Frederick W. Hanson

Acrosome intact and acrosome-reacted human sperm can initiate binding to the zona pellucida.

Mammalian sperm must be acrosome reacted before penetrating the zona pellucida. In some species the sperm undergo the acrosome reaction before binding to the zona pellucida and in other species only acrosome intact sperm can initiate binding to the zona. In this study we addressed the question of acrosomal status and sperm-zona binding with human gametes. Sperm acrosome reactions were induced by treatment with human follicular fluid or N-(6-amino-hexyl)-5-chloro-naphthalene sulfonamide (W-7). The sperm suspensions, containing various percentages of acrosome-reacted sperm, were then incubated with human oocytes for 1 min. The acrosomal status of the sperm population bound to the zona was similar to the acrosomal status of the population of sperm in suspension (R2 = 0.77), regardless of the treatment to induce acrosome reactions. Our interpretation of these results is that both acrosome intact and acrosome-reacted human sperm can initiate binding to the zona pellucida. However, we reported earlier (N. L. Cross, P. Morales, J. W. Overstreet, and F. W. Hanson, 1988, Biol. Reprod. 38, 235-244) that the human zona pellucida is able to induce acrosome reactions. Thus, to exclude the possibility that sperm had undergone the acrosome reaction on the zona within 1 min of binding, sperm were suspended in a nominally calcium-free Tyrodes medium (0 Ca-mTyr) before incubation with oocytes (this medium was supplemented with SrCl2 and spermine to support sperm motility and zona binding). In 0 Ca-mTyr, the proportion of acrosome-reacted sperm on the zona was still highly correlated with the proportion of reacted sperm in suspension, indicating that the sperm were reacted before binding. Evidence that 0 Ca-mTyr effectively inhibited acrosome reactions induced by the zona pellucida was derived from experiments in which sperm were treated with human follicular fluid or control medium and the suspensions were diluted with either 0 Ca-mTyr or control medium.4+ Human oocytes were added for 1 min (pulse) at which time some oocytes were fixed and other oocytes were transferred to sperm-free medium and incubated for 35 min (chase) before fixation. Sperm diluted in control medium, pretreated with either human follicular fluid or control medium, showed a similar increase (40%) in the percentage of acrosome reactions among the zona-bound sperm after the chase. Sperm diluted in 0 Ca-mTyr did not show an increase in the percentage of acrosome-reacted sperm on the zona pellucida after the chase.(ABSTRACT TRUNCATED AT 400 WORDS)

A New Quantitative Test for Sperm Penetration into Cervical Mucus

A new experimental and theoretical procedure is described for characterizing the penetration of spermatozoa into cervical mucus in vitro. Semen is introduced to a prescribed volume of mucus contained in a flat capillary tube. Use of the tube enables the surface area of semen-mucus contact to be fixed, and provides excellent visualization of sperm movement in the mucus. The time interval of semen-mucus contact is also controlled. The number of motile sperm and their swimming speeds are determined in both the semen and mucus by hemocytometer counts and simple time-exposure photomicrography. The assay provides two new measures of the sperm-mucus interaction. The number of successful sperm entries into the mucus is compared with the number of original collisions between seminal sperm and the semen-mucus interface. The comparison is expressed as a ratio of the numbers of sperm in these two groups, viz., PSC = percentage of successful collisions. The vitality of spermatozoa that do succeed in entering the mucus is assessed by comparing their swimming speeds with those of sperm in the semen. This second comparison is also expressed as a ratio, viz., VR = velocity ratio = mean swimming speed in mucus/mean swimming speed in semen. The clinical application of the method to human semen and cervical mucus is described, and sample results are presented.

The interaction of human spermatozoa with cervical mucus in vivo

The interaction between human spermatozoa and cervical mucus was studied during 14 cycles of artificial insemination (AI) with a cervical cup. The concentration of spermatozoa in the inseminate was determined as were the percentage of motility, mean swimming speed, and sperm morphology. The percentage of motility, swimming speed, and morphology of spermatozoa in the mucus were determined at 1 hour and 48 hours after AI. The percentage of motile sperm was always higher in the cervical mucus than in the semen. In some cases, spermatozoa appeared to swim faster in the mucus than in the semen but in other cases the reverse was true. In all 14 cases some spermatozoa could be found in the mucus at 48 hours after AI. In general, the percentage of motility and swimming speeds of the cervical sperm remained unchanged over the 48-hour study interval. The percentage of spermatozoa with normal morphology was higher in the cervical mucus than in the semen. The alteration in the composition of the sperm population appeared to result from exclusion by the mucus of most classes of abnormal sperm. The morphology of the cervical sperm population was similar at 1 hour and at 48 hours after AI.

Amniocentesis before 15 weeks' gestation: Outcome, risks, and technical problems☆

Between January 1, 1979, and May 30, 1986, 4750 amniocenteses were performed by a single physician at the University of California, Davis, Medical Center. Of these procedures 541 (11.4%) were considered early amniocenteses (performed before the fifteenth week since last menstrual period). The medical records were reviewed for maternal age, amniocentesis indication, ultrasound findings, location of the placenta, location of the needle insertion (transplacental versus nontransplacental), color of the amniotic fluid, results of prenatal testing, complications of procedure, and pregnancy outcome. Complete follow-up data were possible for 308 subjects and obtained for 298 (96.8%). There were 11 miscarriages (3.6%), two stillbirths (0.7%), and one neonatal death (0.3%), resulting in a total postprocedural loss rate of 4.7% (14/298). If those patients with a preamniocentesis history of bleeding are eliminated, the total postprocedural rate of loss is 3.3% (7/211). Miscarriage within 2 weeks of amniocentesis occurred in 1.7% of procedures (5/298). No significant difference in pregnancy outcome was noted between transplacental and nontransplacental amniocentesis.

The Importance of Seminal Plasma for Sperm Penetration of Human Cervical Mucus

Experiments were carried out in which semen samples were diluted 1:1 with Tyrodes solution or with their own seminal plasma (obtained by centrifuging another semen aliquot) as a control. Each experiment consisted of a paired comparison of these two sperm suspensions, using a quantitative cervical mucus penetration test with aliquots of the same mucus sample. Videomicrography was used to measure the swimming speeds of spermatozoa in the semen and in the mucus. The spermatozoa swam faster in Tyrodes diluted seminal plasma than in whole seminal plasma, but their swimming speeds in cervical mucus were similar after mucus penetration. Significantly more of the collisions between spermatozoa and the mucus resulted in successful penetration in tests where the sperm were suspended in whole seminal plasma than in tests where they were suspended in Tyrodes diluted seminal plasma. These observations indicate that components of the seminal plasma are important for efficient entry of human spermatozoa into cervical mucus.

Analysis of 2136 genetic amniocenteses: experience of a single physician.

Between 1979 and 1984, 2136 midtrimester genetic amniocenteses were performed by a single physician at the University of California (Davis) Medical Center. The medical records were reviewed for maternal age, amniocentesis indication, ultrasound findings, location of placenta, location of the needle insertion (transplacental versus nontransplacental), color of the amniotic fluid, results of prenatal testing, complications of procedure, and pregnancy outcome. Follow-up data were available for 88% of the pregnancies. There were 38 miscarriages (1.9%), 18 stillbirths (0.9%), and four neonatal deaths (0.2%), resulting in a total postprocedural loss rate of 3.1%. Miscarriage within 2 weeks of amniocentesis occurred in only 0.4% of procedures (seven of 1918). No significant difference in pregnancy outcome was noted between transplacental and nontransplacental amniocentesis. Greenish brown discolored amniotic fluid was found in 3.6% of procedures (76 of 2136). In this group there was a 14.5% fetal loss rate with an overall 22.4% adverse outcome.

A study of the relationship of motile sperm numbers in cervical mucus 48 hours after artificial insemination with subsequent fertility

The records of 159 women who underwent 561 cycles of donor insemination (AID) in the UCD artificial insemination program were analyzed in a study of the relationship of the numbers of motile sperm seen in the cervical mucus 48 hours after AID with subsequent fertility. The numbers of motile sperm were expressed on an integral rank scale (0 to 8). A relationship between the results and subsequent fertility was demonstrated. The distribution of the data on sperm numbers for fertile and infertile cycles were qualitatively and quantitatively different. The latter were consistently skewed toward low values, while the former were not. Overall, mean values were higher for conception cycles. Selective grouping of women to eliminate factors that could potentially have an adverse influence on fertility, such as pelvic abnormality, oligoovulation, or prior pelvic operation, increased the overall conception rate. However, they did not influence the relationship between low and high motile sperm numbers and subsequent conception.

Analysis of the significance of discolored amniotic fluid detected at midtr mester amniocentesis

In 110 midtrimester amniocenteses of the 3349 procedures done at the University of California, Davis, Medical Center between 1979 and 1984, the fluid obtained was noted to be discolored. The significance of this finding was evaluated with review of pregnancy history and outcome and by using biochemical techniques. A significant increase in miscarriage was seen in the pregnancies where discolored fluid was obtained (9%), in comparison with the entire group of pregnancies in which amniocentesis was done at the same institution over the same period (1.6%). This increase was most marked in the pregnancies in which, in addition to discolored fluid, there was a prior history of pregnancy bleeding. Thirty-four discolored fluid samples and 18 clear fluid samples were studied with spectrophotometry, electrophoresis, isoelectric focusing, and chromatography. These studies indicate that in most cases the discoloring pigment is blood rather than fetal bowel contents, as had been suggested by others.

Amniotic Membrane as a Living Surgical Dressing in Human Patients

Preliminary observations on the role of the amniotic membrane devoid of the chorion have been made. Clinical cases have been presented to illustrate that this membrane as an allograft may have a poten

The roles of ultrasonography and amniocentesis in evaluation of elevated maternal serum α-fetoprotein

A total of 681 pregnant women were referred for evaluation of elevated maternal serum alpha-fetoprotein levels. Ultrasonographic examination yielded an explanation for the elevation of maternal serum alpha-fetoprotein in 42% of patients. Diagnoses made by ultrasonography included incorrect fetal dating, multiple gestation, fetal death, open neural tube defect, abdominal wall defect, placental abnormalities, cystic hygroma, renal anomalies, and oligohydramnios. Optimal prenatal diagnosis of fetal anomalies also requires the use of amniocentesis in many patients. Amniocentesis may be obviated if fetal dating is incorrect, if an unsuspected multiple gestation is discovered, or if there is a clear anomaly and the parents do not desire genetic counseling based on karyotype information. If the fetus appears normal, the ultrasonographic results are equivocal, or the parents desire more detailed genetic counseling when an anomaly is found by ultrasonography, then amniocentesis should be performed. Thirteen abnormalities were diagnosed by amniocentesis alone in this group.