0021-972x/92/7506-1487$03.00/0 Journal of Clinical Endocrinology and Metabolism Copyright 0 1992 by The Endocrine Society
Vol. 75, No. 6 Printed in U.S.A.
Renal Clearance, Metabolic Clearance Rate, and HalfLife of Human Growth Hormone in Young and Aged Subjects* MOT01
SOHMIYA
First Division,
YUZURU
AND
Department
of
Medicine,
KATO Shimane
Medical
University,
ABSTRACT We studied the renal clearance, MCR, and half-life of the synthetic 22K form of human GH [recombinant hGH (rhGH)] in seven healthy young and five aged men by means of the constant iv infusion technique. rhGH was infused at a rate of 2 pg/kg ‘2 mL/h for 150 min, after which its disappearance was followed for 50 min. Changes in GH levels in plasma and urine were measured by a highly sensitive enzyme immunoassay. The mean (&SD) renal clearance of GH was significantly greater in the aged group than in the young group (14.3 f 2.5 vs. 4.2 +
Izumo
693, Japan
1.0 FL/min; P < 0.05). The mean MCR was greater in the young group than in the aged group (187.1 f 43.7 us. 120.5 + 39.5 mL/min; P < 0.05), but the MCR adjusted for body weight was not different between the two groups (2.9 f 0.6 us. 2.3 + 0.8 mL/min. kg). No difference was noted in the half-life of GH between the two groups (13.8 f 0.9 us. 14.2 + 0.4 min). These findings indicate that GH reabsorption from the renal tubule may be impaired in elderly subjects, but the disappearance rate of GH is not influenced by age. (J Clin Endocrinol Metab 75: 1487-1490,1992)
C
IRCULATING GH levels are maintained by pituitary GH production, tissuemetabolism, and renal excretion. GH secretion induced by exogenous GH-releasing hormone (GHRH) (1, 2) and insulin hypoglycemia (3) as well as 24-h integrated GH secretion (4-6) are reduced in aged subjects. It was reported that the MCR of GH in adults was greater than that in children (7), although other studies showed no difference in MCR between children and adults (8) or among various age groups in adults (9). Blood GH is filtered into urine at the glomerulus and mostly reabsorbed at the renal tubule (10). However, urinary GH levels were not precisely detectable until the recent introduction of a highly sensitive enzyme immunoassay (11, 12). It is now recognized that urinary GH levels reflect plasma GH concentrations in normal subjects and patients with pituitary disorders (12, 13). It was also reported that urinary GH levels were influenced by the renal function and positively correlated with urinary µglubulin (&MG) and negatively correlated with creatinine clearance (C,,) (14). We previously reported that urinary GH levels were not decreased,but, rather, were increased in aged subjects (15), in whom C,, is decreased and urinary &MG is increased. There are no published data on the renal clearance of GH (C,,) in which the effect of age is examined. In the present study we studied the renal clearance, MCR, and half-life of recombinant human GH (rhGH) in young and aged male subjects. Received October 15, 1991. Address all correspondence and requests for reprints to: Motoi Sohmiya, M.D., First Division, Department of Medicine, Shimane Medical University, Izumo 693, Japan. *This work was supported in part by grants from the Ministry of Education, Science, and Culture, Japan, and the Ministry of Health and Welfare, Japan. A part of the study was presented at the 73rd Annual Meeting of The Endocrine Society, Washington, D.C., June 1991.
Materials
and Methods
Subjects Twelve healthy male subjects, aged 17-76 yr, were included in the present study. They were divided into two groups based on age: a young group (n = 7) with a mean age of 24.6 yr (range, 17-34 yr), and an aged group (n = 5) with a mean age of 66.2 yr (range, 60-76 yr). As shown in Table 1, body weight and body surface area were slightly decreased in aged subjects, but there was no difference in body mass index between the young and aged groups. Renal function, assessed by urinalysis, serum urea nitrogen, serum creatinine, and urinary &MG levels, was within normal limits in all subjects. Informed consents were obtained from all subjects.
Protocol All studies were performed in the morning after overnight fasting in the supine position. After complete voiding of the urine, 500 mL tap water were given orally. Then, an indwelling catheter was placed into a cubital vein for infusion of the 22K form of rhGH (Sumitomo Co., Osaka, Japan). The rhGH was dissolved in 0.9% saline containing 0.3% human albumin and constantly infused for 150 min at a rate of 2 peg/ kg. 2 mL/h using a continuous infusion pump (Atom syringe infusion pump 235, Atom Co., Tokyo, Japan) with a precision of +3.0%. Blood samples were withdrawn through a second iv catheter, placed into the other cubital vein, at intervals of 30 min during the GH infusion and every 5 min after the end of the GH infusion. Urine samples were obtained immediately before and 90 and 150 min after the start of the GH infusion. Blood samples were immediately centrifuged at 4 C, and plasma was kept frozen at -20 C until assayed. Urine samples were added to 0.01 vol 0.01 M sodium phosphate buffer, pH 7.0, containing 10% BSA and 10% NaNa, dialyzed overnight against 0.01 M sodium phosphate buffer, pH 7.0, containing 0.1% NaN3 and 0.1 M NaCl, and kept at 4 C until assayed. All samples were assayed within 1 week.
Assays GH levels in plasma and dialyzed urine were measured in duplicate by a highly sensitive sandwich enzyme immunoassay, using a slight modification of the method described previously (14). For the measurement of GH in plasma and urine samples, anti-GH immunoglobulin Gcoated polystyrene balls were incubated with 100 PL GH standards or 1487
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 06 October 2015. at 02:55 For personal use only. No other uses without permission. . All rights reserved.
1488
SOHMIYA
50. to lOO-PL samples and 50-100 PL buffer in duplicate at 37 C for 6 h with continuous shaking. After removal of the supernatant, the polvstvrene balls were washed twice with 2 mL 150 mM saline and incubated with affinity-purified Fab’-peroxidase conjugate and 150 pg normal rabbit F(ab’h at 4 C for 20 h. then at 20 C for 6 h. After removal of the supernantant, the polystyrene balls were washed with 2 mL saline, incubated with 1 mL saline at 30 C for 10 min, and then washed with 2 mL saline. The balls were moved into a new series of assay tubes and further incubated with 100 WL 0.6 mg 3-(p-hydroxyphenyl)propionic acid as substrate and 50 PL 0.015% Hz02 at 30 C for 90 min. Fluorescence intensity was measured by spectrofluorophotometer (Shimazu RF-5000, Shimazu Co., Kyoto, Japan). The minimal detectable quantity was 0.3 rig/L. The intra- and interassay coefficients of variation were 6.0% and 9.8%, respectively. Creatinine and &MG levels in plasma and urine samples were determined by an autoanalyzer and specific RIA (& RIA Beads, Dainabott Co., Tokyo, Japan), respectively.
Calculations
and data analysis
Ccu was calculated by the following formula: Cc” = (UV/P) X (1.731 A), where U is the urinary GH level in the 150 min urine sample (micrograms per L), I’ is the mean plasma GH at 90, 120, and 150 min (micrograms per L), V is the urine volume from 90-150 min (milliliters per min), and A is the body surface area (square meters; 1.73 m2 is the standard body surface area). The MCR of GH was calculated as follows by the general formula of Tait (16): infusion rate of GH @g/h)/steady state plasma GH (pg/L). The half-life of GH (t& was calculated by means of linear regression analysis (17) of GH levels from 150-200 min in the semilogarithmic system. Differences between groups of data were analyzed by Student’s t test. P < 0.05 was considered significant.
Results As shown in Fig. 1, steady state plasma GH levels were established 90 min after the start of the GH infusion in both age groups. The steady state plasma GH level in each subject was calculated by averaging three consecutive plasma GH values at 90, 120, and 150 min. The mean steady state plasma GH levels were 11.8 f 2.7 pg/L in the young group and 13.6 f 3.6 pg/L in the aged group. The difference was not statistically significant. After stopping the GH infusion, plasma GH levels were monoexponentially decreased for 50 min on a semilogarithmic scale. Mean urinary GH values before the GH infusion was 0.74 f 0.85 ng/mmol creatinine in the young group and 0.83 + TABLE MCR,
1. Characteristics and calculated renal clearance (Cc,), and half-life (tin) of GH in healthy young and aged men Young men 24.6 62.4 1.75 20.7 4.2 127.5 3.2 187.1 2.9 105.5
f&e W BW (kg) Body surface area (m”) Body mass index (kg/m’) CGH bUmin) C,, (mL/min) CCH/Ccr (xlo-5) MCR (mL/min) MCR/BW (ml/min. ke) MCRjbody‘surface a& (mL/min . m”) tl,+ (min) f12MG clearance &L/min) Values
are the mean
7 + f + f k + + + + +
7.1 3.8 0.10 1.8 1.0 29.6 1.8 43.7 0.6 22.3
13.8 + 2.3 56.4 f 47.8 f
SD.
Aged men 66.2 52.5 1.54 20.2 14.3 115.6 13.1 120.5 2.3 79.0
5 + f + + f & f + + +
P
5.7 6.9 0.14 1.3 2.5 25.4 5.6 39.5 0.8 27.5
Vol. 75, No. 6 Printed in U.S.A.
Renal Clearance, Metabolic Clearance Rate, and HalfLife of Human Growth Hormone in Young and Aged Subjects* MOT01
SOHMIYA
First Division,
YUZURU
AND
Department
of
Medicine,
KATO Shimane
Medical
University,
ABSTRACT We studied the renal clearance, MCR, and half-life of the synthetic 22K form of human GH [recombinant hGH (rhGH)] in seven healthy young and five aged men by means of the constant iv infusion technique. rhGH was infused at a rate of 2 pg/kg ‘2 mL/h for 150 min, after which its disappearance was followed for 50 min. Changes in GH levels in plasma and urine were measured by a highly sensitive enzyme immunoassay. The mean (&SD) renal clearance of GH was significantly greater in the aged group than in the young group (14.3 f 2.5 vs. 4.2 +
Izumo
693, Japan
1.0 FL/min; P < 0.05). The mean MCR was greater in the young group than in the aged group (187.1 f 43.7 us. 120.5 + 39.5 mL/min; P < 0.05), but the MCR adjusted for body weight was not different between the two groups (2.9 f 0.6 us. 2.3 + 0.8 mL/min. kg). No difference was noted in the half-life of GH between the two groups (13.8 f 0.9 us. 14.2 + 0.4 min). These findings indicate that GH reabsorption from the renal tubule may be impaired in elderly subjects, but the disappearance rate of GH is not influenced by age. (J Clin Endocrinol Metab 75: 1487-1490,1992)
C
IRCULATING GH levels are maintained by pituitary GH production, tissuemetabolism, and renal excretion. GH secretion induced by exogenous GH-releasing hormone (GHRH) (1, 2) and insulin hypoglycemia (3) as well as 24-h integrated GH secretion (4-6) are reduced in aged subjects. It was reported that the MCR of GH in adults was greater than that in children (7), although other studies showed no difference in MCR between children and adults (8) or among various age groups in adults (9). Blood GH is filtered into urine at the glomerulus and mostly reabsorbed at the renal tubule (10). However, urinary GH levels were not precisely detectable until the recent introduction of a highly sensitive enzyme immunoassay (11, 12). It is now recognized that urinary GH levels reflect plasma GH concentrations in normal subjects and patients with pituitary disorders (12, 13). It was also reported that urinary GH levels were influenced by the renal function and positively correlated with urinary µglubulin (&MG) and negatively correlated with creatinine clearance (C,,) (14). We previously reported that urinary GH levels were not decreased,but, rather, were increased in aged subjects (15), in whom C,, is decreased and urinary &MG is increased. There are no published data on the renal clearance of GH (C,,) in which the effect of age is examined. In the present study we studied the renal clearance, MCR, and half-life of recombinant human GH (rhGH) in young and aged male subjects. Received October 15, 1991. Address all correspondence and requests for reprints to: Motoi Sohmiya, M.D., First Division, Department of Medicine, Shimane Medical University, Izumo 693, Japan. *This work was supported in part by grants from the Ministry of Education, Science, and Culture, Japan, and the Ministry of Health and Welfare, Japan. A part of the study was presented at the 73rd Annual Meeting of The Endocrine Society, Washington, D.C., June 1991.
Materials
and Methods
Subjects Twelve healthy male subjects, aged 17-76 yr, were included in the present study. They were divided into two groups based on age: a young group (n = 7) with a mean age of 24.6 yr (range, 17-34 yr), and an aged group (n = 5) with a mean age of 66.2 yr (range, 60-76 yr). As shown in Table 1, body weight and body surface area were slightly decreased in aged subjects, but there was no difference in body mass index between the young and aged groups. Renal function, assessed by urinalysis, serum urea nitrogen, serum creatinine, and urinary &MG levels, was within normal limits in all subjects. Informed consents were obtained from all subjects.
Protocol All studies were performed in the morning after overnight fasting in the supine position. After complete voiding of the urine, 500 mL tap water were given orally. Then, an indwelling catheter was placed into a cubital vein for infusion of the 22K form of rhGH (Sumitomo Co., Osaka, Japan). The rhGH was dissolved in 0.9% saline containing 0.3% human albumin and constantly infused for 150 min at a rate of 2 peg/ kg. 2 mL/h using a continuous infusion pump (Atom syringe infusion pump 235, Atom Co., Tokyo, Japan) with a precision of +3.0%. Blood samples were withdrawn through a second iv catheter, placed into the other cubital vein, at intervals of 30 min during the GH infusion and every 5 min after the end of the GH infusion. Urine samples were obtained immediately before and 90 and 150 min after the start of the GH infusion. Blood samples were immediately centrifuged at 4 C, and plasma was kept frozen at -20 C until assayed. Urine samples were added to 0.01 vol 0.01 M sodium phosphate buffer, pH 7.0, containing 10% BSA and 10% NaNa, dialyzed overnight against 0.01 M sodium phosphate buffer, pH 7.0, containing 0.1% NaN3 and 0.1 M NaCl, and kept at 4 C until assayed. All samples were assayed within 1 week.
Assays GH levels in plasma and dialyzed urine were measured in duplicate by a highly sensitive sandwich enzyme immunoassay, using a slight modification of the method described previously (14). For the measurement of GH in plasma and urine samples, anti-GH immunoglobulin Gcoated polystyrene balls were incubated with 100 PL GH standards or 1487
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 06 October 2015. at 02:55 For personal use only. No other uses without permission. . All rights reserved.
1488
SOHMIYA
50. to lOO-PL samples and 50-100 PL buffer in duplicate at 37 C for 6 h with continuous shaking. After removal of the supernatant, the polvstvrene balls were washed twice with 2 mL 150 mM saline and incubated with affinity-purified Fab’-peroxidase conjugate and 150 pg normal rabbit F(ab’h at 4 C for 20 h. then at 20 C for 6 h. After removal of the supernantant, the polystyrene balls were washed with 2 mL saline, incubated with 1 mL saline at 30 C for 10 min, and then washed with 2 mL saline. The balls were moved into a new series of assay tubes and further incubated with 100 WL 0.6 mg 3-(p-hydroxyphenyl)propionic acid as substrate and 50 PL 0.015% Hz02 at 30 C for 90 min. Fluorescence intensity was measured by spectrofluorophotometer (Shimazu RF-5000, Shimazu Co., Kyoto, Japan). The minimal detectable quantity was 0.3 rig/L. The intra- and interassay coefficients of variation were 6.0% and 9.8%, respectively. Creatinine and &MG levels in plasma and urine samples were determined by an autoanalyzer and specific RIA (& RIA Beads, Dainabott Co., Tokyo, Japan), respectively.
Calculations
and data analysis
Ccu was calculated by the following formula: Cc” = (UV/P) X (1.731 A), where U is the urinary GH level in the 150 min urine sample (micrograms per L), I’ is the mean plasma GH at 90, 120, and 150 min (micrograms per L), V is the urine volume from 90-150 min (milliliters per min), and A is the body surface area (square meters; 1.73 m2 is the standard body surface area). The MCR of GH was calculated as follows by the general formula of Tait (16): infusion rate of GH @g/h)/steady state plasma GH (pg/L). The half-life of GH (t& was calculated by means of linear regression analysis (17) of GH levels from 150-200 min in the semilogarithmic system. Differences between groups of data were analyzed by Student’s t test. P < 0.05 was considered significant.
Results As shown in Fig. 1, steady state plasma GH levels were established 90 min after the start of the GH infusion in both age groups. The steady state plasma GH level in each subject was calculated by averaging three consecutive plasma GH values at 90, 120, and 150 min. The mean steady state plasma GH levels were 11.8 f 2.7 pg/L in the young group and 13.6 f 3.6 pg/L in the aged group. The difference was not statistically significant. After stopping the GH infusion, plasma GH levels were monoexponentially decreased for 50 min on a semilogarithmic scale. Mean urinary GH values before the GH infusion was 0.74 f 0.85 ng/mmol creatinine in the young group and 0.83 + TABLE MCR,
1. Characteristics and calculated renal clearance (Cc,), and half-life (tin) of GH in healthy young and aged men Young men 24.6 62.4 1.75 20.7 4.2 127.5 3.2 187.1 2.9 105.5
f&e W BW (kg) Body surface area (m”) Body mass index (kg/m’) CGH bUmin) C,, (mL/min) CCH/Ccr (xlo-5) MCR (mL/min) MCR/BW (ml/min. ke) MCRjbody‘surface a& (mL/min . m”) tl,+ (min) f12MG clearance &L/min) Values
are the mean
7 + f + f k + + + + +
7.1 3.8 0.10 1.8 1.0 29.6 1.8 43.7 0.6 22.3
13.8 + 2.3 56.4 f 47.8 f
SD.
Aged men 66.2 52.5 1.54 20.2 14.3 115.6 13.1 120.5 2.3 79.0
5 + f + + f & f + + +
P
5.7 6.9 0.14 1.3 2.5 25.4 5.6 39.5 0.8 27.5
