Semen quality of smoking and non-smoking men in infertile couples in a Swedish population.

A study of semen quality, using conventional semen analysis, was undertaken. 186 male smokers were compared with 164 non-smoking men undergoing infertility investigation. The cigarette smokers were subdivided into three groups according to the number of cigarettes they smoked: less than 10 (n = 27), 10-19 (n = 84) and greater than 19 cigarettes per day (n = 75). No statistically significant effect of cigarette smoking on sperm density, motility or morphologic features of sperm was detected. Nor was any significant difference in sperm quality, except for semen volume and total sperm count, disclosed between men in the different smoking categories or between heavy smokers and non-smoking men. Thus, the present study does not corroborate reports of detrimental effects of cigarette smoking alone on sperm concentration, motility or morphology on the bases of this population of Swedish men in infertile couples.

Dose previous Chlamydia trachomatis infection influence the pregnancy rate of in vitro fertilization and embryo replacement?

The association between previous chlamydial infection, as reflected by the presence of chlamydial antibodies (specific serum immunoglobulin G antibodies with a titer greater than or equal to 32) and pregnancy outcome after in vitro fertilization and embryo replacement was studied in 121 infertile women with tubal damage. The antibody prevalence was 74.4%; the overall pregnancy rate was 26.4%. No difference in seropositivity was detected between those who became pregnant and those who did not (71.9% versus 75.3%). The geometric mean titers were also similar in the two groups. Even after subdivision of the cases into primary or repeated in vitro fertilization attempts, or after stratification of the material according to the number of embryos used for replacement, there was no correlation between chlamydial antibodies and pregnancy rate. Thus, past infection with Chlamydia trachomatis did not influence the outcome of in vitro fertilization and embryo replacement treatment in this study.

Tubal infertility and silent chlamydial salpingitis.

Antibodies to Chlamydia trachomatis (serum IgG antibodies with a titre of at least 32) were detected in 141 (86.0%) of 164 infertile women with tubal infertility (TF group) and in 20 (28.6%) in 70 infertile women with normal tubes (NTF group). The difference was highly significant (P less than 0.001), as was the difference in geometric mean titre of antibody-positive cases, 181 and 87, respectively. Pregnant, age-matched women were used as controls. Significantly fewer of these women (P less than 0.001) had antibodies, when compared with the TF group, 60 (36.6%) of 164, but not when the comparison was made with the NTF group, 31 (44.3%) of 70. There was a history of salpingitis in 64 (39.0%) of the 164 infertile women with damaged tubes. Neither the frequency nor the geometric mean titre of chlamydial antibodies differed between tubal factor infertility patients with and without a history of salpingitis. Previous chlamydial infection, reflected by serological markers, is strongly associated with tubal damage leading to tubal infertility. A large proportion of these cases run a silent course, since a majority of the antibody-positive patients with tubal infertility have never had salpingitis.

A comparison between the hamster egg penetration test and the seminal parameters in men of infertile couples.

The zona-free hamster egg penetration test was performed on 74 random consecutive semen samples from men of infertile couples and on 7 men of proven fertility (semen donors). Of these 74 men, 31 had never before delivered a semen sample (Group I), while the remaining 43 had delivered samples before (Group II). The penetration rates were correlated with the parameters of the semen analysis. In Group I, the mean penetration rate was 59%, in Group II, 42%, and among the fertile donors, 52%. No strong correlation was found between hamster egg penetration rates and the various semen parameters. Only the proportion of motile spermatozoa in Group II correlated significantly with the egg penetration test (P less than .05). Differences in methodology make comparison with the results from different laboratories difficult. It is likely that the egg penetration test measures some separate quality of the spermatozoa that may not be clear from the routine semen analysis. The role of the hamster egg penetration test in the investigation of the causes of infertility should be evaluated further.

Pregnancy rate in relation to number of cleaved eggs replaced after in-vitro fertilization in stimulated cycles monitored by serum levels of oestradiol and progesterone as sole index.

A simple model for monitoring ovarian stimulation in in-vitro fertilization cycles was designed using daily serum levels of 17 beta-oestradiol and progesterone as the only index to determine the day of ovum retrieval. Human chorionic gonadotropin was administered in the evening 2 days after a level of 2500 pmol/l oestradiol in serum was exceeded provided the serum progesterone level was less than 5 nmol/l. Ovarian stimulation was initiated in 128 women scheduled for laparoscopic oocyte retrieval. Twelve cycles (9%) were cancelled before ovum pick up due to sub-optimal hormone levels in serum and five for other reasons. In 107 successful laparoscopies, 616 oocytes (mean 5.8 per laparoscopy) were recovered. The cleavage rate after IVF was 65% in the egg replacement group. In 87 women, a total of 271 cleaved eggs (range 1-6, mean 3.1) were replaced. The most important factor for establishing a pregnancy was the number of eggs replaced at the same time. Clinical pregnancies were achieved after 30% of replacements, increasing to 50% after replacement of 5-6 eggs. The ongoing/delivered pregnancy rate after the 18th week of gestation was 69%. It was concluded that the simple monitoring model used was consistent with a high pregnancy rate and a low rate of cancelled cycles.

Ploidy in human cleavage stage embryos after fertilization in vitro.

Chromosome preparations were made from 15 cleaved human embryos in the 2- to 12-cell stage after in-vitro fertilization. All showed two pronuclei before the first cleavage. Twelve had at least one diploid metaphase, while three had interphase nuclei only. There was no evidence of parthenogenetic haploid cleavage. As expected, the frequency of metaphases increased with duration of colchicine treatment: 25 and 55% of the cells reached metaphase after 6-13 and 16-24 hours' treatment, respectively. Cleaved embryos with 'ideal' blastomere numbers (2, 4 or 8) showed a considerably higher metaphase frequency than others. For a more detailed chromosome analysis the technique will have to be further improved. The asynchronous cleavage of blastomeres makes optimal treatment by mitogens difficult.

Chromosome analysis of human oocytes recovered from preovulatory follicles in stimulated cycles.

To investigate the incidence and types of abnormalities of chromosome number in oocytes, we recovered preovulatory oocytes from 17 women who were undergoing clomiphene stimulation and laparoscopy because of infertility. Twenty-three oocytes were recovered and studied after they had been fixed with a gradual-fixation method: 17 of the oocytes had numbers of chromosomes in the haploid range (19 to 25 second-metaphase chromosomes), 4 had only 1 to 5 chromosomes, 1 was not analyzable, and 1 had 23 chromosome bivalents in the first metaphase. Of the oocytes with chromosome numbers in the haploid range, nine had an apparently normal haploid set of 23 chromosomes. Two had 1 to 2 additional chromosomes, three lacked 2 to 4 chromosomes, and three had totals of chromosomes that were close to 23 but could not be determined with certainty. We conclude that infertile women undergoing clomiphene stimulation have a high proportion (nearly 50 percent) of oocytes with an abnormal karyotype. If this is also true of fertile women, including those not taking clomiphene, it may explain the high frequency of chromosome aberrations occurring in early spontaneous abortion and the low pregnancy rate after in vitro fertilization.

Importance of the use of short and long sperm pre-incubation periods in assessing the fertilizing capacity of human spermatozoa by hamster test.

The fertilizing capacity of semen samples of seven fertile men was studied in vitro using zona-free hamster ova (hamster test). Spermatozoa of each sample were pre-incubated 4 or 24 h before ova were inseminated for 3 h. An extended insemination for an additional 3 h was also performed after the shorter pre-incubation. The overall ovum penetration rate changed from 0 to 100% (mean 52%) after short (4 h) sperm pre-incubation and from 10 to 100% (mean 55%) after long (24 h) pre-incubation. Taking into account the combination of short and long pre-incubation periods, all donors reached a penetration rate of 67 to 100%. The results indicate that both short and long pre-incubation periods must be employed when the hamster test is used diagnostically to assess the fertilizing capacity of human spermatozoa.

Ultrastructural characteristics of human oocytes fixed at follicular puncture or after culture.

The material consisted of 73 oocytes obtained at follicle aspiration in ovarian-stimulated women. Oocytes in various stages of maturation were either immediately fixed or cultured before fixation. Observations by transmission electron microscopy disclosed that 20% of the immature oocytes, which appeared normal in the light microscope, had commenced atresia, and one immediately fixed oocyte surrounded by a normal-appearing preovulatory cumulus mass was found to be degenerated. Further, a cumulus mass judged as preovulatory contained an oocyte possessing a germinal vesicle. Light microscopy is thus not always adequate for judging the condition of ova surrounded by cumulus cells. Cytoplasmic changes which were regarded as being related to oocyte maturation were a decrease in the number of vacuoles and multivesicular bodies, an increase in the volume of the endoplasmic reticulum, a clustering of mitochondria, and the appearance of aggregates of tubuli of the smooth endoplasmic reticulum surrounded by mitochondria and mitochondria-vesicle complexes. Certain features were found to cause misinterpretations at examination of oocytes in a light microscope. For instance, polar bodies can be mimicked by corona cells in the perivitelline space and pronuclei in oocytes by large mitochondria-vesicle complexes in the ooplasm. Consequently, not all oocytes in which polar body-like or pronuclei-like structures are observed are necessarily fertilized oocytes.

A gradual fixation method for chromosomal preparations of human oocytes.

Eleven oocytes were recovered after follicle aspiration carried out in nine women during laparoscopy or laparotomy. All oocytes were classified as preovulatory, judged by the fully dissociated cumulus. After elimination of the cumulus, ten of the oocytes exhibited the first polar body, and one was degenerated. A gradual fixation method was used for the chromosomal preparation of the oocytes. The quality of the preparations was improved, compared with the previously used air-drying technique. Nine of ten preparations from oocytes with first polar body displayed chromosomes. These sets consisted of well-spread meiotic chromosomes, and in six of them the chromosome complements of the first polar body could be seen, although quite distinct from the oocyte complement. Of eight preparations where the chromosomes could be counted, six showed 23 chromosomes, and two were abnormal, showing 21 and 16 chromosomes.

Some clinical results of in vitro fertilization by the Malmö Group, Sweden.

In our in vitro fertilization program, only clomiphene/human chorionic gonadotropin-stimulated cycles have been used, yielding 1.6 preovulatory oocytes/woman at follicle aspiration. The cleavage rate was significantly influenced by the percentage of abnormal spermatozoa and by the percentage of motile spermatozoa in the spermiograms. At embryo replacement, a tilted uterus obtained by a full bladder in the woman simplifies the replacement procedure. In the last 12 months, 47 embryo transfers have been carried out, resulting in five clinical pregnancies. Of these pregnancies, one has proceeded to term, one resulted in a miscarriage, and three resulted in ectopic pregnancies.

Depressed semen quality: a study over two decades.

One hundred-eighty-five consecutive spermiograms from men of infertile couples in 1980-1981 were compared with as many spermiograms of age-matched controls from 1960-1961. The semen parameters analyzed were: volume, sperm density, and sperm morphology. The following significant changes were noted over the time period: the mean seminal volume decreased from 3.79 ml to 3.42 ml (p less than 0.05), the mean sperm count decreased from 125.4 mill/ml to 78.0 mill/ml (p less than 0.001) and the proportion of double sperm heads increased from 0.52% to 1.51% (p less than 0.001). Further analysis showed even more pronounced changes in these parameters for men living in the urban area compared those from the surrounding rural areas.

In vitro fertilization: problems in culture conditions and embryo transfer.

Review about the second and third step of in vitro fertilization and embryo transfer. The success rate is influenced by many factors of culture conditions (water, culture medium, plastics and glassware etc.) and by technical details of embryo transfer.

Scanning electron microscopy of human preimplantation endometrium in normal and clomiphene/human chorionic gonadotropin-stimulated cycles.

Embryo transfer after in vitro fertilization of oocytes obtained from unstimulated or stimulated women is usually carried out earlier than when an embryo developed in vivo reaches the womb. The possible asynchrony between the developmental stage of the embryo and that of the endometrium at transfer might reduce the prospects for implantation; so also might any secondary effect of stimulation. Preimplantation endometrium dated by plasma hormone analyses of 17 beta-estradiol and progesterone in five unstimulated and eight stimulated women was surveyed. Three features were examined, namely, the frequency, distribution, and appearance of ciliated cells, nonciliated cells, and apical protrusion. Wide regional variations were observed, but no continuous or consistent changes or differences were found from ovulation to the time for implantation in unstimulated and stimulated cycles. Therefore, from the morphologic point of view, the surface might be ready to accept a transferred embryo at any time within a range of days after ovulation.

Variation of semen parameters in fertile men.

The spermiograms of 63 fertile men (whose female partners were pregnant at the time of examination) were evaluated. A wide Gaussian distribution of the different semen parameters was found. Both parametric and nonparametric statistical tests were applied and gave almost identical results. In a subgroup of 34 men, a second semen sample was analysed, but a significant difference (p less than or equal to 0.05) between the two samples was found only regarding the proportion of abnormal forms. Between various semen parameters significant correlations were found. There was no association between follicle stimulating hormone (FSH) and sperm density with sperm counts greater than 20 mill/ml.