietic marker genes in the spleen, indicating successful development of the gene-doping model. Next, direct and potentially indirect proof of gene doping were evaluated in whole-blood DNA and RNA by using a quantitative PCR assay and RNA sequencing. Proof of doping could be detected in DNA and RNA samples from one drop of whole blood for approximately a month; furthermore, the overall RNA expression profiles showed significant changes, allowing advanced detection of hEPO gene doping.

| S-EPMC8392868 | biostudies-literature

Integrating whole blood transcriptomic collection procedures into the current anti-doping testing system, including long-term storage and re-testing of anti-doping samples [Long-term]

Project description:Purpose: Recombinant human erythropoietin administration studies involving “omics” approaches have demonstrated a gene-expression signature that could aid detection of blood doping. However, current anti-doping testing does not involve blood collection into tubes with RNA preservative. This study investigated if whole blood in long-term storage could be used for transcriptomic analysis despite lacking RNA preservation. Methods: Whole blood samples were collected from thirteen male healthy individuals. Long-term storage: whole blood collected into Tempus™ tubes and K2EDTA tubes and subjected to long-term (i.e., −80°C) storage and RNA extracted. After storage, Tempus and K2EDTA tubes were thawed and extracted using Tempus™ Spin RNA Isolation Kit (Life Technologies, Carlsbad, CA, USA). Samples from seven subjects that presented higher RIN value (≥7) were selected for RNA_Seq analysis. Results: The experiment provided RNA quality and purity for gene expression analysis. Total of 19239 genes were mapped and the gene expression analysis showed that 658 genes were differentially expressed (which means 3.4% of mapped genes). With 269 being up-regulated and 389 down-regulated. None of the transcripts described in previous studies as biomarkers for blood doping (Durussel et al. 2016; Wang, Durussel, et al. 2017) were differently expressed. Conclusion: RNA quantity, purity and integrity was not significantly compromised from long-term storage in blood storage tubes lacking RNA stabilisation, indicating that transcriptomic/omics analysis could be conducted using anti-doping samples collected or biobanked without RNA preservation.

2021-09-03 | GSE183073 | GEO

Integrating whole blood transcriptomic collection procedures into the current anti-doping testing system, including long-term storage and re-testing of anti-doping samples

Project description:Recombinant human erythropoietin administration studies involving transcriptomic approaches have demonstrated a gene-expression signature that could aid detection of blood doping. However, current anti-doping testing does not involve blood collection into tubes with RNA preservative. This study investigated if whole blood in long-term storage and whole blood leftover from standard haematological testing in short-term storage could be used for transcriptomic analysis despite lacking RNA preservative. Whole blood samples were collected from thirteen and fourteen healthy males, for long-term and short-term storage experiments. Long-term storage: whole blood collected into Tempus™ tubes and K2EDTA tubes and subjected to long-term (i.e., −80°C) storage and RNA extracted. After storage, K2EDTA tubes were thawed and extracted using GeneJET RNA Purification Kit (Thermo Fisher Scientific, Vilnius, Lithuania) or Tempus™ Spin RNA Isolation Kit (Life Technologies, Carlsbad, CA, USA). RNA quality and purity was sufficient for gene expression analysis. Principle Component Analysis of microarray and RNA-seq gene expression data for long-term storage: When comparing gene expression between blood tubes with and without RNA preservation, 6% (4058 transcripts) were differentially expressed. RNA quantity, purity and integrity was not significantly compromised from long-term storage in blood storage tubes lacking RNA preservative, indicating that transcriptomic analysis could be conducted using anti-doping samples collected or biobanked without RNA preservation.

2021-08-13 | GSE181970 | GEO

Gene expression profiling of human whole-blood samples before and after the administration of recombinant human erythropoietin

Project description:As part of cross-platform comparisons of microarray and RNA-seq, this current experiment using the Affymetrix Human Transcriptome Array 2.0 aimed to quantify gene-level expression in subjects administered recombinant human erythropoietin over a 10-week protocol for the identification of gene signatures of blood doping. These results were compared to results obtained from other gene expression quantification platforms using the same experimental cohort, including the Illumina HumanHT-12v4 Expression BeadChips (archived in ArrayExpression; E-MTAB-2874), Illumina NextSeq 500 and MGI DNBSEQ-G400RS.

2021-11-04 | E-MTAB-11080 | biostudies-arrayexpress

RNA-seq of human whole-blood samples before and after the administration of recombinant human erythropoietin

Project description:As part of cross-platform comparisons of microarray and RNA-seq, the current experiment using the Illumina NextSeq 500 and MGI DNBSEQ-G400RS aimed to quantify gene-level expression in subjects administered recombinant human erythropoietin over a 10-week protocol for the identification of gene signatures of blood doping. These results were compared to results obtained from other gene expression quantification platforms using the same experimental cohort, including the Illumina HumanHT-12v4 Expression BeadChips (archived in ArrayExpress; E-MTAB-2874) and Affymetrix Human Transcriptome Array 2.0 (archived in ArrayExpress; E-MTAB-11080).

2021-11-04 | GSE186294 | GEO

Integrating whole blood transcriptomic collection procedures into the current anti-doping testing system, including long-term storage and re-testing of anti-doping samples [Short-term]

Project description:Purpose: Recombinant human erythropoietin administration studies involving “omics” approaches have demonstrated a gene-expression signature that could aid detection of blood doping. However, current anti-doping testing does not involve blood collection into tubes with RNA preservative. This study investigated if whole blood in short-term storage could be used for transcriptomic analysis despite lacking RNA preservation. Methods: Whole blood samples were collected from fourteen male healthy individuals. Short-term storage: whole blood collected into K2EDTA tubes and subjected to short-term (i.e., at 4°C) storage for 6 hours, 12 hours, 24 hours and 48 hours. After storage, blood from K2EDTA tubes were transferred into Tempus™ Blood tubes, and then extracted using Tempus™ Spin RNA Isolation Kit (Life Technologies, Carlsbad, CA, USA). Samples from four subjects of each time point that presented higher RIN value (≥7) were selected for RNA_Seq analysis. Results: The experiment provided RNA quality and purity for gene expression analysis. Considering 6-hours storage as a reference group, the number of differentially expressed genes were 19, 45 and 70 in comparison to 12, 24 and 48-hours, respectively (which means 0.37, 0.88 and 1.37% of mapped genes). Of the 19 differentially expressed genes in the comparison 6 vs. 12-hours, 9 overlapped with the 45 in the comparison 12 vs. 24-hours. Furthermore, 40 of those 45 overlapped with the 70 differentially expressed in the comparison 6 vs. 48-hours. None of the transcripts described in previous studies (Durussel et al. 2016; Wang, Durussel, et al. 2017) were differently expressed. Conclusion: RNA quantity, purity and integrity was not significantly compromised from short-term storage in blood storage tubes lacking RNA stabilisation, indicating that transcriptomic/omics analysis could be conducted using anti-doping samples collected without RNA preservation.

2021-09-03 | GSE183132 | GEO

Transcription profiling of human HepG2 cells treated with UVC (254 nm) for 30 seconds and transduced with Adenoviral vector carrying the Hepatitis B Virus HBx gene vs Adenoviral vector control under UV treatment

Project description:Transcriptional profiling of HepG2 cells treated with UVC (254 nm) for 30 seconds and transduced with Adenoviral vector carrying the Hepatitis B Virus HBx gene vs Adenoviral vector control under UV treatment Experiment Overall Design: One-condition experiment, AdHBx (experimental) UV vs AdEasy (vector control) UV. HepG2 cells were infected with either recombinant HBx adenoviruses (AdHBx) or control adenoviruses (AdEasy). Seventy-two hours later, infected cells were exposed for 30 seconds to UVC (254 nm) irradiation with a germicidal lamp calibrated to deliver 8 or 16 J/m2. Cells were then harvested for either ChIP-chip profiling or expression microarray profiling

2008-10-21 | E-GEOD-11064 | biostudies-arrayexpress

Identification of RNA Markers in Red Blood Cells for Doping Control in Autologous Blood Transfusion

Project description:We conducted an animal experiment and used total RNA sequencing (RNA-Seq) to identify novel RNA markers to detect autologous blood transfusion (ABT) doping within red blood cells (RBCs) as a pilot study before human trials. The total RNA-Seq and bioinformatics were performed using rat RBCs that were stored 10- and 20-day periods with citrate–phosphate–dextrose solution with adenine (CPDA) at 4°C In the results, we identified three significant genes whose expression levels were drastically decreased according to the storage period. Our novel insights may allow future studies to develop a test method for ABT doping.

2022-07-14 | GSE205806 | GEO

Adenoviral vector vaccination induces a conserved program of CD8+ T cell memory differentiation in mouse and man

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2015-09-23 | GSE73314 | GEO

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