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Original Studies |
Medical Department M (Endocrinology and Diabetes) and Center for Clinical Pharmacology, Aarhus University Hospital, Kommunehospitalet, Aarhus, Denmark
Address all correspondence and requests for reprints to: Nina Vahl, M.D., Medical Department M, Aarhus Kommunehospital, DK-8000 C Aarhus, Denmark.
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Abstract |
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 Top Abstract IntroductionSubjects and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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It is known that GH secretion decreases with age (7, 8) and that females secrete more GH than males (9). Furthermore, aging is associated with adiposity, which is known to blunt GH secretion (8, 10). The cause-effect relationship is not know in detail, but we recently showed that intraabdominal fat mass is the major negative determinant of stimulated GH secretion in healthy nonobese adults (11).
The half-life of endogenous GH is reduced with age (7) and obesity (10), whereas discrepancy exists with regard to the existence of sex differences. Little is known about possible age- and sex-related differences in the MCR and apparent distribution volume (Vd) of GH.
Finally, experience with GH substitution in GH-deficient adults unequivocally show that the dose requirements are substantially reduced with age (7, 12). Whether this reflects age-associated changes in GH responsiveness or clearance is unknown.
The present study was designed to investigate the significance of age, sex, and body composition on the pharmacokinetics and metabolic effects of a single exogenous GH pulse of near physiological size in normal adults.
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Subjects and Methods |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Thirty-six healthy adults divided into two age groups were examined: a young group, aged 27–34 yr (mean, 29.6 yr), and an older group, aged 47–59 yr (mean, 51.0 yr). The eight premenopausal female subjects were studied during the early follicular phase of the menstrual cycle (days 3–7). All subjects gave their written informed consent, and the study was approved by the regional ethics committee and conducted according to the Declaration of Helsinki.
Study design
After an overnight fast the subjects were admitted to the hospital at 0700 h, and an iv cannula was inserted in each arm for blood sampling and infusions, respectively. On each occasion, [3-3H]glucose (New England, Nuclear, Boston, MA) was infused as a bolus of 30 µCi, followed by a continuous infusion of 0.3 µCi/min for 7 h. Two hours were allowed for the isotope to equilibrate, after which a bolus of 200 µg biosynthetic human GH (Norditropin, Nordisk Gentofte, Gentofte, Denmark) or saline was administered in an exponentially declining fashion in the course of 8 min. Blood samples were drawn at baseline, just before GH/saline infusion, and then every 5 min for the first hour, every 10 min for the next hour, and every 20 min for the last 3 h when GH was given. In the saline experiment, blood samples were subsequently drawn every 20 min for 5 h. GH was measured in every sample, metabolites were measured at -120, 0, and then every 20 min. Insulin, insulin-like growth factor-binding protein-1 (IGFBP-1), and nonesterified fatty acids (NEFA) were measured at -120, 0, 20, and then every 40 min. Serum insulin-like growth factor I (IGF-I) was measured at -120, 0, and then every 60 min. Indirect calorimetry, using a ventilated hood system (Deltatrac, Datex Instrumentarium, Helsinki, Finland), was performed before (-20 to 0 min) and 2 h after the infusion of GH/saline (120–140 min) to assess resting energy expenditure (REE), respiratory exchange ratio (RER), and rates of lipid and glucose oxidation (13).
Analyses
A double monoclonal immunofluometric assay (Delfia, Wallac, Finland) was used to measure serum GH. The interassay coefficient of variation in samples varied between 1.7–2.4%, the intraassay coefficient of variation varied between 1.9–3.0% for GH concentrations of 12.08 and 0.27 µg/L, and the detection limit was lower than 0.01 µg/L. Serum IGF-I was measured by a noncompetitive time resolved immunofluometric assay (14). Serum IGFBP-1 was measured by a commercial enzyme-linked immunosorbent assay (Medix Biochemica, Kainiainen, Finland), and insulin was determined by conventional in-house RIA (15). NEFA were determined by a colorimetric method employing a commercial kit (Wako Chemicals, Neuss, Germany). Blood 3-hydroxybutyrate, glycerol, alanine, and lactate were assayed by automated fluorometric enzymatic methods (16). Glucose turnover was estimated according to the nonsteady state model of Steele et al. as modified by deBodo et al. (17) based on data from the infused tritiated glucose tracer. Serum GH-binding protein (GHBP) was measured in an immunometric assay as recently described (18). Serum estradiol was measured by a solid phase fluoroimmunoassay (AutoDelphia), based on competition between europium-labeled estradiol and sample estradiol for polyclonal antiestradiol antibodies (derived from rabbits).
Body composition
The muscle/fat ratio of the midthigh region and sc fat, the maximal antero-posterior diameter, and the amount of intraabdominal (visceral) fat at the umbilical level were evaluated by computed tomography. The areas scanned comprised 10 mm cross-sectional slices at the middle of the thigh and at the umbilicus. Total body fat was measured by dual energy x-ray absorptiometry. The data on body composition have been published previously (11).
Metabolic clearance and distribution volume of GH
The MCR and Vd of GH were estimated using the following equations (19): MCR = dose of GH injected/GH area under the curve (AUCGH); Vd = MCR/kß, where kß is the elimination constant determined from the log-linear portion of the plasma concentration time curve (the ß-phase); and kß = ln2/t1/2. This allows estimation of the distribution volume even when drug kinetics are described by more than one body compartment.
Statistical analyses
Differences between gender and age groups in body composition,
basal values of the metabolic parameters, and metabolic response to a
GH bolus were assessed by Student’s t test for unpaired
data. Two-way ANOVA for repeated measures was used to estimate the
changes with time and treatment (GH or saline). Age- or gender-based
differences in the response to GH were detected with multivariate
ANOVA. Intraabdominal fat and total body fat were entered as covariates
one by one to estimate the influence of body fat on age- and
gender-based differences. Pearson’s product-moment correlation was
used to measure the strength of association between the metabolic
response and body composition and age. Multiple linear regression and
forward stepwise regression analyses were used to determine the
strongest predictors among the age and body composition estimates
(independent variables) of selected metabolic response parameters
(dependent variables). Analyses were made on SD scores
(z-scores) where appropriate. The z-score at any time point was
calculated using baseline values as references, i.e. the
concentration at the actual time minus the concentration at time zero
divided by the SD at time zero {z-score =
[C(tn) -
C(t0)]/SD(t0)}. Delta values
expressed in z-scores are differences in z-scores between GH and saline
infusions. Data were log transformed when not normally distributed.
Data are given as the mean ± SE. Statistical
significance was assumed for P < 0.05. When multiple
comparisons were made, a protected P 
0.01 was
employed.
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Results |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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and have been reported previously (11).
In brief, relative adiposity, especially intraabdominal fat mass, was
greater in the older subjects. Body weight was insignificantly higher
in the older group (P = 0.09). In females, total body
fat was higher, whereas intraabdominal fat mass and body weight were
lower than those in males. As shown in Table 2, baseline values of serum IGF-I
(micrograms per L) and REE per kg BW were higher in the younger
subjects. The difference in baseline values of serum insulin
(milliunits per L) approached significance (P = 0.055),
with the older group having the highest values. Females exhibited
higher baseline values of serum IGFBP-1 and lipid intermediates than
males.
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Peak serum GH (Cmax) values after infusion of GH
differed between age groups, but not between sexes (Fig. 1 and Table 3; males, 80.2 ± 10.1 µg/L;
females, 75.2 ± 7.7 µg/L; P = NS). The
AUCGH after GH infusion correlated inversely with age
(r = -0.55; P < 0.001), intraabdominal fat
(r = -0.45; P = 0.009), and waist/hip ratio
(r = -0.43; P = 0.008). Multiple linear
regression analysis revealed that age was the most important predictor
of the AUCGH in all subjects.
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) of serum GH,
as calculated from the first 10 min after peak serum GH, was higher
among young subjects and in males, but the elimination half-life
(t1/2ß) did not differ between age (Table 3
) or gender
(males, 16.8 ± 0.74; females, 17.7 ± 1.02;
P = NS). The MCR and Vd of GH were both
higher in the older group (Table 3), without significant gender
differences. The Vd correlated positively with age (r
= 0.66; P < 0.001) and intraabdominal fat mass (r
= 0.54; P = 0.002). The MCR correlated positively with
age (r = 0.57; P < 0.001), intraabdominal fat
mass (r = 0.54; P = 0.001), sc fat (r = 0.48;
P = 0.005), total body fat (r = 0.47;
P = 0.006), serum insulin (r = 0.49;
P = 0.003), and serum GHBP (r = 0.46;
P = 0.005). Multiple linear and forward stepwise
regression analyses revealed age as the most important predictor of
both MCR and Vd. Lipid intermediates
As shown in Fig. 2, the mean z-score
for NEFA increased after GH from baseline values to a peak of 0.5.
These changes were both GH (P = 0.006) and time
(P < 0.0001) dependent. Serum 3-hydroxybutyrate
increased in a similar GH (P < 0.0001)- and time
(P < 0.0001)-dependent manner from baseline to a mean
z-score of 3.5. Likewise, glycerol increased after GH treatment from
baseline to a mean z-score of 1.4, followed by a decrease to 0.8 after
180 min.
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A time-dependent (P = 0.08) decrease to a mean
z-score of -0.8 in isotopically determined endogenous glucose
production (rate of appearance) was found after both GH and saline
treatment. The disposal rate of glucose equalled the production rate
(Fig. 3). Likewise, plasma levels of
glucose decreased with time to a mean z-score of -0.7
(P < 0.001), with no significant difference between GH
and saline (Fig. 3).
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Serum IGF-I, IGFBP-1, and insulin
No significant changes in serum IGF-I was recorded between the two
situations. A significant decrease in IGFBP-1 to a mean z-score of
-0.5 was seen after GH treatment (Fig. 3). A gradual decline in serum
insulin was observed after saline, whereas serum insulin levels
appeared to reach a plateau after GH administration. With ANOVA,
however, no significant differences between saline and GH could be
detected (P = 0.42; Fig. 3).
Gas exchange
A decrease in RER [saline, 0.82 ± 0.01 to 0.80 ± 0.01 (P = NS); GH, 0.82 ± 0.01 to 0.78 ± 0.01 (P = 0.03)] and an increase in energy expenditure [saline, 22.9 ± 0.4 to 22.8 ± 0.4 Cal/24 h·kg (P = NS); GH, 22.7 ± 0.4 to 23.2 ± 0.4 (P = 0.03)] were found after GH infusion. Glucose oxidation decreased from 5.1 ± 0.3 to 4.3 ± 0.2 mg/kg·min when saline was infused. When GH was infused, the decrease was significantly larger (from 5.1 ± 0.3 to 3.8 ± 0.2 mg/kg·min; P = 0.02). Lipid oxidation increased in both situations, but the increase was greater with GH (saline, 8.2 ± 0.3 to 8.9 ± 0.3 mg/kg·min; GH, 7.8 ± 0.4 to 9.4 ± 0.4; P < 0.001).
Age- and sex-related differences in the metabolic response
Multivariate ANOVA revealed that the young group had a higher
lipolytic response to GH (Fig. 4).
Introducing intraabdominal fat mass and total body fat as covariates
for 3-hydroxybutyrate did not make the age-dependent difference
disappear (Table 4). Gender-based
differences were also found in the GH-induced increase in
3-hydroxybutyrate, with males having a higher lipolytic response (Fig. 4). Correcting for intraabdominal fat or total body fat did not make
the gender-dependent difference disappear (Table 4). Although serum
insulin in the group as a whole did not change significantly after a GH
pulse, compared to that after saline, subdivision according to gender
and age revealed higher (nondeclining) insulin levels during GH
exposure in young subjects and in females. These differences could
largely be accounted for by differences in intraabdominal fat or total
body fat, respectively (Table 4). Analysis of IGFBP-1 dynamics
according to age and sex revealed a GH-induced significant
reduction in older subjects and in females. These age and sex effects
in IGFBP-1 persisted after correction for body composition (Fig. 4 and
Table 4).
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Discussion |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionReferences |
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Surprisingly, the AUC of GH after infusion was much greater in young than in older subjects. Body weight and total body resistance, measured by bioimpedance, did not differ between the age groups (data not shown), indicating no major differences in extracellular volume. Both the MCR and the apparent Vd, calculated from the semilog time-concentration curve, were higher in the older subjects. To our knowledge, no earlier studies have looked at differences in Vd with age, whereas our finding of a higher MCR in the older group is in contrast to the results of earlier studies (22, 23, 24, 25), although higher values of MCR in adults than in prepubertal children have been reported (26), and studies in pigs (27) and heifers (28) reported increased MCR with aging. In the present study, both MCR and Vd were closely related to age and intraabdominal fat mass. It is conceivable, therefore, that GH is distributed in and cleared by adipose tissue. Furthermore, positive correlations were found between GHBP and estimates of body fat (data not shown), which confirms earlier findings (29). As adipose tissue contains receptors for GH (30), it could be speculated that GHBP in serum reflects the amount of GH receptors in adipose tissue. It could be hypothesized that the number of GH receptors is up-regulated in adipose tissue to compensate for the decreased GH secretion even though this is insufficient to impede the formation of additional adipose tissue. Alternatively, the increased MCR in the older subjects could be a primary event, contributing to the hyposomatropinemia of obesity. Our finding is supported by a study in monkeys in which a higher MCR was found in obese compared to lean animals (31).
The increase in Vd with age may in addition have more direct implications. Several hormone analysis programs have been developed over the past years, most of which assume a given distribution space (32, 33). This may tend to underestimate hormone production in subjects with a higher Vd, i.e. the elderly. Our data indicate that individual variations in Vd should be anticipated and accounted for in studies using these techniques.
The observed effects of GH on lipid metabolism are in accordance with the results of previous studies (4, 5, 6). The time lag in the increase in lipid intermediates after GH treatment also resembles that found after a nocturnal peak of endogenous GH in healthy young subjects (34), which underlines that GH plays an important role in the regulation of lipids. Neither intraabdominal nor total body fat accounted for the age- and gender-based differences in the lipolytic GH response.
The gradual decrease in basal endogenous glucose production without any significant effect of GH confirms the findings of earlier studies using similar GH doses (4, 5, 35). The unchanged levels of serum IGF-I throughout both situations are also in accordance with previous findings (4, 35).
It remains uncertain whether aging exerts specific effects on serum IGFBP-1 patterns, and our finding of a suppression of IGFBP-1 shortly after an exogenous GH pulse in older subjects and in females is a new observation. It is generally assumed that circulating IGFBP-1 levels are lowered by a direct suppressive effect of portal insulin on hepatic IGFBP-1 production (36). We only measured peripheral insulin levels, which may not adequately detect small and short-lived secretions from the ß-cell. Regardless of the underlying mechanism, it has been proposed that reductions in circulating IGFBP-1 levels are associated with increased IGF-I activity at target cell levels (36). When applied to our data, this could mean that acute physiological GH exposure may increase IGF-I bioactivity, especially in older subjects and in females.
The decrease in nonprotein RER after GH infusion is in accordance with the findings of other studies (5, 35) and reflects the increase in lipid oxidation and the decrease in glucose oxidation. These studies did not find any effect of GH on REE, whereas in our study an increase in energy expenditure was seen. The latter observation is compatible with a study in GH-deficient adults, in whom overnight GH infusion prompted an increase in REE (3).
This study is to our knowledge the first to look at age- and gender-based differences in the metabolic response to a GH pulse. In a recent study the metabolic actions of recombinant human IGF-I and insulin were compared during euglycemic clamp studies in two large groups of young and older adults (37). They concluded that age did not alter the metabolic response to IGF-I.
In the present study, the lipolytic response was higher in young subjects and among males. By contrast, the GH-induced suppression of IGFBP-1 was more pronounced in older subjects and among females. Although the physiological implications of the latter observation is less clear, it could be speculated that aging is associated with decreased susceptibility to the direct lipolytic effects of GH, whereas the ability of GH to promote IGF-I bioactivity increases with age. It is, however, important to emphasize that we only studied acute effects. As continued GH treatment induces changes in body composition, which, in turn, may modify both the pharmacokinetics and biological effects of GH, it is difficult to make predictions about the physiological and clinical relevance of acute metabolic studies. In all circumstances, our findings suggest that the interactions between GH and aging are highly complex, involving an interplay among pharmacokinetics, body composition, and metabolic responsiveness.
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Acknowledgments |
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Received January 30, 1997.
Revised July 23, 1997.
Accepted August 4, 1997.
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| Endocrinology | Endocrine Reviews | J. Clin. End. & Metab. |
| Molecular Endocrinology | Recent Prog. Horm. Res. | All Endocrine Journals |
, Young; •, older. 
, GH infusion for 8
min. Data are the mean ± SEM.
z-scores (mean ± SEM). 
,
males; 
, females.