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Growth Hormone, Exercise, and Athletic Performance: A Continued Evolution of Complexity

Kraemer, William J.1,2; Dunn-Lewis, Courtenay1; Comstock, Brett A.1; Thomas, Gwendolyn A.1; Clark, James E.1; Nindl, Bradley C.3

Current Sports Medicine Reports: July-August 2010 - Volume 9 - Issue 4 - p 242-252
doi: 10.1249/JSR.0b013e3181e976df
Nutrition & Ergogenic Aids: Section Articles
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Growth hormone (hGH) presents pleiotropic effects in many tissues encompassing a diverse range of physiological actions. Its complexity as a family of hormones with different isoforms and different somatotroph molecular functions continues to challenge the status quo of our understanding of its release, function, and signaling. Owing to the fact that the majority of the literature has viewed hGH from the perspective of the primary 22 kD monomer, further investigation is needed as to the influence and biological activity of other aggregate and splice variant isoforms that are released into circulation. Its role over the life span and with supplementation yields equivocal results with more study needed. Testing for the use of hGH has progressed, and the first positive test was recently documented. Understanding of pituitary function and physiology will remain complex until the use of a broader range of analytical techniques, including assays, becomes mainstream.

1Human Performance Laboratory, Department of Kinesiology, 2Department of Physiology and Neurobiology, University of Connecticut, Storrs, CT; 3Military Performance Division, The U.S. Army Research Institute of Environmental Medicine, Natick, MA

Address for correspondence: William J. Kraemer, Ph.D., FACSM, Professor, Human Performance Laboratory, Department of Kinesiology Unit 1110, 2095 Hillside Road, University of Connecticut, Storrs, CT 06269 (E-mail: William.Kraemer@uconn.edu).

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INTRODUCTION

Within the scope of physiological function, the extensive and interconnected nature of hormonal relationships, the complex web of molecular signaling pathways, and the vast families of genetic interactions have made the study of a single hormone's role daunting at best. Deciphering the results and physiological meaning of one study often can take years, require new technology or analytical methods, and require additional data. While simplistic dismissals for the relative physiological importance of hormonal roles are more palatable, they can obviate the evolution of our understanding and serve to minimize the appreciation for each molecule's role in our complex and integrated biology. A complete picture only emerges with complete and comprehensive analysis.

The complexity of the endocrine system is best understood through the endocrine model. The endocrine model is based upon the basic tenet that perturbations within the physiological system lead to a cascade of events to restore equilibrium. The upper regulatory elements provide an integrated stimulus to all downstream physiological systems and induce adaptations (Fig. 1). Environmental conditions, age, sex, psychological status, nutritional status or intake, and the specific nature of an exercise stimulus are the primary drivers of this process. The integration of adaptations to any exercise or training program is a confluence of these daily signals over time. The dramatic demands on metabolism, cellular repair, and tissue remodeling have made exercise a popular context in which to study the endocrine model and hormones such as human growth hormone (hGH). Of utmost importance, however, is the proper use of appropriate analytical techniques and context in doing so.

Figure 1

Figure 1

In this article, we examine a small portion of the evolving paradigm of hGH with regard to exercise and in the context of analytical techniques used to study it. A biological description of the regulation and action of hGH will be discussed. While many essential aspects of 22 kD hGH regulation and its action are understood and will be detailed, the larger physiological context for growth hormone is, in some senses, still in its infancy. We will discuss how the use of the radioimmunoassay (in most research since the 1960s) has focused research predominantly on the 22 kD monomer of hGH. A host of other biochemical detection systems that detect the rich and diverse nature and function of other hGH isoforms, including bioassays, have been neglected for some time. As a result, the potential importance of all isoforms of hGH to both aerobic and resistance exercise has not been developed fully and will be discussed. As a final example of our need for improved analytical techniques, the modern misconceptions and abuse of exogenous recombinant growth hormone (rGH) in multiple populations will be detailed briefly. Throughout this discussion, the importance of utilizing a broader set of analytical techniques to encompass the complex nature of all growth hormone isoforms will be emphasized.

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SEQUENCE OF PHYSIOLOGICAL EVENTS IN GROWTH HORMONE RELEASE AND REGULATION

The factors involved in the stimulation of 22 kD growth hormone monomer secretion are well known (Fig. 2). A set of upstream regulatory elements surrounding a specific exercise protocol (68) stimulates the release of growth hormone releasing hormone (GHRH) from the hypothalamus into the hypophyseal portal system. GHRH binds with its receptors in the anterior pituitary, leading to the synthesis and release of hGH from their site of production in the somatotrophs of the pituitary into the circulation. Correlations in the response of exercise-induced plasma norepinephrine and hGH levels could suggest that catecholamines drive hGH responses to exercise (82). This basic GHRH model of hGH synthesis and release follows a general endocrine model.

Figure 2

Figure 2

At the same time, the stimulation of hGH may be complicated with an additional pathway. Ghrelin is associated with stimulation of hunger and is produced in the human stomach, the epsilon cells of the pancreas, and the arcuate nucleus of the hypothalamus (53). The discovery of ghrelin as an endogenous ligand for the hGH secretagogue receptor (GHSR) in the anterior pituitary gland demonstrated its ability to stimulate hGH release at the anterior pituitary (34). Although subsequent research has shown that ghrelin directly stimulates growth hormone-releasing neurons in the arcuate nucleus, ghrelin does not appear to be a major factor in the exercise-induced hGH response (8). Nonetheless, it is of interest that hGH may be stimulated both directly by GHRH or indirectly (through increased GHRH or increased release) by ghrelin.

The regulatory restriction of hGH production and release from anterior pituitary somatotrophs is accomplished through both inhibition and negative feedback. The primary inhibitor of hGH secretion is somatostatin [also referred to as growth hormone inhibiting hormone (GHIH) or somatotropin release inhibiting factor (SRIF)] released from the periventricular nucleus. Negative feedback also arises from circulating hGH and insulin-like growth factor-1 (IGF-1, the peripheral hormone for hGH). Thus blood concentrations of hGH are controlled by somatostatin and through negative feedback by hGH and IGF-1.

Like the stimulation pathways, the concept of regulatory control also is continuing to evolve based on inherent differences in the release patterns and contents of somatotrophs (26,28) (Fig. 3). Band 1 somatotrophs show low granularity and contain small molecular weight hGH molecules (e.g., 22 kD). Band 2 cells have much greater granularity and a different cell surface charge, and they contain binding proteins and aggregates of hGH (e.g., 44 kD, dimers, homo- and heterodimeric, tri- to pentameric, and growth hormone binding protein-bound monomeric). An afferent feedback loop from skeletal muscle to the hypothalamic-pituitary axis influences the amount of so-called bioactive, large-molecular weight hGH released, indicating that band 2 cells predominantly were affected (51). Interestingly, reductions in band 2 content of pituitary cells also have been noted with exposure to microgravity with little change in band 1 contents (28). It therefore seems that a regulatory system for hGH may exist within the pituitary gland.

Figure 3

Figure 3

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PHYSIOLOGICAL ACTIONS OF GROWTH HORMONE

While the most dramatic visible action of growth hormone occurs with the development of linear bone growth in childhood, hGH plays several important roles during exercise in adulthood. Growth hormone exhibits pleiotropic effects in many tissues encompassing such diverse physiological actions as growth, reproductive function, immune function, osmoregulation, endocrine function, and neural function. The various forms (variants, aggregates) of hGH circulate to target tissues and interact with receptors to create additional hormonal signal events to the cells, such as peripheral production of IGF-1. The classic "somatomedin hypothesis" states that hGH exerts its primary effects by stimulating IGF-1 secretion from the liver which then mediates somatogenic effects in peripheral tissues, although it appears that direct effects of hGH occur independent of IGF-1 upregulation (67).

The pulsatile release of growth hormone from the anterior pituitary is an important aspect of growth hormone action. Growth hormone pulses are released episodically throughout the day and the majority is released nocturnally. Original work by Isgaard et al. (30) and Maiter et al. (49) in rodent models reported that growth hormone administered in a pulsatile manner had a much more potent effect on both liver IGF-1 and bone IGF-1 than continuous infusion (even when the continuous infusion was five times greater). Conversely, recent work in humans has suggested that continuous growth hormone administration in humans is responsible primarily for augmenting hepatic and muscle IGF-1 while pulsatile growth hormone administration augments adipose tissue lipolysis. Hence, the pulsatile release of growth hormone and pattern of exposure to peripheral tissues determines growth-promoting and metabolic effects in a pattern- and tissue-specific fashion (75).

One key action of hGH is its impact on the metabolism of substrates such as protein. Growth hormone is thought to increase net muscle protein synthesis indirectly by facilitating amino acid transport and availability via both endocrine and locally produced IGF-1. Direct effects on muscle synthesis, unrelated to protein metabolism, also are under investigation (11). This anabolic impact of hGH helps to facilitate the body's exercise responses, but the impact on substrates is not restricted to protein alone.

While often ignored, one of the most potent effects of hGH is its lipolytic actions. hGH increases lipolysis by several mechanisms, including 1) potentiating the sensitivity (via activation of β-adrenergic receptors) of adipose tissue to other lipolytic hormones (i.e., catecholamines); 2) stimulating the primary fat breakdown enzyme (i.e., hormone-sensitive lipase); and 3) inhibiting fat storage enzymes (i.e., fatty acid synthase and acetyl-CoA carboxylase). Through these actions, hGH "diverts" nutrients away from adipose tissue and toward other peripheral tissues. The mobilization of fatty acids during exercise is thought to accommodate the increase in energy demands of the body.

Growth hormone's impact on carbohydrate metabolism causes a net elevation of circulating glucose concentrations (in excess, this may have a diabetogenic effect). It increases gluconeogenesis at the liver, decreases glucose uptake into adipose tissue and skeletal muscle, and decreases utilization of glucose. Recent studies suggest that this may occur not by changes in receptor levels at the plasma membrane (as previously believed) but by impairing the ability of the GLUT-4 receptor itself to transport glucose (65).

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GROWTH HORMONE DETECTION SYSTEMS

The responses to and adaptations of circulatory concentrations of growth hormone (for example, hGH) to exercise and training have been studied extensively over the past 45 yr. Interestingly, as described by Hymer et al. (27), the view of this hormone has been documented primarily through the use of various immunoassays (e.g., EIA, RIA) based on their reduced costs, assay duration, complexity, and/or sensitivity in comparison to other assays (Table 2). One weakness of the immunoassays, however, is that they essentially quantify the 22 kD monomer, the fundamental 191 amino acid product of the hGH1 gene (9). While it is possible that the 22 kD monomer is the primary hGH molecule interacting with target receptors, a paradox exists as the 22 kD does not interact with all target receptors (27). Unfortunately, these immunoassays often have been used to the exclusion of bioassays, which measure concentrations of other hGH isoforms.

TABLE 2

TABLE 2

The differences in the assays used have a significant impact on both research findings and their interpretation. Different assays signal different target end points of interaction and therefore produce divergent values for both the magnitude and response of hGH to exercise or physiological stimuli (13,14,29). Once released from the anterior pituitary, the measured concentration of hGH in the blood via aggregate detection (i.e., tibial line rat assay or bioactive hGH) can be more than a 1000-fold greater than the concentration of 22 kD hGH that is measured by immunoassay (13,29) (Table 1). The principle of growth hormone pulsatility has been shown to exist in immunoreactive samples but it has not been reported with other assays measuring the splice variants, aggregate, or bioactive forms. It is clear that the complexity of pituitary function and the larger context of hGH cannot and should not be reduced to the study of the 22 kD monomer alone.

TABLE 1

TABLE 1

One prominent bioassay that provides important insights into hGH responses is the rat tibial line bioassay. This assay was developed by Greenspan et al. (18) in 1949 and has been used in our laboratory with exercise (18,39,51). Growth hormone concentration differences are determined by the width of the uncalcified growth plate in hyposectomized rats injected with human samples (13,27,29) (Fig. 4). Papkoff and Li (60) indicate that rat weight gain bioassays were the primary hGH detection assay up to the 1960s despite various disadvantages. Bioassays still allow for more information as to the specific receptor end point interactions (as 22 kD has low reactivity in the assay) (14,27). Other bioassays (IM-9 lymphocyte cell line, 3T3-F422A adipocyte) have been used to quantify hGH, but not all in exercise research (14,27). The rat tibial line assay, therefore, is one important alternative to current immunoassay technology.

Figure 4

Figure 4

Several other types of assays have been used in exercise research. The immunofunctional assay has been used to determine the hGH molecules in which both epitopes were available for binding and dimerization (73,74). It has been shown that such signals of hGH are expected to be lower in concentration and also correlate highly to the monoclonal radioimmunoassay in response to exercise (58). The NB-2 node lymphoma assay is a lactogenic assay typically used for determination of large molecular weight prolactin concentrations, but hGH concentrations also can be determined (14). From our laboratory, it appears to have a different concentration signal than the immunoassay response patterns (34a). Similarly, the use of mass spectrometry and a 2D-PAGE method have been used to identify some of the hGH variants as part of a potential strategy to determine ergogenic use of hGH in athletics (33). Each of these assays might very well offer a further dimension to exercise research examining hGH response and adaptations.

As a final point, our picture remains incomplete, in part, because not every assay has been used with exercise. One set of salient examples is related to the available cell proliferation assays that have been used to measure hGH signal in a blood sample. Roswall and colleagues (63) developed the assay using cells from a mouse myeloid leukemia cell line that is transfected with a full length receptor and then incubated with test samples. The 3H-thymidine uptake into DNA was then used as the measure of hGH activity. Wada et al. (79) also used a similar approach to measuring hGH activity of certain variants using Ba/F3-hGHR cells with the intent to gain understanding of the direct biological effects and hGH impact. However, to date these cellular proliferation approaches have not been used to examine exercise responses and may present an opportunity for new discoveries in this area.

Our understanding of hGH has evolved over the past several years based on the results we have been able to obtain from the different types of hGH assays. Over 100 molecular weight forms and two growth hormone binding proteins now have been detected in the blood (3,4,27,45). Growth hormone now appears to be more of a super-family of hGH isoforms and variants (56,59). The complexity of the anterior pituitary cell system, growth hormone isoforms, and targeted function has been described by Hymer et al. (27) and reflects the very challenging and dynamic nature for understanding anterior pituitary function. Investigations that take advantage of the wide variety of assays available will provide a clearer and richer understanding of the physiological complexity of hGH. These assays may have novel stories and may expand the scope of our understanding of hGH behavior in the human body.

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EXERCISE RESPONSES

The potent impact of exercise on the release of hGH into circulation was first documented by Hunter et al. (25) in 1965. Over time, a paradigm for the responses of hGH to both aerobic and resistance exercise has developed, including some appreciation for the influence of the upper regulatory elements. However, studies into hGH forms other than 22 kD have suggested that there is more to the picture that has yet to be uncovered. Taken as a whole, the research that has been done with growth hormone's response and exercise clearly suggests that our knowledge of hGH has been restricted by our preference for specific types of assays. This pattern will be examined for both aerobic and resistance exercise.

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Aerobic Exercise

Research into hGH and aerobic exercise historically has been an attempt to better understand the metabolic roles of hGH in energy production and its responses to exercise intensities (%V˙O2max). Substantial research in the area, particularly recent investigations by Weltman and Veldhuis, has provided a strong understanding of the interaction between acute and chronic aerobic exercise and hGH release, as well as the influences of gender, age, and adiposity (85). At the same time, immunoassays have been the preferred analytical tool for much of our current knowledge in this area.

The acute relationship of hGH to aerobic exercise is well-characterized. The relationship of growth hormone to aerobic exercise time is direct and positive; the rate of growth hormone release increases at or close to the onset of aerobic exercise and peaks at or close to its cessation (7,61,76,83,84,86). Growth hormone concentration also increases linearly (in a dose-response relationship) with aerobic exercise intensity (61,62,71,85). Upper regulatory elements do impact the stereotyped response. Kanaley et al. (32) demonstrated that time of day did not influence the acute exercise response of hGH, but response magnitudes and pulse characteristics potentially were confounded by nutrition, sleep, prior exercise patterns, and body composition. Elevated adiposity suppresses hGH, including the response of hGH to aerobic exercise (31,85). Older individuals have blunted responses of growth hormone to the acute exercise, which may start at middle age and progress (71,85,87). In addition, intensity, sex, adrenal hormonal influences, nutritional intakes, and metabolic factors all impact this exercise response (71).

With chronic training, reductions in the hGH response may be seen. Within 3 wk of chronic aerobic training, there appears to be a down-regulation of exercise-induced hGH release (21,85). When performed at the level of the lactate threshold or above, chronic training may increase total 24-h hGH on both days with and days without training (84). There appears to be no chronic training impact in older individuals or in obese individuals in the absence of weight loss (85). While down-regulation does appear to occur, it may not be at a significant level in all individuals or may require a higher stimulus for some.

Although the relationship of hGH to aerobic exercise seems clear, other research in this area helps to reiterate the importance of bioassays. With submaximal cycle exercise (65% V˙O2max), much of the plasma contains non-22 kD isoforms during recovery (78). Rubin et al. (63) assayed plasma at 90% and 100% V˙O2max and showed that higher plasma concentrations of hGH were observed if the plasma was reduced with glutathione to break down bonds within oligomeres. Such data again point to the importance of alternative analysis techniques to develop the larger context of response patterns.

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Resistance Exercise

The potential role of hGH in the repair and remodeling of skeletal muscle and connective tissue has lead to the study of hGH responses to acute resistance exercise. Research interest began in the 1980s, and more attention was given to this modality in the 1990s and onward. Before the understanding of the importance of the acute program variables (or domains), research into hGH responses did not take such factors (such as gender, metabolic demands, rest period lengths, intensity, amount of tissue activation, volume of exercise, and choice of exercise) into account. However, once the importance of these domains was understood, the acute exercise response patterns of growth hormone in the subsequent studies began to account for the acute program domains. All of these factors contributed to the response patterns of hGH when examining it from an immunoassay signal detection viewpoint (38,41).

An early research study suggested the importance of acute program variables (or domains) such as intensity (68). In an initial study by Vanhelder et al. (78), seven sets of leg lifts at 85% of the subject's 7 repetition maximum increased growth hormone concentrations in the blood. When the load was reduced to one third of the intensity, no changes occurred. The concept that not all resistance training protocols were the same started to gain momentum at this point. Soon, the impact of upstream regulatory elements in the workout design on the hGH response was demonstrated. In 1990, Kraemer and colleagues (37) demonstrated for the first time that rest period lengths, intensity, and volume of work were all potentially modifying factors when examining hGH responses with heavy resistance exercise in men and later in women (35) using immunoassays. Shorter rest periods stimulated dramatically greater hGH responses to the resistance exercise protocol and, as has been shown previously, women during the follicular phase of the menstrual cycle had higher resting growth hormone concentrations then men (36). The potential influence of metabolic signals (with shorter rest and greater total work) may well have influenced the hGH responses observed in these initial studies of the acute resistance exercise response patterns (15,16). Häkkinen et al. (19) eloquently validated the influence of total work by demonstrating dramatically higher growth hormone concentrations with 10 sets at 10 RM compared with 20 sets of 100% of 1 RM. Nindl et al. (57) showed that the pulse characteristics were affected by acute heavy resistance exercise at different times of the day. Different burst and output amounts followed exercise versus control resting conditions over 16 h (including sleep). The complexity of the physiological response of hGH to the acute program variables, therefore, clearly was demonstrated.

In 2000, McCall et al. (51) published data suggesting that hGH responses might be different if viewed from the perspective of a bioassay, opening the door to more complexity in both the regulatory roles and potential influences of higher molecular weight isoforms in biological actions. In 2001, Hymer et al. (29) investigated acute exercise in women during the early follicular phase of the menstrual cycle. He and colleagues showed that the rat tibial line bioassay resulted in much higher values of hGH pre- and postexercise (six sets of 10 RM in Smith-machine squat exercise). However, the values were unchanged by exercise stress, even when fractionated into different molecular weight fractions. Conversely, increases in concentrations were observed concomitantly with the use of immunofunctional assays and the monoclonal RIA, which was higher than the polyclonal RIA. When total samples were reduced with glutathione, an increase in hGH signal was detected, again suggesting that disulfide-linked hormone aggregates/fragments are released into the circulation after the exercise regimen used in this study. Such data also suggested that the exercise response differed by assay perspective, reflecting the potential inherent complexity of anterior pituitary somatotroph function. Bioassays therefore have helped to show that the response of hGH to exercise is not limited to 22 kD.

Resistance training and its impact on hGH adaptations remain in a state of uncertainty because of a number of upstream regulatory elements. Aging can impact the magnitude of the response, but results are equivocal as to the changes in response to a resistance exercise stressor (typically, no changes have been reflected in resting values with training) (41,50). One study examining the role of 6 months of resistance training in women who were tested in the early follicular phase of the menstrual cycle demonstrated even more complexity as hGH. Results differed by training time, acute exercise response, program used, assay type, and assay molecular weight fraction. This showed that typical single radioimmunoassay of the 22 kD molecules could not provide a complete understanding of adaptation; even monoclonal and polyclonal assays showed some differentiation (39). Interestingly, the whole body heavy periodized group was the only group to demonstrate increases after 6 months of training in their postexercise value for the high molecular weight fraction of growth hormone as measured by the rat tibial line bioassay. Another study examining these same assays showed strikingly few differences between assays with the use of oral contraceptives (40). It is clear that a variety of variables related to hGH adaptations are still under investigation.

The significance of aggregates is another area of particular interest in this line of research. One study often lost in this area of study was the finding that stronger women have been shown to have higher concentrations of growth hormone as measured by the rat tibial line bioassay but not RIA. This may suggest that physical characteristics of stronger women in the untrained state are reflected in this aggregate concentration (42). This could relate back to the sophisticated processing of growth hormone in the somatotrophs and the regulation of band 1 and band 2 type cells. Furthermore, the regulation of the aggregate formation on the somatotrophs (potentially by heat shock proteins and chaperone proteins) may be critical in this molecular organization. Investigations into hGH aggregates that look into their significance should be examined with proper assay techniques.

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EXOGENOUS GROWTH HORMONE FOR ANTIAGING AND PERFORMANCE

Overview

Prior to the 1980s, exogenous growth hormone was limited to either human and monkey sources. The synthesis of rGH in the 1980s dramatically increased its availability and allowed for the treatment of a broader range of medical conditions. In concert with the medical implications, this development created an availability of rGH that would allow for its use in otherwise healthy individuals. While availability was no longer a concern, legal considerations became more relevant. Unlike most medications in America, hGH only is approved by the U.S. Food and Drug Administration for a discrete set of medical conditions and cannot be prescribed for off-label. The prescription of rGH for any other purpose is a felony punishable by up to 5 yr in prison, fines, or both. Despite this classification, off-label use of rGH appears to continue in individuals concerned with improvements in body composition, sports performance, or the effects of aging.

The off-label use of growth hormone appears to be prominent in two primary populations: those interested in antiaging and those interested in athletic performance. After 40 yr of age, individuals may expect to lose 5% of their muscle mass every 10 yr. While controversial, sarcopenia, or the decline in muscle mass with age, is a natural extension of frailty and is associated with osteoporosis. While the lay public may not be acutely aware of such factors, they may be concerned with age-associated frailty, decreased independence, and increased rates of illness, disability, and death. The potential for a decrease in these risks may be the cause of the use of rGH in individuals. On the other end of the spectrum, for many elite athletes, any potential to improve performance is worthwhile, regardless of consequences (68). In both cases, the attraction of rGH to healthy populations is based on its perceived ability to improve muscle mass, decrease fat stores, and improve energy regulation.

While still an area of controversy, rGH has been shown to increase uptake and decrease protein oxidation at both the local and central level in healthy individuals (64). Clinically, rGH increases total body water, decreases body fat, and may increase fat-free mass in hGH-deficient patients (70). However, whether those or any results in the literature are relevant in practice with healthy populations is a matter of some debate. In some research, increases in fat-free mass could be attributed to fluid retention and increases in connective tissue (rather than increases in muscle) (22).

One issue with the use of rGH is that, once again, it chiefly is available in the 22 kD form. Given the diverse functions of hGH in the human body, the differential patterns of release seen in acute exercise, and the presence of a broad variety of isoforms within the human body, it is likely that the isoforms do have physiological significance in terms of their function. Growth hormone isoforms do not interact in the same way with all growth hormone receptors. Indeed, the 22 kD form does not interact strongly with the rat tibial line assay (15). In addition, athletes are unlikely to take growth hormone in isolation; much of the time, the concurrent use of anabolic steroids or, at minimum, nutritional supplements limits the applicability of research to athletes (23).

The safety of rGH is another issue of debate. Preliminary studies in mice produced no changes in tumor incidence, longetivity, or survival (70). However, the transfer of any results to humans cannot be generalized. Importantly, the impact of the use of one isoform of growth hormone, the 22 kD version as is marketed with rGH, to the exclusion of all other isoforms cannot be anticipated. In the meta-analysis conducted by Liu et al., side effects in the short-term protocols were common and appeared to be related to salt and water balance at the kidney. These effects included increased sweating, edema, joint pain, and carpal tunnel syndrome. Importantly, the elevation of plasma glucose in response to growth hormone is a concern in older populations in terms of the risk for diabetes (47).

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Efficacy

The antiaging effects of rGH are not understood well. In the animal model, a reduction in the activity of the growth hormone/IGF-1 axis results in an increase in lifespan. This trend also has been seen with caloric restriction, which is associated with a decrease in IGF-1 (although this is neither considered to be safe nor advised in humans). At the same time, some animal studies have shown an increase in mortality when the growth hormone/IGF-1 system was compromised (70,54). Liu et al. (47) reviewed the use of rGH in older populations. In older adults, short-term hGH use may increase lean body mass; however, improvements in strength or quality of life are not observed widely. The combined use of testosterone and hGH only increased lean body mass (6), and rGH and resistance exercise did not increase muscle mass beyond resistance exercise alone (77). However, resistance exercise alone may be an effective method of addressing sarcopenia. In sum, the research on the use of rGH to combat aging is unclear and, in some ways, contradictory at this stage (54).

In 2008, Liu et al. completed a systematic review of 44 articles from 27 unique studies related to rGH for performance enhancement. The 303 total subjects had an average age of 27 yr (46). On average, the daily dose administered was 5 to 10 times higher than the dose administered clinically (36 mcg·kg−1·d−1). The review unfortunately was limited by the available body of knowledge. Recreational athletes completed short-term studies that did not reflect the usage of elite athletes in terms of time, dosage, and use of other substances. Particularly in the areas of strength and exercise capacity, there is little research available. Additionally, the studies were not sensitive enough to detect the minute differences that distinguish first place from last in elite competition.

Despite the limitations in the available research, the results of Liu et al. (46) indicated an increase in lean mass (2.1 kg), no significant change in body mass, and a statistically insignificant decrease in fat mass (−0.9 kg). In two studies evaluating strength changes, no significant differences were seen despite interventions of 42 and 84 d. Results from work on metabolism indicated a slight resting preference for fat over carbohydrate. Respiratory exchange ratios did not change, but higher plasma free fatty acid and glycerol concentrations combined with trends toward higher lactate levels indicated that hGH supplementation may have influenced lipolysis. Measures such as an increased heart rate with exercise could suggest a decrease in work capacity.

A small number of studies do point to a more promising impact from rGH. A carefully controlled 6-d study of 0.058 IU·kg−1·d−1 rGH on regular but abstinent steroid users demonstrated strong results (17). Total protein, albumin, and free thyroxine, and thyroid-stimulating hormone significantly decreased while IGF-1 increased. Fat-free mass, peak V˙O2, peak power, and strength increased and fat mass decreased. A very recent 6-wk study examining recreational athletes also showed positive effects of rGH as an ergogenic aid. Meinhardt et al. (52) demonstrated that growth hormone (2 mg·d−1 subcutaneously) in men and women, growth hormone and testosterone (250 mg·wk−1 intramuscularly), or combined treatments in men resulted in expected hormonal changes as well as increased sprint performance, decreased fat, and increased lean tissue mass. However, these speed changes were not maintained after 6 wk of non-use. Thus, while the majority of research shows little effect of rGH on performance, the concept also is evolving as more research is done. From their article, the authors stated:

"First, body cell mass at baseline was correlated with all measures of physical performance. Second, growth hormone significantly reduced fat mass, increased lean body mass through an increase in extracellular water, and increased body cell mass when given with testosterone. Third, growth hormone led to statistically significant improvements in sprint capacity that were not maintained after a 6-week washout period in a pooled group of men and women, and the improvements were greater when growth hormone was coadministered with testosterone to men. Finally, changes in body cell mass did not correlate with improvement in sprint capacity, except when growth hormone was coadministered with testosterone."

One effect worth discussion is the potential for hGH and IGF impact on connective tissue. The synthesis of collagen, an aspect of connective tissue associated with its strength, is stimulated by exercise (44). Populations deficient in hGH exhibit lower connective tissue deposition and show signs of recovery in this aspect with treatment. The converse also is true in conditions with excess hGH such as acromegaly (10,11). A number of animal and human studies demonstrate strong relationships between plasma growth hormone (or IGF), the effectors of collagen synthesis at both the local and systemic level, and both tendon and muscle synthesis of collagen (10,11). IGF-1 promotes collagen synthesis, including increased cell proliferation and protein synthesis, in the rabbit tendon (1) and increases recovery from anterior cruciate ligament (ACL) injuries in the rat (43). Hasen et al. (20) documented decreased bioavailability of IGF-1 and inhibited collagen synthesis in response to exercise with women taking oral contraceptives (20). Healthy trials in humans have shown increased whole-body collagen synthesis and/or turnover (81), including a dose-dependent response (48) and higher synthesis in an rGH-supplemented exercise group than a placebo group in healthy active men (80); 50 mcg of rGH increased collagen I mRNA and tendon collagen protein synthesis 3.9-fold in younger healthy individuals, although this was not impacted by moderate exercise (10). However, conclusions to be drawn from these data remain speculative (11).

In sum, the available research connecting rGH to antiaging or performance-enhancing effects remains limited. Doessing's laboratory has continued research into effects on connective tissue, but warned in the 2005 review that without a measure of connective tissue breakdown, results must be viewed with scrutiny (11). The conclusion of Liu et al. (46) was that any claim on performance enhancement was premature and that its use was associated with common side effects. In concert, a statement by the Growth Hormone Research Society after an international conference on the use of growth hormone in older adults suggested that until more research was completed, the use of rGH in older populations could not be recommended (70). The complexity of growth hormone and its use as a therapeutic drug or ergogenic aid requires further study and validation of the salient hypotheses (23,54).

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DETECTING rGH USE IN ATHLETES

rGH presents a unique testing challenge on a number of levels. One factor motivating the use of rGH in athletes is that to date no test for it has been proven legally (70). Urine testing, often the preferred method of sampling in athletes, is not possible with growth hormone. Growth hormone is present at levels of less than 0.1% in urine and is highly variable in terms of excretion (5). Blood measures must be used when testing for growth hormone; this necessitates the invasiveness of blood testing and its political ramifications. However, the tests currently available are limited.

It is extremely difficult to accurately determine rGH doping (12,23,24,55,65,66,69,71,72). While many substances may be detected simply by supra-physiological levels in the blood, the pulsatile release of growth hormone, its fast half-life in the body, and its natural fluctuations due to sleep, stress, and exercise, prevent the use of any testing modality based on blood concentration alone. Circadian and ultradian rhythms of secretion must be tracked to obtain accurate measurements because hGH is secreted in a pulsatile manner; the peak for a normal individual may appear to be no different than concentrations observed in an active rGH user. As hGH is released in response to various stresses (exercise and psychosomatic stress included), testing at pre- and postcompetition potentially may confound test results due to stress-induced increases in hGH.

In addition to the fundamental challenges with growth hormone, the blood testing itself presents a number of challenges. rGH is comprised exclusively of the 22 kD isoform and has an amino acid sequence identical to it. There are no differentiating aspects to the molecular sequence of synthetic growth hormone from the 22 kD isoform that would otherwise allow for its detection (5). Growth hormone, regardless of the source, is metabolized rapidly; even individuals who are doping may have undetectable levels of rGH. The fitness level of the athlete, gender, ethnicity, age, nutrition status, and recent athletic performance all may alter the results of the growth hormone doping tests. Further, circulating binding proteins and cross-reactivity may exacerbate confounding factors (24).

The push for testing has resulted recently in positive test results in a British rugby league player and a desire of major sporting leagues in the United States (Major League Baseball, National Football League) to institute rules for testing players for rGH doping. The test for rGH use, as detailed by Saugy et al. (65) and Stow et al. (71), is a two-part analysis of growth hormone and growth hormone-related peptide biomarkers obtained from a blood sample. The first aspect is referred to as the isoform approach and the second is referred to as the marker approach.

The isoform approach is a test of the ratio of the 22 kD isoform (the isoform used in rGH) with other isoforms. Any dramatic increase in the ratio of the 22 kD isoform to the other isoforms could suggest the use of rGH in body. The use of a recombinant assay to detect the 22 kD isoform with a pituitary assay to measure other isoforms helps to determine this ratio. This test cannot account for pituitary-derived growth hormone (which would have natural ratios of all isoforms) or the natural variations in isoforms that occur with developing adolescents. Another weakness of this test is that a short window of time - probably at about 24 h after injection - is required for accurate results. This suggests that unannounced off-season tests would be more effective.

The second part of the test, the marker approach, relies on the factors released in response to growth hormone in the human body that may be detected up to 84 d after rGH administration. Growth hormone-related biomarkers within the same sample, IGF-1 and P-III-P, are analyzed. The results are compared with normative values for gender, ethnicity, and athletic training status (55). To our knowledge, while neither this test nor the isoform approach have been validated in a court of law, some suggest that a complimentary approach utilizing both could present some degree of validity (5). Despite their weaknesses, they have been validated by the World Anti-Doping Agency (WADA) and used in multiple Olympic Games (5).

It has been suggested that pharmaceutical companies add a marker to rGH to reduce the difficulty of doping tests. However, such an approach would not help with the identification of so-called gene doping. This potential for the addition of genes to the human body to produce these hormones, a possibility already tested in animals (2), would present a further and incredible obstacle for testing.

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CONCLUSION

To date, no testing modality has been challenged legally and established to address the use of rGH in sport. The use and abuse of rGH in both athletic and older populations continues despite side effects, the relative paucity of data suggesting benefits, and the unknown impact of the long-term use of the 22 kD form of growth hormone to the exclusion of other isoforms in healthy populations. While we have examined the evolving paradigm of hGH with regards to exercise, the apparent misunderstandings concerning the true physiological role of hGH in the community suggest that this examination will not be complete without further investigations into the varied nature of hGH isoforms, variants, and aggregates. It is the opinion of the authors that the continued development of the larger physiological context and the nature of growth hormone can and should be achieved through the use of a broader range of assay technologies used to partial out the evolving complexities of pituitary function from its somatroph origin, transport, binding, and targeting of hGH in the human body with exercise and cellular and tissue adaptations.

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