title: Gene Expression Profiling of Human Vaginal Cells In Vitro Discriminates Compounds with Pro-Inflammatory and Mucosa-Altering Properties: Novel Biomarkers for Preclinical Testing of HIV Microbicide Candidates.
abstract: BACKGROUND
  Inflammation and immune activation of the cervicovaginal mucosa are considered factors that increase susceptibility to HIV infection. Therefore, it is essential to screen candidate anti-HIV microbicides for potential mucosal immunomodulatory/inflammatory effects prior to further clinical development. The goal of this study was to develop an in vitro method for preclinical evaluation of the inflammatory potential of new candidate microbicides using a microarray gene expression profiling strategy.
  
  
  METHODS
  To this end, we compared transcriptomes of human vaginal cells (Vk2/E6E7) treated with well-characterized pro-inflammatory (PIC) and non-inflammatory (NIC) compounds. PICs included compounds with different mechanisms of action. Gene expression was analyzed using Affymetrix U133 Plus 2 arrays. Data processing was performed using GeneSpring 11.5 (Agilent Technologies, Santa Clara, CA).
  
  
  RESULTS
  Microarraray comparative analysis allowed us to generate a panel of 20 genes that were consistently deregulated by PICs compared to NICs, thus distinguishing between these two groups. Functional analysis mapped 14 of these genes to immune and inflammatory responses. This was confirmed by the fact that PICs induced NFkB pathway activation in Vk2 cells. By testing microbicide candidates previously characterized in clinical trials we demonstrated that the selected PIC-associated genes properly identified compounds with mucosa-altering effects. The discriminatory power of these genes was further demonstrated after culturing vaginal cells with vaginal bacteria. Prevotella bivia, prevalent bacteria in the disturbed microbiota of bacterial vaginosis, induced strong upregulation of seven selected PIC-associated genes, while a commensal Lactobacillus gasseri associated to vaginal health did not cause any changes.
  
  
  CONCLUSIONS
  In vitro evaluation of the immunoinflammatory potential of microbicides using the PIC-associated genes defined in this study could help in the initial screening of candidates prior to entering clinical trials. Additional characterization of these genes can provide further insight into the cervicovaginal immunoinflammatory and mucosal-altering processes that facilitate or limit HIV transmission with implications for the design of prevention strategies.
authors.affiliation: CONRAD, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, Virginia, United States of America.
authors.forename: Irina A
authors.initials: IA
authors.lastname: Zalenskaya
authors.affiliation: CONRAD, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, Virginia, United States of America.
authors.forename: Theresa
authors.initials: T
authors.lastname: Joseph
authors.affiliation: Virginia Bioinformatics Institute, Virginia Polytechnic Institute and State University, Blacksburg, Virginia, United States of America.
authors.forename: Jasmin
authors.initials: J
authors.lastname: Bavarva
authors.affiliation: CONRAD, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, Virginia, United States of America.
authors.forename: Nazita
authors.initials: N
authors.lastname: Yousefieh
authors.affiliation: CONRAD, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, Virginia, United States of America.
authors.forename: Suzanne S
authors.initials: SS
authors.lastname: Jackson
authors.affiliation: Laboratory of Genital Tract Biology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Titilayo
authors.initials: T
authors.lastname: Fashemi
authors.affiliation: Laboratory of Genital Tract Biology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Hidemi S
authors.initials: HS
authors.lastname: Yamamoto
authors.affiliation: Virginia Bioinformatics Institute, Virginia Polytechnic Institute and State University, Blacksburg, Virginia, United States of America.
authors.forename: Robert
authors.initials: R
authors.lastname: Settlage
authors.affiliation: Laboratory of Genital Tract Biology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Raina N
authors.initials: RN
authors.lastname: Fichorova
authors.affiliation: CONRAD, Department of Obstetrics and Gynecology, Eastern Virginia Medical School, Norfolk, Virginia, United States of America.
authors.forename: Gustavo F
authors.initials: GF
authors.lastname: Doncel
mesh_terms: D000890:Anti-Infective Agents / Q000494:pharmacology / Q000627:therapeutic use*; D001419:Bacteria / Q000187:drug effects / Q000254:growth & development; D015415:Biomarkers / Q000378:metabolism; D002460:Cell Line; D016000:Cluster Analysis; D015169:Colony Count, Microbial; D004353:Drug Evaluation, Preclinical; D005260:Female; D020869:Gene Expression Profiling; D053263:Gene Regulatory Networks / Q000187:drug effects; D015658:HIV Infections / Q000188:drug therapy*; D006801:Humans; D007155:Immunologic Factors / Q000494:pharmacology; D007249:Inflammation / Q000473:pathology*; D008826:Microbial Sensitivity Tests; D008954:Models, Biological; D009092:Mucous Membrane / Q000187:drug effects / Q000473:pathology*; D016328:NF-kappa B / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D014158:Transcription, Genetic / Q000187:drug effects; D014621:Vagina / Q000166:cytology* / Q000382:microbiology
chemical_list: D000890:Anti-Infective Agents; D015415:Biomarkers; D007155:Immunologic Factors; D016328:NF-kappa B