Characterization of a Hormone-Responsive Organotypic Human Vaginal Tissue Model: Morphologic and Immunologic Effects.

Estrogen and progesterone regulate proliferation and differentiation of epithelial cells in the female genital tract. We investigated the effects of these hormones on reconstructed human organotypic vaginal epithelial tissue models (EpiVaginal). We ascertained that epithelial cells in the tissue models express estrogen and progesterone receptors. Treatment with estradiol-17ß (E(2)) significantly increased epithelium thickness and transepithelial electrical resistance (TEER), whereas progesterone (P) treatment resulted in thinning of the epithelium and decreased TEER when compared with untreated controls. Exposure to E(2) increased (1) the expression of the progesterone receptor B (PR-B), (2) accumulation of glycogen in suprabasal cells, (3) epithelial differentiation, and (4) the expression of a number of gene pathways associated with innate immunity, epithelial differentiation, wound healing, and antiviral responses. These findings indicate that EpiVaginal tissues are hormone responsive and can be used to study the role of female reproductive hormones in innate immune responses, microbial infection, and drug delivery in the vaginal mucosa.

Applying antithrombotic therapies to improve outcomes in patients with atrial fibrillation.

Approximately 15% to 25% or 75,000 ischemic strokes are attributed to atrial fibrillation annually within the United States. Atrial fibrillation is the most frequently diagnosed cardiac arrhythmia and affects more than 2.66 million Americans. Moreover, atrial fibrillation is associated with a 1.5 to 1.9-fold higher risk of death due to its strong correlation with thromboembolic events. Because of the attributed increased morbidity and mortality, challenges that concern identification of patients at risk for thromboembolic events from atrial fibrillation must be addressed. These challenges include compliance to performance measures, adherence to guidelines, adequate prevention and early control of co-morbidities that affect the progression of atrial fibrillation and associated risks, early initiation of treatment, and successful evaluation of associated risks of bleeding, primary or recurrent stroke, and patient awareness and compliance. This multimedia educational webcast will discuss the state of affairs with respect to antithrombotic therapies and new anticoagulants. The webcast will also review factors influencing physician use of anticoagulation in atrial fibrillation. Clinical decision making and lessons learned from the expert faculty is also included.

Development of an in vitro alternative assay method for vaginal irritation.

The vaginal mucosa is commonly exposed to chemicals and therapeutic agents that may result in irritation and/or inflammation. In addition to acute effects, vaginal irritation and inflammation can make women more susceptible to infections such as HIV-1 and herpes simplex virus 2 (HSV-2). Hence, the vaginal irritation potential of feminine care formulations and vaginally administered therapeutic agents is a significant public health concern. Traditionally, testing of such materials has been performed using the rabbit vaginal irritation (RVI) assay. In the current study, we investigated whether the organotypic, highly differentiated EpiVaginal™ tissue could be used as a non-animal alternative to the RVI test. The EpiVaginal tissue was exposed to a single application of ingredients commonly found in feminine hygiene products and the effects on tissue viability (MTT assay), barrier disruption (measured by transepithelial electrical resistance, TEER and sodium fluorescein (NaFl) leakage), and inflammatory cytokine release (interleukin (IL)-1α, IL-1ß, IL-6, and IL-8) patterns were examined. When compared to untreated controls, two irritating ingredients, nonoxynol 9 and benzalkonium chloride, reduced tissue viability to <40% and TEER to <60% while increasing NaFl leakage by 11-24% and IL-1α and IL-1ß release by >100%. Four other non-irritating materials had minimal effects on these parameters. Assay reproducibility was confirmed by testing the chemicals using three different tissue production lots and by using tissues reconstructed from cells obtained from three different donors. Coefficients of variation between tissue lots reconstructed with cells obtained from the same donor or lots reconstructed with cells obtained from different donors were less than 10% and 12%, respectively. In conclusion, decreases in tissue viability and barrier function and increases in IL-1α and IL-1ß release appear to be useful endpoints for preclinical screening of topically applied chemicals and formulations for their vaginal irritation potential.

Organotypic human vaginal-ectocervical tissue model for irritation studies of spermicides, microbicides, and feminine-care products.

A three-dimensional organotypic vaginal-ectocervical (VEC) tissue model has been developed to test the irritation of topically applied spermicides, microbicides, and vaginal-care products. The in vitro tissue model was reconstructed using normal VEC epithelial cells and is well stratified, containing differentiated basal, suprabasal, intermediate, and superficial cell layers similar to in vivo tissue. The intermediate and superficial cell layers contain glycogen, and the expression of cytokeratins 13 and 14 in the tissue also parallels that of native tissue. The MTT viability assay and histological assessment were used to test inter-lot and intra-lot reproducibility. The MTT average intra-lot coefficient of variation (CV) was less than 10% and the time required to reduce tissue viability by 50% (ET-50) following application of 1% Triton X-100 averaged 1.25+/-0.24h (n=23) upon completion of the 11-day culture period and 1.30 h+/- 0.19 for the same tissues stored overnight at 4 degrees C on agarose gels. The utility of the VEC model for irritation studies was examined by testing commercially available products using the MTT assay and histological assessment. The average ET-50 values ranged between 1.8 and 2.7h for feminine washes, 3.9-6.7 h for spermicides, 6.8-18 h for anti-itch creams, and >18 h for douches, lubricants, and anti-fungal creams. Studies of cytokines released from VEC cultures following product application showed that elevated concentrations of IL-1alpha and IL-1beta were associated with toxicity of test materials. In conclusion, the VEC tissue model is a highly reproducible, non-animal means to assess the irritation of contraceptives, microbicides, and vaginal-care products.