Development of an in vitro alternative assay method for vaginal irritation

Abstract

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. 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-1β, 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-1β 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-1β release appear to be useful endpoints for preclinical screening of topically applied chemicals and formulations for their vaginal irritation potential.

Preclinical evaluation of UC781 microbicide vaginal drug delivery

“…In these experiments, the AP and BL sides were sampled at 0 and 180 min or 30, 60, 120, and 180 min, respectively. Permeability data with these control compounds were consistent with intact, functional tissues [12,13].…”

“…If the ET-50 was ≥ 0.70 h, the tissues were considered acceptable. In both cases (positive and negative controls), the tissues passed these acceptance criteria [12,13].…”

“…While the release of UC781 from the gel prepared with micronized drug was more rapid compared with the nonmicronized UC781, the differences were insignificant. To examine the impact of particle size and drug concentration of delivery into and through tissues, a multilayered human-derived vaginal-ectocervical tissue model (EpiVaginal) was used [12,13]. Preliminary experiments examining tissue delivery of UC781 from gel prepared with UC781 nm or UC781 m at 0.1% and 0.25% concentrations suggested that the nonmicronized form of the drug led to higher tissue concentrations compared with that from the micronized UC781 following 2 h of gel exposure.…”

Impact of Hydroxychloroquine-Loaded Polyurethane Intravaginal Rings on Lactobacilli

“…Although we showed that our HCQ-loaded IVR segments had no significant effects on the viability of vaginal or ectocervical epithelial cells, CD4 ϩ T cells, and lactobacilli, it was not known whether there were any changes in the tight junctions of vaginal epithelial cells, which is a concern since this may increase the opportunities for viruses to penetrate the vaginal mucosal layer and to infect underlying immune cells (38). As a result, we evaluated TEER values for VK2/E6E7 cell monolayers before and after exposure to HCQ alone, drug-free IVR segments, or HCQ-loaded IVR segments.…”

On the Genealogy of Tissue Engineering and Regenerative Medicine

“…102 Specific examples, which have been reviewed extensively here, 100,102,103 include blood vessels for device (stents) 104,105 and drug testing 106 ; bioengineered blood-brain barriers to test drug permeability 107 and disease modelling [108][109][110] ; musculoskeletal tissue to optimize drugs aimed at improving muscular growth and function, 111 as well as to evaluate the impact of prosthetic devices on muscular tissues. 112 Corneal tissue to conduct tests for toxicology and eye irritancy 113 ; skin to test for cytotoxicity, 114 phototoxicity, 115,116 and wound healing 117 ; and reproductive tissues to study sexually transmitted infections, product efficacy, [118][119][120] and, as models of the blood-testis barrier, to predict toxicity and permeability in vivo, 121,122 form prime examples of in vitro models (reviewed here in Refs. 100,102,103 ).…”