The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing
Pre-analytical factors have a significant influence on circulating microRNA (miRNA) profiling. The aim of this study was a comprehensive assessment of the impact of the anticoagulant type in blood collection tubes on circulating plasma miRNA profiles using small RNA sequencing. Blood from ten health...
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2022-09-01
|
| Series: | International Journal of Molecular Sciences |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1422-0067/23/18/10340 |
| _version_ | 1797487472004825088 |
|---|---|
| author | Andrey V. Zhelankin Liliia N. Iulmetova Elena I. Sharova |
| author_facet | Andrey V. Zhelankin Liliia N. Iulmetova Elena I. Sharova |
| author_sort | Andrey V. Zhelankin |
| collection | DOAJ |
| description | Pre-analytical factors have a significant influence on circulating microRNA (miRNA) profiling. The aim of this study was a comprehensive assessment of the impact of the anticoagulant type in blood collection tubes on circulating plasma miRNA profiles using small RNA sequencing. Blood from ten healthy participants (five males and five females from 25 to 40 years old) was taken in collection tubes with four different anticoagulants: acid citrate dextrose (ACD-B), sodium citrate, citrate-theophylline-adenosine-dipyridamole (CTAD) and dipotassium-ethylenediaminetetraacetic acid (K2 EDTA). Platelet-free plasma samples were obtained by double centrifugation. EDTA plasma samples had elevated levels of hemolysis compared to samples obtained using other anticoagulants. Small RNA was extracted from plasma samples and small RNA sequencing was performed on the Illumina NextSeq 500 system. A total of 30 samples had been successfully sequenced starting from ~1 M reads mapped to miRNAs, allowing us to analyze their diversity and isoform content. The principal component analysis showed that the EDTA samples have distinct circulating plasma miRNA profiles compared to samples obtained using other anticoagulants. We selected 50 miRNA species that were differentially expressed between the sample groups based on the type of anticoagulant. We found that the EDTA samples had elevated levels of miRNAs which are abundant in red blood cells (RBC) and associated with hemolysis, while the levels of some platelet-specific miRNAs in these samples were lowered. The ratio between RBC-derived and platelet-derived miRNAs differed between the EDTA samples and other sample groups, which was validated by quantitative PCR. This study provides full plasma miRNA profiles of 10 healthy adults, compares them with previous studies and shows that the profile of circulating miRNAs in the EDTA plasma samples is altered primarily due to an increased level of hemolysis. |
| first_indexed | 2024-03-09T23:48:10Z |
| format | Article |
| id | doaj.art-d1ba5aa7e9b64212b5640dfdd0ca7f61 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-09T23:48:10Z |
| publishDate | 2022-09-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-d1ba5aa7e9b64212b5640dfdd0ca7f612023-11-23T16:40:03ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-09-0123181034010.3390/ijms231810340The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA SequencingAndrey V. Zhelankin0Liliia N. Iulmetova1Elena I. Sharova2Department of Molecular Biology and Genetics, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, 119435 Moscow, RussiaDepartment of Molecular Biology and Genetics, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, 119435 Moscow, RussiaDepartment of Molecular Biology and Genetics, Federal Research and Clinical Center of Physical-Chemical Medicine of Federal Medical Biological Agency, 119435 Moscow, RussiaPre-analytical factors have a significant influence on circulating microRNA (miRNA) profiling. The aim of this study was a comprehensive assessment of the impact of the anticoagulant type in blood collection tubes on circulating plasma miRNA profiles using small RNA sequencing. Blood from ten healthy participants (five males and five females from 25 to 40 years old) was taken in collection tubes with four different anticoagulants: acid citrate dextrose (ACD-B), sodium citrate, citrate-theophylline-adenosine-dipyridamole (CTAD) and dipotassium-ethylenediaminetetraacetic acid (K2 EDTA). Platelet-free plasma samples were obtained by double centrifugation. EDTA plasma samples had elevated levels of hemolysis compared to samples obtained using other anticoagulants. Small RNA was extracted from plasma samples and small RNA sequencing was performed on the Illumina NextSeq 500 system. A total of 30 samples had been successfully sequenced starting from ~1 M reads mapped to miRNAs, allowing us to analyze their diversity and isoform content. The principal component analysis showed that the EDTA samples have distinct circulating plasma miRNA profiles compared to samples obtained using other anticoagulants. We selected 50 miRNA species that were differentially expressed between the sample groups based on the type of anticoagulant. We found that the EDTA samples had elevated levels of miRNAs which are abundant in red blood cells (RBC) and associated with hemolysis, while the levels of some platelet-specific miRNAs in these samples were lowered. The ratio between RBC-derived and platelet-derived miRNAs differed between the EDTA samples and other sample groups, which was validated by quantitative PCR. This study provides full plasma miRNA profiles of 10 healthy adults, compares them with previous studies and shows that the profile of circulating miRNAs in the EDTA plasma samples is altered primarily due to an increased level of hemolysis.https://www.mdpi.com/1422-0067/23/18/10340circulating microRNAplasmaanticoagulantmiRNA-seqextracellular biomarkers |
| spellingShingle | Andrey V. Zhelankin Liliia N. Iulmetova Elena I. Sharova The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing International Journal of Molecular Sciences circulating microRNA plasma anticoagulant miRNA-seq extracellular biomarkers |
| title | The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing |
| title_full | The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing |
| title_fullStr | The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing |
| title_full_unstemmed | The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing |
| title_short | The Impact of the Anticoagulant Type in Blood Collection Tubes on Circulating Extracellular Plasma MicroRNA Profiles Revealed by Small RNA Sequencing |
| title_sort | impact of the anticoagulant type in blood collection tubes on circulating extracellular plasma microrna profiles revealed by small rna sequencing |
| topic | circulating microRNA plasma anticoagulant miRNA-seq extracellular biomarkers |
| url | https://www.mdpi.com/1422-0067/23/18/10340 |
| work_keys_str_mv | AT andreyvzhelankin theimpactoftheanticoagulanttypeinbloodcollectiontubesoncirculatingextracellularplasmamicrornaprofilesrevealedbysmallrnasequencing AT liliianiulmetova theimpactoftheanticoagulanttypeinbloodcollectiontubesoncirculatingextracellularplasmamicrornaprofilesrevealedbysmallrnasequencing AT elenaisharova theimpactoftheanticoagulanttypeinbloodcollectiontubesoncirculatingextracellularplasmamicrornaprofilesrevealedbysmallrnasequencing AT andreyvzhelankin impactoftheanticoagulanttypeinbloodcollectiontubesoncirculatingextracellularplasmamicrornaprofilesrevealedbysmallrnasequencing AT liliianiulmetova impactoftheanticoagulanttypeinbloodcollectiontubesoncirculatingextracellularplasmamicrornaprofilesrevealedbysmallrnasequencing AT elenaisharova impactoftheanticoagulanttypeinbloodcollectiontubesoncirculatingextracellularplasmamicrornaprofilesrevealedbysmallrnasequencing |