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A new method for evaluating cervical penetrability using daily aspirated and stored cervical mucus.

Cervical mucus samples from 33 women undergoing fertility work-up and married to normospermic husbands were aspirated daily with a tuberculin syringe during the periovulatory period. Samples were transferred to polyethylene tubing, sealed, and stored at 4 degrees C for 7 to 14 days. In vitro sperm penetration tests were performed in each case, using only one sample of husband's semen with all of the mucus samples that were collected during one cycle. Preliminary studies with 50 samples of cervical mucus taken at random showed that no difference in physicochemical and penetrability properties existed between fresh mucus and mucus stored for 10 to 14 days. Very often the optimal penetrability of the cervical mucus occurred on unpredictable days. In contrast, many tests performed on days which would have been predetermined for in vivo postocital tests were poor; consequently, those in vivo tests would have been falsely interpreted as negative. This new test using daily aspirated and stored mucus, which is simple to perform, is valuable mainly because it may eliminate the false negative results of other penetration tests. In this way the diagnosis of a truly hostile cervix may be more accurate.

Cervix Mucus

Immunoglobulins, proteinase inhibitors, albumin, and lysozyme in human cervical mucus. I. Communication: hormonal profiles and cervical mucus changes--methods and results.

The serum levels of luteinizing hormone (LH), estradiol-17beta, and progesterone were determined simultaneously with the concentrations of immunoglobulin (Ig) G, IgA, C'3, alpha1-antitrypsin, inter-alpha-trypsin inhibitor, alpha1x-antichymotrypsin, albumin, and lysozyme in cervical mucus during nine ovulatory cycles. Spinnbarkeit and ferning were also assessed, and the basal body temperature was measured and recorded during these cycles. The profiles were synchronized according to the LH peak. The midcycle period, characterized by the rapid increase and decline of estrogen and the beginning rise of progesterone, shows a prounced minimum of immunoglobulins, C'3, proteinase inhibitors, albumin, and lysozyme in cervical mucus, which is known to be most receptive to sperm penetration at this time. Although the variation of cervical mucus values is considerable during the early proliferative and the luteal phases, the midcycle values appear to be constantly low, showing slight differences among the profiles of the different parameters. The statistical evaluation and the assessment of the significance of parameters for ovulation detection and the assessment of the fertile period as well as the correlation of these parameters with basal body temperature will be the subject of the second communication of this series.

Adult

Immunoglobulin concentrations in cervical mucus in patients with normal and abnormal cervical cytology.

The cervical mucus of 31 patients with normal and 16 patients with abnormal cervical cytology was investigated at each stage of the menstrual cycle for immunoglobulin IgG, IgA and IgM. IgG and IgA were present in every mucus sample, while IgM was only occasionally found in trace amounts. IgG and IgA increased towards the last week of the menstrual cycle, the increase being in general more marked for IgA. Patients with abnormal cervical cytology showed increased IgG and, more strikingly, IgA concentrations in their cervical mucus, but there was no correlation between the IgG and IgA concentrations at any stage of the menstrual cycle. Whereas in patients with normal cervical cytology the IgG and IgA concentrations correlated throughout the menstrual cycle.

Adolescent

Antibacterial activity of human cervical mucus.

The antimicrobial property of human uterine cervical mucus was tested in three groups of women. Healthy women, using no contraception, women using an intrauterine device and women receiving hormonal treatment for contraception. Cervical mucus was taken on the 10th, 14th, 18th and 22nd day of the menstrual cycle. Cervical mucus had a strong inhibitory effect on the growth of Micrococcus lysodeicticus in all three groups. The strength of the inhibitory effect on the other microorganisms were in the following order: Staphylococcus albus, Staphylococcus aureus, Proteus mirabilis, Escherichia coli, Candida albicans, Streptococcus haemolyticus, Streptococcus faecalis. Use of an intrauterine device did not affect the antimicrobial effect of cervical mucus. The use of hormonal contraceptive canceled the antimicrobial effect on the series of microorganisms, with the exception of M. lysodeicticus. The maximum inhibitory effect occurred on the 14th day and declined toward the end of the menstrual cycle.

Adult

Significance of timing for the postcoital evaluation of cervical mucus.

This investigation was designed to determine the role of cervical mucus in the dynamic in vivo distirbution of transported spermatozia in the human reproductive tract. Forty-two normal subjects had midcycle cervical mucus collection at various times after insemination. Sperm were identified in the cervical mucus at the level of the internal as shortly after insemination. The numbers of sperm in cervical mucus were directly related to the numbers of sperm in the inseminate and were related to time from insemenation. In order to obtain optimal information, the postcoital examination for the presence of sperm should be performed within two and one-half hours after semen exposure. It was also determined that cervical mucus does serve as a reservoir for those sperm which are transported to the Fallopian tubes.

Adult

The role of acrosin in sperm penetration through human cervical mucus.

Enzymes have been implicated in facilitating cervical mucus penetration by spermatozoa. One of these enzymes in the neutral proteinase acrosin, which is associated with the sperm acrosome. To determine the validity of this hypothesis, human spermatozoa were incubated with the following acrosin inhibitors: p-aminobenzamidine (AB), N-alpha-p-tosyl-L-lysine chloromethyl ketone (TLCK), and p-nitropheyl-p'-guanidino benzoate (NPGB). An in vitro slide test system was developed which allowed inhibitor-treated and control spermatozoa to be evaluated against the same human cervical mucus sample. At inhibitor concentrations far exceeding those necessary for the inhibition of human acrosin, there was no effect on spermatozoal penetration into or through the mucus. These findings indicate that, in man, acrosin activity is neither necessary nor facilitory to sperm penetration of cervical mucus. Evidence is also presented that demonstrates the superiority of the newly developed double-interface slide test, especially for comparative purposes, over the tests currently in use.

Acrosin

Crossed immunoelectrophoresis of sperm antibodies in human serum and cervical mucus.

Sperm agglutinating antibodies are purified from sera and cervical mucus of women with unexplained causes of infertility which were positive in the FD-test. Fractionation was performed by affinity-chromatography in a batch device and the sperm agglutinating activity controlled by the Franklin and Dukes test. This sperm antibody fraction was determined via crossed immunoelectrophoresis by migration into an anti-human serum containing gel. In all cases only one big peak resulted. The negative control serum and mucus samples demonstrated no precipitation peaks. By absorption studies it was shown that the sperm agglutinating antibodies in sera were IgM and in cervical mucus IgA. The concentration of IgA and IgM was determined by comparison with standard human IgA and IgM. Thus only one serum- and one cervical mucus antibody seems to be responsible for agglutination. The number of experiments, however, is still too small for general conclusions. This method is easily and quickly performed and can therefore be used as a routine method for the determination of sperm agglutinating antibodies. Its application for sperm-immobilizing or cytotoxic activity remains to be tested.

Antibodies

Ureaplasma urealyticum (T-mycoplasma) in vaginal fluid and cervical mucus from fertile and infertile women.

Ureaplasma urealyticum (T-mycoplasma) was isolated more frequently and in heavier growth from cervical mucus (49%) than from vaginal fluid (34%). It was isolated in 24% of vaginal fluid samples and in 35% of cervical mucus samples from fertile women, and in 29% of vaginal fluid samples and in 47% of cervical mucus samples from infertile women. The incidence of infection was high following abortion or total hysterectomy and during pregnancy or oral contraceptive use. T-mycoplasma was also isolated from the vaginal fluid and cervical mucus of a woman with tubo-ovarian abscess, but was not present in women with Trichomonas vaginalis infection. U. urealyticum did not alter the physiophysiologic characteristics of vaginal fluid and cervical mucus or the sperm penetration and sperm viability in cervical mucus. Treatment with tetracycline eradicated the organism in 88% of the infected women. Pregnancies were recorded during a 6-month follow-up in 1 of 19 infertile women who were treated with tetracycline.

Cell Survival

The sperm-cervical mucus contact test: a preliminary report.

Infertility in men and women with spermagglutinins is the result of disturbed penetration and migration of spermatozoa in the cervical mucus. In ejaculates with partial spermagglutination caused by autoimmunization, the progressive propulsion of the sperm was changed into stationary, shaking movement the moment the sperm came into contact with cervical mucus. The same alteration in spermatozoal motility pattern also occurred when spermatozoa from a normal, fertile ejaculate came into contact with cervical mucus of a woman whose serum contained sperm antibodies. This shaking phenomenon was visualized in a simple test, the sperm-cervical mucus contact test. We demonstrated that sensitized spermatozoa exhibit the shaking phenomenon after contact with the glycoprotein fraction of the cervical mucus and not after contact with the aqueous fraction. Therefore, the hypothesis is introduced that the shaking phenomenon is due to an interaction between sensitized spermatozoa and the glycoprotein micelles in cervical mucus.

Autoantibodies

Presence of prolactin in human cervical mucus.

The presence of prolactin in human cervical mucus was demonstrated by radioimmunoassay. The levels of prolactin in cervical mucus were significantly higher (260 ng/ml +/- 47.5 SE) than those in sera of normally menstruating women (30.28 ng/ml +/- 2.0 SE).

Cervix Mucus

Human cervical mucus. I. Rheologic characteristics.

The viscoelastic properties of human cervical mucus were subjected to detailed characterization by microrheometry. An introduced to the range of parameters that can be measured and the sensitivity of microroheometry to variations in mucus viscoelasticity was presented, along with an interpretation of results obtained by this approach. Fresh cervical mucus samples were shown to be rheologically heterogeneous, reflecting compositional differences in situ. Mucas viscoelasticity was stable for several hours when samples were stored at ambient temperature in the microrheometer sample holder. Mucus viscoelasticity was strongly dependent upon nondialyzable solids (NDS) concentration. In order to eliminate (NDS) as a variable in comparative studies of mucus, procedures were developed for reconstitution of mucus at identical (NDS) or for normalization of viscoelasticity data derived from fresh samples to 2.5% NDS. The validity of this approach was then examined by applying these procedures to mucus collected during the follicular, ovulatory, and luteal phases of the normal menstrual cycle.

Animals

In vitro action of progestogens on sperm migration in human cervical mucus;.

The presence of progestogens in the cervical mucus suppresses and arrests sperm penetration. Using the Kremer technique, the effects of in vitro released progesterone, d-norgestrel, and cyproterone acetate were studied by inserting silicone rubber threads bearing the respective compounds into capillary tubes containing cervical mucus. Control tubes were fitted with nonmedicated silicone rubber threads. After 24 hours of incubation, the sperm migration test was carried out to evaluate penetration depth, qualitative motility, and proportion of motile forms. Progesterone produced the greatest alterations. Migration was arrested within 30 minutes, the distance reached was consistently less than 2 cm, and sperm were completely immobile at 24 hours. D-norgestrel also exhibited a distinct spermiostatic effect, but it was not as intense as that of progesterone. Cyproterone acetate was practically effective during the first 120 minutes and produced alterations only in the qualitative and proportional motility.

Cervix Mucus

Molecular arrangement of cervical mucus: a reevaluation based on laser light-scattering spectroscopy.

Evidence obtained from laser light-scattering spectroscopy suggests that the molecular arrangement of cow estrous cervical mucus is an ensemble of entangled random-coiled macromolecules rather than a cross-linked macromolecular network, the model heretofore widely accepted. This new model can account for the following phenomena: (1) The viscoelastic properties of the estrous mucus and human mid-cycle cervical mucus are the result of the way in which the glycoproteins are entangled. Variations of these properties during the reproductive cycle may simply be due to changes in the water content of the cervical mucus. (2) Penetration of spermatozoa in mucus may be entirely mechanical without involving any kind of enzymatic lysis of cross-links (this is supported by changes in the pattern of spermatozoa flagellation during sperm penetration of cervical mucus as shown in motion picture films). (3) The orientation of spermatozoa in the cervical muscus need not be due to the presence of 'channels' between micelles but simply to an artifact, the stretching or flow of the mucus, which orients the entangled macromolecules and thus facilitates the penetration of spermatozoa in an axis parallel to them.

Animals

Ultrastructure and variations of human cervical mucus during pregnancy and the menopause.

Ultrastructural aspect and variations of cervical mucus have been studied by scanning electron microscopy in 58 pregnant and 29 menopausal women. In both groups, a striking tightening of the ultrastructural glycoprotein framework was demonstrated. The phenomenon appears to occur very rapidly at the beginning of pregnancy while patients near the menopause show more variable patterns. The results are compared to the known evolution of cervical mucus ultrastructure during the ovarian cycle and discussed from a rheological point of view. Particular attention was paid to the eventual antibacterial role of cervical mucus in pregnancy, which was found to be very plausible.

Adult

Oral contraceptives and human cervical mucus lipids.

Oral contraceptives containing estrogenic and progestational components caused a consistent increase in human cervical mucus lipids during the menstrual cycle. Cyclic variations in cervical mucus lipids occur during the normal cycle. Phospholipids, free cholesterol, and triglycerides in cervical mucus increase with the use or oral contraceptives.

Adult

Effect of human cervical mucus on human sperm motion and hyperactivation in vitro.

The aim of our experiment was to examine the effect of exposure to human cervical mucus on quantitative sperm motility with specific reference to hyperactivated sperm motility. Human spermatozoa were allowed to penetrate cervical mucus for 20 min before swimming into Earle's balanced salt solution tissue culture medium for 25 min. The sperm motion characteristics were compared to those which had been obtained from a direct swim-up for 45 min. Spermatozoa treated with mucus were more 'active' than the control group. Multivariate statistical analysis indicated that cervical mucus promotes hyperactivated motility and that sperm sub-populations exposed to cervical mucus are very heterogeneous, as indicated by the numbers and motility characteristics of spermatozoa.

Cervix Mucus

Cervical mucus changes in infertile ewes previously exposed to oestrogenic subterranean clover.

Characteristics of cervical mucus were compared between groups of ewes which had previously grazed for long periods on either potently oestrogenic clover pasture (group A), mildly oestrogenic pasture (group B) or non-oestrogenic pasture (group C). No phytooestrogens were fed at the time of the study. The chloride content of mucus from group A ewes was significantly decreased, on a dry weight basis. The motility and survival of ram spermatozoa appeared similar in cervical mucus from groups A and B. There was no statistically significant difference between groups of ewes in either the total wet weight of mucus, the proportion of dry weight, or the degree of arborisation of mucus, at the time of oestrus. Mucus from group A ewes had a significantly decreased spinbarkheit, and a decreased ability to orientate sperum migration, when compared with group B. The group B ewes had a significantly smaller spinbarkheit than group C. Measurement of cervical mucus spinbarkheit offers promise as a diagnostic test for permanent clover infertility.

Animal Feed

Cyanogen bromide fragments of bovine cervical mucus glycoprotein.

Treatment of bovine cervical mucus glycoprotein with cyanogen bromide gives four fractions, with the molecular weights of the three major fractions being 230 000, 130 000, and 35 000. The results indicate that, as for other glycoproteins, there are different regions along the protein core, some of which have a high sugar content and others which contain considerably less carbohydrate; it seems likely that the regions of lower sugar content may be important to intermolecular linkages. The data suggest that the basic unit of the glycoprotein has a molecular weight of 550 000-600 000, with its protein core consisting of approx. 1200 amino acid residues.

Amino Acids