The Effects of Hormones and Vaginal Microflora on the Glycome of the Female Genital Tract: Cervical-Vaginal Fluid.

In this study, we characterized the glycome of cervical-vaginal fluid, collected with a Catamenial cup. We quantified: glycosidase levels; sialic acid and high mannose specific lectin binding; mucins, MUC1, MUC4, MUC5AC, MUC7; and albumin in the samples collected. These data were analyzed in the context of hormonal status (day of menstrual cycle, hormonal contraception use) and role, if any, of the type of the vaginal microflora present. When the Nugent score was used to stratify the subjects by microflora as normal, intermediate, or bacterial vaginosis, several important differences were observed. The activities of four of six glycosidases in the samples from women with bacterial vaginosis were significantly increased when compared to normal or intermediate women: sialidase, P = <0.001; α-galactosidase, P = 0.006; ß-galactosidase, P = 0.005; α-glucosidase, P = 0.056. Sialic acid binding sites as measured by two lectins, Maackia amurensis and Sambucus nigra binding, were significantly lower in women with BV compared to women with normal and intermediate scores (P = <0.0001 and 0.008 respectively). High mannose binding sites, a measure of innate immunity were also significantly lower in women with BV (P = <0.001). Additionally, we observed significant increases in MUC1, MUC4, MUC5AC, and MUC7 concentrations in women with BV (P = <0.001, 0.001, <0.001, 0.02 respectively). Among normal women we found that the membrane bound mucin MUC4 and the secreted MUC5AC were decreased in postmenopausal women (P = 0.02 and 0.07 respectively), while MUC7 (secreted) was decreased in women using levonorgestrel-containing IUDs (P = 0.02). The number of sialic acid binding sites was lower in the postmenopausal group (P = 0.04), but the number of high mannose binding sites, measured with Griffithsin, was not significantly different among the 6 hormonal groups. The glycosidase levels in the cervical-vaginal mucus were rather low in the groups, with exception of α-glucosidase activity that was much lower in the postmenopausal group (P<0.001). These studies present compelling evidence that the vaginal ecosystem responds to the presence of different vaginal microorganisms. These effects were so influential that it required us to remove subjects with BV for data interpretation of the impact of hormones. We also suggest that certain changes occurring in vaginal/cervical proteins are due to bacteria or their products. Therefore, the quantitation of vaginal mucins and lectin binding offers a new method to monitor bacteria-host interactions in the female reproductive tract. The data suggest that some of the changes in these components are the result of host processing, such as the increases in mucin content, while the microflora is responsible for the increases in glycosidases and the decreases in lectin binding. The methods should be considered a valid marker for insult to the female genital tract.

Role of Indoleamine 2,3-Dioxygenase and Tryptophan 2,3-Dioxygenase in Patients with Recurrent Miscarriage.

PROBLEM: An indoleamine 2,3-dioxygenase (IDO) and a tryptophan 2,3-dioxygenase (TDO) lead to dysfunction of T cell and immunological tolerance between fetus and mother in early pregnancy. We investigated the role of IDO and TDO in patients with recurrent miscarriage. METHODS OF STUDY: Cervical mucus, decidua, and villi were surgically collected from patients with recurrent miscarriage from April 2010 to March 2013. Samples of cervical mucus were divided into two groups: the delivery group and the miscarriage group. The samples of cervical mucus in the miscarried group and tissue of villi and decidua were divided into normal chromosome group (normal chromosome analysis of villi) and abnormal chromosome group (abnormal chromosome analysis of villi). We performed immunohistochemistry, SDS-PAGE, and Western Blot analysis and measured the activity of IDO and TDO. RESULTS: The activity of IDO and TDO in cervical mucus was not significantly different between the delivery group and the miscarriage group, and between the normal chromosome group and abnormal chromosome group. The expression of TDO in villi and decidua was not significantly different between the normal chromosome group and the abnormal chromosome group. The activity of IDO and TDO in villi and decidua was not significantly different between the normal chromosome group and the abnormal chromosome group. The expression of IDO in villi was significantly higher in the normal chromosome group than in the abnormal chromosome group. CONCLUSION: Our results suggest that the difference of expression of IDO and dysfunctional activation of IDO in villi may play an important role in unexplained recurrent miscarriage.

Matrix metalloproteinases in the cervical mucus plug in relation to gestational age, plug compartment, and preterm labor.

BACKGROUND: High concentrations of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) have been identified in the cervical mucus plug (CMP) at term of pregnancy. Their physiological and pathophysiological implications, however, remain to be elucidated, and CMPs from preterm labor have never been examined. This study was therefore conducted to describe the concentrations of MMP-2, TIMP-1, MMP-8 and MMP-9 in the CMP in relation to gestational age, IL-8 as an indicator of inflammation, compartment of the CMP, and preterm labor. METHODS: An aliquot of the distal plug compartment facing the vaginal microflora (CMP-dist) was collected from non-pregnant (n = 15), early pregnant (n = 15) and term pregnant women (n = 15). Whole CMPs shed during active vaginal term (n = 15) and preterm (n = 4) labor were also included. Protein concentrations were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: MMP-2 was not detectable in the non-pregnant CMP-dists whereas high concentrations were found in early pregnancy followed by an 85% decline at term. High concentrations of TIMP-1 were found in both the non-pregnant and early pregnant CMP-dists with a 90% decline at term. Consequently, the molar TIMP/MMP ratio was 40 in the non-pregnant state and 0.2 at term. The MMP-2 and TIMP-1 concentrations were alike in the CMP-dists and the whole CMPs.MMP-8, MMP-9, and IL-8 were mainly found in the distal CMP compartment. MMP-8 and MMP-9 concentrations were several fold increased in this compartment during pregnancy compared to the non-pregnant state. In the preterm whole CMPs, MMP-8, MMP-9 and IL-8 were 2 to 5 fold increased compared to term whole CMPs. CONCLUSIONS: These results suggest that CMP MMP-2 reflects the non-leukocyte dependent cervical remodeling that occurs in early pregnancy, whereas MMP-8 and MMP-9 are involved in the defense against ascending infections primarily located to the distal compartment of the CMP. The upregulation of MMP-8, MMP-9 and IL-8 in whole CMPs from preterm labor may indicate the involvement of an intrauterine infection.

Increased expression of alpha-enolase in cervico-vaginal fluid during labour.

OBJECTIVES: The aim of this study was (i) to characterise differentially expressed proteins in cervico-vaginal fluid (CVF) at the time of preterm labour onset and (ii) to confirm these studies in human CVF samples taken from women before and during spontaneous labour. STUDY DESIGN: Preterm labour was induced in sheep (n = 5) via fetal dexamethasone infusion (1 mg/24 h). CVF samples were taken prior to dexamethasone infusion (0 h), 28 h after the start of dexamethasone infusion, and immediately prior to delivery. Two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) were used to identify differentially expressed proteins. For the human studies, paired CVF samples were taken 5-9 days before labour and during spontaneous labour onset (n = 7). RESULTS: There was a 4.2-fold increase in α-enolase protein expression in sheep CVF during labour. Likewise, α-enolase protein expression was significantly increased during spontaneous human labour at term. CONCLUSIONS: Alpha-enolase is known to be bound to neutrophils and interact in the immune response, and thus may play a role in inflammation associated with human labour.

Effect of urinary trypsin inhibitor on preterm labor with high granulocyte elastase concentration in cervical secretions.

AIMS: To explore whether intravaginal treatment with urinary trypsin inhibitor (UTI) prevents preterm delivery in patients in preterm labor with increased levels of granulocyte elastase in cervical secretions. METHODS: The subjects were patients in preterm labor with increased levels of granulocyte elastase in cervical secretions from 16 to 33 weeks gestation. Maternal and neonatal outcomes were compared between patients receiving UTI treatment (UTI group; n=33) and those not receiving UTI treatment (control group; n=40). RESULTS: In patients receiving UTI, the mean gestational age at delivery was greater than that in the control group (37.8 vs. 35.6 weeks, p=0.003), and the rates of premature delivery before 34 and 37 weeks gestation were lower (3% vs. 20%, p=0.028; and 18% vs. 47%, p=0.008, respectively). The percentage of neonates weighing more than 2,500 g was significantly higher in the UTI group, with no neonates weighing less than 1,500 g. The neonatal hospitalization rate was lower in the UTI group (9% vs. 42%, p=0.001). CONCLUSION: In patients in preterm labor with a high elastase concentration in cervical secretions, treatment with UTI reduced the risk of preterm delivery and improved neonatal outcomes.

Factors affecting matrix metalloproteinase-8 levels in the vaginal and cervical fluids in the first and second trimester of pregnancy.

BACKGROUND: Cervical ripening during pregnancy resembles an inflammatory process. Matrix metalloproteinases (MMPs), particularly MMP-8, have been linked to inflammatory processes. We studied the concentrations of, and factors associated with, MMP-8 in the lower genital tract fluids in the first and second trimesters. METHODS: In a prospective population-based cohort study, vaginal and cervical swab samples were obtained from 2130 unselected pregnant women undergoing their first and second trimester ultrasound screening. MMP-8 was determined by immunofluorometric assay. Use of antibiotics, history of vaginal bleeding, and history of sexual intercourse were recorded on both occasions. Vaginal smears were obtained for Gram-staining and leukocyte counting. Cervical length was measured by ultrasonography. The main outcome measures were MMP-8 concentrations in the vagina and cervix. RESULTS: The median (range) MMP-8 concentrations in vaginal and cervical samples were 107.4 microg/l (undetectable-2406.6 microg/l) and 318.3 microg/l (0.1-2074.6 microg/l), respectively, in the first trimester, and 112.5 microg/l (undetectable-2093.4 microg/l) and 344.8 microg/l (0.4-1783.5 microg/l), respectively, in the second trimester. Multiparity and vaginal leukocytosis were both associated with increased MMP-8 concentrations in vaginal and cervical samples in both trimesters. Bacterial vaginosis (BV) was associated with increased vaginal and cervical MMP-8 in the first trimester, but only with increased vaginal MMP-8 in the second trimester. A history of sexual intercourse (in the previous 48 h) was associated with lower MMP-8 concentrations in cervical samples in both trimesters. CONCLUSIONS: MMP-8 concentrations were lower in vaginal samples than in cervical samples, and no difference was found between the first and second trimesters. Multiparity, BV and an elevated leukocyte count in the vagina were associated with increased MMP-8 concentrations. Sexual intercourse had an opposite effect. The study suggests that MMP-8 is a physiologic constituent in lower genital tract fluids, where it may be involved in host response to inflammatory and infectious processes.

Balance between matrix metalloproteinases (MMP) and tissue inhibitors of metalloproteinases (TIMP) in the cervical mucus plug estimated by determination of free non-complexed TIMP.

BACKGROUND: The cervical mucus plug (CMP) is a semi-solid structure with antibacterial properties positioned in the cervical canal during pregnancy. The CMP contains high concentrations of matrix metalloproteinase 8 and 9 (MMP-8, MMP-9) and tissue inhibitor of metalloproteinase 1 (TIMP-1). This indicates a potential to degrade extracellular matrix components depending on the balance between free non-complexed inhibitors and active enzymes. METHODS: Thirty-two CMPs collected during active labor at term were analyzed. Twelve CMPs were separated into a cellular and an extracellular/fluid phase and analyzed by gelatin and reverse zymography to reveal MMP and TIMP location. Twenty samples were homogenized, extracted and studied by the TIMP activity assay based on gelatin zymography. Enzyme-linked immunosorbent assay (ELISA) was used to determine TIMP-1, MMP-8 and MMP-9 protein concentrations, and gelatin and reverse zymography used to identify gelatinases and TIMPs, respectively. The Western blotting technique was applied for semi-quantification of alpha2-macroglobulin. An ELISA activity assay was used to detect MMP-8 and MMP-9 activity. RESULTS: ProMMP-2, proMMP-9, TIMP-1 and TIMP-2 were almost exclusively located in the fluid phase compared to the cellular phase of the CMP. All the extracted samples contained MMP-8, MMP-9, TIMP-1, TIMP-2 and alpha2-macroglobulin. Free non-complexed TIMP was detected in all the samples analyzed by the TIMP activity assay and was associated with TIMP-1 protein (R = 0.71, p < 0.001) and with the TIMP/MMP molar ratio (1.7 (1.1-2.5) (mean (95% confidence interval)) (R = 0.65, p = 0.002). The ELISA activity assay showed no activity from MMP-8 or MMP-9. CONCLUSION: Due to their extracellular location, potential proteolytic activity from neutrophil-derived MMPs in the CMP could exert a biological impact on cervical dilatation and fetal membrane rupture at term. The functional TIMP activity assay, revealing excess non-complexed TIMP, and a molar inhibitor/enzyme ratio above unity, indicate that refined MMP control prevents CMP-originated proteolytic activity in the surrounding tissue.

Role of tissue kallikrein-related peptidases in cervical mucus remodeling and host defense.

Human tissue kallikrein-related peptidases (KLKs) are 15 hormonally regulated genes on chromosome 19q13.4 encoding secreted serine proteases. Many KLKs are expressed throughout the female reproductive system and found in cervico-vaginal fluid (CVF). Immunohistochemistry was performed to determine KLK localization in the female reproductive system (fallopian tube, endometrium, cervix and vagina tissues). KLK levels were measured in CVF and saliva over the menstrual cycle to study whether KLKs are regulated by hormonal changes during the cycle. In vitro cleavage analysis was performed to establish whether KLKs may play a role in vaginal epithelial desquamation, mucus remodeling or processing of antimicrobial proteins. KLKs were localized in the glandular epithelium of the fallopian tubes and endometrium, the cervical mucus-secreting epithelium and vaginal stratified squamous epithelium. KLK levels peaked in CVF and saliva after ovulation. In vitro cleavage analysis confirmed KLKs 5 and 12 as capable of digesting desmoglein and desmocollin adhesion proteins and cervical mucin proteins 4 and 5B. KLK5 can digest defensin-1alpha, suggesting it may aid in cervico-vaginal host defense. We provide evidence of potential physiological roles for KLKs in cervico-vaginal physiology: in desquamation of vaginal epithelial cells, remodeling of cervical mucus and processing of antimicrobial proteins.

Menstrual cycle-related sialidase activity of the female cervical mucus is associated with exosome-like vesicles.

OBJECTIVE: To study endogenous sialidase activity in genital tract secretions of pregnant and nonpregnant women. DESIGN: Laboratory study. SETTING: Department of Evolutionary Biology and Department of Obstetrics and Reproductive Medicine, University of Siena, Siena, Italy. INTERVENTION(S): Vaginal and cervical mucus samples were obtained from pregnant and nonpregnant women in different phases of the menstrual cycle and in different weeks of pregnancy. MAIN OUTCOME MEASURE(S): Sialidase activity was assessed by fluorimetric assay and localized by transmission electron microscopy and differential centrifugation. RESULT(S): Sialidase activity in cervical mucus of healthy women reaches a maximum in the ovulatory phase. Cervical mucus from pregnant and nonpregnant women had significant sialidase activity that was associated with membranous vesicles having an exosome-like structure. CONCLUSION(S): Female cervical mucus contains an endogenous menstrual cycle-related sialidase that could be involved in modifying the rheologic properties of mucus to favor sperm progression at fertilization. Its association with exosome-like vesicles also suggests a role in intercellular communication before and after fertilization.

Increased risk of intrauterine transmission of HIV-1 associated with granulocyte elastase in endocervical mucus.

BACKGROUND: One of the remaining challenges in the prevention of mother-to-child transmission of HIV is to reduce the risk of the transmission during pregnancy. It remains to be investigated which factors affect intrauterine HIV transmission and how they can be identified and addressed during pregnancy. METHODS: Granulocyte elastase in the endocervical mucus of HIV-positive pregnant women in Zambia was measured, and its association with intrauterine transmission of HIV-1 from the mother to the fetus was investigated. RESULTS: The intrauterine transmission rate determined by polymerase chain reaction assay of DNA from neonates at birth was 15.3%. The risk for intrauterine transmission was 8.65-fold higher in women who were positive for granulocyte elastase than in those who were negative. CONCLUSION: We suggest that the women showing positive granulocyte elastase at delivery be strongly suspected of having and if having had chorioamnionitis during pregnancy, which could affect the intrauterine transmission of HIV.

Cervical inflammatory cytokines and other markers in the cervical mucus of pregnant women with lower genital tract infection.

OBJECTIVE: To evaluate concentrations of interleukin (IL)-6, IL-8, lactoferrin (LF), and alpha defencine (alpha-DF) in the cervical mucus of pregnant women and analyze their relation to cervicitis and bacterial vaginosis (BV). METHODS: Cervical mucus samples were obtained from August 2003 through May 2004 from 157 women who were between the 6th and 36th week of an uncomplicated singleton pregnancy. All women were delivered at term, 69 without BV or cervicitis, 9 with BV, and 79 with cervicitis. RESULTS: Interleukin 8, LF, and mu-DF concentrations were higher in women with cervicitis (0.81 +/- 0.36 pg/mL, 14.8+/-12.3 microg/mL, and 0.60 +/- 0.49 microg/mL) than in women without BV or cervicitis (0.35 +/- 0.34 pg/mL, 8.0 +/- 11.0 microg/mL, and 0.15 +/- 0.12 microg/mL). Interleukin 6 concentration was higher in women with BV (0.26 +/- 0.32 pg/mL) than in women without BV or cervicitis (0.09 +/- 0.15 pg/mL) or in women with cervicitis (0.12 +/- 0.18 pg/mL). CONCLUSIONS: Higher levels of inflammatory cytokines in the cervical mucus of pregnant women may lead to early detection of lower genital tract infection.

Increased level of granulocyte elastase in cervical secretion is an independent predictive factor for preterm delivery.

OBJECTIVE: The objective of this study was to explore whether increased levels of granulocyte elastase in cervical secretion is an independent predictive factor for preterm delivery before 34 weeks of gestation in the patient with preterm labor. METHODS: One hundred and sixty-one women with preterm labor at 22-28 weeks of gestation were enrolled prospectively. The level of granulocyte elastase in cervical secretions was measured by immunoassay, vaginal secretions were collected for the microscopic evaluation of Gram-stained smears, and the uterine cervix was assessed by transvaginal ultrasonography. RESULTS: Nineteen of 161 patients (12%) delivered before 34 weeks of gestation. Granulocyte elastase assessment had a sensitivity, specificity, positive predictive value, and negative predictive value for preterm delivery of 53, 75, 22 and 92%, respectively. A positive elastase assessment was associated with a relative risk for preterm delivery of 2.9 (95% CI 1.3-6.6), whereas a positive bacterial vaginosis assessment and shorter cervical length less than 25 mm demonstrated a relative risk of 1.9 (95% CI 0.8-4.6) and 1.5 (95% CI 0.6-5.0), respectively. CONCLUSION: The present study demonstrates that the risk of spontaneous preterm delivery before 34 weeks of gestation is increased in the women with preterm labor who are found to have an increased level of granulocyte elastase in cervical secretions.

Enzyme activity in bovine cervical mucus during spontaneous and induced estrus.

The purpose of the present research was to compare the enzyme activity of alkaline phosphatase (ALP), lactate dehydrogenase (LDH), alpha-amylase, alpha-manosidase, beta-N-acetyloglucosaminidase, beta-glucuronidase, and beta-galactosidase in the cervical mucus of cows during spontaneous and induced estrus. Friesian cows (n = 106) were assigned to 4 groups: 1) no treatment; 2) progesterone releasing intervaginal device (PRID) for 12 days plus pregnant mare serum gonadotrophin (PMSG) at the removal of the PRID; 3) PGF2alpha 2 doses 11 days apart; and 4) PRID for 7 days plus PGF2alpha 1 dose, 24 hours before removal of the PRID. Fourteen cows were excluded from the trial because of an inadequate quantity of cervical mucus collected or a lost PRID. The cows from the 3 induced estrus groups were artificially inseminated (AI) twice, while those with spontaneous estrus received only a single AI. Cervical mucus samples were collected from all cows 5 to 30 min before the first AI. The results are summarized as follows: 1) ALP and alpha-amylase activity for spontaneous estrus were similar to those for induced estrus; 2) LDH activity levels during spontaneous estrus were significantly lower (P < 0.001) than that in the P4 and P4+PGF2alpha induced estrus groups; and 3) glycosidases' activity was significantly lower (P < 0.001) in the spontaneous estrus group than that in the induced estrous groups. In conclusion, the activity of most enzymes in the cervical mucus of cows, in the present study, was significantly different between the spontaneous and the induced estrus groups.

Application of a novel human cervical mucin-based assay demonstrates the absence of increased mucinase activity in bacterial vaginosis.

Enzymes produced in bacterial vaginosis (BV) have been proposed as possible mediators of pre-term birth. Most studies have concentrated on mid-trimester measurements of enzyme activity, and utilize synthetic substrates to measure enzyme activity, which may not accurately represent mucinase activity in vivo. We have developed a novel ELISA mucinase assay using biotinylated human cervical mucin as a substrate. The assay is rapid, sensitive and can be used to screen large numbers of samples. The new assay has been used to assess vaginal mucinase activities in 92 women <14 weeks gestational age with and without BV. No differences in mucinase activity were detected between normal and BV groups while significant elevation of sialidase and other glycosidases was confirmed as reported before. This study shows that significant mucinase activity is a normal event in the mucus barrier, but does not reflect changes identified for individual enzyme activities associated with BV.

[Pro- and antioxidant system activity in cervical mucosa]. / Aktywnosc ukladów pro- i antyoksydacyjnego w sluzie szyjkowym szyjki macicy.

The aim of the study was to evaluate the activity of the pro- and antioxidant systems in cervical mucus of healthy women and women with pathological of disorders cervical. After cervical mucus liquefaction, the activity of superoxide dismutase, catalase and xanthine oxidase was determined by means of chemiluminescence assays. The activity of the antioxidant system in cervical mucus of healthy women was higher as compared to the group with pathological cervical disorders. Moreover, in mucus samples obtained from women with dysplasia, the significantly heightened activity of xanthine oxidase (strong pro-oxidant) was observed. The antioxidant system present in cervical mucus may protect both the cells of the uterine cervix and the viable sperm.

Cervical intraepithelial neoplasia is associated with increased polyamine oxidase and diamine oxidase concentrations in cervical mucus.

OBJECTIVE: The aim of this study was to establish whether reactive oxygen species, generated during oxidation of amines, catalyzed by polyamine oxidase (PAO) and diamine oxidase (DAO) in cervical secretions may play a role in the etiology of cervical cancer. METHODS: Cervical mucus was obtained from women attending the gynecological outpatient department: 139 with and 154 without cytological evidence of cervical intraepithelial neoplasia were recruited. The mucus was freeze dried in liquid nitrogen, weighed, and later resuspended for assay of PAO and DAO concentrations using a chemiluminescence method. The two groups were compared by group sequential analysis using PEST3 software. RESULTS: Patients with a colposcopic diagnosis of a high-grade squamous intraepithelial lesion (SIL) had significantly higher enzyme activities than control cases (L(N)PAO 1.37 (0.37) versus 1.18 (0.35): Student t test: P < 0.001; L(N)DAO 1.37 (0.36) versus 1.15 (0.37): Student t test: P < 0.001). CONCLUSION: It is probable that this rise in enzyme activity precedes cytological changes and plays some part in the etiology of cervical cancer, as the cells that undergo premalignant change are normally squamous in origin, whereas mucus is a product of columnar epithelium. Higher enzyme activity in patients with SIL than in controls may be a reflection of higher risk of exposure to amine substrates in semen through multiple sexual partners.

Changes in molecular weight of hyaluronan and hyaluronidase activity in uterine cervical mucus in cervical ripening.

BACKGROUND: This study was undertaken to investigate changes in the forms of hyaluronan and hyaluronidase activity in cervical mucus during cervical ripening. METHODS: Uterine cervical mucus was obtained from 57 pregnant women (25 at preterm gestation, ten at term gestation, 11 within 1 week before labor, and 11 during the first stage of labor). We determined 1) concentration of hyaluronan, 2) hyaluronidase activity, and 3) molecular weight of hyaluronan in cervical mucus. Data are presented as mean and range. RESULTS: The hyaluronan concentration in mucus in the 1st stage of labor (1.58 microg/ml, 0.46-23.96) was significantly (p<0.05) higher than that in all other groups (preterm: 0.29, 0.10-0.88; term: 0.24, 0.11-0.80; within 1 week before labor: 0.30, 0.18-0.62). Hyaluronidase activity both within 1 week before labor group (3.03 min., 1.12-3.95) and in 1st stage of labor group (3.52, 0.43-5.15) was significantly (p<0.05) higher than that in preterm group (1.70, 0.00-5.47). The molecular weight of hyaluronan in cervical mucus in the 1st stage of labor (0.97x106, 0.86-1.41) was significantly (p<0.05) lower than in the preterm and term groups (preterm: 1.60, 1.21-2.20, term: 1.41, 1.21-2.20). There was a significant correlation between hyaluronidase activity and molecular weight of hyaluronan (p<0.05, r=-0.41, n=23). CONCLUSION: These findings suggest that either hyaluronidase or low-molecular weight hyaluronan could be one of the most important regulators in the process of cervical ripening.

Does chondroitin sulfate defend the human uterine cervix against ripening in threatened premature labor?

OBJECTIVE: This study was undertaken to investigate changes in chondroitin sulfate levels in the cervix and the physiologic role of chondroitin sulfate isomers in the process of cervical ripening. STUDY DESIGN: Uterine cervical mucus samples were obtained from 57 women (7 nonpregnant women, 19 at preterm gestation, 9 at term gestation, 16 during the first stage of term labor, and 6 with threatened premature labor). Chondroitin sulfate isomer (chondroitin 0-sulfate, 4-sulfate, and 6-sulfate) concentrations in cervical mucus were measured by high-performance liquid chromatography. The effect of exogenous chondroitin sulfate on hyaluronidase activity in cervical mucus was evaluated by gel permeation chromatography of fluorolabeled hyaluronic acid. RESULTS: Chondroitin sulfate concentrations in cervical mucus were increased significantly (P <. 05) in the threatened premature labor group compared with the preterm and term groups. However, the same finding was not observed in the first stage of term labor. Exogenous chondroitin sulfate inhibited hyaluronidase activity in mucus. CONCLUSION: Chondroitin sulfate may defend against cervical ripening in threatened premature labor.

The usefulness of screening for chlamydial trachomatis infection with cervical mucus leukocyte esterase.

OBJECTIVE: To evaluate the ability of leukocyte esterase dipstick test (LET) performed in cervical mucus to predict direct immunofluorescent (DIF)-verified Chlamydia trachomatis (Ct) cervical infection. METHOD: Eighty women with inflammatory cervico-vaginal smear were submitted to both an endocervical smear, to verify Ct cervicitis by DIF technique; and a cervical mucus sample to perform LET. Results of LET and DIF were matched. RESULTS: Among 30 Chlamydial positive subjects, cervical mucus esterase test was positive in 28 (93.3%); while among 50 Chlamydial negative controls, esterase test was negative in 43 subject (86%). The sensitivity, specificity, positive and negative predictive values for LET were 93.3%, 86%, 80% and 95% respectively. CONCLUSION: Cervical mucus LET is a simple, cost effective screening test, with promising accuracy, for Ct cervical infection among women with inflammatory cervico-vaginal smears.

Binding of a bovine oviductal fluid catalase to mammalian spermatozoa.

Hydrogen peroxide (H2O2) is a reactive oxygen species that at low concentration is toxic to sperm. H2O2 inhibits not only sperm viability but also the acrosome reaction, sperm-egg binding, and oocyte penetration. Catalase activates the decomposition of H2O2 into water and oxygen, thus removing an initiator of free radical chain reactions leading to lipid peroxidation. Since the oviduct is known to enhance sperm survival, we hypothesized that it might secrete catalase. We found that oviductal fluid, harvested from washed cells collected at the slaughterhouse, possessed catalase-specific activity that varied during the estrous cycle. Catalase activity increased during the cycle and reached its maximal level just before ovulation (Days 18-20). No significant difference in activity was seen between fluid from the isthmus and that from the ampulla. Indirect immuno-staining of spermatozoa incubated in the oviductal fluid revealed the association of catalase in the region of the acrosomal cap. Addition of a commercial antibody directed against bovine liver catalase completely inhibited catalase activities from the oviductal fluid. Catalase activity was also detected in porcine oviductal fluid, human oviductal fluid, and cervical mucus. Western blots of oviductal fluid probed with the anti-catalase antibody revealed two major bands at 60 and 40 kDa. An immunoaffinity column was used to purify oviductal catalase, showing a unique band at about 60 kDa when analyzed by SDS-PAGE. The purified protein was incubated with bovine, boar, and human sperm, and Western blots of these sperm after several washes detected a band at 60 kDa, indicating that the protein was bound to sperm membranes. However, bovine liver catalase did not bind to sperm. Since H2O2 is one of the key reactants in the chain reaction of free radical production, this enzyme may play an important role in sperm survival within the female tract.