title: Multifocal vulvar intraepithelial neoplasia grade III and multicentric lower genital tract neoplasia is associated with transcriptionally active human papillomavirus.
abstract: BACKGROUND
  The incidence of vulvar intraepithelial neoplasia Grade III (VIN III) is increasing and is diagnosed at a younger age than previously. VIN III is often multifocal and frequently coexists with multicentric dysplastic lesions in the cervix and vagina. Warty-type VIN III more often has been found to contain human papillomavirus (HPV) DNA than basaloid-type VIN III: The authors performed HPV DNA polymerase chain reaction (PCR) analysis in 48 VIN III biopsies and reverse transcriptase (RT)-PCR in 8 HPV-16 DNA-positive multifocal VIN III biopsies to detect E6/E7 transcripts.
  
  
  METHODS
  Human papillomavirus DNA detection and histologic analysis were performed on alternating slides of paraffin embedded biopsies. Polymerase chain reaction was performed with consensus primers, and HPV typing was performed by direct sequencing. Total RNA was isolated from frozen biopsies by centrifuging a guanidinium thiocyanate (GTC) lysate through a cesium chloride (CsCl) cushion. The RT reaction was performed using a 3' primer, located just downstream of the E7 stop codon, and the PCR reaction was performed using the same 3' primer and a 5' primer located just downstream of the E6 start codon.
  
  
  RESULTS
  The mean age of the 48 patients was 37.7 years. Eighty-one percent had multifocal VIN III: Sixty-six percent had multicentric neoplasia. Forty-six percent of the biopsies were warty-type, 17% basaloid-type, 35% mixed-type and 2% differentiated-type. Ninety-two percent were HPV-positive and 83% contained HPV-16 DNA. Human papillomavirus DNA was more often present in multifocal VIN III lesions than in unifocal VIN III lesions and also more often in VIN III lesions coexisting with other dysplastic multicentric lesions than in unicentric VIN III lesions. Warty-type VIN III more often contained koilocytes than basaloid-type VIN III: A correlation between different morphologic forms of VIN III and the presence of HPV DNA was not found. Both types of VIN III often coexist in one lesion. In all the RT-PCRs, a 593-base-pair fragment was detected, corresponding to the expected length of the major E6*-E7 mRNA.
  
  
  CONCLUSIONS
  The observed high prevalence of transcriptionally active HPV DNA associated with multifocal and multicentric dysplasia suggests a role of HPV in the pathogenesis of these lesions. A positive correlation between different morphologic forms of VIN III and the presence of HPV DNA was not found.
authors.affiliation: Department of Gynecology, University of Amsterdam, The Netherlands.
authors.forename: M
authors.initials: M
authors.lastname: van Beurden
authors.forename: F J
authors.initials: FJ
authors.lastname: ten Kate
authors.forename: H L
authors.initials: HL
authors.lastname: Smits
authors.forename: R J
authors.initials: RJ
authors.lastname: Berkhout
authors.forename: A J
authors.initials: AJ
authors.lastname: de Craen
authors.forename: N
authors.initials: N
authors.lastname: van der Vange
authors.forename: F B
authors.initials: FB
authors.lastname: Lammes
authors.forename: J
authors.initials: J
authors.lastname: ter Schegget
mesh_terms: D000328:Adult; D000368:Aged; D002278:Carcinoma in Situ / Q000473:pathology / Q000821:virology*; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D006801:Humans; D008875:Middle Aged; D027383:Papillomaviridae / Q000235:genetics / Q000302:isolation & purification*; D015533:Transcriptional Activation; D002583:Uterine Cervical Neoplasms / Q000821:virology*; D014625:Vaginal Neoplasms / Q000821:virology*; D014846:Vulvar Neoplasms / Q000473:pathology / Q000821:virology*
chemical_list: D004279:DNA, Viral