Abstract
BackgroundThe development of safe topical microbicides that can preserve the integrity of cervicovaginal tract epithelial barrier is of great interest as this may minimize the potential for increased susceptibility to STI infections. High resolution imaging to assess epithelial integrity in a noninvasive manner could be a valuable tool for preclinical testing of candidate topical agents.MethodsA quantitative approach using confocal fluorescence microendoscopy (CFM) for assessment of microbicide-induced injury to the vaginal epithelium was developed. Sheep were treated intravaginally with one of five agents in solution (PBS; 0.02% benzalkonium chloride (BZK); 0.2% BZK) or gel formulation (hydroxyethyl cellulose (HEC); Gynol II nonoxynol-9 gel (N-9)). After 24 hours the vaginal tract was removed, labeled with propidium iodide (PI), imaged, then fixed for histology. An automated image scoring algorithm was developed for quantitative assessment of injury and applied to the data set. Image-based findings were validated with histological visual gradings that describe degree of injury and measurement of epithelial thickness.ResultsDistinct differences in PI staining were detected following BZK and N-9 treatment. Images from controls had uniformly distributed nuclei with defined borders, while those after BZK or N-9 showed heavily stained and disrupted nuclei, which increased in proportion to injury detected on histology. The confocal scoring system revealed statistically significant scores for each agent versus PBS controls with the exception of HEC and were consistent with histology scores of injury.ConclusionsConfocal microendoscopy provides a sensitive, objective, and quantitative approach for non-invasive assessment of vaginal epithelial integrity and could serve as a tool for real-time safety evaluation of emerging intravaginal topical agents.
Highlights
The development of safe topical microbicides that can preserve the integrity of cervicovaginal tract epithelial barrier is of great interest as this may minimize the potential for increased susceptibility to STI infections
We investigate the ability of a fixed-plane surface imaging confocal fluorescence microendoscopy (CFM) to visualize and quantify epithelial injury in the ovine cervicovaginal tract following treatment with solutions and gels commonly studied in microbicide research
Histology from PBS treated sheep confirmed the presence of a cornified stratified squamous epithelium (Figure 1)
Summary
The development of safe topical microbicides that can preserve the integrity of cervicovaginal tract epithelial barrier is of great interest as this may minimize the potential for increased susceptibility to STI infections. High resolution imaging to assess epithelial integrity in a noninvasive manner could be a valuable tool for preclinical testing of candidate topical agents. Effective preclinical microbicide safety testing is of great importance to insure that only the most promising candidates are advanced to clinical trials. A number of methods are currently used for assessing microbicide effects on the cervicovaginal tract. Colposcopy is implemented and allows repeated evaluation Disadvantages are it only allows visual assessment of the tissue surface albeit with magnification, colposcopy training requirements are intensive, and interpretation is subjective and only modestly quantitative. Histopathology requires biopsy and processing of tissue and results in days to weeks of delay. It is both cumbersome and raises safety issues when used clinically in high-risk populations. Cytokine mapping following vaginal lavage requires offline testing, but has the advantage of being compatible with other methods and can be repeated in longitudinal studies
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