- Email: [email protected]
Pregnancy after intracytoplasmic injection of spermatid
There is really no evidence for a gap, widening or not. Illsley and LeGrand2 have already criticised the use of SMRs in this way but have received no public answer.
Gosfield Hall, Halstead, Essex C09 1SF, UK
2
Phillimore P, Beattie A, Townsend P. Widening inequality of health in northern England. BMJ 1994; 308: 1125-28. Illsley R, LeGrand J. Regional inequalities in mortality. J Epidemiol Commun Health 1993; 47: 443.
Notifying former patients of an infected health-care worker SiR-In January, 1995, a lookback exercise was undertaken after the death of a London dentist with AIDS. Letters were sent from the local health authorities to patients on whom the dentist had performed exposure-prone procedures’ between 1988 and 1993. This letter explained the small risk of infection and offered a confidential and anonymous free helpline telephone service staffed by experienced health advisers providing counselling and offering HIV antibody testing at one of four London centres. At one centre, the Chelsea and Westminster Hospital, we asked callers about their reactions to this approach. Although 74% of 130 callers stated that they were anxious on receipt of the letter, 89% felt that the wording was satisfactory. 96% thought that a letter was the best method of contact, and a similar proportion believed that they had a right to be informed about this matter. One caller would have preferred to be contacted by telephone and another stated a preference for notification by the general
practice. Three weeks after this survey, a High Court judgment to a 1991 lookback of patients of an infected healthcare worker in Manchester, ruled that the health authorities there should have realised that "the best method of informing a patient of the risk of HIV and AIDS, even though the risk is very remote, is face to face and by the patient’s GP or other experienced health worker; secondly that to do so by letter carries a foreseeable risk that some vulnerable individuals may suffer psychiatric injury going beyond the shock and distress which is natural and foreseeable in all or most individuals". This ruling is currently under appeal. There may be major implications depending on the outcome of this case for the method of notifying patients in future lookback exercises; and also for the wider area of notification of patients with sexually transmitted diseases conducted by health advisers in clinics nationally. Indeed, this ruling may call into question whether it is appropriate to notify patients by letter of any potentially adverse news relating to medical matters. The views expressed in our survey of callers challenge the premise that notification by letter is unreasonable in these circumstances.
relating
*James Monteith, Renee Aroney, Simon Barton, Bruce Armstrong, David Hawkins *Health Advisers’ Department, John Hunter Clinic, St Stephen’s Centre, Chelsea and Westminster Hospital, 369 Fulham Road, London SW10 9TH, UK; and St Mary’s Hospital, London W2
1
UK Health
Department. AIDS/HIV infected health care workers: the management of infected health care workers; recommendations of the Expert Advisory Group on AIDS. March, guidance
2
clinically successful report on intracytoplasmic sperm injection (ICSI),’ this technique has been used to achieve conception and delivery of healthy offspring in a variety of causes of severe male-factor infertility, including the use of testicular sperm.3 Recently Vanderzwalmen et a14 reported successful fertilisation and cleavage of an oocyte injected with a "late-stage" spermatid. We report a case in which spermatids were used to achieve conception and subsequent pregnancy. The male was 30 years of age and underwent surgical SiR-Since
R R Gordon
1
Pregnancy after intracytoplasmic injection of spermatid
on
1994. A & B and others
Authority. Jan 31,
Tameside and 1995.
vs
Glossop and Trafford Area Health
the
first
correction of undescended testes at 4 years of age, and a further operation for a bilateral hernia at 5. A testicular biopsy specimen and follicle-stimulating hormone were reported normal, although ejaculates ranged from cryptozoospermia to azoospermia. The female was 27 years of age with no remarkable history, and had undergone routine pituitary desensitisation and follicular stimulation. Initial attempts were made to recover sperm from the epididymes, from which only two grossly abnormal immotile sperm were obtained. We then proceeded to removal of seminiferous tubles which were placed in HEPES-buffered Earle’s medium in a Petri dish. The tissue was extensively dissected with 21 gauge needles. After approximately 2 hours we were able to recover nine "spermatozoa" that had withered and severely abnormally formed pinheads, and truncated and coiled tails. These were unsuitable for ICSI. Further examination of the tissue revealed a scant number of elongated spermatids (Sd 1), looking healthier than the few spermatozoa obtained. Despite intensive investigation of the seminiferous tubules, the outcome of the epididymal aspiration and testicular biopsy appeared incongruous with the previous report. This may be due to focal spermatogenic activity in the testicular material. All sperm were collected with a 14 11 microneedle and placed immediately into 10%
polyvinylpyrolidone. 14 follicles were observed on day 11 of stimulation (day of human chorionic gonadotropin [HCG]), with eight follicles in excess of 14 mm. After extensive discussion with the couple we proceeded to oocyte recovery. 13 oocyte-cumulus complexes were obtained resulting in 12 metaphase II oocytes after cumulus digestion with hyaluronidase 80 IU/mL. The technique for ICSI was identical to that previously published,’ except for the use of a slightly larger needle (>7-8 11) to incorporate the elongated spermatid in which, any event, had to undergo deformation of the large
cytoplasm during aspiration. Only ten spermatids were available for injection, and of the ten oocytes injected only one achieved fertilisation, assessed as the presence of two pronuclei and two polar bodies. 24 h later, this zygote had cleaved to the two-cell stage, scored as a grade 1 embryo with less than 10% cytoplasmic fragments in the perivitelline space. The embryo was transferred and the luteal phase was supported with progesterone, 50 mg intramuscularly. 21 days post-oocyte recovery, the (3-HCG was 823 IU/L; 3 weeks later, ultrasound revealed a singleton pregnancy with an active foetal heart. The pregnancy is continuing. Animal studies have shown the potential of round spermatids to achieve fertilisation and delivery of live, healthy, and fertile offspring. The current and previous reports indicate that the human spermatid is capable of activating and initiating fertilisation in the human oocyte, and providing the necessary male-derived components for cleavage events and implantation. Questions remain about the stages of spermatid development that can be used, and the efficiency, efficacy, and safety of this treatment approach. This gives rise to cautious optimism for the 1641
j 1
treatment
Sertoli-cell We thank assistance.
of
with testicular tubular atrophy, syndrome, and other similar conditions. men
putative
Prof G Potashnik, Dr K Dowell, and Dr G Ndukwe for
*S Fishel, S Green, M Bishop, S Thornton, A Hunter, S S AI-Hassan
Fleming,
*NURTURE, Department of Obstetrics and Gynaecology, University Hospital, Queen’s Medical Centre, Nottingham NG7 2UH, UK; and City Hospital, Nottingham 1
2
3
4
5
Devroey P, Van Steirteghem A. Pregnancies after intracytoplasmic injection of single spermatozoa into an oocyte. Lancet Palermo G, Joris H,
1992; 340: 17. Fishel S, Green S, Dowell K, et al. Fifty-nine cases of extreme malefactor infertility-immotile sperm, Kartagener syndrome, globozoospermia, spermatogenic arrest, testicular sperm-treated by SUZI/ICSI. Human Reprod 1994; 9: 5. Schoysman R, Vanderzwalmen P, Nijs M, et al. Pregnancy after fertilisation with human testicular spermatazoa. Lancet 1993; 342: 1237. Vanderzwalmen P, Lejeune B, Nijs M, Segal-Bertin G, Vandamme B, Schoysman R. Fertilisation of an oocyte micro-inseminated with a spermatid in an in-vitro fertilisation programme. Human Reprod 1995; 10: 502-03. Fishel S, Lisi F, Rinaldi L, et al. Systematic examination of immobilising spermatozoa before intracytoplasmic sperm injection in the human. Human Reprod 1995; 10: 497-500.
We conclude that if an initial examination of a semen sample appears "clear" of sperm the sample must be centrifuged. It should be noted, however, that a single centrifugation step might lead to an unacceptably large detritus deposit which is difficult to examine. Therefore, we suggest a two-stage process with a low (600 g) initial centrifugation for 15 min and examination of the pelleted material. If no sperm are found this should be followed by a second higher (>3000 g) speed centrifugation of the with examination of the deposit. Only when no supernatant are found after a sperm complete and systematic search of 10 multiple uL aliquants of the resuspended second should precipitate samples be classified as azoospermic. *K S Lindsay, I Floyd, R Swan Fertility Laboratory, Queen Charlotte’s and
1
2
3
4
Classification of
azoospermic samples
SiR-There have been reports of failure of vasectomy with comments about small numbers of sperm or apparently no sperm left in the ejaculate after the procedure. 1,2 We have reviewed 300 semen samples that seemed clear on preliminary examination and found that 9-8% had sperm present after centrifugation and examination of the resultant deposit. This demonstrates that a simple examination of a single drop of semen, as done by some laboratories, is flawed as a technique to define azoospermia. Problems in examining a single semen drop include, mixing of a non-homogeneous fluid, examining a small fraction of the total ejaculate (< 1 % if the WH03 recommended 10 ilL volume is used), and conversely, problems with microscope depth of field if a larger volume is selected. According to the WHO, when low numbers of sperm are present, sample examination should be preceded by centrifugation at 600 g for 15 min, removal of the bulk of the seminal plasma, and resuspension of the sample making appropriate volume adjustment. However, the relatively low centrifugal force of 600 g is derived from the possibility of stress damage to sperm at greater than 800 it and even centrifugation at 2500 g will not precipitate all sperm.5 We have subjected semen samples to an initial centrifugation and the resultant supernatants to successive incremental forces up to 3600 g. As can be seen from the table, small numbers of sperm were found in some samples even after centrifugation at over 3000 g. Extending the centrifugation time from 10 to 15 min at either 600g or 1200 g was also found to increase the number of sperm deposited in a standard 1 mL of semen in nine of ten
5
Chelsea Hospital, London W6 OXG, UK
Thompson JA, Lincoln PJ, Mortimer P. Paternity by a seemingly infertile vasectomised man. BMJ 1993; 307: 299-300. Smith JC, Cranston D, O’Brien T, Guillebaud J, Hindmarsh J, Turner AG. Fatherhood without apparent spermatozoa after vasectomy. Lancet 1994; 343: 30. World Health Organization. WHO laboratory manual for the examination of human semen and sperm mucus interaction. 3rd ed. Cambridge: Cambridge University Press, 1992. Fredricsson B, Kinnan S. Vitality and morphology of human spermatozoa studies on the resistance to storage and centrifugation and on the removal of dead spermatozoa. Andrologia 1979; 11: 135-41. Jeulin C, Serres C, Jouannet P. The effects of centrifugation, various synthetic media and temperature on the motility and viability of human spermatozoa. Reprod Nutr Dev 1982; 22: 81-91.
Sudden fall in breast England and Wales
cancer
death rates in
SiR-Breast cancer death rates in England and Wales stopped increasing in the late 1980s and have suddenly started to fall. The death rates have fallen by about 10% since 1985-89 (see figure), with 9517 deaths at ages 20-79 in 1993,’ compared with 10 538 in 1989. The decline has been greater in younger than in older women: since 1985-89 breast cancer mortality rates decreased by 14% in women aged 20-49; by 11% in women aged 50-69; and by 5% in women aged 70-79. Part of the fall between 1992 and 1993 is due to changes in the way of assigning cause of death2 but these changes mostly affected women aged over 80 and make little contribution to the overall downward trend seen in the figure.
samples.
*1 sample of mucoid consistency. Table: Number of samples (n=14) with sperm seen after sequential 10-min centrifugation of semen and resultant
Figure:
supernatants
1950-93
1642
Breast cancer
mortality in England and Wales,
Gosfield Hall, Halstead, Essex C09 1SF, UK
2
Phillimore P, Beattie A, Townsend P. Widening inequality of health in northern England. BMJ 1994; 308: 1125-28. Illsley R, LeGrand J. Regional inequalities in mortality. J Epidemiol Commun Health 1993; 47: 443.
Notifying former patients of an infected health-care worker SiR-In January, 1995, a lookback exercise was undertaken after the death of a London dentist with AIDS. Letters were sent from the local health authorities to patients on whom the dentist had performed exposure-prone procedures’ between 1988 and 1993. This letter explained the small risk of infection and offered a confidential and anonymous free helpline telephone service staffed by experienced health advisers providing counselling and offering HIV antibody testing at one of four London centres. At one centre, the Chelsea and Westminster Hospital, we asked callers about their reactions to this approach. Although 74% of 130 callers stated that they were anxious on receipt of the letter, 89% felt that the wording was satisfactory. 96% thought that a letter was the best method of contact, and a similar proportion believed that they had a right to be informed about this matter. One caller would have preferred to be contacted by telephone and another stated a preference for notification by the general
practice. Three weeks after this survey, a High Court judgment to a 1991 lookback of patients of an infected healthcare worker in Manchester, ruled that the health authorities there should have realised that "the best method of informing a patient of the risk of HIV and AIDS, even though the risk is very remote, is face to face and by the patient’s GP or other experienced health worker; secondly that to do so by letter carries a foreseeable risk that some vulnerable individuals may suffer psychiatric injury going beyond the shock and distress which is natural and foreseeable in all or most individuals". This ruling is currently under appeal. There may be major implications depending on the outcome of this case for the method of notifying patients in future lookback exercises; and also for the wider area of notification of patients with sexually transmitted diseases conducted by health advisers in clinics nationally. Indeed, this ruling may call into question whether it is appropriate to notify patients by letter of any potentially adverse news relating to medical matters. The views expressed in our survey of callers challenge the premise that notification by letter is unreasonable in these circumstances.
relating
*James Monteith, Renee Aroney, Simon Barton, Bruce Armstrong, David Hawkins *Health Advisers’ Department, John Hunter Clinic, St Stephen’s Centre, Chelsea and Westminster Hospital, 369 Fulham Road, London SW10 9TH, UK; and St Mary’s Hospital, London W2
1
UK Health
Department. AIDS/HIV infected health care workers: the management of infected health care workers; recommendations of the Expert Advisory Group on AIDS. March, guidance
2
clinically successful report on intracytoplasmic sperm injection (ICSI),’ this technique has been used to achieve conception and delivery of healthy offspring in a variety of causes of severe male-factor infertility, including the use of testicular sperm.3 Recently Vanderzwalmen et a14 reported successful fertilisation and cleavage of an oocyte injected with a "late-stage" spermatid. We report a case in which spermatids were used to achieve conception and subsequent pregnancy. The male was 30 years of age and underwent surgical SiR-Since
R R Gordon
1
Pregnancy after intracytoplasmic injection of spermatid
on
1994. A & B and others
Authority. Jan 31,
Tameside and 1995.
vs
Glossop and Trafford Area Health
the
first
correction of undescended testes at 4 years of age, and a further operation for a bilateral hernia at 5. A testicular biopsy specimen and follicle-stimulating hormone were reported normal, although ejaculates ranged from cryptozoospermia to azoospermia. The female was 27 years of age with no remarkable history, and had undergone routine pituitary desensitisation and follicular stimulation. Initial attempts were made to recover sperm from the epididymes, from which only two grossly abnormal immotile sperm were obtained. We then proceeded to removal of seminiferous tubles which were placed in HEPES-buffered Earle’s medium in a Petri dish. The tissue was extensively dissected with 21 gauge needles. After approximately 2 hours we were able to recover nine "spermatozoa" that had withered and severely abnormally formed pinheads, and truncated and coiled tails. These were unsuitable for ICSI. Further examination of the tissue revealed a scant number of elongated spermatids (Sd 1), looking healthier than the few spermatozoa obtained. Despite intensive investigation of the seminiferous tubules, the outcome of the epididymal aspiration and testicular biopsy appeared incongruous with the previous report. This may be due to focal spermatogenic activity in the testicular material. All sperm were collected with a 14 11 microneedle and placed immediately into 10%
polyvinylpyrolidone. 14 follicles were observed on day 11 of stimulation (day of human chorionic gonadotropin [HCG]), with eight follicles in excess of 14 mm. After extensive discussion with the couple we proceeded to oocyte recovery. 13 oocyte-cumulus complexes were obtained resulting in 12 metaphase II oocytes after cumulus digestion with hyaluronidase 80 IU/mL. The technique for ICSI was identical to that previously published,’ except for the use of a slightly larger needle (>7-8 11) to incorporate the elongated spermatid in which, any event, had to undergo deformation of the large
cytoplasm during aspiration. Only ten spermatids were available for injection, and of the ten oocytes injected only one achieved fertilisation, assessed as the presence of two pronuclei and two polar bodies. 24 h later, this zygote had cleaved to the two-cell stage, scored as a grade 1 embryo with less than 10% cytoplasmic fragments in the perivitelline space. The embryo was transferred and the luteal phase was supported with progesterone, 50 mg intramuscularly. 21 days post-oocyte recovery, the (3-HCG was 823 IU/L; 3 weeks later, ultrasound revealed a singleton pregnancy with an active foetal heart. The pregnancy is continuing. Animal studies have shown the potential of round spermatids to achieve fertilisation and delivery of live, healthy, and fertile offspring. The current and previous reports indicate that the human spermatid is capable of activating and initiating fertilisation in the human oocyte, and providing the necessary male-derived components for cleavage events and implantation. Questions remain about the stages of spermatid development that can be used, and the efficiency, efficacy, and safety of this treatment approach. This gives rise to cautious optimism for the 1641
j 1
treatment
Sertoli-cell We thank assistance.
of
with testicular tubular atrophy, syndrome, and other similar conditions. men
putative
Prof G Potashnik, Dr K Dowell, and Dr G Ndukwe for
*S Fishel, S Green, M Bishop, S Thornton, A Hunter, S S AI-Hassan
Fleming,
*NURTURE, Department of Obstetrics and Gynaecology, University Hospital, Queen’s Medical Centre, Nottingham NG7 2UH, UK; and City Hospital, Nottingham 1
2
3
4
5
Devroey P, Van Steirteghem A. Pregnancies after intracytoplasmic injection of single spermatozoa into an oocyte. Lancet Palermo G, Joris H,
1992; 340: 17. Fishel S, Green S, Dowell K, et al. Fifty-nine cases of extreme malefactor infertility-immotile sperm, Kartagener syndrome, globozoospermia, spermatogenic arrest, testicular sperm-treated by SUZI/ICSI. Human Reprod 1994; 9: 5. Schoysman R, Vanderzwalmen P, Nijs M, et al. Pregnancy after fertilisation with human testicular spermatazoa. Lancet 1993; 342: 1237. Vanderzwalmen P, Lejeune B, Nijs M, Segal-Bertin G, Vandamme B, Schoysman R. Fertilisation of an oocyte micro-inseminated with a spermatid in an in-vitro fertilisation programme. Human Reprod 1995; 10: 502-03. Fishel S, Lisi F, Rinaldi L, et al. Systematic examination of immobilising spermatozoa before intracytoplasmic sperm injection in the human. Human Reprod 1995; 10: 497-500.
We conclude that if an initial examination of a semen sample appears "clear" of sperm the sample must be centrifuged. It should be noted, however, that a single centrifugation step might lead to an unacceptably large detritus deposit which is difficult to examine. Therefore, we suggest a two-stage process with a low (600 g) initial centrifugation for 15 min and examination of the pelleted material. If no sperm are found this should be followed by a second higher (>3000 g) speed centrifugation of the with examination of the deposit. Only when no supernatant are found after a sperm complete and systematic search of 10 multiple uL aliquants of the resuspended second should precipitate samples be classified as azoospermic. *K S Lindsay, I Floyd, R Swan Fertility Laboratory, Queen Charlotte’s and
1
2
3
4
Classification of
azoospermic samples
SiR-There have been reports of failure of vasectomy with comments about small numbers of sperm or apparently no sperm left in the ejaculate after the procedure. 1,2 We have reviewed 300 semen samples that seemed clear on preliminary examination and found that 9-8% had sperm present after centrifugation and examination of the resultant deposit. This demonstrates that a simple examination of a single drop of semen, as done by some laboratories, is flawed as a technique to define azoospermia. Problems in examining a single semen drop include, mixing of a non-homogeneous fluid, examining a small fraction of the total ejaculate (< 1 % if the WH03 recommended 10 ilL volume is used), and conversely, problems with microscope depth of field if a larger volume is selected. According to the WHO, when low numbers of sperm are present, sample examination should be preceded by centrifugation at 600 g for 15 min, removal of the bulk of the seminal plasma, and resuspension of the sample making appropriate volume adjustment. However, the relatively low centrifugal force of 600 g is derived from the possibility of stress damage to sperm at greater than 800 it and even centrifugation at 2500 g will not precipitate all sperm.5 We have subjected semen samples to an initial centrifugation and the resultant supernatants to successive incremental forces up to 3600 g. As can be seen from the table, small numbers of sperm were found in some samples even after centrifugation at over 3000 g. Extending the centrifugation time from 10 to 15 min at either 600g or 1200 g was also found to increase the number of sperm deposited in a standard 1 mL of semen in nine of ten
5
Chelsea Hospital, London W6 OXG, UK
Thompson JA, Lincoln PJ, Mortimer P. Paternity by a seemingly infertile vasectomised man. BMJ 1993; 307: 299-300. Smith JC, Cranston D, O’Brien T, Guillebaud J, Hindmarsh J, Turner AG. Fatherhood without apparent spermatozoa after vasectomy. Lancet 1994; 343: 30. World Health Organization. WHO laboratory manual for the examination of human semen and sperm mucus interaction. 3rd ed. Cambridge: Cambridge University Press, 1992. Fredricsson B, Kinnan S. Vitality and morphology of human spermatozoa studies on the resistance to storage and centrifugation and on the removal of dead spermatozoa. Andrologia 1979; 11: 135-41. Jeulin C, Serres C, Jouannet P. The effects of centrifugation, various synthetic media and temperature on the motility and viability of human spermatozoa. Reprod Nutr Dev 1982; 22: 81-91.
Sudden fall in breast England and Wales
cancer
death rates in
SiR-Breast cancer death rates in England and Wales stopped increasing in the late 1980s and have suddenly started to fall. The death rates have fallen by about 10% since 1985-89 (see figure), with 9517 deaths at ages 20-79 in 1993,’ compared with 10 538 in 1989. The decline has been greater in younger than in older women: since 1985-89 breast cancer mortality rates decreased by 14% in women aged 20-49; by 11% in women aged 50-69; and by 5% in women aged 70-79. Part of the fall between 1992 and 1993 is due to changes in the way of assigning cause of death2 but these changes mostly affected women aged over 80 and make little contribution to the overall downward trend seen in the figure.
samples.
*1 sample of mucoid consistency. Table: Number of samples (n=14) with sperm seen after sequential 10-min centrifugation of semen and resultant
Figure:
supernatants
1950-93
1642
Breast cancer
mortality in England and Wales,