Until now, blood doping by athletes to increase performance has been difficult to detect by organizations dedicated to doping-free sports

Research into DNA and RNA keeps resulting in potential new opportunities for anatomic pathologists and clinical laboratories to conduct more precise testing. One recent example comes from Duke University (Duke) where researchers announced they’ve created microRNA-based tests that could be used to monitor blood doping in athletes, a news release reported.

According to the researchers, the finding could reveal athletes who removed their blood, took out the red blood cells, and transfused the cells into their bodies before competition. When conducted by medical laboratory professionals, such autologous blood therapies can enhance oxygen intake and increase performance during sports. However, these “self-transfusions” have been difficult to detect using current methods and that highlights the importance of ensuring these procedures are carried out by authorized healthcare facilities.

The researchers published their findings in the British Journal of Haemotology.

Research Focuses on RNA in Red Blood Cells

The World Anti-Doping Agency (WADA), an international organization aimed at research and education for doping-free sport, funded the Duke University research. WADA currently uses the Athlete Biological Passport to assess, over time, competitors’ body chemistries.

As the Duke researchers explored nucleic acids in red blood cells, they found that the cells actually do have a nucleus, contrary to popular belief. From there, they honed in on RNA.

Short RNA pieces, called microRNA (miRNA), control production of proteins in a cell, according to the researchers.

“While once thought to lack nucleic acids, red blood cells actually contain diverse and abundant RNA species,” the scientists noted in their paper. “In addition, proteomic analyses of red blood cells have identified the presence of Argonaute 2 (AGO2), supporting the regulatory function of miRNAs.”

The methodology Duke researchers followed involved these steps, among others:

  • Three units of blood were drawn from volunteers;
  • The researchers removed the white blood cells and about 80% of the plasma;
  • The remaining red blood cells were pure, just as they would need to be by someone doing autologous transfusion;
  • The researchers analyzed cell RNA samples at specific daily intervals: 1, 3, 7, 10, 14, 28, 36, and, 42 days;
  • They then compared samples to day 1 and recorded changes in RNA due to storage.

The researchers found:

  • Two types of miRNA increased during storage and two declined; and,
  • miR-720 had the most dramatic and consistent changes.

They concluded that finding increased miR-720 in athletes’ blood could be used as a biomarker for detecting stored red blood cells, which could indicate blood doping had taken place.

“The difficulty has been that the tests [WADA] have couldn’t tell the difference between a young blood cell and an old one,” Jen-Tsan Ashley Chi, MD, PhD, lead researcher on the study and Duke’s Associate Professor in Molecular Genetics and Microbiology, noted in the news release. “This increase in miR-720 is significant enough and consistent enough that it could be used as a biomarker for detecting stored red blood cells.” Chi is affiliated with Duke’s Center for Genomic and Computational Biology. (Photo copyright: Duke University.)

Implications for Detecting Blood Doping

How does this help clinical laboratories detect blood doping in athletes?

The researchers explained that RNA changes were, indeed, tell-tale signs of old blood cells circulating with normal cells. Those old blood cells could identify an athlete who did a self-transfusion of their blood before a competition.

However, before the test is used in sports more research is needed. Activity by the enzyme angiogenin in stored cells also is worthy of more exploration, as is its role in breaking apart larger RNA, the researchers noted.

“While autologous blood transfusions in athletes is very difficult to identify using conventional tests, it may be detectable based on the presence of red blood cells with levels of miR-720 significantly higher than the normal circulating cells. Further investigations will be necessary to identify the signals during red blood cell storage that stimulate angiogenin activation,” the study paper concluded.

Clinical Laboratories Involved in Sports Testing

In its 2017 Anti-Doping Testing Figures Report, WADA reported 322,050 samples were analyzed, a 7.1% increase from 300,565 samples in 2016. WADA accredits medical laboratories worldwide for conducting such analyses according to the organization’s code. This presents opportunities in sports medicine for medical laboratories to increase revenue through a new line of diagnostic tests.

In fact, the University of California-Los Angeles (UCLA) Olympic Analytical Laboratory is the world’s largest WADA-accredited sports testing facility. Clinical laboratory leaders interested in performing analyses of doping controls for sports—according to WADA’s standards—can contact the organization for its accreditation process.

The Duke study exemplifies how clinical laboratories can extend their services beyond patient care and enter a new realm of leveling playing fields worldwide.

—Donna Marie Pocius

Related Information:

New Finding Could Unmask Blood Doping in Athletes

Angiogenin-mediated tRNA Cleavage as a Novel Feature of Stored Red Blood Cells

Blood and Blood Components

2017 Anti-Doping Testing Figures Report

WADA Accreditation Process