Impact of a hormone-releasing intrauterine system on the vaginal microbiome: a prospective baboon model.

BACKGROUND: Use of a levonorgestrel-releasing intrauterine system (LNG-IUS) in humans may alter vaginal microbial populations and susceptibility to pathogens. This study evaluated the time-dependent effects of an LNG-IUS on the vaginal microbiome of the baboon, a useful animal model for reproductive studies. METHODS: Levonorgestrel-releasing intrauterine systems were inserted into three reproductively mature, female baboons. The animals were evaluated for 6 months by physical examination and Gram-stained cytology. The vaginal microbiota was characterized at each timepoint by culture-independent analysis of the 16S rRNA-encoding gene. RESULTS: Each baboon harbored a diverse vaginal microbiome. Interindividual variation exceeded intra-individual variation. Diversity declined over time in one baboon and showed mild fluctuations in the other two. There were no significant community differences from early to late post-LNG-IUS placement. CONCLUSIONS: The baboon vaginal microbiome is unique to each individual and is polymicrobial. In this pilot study, the vaginal microbiome remained stable from early to late post-LNG-IUS placement.

Novel genital alphapapillomaviruses in baboons (Papio hamadryas anubis) with cervical dysplasia.

Genital Alphapapillomavirus (αPV) infections are one of the most common sexually transmitted human infections worldwide. Women infected with the highly oncogenic genital human papillomavirus (HPV) types 16 and 18 are at high risk for development of cervical cancer. Related oncogenic αPVs exist in rhesus and cynomolgus macaques. Here the authors identified 3 novel genital αPV types (PhPV1, PhPV2, PhPV3) by PCR in cervical samples from 6 of 15 (40%) wild-caught female Kenyan olive baboons (Papio hamadryas anubis). Eleven baboons had koilocytes in the cervix and vagina. Three baboons had dysplastic proliferative changes consistent with cervical squamous intraepithelial neoplasia (CIN). In 2 baboons with PCR-confirmed PhPV1, 1 had moderate (CIN2, n = 1) and 1 had low-grade (CIN1, n = 1) dysplasia. In 2 baboons with PCR-confirmed PhPV2, 1 had low-grade (CIN1, n = 1) dysplasia and the other had only koilocytes. Two baboons with PCR-confirmed PhPV3 had koilocytes only. PhPV1 and PhPV2 were closely related to oncogenic macaque and human αPVs. These findings suggest that αPV-infected baboons may be useful animal models for the pathogenesis, treatment, and prophylaxis of genital αPV neoplasia. Additionally, this discovery suggests that genital αPVs with oncogenic potential may infect a wider spectrum of non-human primate species than previously thought.

Formulation development of retrocyclin 1 analog RC-101 as an anti-HIV vaginal microbicide product.

RC-101 is a synthetic microbicide analog of retrocyclin, which has shown in vitro activity against X4 and R5 HIV-1. In an effort to develop a safe and effective RC-101 vaginal microbicide product, we assessed safety in ex vivo macaque and human models and efficacy using in vitro and ex vivo models. A polyvinyl-alcohol vaginal film containing RC-101 (100 µg/film) was developed. Formulation assessment was conducted by evaluating disintegration, drug content, mechanical properties, and stability. Efficacy was evaluated by in vitro peripheral blood mononuclear cells (PBMC) assay and ex vivo human ectocervical tissue explant model. Ex vivo safety studies were conducted by exposing RC-101 to an excised monkey reproductive tract and excised human ectocervical tissue. RC-101 100 µg films were shown to be safe to human and monkey tissue and effective against HIV-1 in vitro and ex vivo in human ectocervical tissue. The 90% inhibitory concentration (IC90) for RC-101 films at 2,000 µg (IC90=57.5 µM) using an ex vivo model was 10-fold higher than the IC90 observed using an in vitro model (IC90=5.0 µM). RC-101 films were stable for 1 month at 25°C, with in vitro bioactivity maintained for up to 6 months. RC-101 was developed in a quick-dissolve film formulation that was shown to be safe in an ex vivo model and effective in in vitro and ex vivo models. RC-101 film formulations were shown to maintain bioactivity for a period of 6 months. Findings from the present study contribute to the development of a safe and effective topical microbicide product.

Preclinical safety assessments of UC781 anti-human immunodeficiency virus topical microbicide formulations.

The nonnucleoside reverse transcriptase inhibitor UC781 is under development as a potential microbicide to prevent sexual transmission of human immunodeficiency virus type 1 (HIV-1). Two gel formulations of UC781 (0.1% and 1.0%) were evaluated in a range of preclinical safety assessments, including systemic absorption analysis following topical application in the pig-tailed macaque models for vaginally and rectally applied topical microbicides. High-sensitivity high-performance liquid chromatography analysis of serum samples showed that no systemic absorption of UC781 was detected after repeated vaginal or rectal application of either product. However, high levels of UC781 were detectable in the cervicovaginal lavage samples up to 6 h after product exposure. Both formulations were safe to the vaginal microenvironment, even with repeated daily use, as evidenced by colposcopy, cytokine analysis, and lack of impact on vaginal microflora. By contrast, rectal application of the 1.0% UC781 formulation caused an increased expression of numerous cytokines not observed after rectal application of the 0.1% UC781 formulation. These results provide additional support for the continued development of UC781 formulations as anti-HIV microbicides.

Preclinical safety and efficacy assessments of dendrimer-based (SPL7013) microbicide gel formulations in a nonhuman primate model.

Three gel formulations (1%, 3%, and 5% [wt/wt]) of SPL7013, a dendrimer known to have antiviral (anti-human immunodeficiency virus and anti-herpes simplex virus) activities, completed a range of preclinical tests in the pigtailed macaque models for vaginally and rectally applied topical microbicide safety assessments. The vaginal safety profile of the 3% SPL7013 gel formulation was equal to that of the 1% formulation but was superior to that of the 5% formulation. The 3% SPL7013 gel was further evaluated for rectal safety and for antichlamydial efficacy with cervical challenge with Chlamydia trachomatis. This first-generation dendrimer-based product was shown to be safe to the vaginal and rectal microenvironments with repeated daily use. However, a single intravaginal application of the 3% (wt/wt) SPL7013 gel did not provide protection from the acquisition of cervical chlamydial infection.

Safety and efficacy evaluations for vaginal and rectal use of BufferGel in the macaque model.

BACKGROUND: The nonhuman primate model allows for safety and efficacy testing of topical microbicide products. GOAL: The goal of this study was to evaluate the safety and efficacy of vaginal and rectal applications of BufferGel (ReProtect, Inc.). STUDY DESIGN: The safety of repeated product applications was evaluated by microflora, pH, vaginal colposcopy, and rectal lavage. To test efficacy in preventing chlamydia, infection was documented by culture and nucleic acid amplification tests. RESULTS: Repeated vaginal or rectal applications of BufferGel were not associated with significant changes in microflora. BufferGel use had a transient acidifying effect on vaginal and rectal pH. Colposcopic observations remained relatively normal in all test animals. A slightly increased incidence of epithelial desquamation was noted after rectal product use compared with the control group. BufferGel did not prevent cervical or rectal chlamydial infection. CONCLUSION: BufferGel has an acceptable safety profile after repeated vaginal and rectal use, but does not prevent chlamydial infection in the macaque models.

The pig-tailed macaque rectal model: microflora and chlamydial infection.

BACKGROUND: A topical microbicide should protect against acquisition of sexually transmitted infection during both vaginal and rectal intercourse. The rectal microflora of the Macaca nemestrina (pig-tailed macaque) and humans were examined, as well as the histopathology of rectal tissues. In a subset of macaques, a human rectal isolate of Chlamydia trachomatis was inoculated into the rectum to establish rectal chlamydial infection. GOAL: To evaluate the comparability of the pig-tailed macaque rectal model with humans. STUDY DESIGN: Rectal swabs were collected for microbiologic analysis to characterize normal microflora in pig-tailed macaques and humans. Subsequently, 10 macaques received a rectal inoculation with C trachomatis, serovar D, prepared from a clinical rectal isolate. RESULTS: The rectal microflora of pig-tailed macaques (n = 80) were found to be comparable with the rectal flora of humans (n = 40). The prevalence of Lactobacillus in the rectum was higher in the macaques than in humans. Coliform and Enterococcus were decreased in the macaques, as compared with those of humans. In 9 of 10 macaques, rectal chlamydial infection was confirmed by culture or ligase chain reaction on days 2, 7, and 14 after inoculation. The test results were positive for rectal chlamydial infection by ligase chain reaction only for the remaining animal on day 14 after inoculation. CONCLUSIONS: The findings demonstrate that the rectal environment of the pig-tailed macaque is a useful model for further evaluation of newly developed topical microbicides for rectal use. Furthermore, such products can be evaluated for protection against rectal chlamydial infection in this model.

Effects of vaginal intercourse with and without a condom on vaginal flora and vaginal epithelium.

Effects of a single episode of intercourse on vaginal flora and epithelium were examined in subjects randomly assigned to groups that used no condom or lubricated nonspermicide condoms. Subjects were evaluated at visits before (1 month and 1-2 days) and after (8-12 h, 2-3 days, and 6-8 days) an index episode of sexual intercourse. The 22 subjects who used no condoms had significantly more Escherichia coli and a high concentration (> or =10(5) cfu/mL) of E. coli in the vagina (both, P<.001) and urine (all <10(5) cfu/mL; P=.004) at visit 3 than at visits 1 and 2. The 20 subjects who used condoms had a trend toward more vaginal E. coli (P=.06) and a significant increase in other enteric gram-negative rods (P=.001) after intercourse. Intercourse was not associated with gross, colposcopic, or histologic vaginal epithelial abnormalities.

Effects of oral contraceptive pill use on vaginal flora and vaginal epithelium.

The objective of this study was to examine the effect of oral contraceptive (OC) use on vaginal discharge, epithelium, and flora. Thirty women who planned to use OC for contraception were evaluated before and 2 months after the start of OC use. At both visits, genital symptoms and exposures were assessed by questionnaire; vaginal signs were assessed by speculum examination and colposcopy; vaginal microflora was evaluated by quantitative culture; and a vaginal biopsy was obtained for histopathologic evaluation. Variables were compared between the initial visit and after 2 months of OC use. It was found that OC use did not change the gross, colposcopic, or histologic appearance of the vaginal epithelium or characteristics of vaginal or cervical discharge. Vaginal flora essentially remained unchanged after 2 months of OC use, except that a small decrease occurred in the number of subjects with > or =10(5) colony forming units/mL of H(2)O(2) producing Lactobacillus from 16 at baseline to 9 (p = 0.04) and in the total number of subjects with Ureaplasma urealyticum from 17 at baseline to 10 of 29 (p = 0.04). The results indicate minimal effect of OC use on the vaginal epithelium and vaginal and cervical discharge, and a small effect on vaginal flora.

Epithelial cell layer thickness and immune cell populations in the normal human vagina at different stages of the menstrual cycle.

OBJECTIVE: The aim of our study was to examine vaginal tissue during 3 phases of the menstrual cycle for the number of cell layers and epithelial immune cells. STUDY DESIGN: Vaginal biopsies were performed during 3 phases of the normal menstrual cycle (menstrual, days 1-5; preovulatory, days 7-12; and postovulatory, days 19-24) in 74 subjects. A subset of women had vaginal tissues stained with specific monoclonal antibody markers for Langerhans cells (CD1a), macrophages (KP1), T and B lymphocytes (CD4, CD8, CD21) and neutrophils (CD15). The number of cell layers and the number of immune cells in the vaginal tissue biopsy specimen were determined by a single observer who was blinded to clinical data. RESULTS: At 3 phases of the normal menstrual cycle, the mean number of epithelial cell layers underwent a small but statistically significant decrease from 27.8 +/- 0.7 on days 1-5 and 28.1 +/- 0.6 on days 7-12 to 26.0 +/- 0.7 on days 19-24 of the cycle (P =.01). Nonovulating women had a reduced mean epithelial cell layer count on days 7-12 (23.7 +/- 1. 4) compared with the epithelial cell layer count in ovulating women (28.8 +/- 0.7; P =.005). No significant changes were observed in the mean number per high-power field of Langerhans cells, macrophages, CD4 or CD8 lymphocytes, and neutrophil cell populations during the 3 phases of the cycle. B lymphocytes were not observed in the vaginal tissues. CONCLUSION: A small but statistically significant reduction in the number of vaginal epithelial cells was observed over the menstrual cycle. This reduction is not likely to be clinically significant. Immune cell populations in the vaginal tissues appeared stable throughout the menstrual cycle.

Depomedroxyprogesterone-induced hypoestrogenism and changes in vaginal flora and epithelium.

OBJECTIVE: To identify the effects of depomedroxyprogesterone acetate (DMPA) on vaginal microbial flora and epithelium. METHODS: Women who desired DMPA for contraception were evaluated before and at 3 and 6 months after initiation of 150-mg DMPA injections every 3 months. At each visit, we assessed genital symptoms, vaginal signs, vaginal microflora, and histopathology by vaginal biopsies. RESULTS: Among 38 women observed for 6 months, there was significant reduction in mean serum estradiol level (99.9 +/- 9.3 pg/mL to 26.6 +/- 1.6 pg/mL, P <.001). The number of subjects with any Lactobacillus did not change, but the number with hydrogen peroxide (H(2)O(2))-positive Lactobacillus decreased from 20% before to 12% after 6 months of DMPA (P =.005). The log concentration in colony-forming units per milliliter of vaginal fluid of H(2)O(2)-positive Lactobacillus decreased in a linear manner from 4. 0 +/- 0.6 at baseline to 2.5 +/- 0.6 after 6 months of DMPA use (P =. 006). The mean number of cell layers in the epithelium was reduced slightly from 28.1 +/- 0.7 to 25.9 +/- 0.9 (P =.05), epithelial thickness decreased from 1.02 +/- 0.04 mm to 0.89 +/- 0.05 mm (P =. 005), and the glycogen-positive thickness decreased from 0.81 +/- 0. 04 mm at baseline to 0.66 +/- 0.05 after 6 months of DMPA use (P =. 005). CONCLUSION: Depomedroxyprogesterone acetate produced a systemic hypoestrogenic state associated with decreased H(2)O(2)-positive Lactobacillus colonization and slight thinning of the glycogen vaginal epithelial layer. Such changes possibly compromise the vaginal barrier to infection.

Detection of chlamydial antigenic material in ovarian, prostatic, ectopic pregnancy and semen samples of culture-negative subjects.

PROBLEM: The pathogenesis of long-term sequelae in Chlamydia trachomatis infection is poorly understood. While serology indicates previous chlamydial infection, culture studies are frequently negative. We wanted to know whether in chronic cases the bacterium is absent or persists in a dormant state where it evades detection. METHODS OF STUDY: Using immunoperoxidase (IP) staining and in situ hybridization (ISH), we examined tissues of culture-negative subjects. Ovarian biopsy specimens from 19 culture-negative women with pelvic adhesions and/or tubal infertility were analyzed by both methods. Samples of prostates from 10 culture-negative men undergoing prostatectomy for benign hypertrophy, two sets of semen samples from culture-negative sexual partners of 28 women with PID and/or bacterial vaginosis (BV), and ten endometrium-tube sample-pairs from ectopic pregnancies (EPs) were examined by IP only. RESULTS: Seven of the nineteen ovarian specimens tested positive for Chlamydia antigen or deoxyribonucleic acid (DNA) (36%). Of the 10 hypertrophic prostates examined, 4 (40%) were positive. Of the 28 semen samples examined, 10 (35%) tested positive. Tissue samples of 3 cases of EP were positive by IP. CONCLUSIONS: 1. C. trachomatis antigen and nucleic acid can be frequently demonstrated in asymptomatic, culture-negative men and women with chronic infection. 2. Chlamydia antigens may have an etiologic role in benign prostate hypertrophy and EP. 3. Antigenic material may be sexually transmissible. 4. IP and ISH identify temporarily inactive bacteria that may continue to act as immunostimulants and potentially reactivate as Chlamydia infection.

Vaginal myeloperoxidase and flora in the pig-tailed macaque.

Myeloperoxidase (MPO) is an enzyme in neutrophils and monocytes which reacts with H2O2 and chloride to kill microbes after phagocytosis. Instillation of MPO into the vagina may augment vaginal defenses against sexually transmitted diseases, since the normal vaginal flora is characterized by the presence of H2O2-producing lactobacilli. We assessed the menstrual cycle stage, vaginal flora, pH, macroscopic appearance, and endogenous MPO in the adult female pig-tailed macaque (Macaca nemestrina) at baseline (n = 26; 60 observations) and at 0, 4, and 24 hours in untreated animals (n = 6) or in animals treated with intravaginal MPO gel at time 0 (n = 5). Baseline MPO levels were highly variable, and there was no detectable effect of cycle stage. In untreated animals, there was no significant effect of vaginal swab collection on vaginal flora or MPO levels. MPO treatment did not reduce vaginal H2O2-producing organisms, and vaginal MPO levels tended to increase at 4 hours in treated animals. Vaginal/cervical colposcopic changes were not detected in either group.

Influence of the normal menstrual cycle on vaginal tissue, discharge, and microflora.

The objective of this study was to examine genital tissue, vaginal fluid, and vaginal microbial flora at 3 phases of the menstrual cycle in asymptomatic women. Vaginal examinations were performed 3 times in 74 women: at the menstrual phase (days 1-5), the preovulatory phase (days 7-12), and the postovulatory phase (days 19-24). Flora of 50 women without bacterial vaginosis (BV) was analyzed separately from flora of 24 women with BV. The volume of vaginal discharge increased and the amount of cervical mucus decreased over the menstrual cycle. Among subjects without BV, the rate of recovery of any Lactobacillus changed little (range, 82% to 98%; P = .2); however, a small increase occurred in the rate of recovery of heavy (3+ to 4+ semiquantitative) growth of Lactobacillus over the menstrual cycle (P = .04). A linear decrease occurred in the rate of recovery of heavy growth of any non-Lactobacillus species, from 72% at days 1-5 to 40% at days 19-24 (P = .002). A linear decrease also occurred in the rate of recovery of Prevotella species, from 56% on days 1-5 to 28% on days 19-24 (P =. 007), while a small linear increase occurred in the rate of recovery of Bacteroides fragilis (P=.05). Among subjects with BV, the only significant change was an increase in the rate of recovery of Lactobacillus, from 33% at days 1-5 to 54% at days 19-24 (P = .008). Among all subjects, the rate of recovery of heavy growth of Lactobacillus increased over the menstrual cycle and, in contrast, the concentration of non-Lactobacillus species tended to be higher at menses, which is evidence that the vaginal flora becomes less stable at this time.

Antibody response to the chlamydial heat-shock protein 60 in an experimental model of chronic pelvic inflammatory disease in monkeys (Macaca nemestrina).

A primate model of chlamydial pelvic inflammatory disease was used to characterize serum antibody responses to the 60 kDa chlamydial heat shock protein (CHSP60). Forty monkeys were infected in the fallopian tubes with Chlamydia trachomatis and then were treated. Twenty-three (58%) monkeys developed antibodies against CHSP60, of whom 6 (15%) had CHSP60 responses that persisted throughout the study and 17 (42.5%) had a transient response. A persistent CHSP60 antibody response was correlated with being culture- or ligase chain reaction-positive in the fallopian tubes (P=.004), but not in the cervix pretreatment, and with being tubal-positive posttreatment (P=. 02). Compared with tubal-negative monkeys, tubal-positive monkeys had more intense CHSP60 responses (P=.006) that lasted longer (P=. 002). Among CHSP60 responders, an OD>0.5 was correlated with more severe salpingeal pathology before treatment (P=.04). CHSP60 antibody response may be useful as a marker of persistent chlamydial infection in the fallopian tubes.

Effects of multiple applications of benzalkonium chloride and nonoxynol 9 on the vaginal epithelium in the pigtailed macaque (Macaca nemestrina).

OBJECTIVE: Safe and effective vaginally applied microbicides could help to control the continuing spread of sexually transmitted diseases. STUDY DESIGN: This study used nonhuman primates to test the effects of multiple applications of nonoxynol 9, benzalkonium chloride, or a combination on vaginal flora and lower reproductive tract tissues. Fourteen monkeys (Macaca nemestrina) received daily vaginal applications of nonoxynol 9, benzalkonium chloride, or both for 3 to 4 days. Vaginal microflora and colposcopic observations were made at baseline and during and after completion of treatments. Cervical biopsy specimens were collected from a subset of animals. RESULTS: Cervical erythema and vaginal erythema were observed in all 3 treatment groups. Cervical papillae and epithelial disruption were present in both the nonoxynol 9 and the nonoxynol 9 plus benzalkonium chloride groups. Vaginal epithelial disruption was noted in both the benzalkonium chloride and the nonoxynol 9 plus benzalkonium chloride groups. Cervical biopsy specimens from each group revealed acute inflammatory infiltrates with occasional plasma cells and lymphoid follicles. Detection of most microorganisms, including viridans streptococci, decreased in the benzalkonium chloride and the nonoxynol 9 plus benzalkonium chloride groups. Detection of Lactobacillus species decreased in the benzalkonium chloride group. All microflora levels recovered after several days without microbicide use. CONCLUSIONS: Although nonoxynol 9 is currently the only microbicide approved for use as a spermicide in the United States, its repeated use may be detrimental to the epithelial tissues of the female reproductive tract. Benzalkonium chloride, currently approved for use in other countries, not only may damage epithelial tissues but also appears to reduce the population of potentially protective Lactobacillus species in the vagina.

Characterization of ciliary activity in distal Fallopian tube biopsies of women with obstructive tubal infertility.

Biopsy specimens were obtained from the distal end of the Fallopian tubes of 62 women with tubal infertility and examined by light and electron microscopy. Ciliary beat frequency (CBF) measurements were obtained using laser light spectroscopy. Neither demographic nor behavioural characteristics nor serological evidence of past chlamydial infection were associated with CBF measurements. In contrast, CBF were significantly lower (P < 0.05) in tissues with oedema compared to tissues without oedema (6.7 versus 12.9) and in tissues with erythema compared to tissues without erythema (9.2 versus 13.7). Furthermore, CBF measurements did vary by chlamydial serotype pattern, with lower values observed among the tissues of women with antibodies to serotype C or E (without D) as compared to the tissues of women with other serotypes (P < 0.04). However, these data must be interpreted with caution as the numbers of subjects with chlamydial antibodies to serotype C (n = 3) or E without D (n = 5) were few in number and serotyping of IgG antibodies in blood is not as accurate as it is in bacterial isolates. Confirmation of the suggested association between chlamydial serotype and risk of adverse sequelae could indicate potential new avenues for vaccine research.

0.25% chlorhexidine gluconate gel. A protective topical microbicide.

BACKGROUND AND OBJECTIVES: An estimated 4 million new cases of chlamydial infection occur each year. This experiment assessed the effects of a vaginally applied gel formulation of 0.25% chlorhexidine gluconate on chlamydial infection and on the vaginal ecosystem. STUDY DESIGN: Twelve monkeys were treated with a single application of 0.25% chlorhexidine gluconate. These animals were assessed for changes in vaginal flora before and at 30 minutes, 1 day, and 2 days postapplication by microbiologic analysis. Cervical and vaginal tissues were assessed by colposcopy at each time point. Five monkeys received a single application of 0.25% chlorhexidine gluconate gel followed (30 minutes) by a cervical inoculation with Chlamydia trachomatis. Four monkeys were inoculated with Chlamydia only. Cervicovaginal tissues were assessed via modified colposcopy, vaginal swabs were collected for assessment of vaginal flora, and cervical swabs were collected for detection of Chlamydia (culture/ligase chain reaction) at baseline and days 1, 2, and 7 postinoculation. RESULTS: Changes in vaginal flora were minimal in all monkeys. Application of 0.25% chlorhexidine gluconate did not affect adversely vaginal colonization by lactobacilli. All chlamydial infection control monkeys were infected, whereas none of the five monkeys pretreated with chlorhexidine gluconate were positive for C. trachomatis by culture or ligase chain reaction. Colposcopic observations remained largely unchanged in all groups. CONCLUSIONS: A 0.25% chlorhexidine gluconate gel was protective against chlamydial infection in all animals tested, had no adverse effect on the vaginal flora, and had minimal effect on cervicovaginal tissues after a single application.

Killing of Chlamydia trachomatis by novel antimicrobial lipids adapted from compounds in human breast milk.

The development of new methods for prevention of sexually transmitted Chlamydia trachomatis infection is a top public health priority. Topical self-administered vaginal microbicides represent one such approach in which the organism is eradicated at the time of initial exposure. To this end, we examined the activity of five synthetic lipids adapted from naturally occurring compounds found in human breast milk. C. trachomatis serovar D or F elementary bodies were added to serial dilutions of the lipids and incubated for various times. Aliquots were then cultured in monolayers of McCoy cells, and inclusions were counted. A 7.5 mM concentration of 2-O-octyl-sn-glycerol completely prevented growth of C. trachomatis after 120 min of contact with the organism. The remaining lipids, 1-O-octyl-, 1-O-heptyl-, 2-O-hexyl-, and 1-O-hexyl-sn-glycerol, showed less activity. On electron microscopic examination, the lipids were shown to have disrupted the chlamydial inner membrane, allowing leakage of the cytoplasmic contents from the cell. Lipid activity was unaffected by the presence of 10% human blood or alterations in pH from 4.0 to 8.0, conditions reflecting those sometimes found in the vagina. Our results suggest that these lipids, especially 2-O-octyl-sn-glycerol, may be effective as topical microbicides in preventing the transmission of C. trachomatis. Further efficacy and toxicity studies with these lipids and assessment of their activity against other sexually transmitted disease pathogens are in progress.

Lipopolysaccharide binding protein and soluble CD14 receptor protein in amniotic fluid and cord blood in patients at term.

OBJECTIVES: Our purpose was to examine whether lipopolysaccharide binding protein and soluble CD14 are present in amniotic fluid and to determine whether the lipopolysaccharide binding protein and soluble CD14 concentrations are associated with indicators of infection or labor at term. A lipopolysaccharide-lipopolysaccharide binding protein complex activates macrophages through soluble CD14 at lipopolysaccharide concentrations up to 100 times lower than required with lipopolysaccharide alone. Thus lipopolysaccharide binding protein and soluble CD14 in amniotic fluid could explain the high concentrations of cytokines found in amniotic fluid of culture-positive patients and may even explain the presence of cytokines in some culture-negative patients. STUDY DESIGN: Healthy women at term undergoing cesarean section had amniotic fluid, chorioamnion, decidua, and cord blood obtained. Lipopolysaccharide binding protein was measured by enzyme-linked immunosorbent assay. Amniotic fluid was cultured and assayed for cytokines, and the chorioamnion and decidua were cultured and examined histologically. RESULTS: Lipopolysaccharide binding protein and soluble CD14 were present in all amniotic fluids and fetal cord blood. An elevated level of lipopolysaccharide binding protein (270 ng/ml/mg of protein) was present in the amniotic fluid of 12 (36%) of the 33 patients. An elevated level was associated with microorganisms in the chorioamnion and decidua, cytokines (tumor necrosis factor-alpha, interleukin-6, and interleukin-8) in amniotic fluid, histologic chorioamnionitis, and labor. Among patients in labor, the concentration of lipopolysaccharide binding protein appeared independent of microorganisms in the amniotic fluid. CONCLUSIONS: Lipopolysaccharide binding protein and soluble CD14 are present in amniotic fluid, and concentrations of lipopolysaccharide binding protein are elevated in patients in labor with and without evidence of infection. Lipopolysaccharide binding protein and soluble CD14 may mediate intrauterine inflammatory responses at term.