1 Department of Obstetrics and Gynaecology, Royal Infirmary of Edinburgh, 1 Lauriston Place, Edinburgh EH3 9EF and 2 MRC Reproductive Biology Unit, Centre for Reproductive Biology, Edinburgh, UK
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Abstract |
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Key words: dynein arm deficiency/immotile cilia syndrome/immotile spermatozoa/Kartagener's syndrome
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Introduction |
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Until recently there was no treatment available for male infertility due to ICS. The development of assisted reproductive techniques has allowed rational treatment for these patients and to date there have been reported pregnancies using subzonal insemination (Bongso et al., 1989; Wolf et al., 1993
; Nijs et al., 1996
) and intracytoplasmic sperm injection (ICSI) (Brugo et al., 1997
; Chemes et al., 1998
). We report a successful pregnancy achieved by in-vitro fertilization (IVF) using spermatozoa from a patient with Kartagener's syndrome.
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Case report |
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He and his partner presented with a 5 year history of primary infertility to the Infertility Clinic, Royal of Infirmary Edinburgh. His spouse was shown to have hyperprolactinaemia due to a micro-adenoma of the pituitary gland. Following treatment with bromocriptine she had ovulatory levels of progesterone on day 21 of her menstrual cycle. Otherwise she had an uneventful history. Her investigations, including pelvic ultrasound, diagnostic laparoscopy and hydrotubation, were normal. His history provided no further evidence of medical conditions which might affect fertility. On examination he had normal secondary sex characteristics and both testicular volumes were normal. He was noted to have a large right-sided varicocele, which was treated by percutaneous trans-catheter embolization. His serum prolactin, follicle-stimulating hormone (FSH) and testosterone were all within normal limits.
Semen analysis
Viscosity, liquefaction and pH were normal. Semen samples were assessed for ejaculate volume, morphology, concentration and percentage motile spermatozoa using the protocols described by the World Health Organization (WHO, 1992). The results of eight semen analyses assessed over a 4 year period were normal ejaculate volume and sperm concentration but mean total motility was only 25% and mean progressive motility was only 2.5% (Table I). Semen was negative on bacteriological culture and immunological screening.
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On ultrastructural examination sperm heads had intact nuclei and acrosomal regions. However, all flagella examined were grossly abnormal with axonemal disruption surrounded by a thick uncondensed fibrous sheath. Figures 1 and 2 are transverse sections of sperm flagella. In Figure 1
, microtubular pairs can be identified with one set translocated outside the fibrous sheath. Outer dynein arms are mostly absent, though the number of inner dynein arms is reduced. In Figure 2
there is complete disorganisation of the microtubules with no recognizable pattern. Transmission electron microscopy was repeated on two separate occasions and on both occasions all spermatozoa analysed had defects in the dynein arms.
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Sperm preparation was carried out using 50100% discontinuous Percoll gradient (1 ml/1 ml, centrifuged for 17 min at 500 g) followed by two washes (5 min at 230 g). A swim-up from the final pellet was then performed using Earle's medium (Sigma, Bornem, Belgium) supplemented with 0.5% human serum albumin (Irvine Scientific, Zellick, Belgium) in an atmosphere of 5% CO2 for 50 min. After preparation 1.6x106 spermatozoa were present in 0.3 ml culture medium. A total of 88% were motile and 56% had abnormal morphology. No hyperactivated spermatozoa were seen. Spermatozoa were diluted so that ~ 100x103 in 1 ml Ham's F10 medium were added to the oocyte and incubated for 22 h. At this stage two pronuclei were present.
The embryo was transferred without difficulty 49 h following oocyte recovery. Luteal support was carried out with progesterone suppositories (Cyclogest®; Hoechst, Brussels, Belgium) 400 mg for 10 days. Blood was taken for HCG on 14 days post-oocyte recovery and was 147 IU. A pelvic scan was performed 5 weeks following embryo transfer and a single intra-uterine pregnancy was identified. A male infant was delivered at a gestation of 40 weeks weighing 3.9 kg. On examination of the infant there was no evidence of situs invertus and he has remained well to date and is currently over 18 months old.
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Discussion |
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Evidence to date suggests that spermatozoa from men with ICS have a normal ability to capacitate and acrosome react in vitro and can fuse with the vitelline membrane if micromanipulated to lie adjacent to the oocyte (Aitken et al., 1983). In view of the poor sperm motility and in particular the absence of hyperactivation, spermatozoa from men with ICS are unlikely to achieve fertilization in vivo.
The impaired sperm motility in ICS is due to axonemal disruption. The sperm tail has dynein arms, which link microtubular doublets in the axonemal complex (Eliasson et al., 1977). These dynein arms are the site of ATPase activity and mediate the sliding between doublet microtubules, essential for normal ciliary motion (Afzelius, 1986
). When dynein arms are lacking, a variety of ineffective ciliary motion patterns can occur, resulting in poor or no sperm motility. Several other ultrastructural defects have been noted in this condition, including lack of radial spokes (Sturges et al., 1979
), absent central pair of microtubules (Bacetti et al., 1979
), transposition of microtubules (Sturges et al., 1980
) and dysplasia of the fibrous sheath (Chemes et al., 1987
). In this reported case many of these abnormalities could be identified, varying from minor defects with transposition of one doublet of microtubules outside the fibrous sheath and lack of dynein arms to severe defect with no recognizable pattern of microtubules.
Assessment of sperm physiology from men with ICS is necessary in order to devise a rational approach to treatment of this disorder. ICSI would appear to be an appropriate treatment as it avoids the need for spermatozoa to attach and penetrate the zona pellucida. It has been shown that sperm motility does not affect success rates with ICSI (Mansour et al., 1995). Pregnancies have been reported following ICSI in men with totally immotile spermatozoa (Terriou et al., 1993
; Nijs et al., 1996
) and there have been reported pregnancies resulting from ICSI using immature sperm precursor cells (Tesarik et al., 1995
; Barak et al., 1998
). In men with ICS, pregnancy has been reported following subzonal insemination (Wolf et al., 1993
; Nijs et al., 1996
) and there have been live births following ICSI (Brugo et al., 1997
). In a recent publication (Chemes et al., 1998
), a mosaic variant of ICS with dysplasia of the fibrous sheath was treated with IVF and ICSI. There were no pregnancies with IVF, however there was successful fertilization following ICSI in six patients with two resultant pregnancies. However, both subzonal insemination and ICSI require expertise not available in all units and there are concerns over costs and outcomes.
IVF has been previously tried as a treatment for ICS with no success (Chemes, 1993). Our case is the first documented successful IVF treatment of ICS. Unfortunately, only one oocyte was obtained and therefore we are unable to comment on the fertilization rate using this patient's spermatozoa.
It is difficult to understand how spermatozoa with such severe axonemal disruption and impaired motility can penetrate the zona pellucida. Paternity was not proven directly in this case, however routine IVF safety procedures were performed by the embryologist to ensure correct paternity. It could be argued that not all spermatozoa were affected by axonemal disruption in this case. However, in all spermatozoa assessed at repeated electron microscopy there were axonemal defects. When assessed manually and with computer-assisted motility analysis, there were greatly reduced mean motility parameters (total motility, progressive motility, mean path velocity, tract speed, lateral amplitude and beat frequency). There was overlap between the sample population and the normal range, and it is therefore possible that a few spermatozoa had motility parameters within the normal range. However, no spermatozoa displayed hyperactivation. It is possible that despite no hyperactivated motility spermatozoa can penetrate the zona pellucida. Lastly in view of the heterogeneous nature of Kartagener's syndrome, it is difficult to be certain that this case truly has Kartagener's syndrome rather than severe asthenozoospermia with other abnormalities being coincidental. However, this seems unlikely as all the clinical features and most of the laboratory features of Kartagener's syndrome are present in this patient.
There are many incongruities concerning male infertility in ICS. Spontaneous pregnancies have been reported in 12 men affected by Kartagener's syndrome, though paternity was not proven (Rott et al., 1979). Kartagener's syndrome has been diagnosed in men with normal ciliary and flagellar structure (Greenstone et al., 1985; Escudier et al., 1987
), in cilia without dynein arms and normal flagella (Jonsson et al., 1982
) and in flagella without dynein arms and normal cilia (Wilton et al., 1986
). A variant of ICS has been identified (Chemes et al., 1998
), in which there is dysplasia of the fibrous sheath associated with lack of dynein arms in respiratory and sperm flagella. Two subgroups were recognized: a complete form in which all spermatozoa were affected and an incomplete form with alterations in 7080% of spermatozoa. It is clear that ICS is a heterogeneous group of disorders with similar clinical presentations but varied structural defects. This may be due to ciliary and flagellar dyneinmicrotubular binding factors being under separate genetic control or perhaps due to chimeras with mutations occurring during embryogenesis with only certain cells being affected. Lastly, it is possible that in certain cases situs invertus has occurred as an isolated defect and the infertility and chronic respiratory symptoms developed coincidentally due to an unrelated cause.
The heterogeneous nature of ICS makes the planning of treatment difficult. It has been shown that there is a correlation between structural and functional ciliary abnormalities in sperm samples from men with ICS (Afzelius, 1986). Until more is known with regard to the genetic control of ICS, we would suggest that treatment should be individualized depending on sperm motility. In cases where there is no sperm motility, ICSI may be the most appropriate treatment. However, if sperm motility is present a trial of IVF should be considered.
One concern regarding the fertility treatment of men with ICS is the possibility that the resultant child has the risk of being affected by the same condition. Although the exact genetic control of ICS is not understood, it is thought to be an autosomal recessive condition (Afzelius, 1986). Until the genetic basis for this condition is known, it is necessary to counsel couples regarding the possibility of genetic risks and to follow-up children fathered by men affected by ICS.
This case report demonstrates that spermatozoa from a man with ICS can penetrate the zona pellucida and fertilize oocytes in vitro despite impaired motility due to severe axonemal defects. IVF should be considered as a treatment option for infertile men with ICS when sperm motility is present. This case has important implications both for the treatment of infertile men with ICS and to further our understanding of the role of sperm motility in the fertilization process.
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Acknowledgments |
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Notes |
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References |
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Submitted on December 19, 1998; accepted on October 5, 1999.