Estradiol supplementation enhances submaximal feed-forward drive of growth hormone (GH) secretion by recombinant human GH-releasing hormone-1,44-amide in a putatively somatostatin-withdrawn milieu

To test the clinical hypothesis that an estrogen-enriched milieu enhances GHRH action, we administered placebo (Pl) and estradiol-17 beta (E(2)) orally for 23 d to six postmenopausal women in a prospectively randomized, double-masked, within-subject crossover design with 6 wk intervening. The GHRH stimulation protocol entailed consecutive i.v. infusion of L-arginine and a single i.v. pulse of saline or one of five randomly ordered doses of recombinant human GHRH-1,44-amide (0.03, 0.1, 0.3, 1.0, or 3.0 microg/kg) in a total of 12 separate morning, fasting sessions. GH secretion was monitored by sampling blood every 10 min for 6 h; chemiluminescence assay of GH concentrations; deconvolution analysis of stimulated GH release; and nonlinear dose-response reconstruction. Supplementation with E(2), compared with Pl: 1) increased (mean +/- SEM) E(2) concentrations from 18 +/- 3 (Pl) to 164 +/- 12 pg/ml (to convert to picomoles per liter, multiply by 3.57) (P < 0.001); 2) decreased IGF-I concentrations from 181 +/- 14 to 120 +/- 11 microg/liter (P < 0.01); 3) elevated mean GH concentrations from 0.27 +/- 0.06 to 0.59 +/- 0.08 microg/liter (P = 0.014); 4) potentiated GH secretion stimulated by L-arginine alone by 1.43-fold (P = 0.012); 5) reduced the ED(50) of GHRH from 0.27 +/- 0.02 to 0.13 +/- 0.01 microg/kg (P < 0.01), denoting enhanced GHRH potency; and 6) heightened the maximal slope of the dose-response function from 1.1 +/- 0.1 to 1.4 +/- 0.05 [( microg/liter) ( microg/kg)(-1)] (P < 0.05), signifying augmented pituitary sensitivity. The foregoing facilitative mechanisms were specific because E(2) replacement did alter maximal L-arginine/GHRH-induced GH secretion, indicating unchanged secretagogue efficacy. In conclusion, inasmuch as E(2) also attenuates inhibition of GH secretion by infused somatostatin and potentiates stimulation of GH secretion by GH-releasing peptide-2, we postulate that estrogenic steroids drive pulsatile GH production in part via mechanisms that include all three of GHRH, somatostatin, and putatively GH-releasing peptide/ghrelin signaling.     

Estradiol supplementation modulates growth hormone (GH) secretory-burst waveform and recombinant human insulin-like growth factor-I-enforced suppression of endogenously driven GH release in postmenopausal women

The present study tests the mechanistic postulate that estrogen confers resistance to negative feedback by systemic IGF-I. To this end, eight postmenopausal women received a constant iv infusion of recombinant human (rh)IGF-I (10 micro g/kg.h x 6 h) and saline in randomized order on the 10th day of supplementation with oral estradiol (E(2)) and placebo (Pl). GH secretion was quantitated by 10-min blood sampling, immunochemiluminometry assay, and deconvolution analysis. Administration of E(2) compared with Pl followed by saline infusion: 1) stimulated pulsatile GH secretion ( micro g/liter.6 h), viz., 12 +/- 3.3 (Pl) and 18 +/- 4.6 (E(2)) (mean +/- SEM, paired comparison, P < 0.05); 2) halved the time latency (min) to achieve peak GH secretion after GHRH injection, 24 +/- 2.2 (Pl) and 12 +/- 2.1 (E(2)) (P < 0.01); and 3) did not alter the mass of GH secreted ( micro g/liter) in response to a maximally effective dose of GHRH, 30 +/- 7.2 (Pl) and 37 +/- 11 (E(2)). Exposure to E(2) compared with Pl followed by rhIGF-I infusion: 1) accelerated the rate of decline of GH concentrations by 3.3-fold, viz., absolute slope ( micro g/liter.1000 min), 3.8 (range, 2.5-5.0) (Pl) and 12 (range, 10-14) (E(2)) (P < 0.001); 2) augmented the algebraic decrement in GH concentrations ( micro g/liter) enforced by rhIGF-I infusion, 0.73 +/- 0.21 (Pl) and 1.6 +/- 0.25 (E(2)) (P < 0.01); 3) halved the time delay (min) to peak GHRH-induced GH secretion, 20 +/- 1.2 (Pl) vs. 10 +/- 1.3 (E(2)) min (P < 0.01). In contradistinction, E(2) did not alter: 1) the capability of rhIGF-I to suppress GHRH-stimulated GH secretory burst mass significantly, viz., by 50 +/- 8% (Pl) and 52 +/- 14% (E(2)) (P < 0.05 each vs. saline); 2) the hourly rate of rise of infused (total) IGF-I concentrations; and 3) total and ultrafiltratably free IGF-I concentrations ( micro g/liter) attained at the end of the two rhIGF-I infusions. In summary, compared with Pl, E(2) supplementation in postmenopausal women: 1) amplifies endogenously driven GH secretory-burst mass; 2) initiates rapid onset of GHRH-stimulated GH release; and 3) potentiates IGF-I-dependent suppression of unstimulated GH concentrations. Based upon companion modeling data, we postulate that E(2) facilitates the upstroke and IGF-I enforces the downstroke of high-amplitude GH secretory bursts in estrogen-replete individuals.     

Short-term testosterone supplementation does not activate GH and IGF-I production in postmenopausal women

BACKGROUND: Testosterone (Te), oestradiol, GH and IGF-I concentrations fall in postmenopausal women. OBJECTIVE: To test the effect of increased Te availability on impoverished GH/IGF-I secretion in this context. DESIGN/PATIENTS: Placebo (Pl) and a low vs. high dose of Te were administered transdermally for 7 days to each of 15 healthy older women (ages 48-81 years) in a randomized, double-blind crossover design. MEASUREMENTS: Frequent blood sampling, GHRP-2 (Growth hormone releasing peptide-2) infusion, and deconvolution analysis were used to assess GH secretion. RESULTS: Graded Te supplementation increased serum Te concentrations (nmol/l) from 0.87 +/- 0.06 [Pl] to 5 +/- 1 [low] and 7.3 +/- 1.6 [high] (P < 0.001), but did not affect: (i) pulsatile GH secretion (microg/l/12 h; conversion factor: 1 microg/l = 0.33 mU/l), 29 +/- 7.2, 32 +/- 5.6 and 27 +/- 4.9; (ii) GH regularity (approximate entropy), 0.52 +/- 0.05, 0.57 +/- 0.05 and 0.56 +/- 0.05; (iii) IGF-I concentrations (microg/l; conversion factor: 1 microg/l = 0.131 nmol/l), 126 +/- 13, 135 +/- 15 and 141 +/- 13; (iv) IGFBP-3 (mg/l), 3.7 +/- 0.2, 3.6 +/- 0.1 and 3.7 +/- 0.2; (v) IGFBP-1 (microg/l), 41 +/- 2.3, 44 +/- 3.8 and 44 +/- 3.2; and (vi) the mass of GH secreted (microg/l/3 h) after GHRP-2 injection, 123 +/- 27, 146 +/- 32 and 147 +/- 38. CONCLUSION: Up to 8-fold elevation of Te concentrations for 1 week in postmenopausal women does not detectably amplify GH secretion or action, thus indicating that low androgen availability is not a proximate acute mediator of hyposomatotropism in postmenopausal individuals.     

Age and secretagogue type jointly determine dynamic growth hormone responses to exogenous insulin-like growth factor-negative feedback in healthy men

The primary cause of waning GH and IGF-I concentrations in healthy aging adults is not established. To test the postulate that age influences negative feedback by IGF-I in a secretagogue-specific fashion, 17 normal men (nine young and eight older) each completed eight randomly ordered injections of placebo or recombinant human (rh) IGF-I (20 microg/kg sc), followed by saline/rest, aerobic exercise, GHRH (1 microg/kg iv bolus), or GH-releasing peptide-2 (1 microg/kg iv bolus) stimulation. GH secretion was monitored by sampling blood every 10 min for 7 h, high-sensitivity immunochemiluminometric assay, and deconvolution analysis conditioned on prior pulse-onset times and biexponential kinetics. Analysis of covariance showed that age (P = 0.028), secretagogue (P < 0.001), and rhIGF-I (P < 0.005) individually determine pulsatile GH secretion and exhibit a strong 3-fold interaction (P < 10(-5)). Post hoc comparisons revealed that elderly subjects manifest less IGF-I inhibition of a maximal GHRH stimulus (P = 0.013 vs. young), blunted initial IGF-I suppression of fasting GH release (P = 0.038), and impaired IGF-I feedback on the regularity of GH secretion (P = 0.023). Age stratum did not influence peak IGF-I and nadir GH concentrations or rhIGF-I-induced inhibition of GH secretion stimulated by exercise or GH-releasing peptide-2. In summary, experimental elevation of IGF-I concentrations unmasks reduced rhIGF-I-dependent feedback inhibition of fasting and GHRH-stimulated GH secretion in healthy older men, indicating that aging selectively modulates the autoinhibition process.     

Contrasting negative-feedback control of endogenously driven and exercise-stimulated pulsatile growth hormone secretion in women and men

GH represses its own secretion via rapid and reversible feedback exerted at key hypothalamic loci. The primary mechanisms include stimulation of somatostatin release and inhibition of GHRH outflow. Autoinhibition is prominent in the adult male rat but diminutive in the female animal. The sex contrast reflects important differences in central neuropeptide signaling in this species. No comparable insights into gender-specific control of GH autofeedback are available in the human. To examine this issue, we quantitated acute recombinant human (rh)GH-induced inhibition of baseline (resting) and aerobic exercise-stimulated GH secretion in healthy young men (n = 8) and early follicular-phase women (n = 6). Each subject underwent four fasting, morning inpatient infusion studies in a prospectively randomized, placebo-controlled, double-blind, within-subject cross-over design. The feedback paradigm comprised 6-min bolus iv infusion of saline or rhGH (10 microg/kg) followed in 120 min by rest or submaximal aerobic (individually calibrated) bicycle ergometry for 30 min. Concomitantly, blood was sampled every 10 min for 6 h, and sera were submitted to immunochemiluminometric GH assay (sensitivity 0.005 microg/liter). Biexponential deconvolution analysis was applied to estimate stimulated GH secretory-burst mass (microg/liter per 90 min after onset of exercise or rest). Women and men had statistically comparable serum estradiol but unequal testosterone concentrations. Repeated-measures ANOVA documented a significant three-way interaction among gender, stimulus type (rest or exercise), and feedback status (saline or rhGH injection) in determining GH secretory-burst mass (P = 0.008). There were prominent two-factor interactions among gender and exercise (P < 0.001); gender and rhGH-induced negative feedback (P = 0.002); and exercise and rhGH feedback (P = 0.006). Gender comparisons disclosed that women, compared with men, maintain 20-fold higher GH secretory-burst mass at rest (P < 0.001); 40-fold less stimulation of pulsatile GH release by exercise than rest (P < 0.001); and 20-fold greater inhibition of GH secretory-burst mass by rhGH than saline at rest (P < 0.05). Observed feedback contrasts by sex were specific, inasmuch as gender did not affect absolute estimates of exercise-stimulated GH secretion (microg/liter/90 min); nadir GH concentrations (microg/liter) enforced by rhGH infusion; and the time latency (min) to manifest maximal inhibition after rhGH injection. In summary, the present clinical investigation unmasks: 1) markedly greater fractional feedback inhibition of pulsatile GH secretion by rhGH in young women than men; and 2) partial resistance of the aerobic-exercise stimulus to GH autofeedback in both women and men. We postulate that sex-steroid-specific control of somatostatin and GHRH outflow may mediate the former gender contrasts, whereas unknown (gender-independent) factors may determine the capability of exercise to significantly antagonize GH autoinhibition.     

Significance of "abnormal" nadir growth hormone levels after oral glucose in postoperative patients with acromegaly in remission with normal insulin-like growth factor-I levels

Our initial study in postoperative patients with acromegaly identified a group of patients in remission, as defined by normal IGF-I levels, but who had a subtle abnormality of GH suppression after oral glucose. To investigate the significance of this abnormality, we have undertaken further detailed testing of GH secretion and a longitudinal follow-up of some of these patients. Of the 110 postoperative patients with acromegaly evaluated by oral glucose tolerance test, 76 were in remission (i.e. normal IGF-I level), and of these subjects with acromegaly in remission, 50 had normal nadir GH (<0.14 microg/ml) (group I), and 26 had abnormal nadir GH (>0.14 microg/ml) (group II). Fourteen subjects in remission, seven from remission group I and seven from remission group II, underwent additional testing consisting of both hourly GH sampling over 8 h and, on a separate day, arginine stimulation testing. The mean of hourly GH was higher in group II (0.47 +/- 0.04 microg/liter) than in group I (0.19 +/- 0.07 microg/liter; P = 0.002). GH response to arginine was greater in group II than in group I (P < 0.01). Of those patients in remission from the initial cohort studied, 49 (30 subjects from group I and 19 from group II) underwent serial longitudinal oral glucose tolerance testing every 1-2 yr over a 1- to 6.5-yr period (mean follow-up, 3.2 yr). The initial pattern of GH suppression persisted in most patients. IGF-I levels remained normal in all patients in group II, but five subjects from group II developed an elevated IGF-I level and, thus, a biochemical recurrence. The rate of disease recurrence was greater in group II than in group I (P = 0.003). We have found that some postoperative subjects with acromegaly in remission with normal IGF-I levels have persistently abnormal nadir GH levels after oral glucose that may be accompanied by other evidence of greater GH secretion than postoperative patients with normal GH suppression. This abnormal pattern of GH suppression may be associated with increased risk of disease recurrence in some patients.     

Activation of somatostatin-receptor subtype-2/-5 suppresses the mass, frequency, and irregularity of growth hormone (GH)-releasing peptide-2-stimulated GH secretion in men

Somatostatin antagonizes the stimulatory actions of GHRH and GH-releasing peptides (GHRPs). However, precisely how the inhibitory susceptibilities of the two secretagogues differ is not clear. One interpretative difficulty is that native somatostatin activates six different receptor subtypes. The present study adopts the complementary strategy of enforcing feedback inhibition via the preferential somatostatin receptor subtype 2 and 5 (SSTR-2/-5) agonist, octreotide. We postulated that putative SSTR-2/-5 agonism would unmask secretagogue-selective interactions in the control of GH secretory burst mass, frequency, and/or regularity. To this end, 10 healthy men each underwent eight randomly ordered, separate-day, fasting morning infusion sessions. Interventions comprised sc administration of octreotide (1 microg/kg), followed by bolus iv injection of saline, GHRH (1 microg/kg), GHRP-2 (1 microg/kg), or both peptides. Compared with placebo, the SSTR-2/-5 agonist reduced fasting GH concentrations from 0.27 +/- 0.07 to 0.12 +/- 0.02 microg/liter (P = 0.020), GH secretory burst mass from 2.7 +/- 0.65 to 0.55 +/- 0.11 microg/liter (P = 0.013), and basal GH secretion from 0.24 +/- 0.043 to 0.11 +/- 0.015 microg/liter.100 min (P = 0.0063). The foregoing outcomes were selective, because octreotide did not alter GH secretory burst frequency (3.1 +/- 0.5 vs. 3.3 +/- 0.21 events/3 h) or the regularity of the GH release process (approximate entropy, 0.58 +/- 0.048 vs. 0.68 +/- 0.064). In the GHRP-2-stimulated setting, presumptive SSTR-2/-5 agonism suppressed all three GH secretory burst masses, from 28 +/- 3.2 to 18 +/- 2.0 (P = 0.045); GH pulse frequency, from 3.3 +/- 0.30 to 2.0 +/- 0.18 (P = 0.0025); and the irregularity (approximate entropy) of GH release, from 0.648 +/- 0.049 to 0.433 +/- 0.047 (P < 0.01). In contrast, in the GHRH and combined GHRH/GHRP-2-stimulated contexts, octreotide decreased only GH secretory burst mass (P = 0.047). In summary, the present data indicate that GH secretory burst mass, frequency, and orderliness are subject to interactive control by at least SSTR-2/-5-dependent feedback and GHRP-dependent feedforward signals.     

Free rather than total circulating insulin-like growth factor-I determines the feedback on growth hormone release in normal subjects

Pituitary GH secretion is feedback regulated by circulating IGF-I. However, it remains to be determined whether the feedback control is mediated through circulating free or total IGF-I. To study this, we compared the temporal changes in circulating levels of GH vs. free and total IGF-I during fasting. Seventeen healthy normal-weight subjects (body mass index 23.4 +/- 0.6 kg/m(2)) were studied during 80 h of fasting. Serum was assayed for GH every 3 h; total, free, and bioactive IGF-I, IGF binding protein (IGFBP)-1, -2, and -3 as well as IGFBP-1 bound IGF-I were assayed every morning. During fasting, mean 24-h GH levels increased from 1.41 +/- 0.20 to 3.01 +/- 0.46 and 2.09 +/- 0.30 microg/liter (d 1 vs. d 2 and 3; P < 0.03). After 24 h of fasting, free and bioactive IGF-I had decreased by 40 +/- 5 and 17 +/- 5%, respectively (P < 0.02), and both concentrations remained suppressed for the rest of the study. In contrast, total IGF-I remained unchanged until the end of d 3, at which levels were slightly reduced (P < 0.007). IGFBP-1 increased from 38 +/- 2 to 137 +/- 24, 212 +/- 32, and 214 +/- 22 microg/liter (d 1 vs. d 2, d 3, and end of d 3; P < 0.0001), and these changes closely paralleled those of IGFBP-1-bound IGF-I (P < 0.0001). IGFBP-2 increased only transiently at d 2 (P < 0.05), and IGFBP-3 remained unchanged. The increase in mean 24-h GH levels from d 1 to d 2 correlated inversely with the relative reduction in free IGF-I from d 1 to d 2 (r = -0.51; P = 0.04), i.e. the larger the reduction in free IGF-I, the larger the increase in GH. None of the other IGF-related parameters correlated with GH. In conclusion, the temporal relationship between the increase in GH and the reduction in free IGF-I supports the hypothesis that circulating free IGF-I mediates the feedback regulation of GH secretion.     

Accelerated escape from GH autonegative feedback in midpuberty in males: evidence for time-delimited GH-induced somatostatinergic outflow in adolescent boys

A single injected pulse of GH inhibits the time-delayed secretion of GH in the adult by way of central mechanisms that drive somatostatin and repress GHRH outflow. The marked amplification of spontaneous GH pulse amplitude in puberty poses an autoregulatory paradox. We postulated that this disparity might reflect unique relief of GH-induced autonegative feedback during this window of development. The present study contrasts GH autonegative feedback in: 1) normal prepubertal boys (PP) (n = 6; Tanner genital stage I, chronologically aged 8 yr, 9 months to 10 yr, 1 month; median bone age 8.5 yr); 2) longitudinally identified midpubertal boys (MP) (n = 6; Tanner genital stages III/IV, aged 12 yr, 6 months to 15 yr, 6 months; median bone age 15 yr); and 3) healthy young men (YM) (n = 6, aged 18-24 yr; bone age >18 yr). Subjects each underwent four randomly ordered tandem peptide infusions on separate mornings while fasting: i.e. 1) saline/saline infused iv bolus at 0830 h and 1030 h; 2) saline/GHRH (0.3 microg/kg i.v. bolus) at the foregoing times; 3) recombinant human (rh) GH (3 microg/kg as a 6-min square-wave i.v. pulse)/saline; and 4) rhGH and GHRH. To monitor GH autofeedback effects, blood samples were obtained every 10 min for 5.5 h beginning at 0800 h (30 min before GH or saline infusion). Serum GH concentrations were quantitated by ultrasensitive chemiluminometry (threshold 0.005 microg/liter). On the day of successive saline/saline infusion, MP boys maintained higher serum concentrations of: 1) GH ( microg/liter), 2.2 +/- 0.25, compared with PP (0.61 +/- 0.10) or YM (0.88 +/- 0.36) (P = 0.011); 2) IGF-I ( micro g/liter), 493 +/- 49 vs. PP (134 +/- 16) and YM (242 +/- 22) (P < 0.001); 3) T (ng/dl), 524 +/- 58 vs. PP (<20) (P < 0.001); and 4) E2 (pg/ml),19 +/- 3 vs. PP (< 10) (P = 0.030) (mean +/- SEM). Consecutive saline/GHRH infusion elicited comparable peak (absolute maximal) serum GH concentrations (micrograms per liter) in the three study groups, i.e. 18 +/- 5.0 (PP), 9.6 +/- 1.7 (MP), and 14 +/- 5.3 (YM) (each P < 0.01 vs. saline; P = NS cohort effect). Injection of rhGH attenuated subsequent GHRH-stimulated peak serum GH concentrations (micrograms per liter) to 7.8 +/- 1.9 (PP), 5.8 +/- 1.2 (MP), and 4.8 +/- 1.1 (YM) (each P < 0.01 vs. saline; P = NS pubertal effect). GH autofeedback reduced non-GHRH-stimulated (basal) serum GH concentrations by 0.74 +/- 0.28 (PP), 5.7 +/- 1.7 (MP) and 1.4 +/- 0.27 (YM) fold, compared with saline (P = 0.016 for MP vs. PP or YM). In addition to greater fractional autoinhibition, MP boys exhibited markedly accentuated postnadir escape (4.6-fold steeper slope) of suppressed GH concentrations (P < 0.001 vs. PP or YM). Linear regression analysis of data from all 18 subjects revealed that the fasting IGF-I concentration negatively predicted fold-autoinhibition of GHRH-stimulated peak GH release (r = -0.847, P = 0.006) and positively forecast fold-autoinhibition of basal GH release (r = +0.869, P < 0.001). In contrast, the kinetics of rhGH did not differ among the three study cohorts. In summary, boys in midpuberty manifest equivalent responsiveness to exogenous GHRH-stimulated GH secretion; heightened susceptibility to rhGH-induced fractional inhibition of endogenous secretagogue-driven GH release, compared with the prepubertal or adult male; and accelerated recovery of GH output after acute autonegative feedback. This novel tripartite mechanism could engender recurrent high-amplitude GH secretory bursts that mark sex hormone-dependent activation of the human somatotropic axis.     

E2 supplementation selectively relieves GH's autonegative feedback on GH-releasing peptide-2-stimulated GH secretion.

Female gender confers resistance to GH autonegative feedback in the adult rat, thereby suggesting gonadal or estrogenic modulation of autoregulation of the somatotropic axis. Here we test the clinical hypothesis that short-term E2 replacement in ovariprival women reduces GH's repression of spontaneous, GHRH-, and GH-releasing peptide (GHRP)-stimulated GH secretion. To this end, we appraised GH autoinhibition in nine healthy postmenopausal volunteers during a prospective, randomly ordered supplementation with placebo vs. E [1 mg micronized 17 beta-E2 orally twice daily for 6-23 d]. The GH autofeedback paradigm consisted of a 6-min pulsed i.v. infusion of recombinant human GH (10 microg/kg square-wave injection) or saline (control) followed by i.v. bolus GHRH (1 microg/kg), GHRP-2 (1 microg/kg), or saline 2 h later. Blood was sampled every 10 min and serum GH concentrations were measured by chemiluminescence. Poststimulus GH release was quantitated by multiparameter deconvolution analysis using published biexponential kinetics and by the incremental peak serum GH concentration response (maximal poststimulus value minus prepeak nadir). Outcomes were analyzed on the logarithmic scale by mixed-effects ANOVA at a multiple-comparison type I error rate of 0.05. E2 supplementation increased the (mean +/- SEM) serum E2 concentration from 43 +/- 1.8 (control) to 121 +/- 4 pg/ml (E2) (158 +/- 6.6 to 440 +/- 15 pmol/liter; P < 0.001), lowered the 0800 h (preinfusion) serum IGF-I concentration from 127 +/- 7.7 to 73 +/- 3.6 microg/liter (P < 0.01), and amplified spontaneous pulsatile GH production from 7.5 +/- 1.1 to 13 +/- 2.3 microg/liter per 6 h (P = 0.020). In the absence of exogenously imposed GH autofeedback, E2 replacement enhanced the stimulatory effect of GHRP-2 on incremental peak GH release by 1.58-fold [95% confidence interval, 1.2- to 2.1-fold] (P = 0.0034) but did not alter the action of GHRH (0.83-fold [0.62- to 1.1-fold]). In the E2-deficient state, bolus GH infusion significantly inhibited subsequent spontaneous, GHRH-, and GHRP-induced incremental peak GH responses by, respectively, 33% (1-55%; P = 0.044 vs. saline), 79% (68-86%; P < 0.0001), and 54% (32-69%; P = 0.0002). E2 repletion failed to influence GH autofeedback on either spontaneous or GHRH-stimulated incremental peak GH output. In contrast, E2 replenishment augmented the GHRP-2-stimulated incremental peak GH response in the face of GH autoinhibition by 1.7-fold (1.2- to 2.5-fold; P = 0.009). Mechanistically, the latter effect of E2 mirrored its enhancement of GH-repressed/GHRP-2-stimulated GH secretory pulse mass, which rose by 1.5-fold (0.95- to 2.5-fold over placebo; P = 0.078). In summary, the present clinical investigation documents the ability of short-term oral E2 supplementation in postmenopausal women to selectively rescue GHRP-2 (but not spontaneous or GHRH)-stimulated GH secretion from autonegative feedback. The secretagogue specificity of E's relief of GH autoinhibition suggests that this sex steroid may enhance activity of the hypothalamopituitary GHRP-receptor/effector pathway.     

Sustained growth hormone (GH) and insulin-like growth factor I responses to prolonged high-dose twice-daily GH-releasing hormone stimulation in middle-aged and older men

Postulated mechanisms underlying the relative hyposomato-tropism of aging include reduced hypothalamic drive by GHRH. To test this notion, we administered 1 mg (n = 11) vs. 4 mg (n = 11) recombinant human GHRH-1,44-amide s.c. twice daily for 3 months in a double-blind, parallel-cohort design to 22 healthy men (ages, 53-68 yr). After 3 months, GHRH elevated: overnight GH concentrations from 0.71 +/- 0.19 to 1.74 +/- 0.39 microg/liter (P < 0.001; 1 mg) and from 0.80 +/- 0.15 to 5.12 +/- 0.40 microg/liter (P < 0.001; 4 mg) and IGF-I concentrations from 117 +/- 14 to 234 +/- 20 microg/liter (P = 0.007; 1 mg) and from 147 +/- 13 to 286 +/- 22 microg/liter (P < 0.001; 4 mg). Only the higher GHRH dose also increased total body water (tritium space; P = 0.024) and fat-free mass (dual-energy x-ray absorptiometry; P = 0.021), and reduced total abdominal adiposity (computed axial tomography scan; P = 0.042). Both supplementation schedules shortened the time required to walk 30 m and ascend four flights of stairs (P < 0.025 each). Lower extremity strength, aerobic capacity, and bone mineral density did not change. Local injection site reactions were common.We conclude that sc administration of a large dose of GHRH (4 mg) twice daily for 3 months elevates GH and IGF-I concentrations, increases total body water and fat-free mass, reduces total abdominal adiposity, and enhances certain performance measures in healthy aging men but causes local skin reactions.     

Assessment of disease activity in treated acromegalic patients using a sensitive GH assay: should we achieve strict normal GH levels for a biochemical cure?

The definition of a cure for acromegaly is controversial in the absence of a well-defined clinical end-point. Therefore, cure in acromegaly may be arbitrarily defined as a normalization of biochemical parameters. The accepted normal GH levels have been modified over time with the improved sensitivity of GH assays. The objective of the present study was to investigate the suppression of GH levels in the oral glucose tolerance test (oGTT) using a sensitive GH immunoassay in a large group of normal adult subjects and treated acromegalic patients. We evaluated these results in conjunction with IGF-I and IGF binding protein 3 (IGFBP-3) levels. Nadir GH levels after the ingestion of 75 g of glucose, as well as baseline IGF-I and IGFBP-3 levels, were evaluated in 56 normal adult subjects and 32 previously treated acromegalic patients. GH was assayed by an immunofluorometric assay. Normal controls had a mean GH nadir of 0.07 +/- 0.09 microg/liter. Their mean basal IGF-I and IGFBP-3 levels were 160 +/- 58 microg/liter and 1926 +/- 497 microg/liter, respectively. Acromegalic patients had mean GH nadir, IGF-I, and IGFBP-3 levels higher than those of normal subjects (2.6 +/- 7.6 microg/liter, 313 +/- 246 microg/liter, and 2625 +/- 1154 microg/liter, respectively). Considering a GH cut-off value of 0.25 microg/liter, as the normalized postglucose GH upper limit (mean + 2 SD) and, therefore, the target for treated patients, only five patients (15.6%) would have been considered cured. These results suggest that the strict physiological normalization of GH levels after oGTT is not often achieved as a therapeutic endpoint in acromegaly. In addition to the refinement of GH assays, epidemiological studies have suggested that the mean basal GH levels (<2.5 microg/liter) or oGTT-derived GH levels < 2 microg/liter (RIA), or the normalization of IGF-I levels, appear to reduce morbidity and mortality in treated acromegaly. Using this epidemiologically based definition of cure for acromegaly, we reviewed our results obtained with a sensitive GH assay. Twenty-five patients (78%) had oGTT nadir GH < 2 microg/liter. Nineteen subjects had normal age-related IGF-I levels. When the GH nadir cut-off was reduced to 1 microg/liter or less, there was a cure rate of 59.4%. IGF-I and IGFBP-3 levels were normal in 16 and 15 of these 19 patients, respectively. Furthermore, 59.4% of these 32 patients were in remission when age-normalized IGF-I levels were used as a criterion for inactive disease. All but three had GH nadir of 1 microg/liter or less. Finally, the definition of cure may be contradictory in a subgroup (9.4%) of patients with a GH nadir less than 1 microg/liter despite high-for-age IGF-I levels. In conclusion, using a sensitive GH assay it can be seen that the strictly normal postglucose GH values less than 0.25 microg/liter required for biochemical control of acromegaly are not often achieved. Furthermore, the cut-off of GH nadir 1 microg/liter or less is more closely related to normal for age serum IGF-I levels in treated acromegalic patients than 0.25 microg/liter or 2 microg/liter cut-offs. According to previous epidemiological reports, a GH level less than 2.5 microg/liter, determined by RIA, is associated with a reduction of morbidity and mortality. Therefore, our data lead us to postulate that the biochemical criterion of oGTT GH levels 1 microg/liter or less, determined by immunofluorometric assay, is a useful and accurate marker of safe GH secretion in treated acromegaly.     

Lowering total plasma insulin-like growth factor I concentrations by way of a novel, potent, and selective growth hormone (GH) receptor antagonist, pegvisomant (B2036-peg), augments the amplitude of GH secretory bursts and elevates basal/nonpulsatile GH release in healthy women and men.

The present clinical study implements a novel interventional strategy of short-term profound and selective blockade of GH receptors to reduce plasma insulin-like growth factor I (IGF-I) concentrations reversibly in healthy eumetabolic adults. Thereby, we examine the feedback role of systemic IGF-I on GH secretion without introducing the complex metabolic disarray that can otherwise accompany secondary IGF-I deprivation. To this end, we sampled blood at 10-min intervals for 10 h overnight in 8 men (aged 19-46 yr) and 4 women (aged 19-39 yr) to quantitate endogenous GH secretion and half-life 72 h after the prospective, randomly ordered, double blind, and within-subject cross-over administration of pegvisomant (1 mg/kg) or saline (0.5 mL) sc. Pegvisomant is an oligopegylated recombinant human GH peptide mutated to antagonize GH receptor-dependent signaling. Statistical analyses of paired plasma IGF-I concentrations and deconvolution-based quantitation of pulsatile GH secretion revealed that GH receptor blockade 1) suppressed fasting total IGF-I concentrations by 31%, viz. from (mean +/- SEM) 276 +/- 42 (placebo) to 190 +/- 20 microg/L (pegvisomant; P = 0.006) 84 h after drug injection; 2) increased the 10-h mean serum GH concentration by 71% from 1.4 +/- 0.33 (placebo) to 2.4 +/- 0.58 (pegvisomant; P = 0.024); 3) augmented the amplitude of underlying GH secretory bursts by 2.1-fold (i.e. from 0.13 +/- 0.032 to 0.27 +/- 0.076 microg/L.min; P = 0.0088); and 4) elevated the basal/nonpulsatile rate of GH secretion by 2.5-fold (from 2.3 +/- 0.77 to 5.07 +/- 1.8 microg/L.10 h; P = 0.022). The rise in the amplitude of GH secretory bursts correlated with the fall in plasma IGF-I concentrations (r = 0.603; P = 0.038). In contrast, IGF-I depletion did not alter GH secretory pulse frequency, half-duration, interpulse interval, percentage of pulsatile GH release, or half-life of endogenous GH. In summary, selective short-term reduction of systemic IGF-I concentrations in healthy eumetabolic adults drives GH secretion via the specific bipartite neuroregulatory mechanism of amplified GH secretory burst amplitude and elevated basal/nonpulsatile GH release. Endogenous GH half-life and frequency-related features of pulsatile GH secretion are not measurably affected, thus identifying a highly distinctive mode of IGF-I feedback-dependent control of GH output. As the increment in GH secretory burst amplitude correlated with the decrement in plasma IGF-I concentrations, we infer that variations in circulating IGF-I availability within the adult midphysiological range can influence pulsatile and basal GH production by way of negative feedback. Based on data in experimental animals, we speculate that the negative feedback actions of systemic IGF-I on GH secretion are mediated via increased hypothalamic somatostatin release, decreased GHRH (or GH-releasing peptide) secretion, and/or suppressed pituitary GH biosynthesis.     

Growth hormone response during oral glucose tolerance test: the impact of assay method on the estimation of reference values in patients with acromegaly and in healthy controls, and the role of gender, age, and body mass index

CONTEXT: Besides the measurement of IGF-I, GH suppression during an oral glucose tolerance test is recommended to assess the biochemical status in acromegaly. However, the development of highly sensitive and specific GH assays necessitates a critical reevaluation of criteria for diagnosis and follow-up of disease activity. OBJECTIVE: Our objective was to evaluate the between-method discrepancies in GH determinations by different immunoassays considering further confounders like age, gender, and body mass index (BMI). DESIGN, SUBJECTS, AND METHODS: We measured GH during a 75-g oral glucose tolerance test in 46 acromegaly patients (18 controlled, 28 uncontrolled; 19 men; 31-63 yr; BMI 26.4 +/- 0.4 kg/m(2)) and 213 healthy subjects (66 men; 20-76 yr; BMI 30 +/- 0.5 kg/m(2)), using three different commercially available assays [Immulite (Diagnostic Products Corp., Los Angeles, CA), Nichols (Nichols Institute Diagnostika GmbH, Bad Vilbel, Germany), and Diagnostic Systems Laboratories (Sinsheim, Germany)] that were calibrated against the recently recommended GH standards. RESULTS: Results from all assays strongly correlated (r = 0.8-0.996; P < 0.0001). However, the results obtained with the Immulite assay were, on average, 2.3-fold higher than those obtained with Nichols and 6-fold higher than those obtained with Diagnostic Systems Laboratories. Using cutoff limits of 1 microg/liter (Immulite) and 0.5 microg/liter (Nichols) identified 95% of patients with active disease and 78-80% of patients in remission. Basal and nadir GH levels were significantly higher in females than in males (Immulite 2.2 +/- 0.28 microg/liter vs. 0.73 +/- 0.15 microg/liter and 0.16 +/- 0.01 microg/liter vs. 0.08 +/- 0.01 microg/liter; P < 0.001, respectively). In multiple regression analysis, age, BMI, and gender were predictors for basal and nadir GH levels. CONCLUSION: Postglucose GH-nadir values are assay, gender, age, and BMI specific, indicating the need of individual cutoff limits for each assay.     

Testosterone and estradiol regulate free insulin-like growth factor I (IGF-I), IGF binding protein 1 (IGFBP-1), and dimeric IGF-I/IGFBP-1 concentrations

The present study tests the clinical postulate that elevated testosterone (Te) and estradiol (E2) concentrations modulate the effects of constant iv infusion of saline vs. recombinant human IGF-I on free IGF-I, IGF binding protein (IGFBP)-1, and dimeric IGF-I/IGFBP-1 concentrations in healthy aging adults. To this end, comparisons were made after administration of placebo (Pl) vs. Te in eight older men (aged 61 +/- 4 yr) and after Pl vs. E2 in eight postmenopausal women (62 +/- 3 yr). In the saline session, E2 lowered and Te increased total IGF-I; E2 specifically elevated IGFBP-1 by 1.5-fold and suppressed free IGF-I by 34%; and E2 increased binary IGF-I/IGFBP-1 by 5-fold more than Te. During IGF-I infusion, the following were found: 1) total and free IGF-I rose 1.4- to 2.0-fold (Pl) and 2.1-2.5-fold (Te) more rapidly in men than women; 2) binary IGF-I/IGFBP-1 increased 3.4-fold more rapidly in men (Te) than women (E2); and 3) end-infusion free IGF-I was 1.6-fold higher in men than women. In summary, E2, compared with Te supplementation, lowers concentrations of total and ultrafiltratably free IGF-I and elevates those of IGFBP-1 and binary IGF-I/IGFBP-1, thus putatively limiting IGF-I bioavailability. If free IGF-I mediates certain biological actions, then exogenous Te and E2 may modulate the tissue effects of total IGF-I concentrations unequally.     

Ghrelin potentiates growth hormone secretion driven by putative somatostatin withdrawal and resists inhibition by human corticotropin-releasing hormone

CONTEXT: Ghrelin is a 28-amino acid, Ser(3)-octanoylated peptide that stimulates GH secretion in vivo and in vitro. Beyond the capability of ghrelin to synergize with GHRH, little is known about multipeptide modulation of ghrelin's actions in humans. OBJECTIVE: The objective of this study was to test the hypothesis that ghrelin can stimulate GH secretion in the absence or presence of somatostatin withdrawal (induced by l-arginine infusion) and stress-like drive by CRH. DESIGN: This was a randomized, double-blind, placebo-controlled, cross-over interventional study. SETTING: This study was performed at an academic medical center. PARTICIPANTS: Nine healthy postmenopausal women not receiving sex hormones were studied. INTERVENTIONS: Subjects were given an iv infusion of saline and/or l-arginine or human CRH, followed by a bolus iv injection of ghrelin. OUTCOME MEASURES: The outcome measures were pulsatile GH secretion quantified by repetitive blood sampling, immunochemiluminometry, and deconvolution analysis. RESULTS: Consecutive saline/ghrelin infusion increased pulsatile GH secretion from 2.7 +/- 1.0 (saline/saline; mean +/- sem) to 20 +/- 5.0 microg/liter.3 h (P < 0.01). The magnitude of the effect of l-arginine/saline was comparable at 20 +/- 4.5 microg/liter.3 h (P < 0.01). In contrast, sequential l-arginine/ghrelin evoked true synergy of GH release (93 +/- 14 microg/liter.3 h; P = 0.003 vs. l-arginine alone and P = 0.008 vs. ghrelin alone). Human CRH did not affect GH responses to saline/saline (3.9 +/- 1.1 microg/liter.3 h), saline/ghrelin (19 +/- 3.3 microg/liter.3 h), l-arginine/saline (16 +/- 2.7 microg/liter.3 h), or l-arginine/ghrelin (90 +/- 13 microg/liter.3 h). CONCLUSIONS: Assuming that l-arginine reduces somatostatin outflow, we infer that ghrelin can activate hypothalamo-pituitary pathways that are both dependent upon and independent of somatostatinergic restraint even in the face of a strong stress-related signal.     

Short-term testosterone supplementation relieves growth hormone autonegative feedback in men

The present study tests the postulate that testosterone (Te) stimulates GH secretion, in part, by attenuating autonegative feedback. To this end, 13 healthy men (ages 43-71 yr) received three consecutive weekly im injections of placebo (Pl) (n = 7) or Te (200 mg) (n = 6) in a prospectively randomized, double-blind, parallel-cohort design. An iv pulse of saline or recombinant human (rh)GH (3 micro g/kg.6 min) was infused 2 h before bolus saline or GH-releasing peptide (GHRP)-2 (1 micro g/kg) in the fasting state. Blood was withdrawn every 10 min, GH concentrations were quantitated by chemiluminometry, secretion was determined by deconvolution analysis, and outcomes were compared by ANOVA. After Pl, rhGH suppressed basal, pulsatile, and GHRP-2-stimulated GH secretion by 2.6-, 2.4-, and 2.1-fold, respectively (each P < 0.03), and truncated GHRP-2-stimulated GH secretory bursts (P < 0.005). Compared with Pl, Te: 1) stimulated basal and pulsatile GH secretion by 1.9 and 2.4-fold (P < 0.01 and P < 0.02), respectively; 2) reduced feedback on basal GH secretion (P < 0.01); 3) blunted GHRP-2-stimulation by 1.9-fold (P < 0.01); and 4) facilitated initial recovery of rhGH-suppressed GH concentrations (P < 0.005). The foregoing actions were selective, inasmuch as Te did not relieve autoinhibition of pulsatile GH secretion. In summary, short-term Te supplementation decreases rhGH-imposed negative feedback on basal GH secretion and enhances early escape of GH from autoinhibition. In principle, such actions could potentiate the renewal of high-amplitude pulses of GH in androgen-replete individuals.     

Gender- and age-related differences in the endocrine parameters of acromegaly

Acromegaly is a severe slow-developing disease associated with a poor prognosis for cardiovascular disease. To evaluate the impact of age and gender on the severity of the disease, 151 de novo patients with acromegaly (79 women, 72 men, age range 19-77 yr) were included in this open retrospective multi-center cohort study. Basal GH and IGF-I levels, GH response after glucose load and maximal tumor diameter at MRI were measured in all patients at diagnosis. Fasting GH levels and maximal tumor diameter were similar in women and men, while serum IGF-I levels were lower (664.9+/-24.9 vs 755.9+/-32 microg/l; p=0.02) and GH nadir after glucose load was higher (27.5+/-3.7 vs 18.5+/-2.2 microg/l; p=0.04) in women than in men. In both sexes, patients' age was negatively correlated with basal and nadir GH, IGF-I levels and tumor size; fasting GH levels were positively correlated with IGF-I levels and nadir GH after glucose. No interaction between age and gender was found on biochemical and morphological parameters. At diagnosis, elderly patients with acromegaly have lower GH and IGF-I levels, lower GH nadir after glucose load and smaller adenomas than young patients. Women have lower IGF-I levels but higher GH nadir after glucose load than men. These age and gender differences should be considered to appropriately evaluate the activity of acromegaly throughout a life-span.     

Long-term evaluation of postoperative acromegalic patients in remission with previous and newly proposed criteria

Criteria to define remission of acromegaly have changed over years. Since 2000, criteria for cure are normal IGF-I levels and a nadir GH after oral glucose tolerance test (OGTT) of less than 1 microg/liter, although recent studies have suggested to lower this cutoff value. This study reevaluated long-term disease activity of acromegalic patients, who were previously considered in remission, using these criteria. The study included 70 of 146 patients operated on between 1984 and 1996 who were considered cured based on normal IGF-I levels, GH values less than 2.5 microg/liter, and/or disappearance of abnormal GH response to TRH/GnRH. Among these 70 patients, 16 were lost to follow-up, three died, and 11 (one of whom had disease recurrence) only gave a phone interview. Forty patients participated in the study and were reevaluated for IGF-I levels and post-OGTT GH nadir after 14.3 +/- 4.2 (mean +/- sd) yr from surgery. In all patients, normal IGF-I levels and a post-OGTT GH nadir of less than 1 microg/liter were found. In particular, 19 patients had a GH nadir of less than 0.19 microg/liter, i.e. the upper limit (mean + 2 sd) found in 30 controls, whereas 21 patients had a nadir between 0.19 and 0.77 microg/liter. No significant differences in hormonal parameters and comorbidities between the two subgroups were observed. These data showed that lowering the post-OGTT GH cutoff value within the normal range does not seem to better discriminate patients with different disease activity or long-term recurrence risk.     

Unequal impact of short-term testosterone repletion on the somatotropic axis of young and older men

The present clinical study compares the impact of low- and high-dose parenteral testosterone (T) supplementation on daily GH secretory patterns and serum IGF-I, IGFBP-1, and IGFBP-3 concentrations in healthy older (60-82 yr) and young (20-40 yr) men. To this end, we administered three consecutive weekly injections of randomly ordered saline and either a low (100 mg) or a high (200 mg) dose of testosterone enanthate im; namely, saline (n = 17, young and n = 16, older), a low dose (n = 8 young, n = 8 older) and a high dose (n = 9 young, and n = 8 older) of androgen. To monitor somatotropic-axis responses, blood was sampled every 10 min for 24 h for later chemiluminescence-based assay of serum GH, RIA of serum IGF-I, and immunoradiometric assay of serum IGFBP-1 and IGFBP-3 concentrations. Data were analyzed via a nested analysis of covariance statistical design. At baseline (saline injection), older, compared with young, men maintained: 1) similar serum total T, IGFBP-1, and IGFBP-3 but reduced IGF-I concentrations, namely, mean (+/- SEM) IGF-I 160 plus or minus 15 vs. 280 plus or minus 18 microg/liter, (P < 0.001); 2) reduced GH secretory burst mass (0.68 +/- 0.09 vs. 1.2 +/- 0.20 microg/liter, P = 0.031); 3) more disorderly GH release patterns (approximate entropy 0.501 +/- 0.058 vs. 0.288 +/- 0.021, P < 0.001); and 4) blunted 24-h rhythmic GH output (nyctohemeral amplitude 0.25 +/- 0.05 vs. 0.47 +/- 0.08 microg/liter, P = 0.025). Serum T concentrations (ng/dl) did not differ in the two age groups supplemented with either a low dose [550 +/- 50 (young) and 544 +/- 128 (older)] and high [1320 +/- 92 (young) and 1570 +/- 140 (older)] dose of T. The 100-mg dose of androgen exerted no detectable effect on GH secretion in either age cohort but increased the serum IGF-I concentration in young men by 20% (P = 0.00098). The 200-mg dose of T failed to alter daily GH production in young volunteers but in older men stimulated: 1) a 2.03-fold rise in the mean (24-h) serum GH concentration (P = 0.0053, compared with the response to saline); 2) a 1.20-fold increase in basal (nonpulsatile) GH production (P = 0.039); 3) a 2.15-fold amplification of GH secretory burst mass (P = 0.0020); 4) a 2.17-fold elevation of the Mesor of nyctohemeral GH output (P = 0.025); 5) a 1.79-fold enhancement in GH approximate entropy (P = 0.0003); and 6) a 40% increase in the fasting serum IGF-I concentration (P = 0.000005). Multivariate statistical analysis indicated that following high-dose T administration, the E2 increment significantly predicted the IGF-I increment in both age groups combined (P = 0.003); T dose positively forecast the serum total IGF-I concentration (P = 0.0031); and age and T dose jointly determined serum LH concentrations (P = 0.031). In summary, neither a physiological nor a pharmacological dose of T administered parenterally for 3 wk augments daily GH secretion in eugonadal young men. In contrast, a high dose of aromatizable androgen significantly amplifies 24-h basal, pulsatile, entropic, and nyctohemerally rhythmic GH production and elevates the serum IGF-I concentration in older men. The mechanistic basis for the foregoing age-related distinction in GH/IGF-I axis responsivity to T is not known.