Post-Transplant Events

Gonadal shielding to irradiation is effective in protecting testicular growth and function in long-term survivors of bone marrow transplantation during childhood or adolescence

Abstract

An increasing number of long-term surviving bone marrow transplantation (BMT) recipients have recovered from their primary disease but are at risk of developing failure of endocrine organs. We investigated 30 recipients who underwent allogeneic BMT during childhood or adolescence. Testicular growth and function were evaluated by serial measurement of testicular volume, basal luteinizing hormone (LH), basal follicle-stimulating hormone (FSH) and testosterone levels and by gonadotropin-releasing hormone (GnRH) provocative test. Puberty started spontaneously in all patients. However, all except four patients had normal testosterone levels with elevated LH, indicating partial Leydig cell dysfunction. Standard deviation scores of testicular volume at last evaluation were statistically lower in those who had received irradiation without gonadal shield compared to those with (−2.04±0.45 vs −0.30±1.17, respectively, P<0.005), suggesting damage of testicular germinal epithelium owing to gonadal irradiation. Serial measurement of testicular volume showed a tendency of growth to stop at 10 ml in those without gonadal shield. Among the 30 patients, only one patient has fathered a child after reaching spontaneous puberty. These results suggest that gonadal shield is effective to protect testicular growth and function, although the attainment of fertility is difficult to achieve.

Introduction

Large numbers of children with malignant and non-malignant disease have recovered from their primary disease after bone marrow transplant (BMT) and now are surviving long term. However, they are at risk of developing failure of several important organs as a result of toxic effects of the intensive treatment before, during and after BMT.1, 2 Late effects and complications are heterogeneous, and although often not life threatening, they significantly impair the quality of life of survivors.3 Among the endocrine complications following BMT that have been described,4 testicular dysfunction frequently occurs in male long-term survivors.5

Testicular dysfunction occurs both in adolescence and in adult following BMT, mainly owing to chemotherapy6 and/or total body irradiation (TBI) in conditioning regimen,7 and it results in incomplete sexual development at puberty. Recipients transplanted during childhood usually experience spontaneous pubertal development and completion of puberty. However, they might not achieve a normal testicular volume as a result of damage to the testicular germinal epithelium.7 The intensive treatment before BMT may have a direct toxic effect on the Leydig cell, but there is also some evidence that the damage to the germinal epithelium and reduction in testicular volume may affect the Leydig cell function by causing structural changes within the testis.6

Recovery of the testicular function has been reported in less than 30% of patients, and use of increasing doses of TBI without gonadal shield may be associated considerably with lower recovery.8, 9, 10 Only limited data are available on the long-term natural history of BMT-induced testicular dysfunction.

Patients and methods

Patients

We investigated 30 recipients who underwent allogeneic BMT at Tokai University Hospital between 1982 and 1997, and survived at least 7 years after BMT. They were older than 15 years at the time of the last visit and had no history of testicular dysfunction before BMT. At the time of BMT, written informed consent was obtained from either recipients or their parents for the treatment procedure and follow-up. Patient characteristics are shown in Table 1. The median age at BMT was 10.5 years (range 0.9–15.8 years), and the median follow-up duration after BMT was 13.3 years (range 7.6–21.2 years).

Table 1 Patient characteristics

Transplantation procedure

Transplant conditioning regimens are shown in Table 1. Conditioning regimens for 25 patients consisted of irradiation combined with/without cyclophosphamide and/or other drugs. Six-to-12 Gy of TBI was given in 3–6 fractions, and 3–10 Gy of thoraco-abdominal irradiation (TAI) with gonadal shield in one-five fractions. Radiation field was limited to the thoraco-abdominal lymphoid area to reduce regimen-related toxicity in nonmalignant disease group (aplastic anemia, Wiskott–Aldrich syndrome, Gaucher disease, Hurler disease and adrenoleukodystrophy). Furthermore, gonadal area including the testes was shielded in an attempt to prevent testicular damage in recipients receiving TAI+chemotherapy.

Five patients with nonmalignant disease received conditioning without irradiation. The Irradiation (+) Gonadal shield (−) group, the Irradiation (+) Gonadal shield (+) group and the Irradiation (−) group were designated as group A, group B and group C, respectively. Graft-versus-host disease (GVHD) prophylaxis comprised methotrexate and/or cyclosporine.

A 13-year-old boy (UPN 212) with chronic myelogenous leukemia (CML) in the first chronic phase received human leukocyte antigens (HLA)-matched BMT after TBI with chemotherapy-based conditioning regimen in 1996. His gonadal area was shielded from TBI, because we expected that the risk of leukemic relapse in the testes was very low in this case.

Evaluation of pubertal development

Pubertal development was carefully and serially observed and recorded. Onset of puberty was defined by a testicular volume 4 ml.11 Testicular volume was determined using an orchidometer, as described by Prader.12 Testicular measurement using orchidometer was performed by a single investigator (SK). In the supine position, testes were gently held by the fingers of the hand with the scrotal skin smooth but not compressing the testes. Testicular volumes at last evaluation in each patient were evaluated by analyzing with the standard deviation (SDS) of testicular volume according to each age, which was calculated from the data in normal Japanese boys.13

Testicular Leydig cell function and germinal epithelium damage

Testicular Leydig cell function and germinal epithelium damage were evaluated before and annually after BMT by serial measurement of basal serum LH, basal serum FSH and serum testosterone levels. Normal basal serum LH and FSH values in our institute were <5 and <9 IU/l, respectively. In addition, all patients had gonadotropin releasing hormone (GnRH) stimulation test as part of endocrine evaluation after BMT. A dose of 100 μg/m2 body surface area of LH-RH (maximum 100 μg) was given intravenously, and serum LH and serum FSH were measured before and at +30, +60, +90 and +120 min. Normal response to GnRH stimulation should not exceed 30 IU/l in peak serum LH and peak serum FSH according to the data in healthy children who were referred to our pediatric endocrinology clinics for endocrine examination because of too short stature. All measurements were performed in the central laboratory of our hospital. Endocrine tests were undertaken in the morning to avoid diurnal variation of hormones.

Definition of testicular dysfunction

Normal testicular function was defined by the occurrence of spontaneous pubertal development and normal gonadotropin levels. Partial Leydig cell dysfunction was defined by normal serum testosterone levels with increased basal serum LH levels (>15 IU/l) or with elevated peak LH (>40 IU/l) by provocative test; complete Leydig cell dysfunction was defined by low serum testosterone levels with increased serum LH levels or elevated peak LH. Testosterone/LH ratio was also used to assess Leydig cell function.14 Germinal epithelium damage was defined by a partial or complete absence of any increase in testicular volume at pubertal ages, and/or increased basal serum FSH levels (>20 IU/l) or elevated peak FSH level (>40 IU/l).

Testicular biopsy

Testicular biopsy was performed in six patients who received BMT between 1985 and 1990 for evaluation of histological testicular structure. It was performed by open biopsy under general anesthesia.

Statistical analysis

Because the data had a skewed distribution, median and range were used throughout the text, tables and figures. Only for analysis of testicular volumes at last evaluation, SDS of testicular volume was used. The Mann–Whitney test and the Kruskal–Wallis test were used to compare the differences between groups and among groups, respectively. The χ2 test and Fisher's exact probability test were used to assess the association between groups. The results are presented in Box and Whisker plots in the Figure 4. These statistical analyses were carried out using the GraphPad PRISM statistical package. Values of P<0.05 were considered statistically significant.

Figure 4
figure4

Changes in basal FSH (top) and peak FSH (bottom) levels before and after BMT.

Results

Testicular volume

The median age of 30 patients at their last evaluation was 21.9 years (15.8–29.6 years). According to testicular volume, puberty started spontaneously in all patients irrespective of the presence or absence of irradiation in the conditioning for BMT. Changes in testicular volume are shown in Figure 1a. In this study, we focused on testicular growth and function according to the presence of testicular irradiation. Relationship between testicular volume and patient characteristics (sex, age at BMT, disease, testicular irradiation, HLA and GVHD) was evaluated. There was the strongest relationship between testicular volume and testicular irradiation (P<0.0005). Changes of testicular volume were sorted by type of irradiation (Figure 1b–d). Testicular volume at pubertal age (14 years old) is 16.4±4.6 ml according to the reports on testicular growth of Japanese boys from birth to adolescence in cross-sectional series.13 Minimum of the normal range of testicular volume in adulthood is equal to testicular volume at pubertal age.15 Therefore, we used a fixed value after pubertal age (14 years old) for evaluation of testicular volume. Testicular volume of less than 10 ml or less at last evaluation was defined as testicular dysgenesis. The differences in the incidence of testicular dysgenesis among three types of conditioning were statistically significant (Table 2, P<0.05 for group A vs group B and group C and P<0.005 for group A vs group B+C, respectively). In fact, group A showed a significantly smaller size in testicular volume SDS at last evaluation compared to group B and group C (P<0.005 for group A (−2.04±0.45) vs group B (−0.30±1.17); P<0.05 for group A and group C (−0.96±0.57) and P<0.0005 for group A vs group B+C (−0.96±1.01), respectively). It is interesting to note that serial measurement of testicular volume showed a tendency of growth to stop up to 10 ml in group A.

Figure 1
figure1

Changes in testicular volume after BMT. Connecting lines represent data from individuals. Dotted lines represent +2 s.d. (top) and −2 s.d. (bottom) of normal testicular volume in Japanese males, respectively.

Table 2 Testicular volume after BMT

Leydig cell dysfunction

In all patients except three (UPN 1, 18 and 120 in group A), serum testosterone reached adult level at some time points after BMT (data not shown). There was no association between serum testosterone levels and patient characteristics (age at BMT, primary disease, conditioning regimen and type of irradiation). However, serial examination of gonadotropin levels revealed a tendency of basal and peak LH to rise to Leydig cell dysfunctional level around 20 years of age in group A (Figure 2a, d). All patients (16/16) in group A, 6/9 patients in group B and 4/5 patients in group C experienced raised peak LH levels with normal serum testosterone levels indicating the presence of partial Leydig cell dysfunction at some time during the follow-up period (Table 3). The difference between group A and group B was significant (P<0.05), but the difference between group A and group C was not significant (P=0.24). Leydig cell dysfunction was also assessed by the Testosterone/LH ratio (Figure 3). Reduced Leydig cell function was substantiated from diminished testosterone/LH ratio (median ratio 41.5 in group A, 87.6 in group B, 82.1 in group C and 86.9 in group B+C. P<0.005 for group A vs group B, P=0.09 for group A vs group C and P<0.005 for group A vs group B+C, respectively).

Figure 2
figure2

Changes in basal LH (top) and peak LH (bottom) levels before and after BMT.

Table 3 Changes in LH and FSH levels after BMT
Figure 3
figure3

Comparison of the testosterone/LH ratio. The box contains the middle 50th percentile of the value. The lower and upper boundary of the box shows the 25th and the 75th percentile, respectively. The bottom and top of the bar show maximum and minimum values, respectively. Significant differences are shown by *P<0.005 (the Irradiation (+) Gonadal shield (−) group vs the Irradiation (+) Gonadal shield (+) group).

Germinal epithelium damage

Serial examination of gonadotropin levels also revealed a remarkable tendency of basal FSH over rise to germinal epithelium damage level in group A but less in group B or group C around 20 years of age (Figure 4a–c, Table 3). Fifteen out of sixteen patients in group A showed markedly to moderately raised peak FSH levels, whereas 3/9 in group B and 1/5 in group C experienced mildly raised peak FSH levels after puberty, indicating complete damage of testicular germinal epithelium in group A and partial damage in groups B and C (Figure 4d–f, Table 3, P<0.005 for group A vs group B and for group A vs group C and P<0.0005 for group A vs group B+C, respectively).

Testicular biopsy was performed to clarify relationship between endocrinological change and pathological change in only six patients (five in group A and one in group B) after 1 to 6 years post BMT, although number of biopsied recipients was limited. In group A, the testis showed atrophic seminiferous tubules and spermatogenesis was completely absent. No malignancy was noted. However, one group B patient showed well-developed seminiferous tubules with moderate number of spermatogonium.

Effect of testicular shielding from TBI

Although we experienced only one patient who is a 13-year-old boy (UPN 212) with CML in the first chronic phase received BMT after TBI with gonadal shielding, his testicular volume at latest evaluation (17.3 years old) was 20 ml, which is appropriate to a normal Japanese boy. His endocrinological evaluation showed partial Leydig cell dysfunction during follow-up period. However, basal and peak serum FSH levels increased to over 20 and 40 IU/l by 2 months after BMT, respectively, and then returned to the normal range spontaneously. These findings indicated that gonadal shielding to irradiation has possibilities to protect germinal epithelium function.

Fertility

Among the 30 patients, only one patient has fathered a child. A 12-year-old boy with severe aplastic anemia received HLA-matched BMT after TAI with testicular shielding/cyclophosphamide-based conditioning regimen in 1985. As reported previously, he fathered a healthy boy 7 years after BMT16 and paternity was proved by HLA and blood typing of the parents and the baby, which were performed with the informed consent from the parents. His son is healthy and 13 years old at present.

Discussion

Several late complications after BMT have been described,3, 4, 5 but only limited data on testicular growth and function are available in the recipients who were treated in childhood and adolescence. Recipients transplanted during childhood usually spontaneously start and complete puberty. The prognosis for normal pubertal development and testicular function in survivors of childhood leukemia treated with chemotherapy alone is excellent;17 however disturbances in pubertal development in survivors of BMT with TBI are likely to be the result of high-dose chemotherapy6 or of irradiation7 used in preparative regimens.

Although it is very difficult to define puberty in recipients who received BMT, testicular volume in all recipients was over 4 ml at latest evaluation irrespective of the presence or absence of irradiation in the conditioning for BMT and most patients have continued to produce age-appropriate serum testosterone levels. These findings indicate that puberty started spontaneously in all recipients. Our data are similar to the findings of Ogilvy-Stuart et al.,18 who reported normal puberty and testosterone levels in all boys who underwent BMT with TBI at a young age. Although Bakker et al.7 showed that testicular volume was small (mean 10.5 ml) even in adult after BMT only in recipients treated with TBI in childhood, our study examined the differences in the incidence of testicular dysgenesis among three types of conditioning. Testicular volume was significantly smaller in recipients treated with TBI compared to recipients treated with TAI or chemotherapy. In fact, recipients without testicular shield showed a significantly smaller size in testicular volume in adult (median 7 ml) compared to recipients with testicular shield (median 15 ml) or with chemotherapy alone (median 12 ml) as a preparative regimen. Serial measurement of testicular volume showed a tendency of growth to stop at 10 ml in the recipients with testicular irradiation. Endocrinologic evaluation also supported testicular dysgenesis in that only 20–30% of recipients with testicular shield or only chemotherapy experienced raised peak FSH levels, although all recipients without testicular shield except one experienced raised peak FSH levels indicating the presence of germinal epithelium damage. In clinical practice, testicular volume less than 10 ml according to an orchidometer is an important indicator of testicular dysgenesis. These results strongly indicate that testicular shielding from irradiation or chemotherapy alone is effective to retain the ability of testicular growth.

Leydig cell function is relatively resistant to chemotherapy, whereas testicular germinal epithelium is extremely sensitive to several classes of chemotherapeutic agents. Irradiation, on the other hand, can result in testicular germinal epithelium damage, as well as in Leydig cell damage if given in supralethal doses.19 The fact that majority of patients in this study experienced raised peak LH levels with age-appropriate serum testosterone production revealed that subclinical injury to the Leydig cell is common after BMT. Diminished testosterone/LH ratio also demonstrated that recipients without testicular shield subjected to Leydig cell dysfunction compared to recipients with testicular shield or chemotherapy only, and another investigator has found decreased responses to hCG stimulation in patients after BMT with TBI,20 which further supports our data.

The conditioning regimen before BMT will frequently produce infertility, especially if it includes TBI, and the disturbance of the gonadal function is permanent. Although overt Leydig cell failure was rare after BMT, two-thirds of patients treated with irradiation had raised serum levels of FSH, which is indicative of permanent germinal epithelium damage.21, 22 Our observation also showed that 90% of recipients without testicular shield had a high basal and peak FSH levels, although only 30% of recipients with testicular shield or chemotherapy only had germinal epithelium damage. In contrast to other reports,23, 24 we did not find normalization of FSH levels in our patients, even though most recipients received fractionated irradiation, which believed to be less detrimental to the testicular germinal epithelium compared to single-fraction irradiation. One possible explanation is that variations in the exposure to alkylating agents before BMT could be responsible for the difference in testicular recovery, as the cumulative dosage rather than the daily dosage is the most important factor determining gonadotoxicity of cyclophosphamide, also used in the preparative regimens. There are insufficient follow-up data on the effect of irradiation, when given during childhood, on spermatogenesis later in life.

Among the 30 patients in our series, only one patient treated with TAI with testicular shielding/cyclophosphamide based conditioning regimen fathered a child. Testicular biopsy demonstrated that another patient treated with TAI with testicular shield showed well-developed seminiferous tubules with moderate number of spermatogonium. Moreover, the patient with CML receiving BMT after TBI with gonadal shielding, had normal testicular volume and function at latest evaluation. This finding might indicate that gonadal shielding to irradiation could protect germinal epithelium function. In view of the known vulnerability of germinal epithelium to irradiation and the elevated FSH levels, we expect most of the male patients to be infertile later in life.

In conclusion, normal pubertal development was attained in the childhood recipients of BMT, although subtle Leydig cell dysfunction as shown by mildly elevated LH levels and severely affected spermatogenesis was observed. Our data suggest that gonadal shielding to irradiation is effective to maintain testicular growth and function in the long-term surviving male recipients who have undergone BMT during childhood or adolescence. Careful follow-up studies are necessary to determine the natural history of testicular function and the reproductive potential of these subjects.

References

  1. 1

    Shalet SM . Endocrine consequences of treatment of malignant disease. Arch Dis Child 1989; 64: 1635–1641.

    CAS  Article  Google Scholar 

  2. 2

    Kolb HJ, Bender-Gotze C . Late complications after allogeneic bone marrow transplantation for leukaemia. Bone Marrow Transplant 1990; 6: 61–72.

    CAS  PubMed  Google Scholar 

  3. 3

    Duell T, van Lint MT, Ljungman P, Tichelli A, Socie G, Apperley JF et al. Health and functional status of long-term survivors of bone marrow transplantation. EBMT Working Party on Late Effects and EULEP Study Group on Late Effects. European Group for Blood and Marrow Transplantation. Ann Intern Med 1997; 126: 184–192.

    CAS  Article  Google Scholar 

  4. 4

    Brennan BM, Shalet SM . Endocrine late effects after bone marrow transplant. Br J Haematol 2002; 118: 58–66.

    Article  Google Scholar 

  5. 5

    Socié G, Salooja N, Cohen A, Rovelli A, Carreras E, Locasciulli A et al., Late Effect Working Party of the European Group for Blood and Marrow Transplantation. Nonmalignant late effects after allogeneic stem cell transplantation. Blood 2003; 101: 3373–3385.

    Article  Google Scholar 

  6. 6

    Howell SJ, Shalet SM . Testicular function following chemotherapy. Hum Reprod Update 2001; 7: 363–369.

    CAS  Article  Google Scholar 

  7. 7

    Bakker B, Massa GG, Oostdijk W, Van Weel-Sipman MH, Vossen JM, Wit JM . Pubertal development and growth after total-body irradiation and bone marrow transplantation for haematological malignancies. Eur J Pediatr 2000; 159: 31–37.

    CAS  Article  Google Scholar 

  8. 8

    Sanders JE, Hawley J, Levy W, Gooley T, Buckner CD, Deeg HJ et al. Pregnancies following high-dose cyclophosphamide with or without high-dose busulfan or total-body irradiation and bone marrow transplantation. Blood 1996; 87: 3045–3052.

    CAS  PubMed  Google Scholar 

  9. 9

    Jacob A, Goodman A, Holmes J . Fertility after bone marrow transplantation following conditioning with cyclophosphamide and total body irradiation. Bone Marrow Transplant 1995; 15: 483–484.

    CAS  PubMed  Google Scholar 

  10. 10

    Salooja N, Szydlo RM, Socie G, Rio B, Chatterjee R, Ljungman P et al., Late Effects Working Party of the European Group for Blood and Marrow Transplantation. Pregnancy outcomes after peripheral blood or bone marrow transplantation: a retrospective survey. Lancet 2001; 358: 271–276.

    CAS  Article  Google Scholar 

  11. 11

    Tanner JM, Whitehouse RH . Clinical longitudinal standards for height, weight, height velocity, weight velocity, and stages of puberty. Arch Dis Child 1976; 51: 170–179.

    CAS  Article  Google Scholar 

  12. 12

    Prader A . Testicular size: assessment and clinical importance. Triangle 1966; 7: 240–243.

    CAS  PubMed  Google Scholar 

  13. 13

    Fujieda K, Matsuura N . Growth and maturation in the male genitalia from birth to adolescence. I. Change of testicular volume. Acta Paediatr Jpn 1987; 29: 214–219.

    CAS  Article  Google Scholar 

  14. 14

    Chatterjee R, Kottaridis PD, McGarrigle HH, Eliahoo J, McKeag N, Mackinnon S et al. Patterns of Leydig cell insufficiency in adult males following bone marrow transplantation for haematological malignancies. Bone Marrow Transplant 2001; 28: 497–502.

    CAS  Article  Google Scholar 

  15. 15

    Tajima M . Testicular measurement by test size orchidometer. Acta Urol Jpn 1988; 34: 2013–2020.

    CAS  Google Scholar 

  16. 16

    Kubota C, Shinohara O, Hinohara T, Hattori K, Yabe H, Yabe M et al. Changes in hypothalamic-pituitary function following bone marrow transplantation in children. Acta Paediatr Jpn 1994; 36: 37–43.

    CAS  Article  Google Scholar 

  17. 17

    Shalet SM, Hann IM, Lendon M, Morris Jones PH, Beardwell CG . Testicular function after combination chemotherapy in childhood for acute lymphoblastic leukaemia. Arch Dis Child 1981; 56: 275–278.

    CAS  Article  Google Scholar 

  18. 18

    Ogilvy-Stuart AL, Clark DJ, Wallace WH, Gibson BE, Stevens RF, Shalet SM et al. Endocrine deficit after fractionated total body irradiation. Arch Dis Child 1992; 67: 1107–1110.

    CAS  Article  Google Scholar 

  19. 19

    Sarafoglou K, Boulad F, Gillio A, Sklar C . Gonadal function after bone marrow transplantation for acute leukemia during childhood. J Pediatr 1997; 130: 210–216.

    CAS  Article  Google Scholar 

  20. 20

    Shalet SM, Didi M, Ogilvy-Stuart AL, Schulga J, Donaldson MD . Growth and endocrine function after bone marrow transplantation. Clin Endocrinol (Oxf) 1995; 42: 333–339.

    CAS  Article  Google Scholar 

  21. 21

    Shalet SM . Effect of irradiation treatment on gonadal function in men treated for germ cell cancer. Eur Urol 1993; 23: 148–151.

    CAS  Article  Google Scholar 

  22. 22

    Sklar CA, Robison LL, Nesbit ME, Sather HN, Meadows AT, Ortega JA et al. Effects of radiation on testicular function in long-term survivors of childhood acute lymphoblastic leukemia: a report from the Children Cancer Study Group. J Clin Oncol 1990; 8: 1981–1987.

    CAS  Article  Google Scholar 

  23. 23

    Sanders JE . The impact of marrow transplant preparative regimens on subsequent growth and development. The Seattle Marrow Transplant Team. Semin Hematol 1991; 28: 244–249.

    CAS  PubMed  Google Scholar 

  24. 24

    Sklar CA, Kim TH, Ramsay NK . Testicular function following bone marrow transplantation performed during or after puberty. Cancer 1984; 53: 1498–1501.

    CAS  Article  Google Scholar 

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Acknowledgements

We thank all the medical staff in Tokai University Hospital for patient care. This study was partially supported by a Research Grant on Human Genome and Tissue Engineering from the Ministry of Health, Welfare and Labor.

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Ishiguro, H., Yasuda, Y., Tomita, Y. et al. Gonadal shielding to irradiation is effective in protecting testicular growth and function in long-term survivors of bone marrow transplantation during childhood or adolescence. Bone Marrow Transplant 39, 483–490 (2007). https://doi.org/10.1038/sj.bmt.1705612

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Keywords

  • irradiation
  • gonadal shield
  • testicular growth
  • testicular function

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