General Principles for Counseling and Management of Triplet Pregnancies.

Compared with singleton pregnancies, triplet pregnancies are associated with a significantly increased risk of adverse pregnancy outcomes. Early ultrasound examination is the best way to diagnose triplets, establish dating, and determine the number of placentas to provide appropriate counseling and monitoring. Dichorionic placentation adds risks specifically associated with a shared placenta, and limits options for intervention. Multifetal reduction is an option that can significantly improve pregnancy outcomes compared with non-reduced triplet pregnancies. Integration of a Maternal-Fetal Medicine specialist in the prenatal care for a triplet pregnancy reduces the risk of preeclampsia, preterm birth, low birthweight infants, perinatal mortality, and major neonatal morbidity.

Increased Scheduled Intravenous Ketorolac After Cesarean Delivery and Its Effect on Opioid Use: A Randomized Controlled Trial.

OBJECTIVE: To evaluate the efficacy of scheduled ketorolac in reducing opioid use after cesarean delivery. METHODS: This was a single-center, randomized, double-blind, parallel-group trial to assess pain management after cesarean delivery with scheduled ketorolac compared with placebo. All patients undergoing cesarean delivery with neuraxial anesthesia received two doses of 30 mg intravenous ketorolac postoperatively and then were randomized to receive four doses of 30 mg of intravenous ketorolac or placebo every 6 hours. Additional nonsteroidal anti-inflammatory drugs were held until 6 hours after the last study dose. The primary outcome was total morphine milligram equivalents (MME) used in the first 72 postoperative hours. Secondary outcomes included the number of patients who used no opioid postoperatively, postoperative pain scores, postoperative change in hematocrit and serum creatinine, and postoperative satisfaction with inpatient care and pain management. A sample size of 74 per group (n=148) provided 80% power to detect a population mean difference in MME of 32.4, with an SD for both groups of 68.7 after accounting for protocol noncompliance. RESULTS: From May 2019 to January 2022, 245 patients were screened and 148 patients were randomized (74 per group). Patient characteristics were similar between groups. The median (quartile 1-3) MME from arrival in the recovery room until postoperative hour 72 was 30.0 (0.0-67.5) for the ketorolac group and 60.0 (30.0-112.5) for the placebo group (Hodges-Lehmann median difference -30.0, 95% CI -45.0 to -15.0, P <.001). In addition, participants who received placebo were more likely to have numeric rating scale pain scores higher than 3 out of 10 ( P= .005). The mean±SD decrease from baseline hematocrit to postoperative day 1 was 5.5±2.6% for the ketorolac group and 5.4±3.5% for the placebo group ( P =.94). The mean±SD postoperative day 2 creatinine was 0.61±0.06 mg/dL in the ketorolac group and 0.62±0.08 mg/dL in the placebo group ( P =.26). Participant satisfaction with inpatient pain control and postoperative care was similar between groups. CONCLUSION: Compared with placebo, scheduled intravenous ketorolac significantly decreased opioid use after cesarean delivery. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov , NCT03678675.

Effects of prenatal exposure to maternal COVID-19 and perinatal care on neonatal outcome: results from the INTERCOVID Multinational Cohort Study.

BACKGROUND: The effect of COVID-19 in pregnancy on maternal outcomes and its association with preeclampsia and gestational diabetes mellitus have been reported; however, a detailed understanding of the effects of maternal positivity, delivery mode, and perinatal practices on fetal and neonatal outcomes is urgently needed. OBJECTIVE: To evaluate the impact of COVID-19 on fetal and neonatal outcomes and the role of mode of delivery, breastfeeding, and early neonatal care practices on the risk of mother-to-child transmission. STUDY DESIGN: In this cohort study that took place from March 2020 to March 2021, involving 43 institutions in 18 countries, 2 unmatched, consecutive, unexposed women were concomitantly enrolled immediately after each infected woman was identified, at any stage of pregnancy or delivery, and at the same level of care to minimize bias. Women and neonates were followed up until hospital discharge. COVID-19 in pregnancy was determined by laboratory confirmation and/or radiological pulmonary findings or ≥2 predefined COVID-19 symptoms. The outcome measures were indices of neonatal and perinatal morbidity and mortality, neonatal positivity and its correlation with mode of delivery, breastfeeding, and hospital neonatal care practices. RESULTS: A total of 586 neonates born to women with COVID-19 diagnosis and 1535 neonates born to women without COVID-19 diagnosis were enrolled. Women with COVID-19 diagnosis had a higher rate of cesarean delivery (52.8% vs 38.5% for those without COVID-19 diagnosis, P<.01) and pregnancy-related complications, such as hypertensive disorders of pregnancy and fetal distress (all with P<.001), than women without COVID-19 diagnosis. Maternal diagnosis of COVID-19 carried an increased rate of preterm birth (P≤.001) and lower neonatal weight (P≤.001), length, and head circumference at birth. In mothers with COVID-19 diagnosis, the length of in utero exposure was significantly correlated to the risk of the neonate testing positive (odds ratio, 4.5; 95% confidence interval, 2.2-9.4 for length of in utero exposure >14 days). Among neonates born to mothers with COVID-19 diagnosis, birth via cesarean delivery was a risk factor for testing positive for COVID-19 (odds ratio, 2.4; 95% confidence interval, 1.2-4.7), even when severity of maternal conditions was considered and after multivariable logistic analysis. In the subgroup of neonates born to women with COVID-19 diagnosis, the outcomes worsened when the neonate also tested positive, with higher rates of neonatal intensive care unit admission, fever, gastrointestinal and respiratory symptoms, and death, even after adjusting for prematurity. Breastfeeding by mothers with COVID-19 diagnosis and hospital neonatal care practices, including immediate skin-to-skin contact and rooming-in, were not associated with an increased risk of newborn positivity. CONCLUSION: In this multinational cohort study, COVID-19 in pregnancy was associated with increased maternal and neonatal complications. Cesarean delivery was significantly associated with newborn COVID-19 diagnosis. Vaginal delivery should be considered the safest mode of delivery if obstetrical and health conditions allow it. Mother-to-child skin-to-skin contact, rooming-in, and direct breastfeeding were not risk factors for newborn COVID-19 diagnosis, thus well-established best practices can be continued among women with COVID-19 diagnosis.

Diabetes mellitus, maternal adiposity, and insulin-dependent gestational diabetes are associated with COVID-19 in pregnancy: the INTERCOVID study.

BACKGROUND: Among nonpregnant individuals, diabetes mellitus and high body mass index increase the risk of COVID-19 and its severity. OBJECTIVE: This study aimed to determine whether diabetes mellitus and high body mass index are risk factors for COVID-19 in pregnancy and whether gestational diabetes mellitus is associated with COVID-19 diagnosis. STUDY DESIGN: INTERCOVID was a multinational study conducted between March 2020 and February 2021 in 43 institutions from 18 countries, enrolling 2184 pregnant women aged ≥18 years; a total of 2071 women were included in the analyses. For each woman diagnosed with COVID-19, 2 nondiagnosed women delivering or initiating antenatal care at the same institution were also enrolled. The main exposures were preexisting diabetes mellitus, high body mass index (overweight or obesity was defined as a body mass index ≥25 kg/m2), and gestational diabetes mellitus in pregnancy. The main outcome was a confirmed diagnosis of COVID-19 based on a real-time polymerase chain reaction test, antigen test, antibody test, radiological pulmonary findings, or ≥2 predefined COVID-19 symptoms at any time during pregnancy or delivery. Relationships of exposures and COVID-19 diagnosis were assessed using generalized linear models with a Poisson distribution and log link function, with robust standard errors to account for model misspecification. Furthermore, we conducted sensitivity analyses: (1) restricted to those with a real-time polymerase chain reaction test or an antigen test in the last week of pregnancy, (2) restricted to those with a real-time polymerase chain reaction test or an antigen test during the entire pregnancy, (3) generating values for missing data using multiple imputation, and (4) analyses controlling for month of enrollment. In addition, among women who were diagnosed with COVID-19, we examined whether having gestational diabetes mellitus, diabetes mellitus, or high body mass index increased the risk of having symptomatic vs asymptomatic COVID-19. RESULTS: COVID-19 was associated with preexisting diabetes mellitus (risk ratio, 1.94; 95% confidence interval, 1.55-2.42), overweight or obesity (risk ratio, 1.20; 95% confidence interval, 1.06-1.37), and gestational diabetes mellitus (risk ratio, 1.21; 95% confidence interval, 0.99-1.46). The gestational diabetes mellitus association was specifically among women requiring insulin, whether they were of normal weight (risk ratio, 1.79; 95% confidence interval, 1.06-3.01) or overweight or obese (risk ratio, 1.77; 95% confidence interval, 1.28-2.45). A somewhat stronger association with COVID-19 diagnosis was observed among women with preexisting diabetes mellitus, whether they were of normal weight (risk ratio, 1.93; 95% confidence interval, 1.18-3.17) or overweight or obese (risk ratio, 2.32; 95% confidence interval, 1.82-2.97). When the sample was restricted to those with a real-time polymerase chain reaction test or an antigen test in the week before delivery or during the entire pregnancy, including missing variables using imputation or controlling for month of enrollment, the observed associations were comparable. CONCLUSION: Diabetes mellitus and overweight or obesity were risk factors for COVID-19 diagnosis in pregnancy, and insulin-dependent gestational diabetes mellitus was associated with the disease. Therefore, it is essential that women with these comorbidities are vaccinated.

Preeclampsia and COVID-19: results from the INTERCOVID prospective longitudinal study.

BACKGROUND: It is unclear whether the suggested link between COVID-19 during pregnancy and preeclampsia is an independent association or if these are caused by common risk factors. OBJECTIVE: This study aimed to quantify any independent association between COVID-19 during pregnancy and preeclampsia and to determine the effect of these variables on maternal and neonatal morbidity and mortality. STUDY DESIGN: This was a large, longitudinal, prospective, unmatched diagnosed and not-diagnosed observational study assessing the effect of COVID-19 during pregnancy on mothers and neonates. Two consecutive not-diagnosed women were concomitantly enrolled immediately after each diagnosed woman was identified, at any stage during pregnancy or delivery, and at the same level of care to minimize bias. Women and neonates were followed until hospital discharge using the standardized INTERGROWTH-21st protocols and electronic data management system. A total of 43 institutions in 18 countries contributed to the study sample. The independent association between the 2 entities was quantified with the risk factors known to be associated with preeclampsia analyzed in each group. The outcomes were compared among women with COVID-19 alone, preeclampsia alone, both conditions, and those without either of the 2 conditions. RESULTS: We enrolled 2184 pregnant women; of these, 725 (33.2%) were enrolled in the COVID-19 diagnosed and 1459 (66.8%) in the COVID-19 not-diagnosed groups. Of these women, 123 had preeclampsia of which 59 of 725 (8.1%) were in the COVID-19 diagnosed group and 64 of 1459 (4.4%) were in the not-diagnosed group (risk ratio, 1.86; 95% confidence interval, 1.32-2.61). After adjustment for sociodemographic factors and conditions associated with both COVID-19 and preeclampsia, the risk ratio for preeclampsia remained significant among all women (risk ratio, 1.77; 95% confidence interval, 1.25-2.52) and nulliparous women specifically (risk ratio, 1.89; 95% confidence interval, 1.17-3.05). There was a trend but no statistical significance among parous women (risk ratio, 1.64; 95% confidence interval, 0.99-2.73). The risk ratio for preterm birth for all women diagnosed with COVID-19 and preeclampsia was 4.05 (95% confidence interval, 2.99-5.49) and 6.26 (95% confidence interval, 4.35-9.00) for nulliparous women. Compared with women with neither condition diagnosed, the composite adverse perinatal outcome showed a stepwise increase in the risk ratio for COVID-19 without preeclampsia, preeclampsia without COVID-19, and COVID-19 with preeclampsia (risk ratio, 2.16; 95% confidence interval, 1.63-2.86; risk ratio, 2.53; 95% confidence interval, 1.44-4.45; and risk ratio, 2.84; 95% confidence interval, 1.67-4.82, respectively). Similar findings were found for the composite adverse maternal outcome with risk ratios of 1.76 (95% confidence interval, 1.32-2.35), 2.07 (95% confidence interval, 1.20-3.57), and 2.77 (95% confidence interval, 1.66-4.63). The association between COVID-19 and gestational hypertension and the direction of the effects on preterm birth and adverse perinatal and maternal outcomes, were similar to preeclampsia, but confined to nulliparous women with lower risk ratios. CONCLUSION: COVID-19 during pregnancy is strongly associated with preeclampsia, especially among nulliparous women. This association is independent of any risk factors and preexisting conditions. COVID-19 severity does not seem to be a factor in this association. Both conditions are associated independently of and in an additive fashion with preterm birth, severe perinatal morbidity and mortality, and adverse maternal outcomes. Women with preeclampsia should be considered a particularly vulnerable group with regard to the risks posed by COVID-19.

Maternal and Neonatal Morbidity and Mortality Among Pregnant Women With and Without COVID-19 Infection: The INTERCOVID Multinational Cohort Study.

Importance: Detailed information about the association of COVID-19 with outcomes in pregnant individuals compared with not-infected pregnant individuals is much needed. Objective: To evaluate the risks associated with COVID-19 in pregnancy on maternal and neonatal outcomes compared with not-infected, concomitant pregnant individuals. Design, Setting, and Participants: In this cohort study that took place from March to October 2020, involving 43 institutions in 18 countries, 2 unmatched, consecutive, not-infected women were concomitantly enrolled immediately after each infected woman was identified, at any stage of pregnancy or delivery, and at the same level of care to minimize bias. Women and neonates were followed up until hospital discharge. Exposures: COVID-19 in pregnancy determined by laboratory confirmation of COVID-19 and/or radiological pulmonary findings or 2 or more predefined COVID-19 symptoms. Main Outcomes and Measures: The primary outcome measures were indices of (maternal and severe neonatal/perinatal) morbidity and mortality; the individual components of these indices were secondary outcomes. Models for these outcomes were adjusted for country, month entering study, maternal age, and history of morbidity. Results: A total of 706 pregnant women with COVID-19 diagnosis and 1424 pregnant women without COVID-19 diagnosis were enrolled, all with broadly similar demographic characteristics (mean [SD] age, 30.2 [6.1] years). Overweight early in pregnancy occurred in 323 women (48.6%) with COVID-19 diagnosis and 554 women (40.2%) without. Women with COVID-19 diagnosis were at higher risk for preeclampsia/eclampsia (relative risk [RR], 1.76; 95% CI, 1.27-2.43), severe infections (RR, 3.38; 95% CI, 1.63-7.01), intensive care unit admission (RR, 5.04; 95% CI, 3.13-8.10), maternal mortality (RR, 22.3; 95% CI, 2.88-172), preterm birth (RR, 1.59; 95% CI, 1.30-1.94), medically indicated preterm birth (RR, 1.97; 95% CI, 1.56-2.51), severe neonatal morbidity index (RR, 2.66; 95% CI, 1.69-4.18), and severe perinatal morbidity and mortality index (RR, 2.14; 95% CI, 1.66-2.75). Fever and shortness of breath for any duration was associated with increased risk of severe maternal complications (RR, 2.56; 95% CI, 1.92-3.40) and neonatal complications (RR, 4.97; 95% CI, 2.11-11.69). Asymptomatic women with COVID-19 diagnosis remained at higher risk only for maternal morbidity (RR, 1.24; 95% CI, 1.00-1.54) and preeclampsia (RR, 1.63; 95% CI, 1.01-2.63). Among women who tested positive (98.1% by real-time polymerase chain reaction), 54 (13%) of their neonates tested positive. Cesarean delivery (RR, 2.15; 95% CI, 1.18-3.91) but not breastfeeding (RR, 1.10; 95% CI, 0.66-1.85) was associated with increased risk for neonatal test positivity. Conclusions and Relevance: In this multinational cohort study, COVID-19 in pregnancy was associated with consistent and substantial increases in severe maternal morbidity and mortality and neonatal complications when pregnant women with and without COVID-19 diagnosis were compared. The findings should alert pregnant individuals and clinicians to implement strictly all the recommended COVID-19 preventive measures.

Triplet pregnancy: What do we tell the prospective parents.

Experience managing triplet pregnancies has increased over the past few decades as the incidence has changed related to assisted reproductive practices. Physicians caring for women carrying triplets cannot predict an individual outcome or pregnancy course but must educate patients about the challenges related to these high risk pregnancies. Obstetric providers can describe the wide range of risks associated with triplet gestations, and the general plan for management, but ultimately parents must make decisions with potentially lifelong consequences. Here, we present the diagnostic criteria, common complications, and management options for triplet pregnancies, to help obstetricians counsel patients on the medical and psychosocial consequences of triplet pregnancy, potential complications, and multifetal reduction.

Placenta Accreta: A Spectrum of Predictable Risk, Diagnosis, and Morbidity.

OBJECTIVE: Placenta accreta is a feared pathology, in part, because prenatal diagnosis is imperfect. It is not known whether clinical risk factors or sonographic features equally predict the entire graded pathological spectrum of placental overinvasion disease nor whether clinical outcomes differ along the spectrum. STUDY DESIGN: We conducted a mixed methods retrospective study of a cohort of women screened sonographically for placenta accreta, cross-referenced against cases identified by pathological diagnosis (N = 416). Demographic, diagnostic, and outcome information were compared across the spectrum of invasive placentation: percreta, increta, accreta, and focal accreta not requiring hysterectomy. The t-test, chi-square, Mann-Whitney, and Kruskal-Wallis tests were used for statistical analysis across groups. RESULTS: As the depth of invasion decreased, risk factors for placental overinvasion were less common, especially placenta previa and previous cesarean. There was also reduced anticipation by sonographic examination of the placenta. Rates of adverse outcomes were lower among women with focal accreta compared with those with deeper invasion. CONCLUSION: As the depth of invasion decreases, clinical risk factors and sonographic evaluation are less reliable in the antenatal prediction of placenta accreta. The potential for unanticipated morbidity underscores the need for improved diagnostic tools for placenta accreta spectrum.

The Effect of Maternal Obesity on Placental Cell-Free DNA Release in a Mouse Model.

BACKGROUND: The fetal fraction of cell-free DNA (cfDNA) in maternal plasma is decreased in obese women. The underlying mechanism is not well understood. The amount of cfDNA released from the placenta has not been directly examined in maternal obesity. OBJECTIVE: We sought to quantify release of cfDNA from the placenta and fetal membranes in maternal diet-induced obesity using explant cultures in an established mouse model. STUDY DESIGN: C57BL6/J females were fed either 60% high-fat diet or 10% fat-matched control diet for 14 weeks prepregnancy and throughout gestation. Placentas and fetal membranes were collected on e18 and randomly allocated to time 0-, 1-, or 6-hour culture times. The CfDNA was isolated from culture media, quantified, and normalized to tissue weight. RESULTS: Placentas from obese dams released significantly less cfDNA compared to those of lean dams at time 0 (45.8 ± 4.3 ng/mg vs 65.6 ± 7.9 ng/mg, P = .02). Absolute cfDNA levels increased with longer placental culture, with no significant differences between obese and lean dams at 1 and 6 hours. Membranes released significantly less cfDNA than did placentas at every time point. CONCLUSIONS: Maternal obesity is associated with decreased release of cfDNA from the placenta compared to lean controls immediately after tissue harvest. This may provide an alternative explanation for the lower fetal fraction of cfDNA noted in maternal obesity.

Successful Use of Extracorporeal Membrane Oxygenation for Status Asthmaticus in a Woman With a Periviable Pregnancy.

BACKGROUND: Status asthmaticus is a severe asthma exacerbation with persistent airway obstruction despite standard therapy. Use of extracorporeal membrane oxygenation (ECMO) as rescue therapy in pregnancy is exceedingly rare. We describe a case of ECMO for treatment of status asthmaticus in woman with a periviable pregnancy culminating in a term delivery. CASE: The patient was a 33-year-old woman, gravida 3 para 1, admitted at 23 2/7 weeks of gestation with respiratory failure secondary to status asthmaticus. Venovenous ECMO was initiated and continued for 6 days. After hospital discharge, she had no further respiratory issues. She ultimately developed fetal growth restriction and gestational hypertension and underwent a repeat cesarean delivery at 38 weeks of gestation. CONCLUSION: Venovenous ECMO can be used successfully for status asthmaticus during a periviable pregnancy and enable delivery at term.

Thrombin Augments LPS-Induced Human Endometrial Endothelial Cell Inflammation via PAR1 Activation.

PROBLEM: Risk factors for preterm birth include placental abruption, giving rise to excessive decidual thrombin, and intrauterine bacterial infection. Human endometrial endothelial cells (HEECs) express Toll-like receptors (TLRs), and infection-derived agonists trigger HEECs to generate specific inflammatory responses. As thrombin, in addition to inducing coagulation, can contribute to inflammation, its effect on HEEC inflammatory responses to the TLR4 agonist, bacterial lipopolysaccharide (LPS), was investigated. METHOD OF STUDY: HEECs were pre-treated with or without thrombin or specific protease-activated receptor (PAR) agonists, followed by treatment with or without LPS. Supernatants were measured for cytokines and chemokines by ELISA and multiplex analysis. RESULTS: Thrombin significantly and synergistically augmented LPS-induced HEEC secretion of interleukin (IL)-6, IL-8, granulocyte colony-stimulating factor (G-CSF), and growth-regulated oncogene-alpha (GRO-α), and significantly augmented monocyte chemotactic protein (MCP)-1, tumor necrosis factor-alpha (TNF-α), and vascular endothelial growth factor (VEGF) secretion additively. Similar to thrombin, a PAR1 agonist synergistically augmented the LPS-induced HEEC secretion of inflammatory IL-6, IL-8, G-CSF, and GRO-α. CONCLUSION: Thrombin, via PAR1 activation, synergistically augments LPS-induced HEEC production of chemokines involved in immune cell recruitment and survival, suggesting a mechanism by which intrauterine abruption and bacterial infection may together be associated with an aggravated uterine inflammatory response.

Association of bactericidal activity of genital tract secretions with Escherichia coli colonization in pregnancy.

OBJECTIVE: Genital tract secretions exhibit bactericidal activity against Escherichia coli. We hypothesized that this defense may be modulated during pregnancy. STUDY DESIGN: Secretions were collected by vaginal swab from 70 pregnant women (35-37 weeks' gestation) and 35 nonpregnant controls. We mixed E coli with swab eluants or control buffer and colonies enumerated to measure bactericidal activity. Cytokines, chemokines, and antimicrobial peptides were quantified by multiplex or enzyme-linked immunosorbent assay. RESULTS: Pregnant women had significantly greater bactericidal activity, higher concentrations of proinflammatory cytokines, and lower levels of beta defensins compared to controls. Seven (10%) pregnant and 8 (23%) nonpregnant women were vaginally colonized with E coli; colonization was inversely associated with bactericidal activity. CONCLUSION: The soluble mucosal immune environment is altered in pregnancy. We speculate that the observed changes may protect against colonization and ascending infection and could provide a biomarker to identify pregnant women at risk for infectious complications including preterm birth.

Cervical intraepithelial neoplasia is associated with genital tract mucosal inflammation.

BACKGROUND: Clinical studies demonstrate increased prevalence of human papillomavirus (HPV)-associated disease in HIV-infected individuals and an increased risk of HIV acquisition in HPV-infected individuals. The mechanisms underlying this synergy are not defined. We hypothesize that women with cervical intraepithelial neoplasia (CIN) will exhibit changes in soluble mucosal immunity that may promote HPV persistence and facilitate HIV infection. METHODS: The concentrations of immune mediators and endogenous anti-Escherichia coli activity in genital tract secretions collected by cervicovaginal lavage were compared in HIV-negative women with high-risk HPV-positive (HRHPV+) CIN-3 (n = 37), HRHPV+ CIN-1 (n = 12), or PAP-negative control subjects (n = 57). RESULTS: Compared with control subjects, women with CIN-3 or CIN-1 displayed significantly higher levels of proinflammatory cytokines including interleukin (IL)-1α, IL-1ß, and IL-8 (P < 0.002) and significantly lower levels of anti-inflammatory mediators and antimicrobial peptides, including IL-1 receptor antagonist, secretory leukocyte protease inhibitor (P < 0.01), and human ß defensins 2 and 3 (P < 0.02). There was no significant difference in endogenous anti-E. coli activity after controlling for age and sample storage time. CONCLUSION: HRHPV+ CIN is characterized by changes in soluble mucosal immunity that could contribute to HPV persistence. The observed mucosal inflammation suggests a mechanism that may also contribute to the epidemiologic link between persistent HPV and HIV.

Disruption of tight junctions by cellulose sulfate facilitates HIV infection: model of microbicide safety.

BACKGROUND: The lack of biomarkers that are predictive of safety is a critical gap in the development of microbicides. The present experiments were designed to evaluate the predictive value of in vitro models of microbicide safety. METHODS: Changes in the epithelial barrier were evaluated by measuring transepithelial electrical resistance (TER) after exposure of human epithelial cells to candidate microbicides in a dual-chamber system. The significance of observed changes was addressed by challenging cultures with human immunodeficiency virus (HIV) and measuring the ability of virus to cross the epithelium and infect target T cells cultured in the lower chamber. RESULTS: Exposure to nonoxynol-9 (N-9) or cellulose sulfate (CS), but not 9-[2-(phosphonomethoxy)propyl]adenine (also referred to as tenofovir) or PRO2000, resulted in a rapid and sustained reduction in TER and a marked increase in HIV infection of T cells cultured in the lower chamber. Moreover, CS triggered nuclear factor kappaB activation in peripheral blood mononuclear cells and increased HIV replication in chronically infected U1 cells. CONCLUSIONS: Epithelial barrier disruption and enhanced viral replication may have contributed to the increased risk of HIV acquisition observed in phase 3 trials of N-9 and CS. Expansion of in vitro safety testing to include these models would provide a more stringent preclinical assessment of microbicide safety and may prove to be more predictive of clinical outcomes.