A successful strategy for preimplantation genetic diagnosis of myotonic dystrophy using multiplex fluorescent PCR.

The most common form of inherited muscular dystrophy in adults is myotonic dystrophy (DM), an autosomal-dominant disease caused by the expansion of an unstable CTG repeat sequence in the 3' untranslated region of the myotonin protein kinase (DMPK) gene. Expanded (mutant) CTG repeat sequences are refractory to conventional PCR, but alleles with a number of repeats within the normal range can be readily amplified and detected. Preimplantation genetic diagnosis (PGD) of DM has been successfully applied. However, a misdiagnosis using the reported protocol was recently documented. Two new PGD protocols for DM have been developed which utilise multiplex fluorescent PCR. Ideally a linked polymorphic marker, APOC2, is amplified in addition to the normal DMPK alleles, thus providing a back-up diagnostic result. However, the two couples reported in the present study were not fully informative at the APOC2 locus and so an unlinked short tandem repeat (STR) marker, D21S1414, was substituted. The highly polymorphic nature of the D21S1414, DMPK and APOC2 loci means that a very simple genetic fingerprint can be generated by analyses of these loci. This allows most DNA contaminants to be detected. Contamination is a significant problem for PGD and is the primary reason for the inclusion of D21S1414 and APOC2 in this protocol. This paper reports the first clinical experience and pregnancies following PGD for DM using a multiplex fluorescent PCR protocol.

Oocyte morphology predicts outcome of intracytoplasmic sperm injection.

To examine the influence of cytoplasmic morphology on the success rate of intracytoplasmic sperm injection (ICSI), the morphology of 837 metaphase II oocytes was assessed after cumulus stripping. The main abnormalities detected were excessive granularity, cytoplasmic inclusions such as vacuoles, smooth endoplasmic reticulum clustering and refractile bodies. Microinjection was performed in 538 oocytes with normal cytoplasm, 142 out of 161 with excessive granularity and 112 out of 138 with cytoplasmic inclusions. Very poor oocytes were not injected. No difference was found in fertilization rate. The embryos achieved cleaved normally and a similar number of good quality embryos among the three groups was noted. The outcome of transfer of embryos derived solely from normal oocytes (group A: 72 patients, 183 embryos) was compared with those from oocytes with cytoplasmic abnormalities (group B: 34 patients, 85 embryos). In group A, 17 clinical pregnancies (24% per patient, implantation rate 10%) were established. In group B, only one clinical pregnancy (3% per patient, implantation rate 1%) was established, from the transfer of embryos derived from oocytes with homogeneous granularity of the cytoplasm. No pregnancy resulted following the transfer of embryos from eggs with cytoplasmic inclusions. The difference was statistically significant. The outcome of ICSI is dependent on the quality of the oocytes retrieved. Normal fertilization and early embryo development were achieved in oocytes with abnormal cytoplasm morphology, but the resulting embryos failed to demonstrate the same implantation potential as those derived from oocytes with normal cytoplasm.

[Heterotopic ovarian pregnancy after in-vitro fertilization and embryo transfer and contralateral tubal pregnancy after gamete intrafallopian transfer]. / Gravidanza eterotopica ovarica dopo fertilizzazione in vitro ed embryo transfer e gravidanza tubarica controlaterale dopo trasferimento intratubarico dei gameti.

One case of ovarian ectopic pregnancy associated with twin intrauterine viable pregnancy after IVF-ET, and one case of contralateral tubal ectopic pregnancy after GIFT performed with normal fallopian tubes, are reported. The possible pathogenesis and management are discussed.

Oocyte donation in 61 patients.

Oocyte donation was performed by gamete intrafallopian transfer in 61 women, 34 of whom were amenorrhoeic. The mean age of the donors was 32 years (range 23-38). Seventy-five treatment cycles gave 29 clinical pregnancies, of which 21 reached term, 3 continue, and 5 were lost (3 miscarriages, 1 tubal, and 1 cervical). 11 (38%) of the women who became pregnant were over 42 years old. When more than four oocytes were transferred, many of the pregnancies were multiple. 8 (38%) of the pregnancies that came to term were complicated by pre-eclamptic toxaemia. When oocytes are obtained from young women, the fertility potential is high irrespective of the recipient's age.

Unexplained infertility--the value of Pergonal superovulation combined with intrauterine insemination.

Sixty-two women with unexplained infertility were studied. Fifteen (group 1) had timed intrauterine insemination (IUI), 25 (group 2) were treated by Pergonal (Serono Laboratories, Ltd., Welwyn Garden City, England) superovulation, and 22 (group 3) underwent Pergonal superovulation combined with IUI. Where Pergonal treatment was followed by insemination, a significantly greater pregnancy rate per cycle (P less than 0.05) was achieved, whether this group of patients was compared with those treated by IUI alone or with those treated with Pergonal alone. Moreover, the pregnancy rate in group 3 was comparable to that reported following gamete intrafallopian transfer (GIFT). The authors therefore suggest this form of treatment for patients with unexplained infertility prior to their referral to the more invasive procedure of GIFT.

Ovum donation--a simplified approach.

To date, ovum donation (OD) has involved luteinizing hormone (LH) synchronization between recipient and donor for normally cycling women, and a complex steroid replacement regimen given on a sequential and incremental basis for women with primary or secondary ovarian failure. The authors designed a simple hormonal regimen applicable to both normally cycling women starting early in the cycle, and to those with ovarian failure. It consists of administering 2 mg estradiol (E2) valerate orally three or four times daily, augmented with either 100 mg progesterone (P) in ethyl oleate intramuscularly daily or 100 mg oral progesterone (P) orally three times daily, starting on the day preceding the recovery of the donated oocytes. Gamete intrafallopian transfer procedure was undertaken for women with patent tubes and in vitro fertilization for those with obstructed tubes. The authors report their preliminary experience with 17 women who underwent ovum donation.