PGD in 47,XXY Klinefelter's syndrome patients

The use of ICSI has been a major breakthrough in the treatment of male infertility. Even azoospermic patients with focal spermatogenesis in the testis, may benefit from the ICSI technique in order to father a child. As ICSI use has become more common, centres have introduced infertility treatment for Klinefelter patients. To date, 34 healthy children have been born using ICSI without PGD, and the conception of one 47,XXY fetus has been reported. In view of the possible risk of an increased gonosome number in the spermatozoa of Klinefelter patients, a safer approach--offering these couples ICSI combined with PGD--has been used, and has resulted in the birth of three healthy children. Couples in which the male suffered from Klinefelter's syndrome were first treated in 1995; these patients were offered ICSI + PGD using FISH technology, notably to enumerate the X and Y chromosomes. ICSI + PGD was performed in 32 cycles of 20 couples with spermatozoa originating from a fresh ejaculate (n = 1), testicular biopsy (n = 21) or frozen-thawed testicular biopsy (n = 10). Normal fertilization occurred in 56.0 +/- 22.4% of the successfully injected oocytes. On day 3 of development, 119 embryos from 29 cycles were of sufficient quality to undergo biopsy and subsequent PGD; a positive result was obtained in 113 embryos. Embryos were available for transfer in 26 cycles, with a mean of 1.6 +/- 0.6 embryos per transfer. Eight pregnancies were obtained, and five resulted in a delivery. A total of 113 embryos from couples with Klinefelter's syndrome was compared with 578 embryos from control couples with X-linked disease where PGD was used to determine gender. A significant fall occurred in the rate of normal embryos for couples with Klinefelter's syndrome (54.0%) compared with controls (77.2%). Moreover, a significantly increased risk of abnormalities was observed for sex chromosomes and autosomes; for each autosome separately, this reached significance level for chromosomes 18 and 21 only. Hence, a cautious approach is warranted in advising couples with non-mosaic Klinefelter's syndrome. Moreover, the use of ICSI + PGD or prenatal diagnosis should be carefully considered.     

Pregnancies achieved after frozen-thawed pronuclear oocytes obtained by intracytoplasmic sperm injection with spermatozoa extracted from frozen-thawed testicular tissues from non-obstructive azoospermic men.

The use of frozen-thawed testicular tissue as a source of spermatozoa for intracytoplasmic sperm injection (ICSI) in non-obstructive azoospermia yields favourable fertilization and pregnancy rates while avoiding both repetitive biopsies and unexpected cycle cancellations. Spermatozoa were obtained from frozen-thawed testicular biopsy specimens from 67 non-obstructive azoospermic men. Following fertilization, supernumerary two pronuclear (2PN) oocytes were frozen. After thawing, 17 cycles of embryo transfer were carried out with a mean number of 2.7 embryos and a mean cumulative embryo score (CES) of 18.3 per transfer. The clinical pregnancy and implantation rates per transfer in these cycles (23.5 and 8.3% respectively) were comparable to those of fresh embryo transfers (35.7 and 12.7% respectively) with a mean number of 2.7 embryos and a mean CES of 28.7 per transfer. Abortion rates, although higher with cryopreserved 2PN oocytes were not significantly different. With this approach, cryopreservation of supernumerary 2PN oocytes can be used to improve the cumulative pregnancy rates in a severely defective spermatogenetic population. To our knowledge, these are the first pregnancies reported which have been obtained by the transfer of cryopreserved pronuclear oocytes obtained from ICSI using cryopreserved testicular spermatozoa.     

Fine needle aspiration versus open biopsy for testicular sperm recovery: a controlled study in azoospermic patients with normal spermatogenesis

This retrospective controlled study aimed at comparing two techniques for recovering testicular spermatozoa in azoospermic patients undergoing intracytoplasmic sperm injection (ICSI). 102 men suffering from infertility because of obstructive azoospermia had ICSI using testicular spermatozoa recovered either by open excisional biopsy (n = 51), or by fine needle aspiration (FNA) (n = 51). A higher average number of spermatozoa were recovered after open biopsy than after FNA, but no significant differences in either fertilization rates or cleavage rates were observed after ICSI with spermatozoa retrieved by the two techniques. Neither was there any significant difference in ongoing pregnancy and implantation rates: in the FNA group, these figures were respectively 19.6% per cycle and 7.8% per embryo transferred and in the open biopsy group 21.6 and 7.1%. We conclude that ICSI with testicular spermatozoa recovered by FNA yields results comparable to those obtained with spermatozoa recovered by open biopsy in azoospermic patients with normal spermatogenesis. However a prospective study is needed to confirm the present results and to assess recovery rates and patient comfort for the two methods.      

Birth of a healthy neonate following the intracytoplasmic injection of testicular spermatozoa from a patient with Klinefelter's syndrome

Klinefelter's syndrome is one of the known causes of azoospermia or cryptoazoospermia, and it may present in non-mosaic (47,XXY) or mosaic (47,XXY/46,XY) form. The likelihood of finding spermatozoa in the ejaculate or testicular tissue of patients with mosaic Klinefelter's syndrome is low, and with the non-mosaic form, even lower. We describe a patient with non-mosaic Klinefelter in whom initially non-motile spermatozoa were derived from searching the ejaculate. Ten mature oocytes were injected, but none was fertilized. Subsequently, testicular biopsy was undertaken in order to collect spermatozoa for oocyte injection. Fifteen motile sperm cells were found and injected. Nine oocytes were fertilized and cleaved; three embryos were transferred into the uterine cavity. The woman conceived and following a normal pregnancy delivered a healthy child. Genetic analysis of the neonate disclosed a normal 46,XY karyotype. Non-motile spermatozoa in the ejaculate did not prove their fertilization potential, but their presence did not exclude finding motile, fertile spermatozoa in the testicular tissue in a non-mosaic Klinefelter patient. This report is further evidence that normal spermatozoa with fertilization potential are produced in the testes of patients with Klinefelter's syndrome.     

Blastocyst transfer following intracytoplasmic injection of ejaculated, epididymal or testicular spermatozoa

Recent studies indicate a strong paternal influence on embryo development and progression of the embryo to the blastocyst stage. The aim of this study was to compare, during extended culture, the in-vitro development of embryos resulting from intracytoplasmic sperm injection (ICSI) of ejaculated spermatozoa (group 1, n = 347), epididymal (group 2, n = 22) or testicular (group 3, n = 18) spermatozoa from obstructive azoospermic and testicular spermatozoa from non-obstructive azoospermic (group 4, n = 31) subjects. Fertilization and blastocyst formation rates were significantly lower in group 4 (P < 0.05). The incidence of expanded and hatching blastocysts was significantly lower in group 4 (P < 0.05). Overall in 93.2% ejaculate ICSI cycles, blastocysts were transferred on day 5. This was significantly higher than the 62% day 5 transfers in the non-obstructive azoospermic group (P < 0.05). Implantation rate per embryo was significantly higher in the ejaculate ICSI group compared with the other groups (P < 0.05). Clinical pregnancy per transfer was similar between groups; however, significantly fewer multiple pregnancies were encountered in the non-obstructive azoospermic group (P < 0.01). In conclusion, the source of the spermatozoa, most likely to be indicative of the severity of spermatogenic disorder, affects the rate of blastocyst formation and blastocyst implantation. Spermatozoa from non-obstructive azoospermic subjects, when utilized for ICSI, result in embryos that progress to the blastocyst stage at a lower and slower rate and implant less efficiently.     

Different fertilization rates between immotile testicular spermatozoa and immotile ejaculated spermatozoa for ICSI in men with Kartagener's syndrome: case reports

We report two cases of infertility treatment in couples where males suffered from Kartagener's syndrome (KS) and a total absence of motile sperm in the ejaculate. A total of three ICSI cycles was carried out. In all cycles, viable ejaculated or testicular spermatozoa were selected using the hypo-osmotic swelling (HOS) test. Case 1: In the first ICSI cycle total fertilization failure occurred after using ejaculated spermatozoa. In the following cycle testicular spermatozoa were used for ICSI, resulting in 75% fertilized oocytes and a pregnancy. Case 2: In the same ICSI cycle 50% of the oocytes were injected with ejaculated and 50% with testicular spermatozoa. The fertilization rates were 44 and 56% respectively and high quality embryos were achieved in both groups. One single embryo derived from testicular sperm was transferred with a resulting singleton pregnancy. In conclusion, testicular sperm for ICSI seem to have reliable fertilization capacity in men with KS, while ejaculated sperm, even if tested viable, seem more unpredictable. HOS test for selection of viable sperm for ICSI is recommended when ejaculated as well as testicular sperm are used for ICSI.     

Assisted reproduction by intracytoplasmic sperm injection: a survey on the clinical experience in 1994 and the children born after ICSI, carried out until 31 December 1993. ESHRE Task Force on Intracytoplasmic Sperm Injection. European Society for Human Reproduction and Embryology

The second report of the ESHRE Task Force on ICSI describes the outcome of 13,666 ICSI cycles carried out in 1994 by 90 centres in 24 countries. Most cycles used ejaculated spermatozoa (94.4%) while epididymal and testicular spermatozoa were used in 4.1% and 1.5% of the cycles. Outcome measures in the three types of spermatozoa included the number of: (i) intact oocytes after ICSI; (ii) normally fertilized oocytes; (iii) transferred and frozen embryos; (iv) embryo transfers and (v) cycles with positive serum HCG. The evolution of the pregnancies was analysed in terms of pregnancy losses and clinical pregnancies. The results of ICSI with spermatozoa from the ejaculate was analysed according to the year that ICSI started in the different centres. The survey also reports the follow-up of children born after ICSI carried out until 31 December 1993. A total of 455 pre- and postnatal karyotypes revealed the presence of nine abnormal karyotypes. Twenty-four centres reported on 807 ICSI children: 763 using ejaculated spermatozoa, 36 using epididymal spermatozoa and eight using testicular spermatozoa. Sixteen major congenital malformations were reported.      

Azoospermic men with deletion of the DAZ gene cluster are capable of completing spermatogenesis: fertilization, normal embryonic development and pregnancy occur when retrieved testicular spermatozoa are used for intracytoplasmic sperm injection

Some men with non-obstructive azoospermia harbour fully formed spermatozoa within their testicular tissue that can be used to achieve pregnancy via intracytoplasmic sperm injection (ICSI). Recently, Reijo et al. (1995) provided compelling evidence that the DAZ gene cluster is a strong candidate for one of the elusive azoospermia factors (AZF) located on the long arm of the Y chromosome. The DAZ gene cluster is deleted in 13% of azoospermic men and a small percentage of severely oligozoospermic men. Vertical transmission from father to son of AZF region deletions has also been described. Presumably these fathers were oligozoospermic. This led us to ask whether the azoospermic male with deletions of the AZF/DAZ region can also complete minimal spermatogenesis and whether any spermatozoa found could participate in fertilization, embryo development and pregnancy. Three out of six (50%) of the azoospermic men with AZF/DAZ deletions had spermatozoa identified within their harvested testicular tissue. When these spermatozoa were used for ICSI, fertilization occurred in 36% of injected oocytes. This compared favourably with testicular spermatozoa retrieved from non-obstructive azoospermic men without AZF/DAZ gene deletions. In one case, a twin conception resulted, which represents the first term pregnancy reported using spermatozoa from an AZF/DAZ deleted azoospermic male. Therefore it is necessary to take the possibility of transmission of infertility or sterility to our patients' offspring seriously when utilizing today's reproductive technologies, as spermatogenesis in men with AZF/DAZ deletions is by no means an exceptional occurrence.      

In-vitro fertilization treatment for severe male factor: the fertilization potential of immotile spermatozoa obtained by testicular extraction

A retrospective analysis in 50 couples of 53 cycles of intracytoplasmic sperm injection (ICSI) with immotile spermatozoa from testicular-retrieved spermatozoa was performed to evaluate whether total immotile spermatozoa achieved after testicular sperm extraction could fertilize ova and result in pregnancies. We assessed the efficacy of ICSI with totally immotile testicular spermatozoa extracted from the testes of azoospermic patients with severe spermatogenic failure (group 1) and compared these results with those from spermatozoa which were recovered after several hours of incubation and were motile (group 2) at the time of injection. In 19 cycles, only totally immotile spermatozoa were injected at the time of ICSI. For the remaining 34 cycles, at least one motile spermatozoon was found for injection. The oocyte fertilization rates were 51% for group 1 and 62% for group 2 (P < 0.02). Eighteen of 19 cycles in group 1 (90%) and all 34 (100%) cycles in group 2 had embryos for replacement. The mean number of embryos per cycle was 5.2 +/- 0.8 and 7.5 +/- 0.9 in groups 1 and 2 respectively; this and the embryo quality (cumulative embryo scoring = 40 +/- 8 for group 1 and 50 +/- 7 for group 2), and clinical pregnancy rates (15.8% per oocyte retrieval in group 1 and 23.5% in group 2) were not significantly different between groups. Fertilization, cleavage and pregnancy can be achieved with intracytoplasmic testicular sperm injection from patients with immotile spermatozoa, at levels comparable with those of ICSI using motile spermatozoa.     

Genetics: Preimplantation diagnosis for X and Y normality in embryos from three Klinefelter patients

In some 47,XXY Klinefelter patients without evidence of mosaicism,testicular spermatozoa can be successfully recovered and usedfor intracytoplasmlc sperm Injection (ICSI). To ensure the replacementof embryos with a normal X and Y chromosome pattern, preimplantationdiagnosis can be performed. This paper reports on three 47,XXYKlinefelter patients from whom it was possible to retrieve testicularspermatozoa in order to perform ICSI. From their healthy wivesa total of 27 oocyte-cumulus complexes were retrieved from which22 metaphase-II oocytes were obtained and injected; 19 of thesewere intact after injection. Two distinct pronuclel were observedin eight oocytes (42.1%) 18 h after injection. On day 3 of development,five embryos (62.5%) had reached at least the 6-cell stage andwere of sufficient quality to undergo biopsy and subsequentpreimplantation diagnosis for sex chromosome analysis by fluorescentin-situ hybridization. The four embryos diagnosed as normalwere transferred to three respective patients, resulting Inone biochemical pregnancy. The remaining cells of the fifthembryo were analysed afterwards, revealing a normal X and Ychromosome constitution. So far, in the five embryos diagnosed,a normal sex chromosome pattern has been observed.     

The chromosomal constitution of embryos developing from abnormally fertilized oocytes after intracytoplasmic sperm injection and conventional in-vitro fertilization

The aim of this study was to analyse the chromosomal constitution of embryos developing from mono- (1PN) and tripronuclear (3PN) oocytes, after in-vitro fertilization (IVF) and after intracytoplasmic sperm injection (ICSI) into oocytes, by means of the fluorescent in-situ hybridization (FISH) technique with specific probes for the chromosomes X, Y and 18. FISH analysis was carried out on embryos from 3PN oocytes: 106 after ICSI and 71 after conventional IVF. In the 3PN embryos after ICSI, equal ratios of XXX and XXY were observed and no XYY embryos were present. This shows the digynic origin of such 3PN embryos. On the other hand, after conventional IVF, the XYY status indicative of dispermic fertilization was observed in some embryos. After IVF, only 12.7% of the 3PN oocytes developed into embryos with uniformly triploid blastomeres, compared with 55.7% after ICSI (P < 0.001). On the other hand, after ICSI only 16.0% of the embryos developing from 3PN oocytes were mosaic, compared with 42.3% after conventional IVF (P < 0.001). FISH was also carried out on embryos from 1PN oocytes: 61 after ICSI and 115 after conventional IVF. In 35.6% of IVF embryos developing from 1PN oocytes Y-specific hybridization signals were observed. This indicates that in 70-75% of such cases a spermatozoon had penetrated the oocyte and that only 25-30% of them were parthenogenetic. A significantly higher proportion (P < 0.001) of embryos developing from 1PN oocytes were diploid after IVF (48.7%) than after ICSI (27.9%); equal ratios of XX and XY embryos were observed in the two groups. Formation of a single pronucleus in an embryo subsequently shown to be diploid indicates that normal fertilization was followed by asynchronous formation of pronuclei. A significantly (P < 0.001) higher proportion of 1PN oocytes developed into haploid embryos after ICSI (31.2%) than after conventional IVF (13.1%). In both groups most of the haploid embryos were X-bearing (IVF, 93.3%; ICSI, 84.2%) and only a few were Y-bearing (IVF, 6.7%; ICSI, 15.8%). A contribution of normal fertilization and androgenetic activation thus led to 1PN oocytes. Gynogenetic and/or parthenogenetic activation, both leading to indistinguishable chromosomal distributions, also contributed to the formation of 1PN oocytes after ICSI and IVF.      

Outcome of ICSI using fresh and cryopreserved-thawed testicular spermatozoa in patients with non-mosaic Klinefelter's syndrome.

BACKGROUND: Recently, intracytoplasmic sperm injection (ICSI) of testicular spermatozoa retrieved surgically from patients with non-mosaic Klinefelter's syndrome resulted in several deliveries. The aim of this study was to evaluate the outcome of ICSI using fresh and cryopreserved-thawed testicular spermatozoa in these patients. METHODS AND RESULTS: Following informed consent regarding the genetic risks of their potential offspring, mature testicular spermatozoa were found in five out of 12 (42%) patients who underwent testicular sperm extraction, and ICSI was performed while excess tissue was cryopreserved. The mean age of the patients was 28.7 +/- 3.6 (range 23-36 years). Their baseline FSH was elevated (mean 38.3 +/- 11.4; range 22-58 mIU/ml). All patients had small testicles of 2-4 ml in volume. The outcome of ICSI using fresh or cryopreserved-thawed testicular spermatozoa during five cycles in each group, was compared. No statistical significant difference was found in the two pronuclear fertilization rate (66 versus 58%), embryo cleavage rate (98 versus 90%) and embryo implantation rate (33.3 versus 21.4%) for fresh or cryopreserved sperm accordingly. The clinical outcome after using fresh testicular sperm included two singleton pregnancies (one delivered and one ongoing) and a triplet pregnancy resulting in a twin delivery (after reduction of an 47,XXY embryo). After using cryopreserved-thawed testicular spermatozoa, two pregnancies were obtained resulting in one delivery of twins and one early spontaneous abortion. CONCLUSIONS: Outcome of ICSI using cryopreserved-thawed testicular spermatozoa of patients with non-mosaic Klinefelter's syndrome is comparable with that following the use of fresh spermatozoa. The genetic implications for the future offspring should be explained to the patients.     

Fertilization, pregnancy and embryo implantation rates after ICSI with fresh or frozen-thawed testicular spermatozoa

This study aimed to determine whether fertilization and implantation rates after intracytoplasmic sperm injection (ICSI) with fresh or frozen-thawed testicular spermatozoa were comparable. Between 1 January 1996 and 31 December 1996, 65 ICSI cycles with testicular spermatozoa and 35 cycles with frozen-thawed testicular spermatozoa were carried out. In 50 out of 65 ICSI cycles, testicular spermatozoa could be retrieved and in 34 out of 35 cycles carried out with frozen-thawed testicular spermatozoa, motile spermatozoa could be recovered. The fertilization rate after ICSI with frozen-thawed testicular spermatozoa was significantly lower (71.1%; P < or = 0.008) than with fresh testicular spermatozoa (79.3%). The pregnancy rate was similar for both groups (38.2 and 26.5 %). The implantation rate per transferred embryo, however, was significantly lower in the frozen-thawed rather than in the fresh testicular sperm group (9.1 versus 24.6%; P = 0.001). The live birth rate per transferred embryo was also higher in the group in which fresh testicular spermatozoa were used (18.8 versus 7.9% P = 0.043). This retrospective study shows that is possible to achieve a high fertilization rate after ICSI with both fresh and frozen-thawed testicular spermatozoa but implantation and live birth rates per transferred embryo, however, are significantly lower after ICSI with frozen-thawed than with fresh testicular spermatozoa.      

A comparison of ICSI outcomes with fresh and cryopreserved epididymal spermatozoa from the same couples

The published experience with frozen-thawed epididymal spermatozoa and intracytoplasmic sperm injection (ICSI) suggests that fertilization and pregnancy success rates are comparable to those achieved with freshly retrieved spermatozoa. However, no study has exactly compared clinical outcomes between the two IVF/ICSI cycles in the same couples. To formally address this issue, we assessed ICSI outcomes in couples each of whom had had two IVF/ICSI cycles: one using fresh and the second using frozen-thawed epididymal spermatozoa obtained from a single aspiration procedure. From a pool of 101 consecutive patients undergoing IVF/ICSI with epididymal spermatozoa, 19 couples initially used fresh epididymal spermatozoa and subsequently underwent a second IVF/ICSI procedure with frozen-thawed spermatozoa from the same aspiration. Normal (2PN) oocyte fertilization rates, embryo quality and pregnancy rates were compared between the two IVF/ICSI cycles for each couple. In the fresh epididymal sperm group, 58.4% of the injected oocytes fertilized normally compared with 62.0% of the injected oocytes in the frozen-thawed epididymal sperm group, revealing no statistically significant difference. Graded embryo quality also did not differ significantly between the paired IVF/ICSI cycles. The clinical pregnancy rates were 31.6% (6/19) and 36.8% (7/19) in the first and second cycles respectively. All but one pregnancy were singletons. In summary, this study provides strong evidence to support the notion that motile, cryopreserved and thawed epididymal spermatozoa are equal to freshly retrieved spermatozoa for ICSI in couples with obstructive azoospermia.     

Pregnancy achieved following ICSI from a man with Klinefelter's syndrome and spinal cord injury.

Klinefelter's syndrome and spinal cord injury are major causes of male infertility. Intracytoplasmic sperm injection (ICSI) is a relatively new method of assisted reproduction. A testicular biopsy was obtained from a patient with the double complications of non-mosaic 47,XXY Klinefelter's syndrome and spinal cord damage, and motile spermatozoa were collected. ICSI was then performed. Of the four sperm-injected oocytes, three became fertilized and cleaved. Two embryos were implanted, resulting in a single pregnancy with visible evidence of a heartbeat appearing at 6 weeks gestation. The pregnancy is now entering its 20th week. To the best of our knowledge, this is the first case of a pregnancy resulting from the sperm of a patient with double complications.     

Patients with absolutely immotile spermatozoa and intracytoplasmic sperm injection

The microinjection of completely immotile spermatozoa may impair the outcome of intracytoplasmic sperm injection (ICSI). Eleven couples underwent an initial ICSI cycle with 100% immotile freshly ejaculated spermatozoa. Two-pronuclear fertilization ensued in 18 of 145 (12.4%) successfully injected oocytes. None of these cycles resulted in a pregnancy. Nine couples underwent ICSI in subsequent cycles (n = 16). Ejaculated spermatozoa were injected in 15 cycles and testicular spermatozoa in one cycle. In 10 of the 15 cycles, motile spermatozoa were available at the time of injection. Motile testicular spermatozoa could also be injected. In the subsequent cycles, 91 of 176 (51.7%) successfully injected oocytes fertilized normally and four patients became pregnant. In the subsequent cycles where again immotile spermatozoa had to be injected no pregnancies occurred. In four subsequent cycles embryo cryopreservation was carried out. After replacement of two frozen-thawed embryos one additional pregnancy was obtained. In all, five healthy infants were born. It has been ascertained that motile spermatozoa can be detected either in repeated ejaculates or after testicular biopsy. The causes of total asthenozoospermia are variable and the problem is a sporadic rather than a permanent condition.      

A 47,XXY fetus conceived after ICSI of spermatozoa from a patient with non-mosaic Klinefelter's syndrome: case report

The birth of 12 healthy infants to fathers with non-mosaic Klinefelter's syndrome has been reported so far. The spermatozoa for these pregnancies was obtained from frozen-thawed ejaculate in one pregnancy (twins) and from the testis in the remaining 10 infants. All of them had a normal karyotype. We describe a patient with non-mosaic Klinefelter's syndrome from whom a testicular biopsy was obtained and motile spermatozoa were collected. Of 16 oocytes that were injected, 14 fertilized and cleaved. Three embryos were transferred, resulting in a triplet pregnancy. Karyotype analysis from chorionic villous sampling revealed 46,XX, 46,XY and 46,XXY from the three fetuses. The affected 46,XXY fetus was reduced on the 14th gestational week. The pregnancy culminated with the birth of a healthy male and female, on the 36th gestational week, weighing 3600 and 2660 g respectively. This case report proves the presence of hyperploid spermatozoa in the seminiferous lumen, and strengthens the necessity of genetic diagnosis of the embryos or fetuses in such pregnancies to fathers with non-mosaic Klinefelter's syndrome.     

Comparison of the fertilizing capability of spermatozoa from ejaculates, epididymal aspirates and testicular biopsies using intracytoplasmic sperm injection

A prospective study was carried out to compare the fertilizing capability and pregnancy outcome following intracytoplasmic sperm injection (ICSI) using spermatozoa obtained from ejaculates, or surgically from epididymis or seminiferous tubules. A total of 77 ICSI cycles (one per patient) was included. In all, 28 patients had severe oligoasthenoteratozoospermia, 19 patients had obstructive azoospermia and 30 patients had non-obstructive azoospermia. The main outcome measures were fertilization rate per injected metaphase II oocyte and the clinical pregnancy rate per embryo transferred back to the female recipients. In patients with severe oligoasthenoteratozoospermia, the fertilization and pregnancy rates were 79 and 25 %. In patients with obstructive azoospermia, for whom epididymal spermatozoa were used, these were 75 and 28%, and in the non-obstructive group for which testicular spermatozoa were used for injection, they were 69 and 21% respectively. These rates were not significantly different in the three groups (P = 0.85 and P = 0.14 respectively), suggesting that spermatozoa from the ejaculates and epididymal or testicular biopsies are able to fertilize equally by using ICSI. Live birth per embryo transfer was significantly reduced in patients with non-obstructive azoospermia compared to the other two groups. The high abortion rate (50%) in the group in which testicular spermatozoa were used raises doubts about the developmental competence of such embryos.      

一例嵌合型18三体少精子症患者精染色体分析及植入前遗传学诊断

目的 分析1例嵌合型18三体少精子患者精子18、X、Y染色体数目畸变并进行植入前遗传学诊断(preimplantation genetic djagnosis,PGD).方法 采用G带及荧光原位杂交(fluorescence in situ hybridjzation,FISH)对中期分裂相进行分析,应用三色探针CEP18、CEPY、Tel Xq/Yq对患者精子进行FISH分析,同时以1名染色体正常男性的正常精液作为对照,并对嵌合型18三体患者进行PGD.结果 患者精子18二体率、性染色体二体率和二倍体率分别为0.63%、0.94%和0.87%,与对照组相比(0.16%、0.35%、0.31%)差异有统计学意义.患者进行1个PGD周期的治疗、活检4个胚胎,移植正常的XY1818、XX1818各1胚胎后获得临床妊娠.结论 精子FISH分析可为其提供更准确的遗传咨询及指导植入前遗传学诊断,FISH-PGD可有效地应用于嵌合型18三体的植入前遗传学诊断.     

Repetitive ejaculation before intracytoplasmic sperm injection in patients with absolute immotile spermatozoa

Cases with absolute immotile sperm syndrome are rare, and include the genetic defect of immotile cilia syndrome with the absence of dynein arms in the flagellum. We attempted to increase the percentage of viable spermatozoa to improve the outcome of intracytoplasmic sperm injection (ICSI). Three couples in whom repeated analysis of the male partners indicated 100% sperm immotility underwent an in-vitro fertilization (IVF) procedure in which ICSI was performed. On their first ICSI cycle the males produced a single ejaculation while in their successive ICSI cycles they were requested to repeatedly ejaculate (two to four times) and only the last ejaculation was used. The eosin-Y test was performed on each used sample. Following their first treatment, one couple had one repeated treatment cycle, another had two and the third couple had three repeated treatment cycles. The mean percentages of viable spermatozoa were 41+/-7.4 and 71+/-6.9% in the first and repeated cycles respectively (P < 0.01; t-test). Of the 39 oocytes injected in the first ICSI cycles only one (3%) was normally fertilized (2PN) compared with 41 (48%) of the 85 oocytes injected in the repeated ICSI cycles. One (3%) embryo in the first and 35 (41%) embryos in the repeated ICSI cycles respectively were obtained (P < 0.001), enabling their replacement into the uterine cavity in all the repeated cycles. One woman (in a repeated cycle) conceived a twin pregnancy and delivered two healthy babies. The use of spermatozoa from repeated ejaculation is recommended in men with absolutely immotile spermatozoa so as to obtain significantly better viability and fertilizing capacity.