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Birth of healthy children after intracytoplasmic sperm injection in two couples with male Kartagener’s syndrome
FERTILITY AND STERILITYt VOL. 70, NO. 4, OCTOBER 1998 Copyright ©1998 American Society for Reproductive Medicine Published by Elsevier Science Inc. Printed on acid-free paper in U.S.A.
Birth of healthy children after intracytoplasmic sperm injection in two couples with male Kartagener’s syndrome Annette von Zumbusch, M.D.,* Klaus Fiedler, M.D., Artur Mayerhofer, M.D.,† Berthold Jeßberger, M.D.,* Johannes Ring, M.D.,* and Hermann-Josef Vogt, M.D.* Technische Universita¨t Mu¨nchen; Study Group Noxenkatalog-Databank (Hazars in Occupation and Environment); Department of Reproductive Medicine, Frauenklinik Dr. W. Kru¨smann, Munich, Germany
Objective: To describe two cases in which intracytoplasmic sperm injection (ICSI) was successful for patients with infertility due to Kartagener’s syndrome. Design: Case report. Setting: Private hospital for gynecology department of reproductive medicine, and university hospital center for andrology. Patient(s): Two couples with primary infertility due to Kartagener’s syndrome in the male. Intervention(s): ICSI. Main Outcome Measure(s): Pregnancy and birth after ICSI. Result(s): In both couples, ICSI was successful in the first cycle. The uncomplicated pregnancies resulted in the birth of three healthy children. One female and male/female twins.
Received November 10, 1997; revised and accepted June 11, 1998. Presented in part at the “Gemeinsame Tagung der o¨sterreichischen Gesellschaft fu¨r Gyna¨kologie und Geburtshilfe und der Bayerischen Gesellschaft fu¨r Geburtshilfe und Frauenheilkunde,” Graz, Austria, May 29 –31, 1997. Reprint requests: Annette von Zumbusch, M.D., Klinik fu¨r Dermatologie und Allergologie, Biederstein, Technische Universita¨t Mu¨nchen, Biedersteinerstrasse 29, D-80802 Mu¨nchen, Germany (FAX:0049-89334983; e-mail: ANNETTE .VONZUMBUSCH@LRZ .TU-MUENCHEN.DE). * Department of Dermatology and Allergology, Technische Universita¨t Mu¨nchen. † Anatomical Institute, Technische Universita¨t Mu¨nchen. 0015-0282/98/$19.00 PII S0015-0282(98)00246-5
Conclusion(s): In couples with infertility due to Kartagener’s syndrome in the male, ICSI has proved to be a successful therapy resulting in clinically healthy offspring. This knowledge may improve our understanding of the involvement of paternally inherited centrosomes, which nucleate microtubules, in human reproduction. (Fertil Sterilt 1998;70:643– 6. ©1998 by American Society for Reproductive Medicine.) Key Words: Kartagener’s syndrome, immotile cilia syndrome, ICSI (intracytoplasmic sperm injection), centrosome, centriole, microtubule
The terms “immotile cilia syndrome” and “primary ciliary dyskinesia” are used to describe a group of inherited diseases that generally follow an autosomal recessive trait. They are characterized by defective ciliary ultrastructure (axoneme defects, mainly lack of dynein arms and spoke structures) (1) associated with reduced or completely absent ciliary motility. The heterogeneity of the symptoms seen in immotile cilia syndrome may be due to multiple genetic alterations. The molecular nature of such defects has not yet been identified. Kartagener’s syndrome is one of the disorders classified as primary ciliary dyskinesia. In addition to the cilial defects found in this group of diseases, Kartagener’s syndrome is clinically distinguishable by the presence of situs inversus (1, 2). Immotile cilia syndrome results in characteristic clinical symptoms, including chronic pansinusitis, recurrent infections of the respiratory tract, bronchiectasis, and infertility,
particularly in the male. The similarities in ultrastructure between the sperm flagella and cilia of the respiratory epithelial cells may account for the respiratory problems and the impairment or absence of sperm motility in these patients. Female patients, in contrast, are usually fertile, which may be due to muscular motility of the fallopian tubes (2). Intracytoplasmic sperm injection (ICSI) is used in cases in which the male partner has immotile spermatozoa. Stalf et al. (3) reported pregnancy and birth after ICSI with spermatozoa obtained from a patient with tail-stump syndrome. However, in a recent report by Papadimas et al. (4), pregnancy was not achieved when ICSI was performed with spermatozoa from a patient with immotile cilia syndrome. The reasons for this failure are not clear. To our knowledge, we describe the first cases in which ICSI was used successfully to achieve pregnancies in couples who were pre643
viously infertile because of male Kartagener’s syndrome. Two couples were treated with ICSI, which resulted in the birth of three healthy children.
CASE REPORTS
FIGURE 1 Electron micrograph of a sperm flagellum from patient 1. Cross-section of the midpiece of the flagellum shows a 912 arrangement and lack of both the inner and outer dynein arms (bar 5 0.5 mm).
Case 1 A 34-year-old man and his 32-year-old wife were referred to our andrology outpatient department because of a 6-year history of infertility. Gynecologic examination of the wife did not reveal any abnormalities. A semen analysis that was performed in a private practice had revealed that the man had a normal number of immotile spermatozoa. The patient reported no medical problems, and the clinical examination showed no pathologic findings. The patient had three brothers, one of whom had chronic bronchitis and a history of infertility. Radiographic examination of the brother showed situs inversus viscerum. The other brothers had children. A roentgenogram of the patient’s chest showed situs inversus viscerum. Computed tomography of the skull revealed polyposis nasi and pansinusitis. Consanguinity of the male patient’s parents was not known. Semen Analysis Semen was collected by masturbation into a sterile plastic cup. Twenty minutes was allowed for liquefaction, after which a standard semen analysis was performed. The sperm concentration was 75 3 106/mL, and the semen volume was 9.5 mL. The most remarkable feature was the complete absence of motile forms. Staining with aqueous eosin solution showed 90% viable spermatozoa with unremarkable morphology. Transmission Electron Microscopy (TEM) A sperm aliquot was fixed in a solution of 2.5% glutaraldehyde and 0.2 M cacodylate buffer. After centrifugation, the sample was resuspended in 0.2 M sodium cacodylate buffer. The specimen was postfixed in osmium tetroxide, dehydrated through a series of ascending concentrations of ethanol, and then embedded in Araldite (Araldite Kit 2000; Bio-Rad Laboratories, Munich, Germany). Ultrathin sections (50 –70 nm) were cut (LKB III ultramicrotome), mounted on copper grids, and contrasted with uranyl acetate followed by incubation with 2% lead citrate. The sections were examined with a Zeiss transmission electron microscope (Zeiss, Oberkochen, Germany) and then photographed. A total of 150 sperm were evaluated using TEM. Both 912 and 910 configurations were observed. In 90% of the spermatozoa, both inner and outer dynein arms were missing (Fig. 1). The inner dynein arms were present in 10%. In 80% of the sperm, central fibrils were missing or incomplete. Radial spokes were missing in 70% and appeared to be 644
von Zumbusch et al.
ICSI for Kartagener’s syndrome
defective in 10%. Examination of sperm heads revealed normal morphology.
Case 2 A 36-year-old man was referred to our clinic because of infertility. Several members of his large family were known to have immotile cilia syndrome, some of them without situs inversus. This family is being monitored by the Institute for Human Genetics, University of Erlangen, Germany. The patient did not have a history of medical problems. On the basis of situs inversus viscerum and immotile sperm tails, his condition was diagnosed as Kartagener’s syndrome. His wife’s gynecologic examination revealed no abnormalities. Semen analysis previously had demonstrated oligoasthenozoospermia. Semen was collected and prepared as described in case 1. The sperm concentration was 210 3 106/mL and the semen volume was 1 mL. The most noteworthy feature was the complete absence of motile forms. Ninety percent of the sperm were viable, as evaluated with eosin staining, and the morphology was normal. TEM was not performed in this patient because Kartagener’s syndrome had been clinically diagnosed. Vol. 70, No. 4, October 1998
Fertilization Procedure in Both Cases The procedure was approved by the Institutional Review Board at the Frauenklinik Dr. W. Kru¨smann. After down-regulation with a GnRH analogue (triptorelin 0.1 mg, Decapeptyl; Ferring, Kiel, Germany) starting on day 24 of the previous cycle, 150 IU of FSH (Fertinorm HP; Serono, Munich, Germany) was administered daily from the third day of the treatment cycle until the leading follicle reached a mean diameter of 18 mm. Thirty-six hours before oocyte retrieval, 5,000 IU of hCG (Pregnesin 5.000; Serono) was injected. Six metaphase II oocytes were retrieved from both patients by transvaginal ultrasound-guided follicle puncture. Spermatozoa were washed twice by suspension in culture medium (Hams F-10 medium) and were resuspended after centrifugation. The final dilution was placed under a layer of paraffin oil. ICSI was performed in both cases according to the method of Van Steirteghem et al. (5). In patient no. 1, four of six injected oocytes were fertilized. Two days after follicle puncture, two preimplantation embryos at the 4-cell stage were transferred. This resulted in a routine pregnancy and birth of a clinically healthy set of twins (female and male). In patient no. 2, three of six injected oocytes were fertilized and were transferred at the 4- and 6-cell stages 2 days after oocyte retrieval. A normal pregnancy and birth of a healthy girl followed treatment.
DISCUSSION Although the diseases of the immotile cilia syndrome are heterogeneous, they are believed to be due to a microtubuleassociated defect. This defect has not yet been fully characterized (1, 2, 6, 7). Microtubules are cytoskeletal elements essential for many crucial processes in all cells. With respect to germ cells, they are responsible for separation of the chromosomes during mitosis and meiosis, as well as migration of the sperm and egg nuclei during fertilization. The microtubules also form the basis for ciliary processes found in spermatozoa and in respiratory epithelial cells. It is hypothesized that one or more microtubule-associated defects may be detrimental to ciliar motility and/or mitotic or meiotic divisions, therefore hindering fertilization. Thus, a microtubule defect in Kartagener’s syndrome may be responsible for more than ciliary dyskinesia leading to immotile sperm. It is possible that previous attempts at ICSI in cases of Kartagener’s syndrome were unsuccessful because of this defect (4, 8). In this context, it is important to recall that in humans, in contrast to mice (9) or hamsters (10), the spermatozoon introduces the centrosome during insemination into the oocyte. Subsequently, the centrosome nucleates the new microtubule assembly to form the sperm aster (9, 11). The FERTILITY & STERILITYt
resulting microtubule orientation and polarity are determined by the centrosome. Because of the cited species differences, the hamster oocyte penetration test may not be valid for demonstrating reliable fertilizing ability of spermatozoa from patients with immotile cilia syndrome (10). Centrosomes and centrioles are poorly understood cellular structures. They are both self-reproducing organelles, and centrioles merely advertise the presence of centrosomes. Human spermatozoa also have centrioles. A well-defined proximal centriole is present next to the basal plate of the sperm head, whereas the distal centriole (which is a remnant) gives rise to the sperm tail axoneme during spermiogenesis (12). Centrosomes are reduced during gametogenesis. It is assumed that successful fertilization requires centrosome restoration (in which the sperm plays the central role) and microtubule-mediated motility of parental chromosomes at first mitosis. Investigations have shown that defects in microtubule organization result in failure to fertilize. Microtubule inhibitors prevent pronuclear fusion, which underscores the essential role of the sperm astral microtubules for organizing under a paternal direction (12, 13). It is possible that these processes important for fertilization could be defective in some patients with Kartagener’s syndrome. To our knowledge, this is the first report to document the successful use of ICSI in two cases of male Kartagener’s syndrome that resulted in the birth of healthy children of both sexes. Papadimas et al. (4) had limited success treating a male patient with immotile cilia syndrome. After ICSI was performed, 33% of the oocytes were fertilized and subsequently divided but did not implant. Nijs et al. (18) described ICSI in a patient with Kartagener’s syndrome. None of the oocytes fertilized after ICSI, but 3 of 12 oocytes fertilized after subzonal insemination, which suggests normal microtubule function in fertilization. The spermatozoa of the two men in our cases were completely immotile. As noted in case 1, marked structural axoneme defects were present, including the loss of all outer and most of the inner dynein arms. ICSI in both cases was successful in the first cycle. This implies that from the phenotype alone (immotile sperm syndrome and structural defect), no clear prediction of the success of fertilization can be made. It is likely that the active reproducing centrosome is a blend of both maternal proteins and the spermatozoon’s centrosome (12). Whether the maternal or paternal contribution at this point is variable, or whether alternative pathways exist, is a subject worthy of further investigation. The use of ICSI in the two cases of male Kartagener’s syndrome described herein resulted in the birth of three clinically healthy children without situs inversus. These results seem to ethically justify the use of assisted reproductive technology in similar cases. As with all hereditary diseases in which the nature of the defect is unknown and the symp645
toms are heterogeneous, the result of reproductive intervention is ultimately uncertain. In patients with immotile cilia syndrome, ICSI and other procedures leading to conception may result in the birth of seemingly healthy children, but there is a possibility that fertility problems and other medical problems will be diagnosed later. All patients must be informed about these risks and receive appropriate genetic analyses and counseling before the use of ICSI is considered.
4.
5.
6. 7. 8. 9. 10.
Acknowledgment: The authors thank Mrs. Else Marais for editorial assistance and critical review of the manuscript. 11.
References
12.
1. Afzelius BA. The immotile-cilia syndrome: a microtubule-associated defect. CRC 1985;19:63– 87. 2. Rott H-D. Kartagener’s syndrome and the syndrome of immotile cilia. Hum Genet 1979;46:249 – 61. 3. Stalf T, Sanchez R, Kuhn F-M, Schalles U, Kleinstein J, Hinz V, et al. Pregnancy and birth after intracytoplasmic sperm injection with sper-
646
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ICSI for Kartagener’s syndrome
13.
matozoa from a patient with tail stump syndrome. Hum Reprod 1995; 10:2112– 4. Papadimas J, Tarlatzis BC, Bili H, Sotiriadis T, Koliakou K, Bontis J, et al. Therapeutic approach of immotile cilia syndrome by intracytoplasmic sperm injection: a case report. Fertil Steril 1997;67: 562–5. Van Steirteghem AC, Liu J, Joris H, Nagy Z, Janssenswillen C, Tournaye H, et al. Higher success rate by intracytoplasmic sperm injection than by subzonal insemination. Report of a second series of 300 consecutive treatment cycles. Hum Reprod 1993;8:1055– 60. Aitken RJ, Ross A, Lees MM. Analysis of sperm function in Kartagener’s syndrome. Fertil Steril 1983;40:696 – 8. Aitken J. A clue to Kartagener’s. Nature 1991;353:306. Nijs M, Vanderzwalmen P, Vandamme B, Segal-Bertin G, Lejeune B, Segal L, et al. Fertilizing ability of immotile spermatozoa after intracytoplasmic sperm injection. Hum Reprod 1996;11:2180 –5. Schatten H, Schatten G, Mazia D, Balczon R, Simerly C. Behavior of centrosomes during fertilization and cell division in mouse oocytes and in sea urchin eggs. Proc Natl Acad Sci USA 1986;83:105–9. Hewitson L, Haavisto A, Simerly C, Jones J, Schatten G. Microtubule organization and chromatin configurations in hamster oocytes during fertilization and parthenogenetic activation, and after insemination with human sperm. Biol Reprod 1997;57:967–75. Palermo G, Munne S, Cohen J. The human zygote inherits its mitotic potential from the male gamete. Hum Reprod 1994;9:1220 –5. Sathananthan AH, Kola I, Osborne J, Trounson A, Ng SC, Bongso A, et al. Centrioles in the beginning of human development. Proc Natl Acad Sci USA 1991;88:4806 –10. Simerly C, Wu G-J, Zoran S, Ord T, Rawlins R, Jones J, et al. The paternal inheritance of the centrosome, the cell’s microtubule-organizing center, in humans, and the implications for infertility. Nature Med 1995;1:47–52.
Vol. 70, No. 4, October 1998
Birth of healthy children after intracytoplasmic sperm injection in two couples with male Kartagener’s syndrome Annette von Zumbusch, M.D.,* Klaus Fiedler, M.D., Artur Mayerhofer, M.D.,† Berthold Jeßberger, M.D.,* Johannes Ring, M.D.,* and Hermann-Josef Vogt, M.D.* Technische Universita¨t Mu¨nchen; Study Group Noxenkatalog-Databank (Hazars in Occupation and Environment); Department of Reproductive Medicine, Frauenklinik Dr. W. Kru¨smann, Munich, Germany
Objective: To describe two cases in which intracytoplasmic sperm injection (ICSI) was successful for patients with infertility due to Kartagener’s syndrome. Design: Case report. Setting: Private hospital for gynecology department of reproductive medicine, and university hospital center for andrology. Patient(s): Two couples with primary infertility due to Kartagener’s syndrome in the male. Intervention(s): ICSI. Main Outcome Measure(s): Pregnancy and birth after ICSI. Result(s): In both couples, ICSI was successful in the first cycle. The uncomplicated pregnancies resulted in the birth of three healthy children. One female and male/female twins.
Received November 10, 1997; revised and accepted June 11, 1998. Presented in part at the “Gemeinsame Tagung der o¨sterreichischen Gesellschaft fu¨r Gyna¨kologie und Geburtshilfe und der Bayerischen Gesellschaft fu¨r Geburtshilfe und Frauenheilkunde,” Graz, Austria, May 29 –31, 1997. Reprint requests: Annette von Zumbusch, M.D., Klinik fu¨r Dermatologie und Allergologie, Biederstein, Technische Universita¨t Mu¨nchen, Biedersteinerstrasse 29, D-80802 Mu¨nchen, Germany (FAX:0049-89334983; e-mail: ANNETTE .VONZUMBUSCH@LRZ .TU-MUENCHEN.DE). * Department of Dermatology and Allergology, Technische Universita¨t Mu¨nchen. † Anatomical Institute, Technische Universita¨t Mu¨nchen. 0015-0282/98/$19.00 PII S0015-0282(98)00246-5
Conclusion(s): In couples with infertility due to Kartagener’s syndrome in the male, ICSI has proved to be a successful therapy resulting in clinically healthy offspring. This knowledge may improve our understanding of the involvement of paternally inherited centrosomes, which nucleate microtubules, in human reproduction. (Fertil Sterilt 1998;70:643– 6. ©1998 by American Society for Reproductive Medicine.) Key Words: Kartagener’s syndrome, immotile cilia syndrome, ICSI (intracytoplasmic sperm injection), centrosome, centriole, microtubule
The terms “immotile cilia syndrome” and “primary ciliary dyskinesia” are used to describe a group of inherited diseases that generally follow an autosomal recessive trait. They are characterized by defective ciliary ultrastructure (axoneme defects, mainly lack of dynein arms and spoke structures) (1) associated with reduced or completely absent ciliary motility. The heterogeneity of the symptoms seen in immotile cilia syndrome may be due to multiple genetic alterations. The molecular nature of such defects has not yet been identified. Kartagener’s syndrome is one of the disorders classified as primary ciliary dyskinesia. In addition to the cilial defects found in this group of diseases, Kartagener’s syndrome is clinically distinguishable by the presence of situs inversus (1, 2). Immotile cilia syndrome results in characteristic clinical symptoms, including chronic pansinusitis, recurrent infections of the respiratory tract, bronchiectasis, and infertility,
particularly in the male. The similarities in ultrastructure between the sperm flagella and cilia of the respiratory epithelial cells may account for the respiratory problems and the impairment or absence of sperm motility in these patients. Female patients, in contrast, are usually fertile, which may be due to muscular motility of the fallopian tubes (2). Intracytoplasmic sperm injection (ICSI) is used in cases in which the male partner has immotile spermatozoa. Stalf et al. (3) reported pregnancy and birth after ICSI with spermatozoa obtained from a patient with tail-stump syndrome. However, in a recent report by Papadimas et al. (4), pregnancy was not achieved when ICSI was performed with spermatozoa from a patient with immotile cilia syndrome. The reasons for this failure are not clear. To our knowledge, we describe the first cases in which ICSI was used successfully to achieve pregnancies in couples who were pre643
viously infertile because of male Kartagener’s syndrome. Two couples were treated with ICSI, which resulted in the birth of three healthy children.
CASE REPORTS
FIGURE 1 Electron micrograph of a sperm flagellum from patient 1. Cross-section of the midpiece of the flagellum shows a 912 arrangement and lack of both the inner and outer dynein arms (bar 5 0.5 mm).
Case 1 A 34-year-old man and his 32-year-old wife were referred to our andrology outpatient department because of a 6-year history of infertility. Gynecologic examination of the wife did not reveal any abnormalities. A semen analysis that was performed in a private practice had revealed that the man had a normal number of immotile spermatozoa. The patient reported no medical problems, and the clinical examination showed no pathologic findings. The patient had three brothers, one of whom had chronic bronchitis and a history of infertility. Radiographic examination of the brother showed situs inversus viscerum. The other brothers had children. A roentgenogram of the patient’s chest showed situs inversus viscerum. Computed tomography of the skull revealed polyposis nasi and pansinusitis. Consanguinity of the male patient’s parents was not known. Semen Analysis Semen was collected by masturbation into a sterile plastic cup. Twenty minutes was allowed for liquefaction, after which a standard semen analysis was performed. The sperm concentration was 75 3 106/mL, and the semen volume was 9.5 mL. The most remarkable feature was the complete absence of motile forms. Staining with aqueous eosin solution showed 90% viable spermatozoa with unremarkable morphology. Transmission Electron Microscopy (TEM) A sperm aliquot was fixed in a solution of 2.5% glutaraldehyde and 0.2 M cacodylate buffer. After centrifugation, the sample was resuspended in 0.2 M sodium cacodylate buffer. The specimen was postfixed in osmium tetroxide, dehydrated through a series of ascending concentrations of ethanol, and then embedded in Araldite (Araldite Kit 2000; Bio-Rad Laboratories, Munich, Germany). Ultrathin sections (50 –70 nm) were cut (LKB III ultramicrotome), mounted on copper grids, and contrasted with uranyl acetate followed by incubation with 2% lead citrate. The sections were examined with a Zeiss transmission electron microscope (Zeiss, Oberkochen, Germany) and then photographed. A total of 150 sperm were evaluated using TEM. Both 912 and 910 configurations were observed. In 90% of the spermatozoa, both inner and outer dynein arms were missing (Fig. 1). The inner dynein arms were present in 10%. In 80% of the sperm, central fibrils were missing or incomplete. Radial spokes were missing in 70% and appeared to be 644
von Zumbusch et al.
ICSI for Kartagener’s syndrome
defective in 10%. Examination of sperm heads revealed normal morphology.
Case 2 A 36-year-old man was referred to our clinic because of infertility. Several members of his large family were known to have immotile cilia syndrome, some of them without situs inversus. This family is being monitored by the Institute for Human Genetics, University of Erlangen, Germany. The patient did not have a history of medical problems. On the basis of situs inversus viscerum and immotile sperm tails, his condition was diagnosed as Kartagener’s syndrome. His wife’s gynecologic examination revealed no abnormalities. Semen analysis previously had demonstrated oligoasthenozoospermia. Semen was collected and prepared as described in case 1. The sperm concentration was 210 3 106/mL and the semen volume was 1 mL. The most noteworthy feature was the complete absence of motile forms. Ninety percent of the sperm were viable, as evaluated with eosin staining, and the morphology was normal. TEM was not performed in this patient because Kartagener’s syndrome had been clinically diagnosed. Vol. 70, No. 4, October 1998
Fertilization Procedure in Both Cases The procedure was approved by the Institutional Review Board at the Frauenklinik Dr. W. Kru¨smann. After down-regulation with a GnRH analogue (triptorelin 0.1 mg, Decapeptyl; Ferring, Kiel, Germany) starting on day 24 of the previous cycle, 150 IU of FSH (Fertinorm HP; Serono, Munich, Germany) was administered daily from the third day of the treatment cycle until the leading follicle reached a mean diameter of 18 mm. Thirty-six hours before oocyte retrieval, 5,000 IU of hCG (Pregnesin 5.000; Serono) was injected. Six metaphase II oocytes were retrieved from both patients by transvaginal ultrasound-guided follicle puncture. Spermatozoa were washed twice by suspension in culture medium (Hams F-10 medium) and were resuspended after centrifugation. The final dilution was placed under a layer of paraffin oil. ICSI was performed in both cases according to the method of Van Steirteghem et al. (5). In patient no. 1, four of six injected oocytes were fertilized. Two days after follicle puncture, two preimplantation embryos at the 4-cell stage were transferred. This resulted in a routine pregnancy and birth of a clinically healthy set of twins (female and male). In patient no. 2, three of six injected oocytes were fertilized and were transferred at the 4- and 6-cell stages 2 days after oocyte retrieval. A normal pregnancy and birth of a healthy girl followed treatment.
DISCUSSION Although the diseases of the immotile cilia syndrome are heterogeneous, they are believed to be due to a microtubuleassociated defect. This defect has not yet been fully characterized (1, 2, 6, 7). Microtubules are cytoskeletal elements essential for many crucial processes in all cells. With respect to germ cells, they are responsible for separation of the chromosomes during mitosis and meiosis, as well as migration of the sperm and egg nuclei during fertilization. The microtubules also form the basis for ciliary processes found in spermatozoa and in respiratory epithelial cells. It is hypothesized that one or more microtubule-associated defects may be detrimental to ciliar motility and/or mitotic or meiotic divisions, therefore hindering fertilization. Thus, a microtubule defect in Kartagener’s syndrome may be responsible for more than ciliary dyskinesia leading to immotile sperm. It is possible that previous attempts at ICSI in cases of Kartagener’s syndrome were unsuccessful because of this defect (4, 8). In this context, it is important to recall that in humans, in contrast to mice (9) or hamsters (10), the spermatozoon introduces the centrosome during insemination into the oocyte. Subsequently, the centrosome nucleates the new microtubule assembly to form the sperm aster (9, 11). The FERTILITY & STERILITYt
resulting microtubule orientation and polarity are determined by the centrosome. Because of the cited species differences, the hamster oocyte penetration test may not be valid for demonstrating reliable fertilizing ability of spermatozoa from patients with immotile cilia syndrome (10). Centrosomes and centrioles are poorly understood cellular structures. They are both self-reproducing organelles, and centrioles merely advertise the presence of centrosomes. Human spermatozoa also have centrioles. A well-defined proximal centriole is present next to the basal plate of the sperm head, whereas the distal centriole (which is a remnant) gives rise to the sperm tail axoneme during spermiogenesis (12). Centrosomes are reduced during gametogenesis. It is assumed that successful fertilization requires centrosome restoration (in which the sperm plays the central role) and microtubule-mediated motility of parental chromosomes at first mitosis. Investigations have shown that defects in microtubule organization result in failure to fertilize. Microtubule inhibitors prevent pronuclear fusion, which underscores the essential role of the sperm astral microtubules for organizing under a paternal direction (12, 13). It is possible that these processes important for fertilization could be defective in some patients with Kartagener’s syndrome. To our knowledge, this is the first report to document the successful use of ICSI in two cases of male Kartagener’s syndrome that resulted in the birth of healthy children of both sexes. Papadimas et al. (4) had limited success treating a male patient with immotile cilia syndrome. After ICSI was performed, 33% of the oocytes were fertilized and subsequently divided but did not implant. Nijs et al. (18) described ICSI in a patient with Kartagener’s syndrome. None of the oocytes fertilized after ICSI, but 3 of 12 oocytes fertilized after subzonal insemination, which suggests normal microtubule function in fertilization. The spermatozoa of the two men in our cases were completely immotile. As noted in case 1, marked structural axoneme defects were present, including the loss of all outer and most of the inner dynein arms. ICSI in both cases was successful in the first cycle. This implies that from the phenotype alone (immotile sperm syndrome and structural defect), no clear prediction of the success of fertilization can be made. It is likely that the active reproducing centrosome is a blend of both maternal proteins and the spermatozoon’s centrosome (12). Whether the maternal or paternal contribution at this point is variable, or whether alternative pathways exist, is a subject worthy of further investigation. The use of ICSI in the two cases of male Kartagener’s syndrome described herein resulted in the birth of three clinically healthy children without situs inversus. These results seem to ethically justify the use of assisted reproductive technology in similar cases. As with all hereditary diseases in which the nature of the defect is unknown and the symp645
toms are heterogeneous, the result of reproductive intervention is ultimately uncertain. In patients with immotile cilia syndrome, ICSI and other procedures leading to conception may result in the birth of seemingly healthy children, but there is a possibility that fertility problems and other medical problems will be diagnosed later. All patients must be informed about these risks and receive appropriate genetic analyses and counseling before the use of ICSI is considered.
4.
5.
6. 7. 8. 9. 10.
Acknowledgment: The authors thank Mrs. Else Marais for editorial assistance and critical review of the manuscript. 11.
References
12.
1. Afzelius BA. The immotile-cilia syndrome: a microtubule-associated defect. CRC 1985;19:63– 87. 2. Rott H-D. Kartagener’s syndrome and the syndrome of immotile cilia. Hum Genet 1979;46:249 – 61. 3. Stalf T, Sanchez R, Kuhn F-M, Schalles U, Kleinstein J, Hinz V, et al. Pregnancy and birth after intracytoplasmic sperm injection with sper-
646
von Zumbusch et al.
ICSI for Kartagener’s syndrome
13.
matozoa from a patient with tail stump syndrome. Hum Reprod 1995; 10:2112– 4. Papadimas J, Tarlatzis BC, Bili H, Sotiriadis T, Koliakou K, Bontis J, et al. Therapeutic approach of immotile cilia syndrome by intracytoplasmic sperm injection: a case report. Fertil Steril 1997;67: 562–5. Van Steirteghem AC, Liu J, Joris H, Nagy Z, Janssenswillen C, Tournaye H, et al. Higher success rate by intracytoplasmic sperm injection than by subzonal insemination. Report of a second series of 300 consecutive treatment cycles. Hum Reprod 1993;8:1055– 60. Aitken RJ, Ross A, Lees MM. Analysis of sperm function in Kartagener’s syndrome. Fertil Steril 1983;40:696 – 8. Aitken J. A clue to Kartagener’s. Nature 1991;353:306. Nijs M, Vanderzwalmen P, Vandamme B, Segal-Bertin G, Lejeune B, Segal L, et al. Fertilizing ability of immotile spermatozoa after intracytoplasmic sperm injection. Hum Reprod 1996;11:2180 –5. Schatten H, Schatten G, Mazia D, Balczon R, Simerly C. Behavior of centrosomes during fertilization and cell division in mouse oocytes and in sea urchin eggs. Proc Natl Acad Sci USA 1986;83:105–9. Hewitson L, Haavisto A, Simerly C, Jones J, Schatten G. Microtubule organization and chromatin configurations in hamster oocytes during fertilization and parthenogenetic activation, and after insemination with human sperm. Biol Reprod 1997;57:967–75. Palermo G, Munne S, Cohen J. The human zygote inherits its mitotic potential from the male gamete. Hum Reprod 1994;9:1220 –5. Sathananthan AH, Kola I, Osborne J, Trounson A, Ng SC, Bongso A, et al. Centrioles in the beginning of human development. Proc Natl Acad Sci USA 1991;88:4806 –10. Simerly C, Wu G-J, Zoran S, Ord T, Rawlins R, Jones J, et al. The paternal inheritance of the centrosome, the cell’s microtubule-organizing center, in humans, and the implications for infertility. Nature Med 1995;1:47–52.
Vol. 70, No. 4, October 1998