Andrew T. Kicman

Pharmacology of anabolic steroids

Athletes and bodybuilders have recognized for several decades that the use of anabolic steroids can promote muscle growth and strength but it is only relatively recently that these agents are being revisited for clinical purposes. Anabolic steroids are being considered for the treatment of cachexia associated with chronic disease states, and to address loss of muscle mass in the elderly, but nevertheless their efficacy still needs to be demonstrated in terms of improved physical function and quality of life. In sport, these agents are performance enhancers, this being particularly apparent in women, although there is a high risk of virilization despite the favourable myotrophic–androgenic dissociation that many xenobiotic steroids confer. Modulation of androgen receptor expression appears to be key to partial dissociation, with consideration of both intracellular steroid metabolism and the topology of the bound androgen receptor interacting with co‐activators. An anticatabolic effect, by interfering with glucocorticoid receptor expression, remains an attractive hypothesis. Behavioural changes by non‐genomic and genomic pathways probably help motivate training. Anabolic steroids continue to be the most common adverse finding in sport and, although apparently rare, designer steroids have been synthesized in an attempt to circumvent the dope test. Doping with anabolic steroids can result in damage to health, as recorded meticulously in the former German Democratic Republic. Even so, it is important not to exaggerate the medical risks associated with their administration for sporting or bodybuilding purposes but to emphasize to users that an attitude of personal invulnerability to their adverse effects is certainly misguided.

Low-dose MDMA (“ecstasy”) induces vasopressin secretion

p=0·025, one tail). Mean initial cortisol concentration was 331·4 nmol/L (range 208·8–603·4 nmol/L), which increased, although not significantly (p>0·05), to 377·2 nmol/L (range 268·4–583·3 nmol/L) at 2 h. Street “ecstasy” frequently contains more than 100 mg of MDMA. In this study, a single relatively small dose caused an acute rise in AVP concentration at a time of day when it would not be expected to change. The rise in AVP was accompanied by a small fall in plasma sodium concentrations. The hyponatraemic illness experienced by some users is thus likely to be linked to the drug’s ability to stimulate secretion of AVP. Hence, if fluid intake is excessive, even a relatively small dose of MDMA could lead to symptoms of hyponatraemia. The rise in AVP does not seem to be part of a generalised stress response because there was no significant change in plasma cortisol concentration. It therefore seems that MDMA-induced hyponatraemia is unlikely to be due to a rare and idiosyncratic reaction, but results from a pharmacological effect compounded by excessive fluid ingestion. Animal studies show that MDMA stimulates the output of serotonin by serotonergic neurones, and AVP secretion is regulated by serotonergic pathways. The message is that those who take “ecstasy” or similar drugs may be at risk of hyponatraemia and should, therefore, avoid drinking fluid in excess of the body’s requirements. This may be difficult for users to estimate because MDMA reduces perception of thirst and impairs judgment.

Criteria to indicate testosterone administration.

A detection method for testosterone administration was developed using radioimmunoassay to measure the urinary ratios of testosterone (T) to epitestosterone (E) and to luteinizing hormone (LH). A comparative study of the effect on these ratios of a single intramuscular injection of testosterone heptanoate followed by stimulation with human chorionic gonadotrophin (HCG) in three normal men was undertaken. To allow immediate investigation, a commercially supplied epitestosterone antiserum was used. This study showed that both T/E and T/LH ratios could be used to detect testosterone administration, the latter also being an indicator of HCG use due to cross-reactivity with the LH antiserum. Subsequently, an epitestosterone antiserum of superior specificity was raised and used in a study to demonstrate the insignificant effect of exercise on these ratios. Finally, an intramuscular injection of a combined preparation of testosterone/epitestosterone heptanoates resulted in raised ratios of T/LH but not of T/E. This demonstrated the importance of the T/LH ratio in circumstances where the T/E ratio can be easily circumvented.

Detection of ketamine and its metabolites in urine by ultra high pressure liquid chromatography-tandem mass spectrometry

Current analytical methods used for screening drugs and their metabolites in biological samples from victims of drug-facilitated sexual assault (DFSA) or other vulnerable groups can lack sufficient sensitivity. The application of liquid chromatography, employing small particle sizes, with tandem mass spectrometry (MS/MS) is likely to offer the sensitivity required for detecting candidate drugs and/or their metabolites in urine, as demonstrated here for ketamine. Ultra-performance liquid chromatography-mass spectrometry (UPLC-MS/MS) was performed following extraction of urine (4 mL) using mixed-mode (cation and C8) solid-phase cartridges. Only 20 microL of the 250 microL extract was injected, leaving sufficient volume for other assays important in DFSA cases. Three ion transitions were chosen for confirmatory purposes. As ketamine and norketamine (including their stable isotopes) are available as reference standards, the assay was additionally validated for quantification purposes to study elimination of the drug and primary metabolite following a small oral dose of ketamine (50 mg) in 6 volunteers. Dehydronorketamine, a secondary metabolite, was also analyzed qualitatively to determine whether monitoring could improve retrospective detection of administration. The detection limit for ketamine and norketamine was 0.03 ng/mL and 0.05 ng/mL, respectively, and these compounds could be confirmed in urine for up to 5 and 6 days, respectively. Dehydronorketamine was confirmed up to 10 days, providing a very broad window of detection.

Anabolic steroid use: Patterns of use and detection of doping

Anabolic-androgenic steroids (AAS) were the first identified doping agents that have ergogenic effects and are being used to increase muscle mass and strength in adult males. Consequently, athletes are still using them to increase physical performance and bodybuilders are using them to improve size and cosmetic appearance. The prevalence of AAS use has risen dramatically over the last two decades and filtered into all aspects of society. Support for AAS users has increased, but not by the medical profession, who will not accept that AAS use dependency is a psychiatric condition. The adverse effects and potential dangers of AAS use have been well documented. AAS are used in sport by individuals who have acquired knowledge of the half-lives of specific drugs and the dosages and cycles required to avoid detection. Conversely, they are used by bodybuilders in extreme dosages with the intention of gaining muscle mass and size, with little or no regard for the consequences. Polypharmacy by self-prescription is prevalent in this sector. Most recently, AAS use has filtered through to ‘recreational street drug’ users and is the largest growth of drugs in this subdivision. They are taken to counteract the anorexic and cachectic effects of the illegal psychotropic street drugs. Screening procedures for AAS in World Anti-Doping Agency accredited laboratories are comprehensive and sensitive and are based mainly on gas chromatography-mass spectrometry, although liquid chromatography-mass spectrometry is becoming increasingly more valuable. The use of carbon isotope mass spectrometry is also of increasing importance in the detection of natural androgen administration, particularly to detect testosterone administration. There is a degree of contentiousness in the scenario of AAS drug use, both within and outside sport. AAS and associated doping agents are not illegal per se. Possession is not an offence, despite contravening sporting regulations and moral codes. Until AAS are classified in the same capacity as street drugs in the UK, where possession becomes a criminal offence, they will continue to attract those who want to win at any cost. The knowledge acquired by such work can only assist in the education of individuals who use such doping agents, with a view to minimizing health risks and hopefully once again create a level playing field in sport.

The effect of 3,4-methylenedioxymethamphetamine (MDMA, ?ecstasy?) and its metabolites on neurohypophysial hormone release from the isolated rat hypothalamus

Methylenedioxymethamphetamine (MDMA, “ecstasy”), widely used as a recreational drug, can produce hyponatraemia. The possibility that this could result from stimulation of vasopressin by MDMA or one of its metabolites has been investigated in vitro. Release of both oxytocin and vasopressin from isolated hypothalami obtained from male Wistar rats was determined under basal conditions and following potassium (40 mM) stimulation. The results were compared with those obtained for basal and stimulated release in the presence of MDMA or metabolites in the dose range 1 μM to 100 pM (n=5 – 8) using Students t‐test with Dunnetts correction for multiple comparisons. All compounds tested affected neurohypophysial hormone release, HMMA (4‐hydroxy‐3‐methoxymethamphetamine) and DHA (3,4‐dihydroxyamphetamine) being more active than MDMA, and DHMA (3,4‐dihydroxymethamphetamine) being the least active. The effect on vasopressin release was greater than that on oxytocin. In the presence of HMMA the ratio test:control for basal release increased for vasopressin from 1.1±0.16 to 2.7±0.44 (s.e.m., P<0.05) at 10 nM and for oxytocin from 1.0±0.05 to 1.6±0.12 in the same hypothalami. For MDMA the ratio increased to 1.5±0.27 for vasopressin and to 1.28±0.04 for oxytocin for 10 nM. MDMA and its metabolites can stimulate both oxytocin and vasopressin release in vitro, the response being dose dependent for each drug with HMMA being the most potent.

Serum IGF-I and IGF binding proteins 2 and 3 as potential markers of doping with human GH.

IGF‐I and IGF binding protein (IGFBP)‐3 levels in man are positively regulated by GH status; in contrast, evidence suggests an inverse relationship between GH status and IGFBP‐2. We investigated the effects of somatropin administration on the serum concentrations of these analytes, together with serum and urinary concentrations of GH, to evaluate their potential as markers in the development of a test for detecting doping with GH in sports competitors.

Homocysteine induced cardiovascular events: a consequence of long term anabolic-androgenic steroid (AAS) abuse

Objectives: The long term effects (>20 years) of anabolic-androgenic steroid (AAS) use on plasma concentrations of homocysteine (HCY), folate, testosterone, sex hormone binding globulin (SHBG), free androgen index, urea, creatinine, haematocrit (HCT), vitamin B12, and urinary testosterone/epitestosterone (T/E) ratio, were examined in a cohort of self-prescribing bodybuilders. Methods: Subjects (n = 40) were divided into four distinct groups: (1) AAS users still using AAS (SU; n = 10); (2) AAS users abstinent from AAS administration for 3 months (SA; n = 10); (3) non-drug using bodybuilding controls (BC; n = 10); and (4) sedentary male controls (SC; n = 10). Results: HCY levels were significantly higher in SU compared with BC and SC (p<0.01), and with SA (p<0.05). Fat free mass was significantly higher in both groups of AAS users (p<0.01). Daily energy intake (kJ) and daily protein intake (g/day) were significantly higher in SU and SA (p<0.05) compared with BC and SC, but were unlikely to be responsible for the observed HCY increases. HCT concentrations were significantly higher in the SU group (p<0.01). A significant linear inverse relationship was observed in the SU group between SHBG and HCY (r = −0.828, p<0.01), indicating a possible influence of the sex hormones in determining HCY levels. Conclusions: With mounting evidence linking AAS to adverse effects on some clotting factors, the significantly higher levels of HCY and HCT observed in the SU group suggest long term AAS users have increased risk of future thromboembolic events.

Counterfeiting in performance‐ and image‐enhancing drugs

The current drastic escalation in obesity may be contributing to the exponential rise in drugs used for image enhancement. Drugs such as anabolic-androgenic steroids (AAS) are perceived as a viable method of achieving a perfect physique. They are also the most widely abused drugs in sport. The Internet has encouraged the abuse of expensive drugs, particularly human growth hormone (hGH), resulting in increased importation for personal use. The substantial increase in this market has opened up avenues for counterfeiting, estimated as a multi-million pound business. The acute adverse effects from contaminated vials may result in a variety of pathologies including communicable diseases. In 2007, in the UK, a series of intramuscular abscesses, requiring surgical treatment, led us to study samples obtained from the underground market. The analysis of 38 parenteral samples and 19 oral samples of tablets was performed by a World Anti-Doping Agency (WADA) accredited laboratory, in an attempt to establish the extent of available counterfeit products. Fifty-three per cent (20) of the injectable AAS esters and 21% (4) of the oral tablets were counterfeit. Culture and sensitivity revealed the presence of skin commensal organisms, which may have contributed to the development of the abscesses. Users of AAS and hGH for sport, including bodybuilding, are currently risking their health because of counterfeit and poorly controlled products.

Physical effects of short-term recombinant human growth hormone administration in abstinent steroid dependency.

Background/Aims: Recombinant human growth hormone (rhGH) as opposed to cadaver pituitary GH is misused for physical improvement. Six days’ rhGH administration, in abstinent anabolic-androgenic steroid dependents, was compared with controls. Method: Male subjects (n = 48) were randomly divided into two groups: (1): control group (C), n = 24, mean ± SD, age 32 ± 11 years, height 1.8 ± 0.06 m; (2): rhGH-using group (0.058 IU·kg–1·day–1) (GH), n = 24, mean ± SD, age 32 ± 9 years, height 1.8 ± 0.07 m. Physiological measurements included anthropometry, strength, power and peak oxygen uptake (VO2 peak). Biochemical measurements included haemoglobin, packed cell volume, glucose, sodium, potassium, urea, creatinine, total protein, albumin, thyroid function, testosterone, prolactin, cortisol, GH and insulin-like growth factor-I (IGF-I). Results: Strength, peak power output and IGF-I significantly increased and total protein, albumin and free tetra-iodothyronine significantly decreased compared to controls (p < 0.05) and within the GH group (p < 0.017). Fat-free mass index and VO2 peak significantly increased, while body fat and thyroid-stimulating hormone significantly decreased within the GH group (p < 0.017). Conclusions: Short-term rhGH increased strength and power. Of therapeutic value is the possibility that muscle bulk and strength could be increased in patients with muscle-wasting conditions.