J. E ndocrinol. Invest. 13: 1 -7, 1990
Effect of hCG or hCG + treatments in young thalassemic patients with hypogonadotropic hypogonadism R. Balducci*, V. Toscano**, G. Finocchi*, G. Municchi*, A. Mangiantini*, and B. Boscherini* * Clinica Pediatrica 2a Universita di Roma "Tor Vergata" and ** V Clinica Medica, Universita di Roma
"La Sapienza", 00100 Rome, Italy. ABSTRACT. Hypogonadotropic hypogonadism (HH) is common (40%) in beta-thalassemic patients. Taking into consideration that in HH non-thalassemic patients we obtained good results in pubertal development using hCG treatment (1500 IU every 6 days), 10 HH thalassemic subjects (14 5/12 -17 yr, all with bone age greater than 13 6/12) were treated with the same regimen. In 5 of these patients purified FSH (75 IU every 3 days) was added to hCG in order to evaluate the FSH effect on testosterone (T) response (Group 1 was given hCG alone, Group 2 hCG + FSH: Profasi HP and Metrodin Serono). To evaluate the kinetics of testosterone response, plasma level of Twas determined basally and 1, 2, 4 and 6 days after hCG injection. This dynamic study and a clinical examination were carried out at the beginning of treatment and at the 4th and 12th month after. Results obtained in the first group confirmed our previous data from non-thalassemic HH patients: in fact, after 12 months of therapy a stage G2-G3
was reached. In the second group, however, testis size and testosterone secretion were significantly higher than in the first group. At the 4th month, in Group 1 and in Group 2 testis size and the area under the T response curve were 3.18 ± 0.18ml and 13,364 ± 1047 ngldl vs 4.62 ± 0.25 ml (p < 0.02) and 18,045 ± 1110 ng I dl (p < 0.016), respectively. At the 12th month testis size and area under T response curve were increased to 6.12 ± 0.25 ml and 24,017 ± 1176 ng I dl in Group 1 and 8.12 ± 0.6 ml (p < 0.01) and 33,924 ± 2181 ngldl (p < 0.01) in Group 2. After 12 months, a significant comparable increase in growth velocity, without bone age acceleration, was observed in both groups. Our results demonstrate that hCG or hCG + FSH treatments are able to induce a satisfactory sexual development in thalassemic HH patients; moreover FSH in addition to hCG seems to improve not only testis size as expected, but also testosterone production.
INTRODUCTION Life expectancy for patients affected by Thalassemia Major has greatly improved with more intensive transfusion regimens and sc administration of chelant therapy (1-3). The expectations of many thalassemic adolescents, therefore, are now the same as those of their healthy peers with regards to education, employment and a well adjusted social and sexual life. The absence of pubertal development represents one of the major obstacles to the fulfillment of these hopes. Abnormal sexual maturation is present in 79% of boys, and complete
absence of pubertal development is present in at least 40% (4). This hypogonadism is due to a pituitary abnormality, caused by iron deposits (5-14). Pituitary is very sensitive to iron toxicity and it is possible that even small amounts of iron may produce irreversible damage (4). Therefore gonadotropins appear to be the best possible therapy in pubertal induction and sexual development, and have to be preferred to testosterone therapy because of its hepatotoxycity (15-17). The aim of the present investigation was to study the effect of two different regimens of gonadotropin therapy (hCG alone or hCG + FSH) to induce pubertal development in thalassemic HH patients. The choice of the first regimen of treatment (1500 IU Profasi HP Serono every 6 days) was based both on the results obtained in adult subjects (18, 19) showing that hCG alone is able to induce secondary
Key-words. Hypogonadotropic hypogonadism. thalassemia. hCG. FSH. Correspondence 10.' Dr. Vincenzo Toscano. Istituto di V Clinica Medica. PollcllnlCO Umberto I. Universita La Sapienza. 00161 Roma. Italy.
Received March 24. 1989. accepted July 24. 1989.
R. Balducci, V. Toscano, G. Finocchi, et al.
At the start of the study none of the patients presented other endocrine abnormalities and only one (patient no. 9) developed symptomatic diabetes mellitus. The subjects were on a hypertransfusion protocol designed to maintain hemoglobin at a level greater than 11 g/ dl. Transfusions were administered approximately at 4-wk intervals. All patients had been receiving sc deferoxamine mesylate therapy for 2 to 6 5/12 yr beginning at a median age of 5.1 yr. Six of the subjects had undergone splenectomy at a median age of 9.6 yr. The general health of the patients was good and none ot them was underweight for height. The stature at the time of the study was -1.8 ± 0.4 SD (M ± SE). The alanine aminotransferase and the aspartate aminotransferase levels were elevated in 5 of the 8 subjects (ranging from 53 to 128 U/L; normal values ~ 40 U/L). Bilirubin and alkaline phosphatase were within the normal range. Serum ferritin levels were elevated in all subjects (ranging from 500 to 2959 ng/dl; normal values 18-440 ng/ dl).
sex characteristics and on our previous results obtained in hypogonadotropic hypogonadism of other origins (20). The administration of hCG every 6 days instead of the commonly adopted 2-3 day regimen was chosen, since plasma testosterone response to a single im injection of hCG is prolonged over 72 h in prepubertal boys (21 r25), as well in HH patients (26-29). The second regimen (hCG every 6 days and purified FSH every 3 days) was chosen in order to estimate the effect of FSH on testicular response to hCG. It is well known that FSH in addition to its mandatory role in the achievement of a full spermatogenesis, improves in animals the action of LH on testosterone production (3033). MATERIALS AND METHODS Patients The study included 10 thalassemic subjects (14 5/12-17 yr, all with bone age greater than 136/12) with hypogonadism secondary to insufficient gonadotropin secretion. The diagnosis of gonadotropin deficiency was established on the basis of prepubertal testis size (2.12 ± 0.18 ml, M ± SE) and hormone levels and the subsequent finding of subnormal gonadotropin response to GnRH (Table 1). All patients had control visits for at least one yr before starting treatment to confirm subnormal testicular growth.
Study design The patients were assigned at random to one of the two groups. Patients 1,2,3,4 and 5 (Group 1) were given 1500 IU of hCG (Profasi HP Serono Italy) at 08:00 h once every 6 days for 12 months. Patients 6, 7, 8, 9 and 10 (Group 2) were given 1500 IU of hCG once every 6 days and 75 IU of purified FSH
TABLE 1 - Summary of pertinent clinical features of subjects selected for evaluation. Patients (No.)
Chron.
Age Bone (GP)
Height (SOS)
Mean Testis size (ml)
Pubic hair stage
Testosterone*' basal (ng/dl)
LH (mIU/ml) peakbasal
FSH (mIU/ml) peakbasal
GROUP 1 1
154/12
136/12
- 1.9
2
15.2
3.5
6.6
2.6
2
147/12
136/12
- 1.4
1.5
24
2.2
4.6
3.1
4.3
3
147/12
14
- 1.9
2.5
2
13.5
1.5
6.5
1.5
5.5
4
17
14
- 2.5
2
2
25
2.8
4.4
0.5
2.7
5
17
14
-2
2.5
2
10
1.5
1.5
1.5
1.5
6
15
136/12
- 1.4
3
22
3
5.6
3.3
3.9
7
154/12
136/12
- 1.8
2
27
2.5
5.9
0.5
3.8
8
17
14
- 2.2
2.5
24
3
7.4
1.5
4
9
145/12
136/12
- 1.4
1.5
19
1.5
4.5
1.5
2.5
10
164/12
14
-2
2
20
3
6.4
2
2.7
2.8
GROUP 2
2
- After GnRH (100 pg iv). Normal testosterone values (mean ± SE), in our laboratory. in different gonadal stages: Gl = 19 G2-G3 = 116 ± 18 ng/dl: G3-G4 = 164 ± 13.7 ng/dl: Adult = 697 ± 159 ng/dl.
*-
2
±
3.7 ng/dl: G2 = 78
±
9.2 ng/dl:
hCG or hCG + FSH in thalassemic patients
(Metrodin, Serono, Italy) once every 3 days (the FSH was mixed with hCG in order to give only one injection). In order to evaluate the kinetics of testicular response, blood levels of testosterone (T) were determined basally and 1, 2, 4 and 6 days after hCG or hCG + FSH administration. This dynamic study and a clinical examination (including measurement of testicular volume using the Prader orchidometer) were carried out at the beginning, at the 4th and the 12th month of treatment. The study was always performed at least 20 days after administration of a transfusion. The medication was usually self-injected at home, according to a well established schedule and under our constant supervision.
the statistical significance of the results obtained in basal conditions compared to results obtained at each collection time (1, 2, 4 and 6 days after hCG) and compared to basal values (those obtained immediately before hCG injection) at the start and in the 4th and 12th month of treatment. Student's t test was also used to evaluate the statistical significance between the areas evaluated under T responsecurve. This statistical analysis was performed for each group and a comparison was made between the two groups (pC Statistician-Statistical Analysis Software-Human Statistical Oynamics-NorthridgeCA). RESULTS Results are reported in Figures 1 -3.
Hormone measurement Plasma was immediately stored at -20 C until assayed. After ether extraction of 1 ml of plasma, T was measured using Radioimmunoassay after microcolumn Celite (SOH 535) chromatography (with eluting solvent isooctane: benzene 60:40). The specific antibody for T was purchased from Sorin (Saluggia, Vercelli , Italy). The interassay and intraassay coefficients of variation were 8.2% and 2.5%, respectively. The sensitivity of the method was 2 pg/tube. Area under T response curve was geometrically evaluated on each patient.
At the start of treatment Group 1: the mean (± SE) plasma testosterone concentration was 19.1 ± 2.3 ng/dl. After hCG the increase of T level was significant reaching the highest value of 111.2 ± 20.1 ng/dl (p < 0.02) at 96 h. The mean area under T response curve was 5948 ± 694 ng/dl. Mean testicular size was 2 ml 0,2 and pubic hair 1-2. Group 2: the mean testosterone concentration was 22.4 ± 2.2. ng/ dl. After hCG + FSH the increase of T level was significant reaching the highest value of 132.8 ± 9.1 ng/dl (p < 0.001). The mean area under T response curve was 7880. 4 ± 428 ng/dl. Mean testicular size was 2.25 ± 0.32 ml and pubic hair 1-2.
±
Statistical analysis Student's t test for paired data was used to evaluate
"" t
rES lOS TEROtU:
4th _nth
-
D
G""up 1
Fig. 1 - Kinetics of testosterone (T) response (mean ± SE) in Group 1 (hCG alone) and in Group 2 (hCG + ..
1
2
4
,
..
1
2
4
,
..
1
d"IIS
3
2
4
,
FSH). For statistical analysis see text.
R. Balducci, V. Toscano, G. Finocchi, et al.
r ...... 5.
J_ a c..u,
.
Effect of hCG or hCG + treatments in young thalassemic patients with hypogonadotropic hypogonadism R. Balducci*, V. Toscano**, G. Finocchi*, G. Municchi*, A. Mangiantini*, and B. Boscherini* * Clinica Pediatrica 2a Universita di Roma "Tor Vergata" and ** V Clinica Medica, Universita di Roma
"La Sapienza", 00100 Rome, Italy. ABSTRACT. Hypogonadotropic hypogonadism (HH) is common (40%) in beta-thalassemic patients. Taking into consideration that in HH non-thalassemic patients we obtained good results in pubertal development using hCG treatment (1500 IU every 6 days), 10 HH thalassemic subjects (14 5/12 -17 yr, all with bone age greater than 13 6/12) were treated with the same regimen. In 5 of these patients purified FSH (75 IU every 3 days) was added to hCG in order to evaluate the FSH effect on testosterone (T) response (Group 1 was given hCG alone, Group 2 hCG + FSH: Profasi HP and Metrodin Serono). To evaluate the kinetics of testosterone response, plasma level of Twas determined basally and 1, 2, 4 and 6 days after hCG injection. This dynamic study and a clinical examination were carried out at the beginning of treatment and at the 4th and 12th month after. Results obtained in the first group confirmed our previous data from non-thalassemic HH patients: in fact, after 12 months of therapy a stage G2-G3
was reached. In the second group, however, testis size and testosterone secretion were significantly higher than in the first group. At the 4th month, in Group 1 and in Group 2 testis size and the area under the T response curve were 3.18 ± 0.18ml and 13,364 ± 1047 ngldl vs 4.62 ± 0.25 ml (p < 0.02) and 18,045 ± 1110 ng I dl (p < 0.016), respectively. At the 12th month testis size and area under T response curve were increased to 6.12 ± 0.25 ml and 24,017 ± 1176 ng I dl in Group 1 and 8.12 ± 0.6 ml (p < 0.01) and 33,924 ± 2181 ngldl (p < 0.01) in Group 2. After 12 months, a significant comparable increase in growth velocity, without bone age acceleration, was observed in both groups. Our results demonstrate that hCG or hCG + FSH treatments are able to induce a satisfactory sexual development in thalassemic HH patients; moreover FSH in addition to hCG seems to improve not only testis size as expected, but also testosterone production.
INTRODUCTION Life expectancy for patients affected by Thalassemia Major has greatly improved with more intensive transfusion regimens and sc administration of chelant therapy (1-3). The expectations of many thalassemic adolescents, therefore, are now the same as those of their healthy peers with regards to education, employment and a well adjusted social and sexual life. The absence of pubertal development represents one of the major obstacles to the fulfillment of these hopes. Abnormal sexual maturation is present in 79% of boys, and complete
absence of pubertal development is present in at least 40% (4). This hypogonadism is due to a pituitary abnormality, caused by iron deposits (5-14). Pituitary is very sensitive to iron toxicity and it is possible that even small amounts of iron may produce irreversible damage (4). Therefore gonadotropins appear to be the best possible therapy in pubertal induction and sexual development, and have to be preferred to testosterone therapy because of its hepatotoxycity (15-17). The aim of the present investigation was to study the effect of two different regimens of gonadotropin therapy (hCG alone or hCG + FSH) to induce pubertal development in thalassemic HH patients. The choice of the first regimen of treatment (1500 IU Profasi HP Serono every 6 days) was based both on the results obtained in adult subjects (18, 19) showing that hCG alone is able to induce secondary
Key-words. Hypogonadotropic hypogonadism. thalassemia. hCG. FSH. Correspondence 10.' Dr. Vincenzo Toscano. Istituto di V Clinica Medica. PollcllnlCO Umberto I. Universita La Sapienza. 00161 Roma. Italy.
Received March 24. 1989. accepted July 24. 1989.
R. Balducci, V. Toscano, G. Finocchi, et al.
At the start of the study none of the patients presented other endocrine abnormalities and only one (patient no. 9) developed symptomatic diabetes mellitus. The subjects were on a hypertransfusion protocol designed to maintain hemoglobin at a level greater than 11 g/ dl. Transfusions were administered approximately at 4-wk intervals. All patients had been receiving sc deferoxamine mesylate therapy for 2 to 6 5/12 yr beginning at a median age of 5.1 yr. Six of the subjects had undergone splenectomy at a median age of 9.6 yr. The general health of the patients was good and none ot them was underweight for height. The stature at the time of the study was -1.8 ± 0.4 SD (M ± SE). The alanine aminotransferase and the aspartate aminotransferase levels were elevated in 5 of the 8 subjects (ranging from 53 to 128 U/L; normal values ~ 40 U/L). Bilirubin and alkaline phosphatase were within the normal range. Serum ferritin levels were elevated in all subjects (ranging from 500 to 2959 ng/dl; normal values 18-440 ng/ dl).
sex characteristics and on our previous results obtained in hypogonadotropic hypogonadism of other origins (20). The administration of hCG every 6 days instead of the commonly adopted 2-3 day regimen was chosen, since plasma testosterone response to a single im injection of hCG is prolonged over 72 h in prepubertal boys (21 r25), as well in HH patients (26-29). The second regimen (hCG every 6 days and purified FSH every 3 days) was chosen in order to estimate the effect of FSH on testicular response to hCG. It is well known that FSH in addition to its mandatory role in the achievement of a full spermatogenesis, improves in animals the action of LH on testosterone production (3033). MATERIALS AND METHODS Patients The study included 10 thalassemic subjects (14 5/12-17 yr, all with bone age greater than 136/12) with hypogonadism secondary to insufficient gonadotropin secretion. The diagnosis of gonadotropin deficiency was established on the basis of prepubertal testis size (2.12 ± 0.18 ml, M ± SE) and hormone levels and the subsequent finding of subnormal gonadotropin response to GnRH (Table 1). All patients had control visits for at least one yr before starting treatment to confirm subnormal testicular growth.
Study design The patients were assigned at random to one of the two groups. Patients 1,2,3,4 and 5 (Group 1) were given 1500 IU of hCG (Profasi HP Serono Italy) at 08:00 h once every 6 days for 12 months. Patients 6, 7, 8, 9 and 10 (Group 2) were given 1500 IU of hCG once every 6 days and 75 IU of purified FSH
TABLE 1 - Summary of pertinent clinical features of subjects selected for evaluation. Patients (No.)
Chron.
Age Bone (GP)
Height (SOS)
Mean Testis size (ml)
Pubic hair stage
Testosterone*' basal (ng/dl)
LH (mIU/ml) peakbasal
FSH (mIU/ml) peakbasal
GROUP 1 1
154/12
136/12
- 1.9
2
15.2
3.5
6.6
2.6
2
147/12
136/12
- 1.4
1.5
24
2.2
4.6
3.1
4.3
3
147/12
14
- 1.9
2.5
2
13.5
1.5
6.5
1.5
5.5
4
17
14
- 2.5
2
2
25
2.8
4.4
0.5
2.7
5
17
14
-2
2.5
2
10
1.5
1.5
1.5
1.5
6
15
136/12
- 1.4
3
22
3
5.6
3.3
3.9
7
154/12
136/12
- 1.8
2
27
2.5
5.9
0.5
3.8
8
17
14
- 2.2
2.5
24
3
7.4
1.5
4
9
145/12
136/12
- 1.4
1.5
19
1.5
4.5
1.5
2.5
10
164/12
14
-2
2
20
3
6.4
2
2.7
2.8
GROUP 2
2
- After GnRH (100 pg iv). Normal testosterone values (mean ± SE), in our laboratory. in different gonadal stages: Gl = 19 G2-G3 = 116 ± 18 ng/dl: G3-G4 = 164 ± 13.7 ng/dl: Adult = 697 ± 159 ng/dl.
*-
2
±
3.7 ng/dl: G2 = 78
±
9.2 ng/dl:
hCG or hCG + FSH in thalassemic patients
(Metrodin, Serono, Italy) once every 3 days (the FSH was mixed with hCG in order to give only one injection). In order to evaluate the kinetics of testicular response, blood levels of testosterone (T) were determined basally and 1, 2, 4 and 6 days after hCG or hCG + FSH administration. This dynamic study and a clinical examination (including measurement of testicular volume using the Prader orchidometer) were carried out at the beginning, at the 4th and the 12th month of treatment. The study was always performed at least 20 days after administration of a transfusion. The medication was usually self-injected at home, according to a well established schedule and under our constant supervision.
the statistical significance of the results obtained in basal conditions compared to results obtained at each collection time (1, 2, 4 and 6 days after hCG) and compared to basal values (those obtained immediately before hCG injection) at the start and in the 4th and 12th month of treatment. Student's t test was also used to evaluate the statistical significance between the areas evaluated under T responsecurve. This statistical analysis was performed for each group and a comparison was made between the two groups (pC Statistician-Statistical Analysis Software-Human Statistical Oynamics-NorthridgeCA). RESULTS Results are reported in Figures 1 -3.
Hormone measurement Plasma was immediately stored at -20 C until assayed. After ether extraction of 1 ml of plasma, T was measured using Radioimmunoassay after microcolumn Celite (SOH 535) chromatography (with eluting solvent isooctane: benzene 60:40). The specific antibody for T was purchased from Sorin (Saluggia, Vercelli , Italy). The interassay and intraassay coefficients of variation were 8.2% and 2.5%, respectively. The sensitivity of the method was 2 pg/tube. Area under T response curve was geometrically evaluated on each patient.
At the start of treatment Group 1: the mean (± SE) plasma testosterone concentration was 19.1 ± 2.3 ng/dl. After hCG the increase of T level was significant reaching the highest value of 111.2 ± 20.1 ng/dl (p < 0.02) at 96 h. The mean area under T response curve was 5948 ± 694 ng/dl. Mean testicular size was 2 ml 0,2 and pubic hair 1-2. Group 2: the mean testosterone concentration was 22.4 ± 2.2. ng/ dl. After hCG + FSH the increase of T level was significant reaching the highest value of 132.8 ± 9.1 ng/dl (p < 0.001). The mean area under T response curve was 7880. 4 ± 428 ng/dl. Mean testicular size was 2.25 ± 0.32 ml and pubic hair 1-2.
±
Statistical analysis Student's t test for paired data was used to evaluate
"" t
rES lOS TEROtU:
4th _nth
-
D
G""up 1
Fig. 1 - Kinetics of testosterone (T) response (mean ± SE) in Group 1 (hCG alone) and in Group 2 (hCG + ..
1
2
4
,
..
1
2
4
,
..
1
d"IIS
3
2
4
,
FSH). For statistical analysis see text.
R. Balducci, V. Toscano, G. Finocchi, et al.
r ...... 5.
J_ a c..u,
.
