Sunday, July 13, 2008

Does EPO Enhance Performance?

The following is actually a little dated as it is from 2002 when I wrote it then, but the basic info is still valid. I thought it would be appropriate with the Olympics games coming up and the Tour De France.


Mike N

Does EPO improve athletic performance and can it be detected?


EPO is the primary hormone that is responsible for regulation of erythrocyte and hemoglobin production. Endogenous EPO is a 36-kilodalton, 16 amino acid glycoprotein.[1] EPO is manufactured primarily in the kidney which accounts for about 90% of the de novo production and the other 10% synthesis occurs in the liver.[1] The synthesis of erythropoietin is stimulated by tissue hypoxia, which may be brought on by anemia, high altitude (exercise and/or residence), impaired oxygenation of hemoglobin as the result of cardiac or pulmonary disease.[2] The half life of endogenous EPO is about 6 to 9 hours.[3] Erythropoietin stimulates red blood cell formation by the bone marrow. Erythrocyte maturation take about 3 to 5 days from the initial hypoxaemia-induced increase in endogenous EPO.[2] Performance of endurance exercise can be increased by increasing the amount of RBCs. .

DNA recombinant human EPO (rhEPO) is synthesized from Chinese hamster ovary cells and is nearly identical biochemically and immunologically to endogenous EPO.[1] RhEPO became available in 1987 in Europe and 1989 in the US. RhEPO is used clinically to treat anemia due to chronic kidney failure and as a possible therapeutic alternative to blood transfusion.[35] The standard clinical dose of rhEPO is about 200 to 250 U/kg bodyweight (BW).[4] Administration can be either intravenously (IV) or subcutaneous (SC). The half life of rhEPO is 4 to 5 hours when given by IV and 19 to 22 hours when given by SC. The mean elimination half life was calculated to be about 42.0+/- 34.2 hours, and clearence (uncorrected for bioavailability) was calculated to be 0.05 +/- 0.01 h-1 kg-1. [36]

Athletes and rhEPO

Athletes have been using ergogenic aids for decades. There is a constant race between those athletes using illegal ergogenic aids (just take a look at the International Olympic Committee's long list of banned substances) and those in charge of designing tests to catch the cheaters.[5] The use of rhEPO as a means to enhance aerobic performance is believed to be common practice among elite athletes in endurance sports such as cross-country skiing, cycling, triathlon, and distance running. Following the 1984 Summer Olympic Games, the IOC officially banned the use of all forms of blood doping and in 1990 banned the use of rhEPO.[6] Blood doping has gone through a few different revisions, but the current definition is defined by the IOC Medical Commission as, " the use of an artifice, whether substance or method, potentially dangerous to athlete's health and/or capable of enhancing performance, or the presence in the athlete's body of a substance, or the ascertainment of the use of a method on the list annexed to the Olympic Movement Anti-Doping Code. [7]

In addition to being illegal it carries potential health risks. Documented side effects of rhEPO include (i) hyperviscosity (haematocrit >52 % for males and >55% for females); (ii) arterial hypertension; (iii) cerebral convulsion/hypertensive encephalopathy; (iv) thromboembolism; (v) influenza-like syndrome; and (iv) hyperkalaenmia. [8,9,10] About two dozen deaths of European professional and amateur cyclists have been attributed to the illegal use of rhEPO. [11,12,37]

Prevalence of rhEPO misuse

Charles Yesalis, professor of health and human development at Pennsylvania State University says, "There's a small percent of athletes who did not use drugs. There's a small percent of athletes who use drugs and get caught and there is a very large percent who use drugs, who don't get caught." [13] Scarpino et al.[14] interviewed 1015 Italian male and female cyclist regarding the use of EPO or RBC reinfusion. 7% said they were "regular" users and 25% indicated that they were "occasional" users. Anecdotal evidence by international drug control personnel suggest that the top 3-6% of elite endurance sport athletes have used rhEPO at some point in their career. [1] During the 1998 Tour de France, several cyclist and other athletes admitted they took EPO after a Tour de France Festina team car carrying 235 doses of rhEPO was stopped at the France-Belgian border in July 1998.[16] Many teams withdrew from the race due to the enhanced scrutiny with only 96 of original 189 participants finishing.

rhEPO and exercise performance

Birkeland study

Birkeland et al.[17] was the first randomized, placebo controlled study to show that the administration of moderate doses of rhEPO for 4 weeks significantly increased maximal oxygen uptake and time to exhaustion in healthy, well trained endurance athletes. Birkeland et al.[17] studied the effects of rhEPO administration on serum levels of sTfR and cycling performance. A double-blind, placebo controlled study was conducted with the administration of 5000 U of rhEPO (N=10) or placebo (N=10) three times weekly for 4 weeks to male athletes. Haematocrit and concentration of hemoglobin, sTfR, ferritin, EPO were measured. The effects on performance were quantified by measuring time to exhaustion and maximal oxygen uptake on a cycle ergometer.[17]

20 healthy, well trained athletes from cycling, orienteering, running, triathlon, swimming, and cross-country volunteered for the study. Blood samples were obtained immediately before the start of the treatment and thereafter three times weekly during the treatment period, each of he first 5 days after stopping the treatment and two times weekly during the remainder of the 4 week follow-up period. After a warm up for 5 to 10 minutes the subjects cycled to exhaustion. The initial workload was set at 100W and increased by 50 W every 2 minutes until volitional exhaustion. The subjects were instructed to maintain cadence at 85-95 rpm and if the cadence fell below 75 rpm the test was stopped and time to exhaustion was recorded. Expired volume was collected over the last 3-4 minutes of the test. The volume was measured by spirometer and the room air and expired air concentrations were measured by mass spectrometry. Posttest analysis of respiratory quotient and heart rate were used to confirm maximal oxygen uptake had been determined. Treatment with rhEPO was stopped if the haematocrit reached >= 50%. This happened early in two subject who were stopped at day 17 and 23. Haematocrit did not change in the placebo group. By day 30, everyone in the treatment group had reached a haematocrit of 50% Maximal oxygen at baseline for the EPO group was 63.3 +/- 3.9 mL and increased to 68.1 +/- 5.4 mL. Maximal oxygen uptake showed a slight but not significant tendency to increase in the placebo group during the study period. Time to exhaustion increased from 12.8 +/- 1.0 minute at baseline to 14.0 +/- 1.4 minute one day posttreatment in the EPO group (P<.0001) and from 13.1 +/- 1.5 minute to 13.3 +/- 1.5 minutes in the control group (P=0.04).[17] These data in the study indicated a performance advantage for atleast 2 weeks longer than any indication of rhEPO use. It also shows that the use of EPO did enhance exercise performance in both maximal oxygen uptake and time to exhaustion.

Other studies

Balsom et al.[18] examined the effects of rhEPO exercise performance in 6 healthy male physical education students. Subjects received 20 U/kg BW administered SC 3 times per week for 6 weeks. Following rhEPO treatment there was an 8% increase in treadmill VO2 max (pre=4.76 +/- 0.2 L/min, post=5.14 +/- 0.2 L/min). These data also showed an increase in exercise performance.

A limited number of human studies of the effects of EPO in healthy males [18,38,20,21] and in endurance trained athletes[19,17] using SC administered rhEPO ranging from 60 to 232 U/kg BW per week for 4 to 6 weeks. The data from these studies on either healthy or trained humans suggests that moderate rhEPO supplementation results in significant increases in VO2 max (7 to 9%) and endurance performance (9 to 17%).[1] This suggests that the use of rhEPO as an ergogenic aid for performance enhancement is a viable option (although illegal and has health risks associated with its use).

Exercise and endogenous EPO

Another question is, does exercise by itself cause an increase in endogenous EPO? Garaeu et al.[22] reported that serum EPO was not significantly influenced by 30 to 90 minutes of "intense exercise" performed by male cyclist, hockey players, and volleyball players. In more prolonged exercise bouts, no difference was observed in serum EPO in female and male runners before and immediately following a 6 hour race.[23] Bergland et al.[24] reported similar findings that resting serum EPO levels in endurance and strength training in both men and women who were long term residents of moderate altitude (1900m). Roberts et al .[25] observed similar results in elite male swimmers and age matched controls at 1000m above sea level. Based on these data, it appears that serum EPO concentrations are not significantly affected by acute bouts of aerobic exercise

Testing for rhEPO

Wilber et al.[1] describes how testing for EPO can be divided into two categories--indirect and direct methods. Indirect methods would include: macrocytic hypochromatic RBCs (blood), soluble transferrin receptor (blood), and multiple markers of enhanced erythropoiesis (blood). Direct methods would include: electrophoretic mobility (blood and urine), and isoelectric patterning (urine).

Indirect rhEPO testing methods

Macrocytic Hypochromatic Erythrocytes

Casoni et al.[26] concluded from research on the evaluation of the effect of rhEPO on several RBC indices in 20 male aerobic athletes that it is possible to establish a "cut off" value(0.6%) for macrocytic hypochromatic erythrocytes. This method demonstrated good specificity (0% false positives) but relatively poor sensitivity since more than 50% of the rhEPO samples were not detected.

Serum Soluble Transferrin Receptor (sTfr)

STfr is released primarily from erythropoietic progenitors and has been suggested that it may serve as an indirect marker for rhEPO detection. Gareau et al.[22] studied the effect of rhEPO on sTfr concentrations in 24 patients with kidney disease and found that resting sTfr was significantly higher in patients with kidney disease (3135 +/- 312 micro g/L) who received rhEPO than in the in control group with kidney disease (2195 +/- 225 micro g/L). Also, the sTfr was not influenced by 30 to 90 minutes of "intense exercise" performed by hockey and volleyball players; thus helping to reduce false positives. Birkeland et al.[17] showed that sTfr concentration doubled as a result of rhEPO supplementation. Gareau et al.[22] found that the soluble transferring receptor/ferritin ratio (sTfr/fr) is unaffected by hydration level and may be more accurate since the hydration level can affect the haematocrit level. It was concluded that values exceeding 10 micro g/L for sTfr and 403 for sTfr/fr indicated the probable intake (p<0.05) of rhEPO,[28] and Birkeland et al.[17] reported similar findings. Audran et al.[19] evaluated the use of sTfr/serum protein ratio for the detection of rhEPO. The sTfr/protein ratio has an advantage since (i) serum protein controls for possible exercise-induced haemoconcentration, and (ii) serum protein may be a better marker than serum ferritin since iron supplementation is a common practice among endurance athletes and thus may influence serum ferritin levels.[19] These tests show great promised, but as Birkeland et al.[17] demonstrated that although sTfr remain elevated for up to 1 week following rhEPO supplementation, the effects of rhEPO on exercise performance may last up to 3 weeks after last administration.

Multiple Markers for Enhanced Erythropoiesis

The responses of several indirect markers of rhEPO use were measured in whole blood (haematocrit, reticulocyte haematocrit, percentage of macrocytic RBCs) and serum (serum EPO, sTfr, serum ferritin).[29] These measurements were used to derive the ON model. The ON model was approved for use as a primary test for detection of the illicit use of rhEPO by the IOC Medical Commission in August of 200. It was used in conjunction with a confirmatory test (isoelectric patterning).[29]

Direct rhEPO testing methods

Electrphoretic Mobility

Endogenous EPO and rhEPO are almost biochemically and immologically identical. However, the rhEPO molecule is less anionic compared to the endogenous EPO molecule.[1] This difference is exploited by the electrophoretic mobility test. The results of the test suggest that electrophoretic mobility technique cannot accurately detect rhEPO in blood samples obtained 7 or more days after rhEPO use.[30] Out of 45 samples taken there were no false positives observed in either blood or urine.[30]

Isoelectric Pattering

The Isoelectric pattering test in used in conjunction with the ON model by the IOC since the Sidney Olympic games. The test is based on evidence that the rhEPO molecule has a slightly greater number of sialic acids in its glycosylated side chains.[31,32] This small difference allows for differentiation by using the isoelectric patterning method. Isoelectric patterning has two big advantages; (i) it allows the direct measurement of rhEPO, (ii) it is noninvasive (measured in urine). The downside is that the sensitivity of the test may be limited to less than 3 days after supplementation.

Drug Testing

At the Salt Lake City games in 2002, the IOC decided that both the blood and urine samples must return abnormal results before an athlete will be considered positive for rhEPO use.[33] Currently the UCI (oversees Tour De France) only tests for the percentage of RBCs per unit vol, and you can't race if it is above 50%. The duration of the effect on performance is greater than the duration of any hematological changes associated with rhEPO misuse.[17] As a result, an "open window" may exist where there is no evidence of rhEPO misuse but where performance is enhanced. Yesalis says that a urine test is a good start but needs further verification. "It also needs to withstand legal challenge is an athlete opposes the results," he says [16]


EPO is the primary hormone that is responsible for regulation of erythrocyte and hemoglobin production. Performance of endurance exercise can be increased by increasing the amount of RBCs and thus allowing better oxygen delivery. DNA recombinant human EPO (rhEPO) became available in the US in 1989 and is nearly identical both biochemically and immunologically to endogenous EPO. RhEPO is used clinically to treat anemia due to chronic kidney failure and as a possible therapeutic alternative to blood transfusion[35] and can be given by IV or SC.

Athletes have been using ergogenic aids for decades and many are now believed to be using rhEPO but not without possible health affects.[11,12,37] RhEPO became widely known after the Tour de France fiasco in 1989 after several cyclist admitted they took EPO.

RhEPO does appear to increase exercise performance. Birkeland et al.[17] was one of the first to show this using a placebo controlled, double blind study with trained subjects a Audran et al.[19] showed the same trend with trained athletes. Others later showed the same trend of the effects of EPO in healthy males.[18,38,21] Exercise by itself does not seem to increase endogenous EPO.[22,23,24]

There are two main ways to measure rhEP0-indirect and direct methods. Indirect methods would include: macrocytic hypochromatic RBCs (blood), soluble transferrin receptor (blood), and multiple markers of enhanced erythropoiesis (blood). Direct methods would include: electrophoretic mobility (blood and urine), and isoelectric patterning (urine).[1]

At the Salt Lake City games in 2002, the IOC decided that both the blood and urine samples must return abnormal results before an athlete will be considered positive for rhEPO use.[33] Currently the UCI (oversees Tour De France) only tests for the percentage of RBCs per unit volume, and you can't race if it is above 50%.[34] There currently exist an "open window" where there is no evidence of rhEPO misuse but where performance is enhanced. In theory an athlete could time it out so that they do not show up positive on a drug test, but have enhanced performance. It is speculated that some elite athletes have gone beyond EPO and are beginning to experiment with hemoglobin oxygen carries (HBOCs) such as bovine blood and human blood, or even perfluorocarbons (PFCs). [5]

It is a race between the cheaters and those trying to catch them. Even though there has been great advance in detection of banned ergogenic aids, without the use of constant out of competition testing it will be virtually impossible with today's technology to keep all drugs out of elite competition.


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