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- Review -

Physiological and pharmacological basis for the ergogenic effects of growth hormone in elite sports

Christer Ehrnborg, Thord Rosén

Endocrine Section, Sahlgrenska University Hospital, S-413 45 Göteborg, Sweden

Abstract

Growth Hormone (GH) is an important and powerful metabolic hormone that is secreted in a pulsatile pattern from cells in the anterior pituitary, influenced by several normal and pathophysiological conditions. Human GH was first isolated in the 1950s and human derived cadaveric GH was initially used to treat patients with GH deficiency. However, synthetic recombinant GH has been widely available since the mid-1980s and the advent of this recombinant GH boosted the abuse of GH as a doping agent. Doping with GH is a well-known problem among elite athletes and among people training at gyms, but is forbidden for both medical and ethical reasons. It is mainly the anabolic and, to some extent, the lipolytic effects of GH that is valued by its users. Even though GH's rumour as an effective ergogenic drug among athletes, the effectiveness of GH as a single doping agent has been questioned during the last few years. There is a lack of scientific evidence that GH in supraphysiological doses has additional effects on muscle exercise performance other than those obtained from optimised training and diet itself. However, there might be synergistic effects if GH is combined with, for example, anabolic steroids, and GH seems to have positive effect on collagen synthesis. Regardless of whether or not GH doping is effective, there is a need for a reliable test method to detect GH doping. Several issues have made the development of a method for detecting GH doping complicated but a method has been presented and used in the Olympics in Athens and Turin. A problem with the method used, is the short time span (24_36 hours) from the last GH administration during which the test effectively can reveal doping. Therefore, out-of-competition testing will be crucial. However, work with different approaches to develop an alternative, reliable test is ongoing. (Asian J Androl 2008 May; 10: 373_383)

Keywords: growth hormone; IGF-I; doping; doping test; athletes; maximum exercise test; supraphysiological; anabolic androgenic steroids; bone markers

Correspondence to: Dr Thord Rosén, Endocrine Section, Sahlgrenska University Hospital, S-413 45 Göteborg, Sweden.
Tel: ++46-31-342-7055 Fax: +46-31-82-1524
E-mail: thord.rosen@medic.gu.se
Received 2007-12-11 Accepted 2007-12-18

DOI: 10.1111/j.1745-7262.2008.00403.x

1 Introduction

Doping with growth hormone (GH) is a well-known problem in the world of sports and has been known for decades [1]. GH was described as a potent performance-enhancing anabolic agent in The Underground Steroid Handbook first published in California in the early 1980s [2]. Its misuse has since increased, especially since the advent of recombinant GH in the late 1980s. It rapidly gained popularity as it was said to be "efficient, hard to detect and without major side-effects" and users can currently be found both among elite athletes and among people training at gyms [3, 4].

It is mainly the anabolic and, to some extent, lipolytic effect of GH that is valued by users since a reduction in fat mass, especially centrally located fat mass, is a desirable GH effect in a doping situation, especially in body-building competitions where a minimum of visible body fat is rewarded.

2 Historical background

The existence of a growth-promoting substance in the anterior hypophysis was described in animals in the 1920s [5]. Human GH was first isolated by Li et al. [6] in 1956 and, in the early 1970s, the structure of GH was subsequently shown to consist of a single polypeptide chain of 191 amino acids with two disulphide bridges and a molecular weight of 22 kDa [6_8]. It was then stated that 22 kDa GH is the major isoform of GH but a 20 kDa GH variant comprises 5%_10% of the pituitary GH and that a number of other isoforms produced by the pituitary also exist [9, 10]. The gene for GH has been cloned and characterised and synthetic GH is currently produced in bacteria using recombinant DNA technology [11].

Children with GH deficiency (GHD) have been treated with GH since the 1950s, when it was demonstrated that treatment with human GH, purified from cadaver pituitaries increased linear growth [12]. The first paper describing GH treatment in adults was presented in 1962 [13] and described the treatment of a 35-year-old GHD woman with human GH. The patient noticed "increased vigour, ambition and sense of well-being" after two months of treatment [13]. Treatment with GH was however initially restricted by the limited supply of GH and also by the recognition of the risk of Creutzfeldt-Jacobs disease with cadaveric GH, before the advent of widely available recombinant human GH in the mid-1980s. The introduction of recombinant GH made it possible to further study the effects of GH in adults and the consequences of the clinical entity of GHD, including its treatment, have been well described [14_17].

3 Physiology of GH secretion

GH is secreted in a pulsatile pattern from somatotrope cells in the anterior hypophysis, regulated in a complex pattern by two hypothalamic peptides; a stimulating hormone, GH releasing hormone (GHRH), and an inhibiting hormone, somatostatin [18_21].

GH secretion is influenced by several normal and pathophysiological conditions, such as gender, age, sleep, physical exercise, nutritional state and other metabolic factors.

3.1 Gender

A difference between men and women in the GH release at rest, with greater release in young women than in age-matched men, has been described [22_24]. Gonadal steroids interact with GH and the administration of oestrogen increases serum levels of GH [25_29]. Testostereone and GnRH treatment in hypogonadal men has been shown to increase GH secretion [30]. Oestrogens enhance GH secretion, mainly indirectly by inducing GH resistance resulting in higher serum levels of GH in females of reproductive age, a somewhat different secretion pattern and GH production rate [21, 29].

3.2 Age

It has been estimated that there is a 14% decrease in GH secretion per decade of adult life, following a peak during puberty [31].

3.3 Sleep

The GH levels are highest during slow wave sleep and lowest during rapid eye movement sleep [32].

3.4 Physical exercise

Physical exercise has a stimulatory effect on GH secretion [33]. The GH levels rise in response to acute exercise, with a threshold level of approximately 70% of VO₂-max [34], and a twofold rise in GH concentrations after a year of high-intensity aerobic training has been shown in subjects who exercised consistently above the lactate threshold [35].

3.5 Nutritional state and other metabolic factors

Fasting results in enhanced GH production [36], while it is suppressed by glucose [37] and fatty acids [38]. Certain amino acids such as leucine and arginine enhance GH secretion [39, 40].

3.6 Hormones

Hyperthyroidism is associated with increased GH secretion [41], while hypothyroidism is associated with low GH levels [42]. The net effect of corticoids is inhibition of the GH secretion [43].

3.7 Neurotransmitters

Both α₂-adrenergic agonists and cholinergic agents stimulate GH secretion [44], the latter probably via suppression of somatostatin release [45].

4 GH and muscles

GH is an important and powerful metabolic hormone. An anabolic effect by GH in normal adults was demonstrated in 1958 by Ikkos et al. [46] who observed a nitrogen-retention effect after GH administration. Patients with untreated acromegaly have shown a markedly increased body cell mass estimated from assessments of total body potassium [47]. The body cell mass in acromegalic patients decreases in response to surgical treatment [48]. Furthermore, it has been shown that acromegaly causes myopathy with hypertrophic, but functionally weaker muscles [49, 50]. This could indicate that there are negative effects on muscle function following exposure to high levels of GH for a long period of time.

GH promotes the positive protein balance in skeletal muscle by increasing protein synthesis and possibly by inhibiting protein breakdown [51].

Adult GHD patients have a reduced muscle mass, isometric muscle strength and functional exercise capacity compared with healthy controls [52, 53]. Furthermore, isokinetic muscle strength and local muscle endurance are reduced or in the lower range [52, 54_56]. The reduced muscle mass and isometric strength could be an effect of reduced muscle cross-sectional area in GHD patients [57], but it could also be caused by a reduction in the peak torque per muscle area [58], suggesting that contractile properties and neural activation might be responsible for the reduction in muscle strength in adult GHD patients.

GH replacement therapy in GHD adults increases lean body mass (LBM), exercise capacity, muscle volume, muscle mass and maximum voluntary isometric muscle strength [15, 52, 55, 59_63]. The changes in muscle mass and maximum voluntary isometric muscle strength has been shown to become apparent after approximately one year of therapy [55]. However, dynamic muscle strength has not shown any obvious increase in response to GH treatment [60, 64].

The proportion of fast-twitch, type-2 muscle fibres is increased in hypophysectomised rats [65]. However, the histology of muscles from GHD patients does not appear to differ from that of healthy adults [66] and it has not been possible to detect any changes in the proportions of muscle fibres in adult GHD patients receiving GH treatment [66, 67].

5 Lipolytic effects of GH

The lipolytic effects of GH have been known for decades. GH-induced lipolysis was first demonstrated in humans in 1959 [68]. Lipolytic effects have also been demonstrated in GHD and acromegaly patients. GHD patients have increased total body fat and reduced body cell mass and extracellular water (ECW) [69, 70]. GH treatment given to these patients improves the body composition [17, 54, 71]. Furthermore, patients with untreated acromegaly have a marked decrease in adipose tissue mass compared with normal individuals [47] and surgical treatment results in an increase in the fractions of adipose tissue in the subcutaneous trunk and the intra-abdominal depots, while the fractions of adipose tissue in peripheral depots decrease [48].

Meals inhibit GH release, whereas fasting conditions amplify the pulsatile pattern of GH secretion [72], indicating that the main impact of GH is in the fasting state. Dose-dependent action by GH on the induction of lipolysis has been demonstrated, with an elevation of circulatory free fatty acids (FFAs) and glycerol and increased lipid oxidation rates [73]. These effects occur despite increased insulin levels, indicating that relatively low doses of GH can overcome the lipogenic actions of insulin. GH stimulates lipolysis by activating hormone-sensitive lipase activity, with a subsequent increase in lipid oxidation [74].

6 Antinatriuretic effects of GH

A sodium-retention effect with the simultaneous expansion of ECW after GH administration was demonstrated in 1952 [75]. Even though it is not the main focus of attention in a doping situation, the anti-natriuretic effect of GH is still of interest.

The sodium and water-retaining effect of GH is complex. To summarise, both GH and IGF-I are capable of causing fluid retention by stimulating Na⁺K⁺ATPase activity in the distal nephron [76]. However, the stimulation of the renin-angiotensin-aldosterone system (RAAS) [77], the down-regulation of atrial natriuretic peptide (ANP) [78] and increased endothelial nitric oxide (NO) function have also been proposed as possible mechanisms [79].

The anti-natriuretic effects of GH explain the reduction in ECW that is found in adult patients with severe GHD and the marked increase in ECW found in acromegalic patients [47, 69]. However, the exact mechanisms behind these effects are unknown. ECW is increased by as much as 25% in untreated acromegaly patients, an increase that normalises after successful treatment [80]. After treatment, the excess ECW correlates with the GH concentrations [80]. Further studies of acromegalic patients suggest a curve-linear dose-response relationship between GH concentrations and excess ECW [80, 81].

7 GH effects on bone

GH has a stimulating effect on both osteoblasts and osteoclasts and is thereby involved in the regulation of bone metabolism [82_84], leading to both bone formation and resorption [85, 86]. The osteoclasts, osteoblasts and components of the bone matrix release several peptides and proteins into the circulation during bone resorption and formation: peptides and proteins that can be used as biochemical markers of bone metabolism.

In adult GHD patients, both normal [87] and reduced [88] serum levels of biochemical markers of bone turnover have been shown. Several studies of hypopituitary patients have shown that GH treatment accelerates bone turnover [85, 89, 90]. Furthermore, it has been shown that biochemical markers of bone formation and bone resorption increase within a few weeks after the initiation of GH treatment in GHD adult patients [91].

8 GH doping among athletes and in the gyms

GH-doping is mostly seen in sporting disciplines that favour strength and explosivity, that is among weightlifters, body builders, football players (American football) and sprinters, but to some extent also in endurance athletes. It is also used by some female athletes, who want to avoid the androgenic side-effects of the AAS. The exact figures of GH-doping among elite athletes and among people training in gyms are not known and there is a lack of studies on the prevalence of GH usage. One study has reported that 5% of male American high-school students used or have used GH as an anabolic agent [3]. However, this study was published in 1992 and whether the results are valid today is not sure. Furthermore, the figures in this study are based on self-reports in a survey and there might be a risk of an overestimation of the usage, since GH might have been considered as an anabolic steroid, resulting in a higher prevalence. GH is often administered 3_10 mg/day, 3_4 times per week when taken alone, and 1_3 mg daily if combined with AAS, thus well above the doses given in cases of adult GH-deficiency. The administration is usually taken in cycles that vary from 6_12 weeks to 6_12 months in length. However, different patterns and doses have been described.

The fact that most abusers probably use GH in combination with AAS and that this combination possibly has an anabolic effect, due to synergistic actions between the two agents, is important to take into account when discussing GH as a doping agent.

9 Doping with GH

This misuse of GH is forbidden for both medical and ethical reasons. Several issues have made the development of a method for detecting GH doping complicated [92]. It is, for example, not possible to distinguish exogenous recombinant GH from endogenous GH in a blood or urine test. Furthermore, GH secretion is influenced by many different factors such as exercise, food intake, sleep and stress [23]. A test method based on the different GH isoforms has been presented and used in the Olympics in Athens and Turin [93, 94]. However, there are some disadvantages with this method. The previous lack of an official method to discover GH doping, might partly explain the strong position GH is thought to enjoy as a doping agent in elite sports.

The GH-2000 project was initiated by the International Olympic Committee (IOC) with the aim of developing a method for detecting GH doping among athletes. The project was funded by the European Union (EU) BioMed2 Research Programme, with additional support from industry and the IOC.

Prior to the start of the GH-2000 project, Wallace et al. [95_98] studied the effects of exercise and supraphysiological GH administration on the GH/IGF-I axis and bone markers in 17 athletic adult males. To summarise, acute exercise increased all the molecular isoforms of GH, with 22 kDa GH constituting the major isoform, with a peak at the end of acute exercise [95]. The proportion of non-22 kDa isoforms increased after exercise, due in part to the slower disappearance rates of these isoforms. With supraphysiological GH administration, these exercised-stimulated endogenous GH isoforms were suppressed for up to four days [96]. Moreover, all the components of the IGF-I ternary complex transiently increased with acute exercise and GH pre-treatment augmented these exercise-induced changes [97]. Furthermore, acute exercise increased the serum concentrations of the bone and collagen markers, bone-specific alkaline phosphatase, carboxy-terminal cross-linked telopeptide of type I collagen (ICTP), carboxy-terminal propeptide of type I procollagen (PICP) and procollagen type III (P-III-P), while osteocalcin was unchanged. GH treatment resulted in an augmented response to exercise of the bone markers PICP and ICTP [98].

A forthcoming method to discover GH abuse will probably necessitate the use of specific markers of the GH/IGF-I axis and bone markers, with the prerequisite that these variables are more sensitive to exogenous GH administration than to exercise. As a result, it will be important closely to study how the levels of these variables are influenced by a maximum exercise test in comparison to rest and by other factors such as gender, age, fitness, type of sport, medication, menstrual status or illness. Furthermore, it will be important to closely study the effects of different types of injuries, for example fractures, on the specific markers.

10 GH and exercise

Physical training has been shown to change circulating levels of GH and, more inconsistently, IGF-I in normal subjects in relation to improvements in oxygen uptake and muscle strength [99_102].

It is well known that acute exercise above a certain intensity is one of the most potent stimulators of GH secretion and the magnitude of the GH response is closely related to the peak intensity of exercise, rather than to total work output [33, 34, 103]. Exercise not only mediates the acute effects on GH secretion. It has been shown that one year of endurance training above the lactate threshold, increases the basal 24-h pulsatile GH release [35]. Interestingly, subjects training below the lactate level did not show any change in the GH release, indicating that the training intensity may be important in regulating the GH-axis as well as fitness. This physiological GH increase in response to exercise and to other stimuli such as hypoglycemia makes it difficult to use measurements of GH itself in blood as a doping marker, as it would be difficult to discriminate a high exercise-derived endogenous GH level from that resulting from exogenous GH.

In addition, bone markers have been shown to respond to exercise and the effects of low-intensity endurance-type activity or brief high-intensity or resistance exercise have shown no acute change [104_106], increased markers [107, 108] or transient decreased markers [106]. Furthermore, studies of high-intensity exercise showed no rise in PICP or ICTP in response to exercise [104, 106, 108]. This could suggest that the duration of exercise is important in the response by bone markers to acute exercise.

11 Variability

Acute exercise above a certain intensity is one of the most potent stimulators of GH secretion. It is well known that elite athletes are able to train at much higher intensities than the normal population and that, during a training season, there are significant differences in training intensity, which might influence GH secretion. Many of the GH-related mediators, binding proteins and markers do not exhibit the same fluctuations as GH and there is a real lack of knowledge about the seasonal stability of markers of the GH/IGF-I axis and circulating bone markers in athletes. A study of seasonal patterns of sleep stages and secretion of cortisol and GH during 24-hour periods in northern Norway revealed no difference in GH secretion as a function of season of the year [109]. Another study reveals no circannual rhythm of plasma GH in pre-pubescent subjects [110].

There is some evidence that biological rhythms of bone turnover over long periods, such as circannual variation, exist [111_118]. Woitge et al. [118] have shown that seasonal variation contributes to the biological variability in bone turnover and needs to be taken into account when interpreting the results of bone marker measurements.

12 Supraphysiological doses of GH: effects on muscles, power, exercise capacity and body composition

GH plays a regulatory role in the maintenance of normal body composition through its well-known anabolic, lipolytic and antinatriuretic actions. These effects are easily demonstrated when GH-substitution therapy is initiated in patients with GHD, reversing muscle atrophy and decreasing central abdominal adiposity and dry skin, signs typically associated with GHD [15, 17, 119]. The anabolic actions of GH include stimulated protein synthesis through the mobilisation of amino-acid transporters, which is reflected in vivo by an increase in the metabolic clearance rate of amino acids. IGF-I also directly stimulates protein synthesis, albeit to a lesser extent than GH, while insulin inhibits protein breakdown [120_124].

Even though GH has been regarded as an effective ergogenic drug among athletes since the 1980s, only a few controlled studies of the effectiveness of GH in relation to physical performance and the effects on body composition in athletes have been performed. A study of 16 young, healthy adults revealed no differences between the GH or placebo group in terms of muscle size, strength or muscle protein synthesis after GH (40 μg/kg/day) or placebo treatment for 12 weeks combined with heavy-resistance exercise. However, fat free mass (FFM) and total body water (TBW) increased in both groups but significantly more among the GH recipients [125]. Another study of 22 power athletes assigned in a double-blind manner to either GH treatment (30 μg/kg/day) or placebo for a period of six weeks revealed no difference between the groups in terms of maximum voluntary strength (biceps or quadriceps) and no change in body weight or body fat, but a remarkable increase in IGF-I was noted [126]. Crist et al. [127] found increased fat free weight and decreased body fat in eight well-trained adults when given 2.67 mg of GH 3 days/week for six weeks. A study of healthy, experienced male weight-lifters before and at the end of 14 days of subcutaneous GH administration (40 μg/kg/day) revealed no increase in the rate of muscle protein synthesis or reduced whole body protein breakdown, metabolic alterations that would promote muscle protein anabolism [128]. In a placebo-controlled study by Healy et al. [129] including 11 endurance-trained athletes, who received supraphysiological GH-doses (67 μg/kg/day) or placebo during four weeks, it was concluded that GH had a net anabolic effect on whole body protein metabolism at rest and during and after exercise. The whole body protein metabolism was measured with a 1-(¹³C) leucine method. Although no muscle power test was performed in the study, it was thus speculated that the acute excess GH administration might have short-term benefits for physical performance. Finally, in two studies from our own group, we found that treatment with supraphysiological doses of GH during one month given to healthy individuals resulted in a decrease in body fat, an increase in ECW but no visible effect in ICW, indicating limited anabolic effect on muscles. Furthermore, no improvements in power output or oxygen uptake in a bicycle exercise test before and after treatment were observed [130, 131].

Some studies performed on elderly men show the same results. A study of healthy, sedentary men with low serum IGF-I levels who followed a 16-week progressive resistance-exercise program (75%_90% max strength, 4 days/week) after random assignment to either a GH (12.5_24.0 μg/kg/day) or placebo group showed that resistance-exercise training improved muscle strength and anabolism, but these improvements were not enhanced when exercise was combined with daily GH administration [132]. Further supportive findings of the lack of effect are found in elderly but not particularly GH-deficient men. Taffee et al. [133, 134] were unable to see any increase in strength, muscle mass or fibre characteristics after GH treatment during a resistance-exercise training programme.

There is a discrepancy between GH's rumour as a strong anabolic agent and the lack of effects seen in the studies performed. There are some different explanations to this that might be taken into consideration. The doses used in the different studies might be too low, even though being supraphysiological and thought to be equal to the doses used by GH abusers. Furthermore, most abusers probably use GH in combination with AAS and this combination might have an anabolic effect, due to synergistic actions between the two agents.

The reduction in fat mass, especially centrally located fat mass, has also been suggested as a desirable GH effect in a doping situation, especially in body-building competitions where a minimum of visible body fat is rewarded.

13 Side-effects of GH

The fear of the well-known side-effects of GH also reduces its use in the gyms. The fluid retention symptoms with swollen hands and feet and carpal tunnel syndroms reduce the exercise performance, thereby limiting the potential of GH as a powerful doping agent.

Long-term use also increases the risk for ordinary acromegalic symptoms. Due to high costs of the "clean" rhGH, some athletes use the cheaper cadaveric GH, with the potential risk of ending up in fatal Creutzfeldt-Jacobs disease.

14 GH doping: current and future aspects

The use of GH as a doping agent is widespread and doping with GH has become an increasing problem in sports and among young people at ordinary gyms during the last 10_15 years [3, 4, 135]. The actual use of GH as a doping agent is not known, but 5% of male high-school students in the USA have been reported to have used it [3]. The GH abusers primarily aim to benefit from the potential anabolic effects of GH, mostly in combination with AAS, in order to increase muscle mass and muscle power. It has also been popular among female athletes, who wish to avoid the androgenic side-effects of anabolic steroids. However, our own interviews with hormone abusers (to be published elsewhere) indicate a more differentiated pattern of GH doping, revealing that its effect is preferentially on muscle volume, and not on muscle strength, thereby making it more popular among body-builders than among weightlifters.

Although previous reports from GH-abusing athletes uniformly describe the positive effects of GH doping on muscle volume and strength [135], the effectiveness of GH as a doping agent has been questioned during the last few years. There is a lack of scientific evidence that GH in supraphysiological doses has additional effects on muscle exercise performance than those obtained from optimized training and diet itself [125, 126, 128, 134, 136_138]. These data have initiated speculations that the reputation of GH as an effective doping agent is highly exaggerated, at least when taken alone. However, in elite athletes, even a small increase in muscle power or exercise performance might make the difference between a gold or silver medal. Furthermore, there is a great deal of counterfeit, inactive GH present on the black market, complicating the real picture of the effectiveness of GH as a doping agent.

There are risks involved with GH abuse. Several potential risk factors have been suggested by observations in GH-supplemented and acromegalic patients, including carpal tunnel syndrome and pitting edema [139], myocardial hypertrophy, and cancer [140, 141]. Because GH needs to be injected there is also a risk of hepatitis and HIV if the abusers share syringes. Finally, cadaver-derived GH is still present on the black market and abusers using this could acquire the fatal Creutzfelt-Jacob disease [4, 142].

Taken as a whole, there is no published evidence of any anabolic effect on muscles or positive effects on physical performance as a result of GH alone or combined with exercise [136_138].

It has been claimed that one main function of GH is its stimulating effect on collagen synthesis and that this might have positive consequences for athletes [143, 144]. In a review, Doessing and Kjaer [143] concluded that supraphysiological doses of GH do not appear to increase the synthesis of myofibrillar protein, but that it is possible that a supraphysiological GH level has an effect on connective tissue. It is known that the GH/IGF-I level is associated with pathological changes in connective tissue in patients with clinical conditions involving a change in GH activity [145_150]. Furthermore, studies of healthy subjects treated with GH have revealed an increase in levels of whole body collagen synthesis [98, 151], indicating that there is a stimulatory effect on connective tissue in normal healthy subjects.

Tendons heal more rapidly in rats treated with GH [152] and anecdotal reports have suggested that GH prevents tendon and muscle ruptures, especially if combined with AAS [153]. By protecting the myotendinous junction regarded as a "weak link in the chain" in athletes training at high intensity and in those with fast-growing muscles due to heavy resistance training and/or AAS abuse, GH could therefore enable training at even higher intensity with shorter recovery periods.

15 Test methods

Regardless of the effectiveness of GH, there is a huge need for the development of a test method to detect GH doping. The problem of developing a reliable method has been described in several papers and work on different approaches is ongoing [2, 92, 154, 155].

15.1 Isoform method

There are currently two main approaches to developing a method for detecting GH doping. The first is an isoform method, developed by the Strasburger group, based on the knowledge that normal, endogenous GH exists in a variety of isoforms. In the pituitary, the most abundant isoform is 22-kDa GH, but other isoforms (non-22-kDa GH) are present in varying amounts [156]. Recombinant GH, in contrast, is solely made up of the 22-kDa isoform. Wallace et al. [96] have shown that all the measured isoforms of GH increased during and peaked at the end of acute exercise, with 22-kDa GH constituting the major isoform in serum during exercise. They also found that the proportion of non-22-kDa isoforms increased after exercise, due in part to slower disappearance rates of 20-kDa GH and perhaps other non-22-kDa GH isoforms. Furthermore, it has been shown that supraphysiological doses of GH in trained adult males suppressed exercise-stimulated endogenous circulating isoforms of GH for up to 4 days and that the clearest separation of treatment groups required the simultaneous presence of high exogenous 22-kDa GH and suppressed 20-kDa or non-22-kDa GH concentrations [95].

Consequently, even if it is not possible directly to distinguish exogenous recombinant GH from endogenous GH in a blood or urine test, the method developed by the Strasburger group, which was used in the Athens Summer Olympics (2004) and in the Turin Winter Olympics (2006), is based on the change in the normal ratio of 22/non-22-kDa GH isoforms after treatment with exogenous GH. One disadvantage with this isoform-based method, however, is that it is only able to detect GH up to 24 h after the last treatment and, therefore, out-of-competition testing will be crucial.

15.2 GH marker-based method

The other approach to detecting GH misuse is to use longer-lasting GH-dependent markers. This approach, used by the GH-2000 group, involves markers that are more sensitive to exogenous GH substitution than exercise-induced GH increase. Several papers from GH-2000 have been published [129, 151, 157_160] and the results from the GH-2000 group have resulted in mathematical, statistical formulae that have been presented as a potential test for detecting GH doping [160]. This method has then been further validated in a study stating that the test proposed by the GH-2000 study group can be used to detect subjects receiving exogenous GH [161].

However, some questions still remain before an accurate, reliable method is found. The test proposed by the GH-2000 study group requires further evaluation and discussion before it can be finally accepted as a doping test. Any test to detect drug abuse in sport must minimize the risk of a false positive result.

16 Conclusion

The role of GH as an effective anabolic muscle doping agent, when taken alone, is questioned. GH might be effective, in lower doses, when taken together with AAS. GH doping does not seem to have any positive effect on cardiac performance. Fluid retention and other acromegalic side-effects further reduce its use. However, GH seems to stimulate collagen synthesis, which might have a positive, protective effect on ruptures of muscles and tendons, and allow harder training with a shorter recovery period, thus explaining its ongoing use in doping areas. It cannot be excluded that this effect on collagen synthesis might be useful in a few years in clinical practice for the treatment of muscle and tendon ruptures.

References

1 Macintyre JG. Growth hormone and athletes. Sports Med 1987; 4: 129_42.

2 Sonksen PH. Insulin, growth hormone and sport. J Endocrinol 2001; 170: 13_25.

3 Rickert VI, Pawlak-Morello C, Sheppard V, Jay MS. Human growth hormone: a new substance of abuse among adolescents? Clin Pediatr (Phila) 1992; 31: 723_6.

4 Ehrnborg C, Bengtsson BA, Rosen T. Growth hormone abuse. Baillieres Best Pract Res Clin Endocrinol Metab 2000; 14: 71_7.

5 Evans HM, Long JA. The effect of the anterior lobe administered intraperitoneally upon growth maturity, and oestreus cycles of the rat. Anat Rec 1921; 62_3.

6 Li CH, Papkoff H. Preparation and properties of growth hormone from human and monkey pituitary glands. Science 1956; 124: 1293_4.

7 Li CH, Dixon JS. Human pituitary growth hormone. 32. The primary structure of the hormone: revision. Arch Biochem Biophys 1971; 146: 233_6.

8 Niall HD. Revised primary structure for human growth hormone. Nature New Biol 1971; 230: 90_1.

9 Lewis UJ, Dunn JT, Bonewald LF, Seavey BK, Vanderlaan WP. A naturally occurring structural variant of human growth hormone. J Biol Chem 1978; 253: 2679_87.

10 Baumann G, MacCart JG, Amburn K. The molecular nature of circulating growth hormone in normal and acromegalic man: evidence for a principal and minor monomeric forms. J Clin Endocrinol Metab 1983; 5: 946_52.

11 Goeddel DV, Heyneker HL, Hozumi T, Arentzen R, Itakura K, Yansura DG, et al. Direct expression in Escherichia coli of a DNA sequence coding for human growth hormone. Nature 1979; 281: 544_8.

12 Raben MS. Treatment of a pituitary dwarf with human growth hormone. J Clin Endocrinol Metab 1958; 18: 901_3.

13 Raben MS. Growth hormone. 2. Clinical use of human growth hormone. N Engl J Med 1962; 266: 82_6.

14 Cuneo RC, Salomon F, McGauley GA, Sonksen PH. The growth hormone deficiency syndrome in adults. Clin Endocrinol (Oxf) 1992; 37: 387_97.

15 Jorgensen JO, Pedersen SA, Thuesen L, Jorgensen J, Ingemann-Hansen T, Skakkebaek NE, et al. Beneficial effects of growth hormone treatment in GH-deficient adults. Lancet 1989; 1: 1221_5.

16 Rosen T, Bengtsson BA. Premature mortality due to cardiovascular disease in hypopituitarism. Lancet 1990; 336: 285_8.

17 Salomon F, Cuneo RC, Hesp R, Sonksen PH. The effects of treatment with recombinant human growth hormone on body composition and metabolism in adults with growth hormone deficiency. N Engl J Med 1989; 28: 1797_803.

18 Eden S. The secretory pattern of growth hormone. An experimental study in the rat. Acta Physiol Scand Suppl 1978; 458: 1_54.

19 Jansson JO, Eden S, Isaksson O. Sexual dimorphism in the control of growth hormone secretion. Endocr Rev 1985; 6: 128_50.

20 Vance ML, Kaiser DL, Evans WS, Furlanetto R, Vale W, Rivier J, et al. Pulsatile growth hormone secretion in normal man during a continuous 24-hour infusion of human growth hormone releasing factor (1-40). Evidence for intermittent somatostatin secretion. J Clin Invest 1985; 75: 1584_90.

21 Winer LM, Shaw MA, Baumann G. Basal plasma growth hormone levels in man: new evidence for rhythmicity of growth hormone secretion. J Clin Endocrinol Metab 1990; 70: 1678_86.

22 van den Berg G, Veldhuis JD, Frolich M, Roelfsema F. An amplitude-specific divergence in the pulsatile mode of growth hormone (GH) secretion underlies the gender difference in mean GH concentrations in men and premenopausal women. J Clin Endocrinol Metab 1996; 81: 2460_7.

23 Giustina A, Veldhuis JD. Pathophysiology of the neuroregulation of growth hormone secretion in experimental animals and the human. Endocr Rev 1998; 19: 717_97.

24 Stolar MW, Baumann G. Secretory patterns of growth hormone during basal periods in man. Metabolism 1986; 9: 883_8.

25 Massa G, Igout A, Rombauts L, Frankenne F, Vanderschueren-Lodeweyckx M. Effect of oestrogen status on serum levels of growth hormone-binding protein and insulin-like growth factor-I in non-pregnant and pregnant women. Clin Endocrinol (Oxf) 1993; 39: 569_75.

26 Kelly JJ, Rajkovic IA, O'Sullivan AJ, Sernia C, Ho KK. Effects of different oral oestrogen formulations on insulin-like growth factor-I, growth hormone and growth hormone binding protein in post-menopausal women. Clin Endocrinol (Oxf) 1993; 39: 561_7.

27 Dawson-Hughes B, Stern D, Goldman J, Reichlin S. Regulation of growth hormone and somatomedin-C secretion in postmenopausal women: effect of physiological estrogen replacement. J Clin Endocrinol Metab 1986; 2: 424_32.

28 Veldhuis JD, Liem AY, South S, Weltman A, Weltman J, Clemmons DA, et al. Differential impact of age, sex steroid hormones, and obesity on basal versus pulsatile growth hormone secretion in men as assessed in an ultrasensitive chemiluminescence assay. J Clin Endocrinol Metab 1995; 80: 3209_22.

29 Ho KY, Evans WS, Blizzard RM, Veldhuis JD, Merriam GR, Samojlik E, et al. Effects of sex and age on the 24-hour profile of growth hormone secretion in man: importance of endogenous estradiol concentrations. J Clin Endocrinol Metab 1987; 64: 51_8.

30 Liu L, Merriam GR, Sherins RJ. Chronic sex steroid exposure increases mean plasma growth hormone concentration and pulse amplitude in men with isolated hypogonadotropic hypogonadism. J Clin Endocrinol Metab 1987; 64: 651_6.

31 Iranmanesh A, Lizarralde G, Veldhuis JD. Age and relative adiposity are specific negative determinants of the frequency and amplitude of growth hormone (GH) secretory bursts and the half-life of endogenous GH in healthy men. J Clin Endocrinol Metab 1991; 73: 1081_8.

32 Takahashi Y, Kipnis DM, Daughaday WH. Growth hormone secretion during sleep. J Clin Invest 1968; 47: 2079_90.

33 Kjaer M, Bangsbo J, Lortie G, Galbo H. Hormonal response to exercise in humans: influence of hypoxia and physical training. Am J Physiol 1988; 254: R197_203.

34 Felsing NE, Brasel JA, Cooper DM. Effect of low and high intensity exercise on circulating growth hormone in men. J Clin Endocrinol Metab 1992; 75: 157_62.

35 Weltman A, Weltman JY, Schurrer R, Evans WS, Veldhuis JD, Rogol AD. Endurance training amplifies the pulsatile release of growth hormone: effects of training intensity. J Appl Physiol 1992; 72: 2188_96.

36 Hartman ML, Veldhuis JD, Johnson ML, Lee MM, Alberti KG, Samojlik E, et al. Augmented growth hormone (GH) secretory burst frequency and amplitude mediate enhanced GH secretion during a two-day fast in normal men. J Clin Endocrinol Metab 1992; 74: 757_65.

37 Masuda A, Shibasaki T, Nakahara M, Imaki T, Kiyosawa Y, Jibiki K, et al. The effect of glucose on growth hormone (GH)-releasing hormone-mediated GH secretion in man. J Clin Endocrinol Metab 1985; 3: 523_6.

38 Imaki T, Shibasaki T, Shizume K, Masuda A, Hotta M, Kiyosawa Y, et al. The effect of free fatty acids on growth hormone (GH)-releasing hormone-mediated GH secretion in man. J Clin Endocrinol Metab 1985; 60: 290_3.

39 Parker ML, Hammond JM, Daughaday WH. The arginine provocative test: an aid in the diagnosis of hyposomatotropism. J Clin Endocrinol Metab 1967; 27: 1129_36.

40 Bratusch-Marrain P, Waldhausl W. The influence of amino acids and somatostatin on prolactin and growth hormone release in man. Acta Endocrinol (Copenh) 1979; 90: 403_8.

41 Iranmanesh A, Lizarralde G, Johnson ML, Veldhuis JD. Nature of altered growth hormone secretion in hyperthyroidism. J Clin Endocrinol Metab 1991; 72: 108_15.

42 Williams T, Maxon H, Thorner MO, Frohman LA. Blunted growth hormone (GH) response to GH-releasing hormone in hypothyroidism resolves in the euthyroid state. J Clin Endocrinol Metab 1985; 61: 454_6.

43 Wehrenberg WB, Janowski BA, Piering AW, Culler F, Jones KL. Glucocorticoids: potent inhibitors and stimulators of growth hormone secretion. Endocrinology 1990; 126: 3200_3.

44 Muller EE. Neural control of somatotropic function. Physiol Rev 1987; 67: 962_1053.

45 Kelijman M, Frohman LA. The role of the cholinergic pathway in growth hormone feedback. J Clin Endocrinol Metab 1991; 72: 1081_7.

46 Ikkos D, Luft R, Gemzell CA. The effect of human growth hormone in man. Lancet 1958; 1: 720_1.

47 Bengtsson BA, Brummer RJ, Eden S, Bosaeus I. Body composition in acromegaly. Clin Endocrinol (Oxf) 1989; 30: 121_30.

48 Brummer RJ, Lonn L, Kvist H, Grangard U, Bengtsson BA, Sjostrom L. Adipose tissue and muscle volume determination by computed tomography in acromegaly, before and 1 year after adenomectomy. Eur J Clin Invest 1993; 23: 199_205.

49 Brumback RA, Barr CE. Myopathy in acromegaly. A case study. Pathol Res Pract 1983; 1: 41_6.

50 Nagulesparen M, Trickey R, Davies MJ, Jenkins JS. Muscle changes in acromegaly. BMJ 1976; 2: 914_5.

51 Rooyackers OE, Nair KS. Hormonal regulation of human muscle protein metabolism. Ann Rev Nutr 1997; 17: 457_85.

52 Cuneo RC, Salomon F, Wiles CM, Hesp R, Sonksen PH. Growth hormone treatment in growth hormone-deficient adults. I. Effects on muscle mass and strength. J Appl Physiol 1991; 70: 688_94.

53 Jorgensen JO, Pedersen SA, Thuesen L, Jorgensen J, Moller J, Muller J, et al. Long-term growth hormone treatment in growth hormone deficient adults. Acta Endocrinol (Copenh) 1991; 125: 449_53.

54 Beshyah SA, Freemantle C, Shahi M, Anyaoku V, Merson S, Lynch S, et al. Replacement treatment with biosynthetic human growth hormone in growth hormone-deficient hypopituitary adults. Clin Endocrinol (Oxf) 1995; 42: 73_84.

55 Johannsson G, Grimby G, Sunnerhagen KS, Bengtsson BA. Two years of growth hormone (GH) treatment increase isometric and isokinetic muscle strength in GH-deficient adults. J Clin Endocrinol Metab 1997; 82: 2877_84.

56 Wallymahmed ME, Foy P, Shaw D, Hutcheon R, Edwards RH, MacFarlane IA. Quality of life, body composition and muscle strength in adult growth hormone deficiency: the influence of growth hormone replacement therapy for up to 3 years. Clin Endocrinol (Oxf) 1997; 47: 439_46.

57 Sartorio A, Narici MV. Growth hormone (GH) treatment in GH-deficient adults: effects on muscle size, strength and neural activation. Clin Physiol 1994; 14: 527_37.

58 Cuneo RC, Salomon F, Wiles CM, Sonksen PH. Skeletal muscle performance in adults with growth hormone deficiency. Horm Res 1990; 33 (Suppl 4): 55_60.

59 Cuneo RC, Salomon F, Wiles CM, Hesp R, Sonksen PH. Growth hormone treatment in growth hormone-deficient adults. II. Effects on exercise performance. J Appl Physiol 1991; 70: 695_700.

60 Whitehead HM, Boreham C, McIlrath EM, Sheridan B, Kennedy L, Atkinson AB, et al. Growth hormone treatment of adults with growth hormone deficiency: results of a 13-month placebo controlled cross-over study. Clin Endocrinol (Oxf) 1992; 36: 45_52.

61 Christiansen JS, Jorgensen JO, Pedersen SA, Muller J, Jorgensen J, Moller J, et al. GH-replacement therapy in adults. Horm Res 1991; 36 (Suppl 1): 66_72.

62 Johannsson G, Bengtsson BA, Andersson B, Isgaard J, Caidahl K. Long-term cardiovascular effects of growth hormone treatment in GH-deficient adults. Preliminary data in a small group of patients. Clin Endocrinol (Oxf) 1996; 45: 305_14.

63 Rutherford OM, Jones DA, Round JM, Preece MA. Changes in skeletal muscle after discontinuation of growth hormone treatment in young adults with hypopituitarism. Acta Paediatr Scand Suppl 1989; 356: 61_3; discussion 64, 73_4.

64 Degerblad M, Almkvist O, Grunditz R, Hall K, Kaijser L, Knutsson E, et al. Physical and psychological capabilities during substitution therapy with recombinant growth hormone in adults with growth hormone deficiency. Acta Endocrinol (Copenh) 1990; 123: 185_93.

65 Ayling CM, Moreland BH, Zanelli JM, Schulster D. Human growth hormone treatment of hypophysectomized rats increases the proportion of type-1 fibres in skeletal muscle. J Endocrinol 1989; 123: 429_35.

66 Cuneo RC, Salomon F, Wiles CM, Round JM, Jones D, Hesp R, et al. Histology of skeletal muscle in adults with GH deficiency: comparison with normal muscle and response to GH treatment. Horm Res 1992; 37: 23_8.

67 Whitehead HM, Gilliland JS, Allen IV, Hadden DR. Growth hormone treatment in adults with growth hormone deficiency: effect on muscle fibre size and proportions. Acta Paediatr Scand Suppl 1989; 356: 65_7.

68 Raben MS, Hollenberg CH. Effect of growth hormone on plasma fatty acids. J Clin Invest 1959; 38: 484_8.

69 Rosen T, Bosaeus I, Tolli J, Lindstedt G, Bengtsson BA. Increased body fat mass and decreased extracellular fluid volume in adults with growth hormone deficiency. Clin Endocrinol (Oxf) 1993; 38: 63_71.

70 De Boer H, Blok GJ, Voerman HJ, De Vries PM, van der Veen EA. Body composition in adult growth hormone-deficient men, assessed by anthropometry and bioimpedance analysis. J Clin Endocrinol Metab 1992; 75: 833_7.

71 Lonn L, Johansson G, Sjostrom L, Kvist H, Oden A, Bengtsson BA. Body composition and tissue distributions in growth hormone deficient adults before and after growth hormone treatment. Obes Res 1996; 4: 45_54.

72 Ho KY, Veldhuis JD, Johnson ML, Furlanetto R, Evans WS, Alberti KG, et al. Fasting enhances growth hormone secretion and amplifies the complex rhythms of growth hormone secretion in man. J Clin Invest 1988; 81: 968_75.

73 Copeland KC, Nair KS. Acute growth hormone effects on amino acid and lipid metabolism. J Clin Endocrinol Metab 1994; 78: 1040_7.

74 Dietz J, Schwartz J. Growth hormone alters lipolysis and hormone-sensitive lipase activity in 3T3-F442A adipocytes. Metabolism 1991; 40: 800_6.

75 Whitney JE, Bennett LL, Li CH. Reduction of urinary sodium and potassium produced by hypophyseal growth hormone in normal female rats. Proc Soc Exp Biol Med 1952; 79: 584_7.

76 Johannsson G, Sverrisdottir YB, Ellegard L, Lundberg PA, Herlitz H. GH increases extracellular volume by stimulating sodium reabsorption in the distal nephron and preventing pressure natriuresis. J Clin Endocrinol Metab 2002; 87: 1743_9.

77 Moller J, Moller N, Frandsen E, Wolthers T, Jorgensen JO, Christiansen JS. Blockade of the renin-angiotensin-aldosterone system prevents growth hormone-induced fluid retention in humans. Am J Physiol 1997; 272: E803_8.

78 Moller J, Jorgensen JO, Moller N, Hansen KW, Pedersen EB, Christiansen JS. Expansion of extracellular volume and suppression of atrial natriuretic peptide after growth hormone administration in normal man. J Clin Endocrinol Metab 1991; 72: 768_72.

79 Boger RH, Skamira C, Bode-Boger SM, Brabant G, von zur Muhlen A, Frolich JC. Nitric oxide may mediate the hemodynamic effects of recombinant growth hormone in patients with acquired growth hormone deficiency. A double-blind, placebo-controlled study. J Clin Invest 1996; 98: 2706_13.

80 Bengtsson BA, Brummer RJ, Eden S, Bosaeus I, Lindstedt G. Body composition in acromegaly: the effect of treatment. Clin Endocrinol (Oxf) 1989; 31: 481_90.

81 Davies DL, Beastall GH, Connell JM, Fraser R, McCruden D, Teasdale GM. Body composition, blood pressure and the renin-angiotensin system in acromegaly before and after treatment. J Hypertens Suppl 1985; 3: S413_5.

82 Maor G, Hochberg Z, von der Mark K, Heinegard D, Silbermann M. Human growth hormone enhances chondrogenesis and osteogenesis in a tissue culture system of chondroprogenitor cells. Endocrinology 1989; 125: 1239_45.

83 Nishiyama K, Sugimoto T, Kaji H, Kanatani M, Kobayashi T, Chihara K. Stimulatory effect of growth hormone on bone resorption and osteoclast differentiation. Endocrinology 1996; 137: 35_41.

84 Slootweg MC, van Buul-Offers SC, Herrmann-Erlee MP, van der Meer JM, Duursma SA. Growth hormone is mitogenic for fetal mouse osteoblasts but not for undifferentiated bone cells. J Endocrinol 1988; 116: R11_3.

85 Kann P, Piepkorn B, Schehler B, Andreas J, Lotz J, Prellwitz W, et al. Effect of long-term treatment with GH on bone metabolism, bone mineral density and bone elasticity in GH-deficient adults. Clin Endocrinol (Oxf) 1998; 48: 561_8.

86 Ohlsson C, Bengtsson BA, Isaksson OG, Andreassen TT, Slootweg MC. Growth hormone and bone. Endocr Rev 1998; 19: 55_79.

87 Schlemmer A, Johansen JS, Pedersen SA, Jorgensen JO, Hassager C, Christiansen C. The effect of growth hormone (GH) therapy on urinary pyridinoline cross-links in GH-deficient adults. Clin Endocrinol (Oxf) 1991; 35: 471_6.

88 Amato G, Carella C, Fazio S, La Montagna G, Cittadini A, Sabatini D, et al. Body composition, bone metabolism, and heart structure and function in growth hormone (GH)-deficient adults before and after GH replacement therapy at low doses. J Clin Endocrinol Metab 1993; 77: 1671_6.

89 Degerblad M, Bengtsson BA, Bramnert M, Johnell O, Manhem P, Rosen T, et al. Reduced bone mineral density in adults with growth hormone (GH) deficiency: increased bone turnover during 12 months of GH substitution therapy. Eur J Endocrinol 1995; 133: 180_8.

90 Vandeweghe M, Taelman P, Kaufman JM. Short and long-term effects of growth hormone treatment on bone turnover and bone mineral content in adult growth hormone-deficient males. Clin Endocrinol (Oxf) 1993; 39: 409_15.

91 Johannsson G, Rosen T, Bosaeus I, Sjostrom L, Bengtsson BA. Two years of growth hormone (GH) treatment increases bone mineral content and density in hypopituitary patients with adult-onset GH deficiency. J Clin Endocrinol Metab 1996; 81: 2865_73.

92 McHugh CM, Park RT, Sonksen PH, Holt RI. Challenges in detecting the abuse of growth hormone in sport. Clin Chem 2005; 51: 1587_93.

93 Bidlingmaier M, Wu Z, Strasburger CJ. Test method: GH. Baillieres Best Pract Res Clin Endocrinol Metab 2000; 14: 99_109.

94 Wu Z, Bidlingmaier M, Dall R, Strasburger CJ. Detection of doping with human growth hormone. Lancet 1999; 353: 895.

95 Wallace JD, Cuneo RC, Bidlingmaier M, Lundberg PA, Carlsson L, Boguszewski CL, et al. Changes in non-22-kilodalton (kDa) isoforms of growth hormone (GH) after administration of 22-kDa recombinant human GH in trained adult males. J Clin Endocrinol Metab 2001; 86: 1731_7.

96 Wallace JD, Cuneo RC, Bidlingmaier M, Lundberg PA, Carlsson L, Boguszewski CL, et al. The response of molecular isoforms of growth hormone to acute exercise in trained adult males. J Clin Endocrinol Metab 2001; 1: 200_6.

97 Wallace JD, Cuneo RC, Baxter R, Orskov H, Keay N, Pentecost C, et al. Responses of the growth hormone (GH) and insulin-like growth factor axis to exercise, GH administration, and GH withdrawal in trained adult males: a potential test for GH abuse in sport. J Clin Endocrinol Metab 1999; 84: 3591_601.

98 Wallace JD, Cuneo RC, Lundberg PA, Rosen T, Jorgensen JO, Longobardi S, et al. Responses of markers of bone and collagen turnover to exercise, growth hormone (GH) administration, and GH withdrawal in trained adult males. J Clin Endocrinol Metab 2000; 85: 124_33.

99 Eliakim A, Brasel JA, Mohan S, Barstow TJ, Berman N, Cooper DM. Physical fitness, endurance training, and the growth hormone-insulin-like growth factor I system in adolescent females. J Clin Endocrinol Metab 1996; 81: 3986_92.

100 Eliakim A, Brasel JA, Barstow TJ, Mohan S, Cooper DM. Peak oxygen uptake, muscle volume, and the growth hormone-insulin-like growth factor-I axis in adolescent males. Med Sci Sports Exerc 1998; 30: 512_7.

101 Kelly PJ, Eisman JA, Stuart MC, Pocock NA, Sambrook PN, Gwinn TH. Somatomedin-C, physical fitness, and bone density. J Clin Endocrinol Metab 1990; 70: 718_23.

102 Poehlman ET, Copeland KC. Influence of physical activity on insulin-like growth factor-I in healthy younger and older men. J Clin Endocrinol Metab 1990; 71: 1468_73.

103 Kraemer WJ, Gordon SE, Fleck SJ, Marchitelli LJ, Mello R, Dziados JE, et al. Endogenous anabolic hormonal and growth factor responses to heavy resistance exercise in males and females. Int J Sports Med 1991; 12: 228_35.

104 Kristoffersson A, Hultdin J, Holmlund I, Thorsen K, Lorentzon R. Effects of short-term maximal work on plasma calcium, parathyroid hormone, osteocalcin and biochemical markers of collagen metabolism. Int J Sports Med 1995; 16: 145_9.

105 Welsh L, Rutherford OM, James I, Crowley C, Comer M, Wolman R. The acute effects of exercise on bone turnover. Int J Sports Med 1997; 18: 247_51.

106 Virtanen P, Viitasalo JT, Vuori J, Vaananen K, Takala TE. Effect of concentric exercise on serum muscle and collagen markers. J Appl Physiol 1993; 75: 1272_7.

107 Nishiyama S, Tomoeda S, Ohta T, Higuchi A, Matsuda I. Differences in basal and postexercise osteocalcin levels in athletic and nonathletic humans. Calcif Tissue Int 1988; 43: 150_4.

108 Salvesen H, Johansson AG, Foxdal P, Wide L, Piehl-Aulin K, Ljunghall S. Intact serum parathyroid hormone levels increase during running exercise in well-trained men. Calcif Tissue Int 1994; 54: 256_61.

109 Weitzman ED, deGraaf AS, Sassin JF, Hansen T, Godtlibsen OB, Perlow M, et al. Seasonal patterns of sleep stages and secretion of cortisol and growth hormone during 24 hour periods in northern Norway. Acta Endocrinol (Copenh) 1975; 78: 65_76.

110 Bellastella A, Criscuolo T, Mango A, Perrone L, Sinisi AA, Faggiano M. Circannual rhythms of plasma growth hormone, thyrotropin and thyroid hormones in prepuberty. Clin Endocrinol (Oxf) 1984; 20: 531_7.

111 Chapuy MC, Schott AM, Garnero P, Hans D, Delmas PD, Meunier PJ. Healthy elderly French women living at home have secondary hyperparathyroidism and high bone turnover in winter. EPIDOS Study Group. J Clin Endocrinol Metab 1996; 81: 1129_33.

112 Douglas AS, Miller MH, Reid DM, Hutchison JD, Porter RW, Robins SP. Seasonal differences in biochemical parameters of bone remodelling. J Clin Path 1996; 49: 284_9.

113 Hyldstrup L, McNair P, Jensen GF, Transbol I. Seasonal variations in indices of bone formation precede appropriate bone mineral changes in normal men. Bone 1986; 7: 167_70.

114 Overgaard K, Nilas L, Johansen JS, Christiansen C. Lack of seasonal variation in bone mass and biochemical estimates of bone turnover. Bone 1988; 9: 285_8.

115 Storm D, Eslin R, Porter ES, Musgrave K, Vereault D, Patton C, et al. Calcium supplementation prevents seasonal bone loss and changes in biochemical markers of bone turnover in elderly New England women: a randomized placebo-controlled trial. J Clin Endocrinol Metab 1998; 83: 3817_25.

116 Thomsen K, Eriksen EF, Jorgensen JC, Charles P, Mosekilde L. Seasonal variation of serum bone GLA protein. Scand J Clin Lab Invest 1989; 49: 605_11.

117 Vanderschueren D, Gevers G, Dequeker J, Geusens P, Nijs J, Devos P, et al. Seasonal variation in bone metabolism in young healthy subjects. Calcif Tissue Int 1991; 49: 84_9.

118 Woitge HW, Scheidt-Nave C, Kissling C, Leidig-Bruckner G, Meyer K, Grauer A, et al. Seasonal variation of biochemical indexes of bone turnover: results of a population-based study. J Clin Endocrinol Metab 1998; 83: 68_75.

119 Bengtsson BA, Eden S, Lonn L, Kvist H, Stokland A, Lindstedt G, et al. Treatment of adults with growth hormone (GH) deficiency with recombinant human GH. J Clin Endocrinol Metab 1993; 76: 309_17.

120 Bowes SB, Umpleby M, Cummings MH, Jackson NC, Carroll PV, Lowy C, et al. The effect of recombinant human growth hormone on glucose and leucine metabolism in Cushing's syndrome. J Clin Endocrinol Metab 1997; 82: 243_6.

121 Inoue Y, Copeland EM, Souba WW. Growth hormone enhances amino acid uptake by the human small intestine. Ann Surg 1994; 219: 715_22.

122 Umpleby AM, Boroujerdi MA, Brown PM, Carson ER, Sonksen PH. The effect of metabolic control on leucine metabolism in type 1 (insulin-dependent) diabetic patients. Diabetologia 1986; 29: 131_41.

123 Tessari P, Trevisan R, Inchiostro S, Biolo G, Nosadini R, De Kreutzenberg SV, et al. Dose-response curves of effects of insulin on leucine kinetics in humans. Am J Physiol 1986; 251: E334_42.

124 Russell-Jones DL, Umpleby AM, Hennessy TR, Bowes SB, Shojaee-Moradie F, Hopkins KD, et al. Use of a leucine clamp to demonstrate that IGF-I actively stimulates protein synthesis in normal humans. Am J Physiol 1994; 267: E591_8.

125 Yarasheski KE, Campbell JA, Smith K, Rennie MJ, Holloszy JO, Bier DM. Effect of growth hormone and resistance exercise on muscle growth in young men. Am J Physiol 1992; 262: E261_7.

126 Deyssig R, Frisch H, Blum WF, Waldhor T. Effect of growth hormone treatment on hormonal parameters, body composition and strength in athletes. Acta Endocrinol (Copenh) 1993; 128: 313_8.

127 Crist DM, Peake GT, Egan PA, Waters DL. Body composition response to exogenous GH during training in highly conditioned adults. J Applied Physiol 1988; 65: 579_84.

128 Yarasheski KE, Zachweija JJ, Angelopoulos TJ, Bier DM. Short-term growth hormone treatment does not increase muscle protein synthesis in experienced weight lifters. J Applied Physiol 1993; 74: 3073_6.

129 Healy ML, Gibney J, Russell-Jones DL, Pentecost C, Croos P, Sonksen PH, et al. High dose growth hormone exerts an anabolic effect at rest and during exercise in endurance-trained athletes. J Clin Endocrinol Metab 2003; 88: 5221_6.

130 Berggren A, Ehrnborg C, Rosen T, Ellegard L, Bengtsson BA, Caidahl K. Short-term administration of supraphysiological recombinant human growth hormone (GH) does not increase maximum endurance exercise capacity in healthy, active young men and women with normal GH-insulin-like growth factor I axes. J Clin Endocrinol Metab 2005; 6: 3268_73.

131 Ehrnborg C, Ellegard L, Bosaeus I, Bengtsson BA, Rosen T. Supraphysiological growth hormone: less fat, more extracellular fluid but uncertain effects on muscles in healthy, active young adults. Clin Endocrinol (Oxf) 2005; 62: 449_57.

132 Yarasheski KE, Zachwieja JJ, Campbell JA, Bier DM. Effect of growth hormone and resistance exercise on muscle growth and strength in older men. Am J Physiol 1995; 268: E268_76.

133 Taaffe DR, Jin IH, Vu TH, Hoffman AR, Marcus R. Lack of effect of recombinant human growth hormone (GH) on muscle morphology and GH-insulin-like growth factor expression in resistance-trained elderly men. J Clin Endocrinol Metab 1996; 81: 421_5.

134 Taaffe DR, Pruitt L, Reim J, Hintz RL, Butterfield G, Hoffman AR, et al. Effect of recombinant human growth hormone on the muscle strength response to resistance exercise in elderly men. J Clin Endocrinol Metab 1994; 79: 1361_6.

135 Cowart VS. Human growth hormone: the latest ergogenic aid? Phys Sports Med 1988; 16: 175_85.

136 Frisch H. Growth hormone and body composition in athletes. J Endocrinol Invest 1999; 22: 106_9.

137 Rennie MJ. Claims for the anabolic effects of growth hormone: a case of the emperor's new clothes? Br J Sports Med 2003; 37: 100_5.

138 Weber MM. Effects of growth hormone on skeletal muscle. Horm Res 2002; 58 (Suppl 3): 43_8.

139 Lange KH, Isaksson F, Rasmussen MH, Juul A, Bulow J, Kjaer M. GH administration and discontinuation in healthy elderly men: effects on body composition, GH-related serum markers, resting heart rate and resting oxygen uptake. Clin Endocrinol (Oxf) 2001; 55: 77_86.

140 Cittadini A, Berggren A, Longobardi S, Ehrnborg C, Napoli R, Rosen T, et al. Supraphysiological doses of GH induce rapid changes in cardiac morphology and function. J Clin Endocrinol Metab 2002; 87: 1654_9.

141 Colao A, Ferone D, Marzullo P, Lombardi G. Systemic complications of acromegaly: epidemiology, pathogenesis, and management. Endocr Rev 2004; 25: 102_52.

142 Deyssig R, Frisch H. Self-administration of cadaveric growth hormone in power athletes. Lancet 1993; 341: 768_9.

143 Doessing S, Kjaer M. Growth hormone and connective tissue in exercise. Scand J Med Sci Sports 2005; 15: 202_10.

144 Rosen T. Supraphysiological doses of growth hormone: effects on muscles and collagen in healthy active young adults. Horm Res 2006; 66: 98_104.

145 Colao A, Marzullo P, Vallone G, Marino V, Annecchino M, Ferone D, et al. Reversibility of joint thickening in acromegalic patients: an ultrasonography study. J Clin Endocrinol Metab 1998; 83: 2121_5.

146 Colao A, Marzullo P, Vallone G, Giaccio A, Ferone D, Rossi E, et al. Ultrasonographic evidence of joint thickening reversibility in acromegalic patients treated with lanreotide for 12 months. Clin Endocrinol (Oxf) 1999; 51: 611_8.

147 Colao A, Di Somma C, Pivonello R, Loche S, Aimaretti G, Cerbone G, et al. Bone loss is correlated to the severity of growth hormone deficiency in adult patients with hypopituitarism. J Clin Endocrinol Metab 1999; 84: 1919_24.

148 Baroncelli GI, Bertelloni S, Ceccarelli C, Cupelli D, Saggese G. Dynamics of bone turnover in children with GH deficiency treated with GH until final height. Eur J Endocrinol 2000; 142: 549_56.

149 Lange M, Thulesen J, Feldt-Rasmussen U, Skakkebaek NE, Vahl N, Jorgensen JO, et al. Skin morphological changes in growth hormone deficiency and acromegaly. Eur J Endocrinol 2001; 145: 147_53.

150 Scarpa R, De Brasi D, Pivonello R, Marzullo P, Manguso F, Sodano A, et al. Acromegalic axial arthropathy: a clinical case-control study. J Clin Endocrinol Metab 2004; 89: 598_603.

151 Longobardi S, Keay N, Ehrnborg C, Cittadini A, Rosen T, Dall R, et al. Growth hormone (GH) effects on bone and collagen turnover in healthy adults and its potential as a marker of GH abuse in sports: a double blind, placebo-controlled study. The GH-2000 Study Group. J Clin Endocrinol Metab 2000; 85: 1505_12.

152 Kurtz CA, Loebig TG, Anderson DD, DeMeo PJ, Campbell PG. Insulin-like growth factor I accelerates functional recovery from Achilles tendon injury in a rat model. Am J Sports Med 1999; 27: 363_9.

153 Verducci T, Yeager D, Dohrmann G, Llosa LF, Munson L. Totally juiced. Sports Illustrated 2002; 96: 34_7.

154 Rigamonti AE, Cella SG, Marazzi N, Di Luigi L, Sartorio A, Muller EE. Growth hormone abuse: methods of detection. Trends Endocrinol Metab 2005; 16: 160_6.

155 Saugy M, Robinson N, Saudan C, Baume N, Avois L, Mangin P. Human growth hormone doping in sport. Br J Sports Med 2006; 40 (Suppl 1): i35_9.

156 Boguszewski CL, Hynsjo L, Johannsson G, Bengtsson BA, Carlsson LM. 22-kD growth hormone exclusion assay: a new approach to measurement of non-22-kD growth hormone isoforms in human blood. Eur J Endocrinol 1996; 135: 573_82.

157 Dall R, Longobardi S, Ehrnborg C, Keay N, Rosen T, Jorgensen JO, et al. The effect of four weeks of supraphysiological growth hormone administration on the insulin-like growth factor axis in women and men. GH-2000 Study Group. J Clin Endocrinol Metab 2000; 85: 4193_200.

158 Giannoulis MG, Boroujerdi MA, Powrie J, Dall R, Napoli R, Ehrnborg C, et al. Gender differences in growth hormone response to exercise before and after rhGH administration and the effect of rhGH on the hormone profile of fit normal adults. Clin Endocrinol (Oxf) 2005; 62: 315_22.

159 Healy ML, Dall R, Gibney J, Bassett E, Ehrnborg C, Pentecost C, et al. Toward the development of a test for growth hormone (GH) abuse: a study of extreme physiological ranges of GH-dependent markers in 813 elite athletes in the postcompetition setting. J Clin Endocrinol Metab 2005; 90: 641_9.

160 Powrie JK, Bassett EE, Rosen T, Jorgensen JO, Napoli R, Sacca L, et al. Detection of growth hormone abuse in sport. Growth Horm IGF Res 2007; 17: 220_6.

161 Erotokritou-Mulligan I, Bassett EE, Kniess A, Sonksen PH, Holt RI. Validation of the growth hormone (GH)-dependent marker method of detecting GH abuse in sport through the use of independent data sets. Growth Horm IGF Res 2007; 17: 416_23.