William A. Rose

Quantification and comparison of toll-like receptor expression and responsiveness in primary and immortalized human female lower genital tract epithelia.

To better understand innate immune responses to sexually‐transmitted infection (STI) and the appropriateness of epithelial cell (EC) models of the vaginal and cervical mucosa, quantified toll‐like receptor (TLR) expression from a population of women is needed.

Commensal bacteria modulate innate immune responses of vaginal epithelial cell multilayer cultures.

The human vaginal microbiome plays a critical but poorly defined role in reproductive health. Vaginal microbiome alterations are associated with increased susceptibility to sexually-transmitted infections (STI) possibly due to related changes in innate defense responses from epithelial cells. Study of the impact of commensal bacteria on the vaginal mucosal surface has been hindered by current vaginal epithelial cell (VEC) culture systems that lack an appropriate interface between the apical surface of stratified squamous epithelium and the air-filled vaginal lumen. Therefore we developed a reproducible multilayer VEC culture system with an apical (luminal) air-interface that supported colonization with selected commensal bacteria. Multilayer VEC developed tight-junctions and other hallmarks of the vaginal mucosa including predictable proinflammatory cytokine secretion following TLR stimulation. Colonization of multilayers by common vaginal commensals including Lactobacillus crispatus, L. jensenii, and L. rhamnosus led to intimate associations with the VEC exclusively on the apical surface. Vaginal commensals did not trigger cytokine secretion but Staphylococcus epidermidis, a skin commensal, was inflammatory. Lactobacilli reduced cytokine secretion in an isolate-specific fashion following TLR stimulation. This tempering of inflammation offers a potential explanation for increased susceptibility to STI in the absence of common commensals and has implications for testing of potential STI preventatives.

ORIGINAL ARTICLE: Quantification and Comparison of Toll-Like Receptor Expression and Responsiveness in Primary and Immortalized Human Female Lower Genital Tract Epithelia

To better understand innate immune responses to sexually‐transmitted infection (STI) and the appropriateness of epithelial cell (EC) models of the vaginal and cervical mucosa, quantified toll‐like receptor (TLR) expression from a population of women is needed.

FSL-1, a bacterial-derived toll-like receptor 2/6 agonist, enhances resistance to experimental HSV-2 infection.

BackgroundHerpes simplex virus type 2 (HSV-2) is a leading cause of genital ulceration that can predispose individuals to an increased risk of acquiring other sexually transmitted infections. There are no approved HSV-2 vaccines and current suppressive therapies require daily compound administration that does not prevent all recurrences. A promising experimental strategy is the use of toll-like receptor (TLR) agonists to induce an innate immune response that provides resistance to HSV-2 infection. Previous studies showed that anti-herpetic activity varied based on origin of the agonists and activation of different TLR indicating that activity likely occurs through elaboration of a specific innate immune response. To test the hypothesis, we evaluated the ability of a bacterial-derived TLR2/6 agonist (FSL-1) to increase resistance to experimental genital HSV-2 infection.MethodsVaginal application of FSL-1 at selected doses and times was evaluated to identify potential increased resistance to genital HSV-2 infection in the mouse model. The FSL-1 induced cytokine profile was quantified using kinetically collected vaginal lavages. Additionally, cytokine elaboration and organ weights were evaluated after single or multiple FSL-1 doses to establish a preliminary safety profile. Human vaginal EC cultures were used to confirm the mouse model outcomes.ResultsThe results showed that vaginally-applied FSL-1 created an environment resistant to a 25-fold higher HSV-2 challenge dose. Mechanistically, vaginal FSL-1 application led to transient elaboration of cytokines linked to anti-herpetic innate immune responses. No gross local or peripheral immunotoxicity was observed even after multiple dosing. FSL-1 also created an anti-herpetic environment in cultures of human vaginal epithelial cells (EC).ConclusionThe results showed, for the first time, that the bacterial-derived TLR2/6 agonist FSL-1 induced significant resistance to HSV-2 infection when applied in mice or human vaginal EC cultures. Cytokine evaluation illustrated that anti-herpetic activity correlated with induction of a specific profile. The identified anti-herpetic profile provides an invaluable resource for the future design of novel compounds to reduce genital HSV-2 transmission and improves understanding of the complex innate immune response to potential pathogens elicited by the vaginal mucosa.

Scanning Cytometry with a LEAP: Laser-Enabled Analysis and Processing of Live Cells In Situ

Scanning cytometry now has many of the features (and power) of multiparameter flow cytometry while keeping its own advantages as an imaging technology. Modern instruments combine capabilities of scanning cytometry with the ability to manipulate cells. A new technology, called LEAP™ (laser‐enabled analysis and processing), offers a unique combination of capabilities in cell purification and selective macromolecule delivery (optoinjection).

An immunomodulating dipeptide, SCV-07, is a potential therapeutic for recurrent genital herpes simplex virus type 2 (HSV-2)

Herpes simplex virus type 2 (HSV-2) infections produce a recurrent disease state associated with susceptibility to other pathogens, including human immunodeficiency virus (HIV), and cannot be cured by current therapeutic treatments. The HSV-2 epidemic must therefore be addressed by therapeutic strategies that reduce recurrent lesions and ideally lack the possibility for development of drug resistance. To this end, the therapeutic potential of SCV-07 (gamma-D-glutamyl-L-tryptophan), a synthetic dipeptide with potent immunomodulatory and antimicrobial activity, was studied in the guinea pig model of recurrent genital HSV-2. Initial evaluations showed that when delivered orally, but not subcutaneously, SCV-07 significantly reduced recurrent lesions. Oral dose ranging studies indicated that, of the tested amounts, 5microg/kg was optimal when delivered after an overnight fast. Interestingly, fasting induced a significant increase in recurrent lesions in vehicle-treated guinea pigs relative to non-fasted animals. Despite this increase, SCV-07 significantly reduced lesion formation in treated animals but showed no durability following cessation of treatment. In fact, this regimen of SCV-07 treatment produced statistically indistinguishable outcomes compared with those provided by topical aciclovir. These data illustrate that SCV-07 may provide an easily administered alternative or supplemental treatment option for genital HSV-2 recurrent disease.

ORIGINAL ARTICLE: Quantification and Comparison of Toll-Like Receptor Expression and Responsiveness in Primary and Immortalized Human Female Lower Genital Tract Epithelia: HUMAN FEMALE GENITAL EPITHELIA TLR EXPRESSION

To better understand innate immune responses to sexually‐transmitted infection (STI) and the appropriateness of epithelial cell (EC) models of the vaginal and cervical mucosa, quantified toll‐like receptor (TLR) expression from a population of women is needed.